key: cord-285428-fcrunf38 authors: halstead, scott b. title: covid-19: the need for immunoprevention at industrial scale date: 2020-04-08 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0239 sha: doc_id: 285428 cord_uid: fcrunf38 novel coronavirus disease (covid-19): the need for immunoprevention at industrial scale. severe acute respiratory syndrome-cov-2 infections are exacting devastating mortality in elderly persons with preexisting health conditions. this has led to widespread quarantining, suspension of social and business activities, frightening levels of morbidity and mortality, and enormous economic loss. as in the past, the nation has quickly found technical solutions to cope with this challenge. industry has rapidly provided improved tests to detect viral rna, respirator masks to prevent infections, and ventilators for critical intensive care; however, due to the scale of the pandemic, large needs remain. these and other requirements identified by the u.s. coronavirus task force have come from rapid and generous responses by industry. can we prevent these deaths? vaccines, antiviral drugs, and antibodies are our tools of choice. the best option would be a single-dose vaccine. efforts to develop vaccines against coronavirus disease-19 (covid-19) are well advanced, based in part on experience with severe acute respiratory syndrome (sars) and middle east respiratory syndrome (mers). 1 however, before release, vaccines must demonstrate relevant immune responses in human volunteers, absence of unwanted side effects, and protection of humans from disease. these steps take many months to complete. antivirals have already achieved wide attention. a near-term solution is to repurpose existing drugs. remdesivir, a nucleotide analog, is under clinical investigation in china and elsewhere. this drug has shown efficacy against mers disease in a monkey model. 2 antibodies from convalescents or monoclonal antibodies derived from immunes, given early in illness, may reduce disease severity and save lives. 3 convalescent sars antibodies given early in illness have been shown to reduce disease severity. 4 in addition, it is likely that entirely new useful compounds will emerge from the laboratory. prevention of covid-19 may be possible using antibodies. previously in the united states, commercial gamma globulin provided short-term prevention against measles, paralytic poliomyelitis, and hepatitis a. [5] [6] [7] [8] in the 1950s, william mcd hammon at the university of pittsburgh demonstrated that gamma globulin given to 100,000 children in a blinded efficacy trial successfully blunted paralysis attack rates during epidemics of poliomyelitis. 7 gamma globulin prepared from immune donors or protective monoclonal antibodies offer possibilities of short-term protection for care givers and healthcare workers and, in particular, for those at high risk of severe or fatal covid-19. this latter group can be well identified by carefully designed epidemiological studies. potential products must demonstrate protection in animal models and in small clinical trials. to avoid possible enhancement of covid-19, antibodies might be given to prevent sars-cov-2 infections after the removal or inactivation of the igg fc terminus. 9 immunoprotection is an especially forceful solution that could be available soon. efforts are well underway by many groups to derive monoclonal antibodies or manufacture gamma globulin from the huge cohort of covid-19 convalescent immunes. 3, 10 moving new products on line for early and effective use requires commitment and leadership. as suggested by safi bahcall (wall street journal, march 21, 2020), a national science leader should immediately be appointed to organize an initial intervention, the direct immune protection of at-risk persons at industrial scale. immune responses in covid-19 and potential vaccines: lessons learned from sars and mers epidemic prophylactic and therapeutic remdesivir (gs-5734) treatment in the rhesus macaque model of mers-cov infection the convalescent sera option for containing covid-19 the effectiveness of convalescent plasma and hyperimmune immunoglobulin for the treatment of severe acute respiratory infections of viral etiology: a systematic review and exploratory meta-analysis postexposure passive immunisation for preventing measles use of concentrated human serum gammaglobulin in the prevention and attenuation of measles passive immunization against poliomyelitis: the hammon gamma globulin field trials infectious hepatitis: current status of prevention with gamma globulin molecular mechanism for antibody-dependent enhancement of coronavirus entry perspectives on monoclonal antibody therapy as potential therapeutic intervention for coronavirus disease-19 (covid-19) key: cord-257377-x5xijo8m authors: ogoina, dimie title: covid-19: the need for rational use of face masks in nigeria date: 2020-05-15 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0433 sha: doc_id: 257377 cord_uid: x5xijo8m because of the pandemic of covid-19, the federal government of nigeria has instituted a mandatory policy requiring everyone going out in public to wear face masks. unfortunately, the nigeria media is awash with images of misuse and abuse of face masks by the public, government officials, and healthcare workers. medical masks are used widely in community settings amid reported scarcity within healthcare facilities. it is observed that some people wear face masks on their chin and neck, and mask wearers give no attention to covering their mouth and nose, especially when talking. used face masks are kept with personal belongings or disposed indiscriminately in public spaces, leading to self and environmental contamination. inappropriate use and disposal of face masks in nigeria could promote the spread of the novel coronavirus in the country and negate the country’s efforts to contain the covid-19 pandemic. in the implementation of the universal masking policy in nigeria, federal and state governments ought to consider local applicability, feasibility, and sustainability, as well as identify and mitigate all potential risks and unintended consequences. also critical is the need for intensive public sensitization and education on appropriate use and disposal of face masks in the country. the major strategic goals of covid-19 control efforts are to slow or stop transmission and spread of sars-cov-2 and to mitigate the impact of the virus on the health system, social activities, and economies of countries and communities. 1 the scientific information on sars-cov-2 and covid-19 is rapidly evolving, and many countries are adopting preventive measures based on emerging evidence and local applicability. one such strategy, considering evidence of presymptomatic and asymptomatic transmissions of sars-cov-2, is the use of face masks by apparently healthy persons to slow community transmission of the virus. [2] [3] [4] the use of public face masks for the prevention of covid-19 is controversial. the who has indicated that it cannot recommend for or against public use of face masks, as there is yet no clear evidence that this practice is effective in the prevention of covid-19. 5 the who has also emphasized that "the use of a mask alone is insufficient to provide an adequate level of protection, and other measures should also be adopted." however, the who acknowledged potential advantages of the use of masks by healthy people in the community setting to reduce potential exposure from infected persons during the presymptomatic period of infection. it advised a risk-based approach for implementing policies on public wearing of face masks. 5 as part of a comprehensive response to the covid-19 epidemic, the president of the federal republic of nigeria recently announced mandatory wearing of face masks by anyone going out in public. 6 a similar policy is being implemented by almost all state governments of the country. the nigerian centre of disease control (ncdc) indicated that the major rationale for public wearing of face masks is to prevent those who are infected but asymptomatic from spreading the virus. 7 the ncdc has emphasized that wearing of face masks may only be effective in preventing the transmission of sars-cov-2 if they are worn and disposed appropriately, and if mask wearing is combined with other preventive measures such as hand hygiene and social distancing. an advisory on making and proper usage of cloth masks has also been issued by the ncdc; it was recommended that cloth masks be used by the general public, with medical masks reserved for healthcare workers. 8 unfortunately, new face mask policies are leading to widespread misuse and abuse of face masks in nigeria. [9] [10] [11] [12] the nigeria media is awash with images of members of the general public, including healthcare workers and government officials, wearing face masks on their jaws and neck, without covering their mouth or nose, or covering only their mouth while the nose is left opened. many people who use face masks are commonly observed to pull down their mask to their jaw to talk and then pull it back over their mouth and nose after talking. a variety of cloth masks of doubtful efficacy are hawked on the streets and tried by different wearers before deciding on purchase. people are also observed to repeatedly touch the front of their face masks in a bid to adjust the mask, to remove it, or during reflex touching of the face. some wear one mask for prolonged periods, without replacement when it is wet or soiled. furthermore, face masks used for the prevention of covid-19 by the general public are being disposed inappropriately. 11 the rising spate of misuse and abuse of face masks is a source of worry for the nigerian covid-19 presidential task force, which observed "unhygienic and ill-advised use and sharing of masks, especially multiple fittings before buying from vendors." 13 it is noteworthy that medical masks meant for healthcare workers, such as surgical masks and respirators, are being routinely worn by the general public and government officials, when there are complaints that these masks are not available in sufficient quantities in nigerian hospitals. 14 furthermore, n95 respirators with exhalation valves have become the preferred face masks by many, including top government officials, possibly because they are more comfortable. respirators with exhalation valves are, however, not effective in covid-19 source control, as they do not prevent the release of exhaled respiratory particles. although some state governments in nigeria are now producing large quantities of cloth masks for use by the general population, 10 one is concerned if sufficient quantities of these homemade masks can be provided sustainably for a population of more than 200 million. there are also challenges related to enforcement of mandatory usage, access to water and soap to properly wash and reuse homemade masks, and a false sense of security that may lead people to abandon other preventive measures because of the usage of face masks. compliance with and enforcement of social distancing measures have been identified as challenges in nigeria's response to covid-19, and there are reports of people openly flouting lockdown orders and other preventive measures. 15 many nigerians still do not have access to water and basic sanitation, and indices of hygiene are poor in the country. 16 adopting a public face mask strategy may require extra funding to be sustainable, and this strategy could divert scarce resources from other covid-19 preventive measures. another concern is that face masks might become a new medium for propagation of the novel coronavirus in nigeria in view of high risks of self and environmental contamination when masks are used and disposed inappropriately. public mask wearing is most effective at stopping the spread of the novel coronavirus when compliance is high and when masks are used appropriately, especially in combination with other preventive measures such as hand hygiene. 5, 17 in many of its public health advisories, including the advisory on use of face masks by the healthy community, 5 the who cautioned on universal applicability of public health measures without consideration of local context. in the absence of an effective and intensive communication strategy on why, when, and how to use face masks, the strategy of mandatory public face mask use in nigeria carries risks and uncertainties. the federal and state governments ought to consider the local peculiarities, resource requirements, feasibility, sustainability, and potential risks and benefits before and during the implementation of public face mask policies. failure to address these may negate the potential benefits of public face mask policies and inadvertently enhance the spread of sars-cov-2 in nigeria. available at: https:// www.who.int/publications-detail/strategic-preparedness-andresponse-plan-for-the-new-coronavirus. accessed may 5, 2020. 2. world health organisation recommendation regarding the use of cloth face coverings rational use of face masks in the covid-19 pandemic advice on the use of masks in the context of covid-19 covid-19 regulation 2020 nigeria centre for disease control, 2020. advisory on the use of masks by members of the public without respiratory symptoms. available at advisory on use of cloth face masks available at: https:// www.nafdac.gov.ng/covid-19-personal-protection-equipmentppes-masks-and-protective-clothing nids warns against misuse, abuse of face masks to prevent covid-19. independent newspapers nigeria improper use, disposal of facemasks, hand gloves could trigger covid-19 community transmission in nigeria. the guardian newspaper bizzare! these women were seen washing used nose mask in order to resell! naija super fans remarks by the sgf/chairman of the ptf covid-19 at the national briefing of wednesday nma: shortage of protective equipment due to poor funding for health-the whistler ng. the whistler independent newspapers cyclists for flouting lockdown order. independent newspapers nigeria nigeria demographic and health survey covid-19 epidemic: disentangling the re-emerging controversy about medical facemasks from an epidemiological perspective key: cord-318994-1xngjndu authors: lier, audun j.; tuan, jessica j.; davis, matthew w.; paulson, nathan; mcmanus, dayna; campbell, sheldon; peaper, david r.; topal, jeffrey e. title: case report: disseminated strongyloidiasis in a patient with covid-19 date: 2020-08-14 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0699 sha: doc_id: 318994 cord_uid: 1xngjndu the sars-cov-2 virus has emerged and rapidly evolved into a current global pandemic. although bacterial and fungal coinfections have been associated with covid-19, little is known about parasitic infection. we report a case of a covid-19 patient who developed disseminated strongyloidiasis following treatment with high-dose corticosteroids and tocilizumab. screening for strongyloides infection should be pursued in individuals with covid-19 who originate from endemic regions before initiating immunosuppressive therapy. in december 2019, an outbreak of sars cov-2 emerged in wuhan, china-rapidly evolving into a global pandemic of covid-19. a single-center retrospective case series demonstrated bacterial and fungal coinfections among covid-19positive individuals. 1 further research is needed to investigate the relationship between covid-19 and parasitic coinfection, particularly given the fact that patients with covid-19 could receive immunosuppressive treatment, a potential risk factor for severe parasitic infection. this report describes the clinical features of a case of disseminated strongyloidiasis infection and polymicrobial bacteremia in an individual who received immunosuppressive treatment for covid-19. a 68-year-old man presented to our institution with an 8-day history of chills, myalgia, headache, cough, nausea, and worsening dyspnea. the patient denied rhinorrhea, anosmia, dysgeusia, diarrhea, or abdominal pain. past medical history was significant for hypertension and diabetes mellitus complicated by peripheral neuropathy. twenty years before, he had emigrated from sucúa, ecuador, and now resides in connecticut. in ecuador, he worked in a timber industry and also cultivated soil on farms. on admission, temperature was 38.2°c, blood pressure 137/68 mm hg, pulse 97 beats/minute, respiratory rate 24 breaths/minute, and oxygen saturation 96% on 2 l/minute of supplemental oxygen. on physical examination, he had dry mucous membranes and decreased air entry with bibasilar crackles. admission blood work was notable for a white cell count of 10,200/ml 3 (absolute lymphocyte count of 700/ml 3 and absolute eosinophil count of 0/ml 3 ), high-sensitivity c-reactive protein 178.8 mg/l, ferritin 885 mg/ml, d-dimer 1,200 ng/ml, hemoglobin a1c 6.5%. the admission chest x-ray was notable for bilateral patchy airspace opacities in the mid to lower lung zones. sars-cov-2 rna was detected from a nasopharyngeal swab using the cepheid xpert xpress sars-cov-2 assay. sputum culture revealed normal commensal flora. he was admitted to the medicine unit and initiated on 5 days of hydroxychloroquine (400 mg oral twice daily loading dose and then 200 mg oral twice daily). following admission, he developed hypoxemic respiratory failure requiring intubation. tocilizumab (once, intravenous at 8 mg/kg) was given as well as three courses of methylprednisolone (40 mg intravenous every 8 hours) on hospital days 4-6, 8-10, and 12-13 because of persistent hypoxemia and a new fever that manifested on day 9. on hospital day 12, he developed hypotension requiring norepinephrine for blood pressure support. streptococcus constellatus and citrobacter freundii were isolated on blood culture. sputum culture from day 12 grew pseudomonas aeruginosa and methicillin-susceptible staphylococcus aureus. methylprednisolone was discontinued, and he was initiated on intravenous ciprofloxacin, cefazolin, and metronidazole with resolution of fever and hypotension. repeat blood cultures on days 14 and 16 revealed no growth. on hospital day 18, he developed a new fever to 38.8°c while on the aforementioned antibiotic regimen without change to his ventilation settings. absolute eosinophil count was 200/ml 3 . antibiotics were changed to intravenous vancomycin and ciprofloxacin. his oxygenation improved, and he was extubated the following day. sputum culture obtained on the same day grew p. aeruginosa and methicillin-sensitive s. aureus. the next day, serpiginous tracks were noted on a chocolate agar plate ( figure 1 ). gram and iodine stains revealed larvae measuring 280-300 μm with a short buccal canal and prominent genital primordium consistent with strongyloides species. the infectious diseases service was consulted and recommended initiation of ivermectin (200 μg/kg orally daily) for presumed strongyloidiasis and discontinuation of antibiotics. on hospital day 21, his white cell count increased to 39,200/ ml 3 with an absolute eosinophil count of 100/ml 3 . chest x-ray revealed unchanged multifocal bilateral pulmonary opacities. given lack of significant clinical response to ivermectin, albendazole (400 mg orally every 12 hours) was added as adjunctive therapy. the evidence for this combination is low, but some efficacy has been seen in case reports. 2,3 piperacillintazobactam was initiated for suspected nosocomial pneumonia. in the ensuing days, his leukocytosis decreased. strongyloides serum antibody and stool analysis for ova and parasites were negative. hiv and htlv-1 serology were also negative. on hospital day 25, he developed confusion, a new fever (39.4°c), hypotension requiring norepinephrine, and was subsequently reintubated. white cell count was 43,000/ml 3 . blood cultures grew coagulase-negative staphylococcus in one set. repeat sputum culture grew p. aeruginosa, but no larvae were identified. a repeat stool ova and parasite was negative. antibiotics were modified to intravenous vancomycin, ceftazidime, and metronidazole for possible bacterial meningitis associated with disseminated strongyloidiasis, in addition to other possible nosocomial infections. because of hemodynamic instability, a lumbar puncture was deferred. in the ensuing days, his fever and hypotension resolved, mental status improved, and white cell count normalized. notably, the absolute eosinophil count had peaked at 1,900/ml 3 on day 28 and downtrended to 900/ml 3 on day 33. a ct scan on hospital day 30 found widespread peripheral ground-glass opacities and peribronchial consolidation in the right lower lobe. repeat blood and sputum cultures yielded no growth. a third stool ova and parasite was negative. he completed a 2-week course of ivermectin and albendazole as well as a 3-week antibiotic course for suspected gram-negative meningitis. on day 38, repeat strongyloides serology was positive. he currently awaits placement to a skilled nursing facility. strongyloidiasis is caused by the soil-transmitted nematode strongyloides stercoralis, which is endemic in tropical and subtropical regions with an estimated global prevalence of 30-100 million individuals. 4 our patient likely acquired the infection before emigrating to the united states from ecuador in 2000. a recent study demonstrated that 20.7% of 1,418 serum samples from patients in eight provinces of ecuador, one of which is near sucúa, demonstrated positive antibody to s. stercoralis. 5 strongyloides stercoralis filariform larvae can penetrate the skin from the soil, enter the bloodstream, and subsequently migrate into the pulmonary vasculature, alveoli, and lungs. via various mechanisms, larvae enter into the small intestine where they develop into adult female worms and reproduce by parthenogenesis. 4 unique to s. stercoralis among soiltransmitted helminths is that eggs hatch into rhabditiform larvae in the intestine, rather than in the environment, which can reinfect the human host by entering the intestinal wall or perianal skin without exogenous reinfestation. 4, 6 this unique process of autoinfection can yield lifelong, often asymptomatic or subclinical infection. in addition, strongyloidiasis has been associated with immunosuppressive diseases, causing cell-mediated immune deficits including htlv-1 and hiv infection. 4, 6 one of the strongest risk factors associated with strongyloides dissemination or hyperinfection syndrome has been the use of corticosteroids, likely secondary to inhibition of eosinophil and lymphocyte activation. 7 in addition, corticosteroid use increases the fertility of adult female worms, leading to increased numbers of eggs and subsequent rhabditiform larvae. 8 however, there does not appear to be a defined correlation between corticosteroid duration or frequency and development of strongyloidiasis. 7 disseminated strongyloidiasis occurs when larvae invade end organs not classically associated with the normal life cycle of the parasite. gram-negative sepsis and meningitis are attributed to strongyloides larvae assisting enteric bacteria with entering the host's bloodstream via the gut mucosa. 9 our case had several important clinical features. first, in addition to receiving the anti-il-6 receptor antibody tocilizumab, the patient also received three courses of high-dose methylprednisolone to mitigate possible covid-19-related cytokine release syndrome. 10 the most likely predisposing risk factor was receipt of corticosteroids, as tocilizumab alone has not been linked to the development of disseminated strongyloidiasis. a single case report describing disseminated strongyloidiasis after receipt of tocilizumab was in the context of concomitant corticosteroid use (t. t. maffort, unpublished data). a second important clinical consideration is the importance of early suspicion for disseminated strongyloidiasis when associated with gram-negative bacteremia and signs of meningitis. a retrospective study of disseminated strongyloidiasis demonstrated that 23 of 70 cases were associated with sepsis due to enteric organisms. furthermore, 12 of 70 cases had manifestations of meningitis, most frequently with gramnegative organisms, whereas 26 of 70 cases had suppurative culture-negative meningitis. 11 given our patient's epidemiologic risk factor and development of gram-negative bacteremia, the clinical diagnosis of disseminated strongyloidiasis was further supported. finally, our case illustrates the challenge with performing diagnostic testing in immunosuppressed individuals. the initial strongyloides serology on hospital day 21 was negative, but the repeat was positive on day 38. the strongyloides igg serology for the l3 antigen by elisa (new life diagnostics, carlsbad, ca; performed by quest diagnostics, san juan capistrano, ca) has a 92% sensitivity and 100% specificity. 12 consequently, the initial negative serology was likely attributable to the net state of immunosuppression following receipt of multiple courses of high-dose corticosteroids, which has been previously described. 13 in addition, serial stool ova and parasite testing was negative, which may be due to the limited sensitivity of stool testing in the setting of disseminated strongyloidiasis. 13 however, the discovery of tracks of bacterial colonies in sputum culture with identification of rhabditiform larvae with defining morphologic characteristics (appropriate size, short buccal canal, and prominent genital primordium) provided a definitive diagnosis in the clinical context. the covid-19 pandemic has prompted important discussions over its optimal management with the role of corticosteroids and tocilizumab yet to be fully defined. 8 this case highlights important considerations when using immunosuppressive therapies for covid-19 treatment, particularly in patients with risk factors for prior strongyloides infection. clinical suspicion for disseminated strongyloidiasis should be maintained in patients from endemic areas who develop gram-negative sepsis or meningitis. screening is challenging in this clinical setting, given the rapidity with which covid-19 can progress. therefore, before administering immunosuppressive therapy in covid-19 patients, a structured screening mechanism and implementation of a definitive approach may be warranted. we suggest performing strongyloides serology in patients from endemic regions before receiving an immunosuppressive treatment regimen. once the strongyloides serologic status is known, perhaps additional screening with htlv-1 testing and stool analysis could be pursued. clinical features and short-term outcomes of 221 patients with covid-19 in wuhan severe strongyloidiasis: a systematic review of case reports disseminated strongyloidiasis successfully treated with extended duration ivermectin combined with albendazole: a case report of intractable strongyloidiasis mapping the prevalence of strongyloides stercoralis infection in ecuador: a serosurvey human infection with strongyloides stercoralis and other related strongyloides species strongyloides stercoralis in the immunocompromised population dexamethasone effects in the strongyloides venezuelensis infection in a murine model a randomized trial of singleand two-dose ivermectin versus thiabendazole for treatment of strongyloidiasis pharmacologic treatments for coronavirus disease 2019 (covid-19): a review clinical characteristics of disseminated strongyloidiasis severe strongyloidiasis with negative serology after corticosteroid treatment acknowledgments: we would like to acknowledge sid tansey's role in the parasitic diagnosis by noting the track marks in the microbiologic plate. publication charges for this article were waived due to the ongoing pandemic of covid-19. this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-296556-fr8x8j3i authors: chaccour, carlos; hammann, felix; ramón-garcía, santiago; rabinovich, n. regina title: ivermectin and covid-19: keeping rigor in times of urgency date: 2020-04-16 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0271 sha: doc_id: 296556 cord_uid: fr8x8j3i ivermectin and novel coronavirus disease (covid-19): keeping rigor in times of urgency. ivermectin is a widely used drug for the treatment and control of several neglected tropical diseases. 1 the drug has an excellent safety profile, with more than 2.5 billion doses distributed in the last 30 years, and its potential to reduce malaria transmission by killing mosquitoes is under evaluation in several trials around the world. 2 ivermectin inhibits the in vitro replication of some positive, single-stranded rna viruses, namely, dengue virus (dnv), [3] [4] [5] zika virus, 4,6 yellow fever virus, 7,8 and others. 4,7,9 caly et al. 10 recently reported that ivermectin is a potent inhibitor of the severe acute respiratory syndrome coronavirus 2 (sars-cov-2) replication in vitro. given the coronavirus disease-19 pandemic, this has understandably resonated widely in the global press. 11 caly et al. 10 report a 5,000-fold reduction in sars-cov-2 rna levels, compared with those in controls, after infected vero/ hslam cells were incubated for 48 hours with 5 μm ivermectin. the ivermectin ic 50 for the virus was calculated at approximately 2.5 μm. these concentrations are the equivalent of 4,370 and 2,190 ng/ml, respectively, notably 50-to 100-fold the peak concentration (c max ) achieved in plasma after the single dose of 200 μg/kg (14 mg in a 70-kg adult) commonly used for the control of onchocerchiasis. 12 pharmacokinetic studies in healthy volunteers have suggested that single doses up to 120 mg of ivermectin can be safe and well tolerated. 13 however, even with this dose, which is 10-fold greater than those approved by the us food and drug administration, the c max values reported were ∼250 ng/ml, 13 one order of magnitude lower than effective in vitro concentrations against sars-cov-2. these findings may seem to discourage follow-up clinical trials with ivermectin. however, some in vivo effect may be possible even if efficacious in vitro concentrations are physiologically unattainable. a recent phase iii clinical trial in dengue patients in thailand, in which a once-daily dose of 400 μg/kg for 3 days was found to be safe but did not produce any clinical benefit, 14 showed a modest and indirect in vivo effect against dnv. 14 previous work by wagstaff et al. 5 reported inhibition at much higher in vitro concentrations (25 μm) in dnvinfected vero cells. both pharmacokinetic considerations and the relatively long incubation period of dnv might explain the lack of clinical efficacy. until we have a better understanding of ivermectin's antiviral mode of action and of appropriate in vitro systems for testing, we caution against using findings in vero cells as more than a qualitative indicator of potential efficacy. very recently, preliminary findings on a potential effect of hydroxychloroquine combined with azithromycin against sars-cov-2 were widely publicized, 15 leading to a surge in demand and self-medication, which resulted in serious harm in some cases and a stock shortage that jeopardized drug availability for other critical conditions for which hydroxychloroquine or chloroquine is the standard of care, that is, vivax malaria, rheumatoid arthritis, and systemic lupus erythematosus. efficacy claims for hydroxychloroquine against covid-19 have been questioned in follow-up trials using similar dosing regimens, 16, 17 and we await results of randomized, controlled clinical trials exploring treatment efficacy. we believe the recent findings regarding ivermectin warrant rapidly implemented controlled clinical trials to assess its efficacy against sars-cov-2. these trials may open a new field of research on the potential use of avermectin antiparasitic drugs, including compounds with an improved pharmacokinetic profile, as antivirals. 18 however, because of the following points, extreme due diligence and regulatory review are needed before testing ivermectin in severe disease. first, ivermectin, which targets glutamate-gated chlorine channels in invertebrates, may cross-target the gaba-gated chlorine channels present in the mammalian central nervous system (cns) and cause neurotoxicity. 19 this is normally prevented by an intact blood-brain barrier (bbb), but in patients with a hyperinflammatory state, endothelial permeability at the bbb may be increased and cause leaking of drugs into the cns, potentially causing harm. 20, 21 second, boosted antiretrovirals such as lopinavir/ritonavir and darunavir/cobicistat, which have been widely used against sars-cov-2 based on limited evidence, and a number of other drugs, are potent inhibitors of cytochrome p 450 3a4, the main metabolic pathway for ivermectin. concurrent use of these drugs will result in increased systemic exposure to ivermectin. furthermore, ritonavir and cobicistat can readily inhibit one of the main efflux pumps in the bbb, p-glycoprotein, further favoring neurotoxicity. 22, 23 however, it is encouraging that a recent analysis of ivermectin-related neurotoxic adverse events reported to the who program for international drug monitoring found only one case of 1,668 reports in which concomitant use of antivirals was associated with neurotoxicity. 24 third, as earlier, available evidence suggests that levels of ivermectin with meaningful activity against sars-cov-2 would not be achieved without extraordinary, potentially toxic increases in ivermectin dosing levels in humans. however, evidence from animal models showing up to 3-fold higher levels in pulmonary tissue than in plasma 1 week after oral dosing leaves the door open for further research, in particular for the treatment of respiratory viruses. 25, 26 the discovery of ivermectin's activity against sars-cov-2 gives reason for hope, but off-label and compassionate use requires careful risk-benefit considerations, 27 especially in critically ill patients. a path to consider is evaluation first of impacts on virologic outcomes in uncomplicated, low-risk patients early in the course of the disease. well-conducted clinical trials informed by robust pharmacokinetic models should be considered to validate the impact before the use of ivermectin to treat sars-cov-2 is implemented. ivermectin: panacea for resource-poor communities? the ivermectin roadmappers, 2020. a roadmap for the development of ivermectin as a complementary malaria vector control tool nuclear localization of dengue virus (denv) 1-4 non-structural protein 5; protection against all 4 denv serotypes by the inhibitor ivermectin the broad spectrum antiviral ivermectin targets the host nuclear transport importin alpha/beta1 heterodimer ivermectin is a specific inhibitor of importin alpha/betamediated nuclear import able to inhibit replication of hiv-1 and dengue virus a screen of fda-approved drugs for inhibitors of zika virus infection discovery of berberine, abamectin and ivermectin as antivirals against chikungunya and other alphaviruses ivermectin is a potent inhibitor of flavivirus replication specifically targeting ns3 helicase activity: new prospects for an old drug ivermectin inhibits porcine reproductive and respiratory syndrome virus in cultured porcine alveolar macrophages the fdaapproved drug ivermectin inhibits the replication of sars-cov-2 in vitro anti-parasite drug used since 1980s may help stop coronavirus, new study says ivermectin to reduce malaria transmission i. pharmacokinetic and pharmacodynamic considerations regarding efficacy and safety safety, tolerability, and pharmacokinetics of escalating high doses of ivermectin in healthy adult subjects efficacy and safety of ivermectin against dengue infection: a phase iii, randomized, double-blind, placebo-controlled trial. he 34th annual meeting the royal college of physicians of thailand-'internal medicine and one health hydroxychloroquine and azithromycin as a treatment of covid-19: results of an open-label non-randomized clinical trial a pilot study of hydroxychloroquine in treatment of patients with common coronavirus disease-19 (covid-19) no evidence of rapid antiviral clearance or clinical benefit with the combination of hydroxychloroquine and azithromycin in patients with severe covid-19 infection selamectin is the avermectin with the best potential for buruli ulcer treatment relative neurotoxicity of ivermectin and moxidectin in mdr1ab (-/-) mice and effects on mammalian gaba(a) channel activity the blood-brain barrier in systemic inflammation covid-19-associated acute hemorrhagic necrotizing encephalopathy: ct and mri features cobicistat boosts the intestinal absorption of transport substrates, including hiv protease inhibitors and gs-7340, in vitro hiv protease inhibitor ritonavir: a more potent inhibitor of p-glycoprotein than the cyclosporine analog sdz psc 833 serious neurological adverse events after ivermectin-do they occur beyond the indication of onchocerciasis? influence of the route of administration on efficacy and tissue distribution of ivermectin in goat metabolism and tissue residues. campbell wc, ed. ivermectin and abamectin treating covid-19-off-label drug use, compassionate use, and randomized clinical trials during pandemics key: cord-325498-4yciuh1n authors: del brutto, oscar h.; costa, aldo f.; garcía, héctor h. title: incident sars-cov-2 infection and a shared latrine date: 2020-07-22 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0793 sha: doc_id: 325498 cord_uid: 4yciuh1n incident sars-cov-2 infection and a shared latrine. in atahualpa, a rural ecuadorian village, a cohort study has evaluated most adult villagers for factors associated with chronic diseases along the past 9 years. the sars-cov-2 pandemic struck atahualpa in march 2020, as shown by an almost 3-fold excess mortality during april and may, and by a significant number of symptomatic cases. in a recent serosurvey, we found that the use of open latrines (instead of flushing toilet systems) was significantly associated with seropositivity to sars-cov-2 on lateral flowbased antibody testing (biohit health care ltd., cheshire, united kingdom), suggesting a contributory role for fecal-oral transmission of the disease, as previously proposed by others. 1 the baseline study was followed by a second round of testing 4 weeks later to assess the incidence of infection. tests were stored and managed as per manufacturer instructions, and tests were performed in the field by a trained medical doctor. here, we present a cluster of incident cases of sars-cov-2 involving a woman who lived alone (house a), and a five-member family (house b) who were seronegative during the first survey. these families were not related to each other but shared a latrine located between both houses. two weeks after our baseline serosurvey, a 22-year-old grandson of the old woman moved into atahualpa from guayaquil (a heavily infected urban center), staying at her house and using the shared latrine. none of the families refers any other direct interaction or social gathering. during the second serosurvey, the woman in house a seroconverted to positive, and her visiting grandson also resulted seropositive. likewise, four of the five family members of house b became seropositive (figure 1 ). there were no other incident cases in the entire block, where only one person in a distant house had tested positive at baseline, and several other inhabitants of other houses remained seronegative (figure 2, left) . we hypothesize that the visitor was likely infected when he arrived, infected his grandmother by living together, and then the infection was spread to the neighbor's house by fecal contamination of one of the members while using the latrine, which was poorly maintained (figure 2 , right). certainly, we cannot rule out other interpersonal contacts, but-as previously mentioned-neither of the families refer interaction. similarly, we cannot conclude on whether the individual who did not seroconvert was less exposed or had any sort of reduced individual predisposition to infection. this cluster of new infections provides circumstantial evidence that latrines (ergo, fecal contamination) may act as a source of infection. poor hygienic conditions are frequent in rural regions of developing countries and may contribute to the spread of sars-cov-2. 2 e-mail: aldocosva_01@hotmail.com. héctor h covid-19 in latin america: novel transmission dynamics for a global pandemic indigenous communities in brazil fear pandemic's impact ca) of the block where the described cases occurred. yellow arrow points to the shared latrine. the blue cross refers to the visitor (index case) and the green dot in the same house to his seroconverted grandmother. behind house a, house b shows four seroconverted family members (green dots) and the one that remained seronegative (white dot). the other white dots across other houses of the block refer to individuals who were seronegative at the first and the second surveys, and the red dot to a single individual who was seropositive at the first survey this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-340021-pj6fywwc authors: norooznezhad, amir hossein; najafi, farid; riahi, parisa; moradinazar, mehdi; shakiba, ebrahim; mostafaei, shayan title: primary symptoms, comorbidities, and outcomes of 431 hospitalized patients with confirmative rt-pcr results for covid-19 date: 2020-06-24 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0512 sha: doc_id: 340021 cord_uid: pj6fywwc this study aimed to evaluate the primary symptoms, comorbidities, and outcomes of inpatients with confirmed reverse transcription–pcr (rt-pcr) for sars-cov-2 infection among 2077 suspected/diagnosed cases of covid-19. based on the results of least absolute shrinkage and selection operator (lasso) logistic regression, age, and suggestive chest x-ray (cxr) findings for sars-cov-2 infection, cardiovascular diseases, diabetes mellitus, chronic lung diseases, and intensive care units admission had significant associations with positive rt-pcr results for covid-19 infection. also, the highest area under the curve (auc) was related to cough (auc = 0.53, 95% ci: 0.51–0.56), dyspnea (auc = 0.52, 95% ci: 0.50–0.54), and abnormal cxr (auc = 0.52, 95% ci: 0.50–0.54), as significant predictors. this study showed that some symptoms including cough and dyspnea, as well as abnormal cxr, could be proper predictors of positive rt-pcr result for sars-cov-2 infection. it seems that patients with underlying disease(s), such as cardiovascular diseases, diabetes mellitus, and chronic lung diseases, had a higher probability to have positive rt-pcr for sars-cov-2 infection than those with no underlying disease(s). in late 2019, a new coronavirus outbreak started in wuhan (hubei province, china), which rapidly turned into a pandemic emergency according to the who, and, now (10:00 cest, june 12, 2020), more than 7.4 million individuals have emerged as infected cases. 1 the main feature of the disease is pneumonia, 2 and most patients have primary symptoms such as fever, cough, dyspnea, sore throat, myalgia, gastrointestinal penetrations, and rhinorrhea. 3, 4 this study aimed to evaluate the possible predictive value of the most important symptoms and underlying diseases in patients diagnosed with covid-19 using reverse transcription-pcr (rt-pcr) in three agegroups of children and adolescents, adults, and elderly. this analytical cross-sectional study was performed in farabi and imam-reza hospitals as two designated centers for covid-19 patients hospitalization by the ministry of health in kermanshah (a western province), iran. the database used herein was obtained from the vice-chancellor of health in the kermanshah university of medical sciences gathered from 2077 suspected/diagnosed cases of covid-19 from february 20 to april 9, 2020. april 9 is the first day an official statement was made on the first confirmed covid-19 case in iran by the who. 5 the inclusion criteria were defined as 1) individuals with symptom(s) in favor of covid-19 and 2) inpatients with confirmative rt-pcr result for sars-cov-2 infection. also, any lack of information in any patient led to their exclusion from the study. primary data on the hospitalized patients such as biodemographic information, presenting symptoms, underlying disease(s), and abnormal findings in chest x-ray (cxr) were investigated based on the lasso logistic regression by "glmnet" r package. moreover, roc curve analysis was performed for the predicted diagnostic value of the already mentioned variables for prediction of a positive rt-pcr result for sars-cov-2 infection. the data were supported and validated by the vicechancellor of health in the kermanshah university of medical science, and permission was granted to use the mentioned information only for research purposes, not as any official statics for the province. this study was approved by the medical ethics committee of kermanshah university of medical sciences. all the data were encrypted and de-identified when received from the mentioned source. also, all authors declare their adherence to the declaration of helsinki in 1964 and its further revisions. among 2077 suspected/diagnosed cases and considering both inclusion and exclusion criteria, 431 individuals with a definite diagnosis for covid-19 infection based on positive rt-pcr and 1,646 cases with negative rt-pcr results were enrolled in this study. of those 431 inpatients, children and adolescents (aged £ 18 years), adults (aged between 18 and 65 years), and elderlies (aged ³ 65) had the prevalence of 3.5% (15/431), 77.5% (334/431), and 19% (82/431), respectively, among which 29 (6.7%) were admitted to intensive care units (icus). the total fatality rate in rt-pcr positive group was 9% (39/431), in which children and adolescent, adult, and elderly groups have 0% (0/15), 4.8% (16/334), and 28.1% (23/82) proportions, respectively. there was a significant association between variables, including age, abnormal cxr findings, cardiovascular diseases, diabetes mellitus, chronic lung diseases, and icu admission, with confirmative rt-pcr results for sars-cov-2 infection. however, cough, dyspnea, elevated body temperature (both ³ 37.7°c and ³ 38.0°c), and rhinorrhea were among the most prevalent nonsignificant associated features with the confirmed result of rt-pcr for sars-cov-2 infection (p-value> 0.05). also, significantly associated features in the evaluated patients by their age-groups are provided in table 1 . the highest area under the curve (auc) was related to cough (auc = 0.53, 95% ci 0.51-0.56), dyspnea (auc = 0.52, 95% ci 0.50-0.54), and abnormal cxr (auc = 0.52, 95% ci 0.50-0.54), as the most significant predictors of confirmed rt-pcr for sars-cov-2 infection. in adults and elderly patients, the highest auc was related to abnormal cxr (table 2) . in figure 1a , the hierarchical dendrogram, age, and cardiovascular diseases have the strongest interactions compared with the other variables for the prediction of death among positive rt-pcr patients. the color of the connecting line, blue, is indicative of the high degree of redundancy (or most relative). considering the attributes connecting with the green line, these are of a less degree of redundancy in terms of interaction (or middle relative). gold lines are representing the independent (or less relative) attributes, as their interaction coefficient is not significant. also, the circle graph ( figure 1b ) was used as the indicator of interaction network for the death predictors based on the information gain (ig) of each attribute, also considered as the main effect value. as it is shown ( figure 1b) , age (ig = 13.7%), cardiovascular disease (ig = 2.4%), and icu admission (ig = 2.2%) have the most important main effects, and they have interaction effects with abnormal cxr, elevated body temperature, myalgia, sore throat, diabetes mellitus, and malignancy for the prediction of death among positive rt-pcr patients. the results showed that the fatality rate was higher in elderly than other age-groups. also, underlying diseases in the attributes connecting with green line, these are of a less degree of redundancy in terms of interaction (or middle relative). gold lines are representing the independent (or less relative) attributes, as their interaction coefficients are not significant. (b) circle graph of mdr algorithm. circle graph indicator of information gain as the main effects of each variable and the interaction effects between them to prediction of death among positive rt-pcr patients. this figure showed age, cardiovascular disease, and intensive care unit (icu) admission have stronger main effect on the risk of death. also, the interaction among age (older than 65 years), cardiovascular disease, diabetes mellitus, myalgia, malignancy, sore throat, and abnormal chest x-ray (cxr) had stronger effects on death among positive rt-pcr patients. including cardiovascular diseases, diabetic mellitus, and chronic lung diseases were significantly associated with a positive rt-pcr result for sars-cov-2 infection. changes in the importance of variables in each age category showed the importance of each one as clinically important parameters in suspected covid-19 cases. also, table 2 shows that cough, dyspnea, and abnormal cxr seem to be proper tools for the prediction of confirmed rt-pcr in cases suspected of covid-19. we have chosen only rt-pcrpositive cases because at the beginning of the outbreak, details on radiologic findings were not as well known as those at the end of the study, which might lead to falsepositive or negative result in our evaluated patients. thus, including only rt-pcr-positive patients might be one of the reasons why the outcome ratios such as fatality rate or icu admission could not be relied on as full populationbased statics. according to tables 1 and 2, some variables seem to be associated with a positive rt-pcr result in symptomatic cases. on the other hand, it has been shown that rt-pcr is a confirmative diagnostic method for covid-19, with a sensitivity of 71%. considering the study by fang et al., 6 who evaluated symptomatic patients, it has been shown that 36/51 of their cases had the primary confirmed rt-pcr results for sars-cov-2 infection on the first test. they performed second and third tests on days 1-2 and 2-5, which were confirmative for 12/51 (cumulative 48/51) and 2/51 (cumulative 50/51) of their patients, respectively. for the remaining patients, rt-pcr for sars-cov-2 confirmed covid-19 infection on day 7 (cumulative 51/ 51), 6 whereas the cumulative ratio for abnormal chest computed tomography (ct) scan at the first day was 50/ 51 as they have reported. 6 considering their result as well as having a glance at tables 1 and 2, when rt-pcr is negative for patients with already mentioned variables (bolded ones), two pathways could be lead: 1) repeating rt-pcr on the following days and 2) requesting a chest ct scan with a sensitivity of 97% for the diagnosis of covid-19. 7 however, because some patients might have at least an underlying disease, affecting lungs (such as cardiovascular disease and chronic lung disease), we suggest not to miss following up these patients by further rt-pcr tests. 8 regarding the symptoms, a systematic review and metaanalysis on patients diagnosed with covid-19 has shown that fever (88.7%), cough (57.6%), and dyspnea (45.6%) were the most prevalent symptoms in the patients (regardless of age); however, no subgroup for the method of diagnosis has been investigated. from all the symptoms evaluated in the mentioned review, authors have just shown differences of fever and cough among adults and children (fever: 92.8% versus 43.9%; cough: 63.4% versus 22% respectively). however, no other subgroup analysis has been performed neither for children nor for the elderlies. 3 another study evaluating the findings of two groups of young (n = 38) and elderly (n = 18) cases with confirmed covid-19 infection showed that there was no difference between the prevalence of fever, cough, dyspnea, fatigue, rhinorrhea, or vomiting between the two groups. 4 thus, it seems that this study has added some new findings regarding a large number of confirmed covid-19 patients in three age-groups by their symptoms. altogether, we investigated the importance of primary findings in the patients who had a definite laboratory diagnosis for covid-19 (rt-pcr) in three age categories including children and adolescents, adults, and the elderlies. the presence of some presenting symptoms and/or underlying diseases seems to be associated with a confirmed rt-pcr result for sars-cov-2 infection depending on the age category. we are certainly following the performed progress in the diagnosis/outcomes of patients with covid-19 after the end of period for the current study to assess the quality of experiences earned during the time. coronavirus disease 2019 (covid-19) inappropriate antibiotic consumption as a possible cause of inflammatory storm and septic shock in patients diagnosed with coronavirus-19 disease (covid-19) clinical, laboratory and imaging features of covid-19: a systematic review and meta-analysis clinical features of covid-19 in elderly patients: a comparison with young and middle-aged patients coronavirus disease 2019 (covid-19) sensitivity of chest ct for covid-19: comparison to rt-pcr correlation of chest ct and rt-pcr testing in coronavirus disease 2019 (covid-19) in china: a report of 1014 cases chest computed tomography manifestation of coronavirus disease 2019 (covid-19) in patients with cardiothoracic conditions key: cord-255062-7ozdmb09 authors: ogoina, dimie title: improving appropriate use of medical masks for covid-19 prevention: the role of face mask containers date: 2020-08-04 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0886 sha: doc_id: 255062 cord_uid: 7ozdmb09 use of medical masks is a key strategy for covid-19 prevention among healthcare workers. unfortunately, there are global shortages of this essential commodity, and many have resulted in inappropriate usage to conserve supply. this article highlights the likely benefits of face mask containers in promoting safe, appropriate, and extended use of medical masks by healthcare workers in settings where a sustainable supply of medical masks may be limited. public wearing of face masks is now one of the key strategies for containing the current covid-19 pandemic across the globe. 1, 2 the who has also recommended targeted continuous use of medical masks by healthcare workers working in clinical areas in health facilities in geographical areas with community transmission of covid-19. 2 according to the who, face masks are used appropriately when they always cover the mouth and nose, when the front and inside of the face mask is not touched, and when hand hygiene is performed before wearing, and after touching and removal of the face mask. 2 during targeted continuous mask use, healthcare workers are required to wear medical masks throughout their entire shift and advised to replace their medical masks when wet, damp, visibly soiled, damaged, and if the health worker/ caregiver removes the mask (e.g., for eating or drinking or caring for a patient who requires droplet/contact precautions for other reasons). 2 face mask use has been shown to be associated with a large reduction in the risk of covid-19 infection. 3 unfortunately, it is not comfortable to wear face masks for prolonged periods. 2, 4 as an infectious disease physician working in a covid-19 isolation facility in nigeria, i have observed that most healthcare workers do not have adequate supplies of medical masks to replace them each time there is need to temporarily remove their masks to undertake activities such as eating or drinking or when alone in their offices or cars. many have therefore opted to wear masks on their chin and neck or keep potentially contaminated face masks on desks, or inside pockets, or bags in close contact with other personal belongings. 5 in most cases, the masks are handled carelessly and squeezed repeatedly during removal and reuse. the consequence of this inappropriate use of face masks is selfand environmental contamination and increased risk of transmission of covid-19. although there is a growing literature on strategies to decontaminate and reuse single-use medical masks, 4,6 the who currently does not recommend reuse of single-use medical masks. however, if medical masks are to be worn continuously by healthcare workers for up to 8 hours or more every working day, then there should be provision to temporarily and safely store them for extended use during the day, especially when they are not visibly soiled, wet, damp, or damaged. the use of face mask containers could help to promote appropriate and safe storage of face masks and facilitate extended use when medical masks are not due for replacement. we designed a plastic box and a leather pouch as face mask containers (figure 1 ) to store masks temporarily when not in use. within the containers, the masks are secured by their straps and held flat in their natural positions without the risk of squeezing and self-and environmental contamination. this way they can easily be picked up by the straps and worn safely without touching the front or inside of the mask. we also created vents on both sides of the face mask containers to improve ventilation in the containers when closed. in the wake of global supply shortages, 7 appropriately designed face mask containers could be useful in promoting safe extended use of medical masks, especially in resourcelimited healthcare settings. face mask containers could conserve limited supplies of medical masks for sustainable use by frontline healthcare professionals. compared with the chin, neck, pocket, or desk, a face mask container is probably a safer environment to store a medical mask when there is need to remove it temporarily to perform activities such as eating or drinking, or when there is need to temporarily expose the mouth and nose to relieve the discomfort associated with the prolonged use of face masks. face mask containers should however not be used to store medical masks that are due for replacement, especially when they are wet, damp, visibly soiled, or damaged. unfortunately, an unintended risk of these containers is the likelihood of prolonged storage and repeated reuse of masks that should have been discarded or washed before reuse. with the growing call for universal masking as a key costeffective strategy to combat the covid-19 pandemic, it is my view that the benefits of face mask containers in promoting appropriate use of masks and enabling extended and safe use far outweigh the risks. further studies on the benefits and risks of face mask containers would be useful to confirm these assertions. universal masking to prevent sars-cov-2 transmission-the time is now advice on the use of masks in the community, during home care and in healthcare settings in the context of the novel coronavirus (covid-19) outbreak covid-19 systematic urgent review group effort (surge) study authors, 2020. physical distancing, face masks, and eye protection to prevent person-to-person transmission of sars-cov-2 and covid-19: a systematic review and meta-analysis face masks for the public during the covid-19 crisis covid-19: the need for rational use of face masks in nigeria decontamination of surgical face masks and n95 respirators by dry heat pasteurization for one hour at 70°c shortage of personal protective equipment endangering health workers worldwide leather pouch (a) and plastic box (b) designed as face mask containers to safely store face masks to avoid self-and environmental contamination and promote extended use of face masks in the face of scarcity acknowledgments: i acknowledge the support of opaminola okrinya, key: cord-305073-8301zqj9 authors: alhakeem, ayat; khan, muhammad mohsin; al soub, hussam; yousaf, zohaib title: case report: covid-19–associated bilateral spontaneous pneumothorax—a literature review date: 2020-07-14 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0680 sha: doc_id: 305073 cord_uid: 8301zqj9 covid-19 is a pandemic caused by sars-cov-2, primarily affecting the respiratory tract. pulmonary complications of covid-19 may include acute respiratory distress syndrome and pulmonary embolism. pneumothorax has been recently reported in association with covid-19. we report a case of covid-19 pneumonia with bilateral spontaneous pneumothorax with no known underlying lung disease or risk factors. coronaviridae is a family of rna viruses that has captured the attention of epidemiologists, microbiologists, clinicians, and policymakers worldwide. sars-cov-2 has caused a massive impact on the global economy and everyday life, and an unprecedented burden on the healthcare system. this infection has a broad spectrum of presentations that can range from asymptomatic disease to fatal acute respiratory distress syndrome. although most cases are mild, up to 5% of the cases can develop severe illness leading to multi-organ damage. 1 spontaneous pneumothorax is a rare complication of covid-19. most of the reported cases of pneumothorax associated with covid-19 lack traditional risk factors or underlying predisposing lung disease. [2] [3] [4] [5] [6] [7] [8] [9] [10] [11] [12] [13] [14] [15] [16] to date, one case of covid-19-associated spontaneous bilateral pneumothorax in covid-19 has been described to the best of our knowledge. 12 a 49-year-old man, not known to have any chronic medical condition, presented with a fever and dry cough for 7 days. he was a nonsmoker, with a height of 161 cm, weight of 61 kg, and a body mass index of 23.5. he had no known exposure to animals, birds, sick contacts, or toxic fumes. initial physical examination, including chest examination, was unremarkable. chest x-ray showed prominent broncho-vascular markings bilaterally without evidence of consolidation. covid-19 pcr from nasopharyngeal swab was positive. the patient was labeled as mild covid-19 pneumonia and was observed in a quarantine facility. five days from his initial presentation, the patient developed breathing difficulty with desaturation to 85% on room air. chest examination revealed bilateral crackles. a chest x-ray showed bilateral lung infiltrates ( figure 1a ). his laboratory results showed deranged liver enzymes (alkaline phosphatase 197 u/l, alanine aminotransferase 132 u/l, and aspartate aminotransferase 81 u/l), lymphopenia (0.8 × 10 3 /μl), high d-dimers (1.47 mg/l), and raised inflammatory markers (c-reactive protein 133.1 mg/l and ferritin 8,382.0 μg/l). the patient required 15 l of oxygen via a non-rebreather face mask and was cared for in the intensive care unit as a case of severe covid-19 pneumonia. he received covid-19 pneumonia treatment with azithromycin, hydroxychloroquine, ceftriaxone, and lopinavir-ritonavir based on local management guidelines. during his intensive care unit stay, the patient was kept in an awake prone position and received tocilizumab and convalescent plasma. at no point during his stay he required the use of continuous positive airway pressure or bi-level positive airway pressure. his oxygen requirements decreased over the next 5 days, and he was transferred to the medical ward. on day 12 of the initial presentation, he developed sudden shortness of breath and chest pain, with desaturation. examination showed decreased air entry on the right lung with a left-deviated trachea. urgent chest x-ray showed a significant right-sided pneumothorax ( figure 1b) . a right-sided chest tube was inserted and connected to an underwater seal, and his symptoms improved. post-procedure chest x-ray showed a significant reduction in the right pneumothorax with adequate right lung expansion. on day 17, the patient developed severe acute shortness of breath. examination showed decreased air entry in the bilateral lung with deviation of the trachea to the right. urgent chest x-ray showed a significant left-sided pneumothorax, and another chest tube was inserted on the left side ( figure 1c ). highresolution computed tomography (ct) chest showed multiple bilateral bullae in the lungs complicated by the pneumothorax because of rupture ( figure 1d ). the alpha-1 antitrypsin level was normal (187.9 mg/dl), and tuberculosis workup, including sputum culture and acid-fast bacilli smear, was negative. the rightsided chest tube was removed after 6 days of insertion and nearcomplete resolution of the pneumothorax on the ipsilateral side on chest x-ray. the patient continues to receive hospital care at present with near-complete expansion of the left lung. an outbreak of pneumonia with an indeterminate source surfaced in wuhan, china, in december 2019, later known as covid-19. covid-19 has a lower mortality rate in comparison to the other coronaviridae, such as sars-cov-1 and middle east respiratory syndrome, yet higher infectivity. 17 known risk factors for the development of primary spontaneous pneumothorax include male gender, tall stature, thin body habitus, and age-group 10-30 years. 18 secondary causes include infections, smoking, chronic obstructive pulmonary disease, alpha-1 antitrypsin deficiency, and trauma. 19 severe alveolar and airway inflammatory damage from the release of cytokines in covid-19 can lead to weakening of the bronchial walls. edema, vascular congestion, and microthrombi may contribute to the rupture of preexisting bullae. 20 rupture of these bullae can lead to pneumothorax. bullous lung disease is characterized by the development of bullae in otherwise normal lung parenchyma. 21 risk factors for the development of bullae include smoking history, pulmonary sarcoidosis, alpha-1 antitrypsin deficiency, alpha-1 anti-chymotrypsin deficiency, marfan's syndrome, ehlers-danlos syndrome, inhaled fiberglass exposure, and marijuana smoking. 20, 22 the underlying pathophysiology for bullae formation involves inflammatory damage to the bronchiole, leading to trapping of air. interaction of mechanical forces on the weakened tissue may lead to bullae formation. 21, 23 although our patient was a male, he was never smoker, with unrevealing screening for risk factors of bullae formation and pneumothorax. the bullous changes could represent an undiagnosed underlying pulmonary disease, which became apparent after the inflammatory changes and excessive mechanical forces introduced by the sars-cov-2 infection, leading to a unilateral spontaneous pneumothorax, followed by bilateral pneumothorax. review of the literature shows 18 case reports describing covid-19 patients with spontaneous pneumothorax. eight of these patients were managed conservatively, whereas 10 required chest tube insertion. two of the patients required thoracoscopy and bleb resection. one patient developed tension pneumothorax and required emergency needle decompression. only four cases were smokers. three cases were on invasive mechanical ventilation. three had underlying lung disease. one case had bilateral pneumothorax, whereas the rest had unilateral involvement. twelve patients had a favorable clinical course, whereas six patients passed away, resulting in a mortality rate of 33% based on the available literature (table 1 ). [2] [3] [4] [5] [6] [7] [8] [9] [10] [11] [12] [13] [14] [15] [16] our case is the second reported bilateral spontaneous pneumothorax in the literature to the best of our knowledge. acute deterioration in covid-19 patients may be due to primary disease or pulmonary embolism; however, pneumothorax is another important differential. pneumothorax is infrequently associated with covid-19 pneumonia. covid-19-related spontaneous pneumothorax in an otherwise healthy individual may be an underdiagnosed entity. this association could be secondary to underlying undiagnosed characteristics of and important lessons from the coronavirus disease 2019 (covid-19) outbreak in china diagnosis and treatment of severe covid-19 complicated with spontaneous pneumothorax: a case report sars-cov-2 infection associated with spontaneous pneumothorax spontaneous pneumothorax and subcutaneous emphysema in covid-19 patient: case report a case of spontaneous pneumothorax in covid-19 pneumonia covid-19 with spontaneous pneumothorax, pneumomediastinum and subcutaneous emphysema covid-19, chronic obstructive pulmonary disease and pneumothorax: a frightening triad tension pneumothorax in a patient with covid-19 mediastinal emphysema, giant bulla, and pneumothorax developed during the course of covid-19 pneumonia spontaneous pneumomediastinum: a probable unusual complication of coronavirus disease 2019 (covid-19) pneumonia spontaneous pneumomediastinum in a patient with coronavirus disease 2019 pneumonia and the possible underlying mechanism pneumomediastinum and spontaneous pneumothorax as an extrapulmonary complication of covid-19 disease management of persistent pneumothorax with thoracoscopy and blebs resection in covid-19 patients spontaneous pneumomediastinum occurring in the sars-cov-2 infection spontaneous pneumomediastinum in covid-19 sars-cov-2 pneumonia with subcutaneous emphysema, mediastinal emphysema, and pneumothorax: a case report sars-cov-2 shedding and infectivity epidemiology of pneumothorax in england spontaneous pneumothorax bullous lung disease fishman's pulmonary diseases and disorders large lung bullae in marijuana smokers bullous disease of the lung covid-19-associated pneumothorax key: cord-275243-knr3mat0 authors: larsen, kevin; coolen-allou, nathalie; masse, laurie; angelino, alexandre; allyn, jérôme; bruneau, lea; maillot, adrien; lagrange-xelot, marie; vitry, thierry; andré, michel; travers, jean yves; foch, emilie; allou, nicolas title: detection of pulmonary embolism in returning travelers with hypoxemic pneumonia due to covid-19 in reunion island date: 2020-07-01 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0597 sha: doc_id: 275243 cord_uid: knr3mat0 the aim of this study was to evaluate the occurrence of pulmonary embolism in returning travelers with hypoxemic pneumonia due to covid-19. all returning travelers to reunion island with hypoxemic pneumonia due to covid-19 underwent computed tomography pulmonary angiography (ctpa) and were included in the cohort. thirty-five patients were returning travelers with hypoxemic pneumonia due to covid-19 and had recently returned from one of the countries most affected by the covid-19 outbreak (mainly from france and comoros archipelago). five patients (14.3%) were found to have pulmonary embolism and two (5.9%) were incidentally found to have deep vein thrombosis on ctpa. patients with pulmonary embolism or deep vein thrombosis had higher d-dimer levels than those without pulmonary embolism or deep vein thrombosis (p = 0.04). returning travelers with hypoxemic pneumonia due to covid-19 should be systematically screened for pulmonary embolism. an outbreak of covid-19 that started in china in december 2019 began to spread globally, 1 particularly in europe, in january 2020. reunion island (845,000 inhabitants) is a french overseas department located in the indian ocean. it is at a distance of 10,000 km from paris and is connected to metropolitan france by several daily flights (11 hours). between march 11, 2020 and april 20, 2020, 388 cases of covid-19 were reported in reunion island. 2 the covid-19 pandemic is overwhelmingly associated with international travel. 3 long-haul flight travel is a known risk factor of pulmonary embolism. 4 the aim of this study was to evaluate the occurrence of pulmonary embolism in returning travelers with hypoxemic pneumonia due to covid-19. the present observational study was approved by the french ethics committee of infectious disease and tropical medicine and was declared to the commission nationale de l'informatique et des libertés (french data protection agency or cnil, n°2206739). this observational study was conducted between march 11, 2020 (first case of covid-19 in reunion island) and april 20, 2020 at félix guyon university hospital, the only hospital authorized to handle patients with covid-19 in reunion island. in accordance with our protocol, all patients with hypoxemic pneumonia due to covid-19 confirmed by polymerase chain reaction underwent systematic chest computed tomography angiography pulmonary (ctpa). among patients with hypoxemic pneumonia, all returning travelers were included in the cohort study. hypoxemic pneumonia was defined as pneumonia requiring oxygen supplementation to achieve oxyhemoglobin saturation > 94%. all ctpa images were analyzed by two radiologists (t. v. and j. y. t.) and at least two pulmonologists (l. m., e. f., m. a., and n. c. a.) blinded to clinical information. in cases of ctpa contraindications such as contrast allergy or renal failure, patients underwent lung ventilation/perfusion scintigraphy instead. computed tomography angiography pulmonary examinations were performed in multi-detector computed tomography scanners (ge revolution gsi, general electric, milwaukee, wi) by using a standard ctpa protocol. scan parameters were as follows: tube voltage of 100 kv, tube current of 100-300 mas, collimation of 0.625 mm, pitch of 1.375-1.0, table speed of 55 mm/second, and gantry rotation time of 0.5 seconds. images were reconstructed with a thickness of 1 mm and an increment of 1.25 mm. results were expressed as total numbers (percentages) for categorical variables and as medians (25th-75th percentiles) for continuous variables. categorical variables were compared using the chi-square test or the fisher's exact test, as appropriate. a p-value < 0.05 was considered significant. analyses were performed using sas statistical software (8.2, cary, nc). over the study period, 165 of 388 patients (42.5%) who had tested positive for covid-19 were admitted to félix guyon university hospital. of these 165 patients, 44 (26.7%) patients had hypoxemic pneumonia. thromboembolic events occurred in seven of the 35 (20%) returning travelers versus one of the nine (11%) non-returning travelers (p = 0.35). the 35 returning travelers had recently returned from one of the countries most affected by the covid-19 outbreak: 24 from metropolitan france, eight from comoros archipelago, five from spain, two from italy, two from the united states, and one from the united kingdom (some patients had visited several of these countries). patient characteristics at study inclusion are shown in table 1 . the median duration between onset of symptoms and diagnosis of covid pneumonia was 6 (4-9) days. the median number of days between arrival to reunion island and the first day of symptoms was 0 (-3-2) day. of the 35 enrolled patients, 33 (94.3%) underwent one ctpa, one (2.9%) underwent two ctpas, and one (2.9%) underwent one lung ventilation/perfusion scintigraphy. chest examinations were performed on day 7 (5-12) after the onset of symptoms, and the median value of oxygen therapy was 4 (2-13) l/minutes. four of the 35 patients (11.4%) received invasive mechanical ventilation. five patients were found to have pulmonary embolism (14.3%). among the five patients, one had an associated aortic arch thrombosis (figure 1 ). two other patients were (incidentally) found to have extremity deep vein thrombosis (extensive catheter-related jugular thrombosis) on ctpa, for a total of minimum of 20% of the enrolled patients. after univariate analysis, the factors associated with pulmonary embolism and extremity deep vein thrombosis were higher levels of d-dimer (p = 0.04) and absence of low molecular weight heparin prophylaxis (p = 0.02) ( table 1 ). there were no deaths at follow-up (minimum 40 days, and only two remained hospitalized and had been weaned from oxygen therapy). to our knowledge, this is the only study that has consecutively evaluated the occurrence of pulmonary embolism in returning travelers with hypoxemic pneumonia due to covid-19. in our study, the incidence of thromboembolic complications was high despite the fact that the patients had a low severity score with a quick sepsis-related organ failure assessment score of 1 (0-1) (97.1% had a severity score < 2). in the published literature, the incidence of thrombotic complications like pulmonary embolism or deep vein thrombosis in patients with covid-19 pneumonia is highly variable, ranging from 1% to 20%. [5] [6] [7] the incidence of thrombotic complications in our study population may seem relatively high despite that severity score was very low. the incidence that was reported in the retrospective study by chen et al. 6 was lower (< 1%)-though it should be noted that only 25 of the 1,008 patients (2.5%) examined in that study had undergone ctpa. in the other studies published on the topic, the incidence of thrombotic complications was high because only patients hospitalized in intensive care were evaluated. 7, 8 deep vein thrombosis and pulmonary embolism are severe complications of covid-19. it is important that they can be diagnosed early because delay in treatment can be lifethreatening in the short and long term. in our study, patients without pulmonary embolism had a median d-dimer level < 1 μg/ml. in the study by chen et al., 6 the median d-dimer level of patients without pulmonary embolism was 2.44 μg/ml. in the study by cui et al., 7 a level of d-dimer of 1.5 μg/ml was a good cutoff to predict venous thromboembolism (sensitivity of 85.0% and specificity of 88.5%). last, retrospective studies suggest that preventive anticoagulation is associated with decreased mortality in patients with severe covid-19 infection, particularly in those with high d-dimer levels. 8 enhanced preventive anticoagulation should be used in all patients with hypoxemic pneumonia due to covid-19. 9 the main limitations of our study are that the number of evaluated patients and the number of events were relatively small. moreover, it was not possible to perform a control group because the vast majority of covid-19 patients came from europe by long-haul flight (i.e., > 6,000 km). reunion island is located at a distance of 10,000 km from europe. air flight is a well-known risk factor for the occurrence of pulmonary embolism, and this risk increases with the duration of the flight. 10 in conclusion, the incidence of pulmonary embolism and deep vein thrombosis in our study population was relatively high at 20%. returning travelers with hypoxemic pneumonia due to covid-19 should be systematically screened for pulmonary embolism or deep vein thrombosis regardless of the level of d-dimers. first cases of coronavirus disease 2019 (covid-19) in france: surveillance, investigations and control measures suivi de l'épidémie de covid-19 en france travelers give wings to novel coronavirus (2019-ncov) incidence, clinical characteristics, and long-term prognosis of travelassociated pulmonary embolism pulmonary embolism in returning traveler with covid-19 pneumonia findings of acute pulmonary embolism in covid-19 patients prevalence of venous thromboembolism in patients with severe novel coronavirus pneumonia difference of coagulation features between severe pneumonia induced by sars-cov2 and non-sars-cov2 propositions du gfht/gihp pour le traitement anticoagulant pour la prévention du risque thrombotique chez un patient hospitalisé avec covid-19 severe pulmonary embolism associated with air travel acknowledgment: publication charges for this article were waived due to the ongoing pandemic of covid-19.financial support: this work was internally funded.disclosure: the present observational study was approved by the ethics committee of infectious disease and tropical medicine (cer-mit) and was declared to the commission nationale de l'informatique et des libertés (french data protection agency or cnil mr004, n°2 206739). the need for informed consent was waived, as the study was non-interventional and followed our usual protocol. the dataset used in the current study are available from the corresponding author on reasonable request. this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-270495-2u072mtp authors: lokida, dewi; lukman, nurhayati; salim, gustiani; butar-butar, deni pepy; kosasih, herman; wulan, wahyu nawang; naysilla, adhella menur; djajady, yuanita; sari, rizki amalia; arlinda, dona; lau, chuen-yen; karyana, muhammad title: diagnosis of covid-19 in a dengue-endemic area date: 2020-08-05 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0676 sha: doc_id: 270495 cord_uid: 2u072mtp emergence of sars-cov-2 in dengue virus (denv)–endemic areas complicates the diagnosis of both infections. covid-19 cases may be misdiagnosed as dengue, particularly when relying on denv igm, which can remain positive months after infection. to estimate the extent of this problem, we evaluated sera from 42 confirmed covid-19 patients for evidence of denv infection. no cases of sars-cov-2 and denv coinfection were identified. however, recent denv infection, indicated by the presence of denv igm and/or high level of igg antibodies, was found in seven patients. dengue virus igm and/or high igg titer should not exclude covid-19. sars-cov-2 reverse transcription polymerase chain reaction (rt-pcr) testing is appropriate when dengue nonstructural protein 1 (ns1) or rt-pcr is negative. given the possibility of coinfection, testing for both denv and sars-cov-2 is merited in the setting of the current pandemic. emergence of sars-cov-2 in dengue virus (denv)endemic areas has raised concern regarding coinfection with the two viruses. 1, 2 difficulty in distinguishing dengue and covid-19, particularly during the acute stage, can engender inaccurate diagnoses. during the covid-19 pandemic, patients who screen positive on the sars-cov-2 questionnaire (supplemental table 1 ) are tested for sars-cov-2 by rt-pcr. when confirmed, no further investigation for other etiologies is commonly performed. when sars-cov-2 is negative and clinical indication is present (at least fever and thrombocytopenia), denv ns1 antigen and/or igm/igg antibody testing may be performed. clinicians from singapore reported two covid-19 cases that were misdiagnosed as dengue among patients who presented with clinical manifestations and hematology profiles, suggesting dengue infection and false-positive denv igm antibody using a rapid diagnostic test (rdt). 3 this may have occurred because of persistence of denv igm from a prior denv infection. indonesia has experienced a surge in covid-19 cases against the backdrop of dengue endemicity. because the prevalence of denv igg antibodies in singapore is significantly lower than that in indonesia, 4,5 we expect indonesia to face greater challenges with diagnosing sars-cov-2, typically performed by rt-pcr, while denv is co-circulating. to estimate the extent of this problem, we evaluated sera from confirmed covid-19 patients for evidence of denv infection. covid-19 cases were defined as inpatients who met the covid-19 criteria based on a predetermined combination of symptoms, laboratory testing, imaging, and risk exposure at tangerang district hospital, indonesia (see supplemental table 1 ), and had a positive nasopharyngeal or oropharyngeal real-time rt-pcr for sars-cov-2. blood and sera were collected from all suspected covid-19 patients for clinical and research testing. for this study, admission sera for all cases were evaluated for denv ns1 using rdt (panbio ® dengue early rapid, abbot, brisbane, australia) and elisa (dengue ns1 antigen dxselect™, focus diagnostics, cypress, ca) assays, and for denv igm and igg using rdt (panbio ® dengue duo cassette, abbot, sinnamon park, australia) and elisa (focus diagnostics) assays. if available, follow-up sera from 8.5 ± 2.1 days later were evaluated for denv igm and igg by the same rdt and elisa methods. admission sera from cases with positive denv igm were evaluated using rt-pcr. results were not returned in real time for patient care purposes. clinical and laboratory information on admission was obtained by chart review. descriptive statistics were performed to characterize the presentation of covid-19 among these cases and to assess denv infection status. this research was approved by the tangerang district hospital ethics committee. admission sera were available for 42 covid-19 cases. follow-up sera were available for 32 of these patients. the mean age was 43.5 (sd ± 14.1) years, with a male predominance (66.7%). time from the onset of illness to serum collection was 8.1 days (sd ± 4.2) days. the most common signs and symptoms were fever (90.5%); cough (83.3%); fatigue, dyspnea, and dysgeusia (38.1% each); sore throat (33.3%); headache (19%); and anosmia and diarrhea (11.9% each). lymphopenia (< 1,000/mm 3 ), leukopenia (< 4,000/ mm 3 ), and thrombocytopenia (< 150,000/mm 3 ) during admission were found in 26.2%, 11.9%, and 4.8%, respectively. a comparison of signs and symptoms for our patients with covid-19 and dengue patients from a recent fever study 5 is shown in supplemental table 2 . none of the 42 subjects was positive for dengue ns1 or showed seroconversion or increasing denv igm and igg index values, suggesting no acute denv infection among these covid-19 cases. however, both denv igm and high igg titer, as indicated by positive igg rdt, were identified in three patients. dengue virus rt-pcr was negative in all cases. the detection cutoff of igg panbio rdt is set at a high titer (equivalent to a hemagglutination inhibition titer ³ 1,280), which is commonly found in recent secondary denv infection. 6 thus, these cases were probably acute covid-19 cases with recent secondary denv infection. retrospective assessment of exposure history revealed that one patient could have been infected by sars-cov-2 during hospitalization for dengue a week prior at a different hospital. another patient reported that a clinician he visited before hospitalization suspected denv infection clinically, but the ns1 result was negative. the third patient did not recall having a fever before acute covid-19 illness, suggesting asymptomatic or mild dengue, the most common presentation of denv infection. 7 in four patients, denv igm was detected but did not increase in follow-up samples, denv igg was only detected by elisa, and denv rt-pcr was negative, suggesting that denv infection occurred less recently than in the three patients described earlier. detection of igm is plausible as it may be detected until 1 year postinfection. 8 most patients (33, 78.6%) only had denv igg antibodies, implying past denv infection. this is consistent with previous studies conducted in indonesia, which demonstrated that more than 90% of adults aged > 18 years had been infected by denv. 5 no evidence of denv infection was identified in two (4.8%) patients. the distribution of dengue diagnostic results is shown in table 1 . despite concurrent high incidence of covid-19 and dengue in indonesia, acute coinfection with denv was not detected in this cohort of patients identified to have covid-19. this may be because of sars-cov-2 and denv testing practices, which focus on symptomatic cases. it is likely that coinfection is occurring but is often asymptomatic. it is also possible that some patients had already been infected with current circulating denv serotypes and thus had immunity, as indicated by high prevalence of patients with igg antibodies. identification of seven (16.7%) covid-19 cases with denv igm in our cohort may be related to the occurrence of covid-19 during the yearly dengue season. this finding is concerning, particularly because clinicians frequently diagnose dengue based only on denv igm, which may persist for months after resolution of infection. our study demonstrates that adding ns1 to the diagnostic algorithm may reduce dengue overdiagnosis attributable to reliance on igm. missed diagnosis of acute covid-19 due to presumption of dengue can result in inadvertent omission of targeted precautions, which could lead to transmission to contacts, including family, colocated patients, and healthcare workers. a missed diagnosis could also delay the receipt of standard of care covid-19 treatment. missed diagnoses have been reported in singapore due to false-positive denv igm rdt results 3 versus persistence of denv igm. in the setting of the current pandemic and in light of overlapping symptomatology, clinicians should test for both denv and sars-cov-2. it is notable that one of the patients may have contracted sars-cov-2 during hospitalization a week prior. in resourcelimited settings, adequate infection control practices are difficult to implement. hence, nosocomial infection should be considered in the setting of recent contact with the healthcare system, including due to dengue. sars-cov-2 infection control strategies for resource-limited settings are needed. findings from our study should be interpreted with caution. the study population was small and from only one hospital at tangerang district, during march 2020 and april 2020. therefore, results have limited generalizability as they reflect the epidemiology of covid-19 and dengue in that area during the study period. furthermore, as the assays were qualitative (rdt) or semi-quantitative (elisa), increasing antibody titer was only measured by the index value, which may be inaccurate. to reduce inaccuracy, we tested acute and followup specimens simultaneously. in conclusion, our study reaffirms challenges associated with diagnosing covid-19 in areas hyperendemic for tropical infections with overlapping presentations such as dengue. the known potential for repeat dengue infections and the possibility for repeat sars-cov-2 infections add further complication. when molecular diagnostic testing for denv is not available, we recommend the use of a validated ns1 and igm/igg rdt. addition of ns1 will improve the specificity of identifying acute dengue cases. 9 detection of denv igm and/ or high igg titer should not be considered an exclusion of covid-19. past infection with denv with acute covid-19 or even acute denv and sars-cov-2 coinfection would remain possibilities. 10 hence, evaluation for covid-19 should be conducted when dengue ns1 or rt-pcr (when available) is negative. e-mails: unurhayati@ina-respond.net, gsalim@ ina-respond.net, dpepy@ina-respond.net, hkosasih@ina-respond.net, wwahyunawang@gmail.co, amenur@ina-respond.net, yuanita_djajady@ yahoo.com, and rasari@ina-respond.net. dona arlinda and muhammad karyana a novel coronavirus from patients with pneumonia in china coinfection between dengue and covid-19: need for approach in endemic zones covert covid-19 and false-positive dengue serology in singapore force of infection and true infection rate of dengue in singapore: implications for dengue control and management dengue viral infection in indonesia: epidemiology, diagnostic challenges, and mutations from an observational cohort study clinical and laboratory diagnosis of dengue virus infection the epidemiology, virology and clinical findings of dengue virus infections in a cohort of indonesian adults in western java prolonged persistence of igm against dengue virus detected by commonly used commercial assays accuracy of dengue clinical diagnosis with and without ns1 antigen rapid test: comparison between human and bayesian network model decision covid-19 and dengue co-infection in a returning traveller this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-296363-qgoxlqoq authors: khan, yusra habib; mallhi, tauqeer hussain; alotaibi, nasser hadal; alzarea, abdulaziz ibrahim; alanazi, abdullah salah; tanveer, nida; hashmi, furqan khurshid title: threat of covid-19 vaccine hesitancy in pakistan: the need for measures to neutralize misleading narratives date: 2020-06-22 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0654 sha: doc_id: 296363 cord_uid: qgoxlqoq immediately after declaring covid-19 as a pandemic, numerous wild conspiracy theories sprouted through social media. pakistan is quite vulnerable to such conspiracy narratives and has experienced failures of polio vaccination programs because of such claims. recently, two well-known political figures raised conspiracy theories against covid-19 vaccines in pakistan, stating that covid-19 is a grand illusion and a conspiracy against muslim countries. this theory is much discussed in the local community, supporting covid-19 vaccine hesitancy. we urge healthcare authorities in pakistan to take necessary measures against such claims before they penetrate to the general community. anti-vaccine movements could undermine efforts to end the covid-19 pandemic. we believe that ethical and responsible behavior of mass media, a careful advisory from the pakistan electronic media regulatory authority, stern measures from healthcare authorities, effective maneuvers to increase public awareness on covid-19, vigorous analysis of information by data or communications scientists, and publication of counter opinions from health professionals against such theories will go a long way in neutralizing such misleading claims. because pakistan is experiencing a large burden of disease, with a sharp rise in confirmed cases, immediate action is of paramount importance to eradicate any potential barriers to a future covid-19 vaccination program. vaccine hesitancy remains a substantial challenge for pakistan amid various conspiracy theories. the failure to eradicate polio from the country is primarily attributed to such theories. of these, alleged poor quality of vaccines, questioning of dosing recommendations, religious prohibitions ("infidel vaccine"), and rumors related to the presence of active virus in the vaccines are some leading claims obstructing the anti-polio campaign in the country. 1 unfortunately, a conspiracy theory against covid-19 vaccine is currently being spread in pakistan. recently, a renowned political commentator and columnist in pakistan claimed that the virus was a grand illusion to target islamic nations, designed to allow jews to rule the world, and to include nano-chips imbedded in the bodies of people to gain control through 5g towers. 2 a similar theory was presented by an ex-foreign minister of pakistan, accusing the united states of inventing the virus in the united kingdom, with subsequent transfer to china for global spread. 3 these theories are actively discussed in the pakistani community through social media. in the country, where vaccine hesitancy is a prime barrier to curb vaccine-preventable diseases, such conspiracy narratives may plant seeds of resistance against upcoming covid-19 vaccination programs. since a long-term lockdown is not possible for many countries due to economic turmoil, availability of vaccines may be the only way to limit persistence of the pandemic. because of a weak healthcare system, dense population, and poor compliance with hygiene practices in pakistan, the propensity for disease spread is high. as pakistan has already experienced vigorous resistance against polio vaccination, any negative perception among the population toward covid-19 vaccines would have devastating implications regarding efforts to end the pandemic. we urge the government of pakistan to take necessary measures before anti-vaccine campaigns penetrate into the local community. in this context, we share possible measures to neutralize circulating false claims against covid-19 vaccines. because the volume of disparate falsehoods against covid-19 is increasing every day, the primary responsibility lies with pakistani media to play a sensible and professional role during the ongoing health crisis. media should avoid any exaggerated or amplified statements triggering negative perceptions related to covid-19 among the general community. television channels in pakistan should avoid airing unsupported conspiracy theories about covid-19. the most reasonable and ethical approach would be limiting discussions on covid-19 to healthcare professionals, rather than political or business figures. although free speech is a fundamental right of every citizen, public harm associated with false claims must be carefully weighed. another approach that could be useful is debates that offer opinions from researchers or healthcare professionals to counter conspiracy theories. the pakistani mass media is dynamic and has witnessed robust growth in recent years. there are more than 30 news channels in different languages currently aired in pakistan. the pakistan electronic media regulatory authority (pemra), 4 which works in collaboration with the ministry of information, regulates media activities in the country and holds power to suspend or cancel licensure of news channels. moreover, pemra also issues advisories to news channels on airing ethical content and refraining from airing mis-or disinformation. although pemra publishes regular reports on fake and misleading news, we did not come across any action against misleading information related to covid-19. the pakistan electronic media regulatory authority should issue guidelines on statements regarding the covid-19 pandemic. moreover, any person spreading unfounded theories without evidence should be accountable to law enforcement agencies. current conspiracy narratives are tied to religious beliefs. we suggest that religious elements can be addressed by involving enlightened islamic scholars in health promotion and awareness regarding covid-19. a similar approach was adopted by the country when the polio vaccine campaign was hindered by a conspiracy theory claiming that these vaccines were monkey-or pig-derived products, which are forbidden in islam. because most of the population in pakistan regards the advice of islamic scholars highly, the government invited scholars to educate the public on the polio vaccine, particularly in regions highly resistant to vaccination. these scholars highlighted the religious underpinning for the use of preventive medicine according to sharia law. 5 we believe that increased involvement of local religious authorities will facilitate appropriate covid-19 control efforts in pakistan. immunization campaigns in pakistan are controlled by the expanded program on immunization (epi), begun in 1978 in collaboration with the who and unicef. currently, the epi focuses on immunizing all children against eight vaccine-preventable diseases (diphtheria, tetanus, pertussis, polio, measles, tuberculosis, hepatitis b, and haemophilus influenza type b). 6 because the epi staff remain in contact with the public, their role in neutralizing misleading vaccine claims and in maximizing vaccine acceptance is of paramount importance. we suggest that the epi should swiftly respond to any anti-vaccine campaign in the country by providing accurate information to the public. recently, it was documented that confidence about vaccines is directly related to public awareness of infectious diseases. 7 a large survey on "attitudes to vaccines" involving 140,000 participants around the globe showed that countries with active public-awareness campaigns against various infectious diseases achieved very high rates of agreement on vaccine safety, effectiveness, and importance. 8 because conspiracy theories offer ammunition to vaccine deniers, timely intervention is of utmost importance. health authorities in pakistan must ensure stern measures to disseminate accurate and honest information to the public. health authorities should make it clear that they are listening and responding to the public's questions and concerns. keeping in view the large circulating volume of mis-or disinformation, researchers and public health educators need to build a society that is resilient to falsehood about covid-19, a task that will only become more vital as vaccines near. data scientists and communications researchers have the responsibility to analyze data related to such misleading information. it is not possible to stop people from spreading ill-founded rumors. however, analysis of information sources, patterns of spread, and impacts on the general community will foster effective strategies to flatten the curve of the infodemic so that misleading information cannot spread as far and as fast. 9 in the case of vaccines, transparent information on how vaccines are made, how they work, what they contain, how they will be tested, and their effectiveness, possible risks, and side effects will useful to ensure confidence in covid-19 vaccines when they are available. the government of pakistan has not taken a hard line against misleading covid-19 claims. we believe that putting full effort into the implementation of the aforementioned measures will go a long way to mitigating the proliferation of false claims in the country, and thereby help greatly in the control of the covid-19 pandemic in pakistan. polio vaccination controversy in pakistan bill gates' coronavirus vaccine will have nano trackers, will be controlled via 5g satellites to take islam out of muslims: pakistani 'expert' zaid hamid coronavirus is not natural but invented in a laboratory by israel, us and uk: former pakistan foreign minister comes up with a bizarre conspiracy theory pakistan electronic media regulatory authority 2002, as amended by the pemra amendment act polio vaccination in pakistan: by force or by volition? factors associated with non-utilization of child immunization in pakistan: evidence from the demographic and health survey 2006-07 don't demonize parents who are hesitant to vaccinate-discuss their worries instead how does the world feel about science and health? available at the epic battle against coronavirus misinformation and conspiracy theories this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-255940-chb4iuis authors: walton, david a.; ivers, louise c. title: facility-level approaches for covid-19 when caseload surpasses surge capacity date: 2020-06-26 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0681 sha: doc_id: 255940 cord_uid: chb4iuis as covid-19 cases continue to increase globally, fragile health systems already facing challenges with health system infrastructure, sars-cov-2 diagnostic capacity, and patient isolation capabilities may be left with few options to effectively care for acutely ill patients. haiti—with only two laboratories that can perform reverse transcriptase pcr for sars-cov-2, a paucity of hospital beds, and an exponential increase in cases—provides an example that underpins the need for immediate infrastructure solutions for the crisis. we present two covid-19 treatment center designs that leverage lessons learned from previous outbreaks of communicable infectious diseases and provide potential solutions when caseload exceeds existing capacity, with and without access to sars-cov-2 testing. these designs are intended for settings in which health facilities and testing resources for covid-19 are surpassed during the pandemic, are adaptable to local conditions and constraints, and mitigate the likelihood of nosocomial transmission while offering an option to care for hospitalized patients. several low-and middle-income countries have slowed transmission of covid-19 through rapid implementation of physical distancing policies; public health investment in testing, tracing, and isolating; and the mobilization of existing health workers. [1] [2] [3] however, covid-19 cases continue to increase in countries with fragile health systems, and lack of testing capacity and clinical capacities to care for ill patients may become barriers to providing effective care. 4 haiti, a country in which we have both worked as clinicians for over a decade, puts these needs in sharp relief. 5 at the time of this writing, pcr-confirmed cases of covid-19 in haiti are rapidly escalating, and they likely represent only a fraction of actual cases-the only two laboratories in the country with the ability to perform sars-cov-2 reverse transcription-pcr (rt-pcr) being overwhelmed with requests. 6, 7 other diagnostic modalities, such as the sars-cov-2 genexpert test, remain difficult to access, and rapid immunoassays have thus far proven to have limited or no utility for patient care. 8, 9 modeling by cornell university and oxford university projects up to 430,000 people in haiti might require hospitalization, which would require more than 9,000 additional hospital beds 10 ; however, fewer than 300 hospital beds are currently available for covid-19 patients. 11 hospitals are already beginning to report an inability to deal with the significant influx of patients. lack of access to hospital beds, let alone critical care beds, will likely continue to be a major bottleneck to the care of patients. haiti offers one example, but other countries have experienced similar challenges with vulnerable populations, often leveraging unique solutions to add additional bed capacity for covid-19 patients. 12, 13 in march, the who released guidelines on the establishment of treatment centers for severe acute respiratory infections. 14 the document is a thorough and welcome guide, created as a response to the covid-19 pandemic, but the infrastructure solutions proposed are likely to be out of reach for large portions of the global population living where health budgets are woefully inadequate to ensure safe delivery of health services in the midst of this pandemic. we have argued for more than a decade for additional health system investment as an approach to delivering global health equity; however, in haiti, the immediate gap between available hospital beds and the projected need for covid-19 hospitalizations seems insurmountable. any chance at caring for vast quantities of patients sick with respiratory illness will require a simple but effective model that can be implemented by local teams with limited equipment, resources, and testing capacity. to respond to the immediate crisis facing health workers and patients, we propose a covid-19 treatment center design ( figure 1 ) that harnesses lessons learned from other outbreaks and adheres to infection prevention and control principles recommended by the who for the novel coronavirus. although viral hemorrhagic fevers or diarrheal diseases have different transmission dynamics from sars-cov-2, principles of infection control and management of large quantities of patients in low-resource settings can carry through to covid-19. these principles include 1) screen anyone who presents at the facility with any complaint, 2) test patients for infection if they meet certain criteria, 3) isolate individuals with confirmed infection in private rooms or cohort in wards separated from noninfected patients, and 4) have all staff inside the facility in personal protective equipment. cholera treatment centers, typically set up for large outbreaks, cohort patients who meet the clinical case definition for cholera, and maintain stringent borders between the cholera-and non-cholera areas of health facilities. 15 a similar cohorting approach was used for ebola treatment centers in the 2014-2016 ebola epidemic, and although testing for ebola virus disease was an important component of triage, it was not always rapidly available. 16 our design acknowledges a stark reality: in some lowresource settings, the volume of acute respiratory illness cases may surpass surge capacity. the design assumes that two thresholds have been reached: first, the health center no longer has space to individually isolate covid-19 patients, and second, laboratory capacity is limited or surpassed, such that rapid, accurate testing for covid-19 may not be available, as is the reality facing our colleagues in haiti. the covid-19 treatment center is modular for rapid construction and designed to be adaptable to local conditions and constraints. the number of wards and overall footprint is flexible to adapt to the burden of disease and pragmatic budgetary constraints. similarly, the design allows for use of local construction materials: walls can be made with lumber and plywood or with concrete blocks for a more permanent structure; large tents can also be used for wards within the campus plan. nosocomial transmission can be mitigated with natural ventilation, which can often achieve or exceed 12 air changes per hour, and fans can be installed to improve thermal comfort for patients. 17, 18 ideally, but not necessarily, treatment centers would be located adjacent to existing medical facilities to leverage existing water and power supplies, and allow for the ability to care for non-covid-19 patients. in this design, concern for nosocomial transmission would remain high in the "suspect ward," where patients who may ultimately test positive for covid-19 mix with patients who may have malaria, tuberculosis, or another disease that presents similarly. [19] [20] [21] however, the design assumes that the ability to isolate suspect cases in individual rooms has been superseded by the number of cases. the risk of nosocomial infection in such wards can be mitigated (though not eliminated) by spacing beds at least 1.5 m apart and placing protective barriers between beds to reduce the spread of droplet nuclei. 22 if caseload supersedes bed capacity, a common pitfall is to place additional beds in the ward, but this should be avoided and priority given to preserving 1.5 m between beds, which allows the maximum number of beds within the dimensions of the space allocated while aiming to mitigate nosocomial spread. if feasible, patients not requiring oxygen can also wear surgical masks. the risk of nosocomial spread highlights the imperative to increase access to rapid, sensitive, and specific low-cost tests for sars-cov-2. in haiti, testing capacity for rt-pcr has already exceeded capacity. unable to rapidly scale up testing, figure 1 . covid treatment center campus with access to confirmatory sars-cov-2 testing. figure 2. covid treatment center campus without access to confirmatory sars-cov-2 testing. many facilities have started using a clinical case definition for the diagnosis of covid-19 even for hospitalized patients. while triaging testing resources has been used in other settings in which demand for testing exceeds capacity, including new york, california, and washington, having unknown covid-19 status for hospitalized patients presents special challenges. 23, 24 figure 2 demonstrates how the design and flow would change in a situation in which no covid-19 testing is available and cohorting of patients is instead achieved by case definition. with this design, the risk of nosocomial infection remains high but may be reduced with mitigation techniques used in the "suspect ward" (figure 3 ). as we have seen in haiti, the rapid surge of covid-19 in countries with fragile health systems has far surpassed the existing health infrastructure's ability to offer safe spaces to care for patients, with many countries yet to hit their peak caseload. without the ability to effectively isolate covid-19 patients or suspect patients who need hospitalization, we would expect a disproportionately higher morbidity and mortality than in other settings with capacity to receive a surge of cases. as one component of the emergency response, covid-19 treatment centers harness lessons learned from previous infectious disease outbreaks to cohort covid-19 patients who require hospitalization, mitigate (but not eliminate) the risks of nosocomial transmission, and streamline care. covid-19: decisive action is the hallmark of south africa's early success against coronavirus learn from rwanda's success in tackling covid-19 uganda's first 100 covid-19 cases: trends and lessons africa in the path of covid-19 epub ahead of print facing the monster in haiti figure 3. suspect ward design. facility-level approaches for covid-19: situation en haiti covid-19: les gens affluent au centre de dépistage de gheskio msf calls for no patents or profiteering on covid-19 drugs, tests, and vaccines in pandemic advice on the use of point-of-care immunodiagnostic tests for covid-19 entre appelà la solidarité et de sombres prévisions prayer and preparation: how one haiti hospital is confronting covid-19. miami herald perspectives on battling covid-19 in countries of latin america and the caribbean covid-19 and brazilian indigenous populations severe acute respiratory infections treatment centre cholera management and prevention at hôpital albert schweitzer derivation and internal validation of the ebola prediction score for risk stratification of patients with suspected ebola virus disease natural ventilation for infection control in health-care settings natural ventilation for reducing airborne infection in hospitals the 2019 novel coronavirus outbreak -a global threat world health organization differential diagnosis of illness in patients under investigation for the novel coronavirus (sars-cov-2) modes of transmission of virus causing covid-19: implications for ipc precaution recommendations as the coronavirus accelerates in haiti, top doctor says no need to test. miami herald in hard-hit areas, testing restricted to health care workers, hospital patients. the washington post acknowledgments: we would like to acknowledge colleagues at build health international for contributions on design, engineering, and architectural drawings. publication charges for this article were waived due to the ongoing pandemic of covid-19. this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-271906-2nm55024 authors: quincho-lopez, alvaro; quincho-lopez, dania l.; hurtado-medina, fernando d. title: case report: pneumothorax and pneumomediastinum as uncommon complications of covid-19 pneumonia—literature review date: 2020-07-23 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0815 sha: doc_id: 271906 cord_uid: 2nm55024 as the covid-19 pandemic progresses, awareness of uncommon presentations of the disease increases. such is the case with pneumothorax and pneumomediastinum. recent evidence suggested that these can occur in the context of covid-19 pneumonia, even in the absence of mechanical ventilation–related barotrauma. we present two patients with covid-19 pneumonia complicated by pneumomediastinum. the first patient was a 55-year-old woman who developed covid-19 pneumonia. her clinical course was complicated by pneumothorax and pneumomediastinum, and, unfortunately, she died 2 days following the admission. the second patient was a 31-year-old man who developed a small pneumomediastinum and was managed conservatively. he had a spontaneous resolution of the pneumomediastinum and was discharged 19 days later. none of our patients required invasive or noninvasive positive pressure ventilation. we performed a literature review of covid-19 pneumonia cases that developed pneumothorax, pneumomediastinum, or both. the analysis showed that the latter had high mortality (60%). thus, it is necessary to pay attention to these complications as early identification and management can reduce the associated morbidity and mortality. both pneumothorax and pneumomediastinum are known complications of mechanical ventilation due to intubation. 1, 2 nonetheless, even without barotrauma involved, pneumothorax or pneumomediastinum, or more rarely both, can be present in the context of covid-19. 3, 4 herein, we report two cases of patients infected with sars-cov-2, who developed pneumomediastinum, and one of them also presented pneumothorax. we also performed a relevant literature review using the scopus database. the first case was a 55-year-old woman with a past medical history of hypertension, uncontrolled bronchial asthma interspersed with periods of inactivity, and morbid obesity. she presented to the emergency department (ed) with 7 days of marked dyspnea, chest pain, and dry cough. previously, she received outpatient treatment with prednisone and dexamethasone every 8 hours for 5 days. on admission, her vital signs showed tachypnea (22 breaths/minute), with high temperature (38.2°c), increased heart rate (110 beats/minute), and 85% saturation. on physical examination, she had bilateral basal crackles and peripheral cyanosis. laboratory results showed an elevated c-reactive protein (crp) of 2.66 mg/dl (normal range 0-0.50 mg/dl). her blood count showed leukocytosis (30,270 cells/μl) with a lymphocyte count of 1,210 cells/μl. the patient was reactive to the covid-19 igg/igm rapid test. non-contrast chest computed tomography (ct) showed some ground-glass opacities of peripheral subpleural location, associated with multiple areas of consolidation in posterior segments of both lower lobes, with the presence of pneumothorax (approximately 20%) and pneumomediastinum ( figure 1a and b). she received treatment with azithromycin, ceftriaxone, hydrocortisone, and supplemental oxygen with a reservoir mask. she did not receive noninvasive positive pressure ventilation. the pneumothorax and pneumomediastinum were managed conservatively. however, despite the support measures, the patient died from respiratory failure 2 days after admission. the second case was a 31-year-old man with a past medical history of chronic gastritis and hypercholesterolemia in control, who presented to the ed with 6 days of dyspnea, general malaise, dry cough, and continuous fever for 4 days. on admission, tachypnea (22 breaths/minute), high temperature (38.5°c), normal heart rate (94 beats/minute), and saturation of a new ct was performed because of desaturation after the removal of oxygen support from the patient (on day 15 of hospitalization) and showed some foci of consolidation in posterior segments of both lower lobes, associated with parenchymal bands in both hemithoraces, with the presence of laminar air content predominantly on the right side, consistent with pneumomediastinum ( figure 1c ). the patient remained hospitalized for 19 days. a conservative management was chosen because the pneumomediastinum was very small, and its resolution was observed in the subsequent control after 7 days of discharge. the symptoms of sars-cov-2 infection have been widely characterized in large studies, with fever, cough, and dyspnea being the most frequent. these same studies indicate that only 1-2% of patients developed pneumothorax 5, 6 ; although it may occur as the disease progresses, 3 its presentation is still infrequent, like pneumomediastinum. 4 the mechanism is not fully elucidated, although it is probably because of rupture of the alveolar wall due to the increasing pressure difference between the alveolus and the pulmonary interstitium. 7, 8 pneumothorax and pneumomediastinum are defined as the presence of free air in the pleural and mediastinal cavities, respectively. 9,10 spontaneous pneumothorax can be primary or secondary, depending on the absence or presence of an underlying lung disease. 11 by contrast, pneumomediastinum can be primary, or spontaneous, if the cause is idiopathic, or secondary if it responds to a known etiology, whether traumatic or iatrogenic. 10 chest pain and dyspnea are the most common symptoms. 11, 12 an important difference is that pneumothorax occurs mainly at rest, 9 whereas strenuous physical activity has been reported as a triggering event for developing pneumomediastinum. 12 however, both are more frequent in males. 9, 12 drug abuse, asthma, and other lung diseases such as chronic obstructive pulmonary disease and interstitial lung disease are some predisposing factors, with tobacco being the most important risk factor in both. 9,10 none of our patients were active smokers, and only the first one had asthma as a risk factor in developing one of the two complications. chest ct as a diagnostic test to identify patients with covid-19 has a high sensitivity and a high negative predictive value. 13 moreover, depending on the lung involvement, it shows different phases: the early phase, where the groundglass pattern of subpleural distribution uni-or bilateral predominates; the progression phase, where, in addition to ground-glass involvement, there are also paved areas or "crazy paving" with diffuse or multi-lobar distribution; the peak, or the most affected phase, where the affected areas progressively consolidate; and, finally, the absorption phase, where ground glass appears secondary to the absorption of consolidations. 14 table 1 presents a summary of case reports of patients infected with sars-cov-2 who presented pneumothorax, pneumomediastinum, or both. seventeen reports describing 20 patients were found. in total, half of the patients presented a favorable evolution (50%; 10/20), whereas 30% (6/20) died. follow-up was not reported in four patients. it is important to note that subcutaneous emphysema was a radiological finding present in 35% (7/20) of the patients, two in pneumothorax, 15, 16 three in pneumomediastinum, 4, 7, 17 and two in both. 18, 19 the treatment for covid-19 was variable. nevertheless, for the management of pneumothorax or pneumomediastinum, some patients (40%; 8/20) required a chest tube drainage, 8, 16, [19] [20] [21] [22] [23] [24] others (5%; 1/20) also required needle aspiration, 24 and thoracoscopy and bleb resection were required in two (10%) cases of persistent or recurrent pneumothorax. 25 the remaining patients were either managed conservatively or not reported. in those patients with pneumothorax, the majority were male (88.8%; 8/9), and 55.5% (5/9) had some comorbidities. fever was the most frequent symptom on admission (66.6%; 6/9). furthermore, the majority presented a favorable evolution (55.5%; 5/9), with those with the highest number of associated comorbidities having the worst evolution (22.2%; 2/9). followup was not reported in one patient. pneumothorax may also present as a late sequel to covid-19. 21 although most cases report spontaneous pneumothorax, tension pneumothorax is also a possible complication. 23, 24 bulla associated with pneumothorax is reported in two patients. 15 ,24 some authors consider the rupture of a bulla to be the cause of spontaneous pneumothorax. 9 of the pneumomediastinum cases, male gender was the most affected (66.6%; 4/6), and only 33.3% (2/6) presented any associated comorbidity. however, in 50% of the cases, the risk factors were not reported. in 83.3% (5/6), fever was reported as the most frequent symptom, and one patient did not present any symptoms. the evolution was favorable in 50% of the cases. of the patients who presented pneumothorax and pneumomediastinum at the same time, most of the patients were male (60%; 3/5), and 40% (2/5) had some associated comorbidity. fever was the most frequent symptom (100%), followed by dyspnea (80%) and cough (60%). death was inevitable in 60% (3/5) of the patients. some limitations that we can point out are that certain articles that are not indexed in scopus could not be included in our review. however, we decided to use scopus because it contains all the documents included in medline, 26 ensuring not only quantity but also quality of the documents. furthermore, we only considered articles written in english. our literature review was last updated on july 3, finding 32 results, of which only 17 are case reports. to enable the reproducibility of our review, we display our search query: (title-abs {"2019-ncov" or "covid-19" or "ncovid-19" or "hcov-19" or "sars-ncov" or "sars-cov-2" or "severe acute respiratory syndrome coronavirus 2"} or title-abs {coronavirus w/2 [wuhan or china or novel or pneumonia]}) and (title-abs [pneumomediastinum or pneumothorax]) and (limit-to [pubyear, 2020] ). in conclusion, pneumothorax and pneumomediastinum are possible complications of covid-19 pneumonia, causing acute decompensation that can worsen the prognosis of patients, especially those with underlying lung diseases. emergency tracheal intubation in 202 patients with covid-19 in wuhan, china: lessons learnt and international expert recommendations portable chest x-ray in coronavirus disease-19 (covid-19): a pictorial review coronavirus disease 2019 (covid-19): a systematic review of imaging findings in 919 patients covid-19 with spontaneous pneumomediastinum epidemiological and clinical characteristics of 99 cases of 2019 novel coronavirus pneumonia in wuhan, china: a descriptive study clinical course and outcomes of critically ill patients with sars-cov-2 pneumonia in wuhan, china: a single-centered, retrospective, observational study sars-cov-2 pulmonary infection revealed by subcutaneous emphysema and pneumomediastinum pneumomediastinum and spontaneous pneumothorax as an extrapulmonary complication of covid-19 disease spontaneous pneumothorax: epidemiology, pathophysiology and cause spontaneous pneumomediastinum: time for consensus spontaneous pneumothorax systematic review of spontaneous pneumomediastinum: a survey of 22 years' data correlation of chest ct and rt-pcr testing in coronavirus disease 2019 (covid-19) in china: a report of 1014 cases time course of lung changes on chest ct during recovery from 2019 novel coronavirus (covid-19) pneumonia mediastinal emphysema, giant bulla, and pneumothorax developed during the course of covid-19 pneumonia sars-cov-2 pneumonia with subcutaneous emphysema, mediastinal emphysema, and pneumothorax: a case report spontaneous pneumomediastinum in covid-19 covid-19 with spontaneous pneumothorax, pneumomediastinum and subcutaneous emphysema spontaneous pneumothorax and subcutaneous emphysema in covid-19 patient: case report perforated acute abdomen in a patient with covid-19: an atypical manifestation of the disease spontaneous pneumothorax following covid-19 pneumonia sars-cov-2 infection associated with spontaneous pneumothorax secondary tension pneumothorax in a covid-19 pneumonia patient: a case report tension pneumothorax in a patient with covid-19 management of persistent pneumothorax with thoracoscopy and blebs resection in covid-19 patients comparison of pubmed, scopus, web of science, and google scholar: strengths and weaknesses spontaneous pneumomediastinum occurring in the sars-cov-2 infection spontaneous pneumomediastinum in a patient with coronavirus disease 2019 pneumonia and the possible underlying mechanism spontaneous pneumomediastinum: a probable unusual complication of coronavirus disease 2019 (covid-19) pneumonia acknowledgment: publication charges for this article were waived due to the ongoing pandemic of covid-19.financial support: this study was self-financed.disclosure: all patients provided consent to share their cases.authors' addresses: alvaro quincho-lopez, san fernando medical school, universidad nacional mayor de san marcos, lima, peru, e-mail: alvaro209.ql@gmail.com. dania l. quincho-lopez and fernando d. hurtado-medina, diagnostic imaging service, hospital ii lnc luis negreiros vega essalud, lima, peru, e-mails: danita.lizzi.mc@gmail.com and fernando1979sf@hotmail.com. this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-260444-ooi5x9p3 authors: gadelha farias, luís arthur brasil; gomes moreira, ana livia; austregésilo corrêa, eduardo; landim de oliveira lima, cicero allan; lopes, isadora maria praciano; de holanda, pablo eliack linhares; nunes, fernanda remígio; pires neto, roberto da justa title: case report: coronavirus disease and pulmonary tuberculosis in patients with human immunodeficiency virus: report of two cases date: 2020-08-18 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0737 sha: doc_id: 260444 cord_uid: ooi5x9p3 coinfection of sars-cov-2/mycobacterium tuberculosis (mtb) in patients with hiv/aids has not been previously reported. here, we present two cases of coinfection of sars-cov-2 and mtb in patients with hiv. the first case is a 39-year-old patient who was admitted with a 7-day history of fever, myalgia, headache, and cough. the second patient is a 43-year-old man who had a 1-month history of cough with hemoptoic sputum, evolving to mild respiratory distress in the last 7 days. both patients already had pulmonary tuberculosis and subsequently developed sars-cov-2 infection during the 2020 pandemic. nonadherence to antiretroviral treatment may have been a factor in the clinical worsening of the patients. covid-19, caused by sars-cov-2, represents a new fatal disease mainly characterized by a respiratory illness. 1 on march 11, 2020, the who officially declared the infection to have risen to a pandemic state. 2, 3 in brazil, the first case of covid-19 was confirmed on february 26, 2020, and community transmission was recognized on march 20, 2020. 4, 5 on may 24, 2020, there were almost 350,000 confirmed cases and approximately 22,000 deaths, with a mortality rate of 6.9%. the most affected states were são paulo (82,161), rio de janeiro (37,912), and ceará (35,595). 6, 7 the coinfection of sars-cov-2 and mycobacterium tuberculosis (mtb) in patients with hiv infection is a matter of concern and has not been well studied. here, we present two cases of triple coinfections (hiv/sars-cov-2/mtb) in patients admitted to sao josé hospital of infectious diseases, fortaleza, ceará, brazil. both patients provided informed consent to publish their clinical data. nasopharyngeal swab samples were used for covid-19 diagnoses based on the amplification of the betacoronavirus e gene and the specific sars-cov-2 rdrp gene using pcr. sputum samples were used for mtb diagnosis. case report case 1. a 39-year-old italian man who had been living in brazil for more than 20 years arrived at the emergency department (ed) in april 2020 with a known diagnosis of hiv/ aids (hiv viral load 293,313 copies/mm 3 and cd4 cell count 145/mm 3 ) and a 7-day history of sudden fever (temperature 38.0°c), myalgia, headache, and cough. he had poor adherence to antiretroviral therapy and previous use of marijuana and crack. he denied recent travel. he also tested positive for the surface antigen of the hepatitis b virus. he reported previous tuberculosis (tb) treatment for 3 months, although he did not complete the scheme. on physical examination, he was cachectic (weight 38 kg), with mild respiratory distress, a heart rate of 108 bpm, a respiratory rate of 18 rpm, and blood oxygen saturation (spo 2 ) levels of 93% without supplementary oxygen. no lymphadenopathy was observed. a chest examination revealed bilateral crepitations, rhonchi, and wheezes. chest computed tomography (ct) revealed a large cavitation located in the left lung associated with bilateral glass-ground opacities ( figure 1a -d). blood cell count presented low hemoglobin (5.4 g/dl) and hematocrit levels (16.5%), lymphopenia (408/mm 3 ), and normal platelet and white cell count ranges. the c-reactive protein level was markedly elevated (118.8 mg/l). hepatic and renal functions were normal. d-dimer, troponin, aspartate transaminase (ast), alanine transminase (alt), and ferritin levels were at normal range. acid-fast bacilli smears were positive, and mtb dna was detected using pcr (genexpert mtb/rifampicin [rif] assay) without rif resistance. during hospitalization, the patient experienced respiratory distress and required continuous oxygen via nasal cannula 2 l/minute during the first 3 days. admission to the intensive care unit and invasive mechanical ventilation was not required. treatment with isoniazid, ethambutol, pyrazinamide, and rif was initiated. he was also treated with azithromycin (500 mg/day), hydroxychloroquine (hcq) (400 mg/day) for 5 days, and ceftriaxone (2 g/day). antiretroviral therapy was not started to avoid complications with anti-tb treatment. after being hospitalized for 3 weeks, the patient was clinically stable and discharged to home with outpatient follow-up. antiretroviral therapy was planned to be initiated after the first 8 weeks of anti-tb treatment. 8 case 2. a 43-year-old brazilian man arrived at the ed in may 2020 with a history of cough with hemoptoic sputum for more than 1 month, evolving to mild respiratory distress in the last 7 days. he did not show signs of a fever and other symptoms. he had a previous history of hiv/aids with no adherence to antiretroviral therapy, illicit drug abuse, and had generalized anxiety disorder. he had lost clinical and laboratory follow-up in the last 5 years. in 2015, the hiv viral load was 9,054 copies/ mm 3 , and the t-cd4 cell count was 407/mm 3 . on physical examination, his weight was 55 kg, heart rate was 79 bpm, respiratory rate was 18 rpm, and spo 2 was 96%. no lymphadenopathy was observed. blood cell count presented low hemoglobin (9.9 g/dl) and hematocrit levels (29.7%), and normal platelet and white cell count ranges. lactate dehydrogenase and c-reactive protein levels were 312 u/l, above reference limits (140-271 u/l), and 52.3 mg/dl, respectively. the d-dimer level (0.6 mcg/ml) exceeded the normal range. troponin, ast, alt, and ferritin levels were in normal range. chest ct revealed bilateral glass-ground opacities occupying approximately 25% of both lungs (figure 2a-d) . respiratory secretion was collected with a nasopharyngeal swab and tested positive for sars-cov-2. mycobacterium tuberculosis dna was detected using pcr (genexpert mtb/ rif assay) without rif resistance. treatment was initiated with azithromycin (500 mg/day), hcq (400 mg/day) for 5 days, and ceftriaxone (2 g/day). he was treated with isoniazid, ethambutol, pyrazinamide, and rif. antiretroviral therapy was not started to avoid complications with anti-tb treatment. the patient was clinically stable and discharged to home after 1 week. currently, the patient is under follow-up and remains asymptomatic without developing relapses. to the best of our knowledge, this is the first reported case of coinfection of sars-cov-2/mtb in patients with hiv/aids. hiv/mtb coinfection is prevalent worldwide-especially in low-income and developing countries such as brazil-and its clinical consequences are well known. hiv patients with mtb latent infection have a higher risk of reactivation. tuberculosis is the most common opportunistic disease and tends to be more lethal in cases of hiv. extrapulmonary forms and disseminated disease occur more frequently in hiv, especially in those with high viral load and low t-cd4 cell count. however, as the covid-19 pandemic develops, there is a concern regarding its clinical and epidemiological relevance in hiv/ mtb-coinfected patients. covid-19 is an acute viral infection with frequent and severe pulmonary involvement, which can lead to hospitalizations and deaths. the lungs can be directly affected by the virus, leading to viral pneumonia. the immune system is also markedly affected and challenged by sars-cov-2. leukopenia, lymphopenia, and an inflammatory cytokine storm are some of these immunological changes. treatment with immunosuppressive drugs, such as corticosteroids, has been adopted in some cases. in this context, the impact of covid-19 on hiv viral replication in hiv patients with mtb latent infection and in patients with hiv and active tb must be subjects of research. this includes the spectrum of clinical manifestations of the triple coinfections. there are some reports that show that sars/mtb and middle east respiratory syndrome-cov/mtb coinfections can augment other infections. 9 one observational study on sars-cov-2/mtb coinfection concluded that mtb infection possibly increases covid-19 susceptibility and severity. 10 blanco et al. 11 recently published a case series of five hivinfected patients with covid-19, of whom none of them died, although two needed intensive care supportive therapy. gervasoni et al. 12 described 47 hiv-positive patients hospitalized by covid-19 and concluded that hiv patients were not at greater risk of severe disease or death than hiv-negative patients. motta et al. 13 compared 69 patients with sars-cov-2/ mtb coinfections, although they could not show tb as a major predictor of mortality in these patients. this appeared to be true in patients reported in the present study because even with hiv infection association, the patients did not evolve to a greater gravity. however, according to he et al., 14 patients with previous lung disease such as treated or untreated tb could affect the prognosis of patients with covid-19, making greater vigilance necessary during outpatient follow-up. the present article reports two cases of covid-19 in patients with hiv/mtb coinfections. both patients already had active tb before being infected with sars-cov-2. they both had a history of irregular antiretroviral treatment, detectable hiv viral loads, and low t-cd4 lymphocyte counts (< 500). molecular tests (pcr and reverse transcription pcr) were positive for mtb and sars-cov-2 simultaneously. there was an overlap of symptoms commonly seen in active tb and covid-19, such as cough and fever. one of the patients had hemoptoic sputum. chest ct findings showed patterns that could be related to both diseases, although they cannot be differentiated from other pulmonary diseases such as bacterial pneumonia and chronic obstructive pulmonary disease. although the risk factors for covid-19 still need to be fully understood, the two cases presented here may indicate that hiv/mtb coinfection could be another risk factor to be considered when evaluating sars-cov-2-infected patients. nonadherence to antiretroviral treatment may have been a factor in the worsening of mtb/sars-cov-2 coinfection. 15 the possibility of associated bacterial pneumonia could not be ruled out in both cases, and antibiotic coverage with ceftriaxone and azithromycin was chosen. both patients were treated with hcq. thus, the usefulness of this drug is controversial. on june 17, 2020, based on evidence from the solidarity trial, u.k. recovery trial and a cochrane review, who announced that the hcq arm of the solidarity trial to find an effective covid-19 treatment was being stopped. both showed that hcq does not result in the reduction of mortality of hospitalized covid-19 patients. 16 this study has some limitations. herein, we studied only two cases of sars-cov-2 and mtb coinfection in hiv-infected patients. interleukin-6 was not available at our center. it is important to carefully evaluate suspected sars-cov-2 patients in the presence of other infectious diseases, such as tb, especially if cohorting is performed for suspected sars-cov-2 to avoid nosocomial transmission. a pneumonia outbreak associated with a new coronavirus of probable bat origin discurso de abertura do diretor-geral da oms na conferência de imprensa sobre covid-19 how brazil can hold back covid-19 ministério da saúde declara transmissão comunitária nacional.brasília, brazil: ministério da saúde covid-19 and pulmonary tuberculosis in patients with hiv da-saude-declara-transmissao-comunitaria-nacional painel de casos de doença pelo coronavírus 2019 (covid-19) no brasil pelo ministério da saúde. brasília, brazil: ministério da saúde decreto no33.519, de 19 de março de 2020. intensifica as medidas para enfrentamento da infecção humana pelo novo coronavírus. ceará, brazil: editoração casa civil painel de casos de doença pelo coronavírus 2019 (covid-19) no brasil pelo ministério da saúde consolidated guidelines on the use of antiretroviral drugs for treating and preventing hiv infection: recommendations for a public health approach middle east respiratory syndrome coronavirus and pulmonary tuberculosis coinfection: implications for infection control active or latent tuberculosis increases susceptibility to covid-19 and disease severity. medrxiv covid-19 in patients with hiv: clinical case series clinical features and outcomes of hiv patients with coronavirus disease 2019. clin infect dis (epub ahead of print tuberculosis, covid-19 and migrants: preliminary analysis of deaths occurring in 69 patients from two cohorts epub ahead of print the burden of covid-19 in people living with hiv: a syndemic perspective hydroxychloroquine and covid-19 this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-253459-tcn10pho authors: moreau, gregory brett; burgess, stacey l.; sturek, jeffrey m.; donlan, alexandra n.; petri, william a.; mann, barbara j. title: evaluation of k18-hace2 mice as a model of sars-cov-2 infection date: 2020-07-28 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0762 sha: doc_id: 253459 cord_uid: tcn10pho murine models of sars-cov-2 infection are critical for elucidating the biological pathways underlying covid-19. because human angiotensin-converting enzyme 2 (ace2) is the receptor for sars-cov-2, mice expressing the human ace2 gene have shown promise as a potential model for covid-19. five mice from the transgenic mouse strain k18-hace2 were intranasally inoculated with sars-cov-2 hong kong/vm20001061/2020. mice were followed twice daily for 5 days and scored for weight loss and clinical symptoms. infected mice did not exhibit any signs of infection until day 4, when no other obvious clinical symptoms other than weight loss were observed. by day 5, all infected mice had lost around 10% of their original body weight but exhibited variable clinical symptoms. all infected mice showed high viral titers in the lungs as well as altered lung histology associated with proteinaceous debris in the alveolar space, interstitial inflammatory cell infiltration, and alveolar septal thickening. overall, these results show that the k18-hace2 transgenic background can be used to establish symptomatic sars-cov-2 infection and can be a useful mouse model for covid-19. an invaluable step in identifying effective vaccines and therapies to combat covid-19 is the availability of a mouse model of infection. the host receptor for sars-cov-2 is the human angiotensin-converting enzyme 2 (hace2), 1 which was previously identified as the receptor for the sars-cov-1 2 that causes sars, a disease that emerged from china in 2002. 3 the mouse ace2 ortholog, which has significant amino acid sequence variation in the viral receptor binding domain, cannot serve as an efficient receptor for either sars-cov-2 or cov-1. 4 a transgenic mouse model to study sars-cov-1 infection was developed that expresses the hace2 gene under the control of the human cytokeratin 18 promoter. 5 infection of these mice with sars-cov-1 results in a rapidly lethal infection. 5 four other hace2-expressing mouse lines have been created to date and tested for the ability to support sars-cov-2 infection. two lines express the hace2 gene under the control of the mouse ace2 promotor 6, 7 ; one was made using the crispr/cas9 technology. 7 the third strain uses the lung ciliated epithelial cell hepatocyte nuclear factor-3/ forkhead homologue 4 (hfh4) promoter. 8, 9 an additional approach was to transfect wild-type mice with an adenovirus carrying the hace2 gene. 10 overall, with the exception of the hfh4 mice, in which there was some lethality, infection of these three mouse strains with sars-cov-2 results in mild clinical symptoms and no lethality. here, we report the infection of k18-hace2 with sars-cov-2. although this infection resembled that of other strains, we observed variable clinical presentation, with some mice exhibiting more severe symptoms than reported using other models. overall, this work supports the usefulness of k18-hace2 transgenic mice as a model for human covid-19 infections. to investigate the potential of this transgenic mouse strain as a model for covid-19 infection, five k18-hace2 mice were intranasally inoculated with 8 × 10 4 median tissue culture infectious dose (tcid50) of sars-cov-2, and five mice were mock-infected with sterile dulbecco's modified eagle's medium (dmem). mice were followed twice daily for 5 days and scored for clinical symptoms (weight loss, eye closure, appearance of fur [piloerection] and posture, and respiration). the mock-infected mice did not exhibit any clinical symptoms or experience any weight loss throughout the experiment. infected mice did not exhibit any measurable clinical symptoms through day 3. on day 4, no other clinical symptoms other than weight loss were observed. on day 5, all the infected mice had lost around 10% of their original weight ( figure 1a ) and exhibited variability in other clinical signs of infection, with clinical scores ranging from 3 to 9 (maximal score 14) ( figure 1b ). although two of the infected k18-hace2 mice showed only mild symptoms at day 5 (weight loss and reduced activity), two mice exhibited piloerection. the most severe mouse had increased respiration, lethargy, and slight eye closure and met our criteria for euthanasia. because the study was ended on day 5, it is unclear whether the remaining four mice would have recovered if the study was carried past day 5. although the clinical severity was variable between infected k18-hace2 mice, our results suggest that these mice present with more symptomatic disease than other hace2 mouse models of sars-cov-2 infection. in the mouse model expressing hace2 under the mouse ace2 promoter, infected mice did not exhibit any clinical symptoms other than maximal weight loss on day 3 postinfection, and those mice recovered. 7 only mild ruffling of fur and up to 8% weight loss on day 5 were observed in the other model using the mouse ace2 promoter, and once again, all mice recovered. 6 in mice transfected with an adenovirus carrying the hace2 gene, mice exhibited about a 10% weight loss on day 4 postinfection but no lethality. 10 in contrast to these models, in which mice exhibited mild symptoms and recovered, only 60% of the mice survived past day 5 in the mouse strain expressing hace2 under the lung ciliated epithelial cell hfh4 promoter. 9 although this model had higher lethality, weight loss was only about 5% and these mice had no respiratory symptoms. the authors hypothesize that mortality may be due to neuroinvasion because virus was detected in the brain. in k18-hace2 mice infected with sars-cov-1, the course of infection is clearly different; the infection is uniformly fatal, beginning on day 4 postinfection, and mice were symptomatic with labored breathing and lethargy. 5 although the number of mice used in this study was small and we were not able to measure survival, our data support a difference in the disease progression between these two viruses. all mice were euthanized on day 5, and tissue was collected for dissection and enumeration of viral loads. no significant differences in histology of the spleen, small intestine, or liver were observed between infected and mock-infected mice, and these tissues were normal in size and appearance. dissection of the lungs of infected mice revealed a mottled or marbled appearance that was not observed in mock-infected mice (data not shown). lung sections were analyzed after staining with hematoxylin and eosin and scored based on tissue pathology. 11 sars-cov-2-infected mice exhibited significantly higher histopathology scores than mock-infected mice ( figure 2 ). the major histopathology findings in infected mice were proteinaceous debris in the alveolar space, neutrophils in the interstitial space, and alveolar septal thickening ( figure 2 ); these observations were consistent with other hace2 mouse models, which also detected signs of lung injury including interstitial pneumonia, inflammatory cell infiltrates, and alveolar septal thickening. 6, 7 consistent with the observed infiltrating neutrophils, granulocytes and inflammatory monocytes were also elevated in the bronchoalveolar lavage (bal) fluid from the infected mice ( figure 3 ). other hace2 mouse models of covid-19 infection have observed high viral titers in the lungs with limited viral load in organs such as the liver and spleen during intranasal infection. 6,7 although we did not investigate viral load in the liver or spleen, these organs appeared normal by histology, suggesting that there was limited viral titer in these tissues. virus was detected in the lungs of all infected mice, with titers generally in the range of 1 × 10 5 plaque forming units (pfu)/ml (table 1 ). viral titers in the lungs appeared somewhat associated with disease severity: mouse 1390, which had the highest lung titer, had the highest clinical score, histopathology score, and percent weight loss at day 5 (table 1 ) and the highest numbers of neutrophils, monocytes, and eosinophils in the bal (figure 3 ). in addition, mouse 1413, which had the lowest titer, had the lowest clinical score, second lowest percent weight loss at day 5 (table 1) , and lowest number of eosinophils and monocytes in the bal (figure 3 ). of note, mouse 1413 did not have the lowest histopathology score (table 1) . although there were trends toward higher viral titers in the lungs being associated with higher clinical and histopathology scores, these trends were not significant, and viral titer was not a strong predictor of percent weight loss. the power of this analysis is limited by the small sample size, but these results suggest that factors in addition to viral load, such as inflammatory responses, are driving the severity of disease. this would also potentially explain the sudden onset of clinical symptoms at 5 days post-inoculation. in this report, we have described the course of sars-cov-2 infection in k18-hace2 transgenic mice. our findings are consistent with other studies using hace2 mice, which observed successful infection with sars-cov-2 and a milder disease severity compared with sars-cov-1. 6, 7 the onset of symptoms was abrupt, manifesting on day 5. mice exhibited a similar degree of weight loss but a varying degree of symptoms and clinical/histopathological scores. the number of mice used in this study was too small to determine whether this was a result of experimental variability or natural variability in outcomes. the variance in clinical and histopathological scores may be partially explained by viral titer, but there are likely other factors, such as the host immune response, that contribute to the variance observed. the observation of more severe disease in a subset of the k18-hace2 mice is distinct from other hace2-expressing covid-19 models, which typically observed only mild clinical symptoms. 6, 7 this could be due to experimental differences such as strain differences or the challenge dose (table 2 ). to date, little is known about the possibility of virulence differences among isolates. hong kong/vm20001061/2020 and strain 2019n-cov/usa_wa1/ 2020 are closely related and have been classified as type ib. 12 the receptor-binding domains of these strains are 100% identical (data not shown). the phylogeny of hb-01 and wuhan/amms01/2020 has not been reported. the challenge dose used in each experiment is similar; our experiment used the lowest amount of inoculum. the resident microbiota in each mouse strain could also impact outcomes of infection. the other difference between these strains is in the level of hace2 receptor expression or tissue distribution. nonetheless, k18-hace2 transgenic mice may be a particularly useful , and respiratory rate (0-2). all mouse work was approved by the university's institutional animal care and use committee, and all procedures were performed in the university's certified animal biosafety level three laboratory. histology. tissues were fixed in formaldehyde. slides were scanned at ×20 magnification. histopathological scoring for lung tissues was performed according to the guidelines of the american thoracic society. 11 a two-tailed student's t test was used to determined statistical significance. viral titers. the left lobe of the lung was homogenized in 1 ml serum-free dmem with a disposable tissue grinder. plaque assays were performed as described. 13 in brief, vero c1008, clone e6 (atcc crl-1586) cells grown in dmem (gibco 11995-040) with fetal bovine serum (fbs) were seeded into at a concentration of 2 × 10 5 cells/well the night before the assay. serial dilutions were added to the wells. the plate was incubated at 37°c, 5% co 2 for 2 hours, shaking the plates every 15 minutes. after 2 hours, the media was replaced with a liquid overlay of dmem, 2.5% fbs containing 1.2% avicel ph-101 (sigma aldrich, st. louis, mo) and incubated at 37°c, 5% co 2 . after 3 days, the overlay was removed, wells were fixed with 10% formaldehyde, and stained with 0.1% crystal violet to visualize plaques. plaques were counted, and pfus were calculated according to the following equation: average # of plaques/dilution factor × volume diluted virus added to the well. 8, 9 not specified not specified ∼ 5% weight loss, no clinical symptoms, but only 60% survived adenovirus transfection 10 10 5 focus-forming units strain 2019n-cov/usa_wa1/2020 10% maximum weight loss, all recovered hace2 = human angiotensin-converting enzyme 2. this work was supported by nih grants r01 ai124214 to w. a. p., r01 ai146257 to s. l. b., and 5t32ai007496-23 to a. n. d., and the university of virginia's global infectious diseases institute. j. m. s. is a an ithriv scholar, a program supported in part by the national center for advancing translational sciences of the nih under award numbers ul1tr003015 and kl2tr003016 disclaimer: the content is solely the responsibility of the authors and does not necessarily represent the official views of the nih division of infectious diseases and international health receptor recognition by the novel coronavirus from wuhan: an analysis based on decade-long structural studies of sars coronavirus angiotensin-converting enzyme 2 is a functional receptor for the sars coronavirus a cluster of cases of severe acute respiratory syndrome in hong kong structural basis of receptor recognition by sars-cov-2 lethal infection of k18-hace2 mice infected with severe acute respiratory syndrome coronavirus the pathogenicity of sars-cov-2 in hace2 transgenic mice. nature (epub ahead of print a mouse model of sars-cov-2 infection and pathogenesis sars-like wiv1-cov poised for human emergence pathogenesis of sars-cov-2 in transgenic mice expressing human angiotensin-converting enzyme 2 a sars-cov-2 infection model in mice demonstrates protection by neutralizing antibodies an official american thoracic society workshop report: features and measurements of experimental acute lung injury in animals genomic variations of sars-cov-2 suggest multiple outbreak sources of transmission viral concentration determination through plaque assays: using traditional and novel overlay systems acknowledgments: we would like to gratefully acknowledge the advice and assistance of angelina angelucci and young hahn at the university of virginia as well as caitlin woodson and kylene kehn-hall at george mason university. publication charges for this article were waived due to the ongoing pandemic of covid-19. this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-286543-mtjk59rp authors: al-zaman, md. sayeed title: healthcare crisis in bangladesh during the covid-19 pandemic date: 2020-08-20 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0826 sha: doc_id: 286543 cord_uid: mtjk59rp the covid-19 pandemic has had a severe impact worldwide. developed countries, such as the united states, united kingdom, italy, and spain, had their highly efficient medical infrastructure greatly stressed and suffered from high death tolls. similarly, bangladesh, a poverty-stricken south asian country, is losing its battle against the pandemic, but mainly because of its incompetent healthcare system. the casualties are escalating and public sufferings are becoming unimaginable. on this backdrop, this perspective piece discusses the healthcare crisis in bangladesh during the pandemic. this article also identifies three responsible issues for the country’s deteriorating health care: 1) poor governance and increased corruption, 2) inadequate healthcare facilities, and 3) weak public health communication. surprisingly, whereas many developed countries, including the united states, the united kingdom, italy, and spain, have suffered greatly during the covid-19 pandemic even with their highly efficient healthcare systems, many less developed countries with inadequate healthcare systems are surviving the crisis more easily. 1 where is the magic? unfortunately, no magic is there. rather, behind this mysterious façade, the reality tells a different tale. this pandemic reveals the incompetent health care of many less developed countries engulfed by intense corruption, and bangladesh is one of them. as one of the world's most densely populated countries (1,115 people/km 2 ) with 21.8% of people living below the poverty line, 2 bangladesh has a healthcare system that lacks reliability, responsiveness, and empathy, and that has already been proved inadequate to deliver proper health care to the public on many occasions. 3, 4 moreover, medical facilities are concentrated in urban areas that create a healthcare divide depriving rural areas. 3, 5 amid such a situation, the covid-19 pandemic reveals many loopholes in the healthcare system that can be summarized under three themes: 1) poor governance and increased corruption, 2) inadequate healthcare facilities, and 3) weak public health communication. after detecting the first covid-19 case on 8 march, in the following 2 weeks, more than 0.63 million overseas workers entered the country without proper screening, facilitating community transmission. 6 furthermore, a few of the government's delayed decisions were found less effective that worsened the pandemic situation. for example, the government declared a general holiday for 10 days from march 23, 2020 to april 2, 2020 without restricting transportations and public movement. as a result, a crowd of more than 12 million people left dhaka immediately after the notice that expedited the community transmission. 7 moreover, none of the two existing laws, disaster management act 2012 and infectious disease (prevention, control, and elimination) act 2018, were brought into action properly to control such massive mass mobilizations and gatherings. 6 in such a situation, many private medicals around the country were either unwilling or abstained from treating covid-19 patients, and thus healthcare denial intensified. 8, 9 a survey found that 77.3% of patients receive healthcare facilities from private medicals. 6 consequently, a large share of the population suffered, and many of them died without proper medical supports. 10 improper synchronization among the responsible bodies, the ministry of health and family welfare (mohfw), the directorate general of health services, and test laboratories' executives and workers, could be a reason for such mismanagements. 11 with the deteriorating covid-19 situation in bangladesh, corruption surged. as a great initiative, prime minister sheikh hasina promptly declared 19 incentive packages of usd 12.13 billion to ameliorate people's sufferings from the pandemic. 6 but it is a matter of concern how effective these measures could be amid the intensifying nationwide corruption. whereas mismanagements were paralyzing the health sector, increased corruption worsened the situation to a greater degree. 6 the media reported 218 relief-related corruption incidents from march 10, 2020 to june 15, 2020, and most of the convicts were the government officials, public representatives, and ruling party leaders and activists. 12 they either expropriated the public's relief or counterfeited the budgets of medical equipment and health services. 6, 12 in july, a hospital owner along with a few responsible persons was convicted for trading with fake covid-19 test certificates (for more fake covid-19 test news, see https://tinyurl.com/ y23abr3o). more investigations reveal similar incidents, including the case of two doctors who threw thousands of patients' samples in the garbage and provided them imaginary results (for a few notable examples, see table 1 ). 13, 14 despite the rampant irregularities and corruptions, a government promulgation on august 5, 2020 restricted law enforcement's investigations in hospitals: specialists suggest that it would exacerbate health-related corruption. 15 healthcare preparation and capacity against covid-19 might explain the pandemic situation in bangladesh more precisely. the coronavirus testing rate in bangladesh (0.34%) is the second lowest in south asia only after afghanistan, a war-torn nation ( table 2 ). 6 its main reasons could be the limited number of testing laboratories (56 laboratories) and kits, and their uneven distributions across the country, expensive coronavirus tests in private medicals (usd 50-60/test), the fewer number of medical workers, and unregulated testing system (elites get preferences). 1, 6, 11 moreover, many testing kits were preserved by corrupt businessmen to initiate an artificial crisis. 16 recently, the mohfw imposed a fee on covid-19 test in state-run laboratories too that dramatically reduced the average per day tests. 17 thanks to the lower test rate, 1,010 people died undiagnosed with coronavirus symptoms from march 2020 to may 2020. 18 it hints about the possible discrepancies in the coronavirus's official statistics of bangladesh. medical facilities, such as beds, intensive care units, and ventilators, are far fewer than the required amount in both government and private hospitals. 19 therefore, to manage a seat in the country's finest hospitals, patients often need to have connections. also, many patients prefer to remain at home fearing maltreatment in hospitals. 10 a report reveals that 79% of patients stay at home and get treatment over the phone. 20 in bangladesh, only 3.05 physicians and 1.07 nurses serve every 10,000 people on average, 5 which is insufficient for the pandemic situation. moreover, medical workers were provided lower quality medical equipment, such as masks and personal protective equipment. consequently, many doctors got infected and some died, making the doctors' mortality rate of bangladesh highest in the world. 21 inadequate information flow and communication networks make the healthcare system more vulnerable and incompetent. because of health-related uncertainty, information scarcity, the absence of reliable information sources, and defected flow of reliable information amid the pandemic, rumors became prevalent. in such a situation, national media outlets failed to successfully deliver reliable information to a large number of audiences, letting the more personalized and internet-based media occupy the communication space. as a result, around 200 covid-19-related online rumors spread across the country from march 2020 to july 2020 (for the complete list of rumors, see http://bdfactcheck.com, a nonprofit award-winning fact-checking website of bangladesh). as a timely step, the government started detaining rumor-producers and rumorspreaders to reduce the covid-19 crisis to some degree. however, along with the perpetrators, as many human rights activists and organizations believe thanks to a few recent incidents, political dissidents and the government's critics may be suppressed. 22, 23 meanwhile, government officials are ordered "not to like, share or comment on social media posts" that criticize the government's policies. 22 these steps may breed fear among the online communities and hamper the positive health-related communication. in june 2020 after a visit, a team of chinese physicians expressed their concerns about bangladesh's disorganized health sector: this article already discussed the key selected discrepancies. 24 this situation may be controlled by taking a few steps. first, corruption in the health sector is mandatory as this will help improve the proper utilization of allocated resources. second, more tests should be conducted to identify the infected persons to provide them better treatment. third, hospitals should be well equipped with updated and efficient medical supplies such as oxygen and medications to provide supportive treatment for covid-19. fourth, doctors and other medical workers must be protected from infection. moreover, infected doctors and nurses could be super-spreaders of the virus. fifth, thanks to higher population density and lower health awareness, social distancing in public spaces is virtually impossible in bangladesh. therefore, some sort of strict regulations for such spaces may be imposed. sixth, healthy information flow is a must for the current covid-19 situation in bangladesh to reduce the health-related confusions and uncertainties. battling the covid-19 pandemic without organized strategies and an effective healthcare system would be like an attempt to kill a lion with bare hands. received july 9, 2020. accepted for publication august 14, 2020. published online august 20, 2020. the proposed expenditure was usd 20.70 million that was at least 10 times higher than the actual expenditure. also, the products were of poor quality. the proposed expenditure was usd 1.18 million, and the original expenditure was usd 9438. computer software the proposed expenditure was usd 6.49 million, and the original expenditure was usd 0.17 million. the proposed expenditure was usd 59/piece, and the market price was usd 12/piece. audio clips the proposed expenditure was usd 1.36 million, which was unevenly higher than the market price. personal protective equipment the proposed expenditure was usd 52/piece, and the original market price was less than usd 23/piece. five hundred physicians' food and living cost for 1 month why south asia's covid-19 numbers are so low (for now) bangladesh: one in five people live below poverty line diabetes fact: bangladesh perspective patient satisfaction with health services in bangladesh bangladesh. global health workforce alliance the challanges of good governance to combat covid-19 people leave dhaka with high coronavirus risk. new age coronavirus: bangladeshi private hospitals unable to treat coronavirus patients. the business standard covid-19 and healthcare denial covid-19: bangladesh hospitals forced to turn away patients institute for defence studies and analyses bangladesh arrests hospital owner over fake coronavirus results covid-19 fake certificates scam: jkg healthcare's chairman dr. sabrina held how much the permission-based investigation in hospitals would be effective. bbc news bangla coronavirus: testing crisis because of lab and kit scarcity govt imposes fees on covid-19 tests at staterun hospitals unb news, 2020. 1,010 died with coronavirus symptoms: cgs. united news of bangladesh (unb) covid-19: a reality check for bangladesh's healthcare system 79% of patients get treatment over the phone. daily prothom alo coronavirus: doctors' mortality rate in bangladesh 'highest in the world bangladesh: alarming crackdown on freedom of expression during coronavirus pandemic. article 19 bangladeshi lecturer arrested over facebook coronavirus post bangladesh's measures in covid-19 battle disappoint chinese experts acknowledgment: publication charges for this article were waived due key: cord-285490-tpsf05ca authors: solís, josé gabriel; esquivel pineda, alejandra; alberti minutti, paolo; albarrán sánchez, alejandra title: case report: rhabdomyolysis in a patient with covid-19: a proposed diagnostic-therapeutic algorithm date: 2020-07-29 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0692 sha: doc_id: 285490 cord_uid: tpsf05ca covid-19 represents the greatest health challenge of modern years. the spectrum of illness comprises respiratory and non-respiratory manifestations. we report the case of an adult man with covid-19 who presented with rhabdomyolysis as a principal extrapulmonary manifestation. our patient presented with dyspnea, fever, and muscle pain. after a comprehensive approach, the diagnosis of covid-19 and rhabdomyolysis was made. he developed acute kidney injury requiring renal replacement therapy without reversibility, despite optimal treatment. we performed a literature search for similar cases, discuss the potential mechanisms implied, and propose a diagnostic-therapeutic algorithm. introduction covid-19 represents the greatest healthcare challenge of modern years. sars-cov-2 has infected more than 13 million people and caused more than 500,000 deaths worldwide. 1 the spectrum of illness ranges from a mild respiratory infection to severe pneumonia and acute respiratory distress syndrome. there is a wide range of extrapulmonary manifestations such as renal, cardiac, and neurological. 2 we report the case of a patient with confirmed sars-cov-2 infection who presented with rhabdomyolysis as a cardinal manifestation, discuss the possible mechanisms, and propose a diagnostic-therapeutic algorithm. a 46-year-old man presented to the emergency department with a respiratory illness of 5-day evolution characterized by cough, fever, dyspnea, and generalized muscle pain. his medical history was remarkable for chronic myeloid leukemia treated with imatinib, being its last dose 3 months before hospitalization, with an optimal response. he denied recent trauma, use of drugs, or exposure to toxins. on admission, the patient was tachycardiac, tachypneic, and hypoxemic. blood pressure and temperature were normal. physical examination revealed bilateral pulmonary rales and generalized muscle pain. his chest x-ray showed bilateral and diffuse ground-glass opacities with a predominantly peripheral distribution ( figure 1a ). laboratory tests revealed grade 3 acute kidney injury (aki) with a creatinine level of 11 mg/dl (basal value 0.7 mg/dl); increased blood levels of creatine kinase (ck) (> 400,000 u/l), lactate dehydrogenase (ldh), aspartate aminotransferase, alanine aminotransferase; and electrolyte disturbances with hyperkalemia, hyperphosphatemia, hypocalcemia, and severe metabolic acidosis. also, he had lymphopenia, moderate thrombocytopenia, and elevated c-reactive protein and ferritin. his coagulation panel showed elevated fibrinogen levels and d-dimer. his urinary volume in the first 12 hours of hospitalization was 20 ml. urinalysis revealed dark urine, urine dipstick positive for hemoglobin, and a normal sediment, compatible with myoglobinuria. nasopharyngeal swab with real-time reverse-transcriptase polymerase chain reaction (rt-pcr) for sars-cov-2 was positive. real-time reverse-transcriptase pcr for influenza virus was negative. additional investigations included fourth-generation elisa for hiv, hepatitis b surface antigen, and serologic tests for hepatitis c virus, cytomegalovirus, herpes simplex, rubeola, toxoplasma, and epstein-barr virus, all of which were negative. the diagnosis of covid-19 and severe rhabdomyolysis complicated with aki was made. the patient was treated with supplemental oxygen therapy and azithromycin. treatment with intravenous solutions and sodium bicarbonate was administrated without response, persisting with anuria, and developing uremic encephalopathy. continuous renal replacement therapy was instituted with a continuous veno-venous hemodiafiltration modality. on day 5 of hospitalization, the patient developed fever and elevated procalcitonin levels. a new chest x-ray showed new infiltrates and bilateral consolidations, compatible with disease progression or bacterial superinfection ( figure 1b ). broad-spectrum antibiotic therapy was initially administered and suspended after sputum and blood cultures were negative. the patient showed progressive reduction in muscle pain, improvement of strength, decrease in muscle enzyme levels ( figure 2 ), resolution of electrolyte disorders, and stabilization of kidney function. unfortunately, he developed further respiratory impairment and died during his hospitalization. rhabdomyolysis is a life-threatening entity characterized by rapid destruction of skeletal muscle fibers, causing the release of toxic intracellular components into the bloodstream. the diagnosis of this condition requires a high index of suspicion. the most accepted diagnostic criterion is an elevation of ck greater than 1,000 u/l, considering severe rhabdomyolysis with a cutoff value of 5,000-15,000 u/l. 3 the classic clinical triad myalgia, muscle weakness and, pigmenturia is present in less than 10% of cases. in addition to increased amounts of serum ck, other biochemical abnormalities include elevated transaminases, ldh and myoglobin, myoglobinuria, and electrolyte or acid-base disturbances, such as hyperkalemia, hyperphosphatemia, hypocalcemia, hyperuricemia, and metabolic acidosis. 4 the development of rhabdomyolysis is associated with a wide variety of conditions such as trauma, drugs, toxins, autoimmune myopathies and viral infections. although viral infection is the most frequent cause in children, it is seldom reported in adults. influenza virus is the most common viral etiology of rhabdomyolysis; however, many others have been described, including enterovirus, hiv, parainfluenza, adenovirus, cytomegalovirus, coxsackievirus, epstein-barr, herpes simplex, echovirus, varicella-zoster, and dengue virus. 5 sars-cov-2 has shown a wide variability of systemic manifestations. rhabdomyolysis has been described anecdotally with just a few cases reported so far. [6] [7] [8] [9] in addition, there were two patients with rhabdomyolysis in china's first multicenter cohort that included a total of 1,099 patients. 10 rhabdomyolysis can develop either as a first manifestation or as a complication of the disease, independently of the presence of respiratory symptoms. this highlights the need of a high index of suspicion, especially during the actual pandemic. rhabdomyolysis has been previously described in infections caused by other beta-coronaviruses. it developed in up to 10% of patients with severe acute respiratory syndrome (sars) 11 and in 14% of middle east respiratory syndrome (mers). 12 an important issue is that they all received high doses of intravenous steroid and some required neuromuscular blocking agents, contributing factors that were absent in our patient. physiopathology in viral myositis is not entirely known, and the mechanism by which sars-cov-2 can cause rhabdomyolysis has not been studied. direct viral invasion of muscle tissue and toxicity mediated by cytokines or immunological cross-reactivity have been proposed. 13 in a recent study from brazil in which 10 autopsies from patients with covid-19 were performed, histological analysis of skeletal muscle showed myositis in 60% and necrotic fibers in 80% of the patients. 14 however, in patients with sars and rhabdomyolysis, muscle biopsies did not show inflammatory cell infiltration. 15 aki is the most important complication of rhabdomyolysis and occurs in 13-50% of the cases. aki can be secondary to direct tubular injury, tubular obstruction, and intrarenal vasoconstriction. 16 the frequency of aki in covid-19 is variable, ranging from 5% to 60%; however, it has a clear association with mortality. 17 sars-cov-2 can cause aki through diverse mechanisms including virus-mediated injury, cytokine storm, angiotensin ii pathway activation, dysregulation of the complement pathway, hypercoagulation, and microangiopathy, all of which could have contributed in our patient. 18 also, tubular toxicity from rhabdomyolysis has been considered before. 19 in a study of 26 autopsies, the histopathological analysis of 3 patients revealed pigment casts in the renal interstitium, which correlated with elevated levels of ck, probably related to rhabdomyolysis. 20 the management of patients with rhabdomyolysis in covid-19 is challenging. the underlying cause of muscle injury must be identified and treated, which is difficult in patients with covid-19 because there is no specific therapy. other factors that can contribute to muscle injury should be investigated, such as coinfections or drugs. in this case, although our patient was previously treated with imatinib, there was no time relationship with the development of rhabdomyolysis. fluid replacement is the keystone of rhabdomyolysis treatment. other therapies include the use of bicarbonate or mannitol. 4 electrolyte disorders and aki must be detected and treated if required. the indication of renal replacement therapy is based on patients aki and not based on the levels of ck. although there is no specific modality recommended in rhabdomyolysis, continuous veno-venous hemodiafiltration with high permeability membranes seems to be more effective. 21 an algorithm for the detection and management of rhabdomyolysis in covid-19 is proposed (figure 3) . rhabdomyolysis is a rare and probably underdiagnosed complication of sars-cov-2 infection. it should be suspected in patients with disproportionate myalgia, muscle weakness, dark urine, unexplained hyperkalemia, or metabolic acidosis. acute kidney injury can develop as a complication that has implications in treatment and mortality. we recommend measuring ck levels in covid-19 patients with suggestive clinical or analytical alterations, especially in those with aki, in which it can be a pathophysiological mechanism that requires early and aggressive treatment to prevent chronic kidney damage or death. coronavirus cases intensive care management of coronavirus disease 2019 (covid-19): challenges and recommendations beyond muscle destruction: a systematic review of rhabdomyolysis for clinical practice rhabdomyolysis: review of the literature bacterial, fungal, parasitic, and viral myositis rhabdomyolysis as potential late complication associated with covid-19 rhabdomyolysis as a presentation of 2019 novel coronavirus disease a rare presentation of coronavirus disease 2019 (covid-19) induced viral myositis with subsequent rhabdomyolysis weakness and elevated creatinine kinase as the initial presentation of coronavirus disease 2019 (covid-19) clinical characteristics of coronavirus disease 2019 in china rhabdomyolysis associated with acute renal failure in patients with severe acute respiratory syndrome clinical aspects and outcomes of 70 patients with middle east respiratory syndrome coronavirus infection: a single-center experience in saudi arabia productive infection of human skeletal muscle cells by pandemic and seasonal influenza a (h1n1) viruses pulmonary and systemic involvement of covid-19 assessed by ultrasound-guided minimally invasive autopsy rhabdomyolysis associated with probable sars rhabdomyolysis and acute kidney injury kidney disease is associated with in-hospital death of patients with covid-19 acute kidney injury in covid-19: emerging evidence of a distinct pathophysiology kidney involvement in covid-19 and rationale for extracorporeal therapies renal histopathological analysis of 26 postmortem findings of patients with covid-19 in china acute kidney injury due to rhabdomyolysis and renal replacement therapy: a critical review acknowledgments: we would like to thank all our colleagues at the hospital centro médico nacional siglo xxi, imss for the support in facing the constant struggles of this pandemic. publication charges for this article were waived due to the ongoing pandemic of covid-19.financial support: this research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.authors' addresses: josé gabriel solís, alejandra esquivel pineda, paolo alberti minutti, and alejandra albarrán sánchez, internal medicine department, instituto mexicano del seguro social (imss), centro médico nacional siglo xxi, mexico city, mexico, e-mails: gabrielsolismd@ gmail.com, esquivelpinedaalejandra@gmail.com, paolo.alberti@gmail. com, and albarranalejandra@gmail.com. this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-294624-d71za6hi authors: kapepula, paulin m.; kabengele, jimmy k.; kingombe, micheline; van bambeke, françoise; tulkens, paul m.; sadiki kishabongo, antoine; decloedt, eric; zumla, adam; tiberi, simon; suleman, fatima; tshilolo, léon; muyembe-tamfum, jean-jacques; zumla, alimuddin; nachega, jean b. title: artemisia spp. derivatives for covid-19 treatment: anecdotal use, political hype, treatment potential, challenges, and road map to randomized clinical trials date: 2020-07-23 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0820 sha: doc_id: 294624 cord_uid: d71za6hi the world is currently facing a novel covid-19 pandemic caused by sars-cov-2 that, as of july 12, 2020, has caused a reported 12,322,395 cases and 556,335 deaths. to date, only two treatments, remdesivir and dexamethasone, have demonstrated clinical efficacy through randomized controlled trials (rcts) in seriously ill patients. the search for new or repurposed drugs for treatment of covid-19 continues. we have witnessed anecdotal use of herbal medicines, including artemisia spp. extracts, in low-income countries, and exaggerated claims of their efficacies that are not evidence based, with subsequent political controversy. these events highlight the urgent need for further research on herbal compounds to evaluate efficacy through rcts, and, when efficacious compounds are identified, to establish the active ingredients, develop formulations and dosing, and define pharmacokinetics, toxicology, and safety to enable drug development. derivatives from the herb artemisia annua have been used as traditional medicine over centuries for the treatment of fevers, malaria, and respiratory tract infections. we review the bioactive compounds, pharmacological and immunological effects, and traditional uses for artemisia spp. derivatives, and discuss the challenges and controversies surrounding current efforts and the scientific road map to advance them to prevent or treat covid-19. the unprecedented covid-19 pandemic caused by the novel zoonotic pathogen of humans, sars-cov-2, has, as of july 12, 2020, caused 556,335 deaths of 12,322,395 confirmed cases reported by the who. 1 although covid-19 commonly presents as a severe respiratory tract illness, it causes multisystem disease, and deaths have been attributed to cytokine storm, acute respiratory distress syndrome (ards), and excessive aberrant immunological responses. of several recent and ongoing treatment intervention trials, only two randomized controlled trials have to date demonstrated benefits of specific therapies. one study indicated that hospitalized covid-19 patients who received remdesivir had a 31% faster time to recovery than those who received placebo. 2 another trial reported that dexamethasone reduced mortality by one-third in seriously ill patients requiring respiratory support. 3 of note, dexamethasone was previously shown to be effective in the treatment of ards. 4 among other drugs initially considered of promise, trials of chloroquine or hydroxycholoroquine plus lopinavir/ritonavir with or without azithromycin have shown no reduction in mortality in hospitalized patients. 5 despite limited success in finding effective treatments 7 months after the first appearance of sars-cov-2 as a new human pathogen, the desperate quest for new and repurposed drugs to reduce the morbidity and mortality of covid-19 continues. the anecdotal use of artemisia spp. extracts for covid-19 treatment in low-income countries has led to exaggerated and unproven claims of its efficacy in the absence of a scientific basis or results from clinical trials. this highlights the urgent need for further research on herbal compounds to evaluate efficacy through controlled trials, and for efficacious compounds, to establish the active ingredients, develop formulations and dosing, and define pharmacokinetics, toxicology, and safety to enable drug development. we discuss the bioactive compounds, pharmacological and immunological effects, and traditional uses for artemisia spp. derivatives, and discuss the challenges and controversies surrounding current efforts to advance them for potential use to prevent or treat covid-19. traditional herb-and plant-derived medicinal products are being used and trialed for the treatment of covid-19 in china. 6, 7 among many, derivatives from the herb artemisia annua (figure 1 ) have been used as traditional medicine over centuries for the treatment of fevers, malaria, and respiratory tract infections. 8 the "sweet wormwood" plant contains artemisinin (figure 2 ), a medicine developed during the cultural revolution in china. the 2015 nobel prize in physiology or medicine was awarded to professor youyou tu for her key contributions to its discovery. 9 the who recommends artemisinin-based combination therapies (acts) as first-line treatments for uncomplicated plasmodium falciparum malaria. 10 results of a small clinical study from china of artesunate, a hemi-synthetic derivative of artemisinin (figure 2 ), for the treatment of covid-19 reported that artesunate was associated with shorter duration of covid-19 symptoms (3.3 ± 1.9 versus 4.8 ±2.2 days) and hospital stays (16.6 ±3.7 versus 18.0 ±4.0) than standard of care. 8 table 1 shows the concentrations of artemisinin and artesunate that can be achieved in the plasma of patients receiving conventional doses of these drugs, and the concentrations needed to inhibit the replication of other viruses or the inflammatory response in vitro. these data suggest that artesunate may offer both antiviral and anti-inflammatory effects at clinically achievable concentrations. 8, 11, 12 in april 2020, an herbal tonic derived from a. annua extracts by the madagascar institute of applied research and branded "covid-organics" was launched. covid-organics has been promoted as a cure for covid-19. however, reliable pharmacological and efficacy data are lacking, and there is concern that its widespread use for covid-19 could result in reduced access to effective medicines as well as possible selection of p. falciparum resistance to acts by exposing patients to suboptimal concentrations of artemisinin when malaria cases are misdiagnosed as covid-19. of note, the content of artemisinin in a. annua is about 1%, which means that 50 g of plant material is needed to obtain the equivalent of a 500-mg therapeutic dose of artemisinin, if considering a 100% extraction. 13 furthermore, the typical concentration of artemisinin in infusions is around 50 mg/l; 10 l must be ingested to absorb the antimalarial therapeutic dose, which is not feasible. therefore, to avoid the promotion of unproven remedies in this climate of uncertainty and fear, it is important that research into traditional medicinal plants and their derivatives be conducted properly. artemisia annua is an annual herbaceous plant of the asteraceae family native to asia and eastern europe ( figure 1 ). 14 as a. annua is the source for leading whoapproved antimalarials, seed varieties have been adapted by breeding for lower latitudes, and cultivation has been successfully achieved in many tropical countries. 15 artemisia annua is a source of many biologically active compounds, 16, 17 with more than 220 compounds isolated and identified, 18 including at least 28 monoterpenes, 30 sesquiterpenes, 12 triterpenoids and steroids, 36 flavonoids, seven coumarins, and four aromatic and nine aliphatic compounds. 15 it naturally produces and stores artemisinin in the glandular trichomes on its leaves, stems, and flowers. 19 sesquiterpenes, caryophyllene oxide, caryophyllene, farnesene, and germacrene d are the most abundant chemicals identified in the essential oil of the fruits. 20 artemisinin is a sesquiterpene lactone, containing an unusual endoperoxide group (figure 2 ) which is believed to be responsible for its antimalarial activity. artemisia annua extracts are said to contain antiinflammatory, antioxidant, and antimicrobial substances, and to show antiviral activity. [21] [22] [23] the flavonoids casticin and chrysosplenol d, extracted from a. annua, suppressed the expression of inflammatory mediators through the regulation of nf-κb and c-jun in a murine macrophage cell line, suggesting that these components might be useful in the treatment of inflammatory and infectious disorders. 24 the water-soluble fraction of a. annua, after the extraction of artemisinin, was shown to regulate the expression of proinflammatory cytokines, matrix metalloproteinases, and nf-κb; promote cell cycle arrest; drive reactive oxygen species production; and induce bak or bax-dependent or independent apoptosis. 8 artemisia annua extracts significantly inhibited cytopathy caused by sars-cov strain bj001 25 and showed activity against sars-cov-2 in vero-e6 cell-based cytopathic effect screening. 26 since the beginning of the covid-19 pandemic, formulations of a. annua have been used in africa and china for covid-19 prevention and treatment. in the drc, herbal formulations have been used for the prevention and treatment of covid-19 by fumigation, infusion, or decoction. 27 it is important to emphasize that there are no controlled data supporting the use of any of these, and their efficacy for covid-19 is unknown. arguably, natural product research is only relevant to the development of new drugs as a first step to identifying specific molecules with activity. teas cannot function as drugs meeting international standards, as their components are unknown and not standardized. 28 advancing traditional medicines will require identification of active components of plant extracts, methods to yield purified compounds, and determination of compound pharmacology including studies of biological activity, bioavailability, absorption, distribution, metabolism, excretion, and toxicity properties of each molecule. 28 for evaluation of the antiviral effects of herbal formulations, the screening system should meet all requirements of any good assay, including validity, lack of ambiguity, accuracy, reproducibility, simplicity, and reasonable cost. because these requirements are better met by in vitro screening, in vitro bioassays must be used to guide the isolation of active compounds from plant extracts. the antiviral activities of the pure compounds must then be confirmed at a later stage by in vivo assays in appropriate animal models. 23 the who acknowledges that the quantity and quality of safety and efficacy data on traditional medicines are far from sufficient to meet the criteria needed to support their use. the reasons for the lack of research data include inadequate healthcare policies and a lack of accepted research methodology for evaluating traditional medicines. 8, 26, 29 at a time when countries are consumed by their own national interests and agendas, the world is looking to natural products to provide readily available, affordable treatments. a cure or treatment for covid-19 derived from locally grown herbs and plants remains a viable option for some countries and communities. however, to develop drugs rather than crude preparations of herbs and plants, the specific pharmacologically active components need to be isolated, verified through proper pharmacological evaluation, and then possibly optimized through modern (hemi) synthesis strategies before being developed according to rigorous international guidelines for drug development. however, repurposing of available plant-based drugs, for example, artesunate, offers a potential time-and cost-saving approach. indeed, investigators from saudi arabia have registered a placebo-controlled trial (www.clinicaltrials.gov identifier: nct04387240) to evaluate the efficacy of artesunate in adults with mild symptoms of covid-19. as the world desperately searches for new treatments to reduce rates of severe morbidity and mortality from covid-19, the promotion of new drug discovery building on extracts from traditional medicinal plants should be encouraged. the anecdotal and unproven use of a. annua for covid-19 following claims from politicians and others in low-income countries highlights the need for hard data to establish the active ingredients; develop formulations and dosing; define the pharmacokinetics, toxicology, and safety; and evaluate efficacy through controlled trials. coronavirus disease (covid-19) situation report 166 nih clinical trial shows remdesivir accelerates recovery from advanced covid-19. press release low-cost dexamethasone reduces death by up to one third in hospitalised patients with severe respiratory complications of covid-19. recovery trial press release dexamethasone treatment for the acute respiratory distress syndrome: a multicentre, randomised controlled trial no clinical benefit from use of hydroxychloroquine in hospitalised patients with covid-19. press release pay attention to situation of sars-cov-2 and tcm advantages in treatment of novel coronavirus infection traditional chinese medicine for covid-19 treatment anti-malarial drug, artemisinin and its derivatives for the treatment of respiratory diseases the effect of malaria control on plasmodium falciparum in africa between guidelines for the treatment of malaria the antiviral activities of artemisinin and artesunate artemisinin inhibits the replication of flaviviruses by promoting the type i interferon production the content of artemisinin in the artemisia annua tea infusion artemisia annua as a traditional herbal antimalarial flavonoids from artemisia annua l. as antioxidants and their potential synergism with artemisinin against malaria and cancer secondary metabolites of artemisia annua and their biological activity chromatographic fingerprint artemisinin chemical research dried leaf artemisia annua improves bioavailability of artemisinin via cytochrome p450 inhibition and enhances artemisinin efficacy downstream essential oil of artemisia annua l.: an extraordinary component with numerous antimicrobial properties. evid based complement alternat med anti-inflammatory, antioxidant and antimicrobial effects of artemisinin extracts from artemisia annua l antiviral evaluation of herbal drugs. quality control and evaluation of herbal drugs occurrence of some antiviral sterols in artemisia annua flavonoids casticin and chrysosplenol d from artemisia annua l. inhibit inflammation in vitro and in vivo identification of natural compounds with antiviral activities against sars-associated coronavirus a review on identification of antiviral potential medicinal plant compounds against with covid-19 coronavirus (covid-19): a protocol for prevention and treatment (covalyse ® ) the complexity of medicinal plants: the traditional artemisia annua formulation, current status and future perspectives who supports scientifically-proven traditional medicine authors' addresses: paulin m. kapepula and jimmy k. kabengele, key: cord-278283-uoy54dds authors: chen, hualiang; yao, linong; zhang, lingling; zhang, xuan; lu, qiaoyi; yu, kegen; ruan, wei title: malaria in zhejiang province, china, from 2005 to 2014 date: 2015-08-05 journal: am j trop med hyg doi: 10.4269/ajtmh.15-0080 sha: doc_id: 278283 cord_uid: uoy54dds to summarize the changing epidemiological characteristics of malaria in zhejiang province, china, we collected data on malaria from the chinese notifiable disease reporting system (ndrs) and analyzed them. a total of 2,738 malaria cases were identified in zhejiang province from 2005 to 2014, of which 2,018 were male and 720 were female. notably, only 7% of malaria cases were indigenous and the other cases were all imported. the number of malaria cases increased from 2005 to 2007, peaked in 2007, and then decreased from 2007 to 2011. there were no indigenous cases from 2012 to 2014. of all cases, 68% of cases contracted plasmodium vivax, 27% of cases contracted p. falciparum, and two cases contracted p. malariae. about 88% of malaria cases during 2005–2011 occurred yearly between may and october, but the number of malaria cases in different months during 2012–2014 was similar. the median age was 33 years, and 1,892 cases occurred in persons aged 20–50 years. the proportion of businessmen increased and the proportion of migrant laborers decreased in recent years. the median time from illness onset to confirmation of malaria cases was 5 days and it decreased from 2005 to 2014. some epidemiological characteristics of malaria have changed, and businessmen are the emphases to surveillance in every month. despite widespread elimination and control efforts during the 20th century, malaria continues to be the most important parasitic disease known to humankind. according to the world health organization (who) estimate in 2014, there were 3.2 billion people at risk of being infected with malaria and developing disease; 198 million cases of malaria and 584,000 deaths occurred globally in 2013. 1 over the span of the last century, almost half of the world's countries have successfully eliminated malaria. 2 although great success has been achieved since the launch of national malaria control program in 1955, malaria remains a serious public health problem in china. [3] [4] [5] the chinese notifiable disease reporting system (ndrs), which was initiated in the 1950s, is the fundamental communicable disease surveillance system in china. after the outbreak of severe acute respiratory syndrome (sars) in 2003, the chinese government strengthened the construction of public health information system. on january 1, 2004, the real-time notifiable infectious disease reporting system was put into use nationwide, realizing the timely online monitoring of individual cases, which marks a leap in the surveillance of communicable diseases in china. zhejiang province is located in southeastern china, adjacent to anhui province, where malaria is endemic. malaria incidence decreased in recent years in zhejiang province, but there were hundreds of imported malaria cases every year and epidemiological characteristics changed. 6, 7 this stimulated us to analyze the updated epidemiological characteristics of malaria cases in recent years. case definition. indigenous malaria was defined as any case infected within the province where it was diagnosed; in contrast, imported malaria was defined as a malaria case whose origin could be traced to an area of transmission outside the province where the diagnosis of malaria was made. 8, 9 data collection. daily disease surveillance data on malaria from 2005 to 2014 were obtained from the ndrs. information of cases included gender, age, occupation, residential address, type of disease, date of onset, and date of confirmation. data analysis. data were analyzed using the statistical package for the social sciences (spss v19; spss, chicago, il). categorical variables were summarized by frequencies and numerical variables were summarized by means with standard deviations if normally distributed and medians, interquartile ranges (iqrs), and ranges if not normally distributed. ethical approval. experimental research reported in this study has been performed with the approval of the ethics committee of zhejiang provincial center for disease control and prevention (zhejiang cdc). human research was carried out in compliance with the helsinki declaration. all participants provide their written informed consent to participate in this study. a total of 2,738 malaria cases were identified in zhejiang province from 2005 to 2014, of which 2,018 were male and 720 were female. of note, only 7% (183/2,738) of malaria cases were indigenous and the other cases were all imported. as shown in figure 1 of all cases, 68% (1,865/2,738) of reported cases contracted plasmodium vivax, 27% (731/2,738) of reported cases contracted p. falciparum, and two cases contracted p. malariae, which were imported from nigeria and libya, respectively ( table 1) . because of the lack of diagnostic test in some counties before 2009, 140 cases were unclassified. malaria cases were reported in every month; the majority (88%) of malaria cases during 2005-2011 occurred yearly between may and october ( figure 2 ). however, the number of malaria cases of different months during 2012-2014 was similar. the median age of reported malaria cases was 33 (range: 1-84 years) and 1,892 (69%) cases occurred in persons aged 20-50 years ( figure 3 ). migrant laborers, farmers, businessmen, *address correspondence to wei ruan, zhejiang provincial center for disease control and prevention, binsheng road 3399, hangzhou 310051, china. e-mail: zjcdcrw@126.com and urban workers constituted 72% (1,965/2,738) of malaria cases ( figure 4) . moreover, the proportion of businessmen increased and the proportion of migrant laborers decreased in recent years. malaria cases were reported in all 11 cities of zhejiang province from 2005 to 2014. however, malaria cases from ningbo, jinhua, taizhou, wenzhou, and hangzhou accounted for 79% of cases, and these five cities reported 582, 494, 425, 345, and 308 malaria cases, respectively ( figure 5 ). from 2005 to 2009, no information about the origins of imported cases was investigated, but these detailed data were collected from 2010 to 2014. among 808 imported cases, 626 cases were from africa, 117 from asia, 59 from chinese provinces, and only six cases from countries of other continents (table 2) . nigeria, ghana, equatorial guinea, angola, congo, and cameron were the dominant origins of imported cases. anhui province where malaria was prevalent was the main origin of imported cases within china. the median time from illness onset to confirmation of all cases was 5 days (iqrs: 3-8 days; range: 0-332 days). in addition, the median time from illness onset to confirmation decreased from 2005 to 2014 as shown in table 3 . although the elimination of malaria has helped to shrink the global malaria map, this has unveiled a more complex pattern of malaria epidemiology globally. 2 imported cases of malaria into the nonendemic regions are being increasingly recognized as a new public health challenge. 10 in our study, the majority of malaria cases from 2005 to 2011 were imported and all cases from 2012 to 2014 were imported indicating that the control of imported cases was of vital importance for the elimination of malaria in zhejiang province. to eliminate malaria in zhejiang province, plan on elimination of malaria was made by provincial health department, provincial development and reform commission, provincial education department, provincial science and technology department, provincial economic and information commission, provincial public security department, provincial finance department, provincial commerce department, provincial entry-exit inspection and quarantine bureau, provincial radio, film and television (tv) bureau, and provincial tourist administration in 2010. according to the plan, many departments collaborated with each other to eliminate malaria in zhejiang province and comprehensive measures were conducted. first, the control and management of infection source were enhanced. screening of malaria among patients with high fever was conducted in province, city, county, and town levels medical institutions. cases were reported, treated, and managed soon after the confirmation. second, control and prevention of mosquito were enhanced. the patriotic public health campaign was carried and breeding places of mosquito were removed. moreover, residual spray was used to reduce density of mosquito and bed nets were used to prevent mosquito bites. third, health education was enhanced through newspapers, tv, radio, and internet. we also collaborate with entry-exit department and travel bureau to 2005 218 15 0 16 249 2006 341 27 0 44 412 2007 520 34 0 44 598 2008 381 35 0 29 445 2009 172 46 0 3 221 2010 67 60 0 4 131 2011 41 83 0 0 124 2012 46 95 0 0 141 2013 46 156 0 0 202 2014 33 180 2 0 215 total 1,865 731 2 140 2,738 educate methods or prevent malaria among individuals who went to or came from malaria-endemic areas. finally, prevention and control of malaria among immigrants was enhanced. entry-exit departments was in charge of screening of malaria cases among fever patients and the information on patients were reported to medical departments. public security departments helped to trace malaria patients among immigrants. nevertheless, mosquito density was high in malaria areas and businessmen were inevitably bit by mosquito. as a result, decades of malaria cases were imported in zhejiang province. this informed that elimination of malaria in endemic areas would contribute to the reduction of imported cases in nonendemic areas. in our study, all cases contracted p. vivax or p. falciparum except in two cases where p. malariae caused malaria. the reasons might be that the majority of cases were imported and most of the imported cases came from africa. if the majority of the cases were domestic, there might be more vivax cases. furthermore, misdiagnosis might exist because of poor diagnostic capacity of county-level centers for diseases control and prevention. we noted that the majority of cases were p. vivax infection, which was similar to studies from qatar, kuwait, kingdom of bahrain, and the united arab emirates. [11] [12] [13] [14] although p. vivax malaria is commonly considered nonsevere and has been historically termed as "benign tertian malaria," new published studies showed that cases of p. vivax infection often resulted in increased hospitalizations, severe disease, and death than previously expected. 14-17 these informed us that p. vivax malaria was the greatest cause of malaria morbidity in zhejiang province. the majority of malaria cases during 2005-2011 occurred yearly between may and october, but the number of malaria cases in different months during 2012-2014 was similar. the reasons for the temporal pattern of 2005-2011 may include higher mosquito density and more outdoor activities between may and october. therefore, people had more chances to be bitten by mosquito, increasing the number of malaria cases. all malaria cases were imported from 2012 to 2014, the number of cases was mostly related to the frequency of population migration. the age and gender distribution suggested the predominance of male cases aged 20-50 years in zhejiang province from 2005 to 2014. the reasons might include that a large number of nonimmune adults aged 20-50 years such as travelers and laborers moving from low-transmission to hightransmission areas and that men have a higher occupational risk if they work in mines, fields, or forests at peak biting times, or migrate to malaria-endemic areas for work. 18 of note, the proportion of businessmen increased and the proportion of migrant laborers decreased in recent years. this might be due to trade between malaria-endemic areas and zhejiang province was improved in recent years. although malaria cases were reported in all 11 cities of zhejiang province from 2005 to 2014, the majority of cases were reported in ningbo, jinhua, taizhou, wenzhou, and hangzhou. this may be associated with environmental and high frequency of travel and trade in these cities. the growth of rice in these cites provided breeding places for anopheles sinensis, which increased the transmission probability of malaria in these areas. some areas of these five cities were infection focus of malaria in the past. furthermore, the number of imported cases was relative to the frequency of travel and trade to malaria-endemic areas. the frequency of travel and trade of the five cities was significantly higher than that in other areas. early treatment of malaria with an appropriate antimalarial medication is the most important factor in limiting progression to severe or complicated disease, and delayed diagnosis predicted fatal outcome and severe course of falciparum malaria. 19 in our study, the median time from illness onset to confirmation of all cases was 5 days. fortunately, the median time from illness onset to confirmation decreased from 2005 to 2014. this may be related to health education to international traveler, good access to health care, and scaling-up diagnostic testing. in summary, the number of malaria cases decreased in recent years, and no indigenous cases were reported in zhejiang province from 2012 to 2014. some epidemiological characteristics of malaria changed in recent years, the risk of imported malaria was similar among different months, and the businessmen accounted for the highest proportion. these results informed that imported cases were the emphases for the control and prevention of malaria, and measures should be taken in all months in zhejiang province. measures to better intercept imported cases should include health education and preventive medication among travelers to malaria-endemic areas, the screening of malaria among travelers with high fever, and collaborating with the neighboring countries. in addition, other comprehensive measures, such as early case detection and prompt treatment, residual spraying, usage of bed nets, environmental and antilarval management, and monitoring of drug resistance, should also be conducted to achieve the elimination of malaria in zhejiang province. 2 the origins of imported cases from 2010 to 2014 2010 2011 2012 2013 2014 total africa nigeria 10 19 13 33 49 124 ghana 3 4 14 41 13 75 equatorial guinea 2 1 2 2 3 2 2 1 9 7 8 angola 14 15 9 18 20 76 congo 2 10 5 12 15 44 cameron 2 6 1 11 23 43 guinea 1 5 2 3 4 1 5 uganda 4 2 2 3 2 1 3 gabon 1 0 4 this is an open-access article distributed under the terms of the creative commons attribution license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. world health organization shrinking the malaria map: progress and prospects progress in malaria control in china molecular and morphological studies on the anopheles minimus group of mosquitoes in southern china: taxonomic review, distribution and malaria vector status malaria situation in the people's republic of china in 2005 incidence trend of malaria in taizhou municipality in zhejiang province measuring malaria endemicity from intense to interrupted transmission impact of imported malaria on the burden of disease in northeastern venezuela the changing epidemiology of malaria elimination: new strategies for new challenges epidemiology of imported malaria in qatar imported malaria in kuwait (1985-2000) status of malaria in the kingdom of bahrain: a 10-year review epidemiological and clinical characteristics of imported malaria in the united arab emirates demographic risk factors for severe and fatal vivax and falciparum malaria among hospital admissions in northeastern indonesian papua malaria diagnosis and hospitalization trends vivax malaria: neglected and not benign women and malaria-special risks and appropriate control strategy imported malaria in poland 2003 to 2011: implications of different travel patterns key: cord-312841-g948h572 authors: kalkeri, raj; goebel, scott; sharma, guru dutt title: sars-cov-2 shedding from asymptomatic patients: contribution of potential extrapulmonary tissue reservoirs date: 2020-05-13 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0279 sha: doc_id: 312841 cord_uid: g948h572 the ongoing pandemic covid-19, caused by sars-cov-2, has already resulted in more than 3 million cases and more than 200,000 deaths globally. significant clinical presentations of covid-19 include respiratory symptoms and pneumonia. in a minority of patients, extrapulmonary organs (central nervous system, eyes, heart, and gut) are affected, with detection of viral rna in bodily secretions (stool, tears, and saliva). infection of such extrapulmonary organs may serve as a reservoir for sars-cov-2, representing a potential source of viral shedding after the cessation of respiratory symptoms in recovered patients or in asymptomatic individuals. it is extremely important to understand this phenomenon, as individuals with intermittent virus shedding could be falsely identified as reinfected and may benefit from ongoing antiviral treatment. the potential of sars-cov-2 infection to rapidly disseminate and infect extrapulmonary organs is likely mediated through the nonstructural and accessory proteins of sars-cov-2, which act as ligands for host cells, and through evasion of host immune responses. the focus of this perspective is the extrapulmonary tissues affected by sars-cov-2 and the potential implications of their involvement for disease pathogenesis and the development of medical countermeasures. the current pandemic covid-19 caused by sars-cov-2 is rapidly spreading across the globe, with more than 3 million infections and more than 200,000 deaths worldwide. the receptor of sars-cov-2, angiotensin converting enzyme 2 (ace2), is expressed in the lungs, heart, kidneys, intestines, brain, eyes, and testicles. 1, 2 infection of these extrapulmonary organs (eyes, gastrointestinal tract, and brain) 3 has been reported. viral shedding in asymptomatic individuals and recovered patients after the cessation of respiratory symptoms 4, 5 has been documented. although sars-cov-2 positivity of recovered patients may be interpreted as reinfection, failure to reinfect monkeys in the laboratory setting 6 argues against the possibility of reinfection and suggests the likelihood of extrapulmonary reservoirs in the infected individuals. considering this possibility, this perspective is focused on extrapulmonary organs affected by sars-cov-2 and the implications of their involvement for disease transmission, clinical management strategies, and medical countermeasure discovery and development. sars-cov-2 and extrapulmonary tissues and organs. in addition to the primary respiratory route of infection via droplets or contact with fomites, the expression of ace2 in aqueous humor 7 and neural tissue of the retina 8 suggest a potential role of transmission via an ocular route. the ocular reservoir can harbor low viral load, even before transmission to other organs such as the throat or lungs, as 75% of tears drain into the inferior meatus of the nasal cavity and to the back of the throat. 9 red eyes, conjunctivitis, conjunctival hyperemia, chemosis, epiphora, or increased secretions are observed in a minority of patients, along with detectable sars-cov-2 rna in tears. 10, 11 although viral rna is infrequently detected (1-5%) in tears, ocular manifestations are relatively common in covid-19-positive patients (10-30%). this could be due in part to timing of sample collection, fluctuations in virus shedding, and variability in testing methods. standardized approaches for sample collection along with more sensitive testing methods may yield more robust data. additional research is needed to confirm the temporal correlation between conjunctivitis and viral shedding in covid-19 patients. the gastrointestinal tract is also affected by sars-cov-2. diarrhea and shedding of sars-cov-2 in stool are reported in the literature. 12, 13 currently, transmission through the fecaloral route is not documented. however, it remains a possibility considering the detection of sars-cov-2 rna in wastewater and municipal sewage. 14 fecal shedding also increases the risk of creating a new intermittent animal reservoir and emergence of new viral strains through recombination, which could serve as starting points of new outbreaks. neurological manifestations (headache, loss of taste and smell, dizziness, impaired consciousness, and epilepsy) are reported in some covid-19 patients. 15 sars-cov-2 rna was also detected in the cerebrospinal fluid of a patient diagnosed with covid-19 and viral encephalitis. 16 it is postulated that coronaviruses can enter the central nervous system (cns) via olfactory nerve, blood circulation, and neuronal pathways, leading to neurological abnormalities and symptoms. 17 liver, kidney, and heart abnormalities are also observed in covid-19 patients, 18, 19 and although sars-cov-2 rna is not reported in these tissues after autopsy, the detection of viral rna in the liver of the hamster model 20 suggests the infection of these organs in patients. although sars-cov-2 rna is detected in the blood (1% of patients), 3 at present, it is unknown if the virus is shed in breast milk, semen, or vaginal fluid. extrapulmonary complications in covid-19 patients include diarrhea (gastrointestinal tract), confusion (cns), hepatic, and renal injury. 21 some of these complications may also be due to compromised pulmonary function. extrapulmonary tissues affected by sars-cov-2 are listed in table 1 . currently, it is unknown if sars-cov-2 can replicate in non-respiratory tissues (eyes, liver, and cns) to produce infectious virus. however, sars virus has been shown to replicate in human kidney (hek293) and hepatic (huh7 and hepg2) 22 cell lines and detected in the liver and brain of patients. 23, 24 experimental infection of primary tissue cells with sars-cov-2 and longitudinal studies in infected patients and animal models can promote a greater understanding of the role of these tissues in the infection. ocular and cns tissues are considered immune-privileged sites. 25, 26 for other pathogens such as cytomegalovirus (cmv), zika virus, ebola virus, and other beta coronaviruses (table 2 ), these organs have been shown to serve as reservoirs, facilitating viral persistence. 27 many covid-19 patients test positive even after discharge from the hospital. 28, 29 in one report, sars-cov-2 rna was detected up to 60 days after the onset of symptoms and 36 days after complete resolution of symptoms in the patient's nasopharyngeal and/or oropharyngeal swabs. 30 another study reported undetectable viral load on days 21 and 22 after symptom onset in oropharyngeal saliva samples of a covid-19 patient, followed by viral rna detection on days 23 and 24, without any detectable virus for the next 5 days. 31 taken together, reports of prolonged incubation periods where virus is shed from asymptomatic infected persons 4 or recovered patients several days after disease symptoms with an intermittent period of shedding, 31 along with the detection of sars-cov-2 in the extrapulmonary tissues, strongly suggest the presence of extrapulmonary sars-cov-2 tissue reservoirs. these extrapulmonary virus tissue reservoirs in infected patients may also explain the highly variable incubation period associated with the onset of symptoms after an initial exposure as well as the duration of time for complete viral clearance. role of sars-cov-2 proteins in immune evasion. nonstructural proteins (nsp1, 3, and 16) and accessory proteins (orf 3a, 6, and 9b) of sars-cov-2 are thought to play a role in the evasion of host immune responses (table 3) . a recent report also predicted a potential role of sars-cov-2 nsp5 and nsp13 interfering with the host immune response. 32 considering the substantial sequence similarity of more than 80% between sars and sars-cov-2 proteins (table 3) , it is quite possible that sars-cov-2 can also escape the host immune response using similar mechanisms in nonrespiratory tissues such as the liver and kidneys. implications of sars-cov-2 infection in extrapulmonary tissues. the presence of extrapulmonary tissue reservoirs enhances the risk of organ malfunction, such as abnormal liver or kidney functions and impaired nervous system, leading to exacerbated disease complications and delayed recovery time in covid-19 patients. tissue reservoirs in immunocompromised patients are a major concern as the virus could spread to the respiratory system at an opportune time, exerting a more aggressive clinical course. reports of continued or delayed virus shedding up to 36 days after cessation of symptoms 30, 33 suggest that longer term monitoring of recovered covid-19 patients and improved virus containment strategies will be required to mitigate further community transmission. currently, the amount of virus present in the extrapulmonary reservoirs relative to the amount of virus shed, such as in aerosol droplets, is unknown. as different viral loads have been observed in various bodily fluids (saliva, tears, feces, throat, or nasal discharge), longitudinal testing of "paired samples" collected from these different sites may be needed. the proportion of asymptomatic carriers potentially shedding the virus from both pulmonary and extrapulmonary virus reservoirs is estimated to be between 17.9% 34 and 30.8%, 35 suggesting the importance of population-based screening using sensitive and robust assays. for other viral diseases such as measles and norovirus infection, viral transmission from asymptomatic carriers is well documented. 36, 37 hence, global harmonization of the sensitivity and robustness of sars-cov-2 detection kits and screening of populations at risk might ensure identification of asymptomatic carriers of infection. potential antiviral drugs against sars-cov-2 may need to demonstrate bioavailability in extrapulmonary tissue reservoirs outside of the lungs, raising concerns of adverse events. achieving efficacious levels of therapeutics in some of these tissues may be challenging because of the presence of blood-brain and blood-retina barriers. vaccine and antiviral candidates may also need to demonstrate efficacy in the prevention of tissue reservoirs, which could introduce additional stringency requirements for clinical trials. development of appropriate animal models can address some of these questions. golden syrian hamsters infected with sars-cov-2 exhibited contact transmission, weight loss, lung damage, intestinal mucosal inflammation, lymphoid acute respiratory distress syndrome eyes 39 conjunctivitis liver 40, 41 liver injury systemic circulation (blood) 3 thrombosis kidney 19, 18 renal injury brain/cns 1, 42 cns symptoms gi tract 13, 12 diarrhea cns = central nervous system; gi = gastrointestinal. sars-cov-2 shedding from the asymptomatic patients atrophy, myocardial degenerative changes, and expression of viral nucleocapsid in lungs and intestines. 20 interestingly, viral rna could be detected in extrapulmonary tissues such as the liver, heart, spleen, kidneys, brain, and salivary glands, confirming the extrapulmonary manifestation of sars-cov-2 disease. although hamsters could be a cost-effective animal model for sars-cov-2, lack of hamster-specific immunological reagents and unknown utility for testing medical countermeasures could limit their role in sars-cov-2 preclinical studies. rhesus monkeys have been successfully infected with sars-cov-2. 6 viral replication was observed in extrapulmonary tissues (gut, spinal cord, heart, skeletal muscles, and bladder). reexposure of previously infected monkeys elicited no signs of viral replication in extrapulmonary tissues, suggesting it could be a useful animal model to study sars-cov-2 tissue reservoirs and efficacy of vaccines. however, it is also important to note the importance of inoculation dose, age of animals, and route of challenge (ocular, intranasal, or oral) in the development and utility of animal models to address different research questions. several scientific questions remain to be addressed to fully understand covid-19 clinical disease progression, including potential differences in extrapulmonary tissue infections with respect to age or ethnicity. it will also be necessary to consider the kinetics and duration of viral shedding, which could be impacted by viral bio-distribution within and among different tissue reservoirs. in addition, the role of host immune responses and the expression of host factors must be considered as dynamic forces in driving genotypic or virologic differences among viral quasi-species isolated from different reservoirs. the identification of non-respiratory tissue reservoirs of sars-cov-2 suggests that further studies are needed to address implications for covid-19 disease progression, effects on extrapulmonary tissues harboring the virus, and development of optimal medical countermeasures and disease management strategies. evidence of the covid-19 virus targeting the cns: tissue distribution, 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excrete the virus for about two weeks after recovering from diarrhea: a case report sars-cov-2 in wastewater: potential health risk, but also data source neurological manifestations of hospitalized patients with covid-19 in wuhan, china: a retrospective case series study a first case of meningitis/encephalitis associated with sars-coronavirus-2 nervous system involvement after infection with covid-19 and other coronaviruses liver and kidney injuries in covid-19 and their effects on drug therapy; a letter to editor clinical characteristics of 113 deceased patients with coronavirus disease 2019: retrospective study simulation of the clinical and pathological manifestations of coronavirus disease 2019 (covid-19) in golden syrian hamster model: implications for disease pathogenesis and transmissibility clinical characteristics of coronavirus disease 2019 in china sars-associated coronavirus replication in cell lines sars-associated viral hepatitis caused by a novel coronavirus: report of three cases detection of severe acute respiratory syndrome coronavirus in the brain: potential role of the chemokine mig in pathogenesis immunologically privileged sites cns immune privilege: hiding in plain sight zika virus reservoirs: implications for transmission, future outbreaks, drug vaccine development positive rt-pcr test results in patients recovered from covid-19 clinical characteristics of 24 asymptomatic infections with covid-19 screened among close contacts in nanjing case report: viral shedding for 60 days in a woman with novel coronavirus disease (covid-19) temporal profiles of viral load in posterior oropharyngeal saliva samples and serum antibody responses during infection by sars-cov-2: an observational cohort study a sars-cov-2-human protein-protein interaction map reveals drug targets and potential drugrepurposing presumed asymptomatic carrier transmission of covid-19 estimating the asymptomatic proportion of coronavirus disease 2019 (covid-19) cases on board the diamond princess cruise ship estimation of the asymptomatic ratio of novel coronavirus infections (covid-19) estimating the asymptomatic ratio of norovirus infection during foodborne outbreaks with laboratory testing in japan transmission potential of modified measles during an outbreak rapid progression to acute respiratory distress syndrome: review of current understanding of critical illness from covid-19 infection can the coronavirus disease 2019 (covid-19) affect the eyes? a review of coronaviruses and ocular implications in humans and animals liver injury in covid-19: management and challenges liver injury during highly pathogenic human coronavirus infections the neuroinvasive potential of sars-cov2 may play a role in the respiratory failure of covid-19 patients mechanisms of host defense following severe acute respiratory syndrome-coronavirus (sars-cov) pulmonary infection of mice middle east respiratory syndrome coronavirus causes multiple organ damage and lethal disease in mice transgenic for human dipeptidyl peptidase 4 neurotropism of human coronavirus 229e axonal transport enables neuron-to-neuron propagation of human coronavirus oc43 comparison of mouse hepatitis virus strains for pathogenicity in weanling mice infected by various routes isolation of a coronavirus from kidney biopsies of endemic balkan nephropathy patients species-specific clinical characteristics of human coronavirus infection among otherwise healthy adolescents and adults severe acute respiratory syndrome coronavirus nsp1 protein suppresses host gene expression by promoting host mrna degradation severe acute respiratory syndrome coronavirus nsp1 suppresses host gene expression, including that of type i interferon, in infected cells the papain-like protease of severe acute respiratory syndrome coronavirus has deubiquitinating activity the conserved coronavirus macrodomain promotes virulence and suppresses the innate immune response during severe acute respiratory syndrome coronavirus infection attenuation and restoration of severe acute respiratory syndrome coronavirus mutant lacking 2'-o-methyltransferase activity the sars coronavirus 3a protein causes endoplasmic reticulum stress and induces ligand-independent downregulation of the type 1 interferon receptor sars coronavirus and innate immunity sars-coronavirus open reading frame-9b suppresses innate immunity by targeting mitochondria and the mavs/traf3/traf6 signalosome sars-cov-2 shedding from the asymptomatic patients key: cord-280164-dukccrjb authors: nachega, jean b.; mbala-kingebeni, placide; otshudiema, john; mobula, linda m.; preiser, wolfgang; kallay, oscar; michaels-strasser, susan; breman, joel g.; rimoin, anne w.; nsio, justus; ahuka-mundeke, steve; zumla, alimuddin; muyembe tam-fum, jean-jacques title: responding to the challenge of the dual covid-19 and ebola epidemics in the democratic republic of congo—priorities for achieving control date: 2020-06-19 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0642 sha: doc_id: 280164 cord_uid: dukccrjb as of june 11, 2020, the democratic republic of the congo (drc) has reported 4,258 covid-19 cases with 90 deaths. with other african countries, the drc faces the challenge of striking a balance between easing public health lockdown measures to curtail the spread of sars-cov-2 and minimizing both economic hardships for large sectors of the population and negative impacts on health services for other infectious and noninfectious diseases. the drc recently controlled its tenth ebola virus disease (evd) outbreak, but covid-19 and a new evd outbreak beginning on june 1, 2020 in the northwest équateur province have added an additional burden to health services. although the epidemiology and transmission of evd and covid-19 differ, leveraging the public health infrastructures and experiences from coordinating the evd response to guide the public health response to covid-19 is critical. building on the drc’s 40 years of experience with 10 previous evd outbreaks, we highlight the drc’s multi-sectoral public health approach to covid-19, which includes community-based screening, testing, contact-tracing, risk communication, community engagement, and case management. we also highlight remaining challenges and discuss the way forward for achieving control of both covid-19 and evd in the drc. the spread of covid-19 compounds the burden on health services in african countries that have experienced recurrent outbreaks of deadly zoonotic diseases in recent years. as of june 8, 2020, the who africa region has reported 135,412 covid-19 cases, with 3,236 deaths from 45 countries. 1 most african countries are facing difficult decisions as they attempt to balance efforts to limit the spread of sars-cov-2, control local outbreaks of other infectious diseases, and lessen economic hardships and food insecurity for large sectors of the population. [2] [3] [4] the democratic republic of the congo (drc) recently experienced its tenth ebola virus disease (evd) outbreak, the second largest globally after the 2014-2016 west african epidemic, which was recently brought under control. the lessons learned, coordination mechanisms developed, and public health infrastructures put in place for evd are guiding the public health response to covid-19 in the drc, although the two diseases are fundamentally different. 5 building on four decades of experience with evd, we discuss the drc's response to covid-19 and associated challenges, priorities, and innovations for disease control. early covid-19 cases in africa were mostly due to air travel of infected individuals from europe. 2, 3 the democratic republic of the congo confirmed its first case of covid-19 on march 10, 2020. two days after returning from france, an adult male with cough and fever tested positive in the capital city of kinshasa. the subsequent early index cases in kinshasa also occurred among young affluent adult travelers from europe. 3 the drc declared a state of emergency that included travel bans on march 24, and on april 6, a lockdown of the initial covid-19 hotspot, gombe, an affluent health zone in kinshasa, and other selected regions of the country, was instituted. since then, the number has increased to 4,258 covid-19 cases, with 90 deaths (case fatality rate of 2.1%) as of june 11, 2020. 1 to date, the disease has spread to 11 provinces and 54 (10% of total) health zones in the drc (figures 1 and 2) . as in other african countries, the travel bans and lockdowns have had negative socioeconomic impacts on the population, most of whom live below the poverty line. a multi-sectoral national committee to organize the covid-19 response was created following the diagnosis of the first confirmed cases ( figure 2 ) using lessons learned from the tenth evd outbreak. the committee, which includes a presidential task force that liaises with the president's office and a strategic and operational management task force comparable to that of the ebola incident management system, has support from the who, u.s. and africa cdc, world bank, and u.k. department for international development as part of the fourth strategic response plan. the committee's secretariat is made up of five sections with distinct responsibilities (supplemental figure 1 ). similar organizations have been set up for the management of the response in the different provinces under the coordination of each governor and provincial minister of health. the drc government's covid-19 task response structure was incorporated into existing health system structures for hiv, tuberculosis, malaria, and other noncommunicable diseases. the tenth evd outbreak in the drc was announced by the ministry of health on august 1, 2018, ten days after the ninth outbreak was declared over. since june 2018, approximately 300,000 people in evd-affected health zones have been immunized with the merck rvsv-ebov vaccine and another 20,000 in goma with the janssen ad26.zebov vaccine. 6 in addition, investigational drugs were provided to nearly all patients confirmed to have evd either through the monitored emergency use of unregistered investigational drugs protocol (allowing patients to receive investigational drugs under compassionate use) or as part of the pamoja tulinde maisha (palm ["together save lives" in kiswahili]) randomized controlled trial. 7 a multi-sectoral response, including standard public health measures (surveillance, contact-tracing, active case finding, infection prevention and control, risk communication, community engagement, and safe burials) coupled with community-based interventions such as cash-for-work and water and sanitation hygiene projects, as well as security, eventually controlled a complex outbreak that lasted almost 2 years. unfortunately, hope held by health officials to declare the outbreak over on april 12, 2020 vanished as a new case was confirmed on april 10, thereby resetting the countdown clock. this was the first time an ebola outbreak occurred in a conflict zone with an ongoing humanitarian emergency. numerous factors contributed to the 2-year-long evd outbreak in the eastern drc, including a fragile and fragmented health system, population displacement, movement of contacts, disenfranchisement of the community, mistrust, and ongoing armed conflicts. 8 the drc continues to face the challenge of having back-to-back evd outbreaks with limited funding for existing needs. on june 1, 2020, the drc government announced an eleventh evd outbreak occurring in the northwestéquateur province. the drc ministry of health, in close collaboration with who teams who were already on the ground in mbandaka as part of capacity building, deployed additional multidisciplinary rapid response teams from goma and kinshasa to support local teams. according to the who, as of june 9, 2020, a total of 12 evd cases (nine confirmed and three probable) including nine deaths (case fatality rate 75%) were reported in three affected health zones (wangata, mbandaka, and bikoro). overall, 85.3% (521/611) of contacts were traced, but none turned out to be a suspected evd case. also, 1,495 people, including 436 frontline health professionals and close contacts, were vaccinated using the rvsv-zebov-gp vaccine since the beginning of this outbreak. the inability to act rapidly and diagnose and isolate cases of evd was an important factor in the large-scale progression of the 2014-2016 ebola outbreak in west africa. a range of novel ebola diagnostic tools were trialed and introduced, including automated pcr machines and rapid test kits for point-of-care diagnosis. system-wide support was put in place for safely transporting samples, sourcing reagents, disposal of hazardous materials, and rapid feedback of diagnostic data into public health and clinical decision-making. although the global covid-19 pandemic presents unique challenges, several lessons from the evd outbreaks are informing the covid-19 response. 6 first, the ebola standard operating procedures (sops) have been used as a starting point to speed the development and updating of covid-19 sops. second, ebola contact follow-up approaches have been leveraged for the follow-up of covid-19 contacts, with the difference that in this case the duration of follow-up is 14 days, compared with 21 days for evd. because persons with covid-19 may be asymptomatic, contact-tracing includes the collection of respiratory samples on days 7 and 12 from all high-risk contacts of a confirmed case, regardless of symptoms. third, the follow-up of covid-19 contacts is modeled after our evd experience using contact-tracers and community health workers (chcws) at the peripheral level: health areas, neighborhoods, and villages. fourth, the evd response established mobile laboratories in target provinces. currently, the covid-19 response is planning to setup such provincial laboratories for point-of-care (poc, e.g., genexpert) covid-19 testing. finally, based on the critical importance of community engagement and feedback during the ninth and tenth evd outbreaks in the drc, a mechanism to collect feedback from communities was put in place from the beginning of the covid-19 response. community-based covid-19 screening, testing, and contact-tracing as of may 27, 2020, there were 6,389 contacts of the 2,659 confirmed cases traced, resulting in a daily contact-tracing proportion of 92% (table 1) . among reported confirmed globally, the current gold standard test for the diagnosis of sars-cov-2 infection is detection of viral rna in a sample from the respiratory tract by rt-pcr. [9] [10] [11] laboratories with skilled staff and the rt-pcr equipment to perform these tests are scarce in the drc (table 2 ); all covid-19 testing is performed at the national institute of biomedical research in kinshasa, a national referral laboratory. because of sample transport from the provinces, turnaround times are lengthy for samples collected from provinces, causing delays in diagnoses. of note, poc or near-patient solutions would be preferable. 12, 13 the genexpert platform, already in place for tb testing across africa, is an attractive option, but drawbacks include cost and limited supplies of sars-cov-2 cartridges. poc viral antigen detection is not yet sufficiently sensitive. 14 serological testing for antiviral antibodies is unavailable in the drc and is unsuitable for diagnosing active covid-19 cases. 15 treatment of moderate and severe cases of covid-19 requires hospitalization for supportive care, oxygen, and anticoagulation as per who guidelines. 16 remdesivir, which has been shown to be effective in reducing the length of hospitalization for moderately severe cases, is not yet available in the drc. 17 weak health systems in the drc, with limited intensive care beds, oxygen supply, ventilators, and trained staff, remain a key challenge in the management of covid-19, especially as case numbers rise. several hospitals were identified as reference centers for the treatment of covid-19 as part of the national plan. furthermore, building on an existing innovative tele-mentoring program developed to capacitate nurses and other frontline healthcare workers, a series of in-service covid-19 training modules covering triage, infection prevention and control, testing, maintenance of essential services, and other topics was developed. at the start of the outbreak, only 60 ventilators were available countrywide and oxygen supplies were limited, and there was minimal technical capacity to provide intensive care. a clinical protocol was developed by the case management commission with support from technical partners. bilateral and multilateral partnerships are scaling-up donations including medical and personal protective equipment to the reference hospitals, and training has been provided to clinical staff to ensure optimal care and prevention of infection of healthcare workers. 18 remaining challenges and priority solutions are listed in table 2 . as the drc decides how best to control the covid-19 pandemic, it is essential to reflect on lessons learned from past and current evd outbreaks. the drc must adapt the available infrastructure and protocols to covid-19 while embedding community needs and concerns into its response. the country must also significantly invest in its fragile health systems to ensure equity, stability, and global health security. control of the covid-19 pandemic in the drc will be possible only with efficient community screening, testing, and contact-tracing as well as behavioral modification, all of which require adequate local and national resources and enough trained and protected personnel. by addressing the challenges, the drc and other countries in africa can limit the impact of the covid-19 pandemic on the health of its already vulnerable citizens. coronavirus disease (covid-2019) situation reports the late arrival of covid-19 in africa -mitigating pan-continental spread limiting the spread of covid-19 in africa: one size mitigation strategies do not fill all countries from easing lockdowns to scaling-up community-based covid-19 screening, testing, and contact tracing in africa -shared approaches, innovations, and challenges to minimize morbidity and mortality world health organization recommendations for the covid-19 response at the national level based on lessons learned from the ebola virus disease outbreak in the democratic republic of the congo a rendomized trial of ebola virus disease therapeutics the ebola outbreak in the democratic republic of the congo: why there is no 'silver bullet laboratory testing for 2019 novel coronavirus (2019-ncov) in suspected human cases. interim guidance. who/covid-19/laboratory/2020 laboratory testing strategy recommendations for covid-19: interim guidance detection of 2019 novel coronavirus (2019-ncov) by real-time rt-pcr 13. foundation for innovative new diagnostics (find), 2020. sars-cov-2 diagnostics: performance data. available at scientific brief: advice on the use of point-of-care immunodiagnostic tests for covid-19 severe acute respiratory syndrome coronavirus 2-specific antibody responses in coronavirus disease 2019 patients clinical management of covid-19 remdesivir compassionate use of remdesivir for patients with severe covid-19 h5n1, 2020. covid-19 in the drc: dr muyembe fears large-scale contamination of healthcare workers. available at this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-285369-ktg2b9jb authors: mohamed, mouhand f. h.; al-shokri, shaikha; yousaf, zohaib; danjuma, mohammed; parambil, jessiya; mohamed, samreen; mubasher, mahmood; dauleh, mujahed m.; hasanain, bara; alkahlout, mohamed awni; abubeker, ibrahim y. title: frequency of abnormalities detected by point-of-care lung ultrasound in symptomatic covid-19 patients: systematic review and meta-analysis date: 2020-06-02 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0371 sha: doc_id: 285369 cord_uid: ktg2b9jb the covid-19 pandemic has resulted in significant morbidity, mortality, and strained healthcare systems worldwide. thus, a search for modalities that can expedite and improve the diagnosis and management of this entity is underway. recent data suggested the utility of lung ultrasound (lus) in the diagnosis of covid-19 by detecting an interstitial pattern (b-pattern). hence, we aimed to pool the proportion of various reported lung abnormalities detected by lus in symptomatic covid-19 patients. we conducted a systematic review (pubmed, medline, and embase until april 25, 2020) and a proportion meta-analysis. we included seven studies examining the role of lus in 122 covid-19 patients. the pooled proportion (pp) of b-pattern detected by lung ultrasound (us) was 0.97 (95% ci: 0.94–1.00 i(2) 0%, q 4.6). the pp of finding pleural line abnormalities was 0.70 (95% ci: 0.13–1.00 i(2) 96%, q 103.9), of pleural thickening was 0.54 (95% 0.11–0.95 i(2) 93%, q 61.1), of subpleural or pulmonary consolidation was 0.39 (95% ci: 0.21–0.58 i(2) 72%, q 17.8), and of pleural effusion was 0.14 (95% ci: 0.00–0.37 i(2) 93%, q 27.3). our meta-analysis revealed that almost all sars-cov-2–infected patients have abnormal lung us. the most common abnormality is interstitial involvement depicted as b-pattern. the finding from our review highlights the potential role of this modality in the triage, diagnosis, and follow-up of covid-19 patients. a sizable diagnostic accuracy study comparing lus, computed tomography scan, and covid-19–specific tests is warranted to further test this finding and to delineate the diagnostic and prognostic yield of each of these modalities. the covid-19 pandemic has put enormous pressure on healthcare systems all around the globe. 1 since its advent, there has been a quest for aiding symptoms, signs, laboratory, and imaging modalities that assist in triaging and prioritizing patients for testing and isolation. this is of exceptional value when dealing with atypical presentations of covid-19 or when working in resource-depleted settings. computed tomography (ct) scan of the chest has emerged as the imaging modality of choice in the diagnosis of this disease. 2 the main findings are that of interstitial involvement. 2 however, the difficulties associated with the transfer of infectious and potentially sick patients, disinfecting the machine, ionizing radiation exposure, immediate availability concerns, and the need for lesions follow-up made it less appealing as a triaging tool for clinicians, especially those working in the front line. 3, 4 lung ultrasound (lus) or point-of-care ultrasound (pocus) has gained popularity in the triage, diagnosis, and follow-up of various lung lesions and is considered an alternative to chest x-ray (cxr) and ct scan. 5, 6 it is used routinely by critical care specialists, emergency physicians, and, recently, internists. 7 it demonstrated a better diagnostic yield than a cxr in the early diagnosis of h1n1 2009 pandemic viral pneumonia. 8 recent data suggested the potential utility of lung us in the diagnosis of covid-19, depicting interstitial phenomenon as evident by b-lines. 4, 9, 10 lung us is a tempting modality, given the ease of use, availability in many emergency departments, relative ease of disinfection, and potential role in the follow-up. 11, 12 thus, we aimed to explore the potential utility of this modality by systematically reviewing the literature and describing the frequency of b-pattern detected by lung us. in addition, we describe the frequency of other lung abnormalities detected by this modality. this is a systematic review and a meta-analysis keeping with prisma guidance. 13 study eligibility criteria. we included case series and observational studies guided by the following inclusion criteria: search strategy. we performed a comprehensive literature search of pubmed, medline, and embase since their inception, with no limitations. the search was updated on april 22, 2020. example of a database search strategy is as follows: ("lus" or "point of care ultrasound" or pocus or ultrasound or "ultrasound"/exp/mj or "point of care ultrasound"/ exp/mj) and ("covid-19" or (sars and cov and 2) or "covid 19" or "covid 19"/exp/mj or "covid-19"/exp/mj). besides, we performed a manual reference search and freetext search on google and google scholar to further add to the search comprehensiveness. screening and data extraction. initial title and abstract screening were conducted by two reviewers (m. f. h. m. and s. a.). potentially eligible articles were imported for full-text review and assessed for inclusion. a third reviewer (i. y. a.) adjudicated discrepancies guided by the protocol whenever disagreement arose that was not settled by discussion. we extracted data using an excel sheet. examples of data collected are author, year of publication, study type, type of probe, frequency of various lung abnormalities, and the severity of the illness. outcome. we performed a scoping search and reviewed some of the constituent studies to identify the commonly reported outcomes. 10 this was done at the design phase before proceeding with the actual search. we opted to summarize the pooled proportion (pp) of various lung abnormalities detected by lus. these abnormalities are as follows: 1. b-pattern (positive if three or more b-lines were present in a lung region, confluent b-lines, or white lung appearance). 2. pleural line abnormalities: some of the constituent studies did not use a uniform description when referring to pleural changes. hence, we pooled the higher frequency of either pleural thickening or pleural line irregularities. 3. pleural thickening was solely pooled. 4. consolidations: the reporting of consolidation was incomplete. so, we chose to combine subpleural and pulmonary consolidations and considered the higher frequency of the two. study quality and risk of bias assessment. we used the qudas 2 quality assessment score to judge the quality of the included studies in our review. 14 statistical analysis. we used a proportion meta-analysis to summarize or pool the frequency of various findings on lung us (based on our scoping review, we concluded that the sensitivity, specificity, and diagnostic accuracy could not be computed from the constituent studies). we used the random-effects model (double arcsine transformation and back transformation). i 2 was used to adjudicate heterogeneity (> 50% was considered marked). the analysis was conducted via metaxl version 5.3 (epigear international, sunrise beach, queensland, australia). our initial database search has retrieved 107 potentially relevant articles. finally, after duplicate removal and full-text screening (all articles excluded were duplicates, reviews, opinions, or case reports), seven articles were included in our quantitative synthesis ( figure 1 flow diagram). 9,10,12,15-18 a total of six observational studies and a case series describing a total of 122 patients constituted our review population ( table 1 presents a summary of the included studies). lung zones examined. five studies reported on the number of lung zones examined. twelve zones were examined in four studies, whereas in one study, 10 zones were examined ( table 1) . the proportion of b-pattern. all seven studies reported on the frequency of b-pattern. the pp of b-pattern in the review population is 0.97 (95% ci: 0.94-1.00 i 2 0%, q 4.6). the results were homogenous and consistent among studies (figure 2) . the proportion of pleural line abnormalities. five studies reported the frequency of pleural line abnormalities. the frequency of these abnormalities ranged between 10% and 100% ( table 1 ). the pp is 0.70 (95% ci: 0.13-1.00 i 2 96%, q 103.9) (figure 3 ). one study did not report the exact frequency but stated that most patients had pleural thickening; hence, pooling this additional study may have led to a slight increase in the pp. the proportion of pleural thickening. five studies reported the frequency of pleural thickening. the pp of pleural thickening is 0.54 (95% 0.11-0.95 i 2 93%, q 61.1). this was less than the pp of pleural line abnormalities because of one study that reported a 35% frequency of pleural thickening while reporting a 100% frequency of pleural line irregularities. there was marked heterogeneity evident by the high i 2 ( figure 2 ). as explained earlier, the study by peng et al. 10 reported that pleural thickening has occurred in most patients without specifying the frequency, hence excluded from the computation of this pp. the proportion of consolidations. six studies reported on the frequency of subpleural or pulmonary consolidations detected by lung us. the frequency ranged from 20% to 75%. the pp is 0.39 (95% ci: 0.21-0.58 i 2 72%, q 17.8) (figure 2) . i 2 indicated a marked heterogeneity. the proportion of pleural effusion. five studies reported the frequency of pleural effusion detected by lung us. it ranged from 0% to 12.5% in four studies, whereas in one study, 60% of the patients had pleural effusion. the pp of this finding was the lowest at 0.14 (95% ci: 0.00-0.37 i 2 93%, q 27.3). the results of these studies were significantly heterogeneous ( figure 3) . risk of bias assessment. the funnel plot depicted moderate to marked asymmetry, suggesting potential publication bias (supplemental figure 1) . for comparison, we populated doi plots with lfk indices to ascertain the publication bias. using these additional measures, only pleural line abnormalities, finding remained at high risk of publication bias. we used the quadas2 tool to assess the study quality and risk of bias; the included studies mostly revealed an unclear or moderate risk of bias (supplemental material table 1 ). there was marked heterogeneity with regard to pooling the proportion of pleural thickening, consolidation, or pleural effusion. covid-19 has struck the world with surprise, resulting in elated morbidity and mortality, strained the healthcare system, and depleted the resources even in resource-rich settings. 19 up to the date of submitting this manuscript, covid-19 has affected 3 million individuals (confirmed cases) and resulted in more than 200,000 deaths worldwide. 20 tools to aid in the early identification and follow-up are needed in an attempt to provide appropriate care and to allocate resources better. 21, 22 computed tomography scan has surfaced as a useful imaging modality in the diagnosis and follow-up of covid-19. 23, 24 although useful, its use is limited, as explained earlier. the recent interest in point-of-care ultrasound (pocus) of the figure 2 . forest plot presenting (a) the pooled proportion of b-pattern and (b) consolidation (the higher frequency of subpleural or pulmonary consolidations reported by the primary study) detected by lung ultrasound in symptomatic covid-19 patients. *i 2 is 0% for b-pattern proportion, suggesting homogeneity of data. there is marked heterogeneity depicted by extremely high i 2 for the finding of consolidation. lungs is due to its portability, steep learning curve, a relatively easier sterilization process, absence of ionizing radiation exposure, and its role in the follow-up. moreover, it has an excellent correlation with ct scan in various pulmonary diseases (b-lines, subpleural consolidations, and irregular pleural line). 4, 25 the use of pocus is of greater value in resourcelimited settings, for example, in some tropical areas where other diagnostic modalities may not be readily available, and testing resources may be limited. in these settings, basic ultrasound image acquisition and interpretation skills can be taught to healthcare providers of varying experiences, following a brief training course. 26 in our review, b-pattern predominated, occurring with a pooled frequency (pf) of 97% (94-100%). the results were consistent and homogenous across all the constituent studies. pleural line abnormalities were present in two-thirds of the cases (pf 70%, 95% ci: 13-100%), the frequency of other findings was less in our review, and the results were extremely heterogeneous. this heterogeneity is likely owing to differences in the settings, patient populations, number of lung zones examined, the level of operator expertise, ultrasound machine or probe used, stage and severity of the illness. hence, the presence of these findings (consolidations, pleural thickening, or pleural effusion) may be useful in the triage, prognosis, and follow-up. however, their absence cannot be used to rule out covid-19. two of the constituent studies (guorong et al. 18 and poggiali et al. 9 ) demonstrated a good correlation between the lus and ct scan findings. furthermore, guorong et al. demonstrated that the lus findings improve synchronously with clinical improvement, suggesting a potential role of lus in the clinical follow-up. however, this role needs to be supported by future studies. 9, 18 it is worthy to note two studies that were excluded from our review. the first study examined the role of lung us in asymptomatic patients with covid-19, hence excluded. 27 in their retrospective analysis of nine asymptomatic patients, lus revealed abnormalities in 22% (n = 2/9). one patient had b-pattern, and the other patient had pulmonary consolidations. whereas the frequency of b-pattern was low in this study, ct scan did similarly depict abnormalities in only 33% (n = 3/9), indicating a possible low yield of various imaging modalities in asymptomatic covid-19 patients. the second study was excluded as it was limited to a pediatric cohort. they retrospectively analyzed the data of eight covid-19-infected children; 88% (n = 7/8) had abnormalities on lung us (b-pattern n = 5/8, consolidations n = 2/8). on clinical improvement, all the lesions radiologically improved either partially or entirely, hinting toward a potential role of us in the clinical follow-up. 28 in the 2009 h1n1 pandemic, lus findings were used to differentiate between viral and bacterial pneumonia with an excellent interobserver agreement. bacterial pneumonia findings were lung consolidations with sonographic air bronchograms. however, the findings noted in cases of viral pneumonia were similar to our findings (b-pattern, pleural line abnormalities, or subpleural consolidations). 8, 26 it may be argued that lus findings may not enable clinicians to differentiate covid-19 from other viral lung infections; however, having such a prevalent finding amid a pandemic will lead to faster diagnostic and therapeutic decisions and better resource allocation. our review aimed to pool the reported proportions of various findings detected by pocus lungs and is the first metaanalysis aimed at assessing the role of lung us or pocus in the diagnosis of covid-19. the authors are of the view that this will be of value to frontline clinicians. we believe that the findings from our review will assist in the integration of this useful modality in the triage, diagnosis, management, and follow-up of covid-19 patients. our review is limited by a small number of constituent studies, a small number of patients, unclear bias risk, and inability to rule out publication bias. also, there is a lack of unifying definitions and inconsistencies in the reporting of various lung abnormalities. inadequate reporting of the extensiveness of lus findings (lung areas involved or a representative lus score) may limit its role in the temporal follow-up and its prognostic value. finally, we were not able to calculate the sensitivity and specificity owing to the absence of data necessary for their computation. a well-conducted diagnostic accuracy study comparing lus, ct scan, and various specific tests for covid-19 (pcr, igm, and igg on serial measurements) to ascertain the sensitivity, specificity, and diagnostic accuracy of each modality in the diagnosis of covid-19 is needed. in addition, we suggest studying lus on various severity spectrum of the disease to identify findings that correlate with disease severity. the results of the recently planned and ongoing trials, such as pocusco, echovid-19, pocusars-cov-2, virus, and covilus, will address some of the aforementioned limitations. [29] [30] [31] [32] [33] conclusion evidence of interstitial lung involvement, as depicted by b-pattern, is the most common and consistent finding on lung us in covid-19 patients. although nonspecific, the presence of this finding amid the covid-19 pandemic, in addition to other characteristic symptoms, will increase the disease likelihood. thus, pocus will likely play a vital role in the future triage, diagnosis, management, and follow-up of covid-19 patients. all together to fight novel coronavirus disease (covid-19) the characteristics and clinical value of chest ct images of novel coronavirus pneumonia lung ultrasound for daily monitoring of ards patients on extracorporeal membrane oxygenation: preliminary experience ultrasound in covid-19: a timeline of ultrasound findings in relation to ct point-of-care ultrasonography in patients admitted with respiratory symptoms: a single-blind, randomised controlled trial lung us surface wave elastography in interstitial lung disease staging point-of-care ultrasound (pocus) for hospitalists and general internists early recognition of the 2009 pandemic influenza a (h1n1) pneumonia by chest ultrasound can lung us help critical care clinicians in the early diagnosis of novel coronavirus (covid-19) pneumonia? findings of lung ultrasonography of novel corona virus pneumonia during the 2019-2020 epidemic challenges and opportunities for lung ultrasound in novel coronavirus disease (covid-19) 2020: point-of-care lung ultrasound findings in patients with novel coronavirus disease (covid-19) pneumonia preferred reporting items for systematic reviews and meta-analyses: the prisma statement quadas-2: a revised tool for the quality assessment of diagnostic accuracy studies covid-19 pneumonia manifestations at the admission on chest ultrasound, radiographs, and ct: single-center study and comprehensive radiologic literature review analysis of ultrasonic manifestations of pulmonary lesions in patients with covid-19 a clinical study of noninvasive assessment of lung lesions in patients with coronavirus disease-19 (covid-19) by bedside ultrasound transthoracic ultrasound evaluation of pulmonary changes in covid-19 patients during treatment using modified protocols two for one with split-or co-ventilation at the peak of the covid-19 tsunami: is there any role for communal care when the resources for personalised medicine are exhausted coronavirus cases wuhan and hubei covid-19 mortality analysis reveals the critical role of timely supply of medical resources global epidemiology of coronavirus disease 2019 (covid-19): disease incidence, daily cumulative index, mortality, and their association with country healthcare resources and economic status imaging manifestations and diagnostic value of chest ct of coronavirus disease 2019 (covid-19) in the xiaogan area ct characteristics of patients infected with 2019 novel coronavirus: association with clinical type identification of lung sliding: a basic ultrasound technique with a steep learning curve skills acquisition for novice learners after a pointof-care ultrasound course: does clinical rank matter? application value of lung ultrasound in asymptomatic patients with confirmed covid-19 lung ultrasound in children with covid-19 coronavirus (covid-19) diagnostic lung ul-trasound study -full text view -clinicaltrials lung ultrasound to diagnose covid-19 -full text view -clinicaltrials accuracy of lung ultrasound in the diagnosis of covid19 pneumonia -full text view -clinicaltrials the role of ultrasound in covid-19 -full text view -clinicaltrials point of care ultrasonography for riskstratification of covid-19 patients -full text view -clinical-trials acknowledgments: we thank the reviewers for their constructive feedback that led to improving the manuscript. publication charges for this article were waived due to the ongoing pandemic of covid-19.disclosure: no ethical approval was sought, given that this is a secondary synthesis of the already available literature. the data used in this review are available on reasonable request. this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-285486-99trkti1 authors: abd-elsalam, sherief; esmail, eslam saber; khalaf, mai; abdo, ehab fawzy; medhat, mohammed a.; abd el ghafar, mohamed samir; ahmed, ossama ashraf; soliman, shaimaa; serangawy, ghada n.; alboraie, mohamed title: hydroxychloroquine in the treatment of covid-19: a multicenter randomized controlled study date: 2020-08-14 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0873 sha: doc_id: 285486 cord_uid: 99trkti1 the covid-19 pandemic is showing an exponential growth, mandating an urgent need to develop an effective treatment. indeed, to date, a well-established therapy is still lacking. we aimed to evaluate the safety and efficacy of hydroxychloroquine (hcq) added to standard care in patients with covid-19. this was a multicenter, randomized controlled trial conducted at three major university hospitals in egypt. one hundred ninety-four patients with confirmed diagnosis of covid-19 were included in the study after signing informed consent. they were equally randomized into two arms: 97 patients administrated hcq plus standard care (hcq group) and 97 patients administered only standard care as a control arm (control group). the primary endpoints were recovery within 28 days, need for mechanical ventilation, or death. the two groups were matched for age and gender. there was no significant difference between them regarding any of the baseline characteristics or laboratory parameters. four patients (4.1%) in the hcq group and 5 (5.2%) patients in the control group needed mechanical ventilation (p = 0.75). the overall mortality did not differ between the two groups, as six patients (6.2%) died in the hcq group and 5 (5.2%) died in the control group (p = 0.77). univariate logistic regression analysis showed that hcq treatment was not significantly associated with decreased mortality in covid-19 patients. so, adding hcq to standard care did not add significant benefit, did not decrease the need for ventilation, and did not reduce mortality rates in covid-19 patients. coronaviruses are a large family, which may cause illness in animals or humans. in humans, several coronaviruses are known to cause respiratory infections, ranging from common cold to more severe diseases such as middle east respiratory syndrome and sars. [1] [2] [3] [4] [5] [6] the most recently discovered coronavirus is sars-cov-2 which causes covid-19. as cases of covid-19 continue to rise in different countries, health systems are facing enormous pressure to manage covid-19 patients. by august 2, 2020, covid-19 has been confirmed in about 17,660,523 million individuals worldwide and has resulted in more than 680,894 deaths. these numbers are still increasing. more than 180 countries have reported laboratoryconfirmed cases of covid-19 on all continents, except antarctica. [1] [2] [3] [4] in egypt, the official number of infected patients was 94,316, with 4,834 deaths as of august 2, 2020. [1] [2] [3] [4] [5] [6] [7] [8] [9] [10] [11] although many vaccines are in development, effective therapy is needed to treat currently infected patients and prevent mortality. chloroquine (cq) and hydroxychloroquine (hcq) have been used for decades in the treatment and prophylaxis of a number of conditions including malaria. the ability of these drugs to inhibit other coronaviruses, such as sars-cov-1, has been explored. although generally considered safe, there are potential risks associated with taking these medications, including cardiac arrhythmia. [7] [8] [9] [10] [11] although an initial study in france found encouraging results for the treatment of covid-19 with hcq, the study was later criticized for its methodological problems, leading to skepticism about the validity of its results. other similar results were not represented in any further subsequent studies, but even reported deleterious clinical outcomes especially cardiac adverse events like prolongation of qt interval. 8 on march 28, 2020, the food and drug administration (fda) granted an emergency use authorization for use of oral formulations of cq and hcq in the treatment of covid-19. 7-11 based on emerging data showing cq and hcq as unlikely to be effective in the treatment of covid-19, 12,13 the fda revoked its previous emergency use authorization for both drugs on june 15, 2020. in this study, we aimed to evaluate the safety and efficacy of hcq added to the standard of care versus the standard of care alone in patients with covid-19. patients admitted to three tertiary referral centers (n = 194) managing patients with suspected and confirmed covid-19 in egypt in the period between march and june 2020 were enrolled. the patients were clinically stratified into mild, moderate, and severe disease according to the who interim guidelines published on march 13, 2020. mild cases represented patients with uncomplicated upper respiratory tract viral infection, moderate cases represented patients with pneumonia but without need for supplemental oxygen, whereas severe disease represented cases with fever or suspected respiratory infection, plus one of the following: respiratory rate > 30 breaths/min, severe respiratory distress, or spo 2 £ 93% on room air. 14 the egyptian ministry of health (moh) adopted a standard of care treatment protocol for covid-19 patients. it included paracetamol, oxygen, fluids (according to assessment), empiric antibiotic (cephalosporins), oseltamivir if needed (75 mg/ 12 hours for 5 days), and invasive mechanical ventilation with hydrocortisone for severe cases if pao 2 < 60 mmhg, o 2 saturation < 90% despite oxygen or noninvasive ventilation, progressive hypercapnia, respiratory acidosis (ph < 7.3), and progressive or refractory septic shock. 15 patients were randomized into two groups using a computerized random number generator using simple randomization with an equal allocation ratio. during randomization, the proportional allocation of each clinical stratum was equalized in both groups. study groups. 1. hydroxychloroquine group: this group included 97 patients who received hcq 400 mg twice daily (in day 1) followed by 200 mg tablets twice daily added to the standard of care treatment adopted by the egyptian moh for 15 days. 2. control group: this group included 97 patients who received only the standard of care treatment adopted by the national moh for 15 days. all the patients were followed up for 4 weeks. the study included all patients admitted with sars-cov-2 infection and enrolled both genders. patient who had allergy or contraindication to hcq, pregnant and lactating females, and patients with cardiac problem (chronic heart failure or prolonged qt interval on electrocardiogram [ecg]) were excluded from the study. informed written consent was obtained from each participant, and the study was approved by the ethics committee of the faculty of medicine, tanta university. privacy of the participants and confidentiality of the data were assured. risks and benefits were explained to the patients. the study was registered on clinicaltrials.gov with registration number nct04353336. all the participants were subjected to thorough history taking and full clinical examination including age, gender, weight and height measurements, and calculation of body mass index (bmi); medication history; and investigations in the form of complete blood picture, liver function tests, computed tomography of the chest (ct chest), and sars-cov-2 detection in nasopharyngeal swabs using pcr and ecg. assessment of the studied medication side effects was performed using a questionnaire. statistical analysis. data were analyzed using statistical package for social sciences v. 23 and were expressed in number, percentage (%), mean (x̅ ) and sd. the variables were tested for normality by the shapiro-wilks test. student's t-test was used for normally distributed quantitative variables and mann whitney's test for not normally distributed ones. chi-square test (χ 2 ) was used to study association between qualitative variables, and whenever any of the expected cells were less than five, fischer's exact test was used. binary logistic regression was used to ascertain the effect of the potential risk factors on the patients' mortality. a two-sided p-value of < 0.05 was considered statistically significant. post hoc power analysis. considering the percentage of recovery as a primary endpoint and by using g*power program, post hoc power analysis revealed a sample power of 80.6% with the following input parameters: two-tailed α error 0.05, 54.0% recovery rate in the hcq group, 34.0% recovery in the control group, and 97 sample size in each group. 16 at the time of presentation, interrupted fever was present in 44.6%, continuous fever in 22.3%, headache in 42.9%, sore throat in 25.7%, anorexia in 33.1%, anosmia in 26.9%, pallor in 3.4%, cyanosis in 4.6%, fatigue in 49.0%, vomiting in 13.7%, diarrhea in 35.0%, abdominal pain in 19.4%, cough in 61.3%, and dyspnea in 24.2% of the included patients. oxygen saturation between 95 and 90 was present in 16.0%, 90-85 in 7.4%, and less than 85 in 6.9% of all the participants. the computed tomography chest scans were normal in 33.1%, ground-glass opacities in 23.4%, confluent opacities in 25.7%, consolidation in 10.9%, extensive consolidation in 6.3%, and emphysema in only 0.6%. the two groups were matched for age and gender, with no significant difference between them. they had no significant difference regarding bmi, residence, smoking, pregnant females, or the presence of comorbidities. the patients were randomized equally between the two groups regarding the disease severity (table 1) . there was no significant difference between the two groups regarding laboratory parameters (table 2) . mechanical ventilation was needed in four patients (4.1%) in the hcq group and 5 (5.2%) in the control group, with no significant difference between the two groups (p = 0.75). six patients (6.2%) died in the hcq group, and five patients (5.2%) died in the control group without any significant difference between the two groups either (p = 0.76). eleven patients (11.3%) in the hcq group needed intensive care unit (icu) admission, and 13 patients (13.4%) in the control group needed the same (p = 0.83). the mean duration to negative pcr was 17 ± 3 days in the hcq group and 18 ± 2 in the control group (p = 0.11). the hcq group had a mean of 9 ± 2 days to show clinical improvement and 11 ± 3 days to hospital discharge, whereas the control group had a mean of 10 ± 3 to clinical improvement and 11 ± 2 to hospital discharge (p = 0.80 and 0.52, respectively) ( table 3) . after 28 days, there was no significant difference between the two groups regarding the clinical outcome (p = 0.07). complete recovery was achieved in 52 cases (53.6%) of the hcq group, whereas 23 cases (23.7%) were in mild, 8 (8.2%) were in moderate, 8 (8.2%) in severe disease status, and six patients (6.1%) died. among the control group, 33 patients (34.0%) recovered completely, 39 (40.2%) were in mild, 10 (10.3%) were in moderate, 9 (9.2%) were in severe disease status, and five patients (5.1%) died. by logistic regression, the overall mortality was not significantly associated with hcq therapy; however, it was significantly related to the patient's age, alanine aminotransferase, serum creatinine, serum ferritin, c-reactive protein, oxygen saturation, and the presence of diabetes mellitus (table 4) . chloroquine and hcq are well-known drugs and have been used for decades as antiparasitic and anti-inflammatory drugs to treat malaria and rheumatological disorders. chloroquine was shown to be effective against sars-cov in invitro studies. this may be because of disruption of viral replication, changing immune system activity in addition to its inflammatory effect. 17 the two drugs have been tried earlier for the treatment of sars infection and showed promising efficacy. with the emergence of sars-cov-2 pandemic, they have been suggested as potential treatment for the new coronavirus 2019 based on the previous evidence from different coronavirus strains. 18 although cardiac toxicity is a known adverse event requiring monitoring during treatment, hcq showed promise in treating sars-cov-2-infected patients with multiple comorbidities including coronary artery disease. a large trial from india showed that hcq can decrease time to recovery both in symptomatic and in asymptomatic patients with no effect on mortality. 19 at the beginning of the pandemic in europe, a small series of covid-19 patients treated in france with hcq showed improved decline in sars-cov-2 viral load compared with controls, which was augmented by the addition of azithromycin. 7 however, this study had serious methodological flaws and could not be considered as a good evidence in the favor of hcq use. [8] [9] [10] [11] many other conflicting trials have been published in the past few months leading initially to emergency use authorization for hcq use in the treatment of covid-19 and later on withdrawal of this authorization by the fda. initial observational trials of hcq use in hospitalized patients showed that there were no increased risks of mortality or intubation in groups receiving hcq or the control group who received only standard of care although patients who received hcq were more critically ill. 20 however, many published trials had some methodological flaws and missed important patient outcomes urging the need for properly designed, adequately powered trials to support clinical decisions of hcq use in treating covid-19 patients. 21 administration of hcq did not result in a significantly higher probability of conversion from positive to negative pcr than standard care alone in patients admitted to hospital with nonresponsive mild-to-moderate covid-19 in china. adverse events were more frequent in hcq recipients than in non-recipients. 22 a meta-analysis included seven studies with a large number of patients showing that treatment with hcq was associated with faster improvement of fever, cough, and less radiological progression of lung lesions. however, there was no difference in the virological cure, clinical improvement, or mortality. 23 many subsequent trials did not show benefit for hcq use in covid-19, with some of them suggesting more adverse events associated with its use. [22] [23] [24] a recent clinical trial by skipper et al. 12 studied the change in symptom severity over 14 days in nonhospitalized patients between hcq and control groups and did not find any significant difference (p = 0.12). another trial by cavalcanti et al. 13 compared three groups; standard care group, standard care plus hcq, and standard care plus hcq and azithromycin. the clinical status at 15 days assessed by a seven-level ordinal scale did not show any significant difference among the three groups. moreover, elevated liver enzymes and prolonged qt intervals were more frequent among patients who used hcq. in our study, adding hcq to standard care did not add an extra benefit for the patients. hydroxychloroquine arm was similar in all outcomes. moreover, hcq was not effective as postexposure prophylaxis against covid-19 when administered within 4 days after exposure. [25] [26] [27] [28] [29] limitations of the study include small sample size which was not adequately powered for survival endpoint. the number of the included patients was limited because in egypt, tertiary care hospitals were assigned lately to deal with covid-19 patients and had many regulations by the egyptian moh. the study lacks long-term follow-up which could be addressed in a prospective trial. the utility of hcq should be evaluated in larger multicenter trials either alone or in combination with other drugs/lines of treatment. the role of hcq as a prophylaxis against sars-cov-2 infection should be among the future trials also. in conclusion, our trial adds extra evidence from egypt that hcq may not be beneficial as a treatment for covid-19. a novel coronavirus from patients with pneumonia in china mers, sars and other coronaviruses as causes of pneumonia a pneumonia outbreak associated with a new coronavirus of probable bat origin lopinavir/ritonavir combination therapy amongst symptomatic coronavirus disease 2019 patients in india: protocol for restricted public health emergency use case of the index patient who caused tertiary transmission of covid-19 infection in korea: the application of lopinavir/ritonavir for the treatment of covid-19 infected pneumonia monitored by quantitative rt-pcr a trial of lopinavir-ritonavir in adults hospitalized with severe covid-19 hydroxychloroquine and azithromycin as a treatment of covid-19: results of an open-label nonrandomized clinical trial no evidence of rapid antiviral clearance or clinical benefit with the combination of hydroxychloroquine and azithromycin in patients with severe covid-19 infection identification of falsified chloroquine tablets in africa at the time of the covid-19 pandemic insight into 2019 novel coronavirus -an updated interim review and lessons from sars-cov and mers-cov chloroquine and hydroxychloroquine for the prevention or treatment of covid-19 in africa: caution for inappropriate off-label use in healthcare settings hydroxychloroquine in nonhospitalized adults with early covid-19: a randomized trial hydroxychloroquine with or without azithromycin in mildto-moderate covid-19 clinical management of severe acute respiratory infection (sari) when covid-19 disease is suspected interim guidance egyptian ministry of health and population (moh), 2020. diagnosis and treatment protocol for covid-19 g*power 3: a flexible statistical power analysis program for the social, behavioral, and biomedical sciences chloroquine or hydroxychloroquine for prophylaxis of covid-19 hydroxychloroquine for covid-19: what is our current state of knowledge? characteristics, treatment outcomes and role of hydroxychloroquine among 522 covid-19 hospitalized patients in jaipur city: an epidemio-clinical study observational study of hydroxychloroquine in hospitalized patients with covid-19 covid-19 coronavirus research has overall low methodological quality thus far: case in point for chloroquine/hydroxychloroquine hydroxychloroquine in patients with mainly mild to moderate coronavirus disease 2019: open label, randomised controlled trial virological and clinical cure in covid-19 patients treated with hydroxychloroquine: a systematic review and meta-analysis effect of hydroxychloroquine in hospitalized patients with covid-19: preliminary results from a multi-centre, randomized a randomized trial of hydroxychloroquine as postexposure prophylaxis for covid-19 hydroxychloroquine as postexposure prophylaxis for covid-19 sars-cov-2: the path of prevention and control apasl covid task force, apasl covid liver injury spectrum study, 2020. pre-existing liver disease is associated with poor outcome in patients with sars cov2 infection; the apcolis study (apasl covid-19 chloroquine as chemoprophylaxis for covid-19: will this work? key: cord-310868-rqmny4r2 authors: tesfaye, wubshet; abrha, solomon; sinnollareddy, mahipal; arnold, bruce; brown, andrew; matthew, cynthia; oguoma, victor m.; peterson, gregory m.; thomas, jackson title: how do we combat bogus medicines in the age of the covid-19 pandemic? date: 2020-08-18 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0903 sha: doc_id: 310868 cord_uid: rqmny4r2 the covid-19 pandemic has brought concurrent challenges. the increased incidence of fake and falsified product distribution is one of these problems with tremendous impact, especially in lowand middle-income countries. up to a tenth of medicines including antibiotics and antimalarial drugs in the african market are considered falsified. pandemics make this worse by creating an ecosystem of confusion, distraction, and vulnerability stemming from the pandemic as health systems become more stressed and the workload of individuals increased. these environments create opportunities for substandard and falsified medicines to be more easily introduced into the marketplace by unscrupulous operators. in this work we discuss some of the challenges with fake or falsified product distribution in the context of covid-19 and proposed strategies to best manage this problem. covid-19 has revealed weaknesses and vulnerabilities in global and regional health governance, with the proliferation of "fake health news" and the surgisphere scandal exacerbating the dissemination of fake medicines. [1] [2] [3] [4] these weaknesses are of particular concern in developing countries with inadequate health infrastructures. 5, 6 research into finding an effective treatment for covid-19 has continued unabated. although some observational studies showed early promise with chloroquine and hydroxychloroquine, subsequent randomized trials revealed the lack of clinical benefit on these drugs. 7, 8 the discontinuation of the major trials on hydroxychloroquine-recovery 9 and solidarity 10 -further strengthens the notion that it may not be effective, especially in severe covid-19 cases. subsequently, randomized trials revealed positive outcomes associated with remdesivir and dexamethasone use in people with severe covid-19. 11, 12 remdesivir was associated with shorter time to recovery, 11 whereas dexamethasone significantly cut mortality rates in people on oxygen support. 12 covid-19 management protocols in different jurisdictions are modified accordingly, although firm evidence is needed to clearly understand the benefit of these drugs and to whom. on top of that, accelerated vaccine developments are underway in various countries, with 21 of them currently under clinical evaluation and one in phase iii testing. given the urgency associated with finding the magic bullet for covid-19, research has become increasingly challenging and is resulting in self-examination by the scientific community and the wider public about the governance of fast-tracked research. 13 the hype with chloroquine and hydroxychloroquine at the beginning was associated with misinformation 14 -sometimes in the form of "fake health news"-and incidents of quarantinebased supply chain disruption and stockpiling. 15 for the global south, the pandemic exacerbates the existing proliferation of substandard and falsified medical products, and these trends are likely to continue as effective drugs continue to emerge from ongoing trials. the who defines substandard products as "authorized medical products that fail to meet quality standards or specifications, or both" and falsified products as "products that deliberately/ fraudulently misrepresent identity, composition, or source." 16 considering less than 30% of regulatory agencies in the world can ensure the adequacy of medicines and vaccines, 17 pandemics of covid-19 proportion would inevitably present enormous regulatory challenges. health emergencies can create an environment of confusion, distraction, and vulnerability stemming from the pandemic, as health systems become more stressed and the workload of individuals increased. these environments create opportunities for substandard and falsified medicines to be more easily introduced into the marketplace by unscrupulous operators. it is worth noting that supply chains are fragile even in normal times, 18 and the covid-19 pandemic made the vulnerabilities with the supply chains even more noticeable. the supply chain disruption caused by the reduced production and export of some medicines owing to closure of china's active pharmaceutical ingredient and raw material supply revealed this vulnerability. 19 this has stimulated debate on the need to diversify the sources of pharmaceutical supply to ensure continuity of supply across the globe. 20 there are ongoing industry efforts to support initiatives to maximize domestic productions of raw materials to prevent disruption to the supply chain in the future. [19] [20] [21] these problems particularly need further attention in fragile healthcare systems and are highly dependent on the import of medicines to cover their domestic demand. falsified medical products may contain no active ingredient, the wrong active ingredient, or the wrong amount of the correct active ingredient. 16 the prevalence of falsified medicines spans from 1% in developed settings to 10% in the global south. 22, 23 based on more than $4.3 billion worth of substandard and falsified products seized by border security agents from 2014 to 2016, over 35% of the products were antibiotics and 9% were antimalarials. 24 counterfeiters also target treatments for diabetes, epilepsy, heart diseases, allergy, blood pressure, cancer, and stomach ulcers. 25 falsified and substandard medicines are deliberately produced mostly for monetary gain and do not produce the required therapeutic benefits. 16, [26] [27] [28] evidence shows that the use of falsified medicines is associated with serious health outcomes, including mortality both in developing and developed countries. 27 in low-and middle-income countries (lmics) alone, falsified medicines cost economies up to us$200 billion for additional care following treatment failure. 26, 29 nearly half of the who reports on falsified medical products (42%) originate from the african region, and many are linked with antimalarial drugs. in sub-saharan africa, substandard and falsified antimalarials are estimated to cost us$38.5 million (us$21.4-us$52.4 million) and 116,000 (64,000-158,000) additional deaths 16, 29 -which can be prevented. falsified products in the context of covid-19. although a true estimate on the impact of falsified medical products during covid-19 is yet to be made, the pandemic has fueled a global surge in the production and distribution of falsified medicines and medical supplies 25 -resulting in a multifold global crisis. this is evident from recent operations by the international criminal police organization that captured millions of units of counterfeit pharmaceuticals (antiviral medications, antimalarial chloroquine, vitamin c, painkillers, and antibiotics), coupled with several reports on the counterfeit medical supplies (mostly surgical masks and coronavirus testing kits) from around the globe. similarly, in africa, the early days of the pandemic have seen a spike in the demand for chloroquine and hydroxychloroquine that subsequently led to a surge in their falsified version. 30 the who global surveillance and monitoring system on substandard and falsified medical products received nine reports of confirmed falsified chloroquine products from three countries-cameroon, democratic republic of congo, and niger between march 31, 2020 and april 2, 2020. 31 this is a worrisome development for a continent with a long-standing problem of falsified antimalarial drugs. 32 looking at the trends of a surge in falsified products and in the absence of proper regulation, drugs such as dexamethasone may have a similar fate. therefore, regulatory agencies should have additional oversight on the distribution of products containing this drug to ensure their quality, particularly in settings where product regulation is weak. the trend is also likely to be much higher this year as the demand for important drugs exceeds the supply, following covid-19-led lockdowns in india and china (two largest producers of medical products). 8, 30 african nations are largely spending their scarce resources on covid-19 containment measures, aiming to prevent largescale disease dissemination that can lead to huge health, social, and economic crises. this will undoubtedly leave them with limited capacity to tackle other parallel emergencies. falsified medicines tend to emerge when access to medicines is constrained, when the pharmaceutical governance is weak, where there is limited technical capacity to monitor products throughout the supply chain, and when there is lack of adequate financial and political commitments. 33 these standards come under the purview of the national regulatory system. a looming recession on the horizon is also likely to stimulate social tolerance for substandard and falsified medical products. strong regulatory frameworks are, therefore, essential in enabling regulatory authorities to effectively combat the distribution of falsified medicines. corrupt and under-resourced systems, coupled with limited capacity for oversight, allow falsified medicine to easily reach the end user. 32 high-income countries have a strict regulatory framework, technological means, and financial resources to detect and limit the distribution of falsified and counterfeited products. the penetration of counterfeit products is generally higher in lmic settings. there is a risk that when the covid-19 pandemic further spreads in lmics, there will be an increased potential for the distribution of falsified and counterfeit medicines, something the international community needs to be alert to and work against. ongoing efforts to tackle distribution of falsified products. given the complexity of the pharmaceutical distribution and supply chain, vulnerabilities could occur at any stage across the supply chain, endangering patients around the world. 34 building collaborations across countries and private and public agencies is required to strengthen the regulatory footprint at national and international levels. the u.s. food and drug administration and the european medicines agency are trialing to implement various technologies to track the steps involved in the supply chain. in africa, a health information technology/big data startup rxall has come up with a handheld scanner, which collects the spectral signature of a drug and then compares it with a cloud-based database and sends information to an application on the phone. although field detection technologies could play a role in intercepting substandard and falsified products, their cost feasibility and when to effectively use them across the supply chain continuum remain to be understood. 35, 36 also, this seems to be a palliative approach to a multifactorial systemic problem. there are some global efforts to improve access and quality of medicines. the united states agency for international development (usaid) global health supply chain program is exploring innovations based on prior experiences to devise more efficient supply chains aiming to implement global supply chain standards (gs1) for improved cost, efficiency, and availability of health commodities worldwide. 37 furthermore, some programs specifically aim to strengthen regulatory agencies in lmics with the ultimate goal to ensure sustainable access to appropriate, safe, affordable, and quality-assured medicines. 38, 39 this also includes supply chain strengthening activities undertaken by usaid, unicef, the united nations population fund, the global fund, bill & melinda gates foundation, and others that focus on supporting the end to end product traceability supported by the use of novel technologies. however, recent attempts to halt funding arrangements for the who could be a serious threat to global health security, especially in the middle of a catastrophic global pandemic. in summary, the availability and distribution of falsified products is an age-old problem, with higher penetration in lmics, a situation that is likely to get worse as disruptions place greater stresses on regulatory and supply chain processes. in the age of covid-19, there is an ongoing need to ensure adequate supplies of medicines and medical equipment, not only for meeting the needs of the pandemic but also for other health needs. in meeting these needs, governments, pharmaceutical regulatory agencies, and associate supply chains must have practical and financially supported strategies to ensure quality-assured medicines are made available for the determined need. more importantly, it is imperative medicines regulatory authorities and relevant stakeholders implement robust authentication and procurement processes to ensure quality medicines supply. 40 in line with this, we propose the following set of strategies can be cascaded to relevant stakeholders (table 1) to ensure the availability of quality medicines during the time of covid-19 and beyond. strengthening national and international pharmaceutical governance (agencies such as interpol have intercepted significant falsified products, which should continue) adequate funding for monitoring distribution of falsified products initiation and/or assistance in coordinating the information-sharing process among regulators in different regions and countries supply agencies ensuring security of supply chains through implementation of good distribution and warehousing practice. adequate and continuous supply of standard drugs can avert the bad ones from filling the vacuum in times where there is a heightened fear and desperation for treatments good procurement practice with an extra focus on quality local regulatory agencies regulatory agencies are expected to fulfill the global benchmarking tool, which has more than 200 indicators to measure effectiveness of regulatory functions in a country. at this stage, tanzania is the only country that achieved such a milestone in africa, 17 emphasizing on the need for other african nations to leverage this local experience for implementation in their respective settings have the responsibility to make sure there is a routine inspection on their borders in relation to the entry of new medicines and medical products implementation of legislative changes restricting counterfeit drugs improved investment capital and infrastructure to encourage small-and medium-sized drug manufacturing companies to meeting international standards strong political will to enforce regulations health ministries creating awareness among citizens regarding medicine quality through their public health section technological companies implementation of detection technologies to enable easy identification of falsified products, when affordable and appropriate social media platforms should be held accountable for the dissemination of fake health news, with a more proactive identification and removal of fake health news provided by armchair epidemiologists, vendors of bogus cures, and citizen journalists. this requires working closely with consumer protection agencies across the globe and with qualified public health officials to take quick and decisive steps to remove inappropriate and inaccurate content, and to prevent it appearing in the first place researchers and health professionals the hype associated with finding a cure for covid-19 has changed the way research is communicated to the public, with potentially flawed or premature research findings being hastily released. researchers and health professionals need to play a greater role in controlling the narrative surrounding covid-19 research continued engagement with the communities through education, training, and public discourse can reduce harm and save lives health professionals should always be vigilant of newer medicines joining the market, especially those with increased relevance due to the pandemic citizens the pandemic response cannot be effective without active participation of the people. it is important to heed advices by health experts, especially in relation to drugs, and to differentiate between fact-based reporting and nonfactual claims particularly when it comes to medicine-related information. 41 high-profile coronavirus retractions raise concerns about data oversight the pandemic's first major research scandal erupts pharmaceutical anti-counterfeiting: combating the real danger from fake drugs pollution of health news phake: the deadly world of falsified and substandard medicines a growing headache: the prevalence of international counterfeit pharmaceutical trade in developing african nations a randomized trial of hydroxychloroquine as postexposure prophylaxis for covid-19 hydroxychloroquine in patients with mainly mild to moderate covid-19: open label, randomised controlled trial preliminary results from recovery trial on the use of hydroxychloroquine in hospitalised patients with covid-19 who discontinues hydroxychloroquine and lopinavir/ritonavir treatment arms for covid-19 remdesivir for the treatment of covid-19-preliminary report dexamethasone in hospitalized patients with covid-19-preliminary report covid-19: validity of key studies in doubt after leading journals issue expressions of concern when fear and misinformation go viral: pharmacists' role in deterring medication misinformation during the 'infodemic' surrounding covid-19 potential shortages of hydroxychloroquine for patients with lupus during the covid-19 pandemic. paper presented at: jama health forum substandard and falsified medical products tanzania is first african country to reach an important milestone in the regulation of medicines medical supply chains are fragile in the best of times and covid-19 will test their strength. the conversation covid-19 is reshaping the pharmaceutical supply chain covid-19 has exposed cracks in the global medicines supply chain. we need to fix them covid-19 pandemic: knock-on effects for pharma supply chains a study on the public health and socioeconomic impact of substandard and falsified medical products. geneva, switzerland: world health organization poor-quality and counterfeit drugs: a systematic assessment of prevalence and risks based on data covid-19 crisis underscores need to address trade in fake pharmaceuticals falsified and substandard drugs: stopping the pandemic the health consequences of falsified medicines-a study of the published literature are we doing enough to prevent poor-quality antimalarial medicines in the developing world? 10 medical products in developing countries is substandard or falsified the consequence of covid-19 on the global supply of medical products: why indian generics matter for the world? falsified chloroquine products circulating in the who region of africa committee on understanding the global public health implications of substandard, falsified, and counterfeit medical products, institute of medicine identifying market risk for substandard and falsified medicines: an analytic framework based on qualitative research in china an empirical review of antimalarial quality field surveys: the importance of characterising outcomes field detection devices for screening the quality of medicines: a systematic review global landscape assessment of screening technologies for medicine quality assurance: stakeholder perceptions and practices from ten countries usaid global health supply chain program. global standards [web site usp, 2020. promoting the quality of medicines (pqm+) program. available at projects/the-medicines-technologies-andpharmaceutical-services-mtaps-program covid-19 and risks to the supply and quality of tests, drugs, and vaccines fake news: medicines misinformation by the media key: cord-306149-sd0s0jup authors: ratanarat, ranistha; sivakorn, chaisith; viarasilpa, tanuwong; schultz, marcus j. title: critical care management of patients with covid-19: early experience in thailand date: 2020-05-18 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0442 sha: doc_id: 306149 cord_uid: sd0s0jup since late december 2019, the world has been challenged with an outbreak of covid-19. in thailand, an upper middle–income country with a limited healthcare infrastructure and restricted human resources, nearly 3,000 confirmed covid-19 cases have been reported as of early may 2020. public health policies aimed at preventing new covid-19 cases were very effective in halting the pandemic in thailand. case fatality in thailand has been low (1.7%), at least in part due to early stratification according to risk of disease severity and timely initiation of supportive care with affordable measures. we present our initial experience with covid-19 in thailand, focusing on several aspects that may have played a crucial role in curtailment of the pandemic, and elements of care for severely ill covid-19 patients, including stratification, isolation, and affordable diagnostic approaches and supportive care measures. we also discuss local considerations concerning some proposed experimental treatments. covid-19, a respiratory infection that originated in wuhan, china, in late 2019, by now has spread to almost all countries worldwide. 1 the numbers of cases are still rising in most countries, at an unprecedented and worrisome speed. 2 thailand, an upper middle-income country in southeast asia, was one of the first countries outside of china to report covid-19 cases. the first case was a traveler from wuhan, china, 3, 4 followed by local transmission to a public transport driver in bangkok. 5 since then, nearly 3,000 confirmed covid-19 cases have been reported in thailand. 2 among several problematic aspects of covid-19, its transmissibility may be most challenging. infected individuals produce large quantities of virus in their upper airways in the prodromal phase, during which they remain mobile and able to continue their usual activities, and thus able to transmit the disease to many contacts. this is very different from the earlier sars, in which viral shedding occurred mainly when patients manifested severe illness. 6 the higher transmissibility of covid-19 makes the outbreak much more difficult to control, thus increasing the size of the pandemic. as of early may, however, thailand's response to covid-19 seems successful. public health measures, 7, 8 together with assistance from village health volunteers and the general public, limited the number of covid-19 cases, and the pandemic is by now, for the most part, halted in thailand. 9 since soon after the start of the outbreak, a tremendous volume of reports on clinical management of covid-19 has been published, mainly from and for high-income countries. some of the suggestions and recommendations in those reports are rather unsuitable for middle-income countries, where resources are usually severely limited, as in thailand. the objective of this report is to outline our initial experience with the management of covid-19 patients requiring intensive care unit (icu) admission. after highlighting the most important measures taken in thailand, we discuss affordable diagnostic approaches and supportive care measures taken in thai hospitals. we also discuss local considerations regarding experimental treatments. in thailand, as in wuhan, china, 10 several measures may have limited the progression of covid-19 (table 1) , possibly reducing the need for hospitalization and eventually icu admission for ventilatory support. case definition. in thailand, the definition of "patients under investigation" (puis) was updated frequently, to improve early identification of covid-19 cases. the current definition includes 1) a history of fever or documented temperature ³ 37.5°c and/or any respiratory symptom accompanied by a history of high risk for covid-19 infection in the 14 days before the onset of symptoms, 2) cases of suspected pneumonia with unknown etiology, 3) clusters of sick individuals, and 4) suspected cases among healthcare workers. 8 laboratory screening and isolation. in thailand, the diagnosis of covid-19 is confirmed by reverse transcriptase-pcr (rt-pcr). in critically ill patients, sputum or tracheal suction specimens are preferred over nasopharyngeal or throat swabs for testing because the lower respiratory tract samples are most often testing positive for the virus. 11 it is recommended that every confirmed case, with or without symptoms, is hospitalized or kept under observation at a designated isolation facility. after the clinical condition has improved, and if no complications are observed, cases are considered for transfer to a designated hospital or temporary patient ward for covid-19, continuing isolation for at least 14 days from the date of onset. for asymptomatic infection, which is usually identified by contact tracing, the isolation duration will be counted from the day of diagnosis. after that, it is recommended that, for an additional 2 weeks, patients 1) wear a surgical mask at all times and pay extra attention to respiratory hygiene, such as coughing or sneezing in a flexed elbow or tissue, and disposing of used tissue into a closed bin; 2) avoid close contact with vulnerable populations; and 3) seek medical care immediately whenever respiratory symptoms recur or worsen. lung imaging. a chest radiograph (cxr) in patients with moderate covid-19 may show nonspecific multi-lobar opacities that rapidly progress over the first days of illness. chest computed tomography (ct) findings in covid-19 patients are more specific, showing bilateral, multi-lobar "ground-glass opacification" with the so-called crazy paving. 12 because of lack of ct scanners in most thai hospitals, cxr remains the first imaging modality for diagnosis. it should be noted that both "ground-glass opacification" and "crazy paving" cannot be seen on a cxr, and also that a cxr may not show abnormalities in the early stages of this disease. 13 lung ultrasound (lus) is an attractive alternative for screening and monitoring covid-19. 14 in our experience, lus is able to detect covid-19 earlier than a cxr, and daily lus combined with physical examination facilitates early detection of progression of the disease. however, lus requires specific training, which is a limitation in settings where ultrasound is not yet as extensively used as in thai hospitals. we also have promising experience with the "kigali modification of the berlin definition for acute respiratory distress syndrome (ards)" in the early detection and management of covid-19 patients. 15 the kigali modification allows the use of pulse oximetry instead of blood gas analysis and lus instead of chest ct to detect and define the severity of ards. antiviral treatment before icu admission. thailand has set national guidelines for antiviral treatment. patients with mild symptoms receive chloroquine or hydroxychloroquine plus a boosted protease inhibitor, lopinavir or darunavir plus ritonavir. favipiravir is not recommended in mild cases because of its limited availability. 8 favipiravir is an antiviral rna polymerase inhibitor for which most preclinical data are derived from its influenza and ebola activity. 16 it is given to all patients with proven covid-19 who have symptoms or signs consistent with pneumonia, or when there is hypoxemia (spo 2 < 95% on room air). 8 in thailand, icu management of covid-19 is limited to affordable measures (table 2) . infection control in the icu. availability and proper use of isolation and personal protective equipment (ppe) are essential to protect frontline healthcare workers, as well as other patients without covid-19. as in other countries, thailand has a shortage of negative pressure rooms and ppe. only severe cases or cases that undergo aerosolgenerating procedures can be placed within negative pressure rooms, if available. aerosol-generating procedures, such as collection of respiratory specimens, bronchoscopy, and cardiopulmonary resuscitation, should be minimized or avoided. metered dose inhalers are used for inhaled medications, including for intubated patients. disposable filtering facepiece respirators (ffrs) are not approved for routine decontamination and reuse as standard of care. however, ffr decontamination and reuse is considered a crisis capacity strategy. respiratory support. supplementary oxygen is a first step in respiratory support of covid-19 patients with hypoxemia. oxygen is provided either by nasal prongs or a non-rebreather mask. the use of high-flow nasal oxygen (hfno) and noninvasive ventilation (niv) is not supported when airborne infection isolation rooms (aiirs) are not available because of infection control concerns. in case an aiir is available, hfno can be used in younger patients without comorbidities who do not tolerate nasal cannula, or with niv with a dual limb system in morbid obesity or copd patients ( figure 1 ). their use is weighed against the assumption that these therapies often fail to prevent the need for invasive ventilation in covid-19 patients. in our experience, hypoxemia is often remarkably well tolerated with covid-19 ("happy hypoxia"), in particular in younger patients. this is notably different from the case with other causes of severe pneumonia and ards, 17 and hypoxemia alone, even if severe, is not seen as a valid reason to intubate a patient for invasive ventilation. awake covid-19 patients requiring > 2 l/minute of oxygen to maintain spo 2 > 94% or with a pao 2 /fio 2 < 200 mmhg are considered candidates for awake prone positioning when there is no contraindication. 18 this approach may improve oxygenation without the need for additional resources and is of immense value during a surge of covid-19 patients, especially in resource-limited settings. however, because awake prone positioning may not be tolerated for a prolonged period of time, we frequently use a lateral recumbent or three-quarters prone (recovery) position 19 for 2-4 hours, alternating with prone positioning. intubation is not to be delayed until the patient acutely decompensates, with spontaneous vigorous inspiratory effort that may cause self-induced lung injury. 20 we have a low threshold to intubate those who fail to improve or rapidly progress over a few hours despite oxygen supplementation, or who develop hypercapnia, hemodynamic instability, or multi-organ failure (figure 1) . invasive ventilation in patients with critical covid-19 differs in several aspects from that in patients with ards from other causes. one important difference is the coexistence in covid-19 of severely affected noncompliant lung tissue adjacent to relatively compliant unaffected areas. whereas affected areas cannot be opened, or are very difficult to open with recruitment maneuvers and higher positive endexpiratory pressure (peep), unaffected areas are at risk of overdistension by such pressure levels. in these patients, strategies to prevent atelectrauma with high levels of peep could in fact be harmful. 21 however, this may not be true for all patients. it has been suggested that a subset of patients may have recruitable covid-19 lesions that respond to higher peep. 22, 23 this may be evaluated with a chest ct scan at two different peep levels, although this approach is difficult in settings where resources are limited. our initial response to intubated patients with covid-19 is to set peep at ∼10 cm h 2 o and to do a trial with higher peep only in cases with a high driving pressure. ventilation is continued with higher peep only when this results in a drop in the driving pressure. fluid management. a restrictive fluid strategy may be important to avoid aggravation of pulmonary edema. in our patients, if organ perfusion is appropriate, fluid boluses are avoided. it has been suggested that some patients develop the so-called cytokine storm or hyperinflammatory phase, in which hypotension and hypoperfusion may respond well to fluid administration. we continue a restricted fluid approach also in cases of hypotension ( figure 2 ). early application of vasopressors concurrent with proper fluid resuscitation is used to provide good tissue perfusion with limited fluid therapy. loop diuretics are recommended in case the fluid balance becomes too positive, and renal replacement therapy may be necessary to correct the fluid balance in some patients. secondary infections. covid-19 patients frequently need prolonged intubations and stays in the icu, and thus may develop typical nosocomial complications such as ventilatorassociated pneumonia and sepsis. in this setting, these infectious complications are diagnosed and treated as for any hospitalized patient. clinical trials evaluating the effects of favipiravir, hydroxychloroquine, chloroquine, lopinavir or darunavir plus ritonavir, remdesivir, and several other compounds in covid-19 are planned or currently underway. 16 currently, there is no evidence from randomized clinical trials that any of these adjunctive therapies improve outcomes in patients with severe covid-19. several of these therapies have been included in the thai national guidelines, 8 albeit at a limited level (tables 3 and 4 ). available evidence may be insufficient to make a recommendation, as is a usual practice in highincome countries. antiviral and antimalarial strategies. in thailand, awaiting definitive results of randomized clinical trials, critically ill covid-19 patients receive combination therapies with at least three different mechanisms of action, including favipiravir for 10 days, depending on clinical symptoms. because of availability and affordability, the combination typically includes favipiravir, chloroquine or hydroxychloroquine, and darunavir/ritonavir or lopinavir/ritonavir. adding azithromycin is optional. 24 patients are closely monitored for side effects such as diarrhea, nausea, and hepatitis, as well as potential drug interactions. patients who receive darunavir/ritonavir or lopinavir/ ritonavir in combination with azithromycin for more than 5 days are monitored by daily electrocardiogram (ecg). if the ecg reveals a qtc > 480 milliseconds, discontinuation of darunavir/ritonavir, lopinavir/ritonavir, or azithromycin should be considered. because favipiravir may be teratogenic, this agent should not be given to women in the reproductive age. 8 immunomodulatory therapies. immunomodulatory therapies with promising results are corticosteroids, 25 a monoclonal antibody interleukin 6 (il-6) receptor antagonist, 26 immunoglobulin therapy, 27, 28 and cytokine adsorption therapy. 29 the rationale for their use is that the underlying pathophysiology of significant organ damage in the lungs and other organs is caused by a cytokine storm. 30 the potential benefit of corticosteroid therapy 16,31 may be outweighed by its adverse effects, including delayed viral clearance 32 and an increased risk of secondary infections. despite the finding that administration of methylprednisolone was associated with a decreased risk of death in one retrospective study of covid-19 patients in china, 25 in thailand, routine use of corticosteroids is not recommended. 33 in cases with progressive opacities on cxr where bacterial infection is considered unlikely, we consider methylprednisolone for 3-5 days (figure 2 ). it is to be used with caution and stopped immediately in case of suspected or confirmed bacterial superinfection. tocilizumab (tcz) is a monoclonal antibody against the il-6 receptor that possibly mediates sars-cov-2-associated inflammation. tocilizumab has been used to treat rheumatoid arthritis and approved by the fda for treating cytokine release syndrome. 34 in one retrospective study in china, in covid-19 patients with respiratory distress, hypoxemia, or requiring icu support, a single dose of tcz was associated with clinical improvement. 35 because of costs and limited availability, tcz is only occasionally used in thailand. blood purification therapy may have beneficial effects. 29 in thailand, hemoperfusion with cytokine adsorbent is sporadically used in patients who were unresponsive to or had contraindication for corticosteroids ( figure 2) . anticoagulant therapy. with emerging evidence that peripheral thrombosis and pulmonary embolism are very common in covid-19 patients and that microthrombi may be responsible for much of the pathophysiology, 36 we strongly favor the use of low molecular weight heparin as a prophylactic therapy. in case of confirmed peripheral thrombosis and pulmonary embolism, aggressive anticoagulant therapy should be started immediately. 6 (46) course of interleukin 6 inhibitor (tocilizumab), n (%) 3 (23) hemoperfusion with cytokine absorber, n (%) 3 (23) intensive care unit length of stay (days) 17 ± 9 mortality at the longest follow-up, n (%) 0 (0) * all patients received combined antiviral medications as clinical practice guideline for covid-19 in thailand, and none required extracorporeal membrane oxygenation. the covid-19 pandemic represents the greatest global public health crisis of our generation. to date, no specific therapies have been shown effective. although public health policies aimed at preventing new covid-19 cases, including assistance from the general public, are far more important than advanced medical technologies, governments, hospital administrators, and policy-makers must collaborate with icu practitioners to tackle the challenges of icu care for covid-19. the approach presented here may serve as an example for other countries and regions facing similar restrictions in care. who director-general's opening remarks at the media briefing on covid-19-1 covid-19 dashboard by the early transmission patterns of coronavirus disease 2019 (covid-19) in travellers from wuhan to thailand case report: walking pneumonia in novel coronavirus disease (covid-19): mild symptoms with marked abnormalities on chest imaging journey of a thai taxi driver and novel coronavirus the severe acute respiratory syndrome the importance of diagnostic testing during a viral pandemic: early lessons from novel coronavirus disease (covid-19) guidelines for health care and public health workers the new york times, 2020. coronavirus map: tracking the global outbreak. the new york times. available at favipiravir versus arbidol for covid-19: a randomized clinical trial detection of sars-cov-2 in different types of clinical specimens coronavirus disease 2019 (covid-19): a systematic review of imaging findings in 919 patients novel coronavirus pneumonia emergency response epidemiology team, 2020. the epidemiological characteristics of an outbreak of 2019 novel coronavirus diseases (covid-19) in china early lessons on the importance of lung imaging in novel coronavirus disease (covid-19) diagnosing acute respiratory distress syndrome in resource limited settings: the kigali modification of the berlin definition the search for optimal oxygen saturation targets in critically ill patients: observational data from large icu databases guidance for prone positioning of the conscious covid-19 patient 2020 erc guidelines recommended to continue monitoring the patient placed in the recovery position mechanical ventilation to minimize progression of lung injury in acute respiratory failure respiratory support in novel coronavirus disease (covid-19) patients, with a focus on resource-limited settings covid-19 pneumonia: different respiratory treatments for different phenotypes? management of covid-19 respiratory distress hydroxychloroquine and azithromycin as a treatment of covid-19: results of an open-label non-randomized clinical trial risk factors associated with acute respiratory distress syndrome and death in patients with coronavirus disease 2019 pneumonia in wuhan, china clinical course and risk factors for mortality of adult inpatients with covid-19 in wuhan, china: a retrospective cohort study treatment of 5 critically ill patients with covid-19 with convalescent plasma high-dose intravenous immunoglobulin as a therapeutic option for deteriorating patients with coronavirus disease 2019 potential effect of blood purification therapy in reducing cytokine storm as a late complication of severe covid-19 covid-19: consider cytokine storm syndromes and immunosuppression clinical evidence does not support corticosteroid treatment for 2019-ncov lung injury factors associated with prolonged viral rna shedding in patients with covid-19 management of critically ill adults with covid-19 cytokine release syndrome effective treatment of severe covid-19 patients with tocilizumab abnormal coagulation parameters are associated with poor prognosis in patients with novel coronavirus pneumonia key: cord-308680-moligska authors: wong, justin; chaw, liling; koh, wee chian; alikhan, mohammad fathi; jamaludin, sirajul adli; poh, wan wen patricia; naing, lin title: epidemiological investigation of the first 135 covid-19 cases in brunei: implications for surveillance, control, and travel restrictions date: 2020-08-14 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0771 sha: doc_id: 308680 cord_uid: moligska studies on the early introduction of sars-cov-2 in a naive population have important epidemic control implications. we report findings from the epidemiological investigation of the initial 135 covid-19 cases in brunei and describe the impact of control measures and travel restrictions. epidemiological and clinical information was obtained for all confirmed covid-19 cases, whose symptom onset was from march 9 to april 5, 2020. the basic reproduction number (r0), incubation period, and serial interval (si) were calculated. time-varying r was estimated to assess the effectiveness of control measures. of the 135 cases detected, 53 (39.3%) were imported. the median age was 36 (range = 0.5–72) years. forty-one (30.4%) and 13 (9.6%) were presymptomatic and asymptomatic cases, respectively. the median incubation period was 5 days (interquartile range [iqr] = 5, range = 1–11), and the mean si was 5.4 days (sd = 4.5; 95% ci: 4.3, 6.5). the reproduction number was between 3.9 and 6.0, and the doubling time was 1.3 days. the time-varying reproduction number (rt) was below one (rt = 0.91; 95% credible interval: 0.62, 1.32) by the 13th day of the epidemic. epidemic control was achieved through a combination of public health measures, with emphasis on a test–isolate–trace approach supplemented by travel restrictions and moderate physical distancing measures but no actual lockdown. regular and ongoing testing of high-risk groups to supplement the existing surveillance program and a phased easing of physical distancing measures has helped maintain suppression of the covid-19 outbreak in brunei, as evidenced by the identification of only six additional cases from april 5 to august 5, 2020. the global spread of covid-19 and the lack of an effective vaccine or therapeutic options pose challenges for disease control 1 and travel health. 2 importation events in a country with no cases can lead to an exponential increase in case numbers within a short time period. 3 as such, countries have implemented travel restrictions in response to the global rise; however, their effectiveness is debatable. 4, 5 studies on the early introduction of the virus in a naive population can provide insight into the natural history of the disease and have implications for control measures. brunei darussalam (pop. 459,500), a well-connected country vulnerable to multiple importation events, detected its first covid-19 imported case on march 9, linked to an international super spreading event in kuala lumpur, malaysia: the sri petaling mosque tablighi jamaat cluster. this 4-day event (february 27-march 1, 2020) was attended by more than 16 ,000 members of the tablighi, an apolitical islamic group, from various parts of the world, including brunei. 6, 7 given the absence of widespread community transmission and a slowing in the trajectory of spread since the 100th confirmed case, brunei's response to this first wave of the pandemic has been generally successful. 8 to maintain this, a number of questions need to be addressed. first, what were the epidemiological characteristics of the cases observed so far? second, what was the role of travel-related cases in driving the disease? third, how infectious were the cases and how effective were the approaches to reducing transmission? here, we report findings from the first 135 covid-19 cases, detected within the first 5 weeks of the local epidemic, along with their epidemiological, clinical, and transmission characteristics. as jurisdictions that have implemented lockdowns begin to bring the epidemic under control, 9 our findings will be important to calibrate detection and response efforts in potential future waves of the pandemic. case identification and contact tracing. the brunei surveillance and contact tracing strategy has previously been described. 7 since january 23, clinical and laboratory surveillance has been implemented across the country, and testing criteria have progressively expanded in scope (supplemental table 1 ). a confirmed case is someone who tested positive for sars-cov-2 through real-time reverse transcriptase-pcr (rt-pcr) test on nasopharyngeal (np) swab. all laboratory-confirmed covid-19 cases with symptom onset from march 9 to april 5, 2020 were included in this study and followed up until recovery or death. epidemiological investigation was conducted for each confirmed case, and information was collected on demographic characteristics, clinical symptoms, travel history, activity mapping 2 days before the onset of symptoms (or swab date for asymptomatic cases), and contact history. a close contact was any person living in the same household, or someone within 1 m of a confirmed case in an enclosed space for more than 15 minutes. all close contacts underwent rt-pcr testing. those who tested negative were quarantined at home for 14 days from last exposure, and those who later developed symptoms were retested. all confirmed covid-19 cases were treated and isolated at the national isolation centre. cases were discharged following two consecutive negative sars-cov-2 np swabs collected at ³ 24-hour intervals. we categorized cases into two groups: imported cases (defined as individuals presumed to have acquired the infection outside brunei) and locally transmitted cases (defined as those without a travel history). statistical analysis. an epidemic curve was constructed based on the date of symptom onset (for symptomatic and presymptomatic cases) or the date of np swab collection (for asymptomatic cases). the duration between symptom onset to diagnosis date was calculated. the exposure period for imported cases and their close contacts was calculated as the duration between their return date to brunei and their diagnosis/swab collection date. the incubation period was calculated as the duration between the known exposure date of confirmed cases and their symptom onset dates. the serial interval (si) was calculated as the duration between the symptom onset dates for the primary case and that of a secondary case. only symptomatic or presymptomatic infectorinfectee pairs with clear epidemiological links were included in the si calculation. group comparison was performed between the imported and local contact cases, using chi-square, fisher's exact, or mann-whitney's tests as appropriate. the estimation of the reproduction number (r0) (defined as the expected number of secondary cases infected by a primary case) was performed using two methods. the basic r0 was estimated from the mean si and the exponential growth rate of the cumulative number of cases in the epidemic λðtþ = lnðy½tþ=t, using the formula r0 = 1 + siλ + fð1 à fþðsiλþ 2 , where f is the ratio of the infectious period to the si. this assumes an exponential distribution, allowing for a range of reported values. 10 the exponential growth phase between march 5 and 10 was chosen for the r0 estimation because it represents the initial growth of the epidemic curve (see figure 1 ), and also because control measures were enhanced after march 9. thus, this 6-day period represented more accurately the nature of sars-cov-2 transmission, in the absence of heightened measures. the epidemic growth rate and doubling time were also calculated, based on established formulas. 11 we also estimated the time-varying reproduction number (rt) to assess the effectiveness of epidemic control measures, based on methods proposed by thompson et al. 12 and using the epiestim 2.2 package. this rt estimation uses case incidence data and si distribution, and distinguishes between imported and locally transmitted cases. following studies that use a nonnegative si distribution to approximate the distribution of the generation time, 13, 14 we used a gamma distribution and incorporated uncertainty in the parameters (sd of mean = 1, sd = 0.5). the median rt and 95% credible intervals for each day were estimated jointly from incidence data and from the posterior si distribution, using a 6-day sliding window. this 6-day window reduces the bias associated with early estimations of rt, as at least one average si has passed. 15 all analyses were conducted using microsoft excel (microsoft corp., redmond, wa) and r (ver. 3.6.3, r core team, vienna, austria). 16 a p-value < 0.05 was considered as statistically significant. ethics approval was granted by the university research ethics committee, universiti brunei darussalam (ref: ubd/oavcr/urec/apr2020-05). a total of 135 cases were detected in the first 5 weeks, comprising 53 (39.3%) imported and 82 (60.7%) locally transmitted cases ( table 1 ). the median age was 36 years (ranging from 6 months to 72 years). fifty-three (39.3%) cases were female. locally transmitted cases were significantly younger (p = 0.002) than imported cases and tend to be females (p = 0.008). eighty-one (60%) cases developed symptoms, reported either during or before np sample collection. notably, we observed high proportions of presymptomatic and asymptomatic cases-41 (30.4%) and 13 (9.6%), respectively. in total, 61.5% of the cases were detected within 2 days of symptom onset or np swab date. among them, 32.6% (n = 44) were detected on or before the day of symptom onset. no significant differences were observed between the imported and locally transmitted cases ( table 1 ). the most common reported symptoms were fever (62.2%), sore throat (62.2%), and cough (59.3%) (supplemental table 2 ). among these 135 cases, three subsequently died from covid-19 complications, giving a case fatality rate of 2.2%. all three deaths were in men, aged 64, 56, and 67 years, respectively. impact of travel restrictions. since late-january, travel restrictions were progressively implemented in response to the emerging regional and later global situation. initially, travelers from hubei province, china, were restricted, whereas those from other parts of mainland china underwent 14 days home quarantine. these restrictions were gradually tightened, first to travelers from iran and italy (the emerging epicenters) and then mandating quarantine for travelers from china and south korea. these restrictions had a considerable impact on arrivals into brunei. immigration data indicate a 20.9% decrease in arrivals from january to march 2020 (872,315 people), compared with the same period in 2019 (1,103,028 people). outbound travel was restricted for all brunei residents on march 15, and a ban on all foreign citizens entering the country was enacted on march 23. beginning on march 20, all the origin country of imported cases changed over time. cases were initially from malaysia (20 cases), and, as the epidemic progressed globally, and overseas brunei citizens were returning, imported cases were identified in travelers and returning residents from indonesia (n = 14), united kingdom (n = 11), thailand (n = 2), the united states (n = 2), austria (n = 1), cambodia (n = 1), australia (n = 1), and the philippines (n = 1). figure 1 shows the epidemic curve, by the date of symptom onset or np swab date for asymptomatic cases. detection of positive cases among local close contacts occurred very early in the epidemic. coupled with the early detection of further generations (supplemental figure 1 ), this suggests a short time interval for transmission within the community. epidemic characteristics over time. the mean duration from symptom onset to diagnosis for local transmitted cases decreased from 9 days in the first week to −1.7 days in the fifth week of the epidemic (figure 2a ). among imported cases, this reduction occurred between the third and fourth weeks of the epidemic (from 7.3 to 1.3 days, respectively), coinciding with implementation of quarantine and testing of all arrivals beginning march 20 (figure 2a) . this demonstrates the impact of increasingly stringent travel restrictions. there were no local infections linked to imported cases in the last 2 weeks of the epidemic. using the known return dates from imported cases (n = 53), the median duration in the exposure period to other contacts was 7 days (iqr = 5), ranging between 1 and 16 days. the reduction in the median exposure period between imported cases and their close contacts from their return to diagnosis was apparent between the third (8.5 days) and fourth weeks (4.0 days; figure 2b ). summary statistics and distribution remained unchanged when the 15 local contact cases with known exposure dates were included. among 82 local contact cases, 15 of them (18.3%) had known dates of exposure to confirmed imported cases. using these dates, the median incubation period was 5.0 days (iqr = 5), ranging between 1 and 11 days. based on 59 symptomatic and presymptomatic infectorinfectee pairs, the mean si was 5.4 days (sd = 4.5; 95% ci: 4.3, 6.5 [approximated using normal distribution]). the range for the si was between −4 and 20 days ( figure 2c ). four pairs (6.8%) had negative si values. the median si was relatively constant throughout the 4 weeks of the epidemic ( figure 2d ). using the calculated mean si and the 6-day growth phase of the epidemic, the growth rate was 0.54/day, and the r0 in the early phase of the epidemic ranges between 3.9 and 6.0. the doubling time was 1.3 days. figure 3 shows the estimated rt and the timing of the control measures implemented after march 11. the initial median reproduction number was estimated to be 2.2 (95% credible intervals: 0.86, 5.0) on the seventh day of the epidemic (march 11). the time-varying reproduction number gradually decreased after several control measures were put in place and was below one on the 13th day (march 17) and the bold values indicate findings with p-value is < 0.05. * cases were classified as follows: 1) symptomatic, if symptoms were reported on or before np swab collection day; 2) presymptomatic, if symptoms were reported after np swab sample was taken but during admission; and 3) asymptomatic, if no symptoms were reported since np swab collection day until the date of hospital discharge. † this includes the asymptomatic cases, from whom the symptom onset date was replaced by the date of swab collection. ‡ severity was classified as 1) asymptomatic, for those with no symptom throughout their disease; 2) mild, for patients who had uncomplicated upper respiratory tract infection symptoms and no radiological changes; 3) moderate, for patients with radiological changes but did not require supplemental oxygen; 4) severe, for patients who showed signs of severe pneumonia including tachypnea > 30/minute, spo 2 of £ 93% on room air, or abnormal arterial blood gases, as well as patients showing signs of sepsis with evidence of organ dysfunction; and 5) critical, for patients who developed septic shock, that is, persistent hypotension requiring vasopressors support to maintain mean arterial pressure ³ 65 and those who developed acute respiratory distress syndrome requiring ventilatory support. reached 0.20 (95% credible intervals: 0.06, 0.49) on the 30th day (april 3). however, because these control measures were implemented quickly over a short time period, it is difficult to attribute the observed reduction in rt to a specific intervention. key findings. a total of 135 cases (53 imported and 82 locally transmitted) were reported from the start of the epidemic on march 5 until the first week of april. all locally transmitted cases could be traced to an importation event, and there were no cases detected without a clear epidemiological link. brunei managed to successfully control the first wave of the covid-19 pandemic. the r0 was between 3.9 and 6.0, and the epidemic had a doubling time of 1.3 days during the exponential phase. there was a rapid decline such that by day 13, the rt was under one. estimating the reproduction number. assuming the ratio of the infectious period to the si (f) is 0.3, the r0 was 5.6, higher than those estimated by the who (1.9-2.5), but within those estimated in other studies from china [17] [18] [19] and europe. 20 our observations of negative sis pose challenges for specifying f. hence, we reported r0 as a range covering all plausible values of f, using the mean si value from our data and with assumption of exponential growth. then, rt was estimated from the 7th to the 30th day. the 95% credible intervals of the initial rt value (0.86-5.0) are consistent with those of the r0 estimate. test, isolate, and trace. we observed that the approach of isolation of confirmed cases, as well as tracing and quarantine of their contacts, was associated with a significant reduction in the effective reproduction number. by the time the early physical distancing measures had been implemented, the rt had reduced from 5.6 (ranging between 3.9 and 6.0) at the start to 2.2 (95% credible bands: 0.86, 5.0) by the seventh day (march 11). our findings on the importance of the test, isolate, and trace approach are consistent with observational studies from hong kong 21 and singapore. 22 we highlight three characteristics of the approach implemented in brunei. first, testing was conducted on a significant scale. even before detection of the first case, testing was already conducted for all inpatient pneumonia cases, in addition to those who met the suspect case definition; 24-hour testing centers were established within 2 days of the detection of the first case (march 11) , and all contacts were tested regardless of symptom status. 8 the relatively high proportion of asymptomatic (9.6%) and presymptomatic (30.4%) cases identified demonstrates both the breadth of testing and the speed at which cases were identified. second, all confirmed cases (regardless of disease severity and symptom status) were isolated in a dedicated isolation facility and remained isolated until two negative results were obtained from rt-pcr specimens at 24-hour intervals. this reduced the chance of household transmission, which could not be excluded if cases were allowed to isolate at home. 23 finally, contact tracing was conducted for each case, using a variety of tools including case interview, workplace assessment, and mobile phone data. contacts were placed on 14-day home quarantine, and inperson spot checks with penalties for noncompliance were also conducted. travel restrictions and other non-pharmaceutical interventions. we report several characteristics of sars-cov-2 that make effective isolation and contact tracing challenging, including high transmissibility, a relatively short si (mean si = 5.39 days), and a high proportion of asymptomatic and presymptomatic cases suggesting the potential for silent transmission. as such, the test, isolate, and trace approach was supplemented with physical distancing measures to increase the likelihood of achieving sustained control. 24 restricting travel is one measure by which countries have responded to the covid-19 pandemic. 25 by the time brunei implemented an exit travel ban and restricted the entry of foreign citizens in the country, the rt was already decreasing and had neared one. nonetheless, we suggest that reducing ongoing vulnerabilities to importation events, through restrictions on incoming travelers, and requirements for testing and quarantine for all arrivals in the country limited additional spread, consistent with a reduction in mean time from symptom onset to diagnosis observed for imported cases following implementation of mandatory quarantine and testing for all arrivals. modeling studies suggest a role for travel restrictions in containing the epidemic, with one model estimating that travel restrictions in wuhan reduced case importations elsewhere by nearly 80% through mid-february. 26 although various other physical distancing measures were implemented in brunei, including school closures; prohibition on mass gatherings, cinemas, and religious services; and dine-in restrictions, importantly, no lockdown was implemented, and there were no generalized stay-at-home orders. most businesses and government agencies were able to operate. these suggest that effective test, isolate, and trace approaches were able to control the epidemic with moderate levels of physical distancing. 27 this finding corroborates the experience of other countries. in hong kong, case isolation and contact tracing were combined with other physical distancing measures (but no lockdown), which resulted in an estimated effective reproduction number near 1 for 8 weeks. 28 in south korea, testing and tracing have been combined with school closures and remote working. 29 even with the best efforts at testing, case identification, and quarantine, the potential for widespread community transmission of covid-19 is clear. once the disease is established, suppression may require the implementation of severely disruptive social distancing measures. 30, 31 limitations. our study had several limitations. first, although we can be reasonably confident of having identified most cases since march, given more restrictive testing criteria in january and february, we were unable to account for potential importation events that may have occurred before detection of the first case. second, the generalizability of our results are limited because of lack of community transmission, small number of cases, and a lack of cases in settings such as residential care facilities and dormitories. third, due to the potential for presymptomatic infection of sars-cov-2, using the si distribution to approximate the generation time distribution is problematic. we had not accounted for negative sis with the use of the gamma distribution, and thus may have overestimated rt to fit incidence data. one way to account for negative si is to use a deconvolution approach using the incubation period distribution to recover the generation time distribution. 32 however, this assumes that the generation time and incubation period distributions are independent, which may not be appropriate. finally, given the limited data available and the analytic methods used, we could not directly estimate the effectiveness of other non-pharmaceutical interventions, such as face mask wearing, personal hygiene practices, and voluntary reductions in mobility. swift control of covid-19 in brunei was achieved through a combination of public health measures, focusing on a testisolate-trace approach supplemented by travel restrictions and general physical distancing measures, but no actual lockdown. as of august 5, 2020, brunei has recorded a total of 141 confirmed covid-19 cases, with the last case reported on may 6. on may 18, brunei commenced its de-escalation plan in a stepwise approach including reopening of mosques, schools, and other indoor facilities in phases, while still maintaining its border control and enhancing the surveillance program (including testing of high-risk groups such as those working in residential institutions, healthcare workers, and also individuals at relatively high risk of developing severe complications). in conjunction with easing of physical distancing restrictions, a mobile application was rolled out on may 14 to speed up the process of contact tracing. its public uptake among the general adult population was > 90%. along with timely reimposition of physical distancing measures, if necessary, these can help maintain suppression of a second wave in brunei. e-mail: sirajul.jamaludin@moh.gov.bn. wan wen patricia poh, department of dental services, ministry of health covid-19: what is next for public 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outbreaks estimation of the time-varying reproduction number of covid-19 outbreak in china evolving epidemiology and transmission dynamics of coronavirus disease 2019 outside hubei province, china: a descriptive and modelling study a new framework and software to estimate time-varying reproduction numbers during epidemics r: a language and environment for statistical computing estimation of the transmission risk of the 2019-ncov and its implication for public health interventions effects of media reporting on mitigating spread of covid-19 in the early phase of the outbreak modelling the epidemic trend of the 2019 novel coronavirus outbreak in china. biorxiv monitoring transmissibility and mortality of covid-19 in europe suppressing covid-19 transmission in hong kong: an observational study of the first four months evaluation of the effectiveness of surveillance and containment measures for the first 100 patients with covid-19 in singapore feasibility of controlling covid-19 outbreaks by isolation of cases and contacts early dynamics of transmission and control of covid-19: a mathematical modelling study estimating covid-19 outbreak risk through air travel the effect of travel restrictions on the spread of the 2019 novel coronavirus (covid-19) outbreak reduction in effective reproduction number of covid-19 is higher in countries employing active case detection with prompt isolation impact assessment of non-pharmaceutical interventions against coronavirus disease 2019 and influenza in hong kong: an observational study flattening the curve on covid-19-how korea responded to a pandemic using ict only strict quarantine measures can curb the coronavirus disease (covid-19) outbreak in italy the positive impact of lockdown in wuhan on containing the covid-19 outbreak in china estimating effective reproduction number using generation time versus serial interval, with application to covid-19 in the greater toronto area this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-287247-vv0zc0gd authors: gutman, julie r.; lucchi, naomi w.; cantey, paul t.; steinhardt, laura c.; samuels, aaron m.; kamb, mary l.; kapella, bryan k.; mcelroy, peter d.; udhayakumar, venkatachalam; lindblade, kim a. title: malaria and parasitic neglected tropical diseases: potential syndemics with covid-19? date: 2020-06-01 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0516 sha: doc_id: 287247 cord_uid: vv0zc0gd the covid-19 pandemic, caused by sars-cov-2, have surpassed 5 million cases globally. current models suggest that lowand middle-income countries (lmics) will have a similar incidence but substantially lower mortality rate than high-income countries. however, malaria and neglected tropical diseases (ntds) are prevalent in lmics, and coinfections are likely. both malaria and parasitic ntds can alter immunologic responses to other infectious agents. malaria can induce a cytokine storm and pro-coagulant state similar to that seen in severe covid-19. consequently, coinfections with malaria parasites and sars-cov-2 could result in substantially worse outcomes than mono-infections with either pathogen, and could shift the age pattern of severe covid-19 to younger age-groups. enhancing surveillance platforms could provide signals that indicate whether malaria, ntds, and covid-19 are syndemics (synergistic epidemics). based on the prevalence of malaria and ntds in specific localities, efforts to characterize covid-19 in lmics could be expanded by adding testing for malaria and ntds. such additional testing would allow the determination of the rates of coinfection and comparison of severity of outcomes by infection status, greatly improving the understanding of the epidemiology of covid-19 in lmics and potentially helping to mitigate its impact. the covid-19 pandemic caused by sars-cov-2, a novel coronavirus, has now reached all corners of the world, and cases have surpassed 5 million. 1 sars-cov-2 is currently spreading in low-and middle-income countries (lmics) that experience the highest rates of malaria and neglected tropical diseases (ntds). neglected tropical diseases refer to a diverse group of communicable diseases caused by parasites, fungi, bacteria, and viruses that occur primarily in tropical and subtropical climates; only parasitic ntds are considered here (table 1) . with many lmics implementing movement restrictions or ordering their populations to stay at home to limit sars-cov-2 transmission, the threat to essential health services is likely to be immediate, causing delays to diagnosis and treatment for other diseases, including malaria and ntds. during the ebola epidemic in west africa, there were substantial reductions in all-cause outpatient visits and patients treated with antimalarial drugs 2 ; modeling the potential for similar disruptions in malaria control due to covid-19 suggests that there could be up to an estimated 769,000 deaths due to malaria in 2020 (approximately double the number seen in 2018), mostly among children younger than 5 years. 3 countries working toward the elimination of malaria or ntds may face setbacks. less obvious, but potentially important, is the possibility of sars-cov-2 interacting with parasitic infections and changing the rate of severe outcomes, particularly among younger populations that have been relatively less affected by covid-19 to date. 4 under the assumption that public health and social distancing measures are used to mitigate the epidemic, the modeled estimates for sars-cov-2 infection incidence rates for lmics, assuming comorbidity rates for all countries similar to what was seen in wuhan, china, are projected to be around 600 infections per 1,000 population, similar to the rate anticipated for high-income countries. however, the mortality rate for lmics (∼2 per 1,000) is projected to be about half that of the high-income countries (∼4 per 1,000). 5 the difference in predicted mortality rates between lmics and high-income countries is largely due to the younger age structure in lmics; in 2020, the median age in sub-saharan africa is 18.7 years, compared with 38.4 years in china. 6 syndemics, or synergistic epidemics, occur when two or more concurrent epidemics have a deleterious interaction, 7 that is, when coinfections result in a worse overall outcome than for either individual infection. there are many examples of important interactions between malaria and ntds and other infectious diseases. for example, malaria plays a role in epstein-barr virus (ebv) infection, leading to burkitt's lymphoma by contributing to b-cell proliferation and increasing ebv loads 8 ; hiv-infected individuals experience a greater frequency of severe malaria and increased hiv viral load following infection with plasmodium falciparum 9 ; several parasite-hiv coinfections are associated with increased hiv viral load and worsened immunosuppression 10 ; and schistosome infections are associated with increased transmission of hiv, 11 whereas deworming is associated with decreased hiv viral load and improved cd4 counts among hiv-infected individuals. 12 biological interactions between coinfecting pathogens could involve changes in host pathology related to indirect immune effects. 12 the interplay of coinfections hinges on several host-pathogen factors and host immunodynamics. low-and middle-income countries in africa suffer the greatest burden of malaria; in 2018, there were more than 200 million cases per year, with an annual incidence of 229 per 1,000 persons. 13 despite substantial progress in reducing malaria mortality over the past two decades, more than 400,000 malaria deaths (> 90% in sub-saharan africa) were estimated to have occurred in 2018. 13 outside of africa, india has the greatest burden of malaria cases, accounting for 3% of the global burden. 13 globally, ntds affect more than 1 billion people, especially those living in poverty, who often lack access to clean water and adequate sanitation. 14 africa has a disproportionate burden of ntds and malaria, with a significant geographical overlap. 14, 15 with rapid transmission of sars-cov-2, many people in lmics, particularly in africa, soon will be coinfected with sars-cov-2 and plasmodium spp. or one or more ntd pathogens; cases of covid-19 in the africa region will soon surpass 100,000. 1 preexisting infection with any of these parasitic infections may lead to changes in susceptibility and/or severity of covid-19. it is unclear whether immunomodulation caused by malaria and ntds will be beneficial or harmful when hosts are coinfected with sars-cov-2, but even small changes in the risk of severe outcomes due to coinfections could result in substantial changes in the impact and epidemiology of covid-19 in lmics. sars-cov-2 infection. common symptoms of infection with sars-cov-2 include fever, cough, shortness of breath, chills, myalgia, headache, sore throat, and new loss of taste or smell 16 ; the onset of symptoms generally occurs 4-5 days after infection, although it can be as late as 14 days, [17] [18] [19] and not all infected people develop symptoms. [20] [21] [22] approximately a week after the development of symptoms, some patients experience an acute worsening, with a pronounced systemic increase of inflammatory mediators and cytokines. 23 the severe systemic inflammatory response, referred to as a "cytokine storm," is characterized by markedly increased levels of interleukins (il) and tumor necrosis factor (tnf)alpha, and is associated with the development of acute respiratory distress syndrome (ards). 24 among 72,314 cases reported from china, 14% were rated as severe and 5% were critical (respiratory failure, septic shock, and multiple organ dysfunction or failure). 17 case fatality ratios (cfrs) ranged from 2.3% to 7.2%, with higher cfrs among older adults (8.0-12.8% among those aged 70-79 years and 14.8-20.2% among those 80 years and older, versus £ 0.4% among those younger than 50 years). 17, 25 hypertension, diabetes, cardiovascular disease, preexisting respiratory disease, and obesity were common comorbidities 26, 27 ; in a meta-analysis of 1,576 patients in china, all but diabetes and obesity were associated with increased risk of severe disease. 27 potential plasmodium spp.-sars-cov-2 interactions. of the five parasitic species that cause malaria in humans (table 1) , p. falciparum accounts for most morbidity and mortality, followed by plasmodium vivax. 13, 15 clinical illness arises from asexual parasite replication within erythrocytes. infected erythrocytes lyse and release merozoites into the circulation, causing activation of the immune system and leading to the release of pro-inflammatory cytokines including tnf-alpha, interferon-gamma, il-6, and il-12. 28 this cascade of cytokines leads to symptoms of uncomplicated malaria, including periodic fever, which, if left untreated, can progress to severe disease. severe disease manifests as severe anemia, respiratory failure, cerebral malaria, acidosis, and renal failure. children and infants are at greatest risk for severe malaria; 67% of malarial deaths are estimated to occur among african children younger than 5 years. 13 as with covid-19, cellular immune responses in malaria involving the cytokine cascade must be carefully regulated to achieve a protective response without causing adverse impact on the host. studies in malaria-endemic regions have found that it is important to have a balance between a host pro-inflammatory, th1 response (e.g., tnf-alpha, il-6, il-12, and interferon-gamma) and anti-inflammatory, th2 response (il-4, il-10, and others) 29, 30 ; severe manifestations of malaria are often due to excessive pro-inflammatory responses. the same appears to be true in at least some cases of covid-19, 18 suggesting that a coinfection that also leads to excess proinflammatory responses might result in more severe manifestations and poor prognosis. malaria-induced immunosuppression has also been observed in many coinfections, significantly inhibiting immune responses to the other infection (e.g., to salmonella spp.). 31, 32 however, malaria-induced immunomodulation has been shown to be protective against severe manifestations of some respiratory viruses. in kenya, hospitalized children diagnosed with influenza and malaria were less likely to experience respiratory distress than those with influenza alone. 33 coinfection with plasmodium spp. could suppress the production of pulmonary cytokines and decrease the recruitment of cellular inflammatory components to the lungs, leading to reduced clinical symptoms and inflammation, as was found during pneumovirus infections in a murine model. however, in the murine model, viral control was also impaired, leading to increased viral dissemination. 34 similar dynamics could occur during plasmodium-sars-cov-2 coinfection; malaria-induced immunosuppression might lead to milder manifestations of covid-19 but simultaneously decrease viral control, potentially increasing or sustaining viral loads, which could increase the potential for viral transmission. age-related vulnerability to malaria and covid-19. susceptibility to malaria in highly endemic areas differs by age: younger children are more vulnerable to malaria infections and at a higher risk for severe malaria. 13 for covid-19, children are less likely to develop severe disease, whereas older populations are disproportionately affected, with a higher risk of severe disease and death. 35 this may be due to the fact that children are more likely to produce t-regulatory cytokines (il-10, il-23, and il-6) and have less inflammation (because of their immature immune systems) than older people who mount a more proinflammatory cytokine cascade, potentially contributing to pathogenesis. 35 how age-related susceptibility to covid-19 will play out in africa, where many children are immunologically stimulated by several infections in addition to malaria, is not clear. importantly, malaria infections in endemic areas frequently result in chronic, afebrile disease in older children and adults. 36 it remains unknown whether this underlying infection will alter susceptibility to or severity of covid-19 in these populations; it is important that surveillance systems be modified to collect data to inform our understanding of this issue. respiratory distress and ards. respiratory distress, observed in up to 25% of adults and 40% of children with severe p. falciparum malaria, has several causes, including severe anemia, metabolic acidosis, cytoadherence of infected erythrocytes in pulmonary vasculature, and coinfections with pneumonia-causing pathogens. 37 the clinical spectrum varies from mild upper respiratory symptoms to acute lung injury and fatal ards. acute respiratory distress syndrome is rare in young children with malaria but occurs in 5-25% of adults and 29% of pregnant women with severe p. falciparum infections, and less commonly with p. vivax malaria. 37 in both malaria and covid-19, ards is linked to inflammatory cytokine-mediated increased capillary permeability or endothelial damage, which results in major alveolar damage. [38] [39] [40] given this situation, plasmodium spp.-sars-cov-2 coinfections may result in particularly rapid deterioration, with a poor prognosis. as the inflammatory-mediated alveolar damage in malaria-induced ards progresses even after treatment and parasite clearance, 37 coinfected individuals may be prone to severe covid-19. because both malaria and covid-19 can lead to similar clinical manifestations, including fever and respiratory symptoms, one or the other may be overlooked in a differential diagnosis of respiratory distress, leading to increased fatalities. as sars-cov-2 transmission increases in lmics, particularly in africa and india, clinicians should keep this in mind. in addition, documenting the frequency, distribution, and outcomes of these coinfections is important. anemia. anemia is highly prevalent in lmics and results from multiple causes. in cross-sectional household surveys in sub-saharan africa, 61%, 33%, and 3% of children younger than 5 years had any anemia, moderate anemia, and severe anemia, respectively. 13 more than one-fifth of children with malaria develop sma, with a cfr of 8.4%. 41 whereas the hematologic sequelae of covid-19 are still being elucidated, a meta-analysis describing 1,210 covid-19 patients from four studies found that hemoglobin values were 0.71 g/dl (95% ci: 0.59-0.83 g/dl) lower in individuals with severe disease versus milder disease. 42 whether lower hemoglobin is a risk factor or a sequela of severe covid-19 disease is unknown. however, because of limited reserves, even small perturbations in oxygen-carrying capacity in individuals with preexisting malarial anemia may result in insufficient tissue oxygenation in the midst of covid-19-induced respiratory failure. pro-coagulant state. numerous viral infections, including sars-cov-2, induce a pro-coagulant state through the induction of tissue factor expression, endothelial dysfunction, von willebrand factor elevation, and toll-like receptor activation. 43, 44 markers of a hypercoagulable state, including increased d-dimer and fibrin degradation product levels, and prolonged prothrombin time are associated with a poor prognosis. 45 clinically, the hypercoagulable state manifests with a high rate of venous thromboembolism and arterial thrombotic complications (including pulmonary embolism and stroke). 46,47 covid-19 patients are at risk for developing disseminated intravascular coagulation (dic), 45, 48 and autopsy findings have included both pulmonary hemorrhage and thrombosis. 49 thrombocytopenia is another potential feature of covid-19, thought to be due to excessive activation of the coagulation cascade, leading to platelet activation and subsequent consumption, 44 and is associated with worse outcomes. 50 malaria is also associated with a pro-coagulant state, with activation of the coagulation cascade, mediated by tnf-alpha and il-6, proportional to disease severity. 51 whereas microthrombotic complications are most commonly described, thrombosis of large vessels, including cerebral venous thrombosis, and pulmonary embolism have been described. 52, 53 thrombocytopenia develops in 60-80% of malaria cases. 51 although bleeding and dic are rarely seen, occurring only in severe malarial cases accompanied by coagulopathy, 54 they are associated with high mortality. 51 lysis of activated platelets, along with tissue factor released from damaged vascular endothelial cells, promotes the pro-coagulant state, 54 similar to the proposed mechanism in covid-19. thus, plasmodium spp.-sars-cov-2 coinfection could lead to even greater degrees of coagulopathy and more severe disease than with either infection alone. potential interactions between ntds and covid-19. helminths, including stool-transmitted helminths (sth), schistosomes, and filariae, typically push the immune system toward anti-inflammatory th2 pathways through a variety of regulatory mechanisms. 55, 56 protozoal parasites, such as trypanosomes or leishmania spp., are more likely to induce a th1, pro-inflammatory response. however, there are many deviations from this characterization. some helminths induce th1 responses in some stages of the life cycle (e.g., microfilariae of filarial parasites or schistosome eggs), resulting in symptomatic disease, but th2 responses in other stages (e.g., adults of both filarial parasites and schistosomes). the downregulation of the inflammatory response associated with helminths may reduce the development of immunity or response to vaccines, decrease inflammation associated with autoimmune diseases, reduce the ability to control mycobacterium tuberculosis and mycobacterium leprae coinfections, and reduce the severity of malarial coinfection. the pro-inflammatory effects of some protozoal infections may worsen the severity of some, but not all, viral infections. 55, 57 in addition, polyparasitism is quite common, and the overall impact on inflammation depends on the sequence of infections and burden of each. 58 thus, coinfection with parasitic ntds could result in altered risks and severity of clinical manifestations of sars-cov-2 infection, with the potential for decreased development of immunity with increased viral loads. the severity of covid-19 has been associated with underlying health conditions that usually occur with advancing age. several ntds, if left untreated, can result in chronic sequelae in much younger populations. for example, because acute trypanosoma cruzi infection is typically asymptomatic or results in a mild, self-limited illness, it is frequently undetected and left untreated. yet, in young or middle adulthood, 20-30% of persons chronically infected with t. cruzi develop cardiac manifestations, commonly a complex, dilated cardiomyopathy. 59 for these individuals, coinfection with sars-cov-2 could be lifethreatening. sth infections may result in anemia 60 ; if, as described previously, anemia predisposes individuals to more severe outcomes, then coinfection of sths and sars-cov-2 in children and pregnant women could be problematic. malnutrition and covid-19. chronic malnutrition is associated with both malaria 61 and ntds, 62 and is relatively common among children in sub-saharan africa as well as parts of latin america and asia. prealbumin, a marker for protein malnutrition, 63 was found to be lower on admission in patients with covid-19 who developed ards than on those who did not. 64 although lower prealbumin may be a marker for more severe disease, immunosuppression associated with undernutrition preceding infection with sars-cov-2 could exacerbate the severity of covid-19. 65, 66 undernutrition is thought to have led to excess mortality with both the 1918 and h1n1 influenza pandemics. 67, 68 given relatively high rates of undernutrition among children in lmics (12.3%), 69 an association between undernutrition and clinical severity of covid-19 could increase the proportion of severe illness above current predictions, particularly among children. although sars-cov-2 has spread globally, our understanding of the epidemiology and clinical course of covid-19 in countries with substantial burdens of malaria and ntds is just beginning, in part because community transmission generally started later in these countries and because testing for sars-cov-2 is limited in most lmics. although current predictive models suggest lower mortality rates in lmics than in high-income countries, if coinfections with malaria or parasitic ntds increase complications with sars-cov-2 infections and there is a shift in the age pattern of comorbidities to younger ages, then the burden of covid-19 in lmics may be substantially worse than predicted, and potentially higher than the burden in high-income countries. 70 if a shift to a th2 response is more common, and if that shift provides some protection from severe disease while reducing long-term immunity or increasing the time frame of viral shedding, the epidemiology of covid-19 in lmics could be substantially different from what has been seen elsewhere. rapidly developing surveillance platforms to monitor signals of sars-cov-2 coinfection with malaria or other ntds will be critical. one early indication of a potential interaction would be a shift in the age pattern of severe covid-19, with higher rates of clinical disease in children than has been observed in china, europe, or north america. however, more definitive information on coinfections and outcomes will be needed to interpret such shifts. efforts to characterize covid-19 cases in lmics, such as the who first few x cases protocol, 71 and addition of sars-cov-2 testing to influenza sentinel surveillance 72 could be expanded, based on local prevalence of malaria and ntds, to include testing for malaria and ntds. such additional testing could help determine rates of coinfection and compare severity of outcomes by infection status. additional efforts to more carefully describe the clinical impacts of coinfections can follow. these efforts are important to understanding the potential impact of covid-19 on lmics and for mitigating against the worst outcomes. who coronavirus situation report-130 effect of the ebola-virus-disease epidemic on malaria case management in guinea, 2014: a crosssectional survey of health facilities the potential impact of health service disruptions on the burden of malaria: a modelling analysis for countries in sub-saharan africa systematic review of covid-19 in children shows milder cases and a better prognosis than adults report 12: the global impact of covid-19 and strategies for mitigation and suppression united nations department of economic and social affairs population division introduction to syndemics: a critical systems approach to public and community health sickle cell trait is not associated with endemic burkitt lymphoma: an ethnicity and malaria endemicity-matched case-control study suggests factors controlling ebv may serve as a predictive biomarker for this 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laboratory-confirmed conronavirus disease 2019 -covid-net, 14 states estimation of the asymptomatic ratio of novel coronavirus infections (covid-19) estimating the asymptomatic proportion of coronavirus disease 2019 (covid-19) cases on board the diamond princess cruise ship asymptomatic and presymptomatic sars-cov-2 infections in residents of a long-term care skilled nursing facility the pathogenesis and treatment of the 'cytokine storm' in covid-19 il-19 induces production of il-6 and tnf-alpha and results in cell apoptosis through tnf-alpha case-fatality rate and characteristics of patients dying in relation to covid-19 in italy obesity in patients younger than 60 years is a risk factor for covid-19 hospital admission prevalence of comorbidities in the novel wuhan coronavirus (covid-19) infection: a systematic review and meta-analysis setting the stage: the initial immune response to blood-stage parasites a low interleukin-10 tumor necrosis factor-alpha ratio is associated with 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malaria-associated acute respiratory distress syndrome covid-19: consider cytokine storm syndromes and immunosuppression standardized data collection for multi-center clinical studies of severe malaria in african children: establishing the smac network hemoglobin value may be decreased in patients with severe coronavirus disease 2019 procoagulant activity of endothelial cells after infection with respiratory viruses coagulation disorders in coronavirus infected patients: covid-19, sars-cov-1, mers-cov and lessons from the past abnormal coagulation parameters are associated with poor prognosis in patients with novel coronavirus pneumonia incidence of thrombotic complications in critically ill icu patients with covid-19 large-vessel stroke as a presenting feature of covid-19 in the young clinical characteristics of coronavirus disease 2019 in china vander heide rs, 2020. pulmonary and cardiac pathology in african american patients with covid-19: an autopsy series from new orleans 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disease 2019 pneumonia in wuhan, china immune dysfunction as a cause and consequence of malnutrition malnutrition and infection: complex mechanisms and global impacts the 1918-1919 influenza pandemic-the indian experience population-based surveillance for 2009 pandemic influenza a (h1n1) virus in guatemala sustainable development goals: indicator 2.1.1 -prevalence of undernourishment age-dependent effects in the transmission and control of covid-19 epidemics. supplementary information. london, united kingdom: london school of hygiene and tropical medicine world health organization, 2020. the first few x cases and contacts (ffx) investigation protocol for coronavirus disease 2019 (covid-19) world health organization, 2020. operational considerations for covid-19 surveillance using gisrs report of the committee on infectious diseases this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-264140-5cxzc3z8 authors: tam, clarence c.; anderson, kathryn b.; offeddu, vittoria; weg, alden; macareo, louis r.; ellison, damon w.; rangsin, ram; fernandez, stefan; gibbons, robert v.; yoon, in-kyu; simasathien, sriluck title: epidemiology and transmission of respiratory infections in thai army recruits: a prospective cohort study date: 2018-09-04 journal: am j trop med hyg doi: 10.4269/ajtmh.18-0219 sha: doc_id: 264140 cord_uid: 5cxzc3z8 military recruits are at high risk of respiratory infections. however, limited data exist on military populations in tropical settings, where the epidemiology of respiratory infections differs substantially from temperate settings. we enrolled recruits undertaking a 10-week military training at two royal thai army barracks between may 2014 and july 2015. we used a multiplex respiratory panel to analyze nose and throat swabs collected at the start and end of the training period, and from participants experiencing respiratory symptoms during follow-up. paired sera were tested for influenza seroconversion using a hemagglutinin inhibition assay. overall rates of upper respiratory illness and influenza-like illness were 3.1 and 2.0 episodes per 100 person-weeks, respectively. a pathogen was detected in 96% of samples. the most commonly detected microbes were haemophilus influenzae type b (62.7%) or non–type b (58.2%) and rhinovirus (22.4%). at baseline, bacterial colonization was high and included h. influenzae type b (82.3%), h. influenzae non–type b (31.5%), klebsiella pneumoniae (14.6%), staphylococcus aureus (8.5%), and streptococcus pneumoniae (8.5%). at the end of follow-up, colonization with h. influenzae non–type b had increased to 74.1%, and s. pneumoniae to 33.6%. in the serology subset, the rate of influenza infection was 3.4 per 100 person-months; 58% of influenza infections resulted in clinical disease. our study provides key data on the epidemiology and transmission of respiratory pathogens in tropical settings. our results emphasize the need for improved infection prevention and control in military environments, given the high burden of illness and potential for intense transmission of respiratory pathogens. military recruits are at high risk of respiratory infections. 1 the congregation of individuals from diverse geographic locations in semi-closed settings, together with high levels of close contact, provide conditions that favor the introduction and transmission of respiratory pathogens. 2 studies among military recruits have found high rates of illness and infection with respiratory viruses. a longitudinal study in singapore found that 39.4% of military personnel had evidence of seroconversion against influenza a/h1n1 during the 2009 pandemic, compared with 13.5% of individuals in the community and 6.5% of hospital staff. 3 numerous outbreaks of respiratory infections among military recruits have been reported in the literature, including outbreaks of influenza, adenovirus, 4-6 and pertussis. 7 resumption of vaccination against adenovirus types 4 and 7 in the u.s. military is estimated to have resulted in a 7-fold decrease in acute respiratory disease 8 and a shift in the predominant adenovirus types from types 4, 3, and 7 to types 1 and 2. the increasing availability of multiplex molecular diagnostic assays can yield data on multiple pathogens that would not routinely be detected. 9, 10 studies in military populations can provide valuable information regarding the epidemiology and transmission of respiratory infections in adults because of the availability of well-defined populations that can be followed up over time. despite this, data from tropical settings, in which the epidemiology of influenza and other respiratory infections differs substantially from temperate settings, are limited. increased understanding of the etiology of respiratory infections in these settings is important to identify opportunities for disease control in high-risk military populations and to identify risks of pathogen emergence with potential for spread into the wider community. we present the results of a longitudinal study of the burden and etiology of respiratory infections in thai military recruits. the study setting and procedures have been previously described. 11 between may 2014 and may 2015, we enrolled participants from consecutive cohorts of recruits undertaking basic military training at two royal thai army (rta) barracks in bangkok. trainees entered the camps at the start of may and november each year and remained in the camps for 10 weeks. the camps consist of large, common sleeping quarters with beds arranged foot-to-foot in long rows. trainees share meals in a common canteen and train in a large field in the center of the barracks and a number of ancillary buildings. each camp has its own medical unit. individuals were eligible for enrollment if they were aged ³ 18 years and were entering one of the two army barracks involved in the study. suspected tuberculosis cases or individuals with immune deficiencies, such as acquired immune deficiency syndrome, leukemia, or lymphoma, were excluded. individuals providing informed consent were enrolled in the study and a demographic and clinical questionnaire was administered. participants were then followed up for symptoms of respiratory illness until the end of the 10-week training period. at one camp, nasal and throat swabs were obtained at the time of enrolment and at the end of the follow-up period (10 weeks later) from consenting participants. participants experiencing respiratory symptoms were asked to consult the medical unit, where medical staff took a history, conducted a medical examination, and recorded symptoms of upper respiratory illness (uri) or influenza-like illness (ili). upper respiratory illness was defined as an illness with at least two of the following: 1) runny nose or sneezing; 2) nasal congestion; 3) sore throat, hoarseness, or difficulty swallowing; 4) cough; 5) swollen or tender glands in the neck; and 6) fever (oral temperature > 38°c) at the time of presentation. influenza-like illness was defined as a respiratory illness with acute onset presenting with fever and cough or sore throat. all ili cases also met the case definition for uri. non-ili cases were defined as uri cases not meeting the case definition for ili. additional nasal and throat swabs were requested from participants presenting with acute ili or uri. laboratory investigations. nasal and throat swabs were placed in viral transport media and stored at −20°c until transfer to the armed forces research institute of medical sciences for further testing. we tested acute swabs for influenza virus using reverse transcription polymerase chain reaction (rt-pcr) following the u.s. centers for disease control protocol. 12, 13 we also tested acute samples (from both camps) and the routine enrolment and follow-up specimens (from one camp) using a multiplex real-time pcr assay comprising 33 bacterial, viral, and fungal targets (ftd33 kit; fast track diagnostics, esch-sur-alzette, luxembourg). a cycle threshold value of < 33 was considered a positive result. previous studies have shown high agreement between ftd33 and other commercial multiplex assays with specificities > 95% for most viral targets. 14,15 sensitivity for influenza a and b, and human coronaviruses is high (> 90%), but is lower and more variable for other viral targets, including human bocavirus (80-92%), rhinovirus (61-75%), respiratory syncytial virus (50-72%), and human metapneumovirus (74-100%). 14, 15 in a subset of recruits, we measured influenza antibody titers in paired sera taken at enrollment and at the end of followup using a hemagglutinin inhibition assay. seroconversion was defined as a 4-fold rise in influenza subtype-specific antibody titers. statistical analysis. rates of illness. individuals were considered at risk from the date they entered the camp for a period of 10 weeks. we computed the rates of uri and ili as the number of cases of each outcome divided by the total person-time at risk, using robust standard errors in the calculation of 95% confidence intervals (cis) to allow for clustering of illness episodes by camp and cohort. disease etiology. we determined the percentage of uri and ili cases positive for each target pathogen, the percentage positive for more than one pathogen, and the percentage with no pathogen identified. influenza seroconversion. we calculated the percentage of recruits who seroconverted, overall and by influenza subtype, based on a 4-fold rise in antibody titer between baseline and end of follow-up samples. bacterial colonization. we determined the percentage of recruits with a positive identification of a bacterial pathogen at recruitment, the overall change in colonization prevalence between baseline and end of follow-up for each bacterial species, and the risk of acquisition of bacterial pathogens over the follow-up period among those initially negative. antibiotic use. we calculated the percentage of uri and ili cases that were prescribed antibiotics and investigated clinical signs and symptoms associated with antibiotic prescription using logistic regression. among those prescribed antibiotics, we determined the fraction of cases with a likely viral etiology based on the available fast track multiplex pcr results. analyses were conducted using stata 11 (statacorp, college station, tx) and r 3.2.2. ethical approval. the study was approved by the institutional review boards of the rta in bangkok, thailand, the walter reed army institute of research, and the london school of hygiene & tropical medicine. all participants provided written informed consent. three cohorts of army recruits undertook basic military training at the two camps during the study period, in may 2014-july 2014, november 2014-january 2015, and may 2015-july 2015. in total, 933 recruits undertook training during these three periods. of these, 867 (93%) were enrolled in the study. of the 867 participants, five participants withdrew from the study and 44 participants recruited in november 2014 were transferred out of the camp to complete their training elsewhere. the remaining 818 (94%) completed the 10-week follow-up and were included in the analysis. all participants were male and the median age was 21 years (range: 20-29 years). details of recruitment and follow-up completion for each cohort are given in supplemental figure 1 . rates of illness. there were a total of 248 episodes of uri and 8,063 person-weeks of follow-up, yielding an overall uri rate of 3.1 per 100 person-weeks. upper respiratory illness rates ranged from 2.6 to 4.6 per 100 person-weeks across cohorts, with the exception of the camp a may 2014 cohort, in which no uri episodes were reported ( figure 1 ). there were 160 ili episodes, giving an overall ili rate of 2.0 per 100 person-weeks, ranging from 0 to 4.1 per 100 person-weeks across cohorts. symptoms and time off work. fever was present in nearly two-thirds of uri episodes. cough and sore throat occurred in > 90% of uri episodes, nasal congestion and headache in > 80% of episodes, and malaise and breathing difficulty in > 60% of episodes. all these symptoms were significantly more common in ili compared with non-ili episodes ( table 1) . influenza-like illness cases were also more likely to take time off work than non-ili cases (57.5% versus 13.6%, p < 0.001), although the length of time taken off work was short (median 1 day). there was one hospitalization in an individual diagnosed with pneumonia who was off work for 7 days. disease etiology. among the 134 uri and ili samples tested by multiplex pcr, a pathogen was detected in 128. a single pathogen was detected in 26% of samples, two pathogens in 31%, three pathogens in 26%, and four pathogens in 12%. among uri pathogens, the most common viruses detected were rhinoviruses (22.4%), influenza b (11.9%), coronavirus 229 (5.2%), adenovirus (3.0%), and influenza a/h3 (3.0%). other viruses accounted for < 3% of uri cases each. of 133 samples tested for influenza by both rt-pcr and the fast track multiplex assay, 20 (15%) tested positive by at least one assay, and all of these were also positive by rt-pcr. of the 20 rt-pcr positives, four were positive for influenza a/h3, of which one was also positive by the multiplex assay. of the remaining 16 positive for influenza b by rt-pcr, 14 were also positive by the multiplex assay. there were no samples that tested positive by the multiplex assay and negative by rt-pcr. among the bacteria, the most commonly detected species were haemophilus influenzae type b (62.7%), h. influenzae non-type b (58.2%), streptococcus pneumoniae (16.4%), and klebsiella pneumoniae (16.4%). however, these were commonly found in combination with other pathogens. when considering only samples in which a single organism was found, h. influenzae type b was found in 13.4% of samples, h. influenzae non-type b in 9.0%, and s. pneumoniae, k. pneumoniae, and legionella pneumophila in 0.7% each. there was little evidence to suggest that etiological agents differed between ili and non-ili cases, with the exception of h. influenzae non-type b and coronavirus 229, which occurred more frequently in non-ili cases, and influenza a/h3 and coronavirus hku1, which occurred more frequently in ili cases ( table 2) . analysis of uri cases by time since the start of follow-up indicated strong temporal clustering of cases, with each cluster involving several respiratory pathogens (figure 2 ). for example, the single cluster in camp a during the november 2014 cohort resulted in identification of both rhinovirus and coronavirus 63, as well as numerous colonizing bacteria. in other clusters, rhinovirus was also identified in combination with parainfluenza viruses, adenovirus, and influenza b virus ( figure 2) . influenza vaccination and seroconversion. at enrollment, 2.4% of recruits reported having received influenza vaccination in the previous 12 months. in the serology subset, table 3) . thirteen of 169 (8%) recruits seroconverted over the 10-week training period. seroconversion was most common against influenza b/massachusetts (n = 7) and influenza a/h3 (n = 3) (table 3) colonization. at baseline, 82.3% of recruits were colonized with h. influenzae type b and 31.5% with h. influenzae non-type b. other common colonizing bacteria were k. pneumoniae (14.6%), staphylococcus aureus (8.5%), and s. pneumoniae (8.5%). at the end of follow-up, colonization with h. influenzae non-type b had increased to 74.1%, and s. pneumoniae to 33.6%, whereas colonization with h. influenzae type b and k. pneumoniae had decreased to 67.2% and 8.4%, respectively. in addition, moraxella catarrhalis was found in four (3.1%) and l. pneumophila in two (1.5%) end of follow-up samples. no change in s. aureus colonization was found. in a subset of paired baseline and end of follow-up samples initially negative for h. influenzae non-type b (n = 89), the risk of h. influenzae non-type b acquisition was 70.1% (95% ci: 54.4-90.6%). among those negative for s. pneumoniae at baseline (n = 119), the risk of acquiring s. pneumoniae during the follow-up period was 31.9% (95% ci: 22.6-43.8%). use of medications. of 248 uri episodes, 39.1% were prescribed antibiotics. of these, all but one were prescribed amoxicillin or azithromycin. in multivariable logistic regression, cases were more likely to be prescribed antibiotics if they presented with tonsillitis (or = 5.10, 95% ci: 2.35-11.10), chills (or = 3.91, 95% ci: 1.44-10.62), and swollen lymph nodes (or = 2.54, 95% ci: 1.07-6.04). antibiotic prescription was less likely if patients presented with nasal congestion (or = 0.26, 95% ci: 0.12-0.54). there was no difference in prescribing between ili and non-ili patients. of 35 uri cases prescribed antibiotics and with available multiplex pcr results, 16 (45.6%) had viral pathogens detected and eight (22.9%) had a bacterial target detected in the absence of other pathogens. we found a high incidence of uri in thai army recruits, indicating that approximately 30% of recruits experienced a uri episode over the 10-week training period. comparison with other military cohorts is not straightforward because of differences between settings in terms of vaccination policies and seasonality of key pathogens such as influenza. the uri and ili rates in our study are nonetheless comparable with those reported by other studies in the united states. [16] [17] [18] despite this, we found low rates of clinical influenza a in this minimally vaccinated population. possible reasons for this could be exposure to circulating influenza strains before recruits entering the training camps. thai national influenza surveillance data indicate high influenza activity in the first half of 2014, with influenza a/h1n1 and influenza b predominating. 19 concomitantly, our data indicated that 55% of recruits with available serological information had baseline influenza a/h1n1 titers ³ 1:40. titers greater than this value have been shown to be associated with a 50% reduction in risk of clinical influenza from homologous strains, for both influenza a and b viruses. [20] [21] [22] [23] of interest, 86% of recruits also had baseline titers against influenza b/massachusetts greater than this level. the higher baseline titers against influenza b/massachusetts are consistent with the frequent detection of influenza b isolates from the yamagata lineage in clinical specimens from thailand in the first half of 2014. 19 in our study, the fast track multiplex assay appeared to have lower sensitivity for influenza compared with the cdc rt-pcr assay. we did not specifically investigate reasons for these discrepancies, although it should be noted that the two assays were not performed at the same time, as the multiplex assays were all performed at the end of data collection. we therefore cannot discount the possibility that sample degradation might have influenced the relative performance of the fast track assay. although influenza incidence was low, pathogen detection with sensitive multiplex diagnostics indicated high frequencies of respiratory pathogens, with co-circulation of multiple viruses including rhinovirus, adenovirus, and coronaviruses as well as influenza. in addition, we found high levels of colonization with numerous bacterial species, including both type b and non-type b h. influenzae, k. pneumoniae, and s. pneumoniae. although we did not specifically study transmission chains or epidemiologic links between cases of illness, in the context of our study, recruits did not leave the camps during the 10-week training periods, such that the occurrence of respiratory pathogens is most likely to have resulted from transmission within the camps rather than from external sources. intense transmission of bacterial species was apparent, as the risks of acquiring h. influenzae non-type b and s. pneumoniae among initially non-colonized individuals were 70% and 31%, respectively. bacterial species were also commonly found in samples from uri cases. the colonizing nature of these bacterial species makes it difficult to determine the clinical relevance of these detections. however, h. influenzae type b and non-type b were identified in 13% and 9% of clinical samples in the absence of any other pathogens, suggesting that these pathogens could be responsible for illness in a fraction of adult uri cases. interestingly, h. influenzae type b has not been previously associated with a significant burden of uri in adults. 24 it is possible, however, that these cases were caused by other pathogens not included in the multiplex assay. high levels of colonization with bacterial species also point to the need for judicious use of antibiotics to treat uri in military populations. a quarter of uri cases in our study were prescribed broad-spectrum antibiotics, of whom 50% were more likely to have infections caused by viral pathogens based on multiplex pcr results. although we lacked additional information to determine whether this resulted in adverse antibiotic resistance patterns, high levels of antibiotic use in a population with a high background of bacterial colonization presents a risk for development of antibiotic resistance, and highlights the importance of improved rapid diagnostics to rule out viral etiologies to aid clinical management. in addition, it points to the need for increased antibiotic stewardship and health-care provider education to reduce unnecessary use of antibiotics in military settings. our study also emphasizes the importance of systematic microbiological surveillance of uri in military populations, to allow prompt detection and control of outbreaks and early identification of emerging pathogens that may be circulating or could be introduced in the wider population. this is particularly important as infection control options in such closed settings are limited and vaccination coverage for key pathogens is low. limited resources meant it was not possible to conduct multiplex diagnostics for all baseline, acute, and end of followup samples, as well as influenza serology. as a result, there was limited power to investigate whether colonization influenced risk of illness, or factors associated with colonization and acquisition of bacteria. in addition, co-circulation of multiple pathogens makes it difficult to establish chains of transmission, which would require more detailed microbial sequencing information. clinical signs and symptoms were available at presentation only, and we did not have information regarding the duration of individual symptoms or additional clinical investigations conducted longitudinally during the course of illness, as these were not routinely performed. the specific barracks setting and population characteristics limit the generalizability of our findings to other military settings with similar conditions. nevertheless, our study provides key data regarding the epidemiology and transmission of respiratory pathogens in military populations in tropical settings. our results emphasize the need for improved infection prevention and control strategies in military settings, given the potential for intense transmission of a wide range of respiratory pathogens. financial support: this work was supported by the united states department of defense-global emerging infectious disease surveillance (dod-geis), protocol 989a. disclosures: the authors declare that they have no competing interests. material has been reviewed by the walter reed army institute of research. there is no objection to its presentation and/or publication. the opinions or assertions contained herein are the private views of the author, and are not to be construed as official, or as reflecting true views of the department of the army or the department of defense. the investigators have adhered to the policies for protection of human subjects as prescribed in ar 70-25. respiratory tract infections in the military environment environmental factors, immune changes and respiratory diseases in troops during military activities 2009 influenza a(h1n1) seroconversion rates and risk factors among distinct adult cohorts in singapore epidemiology and transmission characteristics of human adenovirus type 7 caused acute respiratory disease outbreak in military trainees in east china febrile respiratory illness associated with human adenovirus type 55 in south korea military outbreak of acute respiratory disease caused by human adenovirus type 7 in a military training camp in shaanxi pertussis outbreak among soldiers during basic training: the need for updated protocols acute respiratory disease in us army trainees 3 years after reintroduction of adenovirus vaccine the increasing application of multiplex nucleic acid detection tests to the diagnosis of syndromic infections advances in multiparametric molecular diagnostics technologies for respiratory tract infections elevated transmission of upper respiratory illness among new recruits in military barracks in thailand cdc protocol of realtime rtpcr for influenza a(h1n1) cdc realtime rt-pcr (rrtpcr) protocol for detection and characterization of influenza evaluation of four commercial multiplex molecular tests for the diagnosis of acute respiratory infections comparison of fast-track diagnostics respiratory pathogens multiplex real-time rt-pcr assay with in-house singleplex assays for comprehensive detection of human respiratory viruses reduction in acute respiratory infection among military trainees: secondary effects of a hygiene-based cluster-randomized trial for skin and soft-tissue infection prevention comparison of the effectiveness of trivalent inactivated influenza vaccine and live, attenuated influenza vaccine in preventing influenza-like illness among us military service members handwashing and respiratory illness among young adults in military training influenza activity in thailand and occurrence in different climates the role of serum haemagglutination-inhibiting antibody in protection against challenge infection with influenza a2 and b viruses determinants of immunity to influenza infection in man estimation of the association between antibody titers and protection against confirmed influenza virus infection in children relationship between haemagglutination-inhibiting antibody titres and clinical protection against influenza: development and application of a bayesian random-effects model haemophilus influenzae infections in the h. influenzae type b conjugate vaccine era acknowledgments: we are grateful to the military recruits who participated in the study, the royal thai army, and the clinical, laboratory, and administrative personnel at afrims for their support. this is an open-access article distributed under the terms of the creative commons attribution license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-003360-mkv1jc7u authors: chen, yirong; badaruddin, hishamuddin; lee, vernon j.; cutter, jeffery; cook, alex r. title: the effect of school closure on hand, foot, and mouth disease transmission in singapore: a modeling approach date: 2018-10-22 journal: am j trop med hyg doi: 10.4269/ajtmh.18-0099 sha: doc_id: 3360 cord_uid: mkv1jc7u singapore implements a school closure policy for institutional hand, foot, and mouth disease (hfmd) outbreaks, but there is a lack of empirical evidence on the effect of closure on hfmd transmission. we conducted a retrospective analysis of 197,207 cases of hfmd over the period 2003–2012 at the national level and of 57,502 cases in 10,080 institutional outbreaks over the period 2011–2016 in singapore. the effects of school closure due to 1) institutional outbreaks, 2) public holidays, and 3) school vacations were assessed using a bayesian time series modeling approach. school closure was associated with a reduction in hfmd transmission rate. during public holidays, average numbers of secondary cases having onset the week after dropped by 53% (95% credible interval 44–62%), and during school vacations, the number of secondary cases dropped by 7% (95% credible interval 3–10%). schools being temporarily closed in response to an institutional outbreak reduced the average number of new cases by 1,204 (95% credible interval 1,140–1,297). despite the positive effect in reducing transmission, the effect of school closure is relatively small and may not justify the routine use of this measure. hand, foot, and mouth disease (hfmd) is a common pediatric disease that is endemic in east and southeast asia 1,2 and increasingly found in north america 3, 4 and europe. 5 when caused by coxsackieviruses, hfmd usually leads to relatively mild infections with symptoms that are self-limiting. 6 however, hfmd caused by human enterovirus 71 may lead to complications involving the nervous system and result in reduced cognitive function, delayed neurodevelopment, and motor impairment, or death [7] [8] [9] ; in china, an estimated 350-900 children die of hfmd annually. 10 because of this potential severity, the ministry of health in singapore imposes strict control policies, especially in preschools. in preschools, daily routine health checks for all children and isolation of suspected cases are implemented for early detection and control of outbreaks. 11 should transmission continue within an outbreak, school closure is enforced. 12 this policy was implemented following an outbreak in 2000 in which several children died from enterovirus 71 complications 13, 14 ; the details have evolved over time, but until recently, if a school has more than 16 cases or an attack rate more than 23% with a transmission period more than 24 days, the school will be required to close for a period of 10 days. 15 in the most updated guideline, the policy has been relaxed somewhat, and ministry of health will consider the predominant circulating strain when assessing closure in addition to the trigger. this change provides an avenue to assess the impact of this policy. school closure as a form of social distancing intervention to mitigate transmission during an infectious disease outbreak 16 is often found in countries' pandemic preparedness plans. 17 the influenza literature shows that school closure may effectively reduce the spread, [18] [19] [20] and school vacations have a significant impact in limiting transmission. 21, 22 analyses also suggest that school closure combined with the use of antiviral agents is cost-effective and is a justifiable strategy for mitigating influenza pandemics. 23, 24 there is, however, a lack of empirical evidence on the effect of school closure on hfmd transmission: one exception is a review of hfmd in hong kong that revealed fewer hfmd consultations than expected during the 2003 severe acute respiratory syndrome and the 2009 influenza pandemic, which was attributed to various control measures including school closure. 25, 26 this article aimed to assess the effect of school closure on hfmd transmission. in singapore, hfmd is endemic with year-round transmission and is legally notifiable by physicians and childcare teachers, as well as actively screened for in preschool-aged children. these policies provide data that enable us to obtain three sources of information on the effect of school closure: 1) the reduction in the numbers of cases after a public holiday, when childcare centers and schools close; 2) the reduction during school vacations; and 3) the impact within childcare centers of school closure in response to an ongoing outbreak. singapore's school closure policy for hfmd, which has been implemented for over a decade, in tandem with a comprehensive hfmd surveillance system, therefore, provides a unique opportunity to assess the impact of this important method of outbreak control. source of data. ministry of health, singapore, actively monitors and publishes the incidence of hfmd, which was made a notifiable disease in the year 2000. 27 two sets of data on hfmd were extracted from the ministry's records for this study. the first dataset contains aggregate reported hfmd cases from 2003 to 2012, with the number of daily cases with onset of symptoms, stratified by age. the second dataset contains information on all hfmd outbreaks in childcare centers and kindergartens in singapore, during the period 2011-2016. this provides the cumulative number of cases per day in each preschool with an outbreak, together with the school type (childcare centers or kindergarten), enrollment size, whether the schools were closed because of the outbreak, and, if so, dates of closure and reopening. data were retained at a daily resolution for two analyses but aggregated to weekly for the analysis of vacations. data were collected under singapore's infectious disease act, and because aggregate non-identifiable data were used, institutional review board approval was not deemed necessary for this study. statistical analysis. separate statistical analyses were performed to investigate the 1) public holiday effect, 2) school vacation effect, and 3) school closure effect on hfmd transmission, as described in the following paragraphs. public holiday effect. there are typically 11 public holidays in singapore each year, as detailed in supplemental table 1 ; these are a mix of secular and religious holidays, some of which rotate around the year following the lunar calendar. if a public holiday falls on sunday, the following monday will be a public holiday. we derived the dates of all public holidays from 2003 to 2012 from the official listing of the ministry of manpower. the effect of public holiday on hfmd transmission was measured by quantifying the reduction attributable to the public holiday. because time points not immediately preceding or following a public holiday contribute little information to the effect of the public holiday, rather than considering a time series model, we developed a bayesian model of the time points surrounding public holidays. the number of cases in the week before the holiday i is modeled as x i ∼ poissonðα i × 7μþ, and the number the week after as y i ∼ poissonðα i × ½6μ + θμþ, or y i ∼ poissonðα i × ½5μ + 2θμþ for two-day-long holidays. here, μ represents the average number of cases on a typical day; estimates of θ and a 95% credible interval provide a measure of the public holiday effect (i.e., 1 à θ is the reduction in the number of infections on a public holiday compared with that on a normal day); and α i ∼ γða,aþ is an individual week effect which has an expected value of 1 and allows for autocorrelation in the time series. the window length of 1 week was selected to ensure balance in the number of weekdays in each window and to correspond roughly to the assumed incubation period of hfmd of around 3-7 days. 2 this model assumes that the difference between the number of cases prior or after the public holidays is purely because of the difference in the number of infections happening on the public holiday, which is unobservable but indirectly represented by the change in the number of symptomatic cases in the following weeks. to assess whether there was any longer term impact, we repeated the analysis comparing incidence in the second week (days [8] [9] [10] [11] [12] [13] [14] following the holiday with the week before it, as well as in the third week (days [15] [16] [17] [18] [19] [20] [21] . the main analysis does not treat public holidays that were less than 7 days apart specially. in sensitivity analysis, we removed data corresponding to both holidays from the fitting procedure if they were less than 7 days apart. sensitivity analysis was also conducted by applying different forms of the non-informative prior distributions for the parameters to test the robustness of the inference. school vacation effect. in singapore, kindergartens, primary schools, and secondary schools have school vacations in march for 1 week, in june for 4 weeks, in september for 1 week, and in november and december for 5-6 weeks. to measure the effect of school vacations on hfmd transmission, we built time series models, fit bayesianly, for the weekly number of children with hfmd aged 12 years and younger. this has some similarities with an autoregressive time series, but the bayesian approach afforded greater flexibility in the model specification. we assumed a negative binomial distribution for the number of hfmd cases, y t , observed in week t, with mean and shape parameter p, where h t = 1 if week t is a school holiday and 0 otherwise, and a, b, d, p, are model parameters that needed to be estimated: b accounts for autocorrelation, whereas d determines the effect of school vacation. a negative binomial distribution is used to allow more flexibility to capture the observed variability in the data than that obtained using the poisson model originally considered. a constant b that does not vary with time was used because singapore has very little seasonality that would add forcing to the timing of epidemics of hfmd, 28 and because it would hamper identifiability of the vacation effect. the adequacy of using a constant b was assessed and confirmed by examining the distribution of the residual of 1-week-ahead model forecasts. the model was fit using bayesian methods (markov chain monte carlo) with non-informative prior distributions, as described in the supplemental file. for each draw from the posterior distribution, we iteratively derived the median number of cases for the following week based on the number of cases in the current week using the formula in the aforementioned model. the number of cases in the first week of the simulations was taken to be the observed average weekly number of cases across the time horizon. this was done for 104 weeks (2 years) and only the latter 52 weeks were kept as the number of weekly cases for a typical year. for each set of posterior values, 1,000 such simulations were performed. based on these 1,000 simulations of a typical year scenario using each of the 10,000 sets of posterior values, 95% prediction and credible intervals were derived for each of the 52 time points. median values for all 1,000 simulated numbers for each set of posterior values (10,000 median values in total) were calculated. school closure during outbreaks effect. for each preschool that had an outbreak during the time horizon 2011-2016, a period of 50 days from the first day of case onset was considered; this was enough to span the duration of most outbreaks. the number of incident cases on day j in outbreak i, x i,j , was modeled by where c i,j is the cumulative number of cases within 1 week before day j in outbreak i, s i,j is the number of children not yet symptomatically infected during the current outbreak by day j of outbreak i, and w i,j is the indicator for school closure on day j in outbreak i (w i,j = 1 if the school was closed on day j in outbreak i, and 0 otherwise). we smoothed the effect λ k of the current outbreak size k using the formula the posterior samples were used to project the number of cases that were avoided by forcing schools to close when they hit the trigger. for each draw from the posterior, 100 simulations were performed to each of the schools with closures in our study period. in each simulation, the number of new incident cases on each day after the closure day was calculated using the model formulation and w i,j was set to be 0. the total number of cases was then compared with the observed number of cases when schools close and the total number of additional cases due to not closing schools for each simulation/set of posterior values was recorded. median values for all 100 simulated numbers for each set of posterior values (10,000 median values in total) were calculated. we use bayesian methods to fit the models as the bayesian paradigm was considered more flexible than its frequentist analog. throughout, non-informative prior distributions were selected (detailed in supplemental file 1) and models were fit, using markov chain monte carlo algorithms with burn-in periods of 1,000 and 10,000 iterations, in the r statistical environment. 29 the posterior samples were then used to obtain posterior distributions of derived quantities. throughout, equal-tailed 95% credible intervals are used. model convergence was evaluated by using the gelman diagnostic tests 30 and model validity assessed by comparing posterior predictions or simulations with the actual observed data. a total of 197,207 hfmd cases were notified to the ministry of health, singapore, from 2003 to 2012. table 1 shows the demographics of all notified cases. children aged less than 12 years accounted for about 90% of all hfmd cases, and about 70% of all children with hfmd were aged 5 years and younger; 57% of the cases were male, whereas 74% were ethnic chinese, 15% malay, 3% indian, and 8% others: the indian ethnic group is substantially underrepresented among cases compared with the general population (11% of singapore residents aged less than 12 years have indian ethnicity). table 2 shows the summary statistics for the bayesian models for the public holiday effect: the average of 50 hfmd cases a day was reduced by 53% (95% confidence interval [ci]: [44%, 62%]) in the 1 week following a public holiday, but only by 34% (95% ci: [25%, 43%)]) in the second week, whereas there was no reduction in the third week (0%, 95% ci: [−10%, 9%]). alternative forms of non-informative prior distribution did not affect the results (not shown). during school vacations, weekly number of cases were modeled to be reduced to 93% ( figure 1 shows the simulations for a typical year based on the modeled effects of school vacations on hfmd transmission, using typical timing of school holidays. both overall analysis and separate models for different age groups showed similar results and are consistent with the patterns of the data for the 10-year period. the temporally structured distribution of residuals in the 1-week-ahead model forecasts is shown in supplemental figure 1 . no trends were observed, suggesting that the model with a constant b is adequate. from 2011 to 2016, there were totally 10,080 school-level outbreaks involving a total of 57,502 hfmd cases in childcare centers and kindergartens, of which 105 led to closure (table 3 ). in all, 9,903 outbreaks including 100 closures were included in our model: 177 schools without accurate enrolment sizes were removed and four schools with closure falling beyond 50 days after the first day of outbreak were categorized as no closure (detailed in supplemental file 1) . the bayesian poisson model shows that the expected number of new cases decreases to 26% (95% ci: 21-32%) if a school was closed on that day compared with a normal school day, after adjusting for size and duration of the outbreak. one-dayahead predictions based on the previous day's observed number of cases, shown in figure 2 for arbitrarily selected outbreaks (more outbreaks are presented in supplemental file 2), both with and without short-term school closure, demonstrate that the fitted model adequately captures the observed outbreak patterns. figure 3 show the effect of school closure by showing the cumulative number of cases if there were no school closure for four arbitrarily selected outbreaks with closure (more outbreaks are presented in supplemental file 3). the percentage of cases avoided from school closure for all closures is shown in figure 4 . the majority of school closure events (> 80%) were associated with a less than 5% difference between observed and modeled cases, regardless of outbreak size at closure. closures that prevented more than 10% of total school size were generally in bigger schools (of size more than 200). overall, the modeled number of infections prevented through the school closure policy from 2011 to 2016 was 1,204 (95% ci: [1, 140, 1, 297] ), that is, an average 2% reduction compared with the total number of cases during that period. convergence of all bayesian models used was achieved according to the gelman diagnostic test. model validity assessed by comparing posterior predictions or simulations school closure is a common control measure in pandemic preparedness plans and in response to actual outbreaks, which may possibly be a high impact method of controlling an outbreak of a severe or potentially severe infectious disease. its effectiveness has been assessed in modeling studies, 19, 20, 22 which posit that because children have closer social networks and high contact rates in schools, 31 closing schools may substantially reduce transmission, if they do not compensate by having greater contact outside of school during a closure. the burden of school closure on families which may need to make alternative childcare arrangements means that it is imperative to have real-world evidence supporting its effectiveness. relatively little such evidence is available, though some studies in japan, france and the united states have assessed school closures in response to influenza outbreaks. 18, 32 in light of this, the evidence on the effect of closure from singapore's long-standing routine school closures to control hfmd may prove valuable. this study demonstrated a consistent reduction in average numbers of hfmd cases when schools were closed, regardless of the reason for the closure. there was an estimated decrease of 7% in the number of cases in the week after a public holiday than the week before it, corresponding to a reduction in the number of infections that occurs on the holiday itself of 53%, although the effect of the holiday declines in the weeks following as infections return to baseline levels. a similar reduction in the number of infections during school vacations was also estimated (also about 7%). this may be an underestimate of the actual effect size because many preschool-aged children still attend preschools during school holidays. this hypothesis is supported by the slightly accentuated effect in children who have started formal schooling, with a reduction in risk around 10% for children aged 6 years and older. although the reduction was statistically significant for all age groups, the difference in effect between age groups was not statistically significant. the data on school closure in response to outbreaks allowed a quasi-experimental analysis because recently the school closure policy has been reinterpreted to allow more discretion by the ministry of health whether to close the affected school or not. this led to a decrease in the number of school closures in the later periods of the study (table 3) . although not randomized, this change allowed some overlap in the exposure (closure) and response (outbreak growth). the analysis showed that as with holidays and public holidays, school closure during an outbreak had the intended effect of orange line shows the cumulative number of cases if the school was not closed during the closure period. light pink and dark pink shades are 95% and 70% confidence intervals, respectively. this figure appears in color at www.ajtmh.org. mitigating transmission, but the effect was relatively small, and we estimated that only ∼1,200 cases were averted over the 6 years analyzed. the high asymptomatic rate 33, 34 and seroprevalence of the main causative viruses 35 may mean that a substantial fraction of children are no longer susceptible by the time of closures in response to large outbreaks, which is currently triggered when more than 16 cases or 23% of children are symptomatic. the fraction of cases prevented by outbreak-induced closure was modeled to be small in general, but larger for the larger preschools, which we attribute to the structure of the thresholds for closure: this is effected when either the fraction or the number of children in the school is notified, meaning that closure of larger schools occurs when the threshold for the number of cases is hit but the fraction is still low. given the disruptions to parents/families from unplanned closures, 11,36 outbreak-driven closures may, therefore, cause more problems than closure because of holidays. in light of the limited effect, it is not clear that this policy should be continued to be used routinely for hfmd outbreaks, but the effect of closures because of holidays suggests that a school closure policy may still be valuable for pandemic preparedness plans and needs to be carried out when there are serious outbreaks or when the outbreak is due to novel pathogens of unknown severity. although this was a retrospective study, with hfmd being a legally notifiable disease in singapore, incident data were collected in a standard way through preschools, clinics, and hospitals, and as such, the data used for this study are largely reliable indicators of symptomatic cases. however, some other limitations exist. several assumptions were made in the modeling process: the public holiday effect analysis assumed that the observed difference in the number of cases before and after the public holiday was solely attributable to the public holiday. in temperate countries, this assumption may not be tenable because of seasonal changes that drive transmission. singapore, however, lies close to the equator and has almost no seasonality-having year-round high temperature and humidity-with the effect that some virus transmission is effectively aseasonal. 28 for the same reason, the assumption that there are no other temporal effects in the school vacation analysis may be more robust than it would in a temperate setting. these three analyses provide evidence that school closure-whether in response to an outbreak, or because of a public holiday or longer school vacation-reduces the transmission of hfmd. although we found that school closure in response to outbreaks does have a positive effect in reducing transmission, the effect is small, and given the high incidence and the fact that most of the cases of hfmd are mild and selflimiting, the evidence from this study suggests that policymakers in singapore should evaluate whether routine use of school closure outside of public health emergencies justifies the impact on families. epidemiology and control of hand, foot and mouth disease in singapore the epidemiology of hand, foot and mouth disease in asia: a systematic review and analysis notes from the field: severe hand, foot, and mouth disease associated with coxsackievirus a6-alabama hand, foot, and mouth disease caused by coxsackievirus a6 avoort h; dutch working group for clinical virology cdc. hand foot and mouth disease (hfmd)-signs and symptoms. available at long-term cognitive and motor deficits after enterovirus 71 brainstem encephalitis in children neurodevelopment and cognition in children after enterovirus 71 infection outbreak study group. deaths of children during an outbreak of hand, foot, and mouth disease in sarawak, malaysia: clinical and pathological characteristics of the disease 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modelling of costs and health consequences of school closure during an influenza pandemic acknowledgments: we thank the data extraction team from communicable disease division, ministry of health, singapore, for their help in extracting data for our analysis. authors' addresses: yirong chen and alex r. cook, saw swee hock school of public health, national university of singapore and national university health system, singapore, e-mails: chenyirong89@gmail. com and alex.richard.cook@gmail.com. hishamuddin badaruddin, vernon j. lee, and jeffery cutter, ministry of health, singapore, e-mails: drphisham@gmail.com, vernonljm@hotmail.com, and jeffery_cutter@ moh.gov.sg.this is an open-access article distributed under the terms of the creative commons attribution license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-260871-dtn5t8ka authors: silva, marcus tulius t.; lima, marco; araujo, abelardo q.-c. title: sars-cov-2: should we be concerned about the nervous system? date: 2020-07-17 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0447 sha: doc_id: 260871 cord_uid: dtn5t8ka the covid-19 pandemic has proved to be an enormous challenge to the health of the world population with tremendous consequences for the world economy. new knowledge about covid-19 is being acquired continuously. although the main manifestation of covid-19 is sars, dysfunction in other organs has been described in the last months. neurological aspects of covid-19 are still an underreported subject. however, a plethora of previous studies has shown that human covs might be neurotropic, neuroinvasive, and neurovirulent, highlighting the importance of this knowledge by physicians. besides, several neurological manifestations had been described as complications of two other previous outbreaks of cov diseases (sars ad middle east respiratory syndrome). therefore, we should be watchful, searching for early evidence of neurological insults and promoting clinical protocols to investigate them. our objectives are to review the potential neuropathogenesis of this new cov and the neurological profile of covid-19 patients described so far. the world has been facing a pandemic for less than 6 months that has already resulted in thousands of deaths and is paralyzing the world economy. in december 2019, severe pneumonia cases of unknown origin were seen in wuhan, china. 1, 2 the pathogen was rapidly identified as a novel enveloped rna β-cov, named sars-cov-2. sars-cov-2 quickly spread to other parts of china and later to all continents. thenceforth, in march 2020, the who declared covid-19 a new pandemic. several neurological manifestations were described as complications of two other previous outbreaks of cov diseases, namely, sars and the middle east respiratory syndrome (mers). [3] [4] [5] [6] [7] furthermore, recent clinical observations had stressed the possibility of neurological diseases also in the context of covid-19. [8] [9] [10] [11] therefore, we should be vigilant, searching for early evidence of neurological insults and promoting clinical protocols to investigate them. for instance, considering that encephalitis is associated with high mortality and morbidity, early diagnosis and management may contribute to better outcomes. several clinical and laboratory studies have shown that human coronaviruses (hcovs) might be neurotropic, neuroinvasive, and neurovirulent. [12] [13] [14] [15] the objectives of this article are to review the potential neuropathogenesis of this new cov and the neurological profile of covid-19 patients described worldwide. viral respiratory infections in humans are generally secondary to the human respiratory syncytial virus, influenza virus, hcov, measles virus, rhinovirus, adenovirus, and human metapneumovirus. 16 transmission of these agents occurs mainly by contact with fomites or suspension droplets. 17 all these viruses can produce bronchiolitis and pneumonia, being responsible for a large number of hospitalizations every winter season. 16, 18, 19 in some cases, central nervous system (cns) diseases can also be seen. it is well known that viral respiratory infection can result in several neurological disorders such as seizures, status epilepticus, encephalopathy, and encephalitis (table 1) . respiratory viruses, in general, can invade the cns through three main routes: the hematogenous route, through the infection of the endothelium or by transendothelial mechanisms; the "trojan horse" mechanism, by which viruses in the bloodstream infect leukocytes that can transmigrate across the permeable blood-brain barrier; and through the olfactory nerves by axonal transport via olfactory neurons. this last example is an elegant mechanism to access the cns for a virus that enters the body intranasally, such as most of the respiratory viruses highlighted before. 20, 21 furthermore, it is tempting to associate this route with anosmia, a frequent symptom of covid-19. 22, 23 however, in experimental work, the probable route for brain infection in macaques was the hematogenous one. 24 cov rna and cov antigen were detected in nonhuman primates' brain after intranasal, intraocular, or intravenous inoculation of murine cov jhm omp1. in this animal model, both the lack of detection of virus products in the trigeminal ganglia or olfactory bulbs and the presence of viral antigen in vessels and perivascular regions suggest that cov entered the cns through vascular endothelium. covs are widespread and infect different species, generally causing mild, uncomplicated respiratory and enteric diseases. usually, they infect the upper respiratory tract, being mainly associated with the common cold. however, in some patients, they can reach the lower respiratory tract, causing interstitial pneumonia, exacerbations of asthma, respiratory distress syndrome, or even sars. 25 covs are enveloped positive-sense rna viruses characterized by club-like spikes that project from their surface, an unusually large rna genome, and a unique replication strategy. covs are classified into four different groups; α-, β-, γ-, and δ-cov. 26 sars-cov-2 is an rna β-cov with a characteristic crown-shaped appearance, grouped within the family coronaviridae, order nidovirales. sars-cov-2 shares significant genetic homology with sars-cov, a virus associated with the pandemic of sars that occurred in 2003. similar to sars-cov, angiotensinconverting enzyme 2 (ace2), an enzyme that physiologically counters renin-angiotensin-aldosterone system activation, is the functional receptor to sars-cov-2. 27 it is known that the ace2 receptor is also expressed in the brain. 28 several hcovs are pathogenic to humans, such as hcov-oc43, hcov-229e, mers-cov, and sars-cov, all of them having different genotypes. [29] [30] [31] neurotropic and neuroinvasive abilities of hcov have been described both in animals and humans, and is implicated in conditions such as multiple sclerosis and encephalomyelitis. 5, 18 interestingly, the first detection of hcov in the human brain was made at autopsy cases of multiple sclerosis in the early eighties. 32 more recently, arbour et al. 13 detected the presence of the hcov-oc43 in brain parenchyma samples of 35.9% patients with multiple sclerosis, compared with 13.7% of controls. also, murine hepatitis virus, another cov, has been linked to chronic inflammation and demyelination of the cns in animal models. 33 the viral glycoprotein s (spike) has an essential role for the neurovirulence, especially for the hcov-oc4. 34 sars-cov. sars was a novel zoonotic infectious disorder associated with sars-cov. it was first diagnosed in china, in november 2002. the comparison of sars-cov sequences isolated from civets and patients supported the concept of transmission from these animals to humans. 35, 36 phylogenetic analysis showed that sars-cov was not a novel cov, but a branch of the β-cov. [37] [38] [39] typically, sars patients exhibited a triphasic pattern of disease, initially presenting with fever, a nonproductive cough, sore throat, and myalgia. 40 generally, dyspnea does not become a prominent feature until the second week of illness. in the second phase, dyspnea and hypoxia with fever become more prominent. some patients progress to acute respiratory distress by the third week, often requiring mechanical ventilation. the severity of the disease was correlated with increasing age, and the mortality can reach 50% for patients older than 60 years. 40, 41 during the sars-cov outbreak, several neurological diseases were reported. peripheral nervous system manifestations associated with sars were described in four patients, including both axonal polyneuropathy and myopathy. 4 the neuromuscular disorders developed approximately 3 weeks after the onset of sars, and the prognosis was good. interestingly, olfactory neuropathy was described during the sars outbreak. 42 this finding is relevant because, nowadays, many patients with sars-cov-2 infection have reported anosmia. 23 a neuroinvasive behavior of sars-cov could be found in some other reports. sars-cov rna was present in both the serum and cerebrospinal fluid (csf) from a patient with status epilepticus and sars during the 2003 outbreak. 43 in another report, sars-cov rna was recovered from a csf sample of an infected patient admitted because of generalized seizures. 5 in addition, a 39-year-old patient died of sars after a severe chronic progressive viral cerebritis. 44 neuropathological examination showed gliocyte hyperplasia, neuron denaturation, and necrosis coupled with striated encephalomalacia. neuroinvasion by sars-cov was confirmed by the typical viral morphology observed under electron microscopy, by genetic identification, and by the detection of the viral antigen (n protein) in the brain. in a neuropathological study of patients with sars, performed during the 2003 chinese outbreak, sequences of sars-cov genome were detected in all brain samples, but in none of the control cases. 45 the authors stressed that the infection of neurons occurred in selected areas such as the hypothalamus and cortex. the degeneration seen in these cases was probably secondary to neuronal hypoxia/ischemia. it is hypothesized that the infection of neurons may explain a higher than usual percentage of neurological and psychological abnormalities observed in patients with late-stage sars. 46 therefore, those neuropsychiatric symptoms could not be merely attributed to negative social pressure during the epidemic. interestingly, it is not uncommon for survivors of sars to report neuropsychiatric symptoms, such as lethargy, malaise, orthostatic dizziness, apathy, depression, and anxiety. recently, studies have linked neuroendocrine aberrations to some neuropsychiatric conditions. dysfunction in the hypothalamic-pituitary-adrenal axis was reported in survivors of sars by leow et al. 47 in a cohort of 61 sars patients, 39.3% had a central hypocortisolism. the authors speculated that either a hypophysitis or a direct viral hypothalamic infection could be the cause of these extrapulmonary symptoms. all these clinical, laboratory, and neuropathological evidence suggest that cns infection is possible in patients with sars. most hcovs share similar viral structures and mechanisms of infection and have documented neurotropism. therefore, infectious mechanisms previously found in other hcovs may also apply to sars-cov-2. 48, 49 taking these observations into account, a proactive search for neurological symptoms and signs could elucidate if the same occurs in sars-cov-2 infection. sars-cov-2. sars-cov-2 is a new virus that shares almost 80% genomic homology with sars-cov. however, the highest level of similarity is with a horseshoe bat cov. 50 therefore, it is believed that sars-cov-2 is a recombinant virus, transferred from bats to human hosts via an intermediate host. 51 because it is an rna virus with an rna-dependent rna polymerase-based replication, mutation and recombination are not uncommon events. sars-cov-2 has been recently associated with severe interstitial pneumonia, called covid-19. 1,2 covid-19 is an acute viral pneumonia, potentially lethal in many cases. characteristics of patients with severe evolution are the rapid progression to respiratory failure; it is estimated that among those with respiratory difficulties, 50% have to be admitted to intensive care units, and, of these, 46-65% worsen in a short period and may die in a few days. although sars-cov and sars-cov-2 use the same receptor to enter human cells, the ace2 binding affinity of the sars-cov-2 spike protein is 10-to 20-fold higher than that of the sars-cov spike protein. 52 once within the cns environment, its interaction with ace2 receptors expressed in neurons can initiate a cycle of viral budding accompanied by neuronal damage, without substantial inflammation, as seen in the past in cases of sars-cov. 53 it is also important to mention that long before the neuronal damages occur, endothelial tears in cerebral capillaries, followed by bleeding within the cerebral tissue, can have fatal consequences in patients with covid-19. 54 experimental studies with hcov and other viruses have shown the presence of viral particles in the brain, especially in the brainstem. viral antigens of influenza and pseudorabies virus have been detected in the nucleus of the solitary tract and the nucleus ambiguous. 55, 56 the nucleus of the solitary tract receives sensory information from the mechanoreceptors and chemoreceptors of the lung and upper and lower airways, whereas the efferent fibers from the nucleus ambiguous and the nucleus of the solitary tract provide innervation to airway smooth muscle, glands, and blood vessels. such neuroanatomical interconnections suggest that the death of many infected animals and even patients may be due to a dysfunction of the cardiorespiratory center in the brainstem. 53, 57 likewise, experimental studies using transgenic mice have demonstrated brainstem infection by sars-cov and mers-cov. 53, 57, 58 whether the neuroinvasion of sars-cov-2 has a role in the development of respiratory failure in covid-19 patients is still a matter of speculation. this information is essential for the prevention and better treatment of sars-cov-2-induced respiratory failure. several reports discussed neurological complications of covid-19. in a clinical series, giacomelli et al. 23 described olfactory and taste disorders in 20 of 59 sars-cov-2-infected individuals. lechien et al. 59 studied 417 patients and found olfactory dysfunction in 85.6% and gustatory dysfunction in 88%. in 11.8% of patients, smell loss was the first symptom of covid-19. because the ace2 receptor is widely expressed on the epithelial cells of the mucosa of the oral cavity and sars-cov exhibits a transneural penetration into the olfactory bulb, the pathogenetic mechanism of taste and olfactory disorders in sars-cov-2 infection could be justified. 53, 60 a case of a 56-year-old patient with covid-19 and encephalitis was recently described; csf tested positive for sars-cov-2 by gene sequencing. 61 in another report, a woman with covid-19 and acute necrotizing encephalopathy (ane) was described. 9 acute necrotizing encephalopathy is a rare and potentially severe neurological complication of some viral infections, such as influenza. it has been associated with an exaggerated inflammatory response in the cns named cytokine storm, responsible for a blood-brain barrier breakdown. characteristic imaging features of ane include symmetrical, multifocal lesions with invariable thalamic involvement. lesions appear hypodense on computador tomography images, and magnetic resonance imaging (mri) demonstrates t2/fluid attenuation inversion recovery hyperintense signal with internal hemorrhages. as in other severe viral infections, covid-19 has been associated with cytokine storms. 62 a case of guillain-barre syndrome (gbs) was recently published by zhao et al. 11 the authors described a 61-yearold woman infected by sars-cov-2 who developed acute weakness in both legs and severe fatigue, progressing within 1 day. although it is not possible to exclude an epiphenomenon between sars-cov-2 infection and gbs, considering the temporal association, the authors speculate that sars-cov-2 might have been responsible for the development of gbs in this patient. in this report, the disease followed a parainfectious profile, instead of the classic postinfectious pattern, reported in gbs associated with other pathogens. more recently, toscano et al. 63 described a clinical series of five patients with gbs in italy. the interval between the onset of symptoms of covid-19 and the first neurological symptoms ranged from 5 to 10 days. csf samples of all patients were negative to sars-cov-2. in three patients, the findings were consistent with an axonal variant of gbs and with a demyelinating process in two. all patients were treated with intravenous immune globulin. in addition, two patients with sars-cov-2 infection were reported with miller-fisher syndrome and polyneuritis cranialis, respectively. 64 in the first patient, vertical diplopia, perioral paresthesia, and gait ataxia were noted 7 days following covid-19 symptoms (fever, anosmia, ageusia, low back pain, and malaise). this patient was treated with intravenous immune globulin, with the resolution of the neurological symptoms, except for anosmia and ageusia. goh et al. 65 recently described a 27-year-old man who developed isolated peripheral facial palsy on the sixth day of sars-cov-2 infection. mao et al. 8 have published a clinical series on neurological aspects of covid-19. among 214 covid-19 patients, 37% had some neurological manifestation, with nearly 50% having severe covid-19 disease ( table 2 ). the authors drew attention to the fact that in patients with severe infection, the neurological involvement was more frequent, and included acute cerebrovascular diseases, disturbances of consciousness, and skeletal muscle injury. in this seminal series of cases, the nervous system involvement was associated with a poorer prognosis. the authors described mental status alterations in 15% of severe cases and nonspecific symptoms, including headache and dizziness, in nearly 20%. the authors referred to another common finding: "skeletal muscle injury" (creatine kinase > 200 iu/ml) that was seen in approximately 20% of severe cases. unfortunately, the authors did not describe whether there were clinical manifestations suggesting myositis or myopathy, or even signs of acute motor neuron injury. in another clinical series of neurological patients with covid-19, helms et al. 66 described the following neurological disturbances: agitation in 40 of 58 patients (69%), corticospinal tract signs in 39/58 (67%), and dysexecutive syndrome in 14/39 (36%). brain mri findings could be summarized as leptomeningeal enhancement in eight of 13 patients (62%), perfusion abnormalities in 11/11 (100%), and ischemic stroke in 3/13 (23%). these data should be interpreted with caution because they did not allow the determination of which of these clinical and radiological characteristics were due to critical illness-related encephalopathy, brain cytokine effects, or the effect of withdrawal of medication, and which features were directly associated with sars-cov-2 infection. stroke is one of the most frequent neurological diseases associated with sars-cov-2 infection, 8 and large-vessel stroke in younger patients was recently reported in five patients. 67 the mean national institutes of health stroke scale score in these patients was 17 (scores range from 0 to 42; the higher the score, the greater the severity of stroke). intravascular coagulation is the most likely factor in causing ischemic stroke in these patients; a high d-dimer is associated with increased risk for thrombosis and with a poor prognosis in covid-19 patients. 68, 69 finally, the presence of sars-cov-2 in the human brain was recently documented in the frontal lobe. 70 a patient with parkinson's disease died 11 days after sars-cov-2 infection because of cardiac and pulmonary complications. transmission electron microscopy showed viral particles in frontal lobe sections. noteworthy, virus-like particles were also found in the capillary endothelium and actively budding across endothelial cells, which could suggest a hematogenous route for sars-cov-2 entry into the cns. the real spectrum of neurological manifestations of covid-19 is an ongoing story. as the virus spreads to all continents, we may observe different manifestations in populations with diverse genetic and environmental backgrounds. besides, rna viruses such as sars-cov-2 suffer frequent mutations, which can be associated with new and unidentified neurological manifestations. close epidemiological surveillance is necessary to follow gbs's frequency and acute demyelinating encephalomyelitis, autoimmune conditions that usually follow viral infections. another critical aspect of the covid-19 pandemic is the collapse of many national health services worldwide. many neurological diseases require continuous follow-up and regular outpatient visits, and patients with stroke and epilepsy, among other neurological conditions, frequently present to the emergency every day. the impact of the pandemic in the care of patients with other neurological diseases has already been observed in many countries, raising the fear of additional load over an already overburdened health system. neurological societies are urged to devise guidelines and recommendations based on the best current information available on how neurologists should manage patients with neurological conditions that can be directly affected by covid-19, such as multiple sclerosis, epilepsy, or myasthenia gravis. another exciting field of research is about the long-term neurological consequences of sars-cov-2 infection. because a vast number of people worldwide will be infected, the inflammatory response elicited by sars-cov-2 may trigger or accelerate via impaired blood-brain barrier function some subclinical mechanisms that underlie the earliest stages of some neurodegenerative disorders. 71 prospective studies on this topic could answer this question in the future. although covid-19 does not appear to have exuberant neurological manifestations like the recent epidemics of the zika virus, hcovs have previously demonstrated the potential to invade and damage the nervous system. therefore, physicians should be alert to new clinical syndromes related to infection and be prepared to face worsening clinical control of preexisting neurological diseases in patients with covid-19. clinical characteristics of 138 hospitalized patients with 2019 novel coronavirus-infected pneumonia in wuhan, china clinical characteristics of coronavirus disease 2019 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due to respiratory virus infections key: cord-256852-lrz17bdx authors: nayyar, gaurvika m. l.; attaran, amir; clark, john p.; culzoni, m. julia; fernandez, facundo m.; herrington, james e.; kendall, megan; newton, paul n.; breman, joel g. title: responding to the pandemic of falsified medicines date: 2015-06-03 journal: am j trop med hyg doi: 10.4269/ajtmh.14-0393 sha: doc_id: 256852 cord_uid: lrz17bdx over the past decade, the number of countries reporting falsified (fake, spurious/falsely labeled/counterfeit) medicines and the types and quantities of fraudulent drugs being distributed have increased greatly. the obstacles in combating falsified pharmaceuticals include 1) lack of consensus on definitions, 2) paucity of reliable and scalable technology to detect fakes before they reach patients, 3) poor global and national leadership and accountability systems for combating this scourge, and 4) deficient manufacturing and regulatory challenges, especially in china and india where fake products often originate. the major needs to improve the quality of the world's medicines fall into three main areas: 1) research to develop and compare accurate and affordable tools to identify high-quality drugs at all levels of distribution; 2) an international convention and national legislation to facilitate production and utilization of high-quality drugs and protect all countries from the criminal and the negligent who make, distribute, and sell life-threatening products; and 3) a highly qualified, well-supported international science and public health organization that will establish standards, drug-quality surveillance, and training programs like the u.s. food and drug administration. such leadership would give authoritative guidance for countries in cooperation with national medical regulatory agencies, pharmaceutical companies, and international agencies, all of which have an urgent interest and investment in ensuring that patients throughout the world have access to good quality medicines. the organization would also advocate strongly for including targets for achieving good quality medicines in the united nations millennium development goals and sustainable development goals. malaria is a devastating illness, particularly to young children and pregnant women in tropical countries. a recent review reported that the active pharmaceutical ingredient (api) was absent in over one-third of close to 4,000 antimalarial drug samples tested from pharmacies in seven southeast asian and 21 sub-saharan african countries 1 ; over 40% of the alleged artemisinin-containing drugs were falsified, outright fakes. a wide variety of falsified brand name and generic medicines and even falsified raw ingredients for several essential pharmaceuticals have been found in rich and poor countries. [2] [3] [4] [5] [6] [7] such drugs are often used for acutely ill patients, many of whom would die or suffer prolonged illness without proper treatment. in addition to patients' loss of confidence in the health-care delivery system, microbial resistance to the drug may develop and spread if medicines contain subtherapeutic doses or no api. the increasing global scientific and public awareness and epidemic proportions of the spreading problem are reflected in the number of articles on "fake drugs" cited in pubmed: 27 until recently, there were a paucity of reports from pharmaceutical companies on the type and quantity of drugs that were fraudulently compounded or transferred by criminals. data are emerging from the pharmaceutical security institute (psi), a not-for-profit membership organization of pharmaceutical security directors, indicating that a large number of companies, products, and countries are targeted. 8 for instance, since 2008, pfizer pharmaceuticals (pfizer global security, new york, ny) has identified a rapidly increasing number of falsified products, countries reporting falsified drugs, and breaches of the legitimate supply chain national entry points (table 1) ; the increases have been from 40% to over 100%. of pfizer products, those for erectile dysfunction are most frequently falsified 9 ; other such products target patients with alzheimer's disease, cancer, high cholesterol, hypertension, malaria, and anxiety disorder. facilities where fake drugs were made or compounded were discovered with moldy walls, dirty equipment, and infested with rodents and insects ( figure 1 ). falsifiers have created products that are visually indistinguishable from the genuine product, clearly demonstrating criminal intent to deceive. this increasingly recognized problem on virtually all continents is a pandemic defined as "an epidemic occurring over a very wide area, crossing international boundaries, and usually affecting a large number of people." 10 definitions despite increasing awareness of the fraudulent drug epidemic, efforts to quantify and stop this peril have been stymied by multiple obstacles, not the least of which is agreement on definitions. 4, 11, 12 poor quality drugs include substandard/ spurious/falsely labeled/falsified/counterfeit (ssffc) medical products. 4 falsified (also commonly called fake or counterfeit products are intentionally and fraudulently produced and contain no api, the incorrect dose of the api, or the incorrect api. substandard medicines are caused by unintentional or negligent errors of manufacturing or by degradation after manufacturing resulting in insufficient api, poor dissolution properties, or degradation products. the nomenclature used by the world health organization (who), the world trade organization, the united nations (u.n.) office on drugs and crime, interpol, and others can be confusing; hence, we are using terms agreed upon by who. 4 there are also properly manufactured medicines that are unlawful for reasons apart from their quality. these can be unregistered with company branded or generic medicines that, for reasons of theft or accidental or intentional diversion, do not have the legally required marketing authorization of the country's regulators to be imported or sold there, and medicines that infringe the trademark of a legal product. relatively little is known about medicines that have expired and are repackaged with a new date; these topics along with diverted products are beyond the realm of this article. this article focuses mainly on medical and public health considerations of falsified medicines that are particularly widespread in low-and middle-income countries. [13] [14] [15] although there are increasing reports of detection of a variety of fake drugs from around the world, paradoxically, there are virtually no reports from middle-or high-income countries quantifying the state of poor quality medicines, only anecdotal case citations. governments have been hampered by a confusing array of expensive detection technologies. few functional national regulatory authorities exist in low-income nations that lack trained staff and suitably equipped laboratories to test drug quality centrally or in peripheral pharmacies or markets. 13, 14 furthermore, the variability or absence of national and international criminal statutes, lack of an international agreement against trafficking of poor quality medicines, and inadequate punishments for convicted offenders reflect the weak legal framework for confronting drug fraud. one of the biggest obstacles in provision of quality-assured pharmaceuticals is the lack of effective manufacturing, regulatory, and quality processing in india and china. in 2013, 10 global public health agencies including providers, foundations, and research institutions contributed to developing an advocacy campaign to address falsified medicines, particularly in china. this campaign called fight the fakes is a step toward raising awareness about the problem, but legal action has to follow along with more public and political awareness. 15 the u.s. institute of medicine (iom) has published a report "countering the problem of falsified and substandard drugs." 16 the iom recommendations to "stem the global trade" in such products are laudable in advising that the u.s. food and drug administration (fda), the national institute of standards and technology, and other u.s. and international pharmaceutical and financing agencies be more actively involved in setting standards and financing improvements; yet this report falls far short of making a strong call for standardized, agreed-upon quality assessment technologies; an international law convention; and a more activist, internationally recognized lead organization, all three of which are essential for stopping the many health threats of fake drugs. global leadership to date has devolved in parts to the who, the u.n. office on drugs and crime, and interpol, each with diverse missions, responsibilities, limited authorities, and their own collaborations, funding networks, cultures, and languages. 17 no organization is leading assertively. of the three areas listed, an international convention and improved national regulations are likely to have the most enduring value in concert with effective leadership and other innovations. the focus of all actions tied to drug quality must be on public and individual health, and strengthening national capacities to improve the health of their citizens. detection methods and technology. a major hindrance to understanding the types, names, extent, and amount of poor quality drugs nationally and globally has been 1) the lack of agreed-upon field survey approaches 18 and 2) available lowcost tools to detect and classify bad drugs quickly at points of entry into countries, at public and private pharmacies, and in health units. in 2014, the who published draft guidelines for surveys of medicine quality that are currently being revised. 19 two or more levels of drug quality tests exist: 1) methods useable in the field that are quick, inexpensive, and easy to use and teach; these methods are targeted mainly to examine packaging and detect drug contents and 2) technologies requiring a laboratory equipped for exhaustive chemical analysis. these approaches are summarized in table 2 , deriving from the iom report 16 and a recent analysis by green and others 20 from the centers for disease control and prevention reference laboratory. within each method there are numerous tools and prototypes being used and new ones tested. current technologies for field use rely on visual packaging inspection, lot number reporting via mobile phones, thin-layer chromatography, colorimetric tests, and simplified spectroscopic methods. gas and liquid chromatography and mass spectrometry are some of the more advanced and complex techniques for investigating drug quality in central laboratories. qualitative or semiquantitative tests for an api are not substitutes for proper manufacturing control, dissolution studies, pharmacokinetics equivalence, and supply chain integrity. a very promising recent development has been the u.s. pharmacopeia promoting the quality of medicines (pqm) program in several african, asian, and latin american countries using the "minilab" (global pharma health fund e.v., giessen, germany). [21] [22] [23] this training program supported by the u.s. agency for international development (usaid) and the president's malaria initiative (pmi) has trained several hundred persons in rapid chemical analysis of drugs taken from public and private pharmacy stocks. 23 a major reference training center has recently been opened in accra, ghana, with usaid and u.s. pharmacopeia support as a referral testing and regional training center. important also is the development of the counterfeit detection (cd)-3 (us food and drug administration, forensic chemistry center, cincinnati, oh), a promising handheld electronic device for peripheral use that detects fake packaging at point of sale with images and videos of the suspect samples. 24, 25 the fda, skoll foundation, and other partners are supporting expansion of testing and use of this device. we recommend that a precertification of essential diagnostics, drugs, and vaccines should be required for specific regional and global control, elimination, and eradication programs and campaigns. more information is needed to confirm that precertification of products is occurring for the pmi and the global fund to fight aids, tuberculosis and malaria, and products purchased by u.n. international children's emergency fund (unicef) and who. essential drugs designated by who should also be targeted for special vigilance by quality assurance mechanisms. no independent agency has inventoried and performed comparative quality assessments of these packaging and drug-testing devices and made recommendations to countries for their use. objective comparisons are needed of the diversity of field methods in terms of accuracy, reliability, costs of equipment and supplies, level of training needed, ease of use, spare part availability, and maintenance requirements. simplified standard survey protocols and methods for sampling drugs at country entry points (seaports, airports, and roads); at major pharmacy depots; in health units (public and private hospitals and clinics); and at more peripheral distribution sites (district and village pharmacies and individual vendors) are also needed. 19 low-cost, portable detection tools would empower pharmaceutical inspectors in numerous countries that have oversight of the medicine supply. results would be available promptly rather than delayed when samples are sent to national or international laboratories as occurs now; lamentably, intervals of several years have occurred from the time specimens were collected to the time the results were available to those needing to take action. 1 ideally, central reference laboratories vetted by who, fda, or another agency would back up spot checks and random sampling of pharmaceuticals at the periphery. good quality medicines by law. falsified medicines are ultimately a problem that impacts public health. the solution needs to reflect various incentives, either via financial gain, avoidance of punishment or both. a multi-sectorial effort is essential for taking into account how this illegal market is interwoven with world trade agreements, business models, and associated legal ramifications. globalization has enlarged the international trade in medicines. for example, india exports over us$15.5 billion in pharmaceuticals, which are among their most important exports. 26 as of 2011, 40% of drugs and 80% of apis for drugs in the united states are imported from foreign countries. 27 an international law convention against substandard and falsified medicines would address both regulatory and criminal international governance challenges simultaneously through technical, legal, and financial mechanisms. how would the convention work and what national benefits would it bring? a convention would provide four legal underpinnings that do not exist, that together would advance patient safety and access to quality medicines. 12 first, a convention would define the various sorts of wrongful medicines accurately and thereby avoid misunderstandings caused by today's problematic or vague terminology (e.g., where countries seized good quality generic medicines as "counterfeit"). second, a convention would promote the requirement that signatory countries enact national laws to designate wrongful actssuch as the intentional manufacture, trafficking, or selling of falsified medicines-as criminal offences, with attendant obligations to alert health-care workers and to prosecute or extradite the offenders to justice promptly. third, a convention would provide the legal and institutional framework for participating countries to agree, implement, and evolve convergent standards of medicine regulation, so as to reduce poor quality medicines in international trade. fourth, and for lower income countries particularly, a convention would contain mechanisms for financial and technical assistance, and, to join local and regional networks. these actions would help build national and regional medicine regulatory authorities (mras) to a point where patients' access to quality medicines is protected. some have said that establishing recommended codes of practice that are nonbinding (soft law) are better than international norms and regulations that are binding (hard law). 28 we disagree with soft law in regard to controlling the current fake-drug pandemic. there are precedents for using international law in this way. a 1929 treaty that internationally criminalizes counterfeit banknotes provides an analogy for falsified medicines. in the health field, there are treaties specifically addressing the illicit traffic of certain narcotic drugs and treaties to prevent harm-particularly, the framework convention on tobacco control and its associated protocols to stanch illicit trade. that convention has brought over us$250 million new funding to global tobacco-control efforts, demonstrating that international law need not compete for resources, but can increase them. 29 the u.n. office on drugs and crime has been developing "draft model legislative provisions on fraudulent medical products" for several years but there has been no agreement on final text; the focus appears to be on criminal and judicial issues. challenges ahead. information is accruing that large quantities of falsified drugs are being manufactured in asian countries. china and india are two of the largest producers of good quality drugs and vaccines, many of which are purchased or funded by the usaid; unicef; global fund to fight aids, tuberculosis and malaria; who; and other organizations, charities, and national agencies for global disease control and eradication programs. however, according to the world customs organization, in 2006, 54% and 21% of unlawful drugs of all sorts confiscated worldwide were manufactured in india and china, respectively. 30 the circuitous travel itineraries of fake medicines have been traced across continents, such that the unsuspecting recipient countries assume a bona fide origin. a particularly heinous example is that of multiples instances of the production, marketing, and international travel of falsified bevacizumab (avastin ), a cancer medicine; the fake drug closely matched the appearance of the real medicine, but tests indicated salt, starch, and various cleaning solvents instead of the active ingredient with resulting endophthalmitis. 31 the internet has opened up an unregulated opportunity for criminals to promote and sell fake drugs to unsuspecting vulnerable populations, often the aged and others seeking convenience and low cost. a recent survey of over 10,000 online pharmacies found that 96% operated outside legal regulations and a large percentage closed operations within 3 years of operation. 32, 33 the fda and other organizations participate with interpol in annual international actions (operation pangea) to shutdown illegal pharmacy websites selling potentially counterfeit and illegal medical products. more than 18,000 such illegal websites were closed during one week in 2012 with seizure of us$10.5 million of pharmaceuticals worldwide. 34 leadership, collaboration, and national strengthening. arguably, the major obstacle to solving the problem of poor quality medicines has been the lack of a clearly identified lead organization with a plan of action developed in concert with countries, pharmaceutical companies (multinational corporations and innovator/biotechnology enterprises), and national and international agencies-and a sense of urgency to implement the plan with resources and partners-including pharmaceutical companies in low-income countries. who has estimated that 30% of countries have inadequate medicines regulation authorities (mras) or none at all. moreover, who has found that 90% of african mras lack the capacity to undertake medicine regulatory functions and therefore cannot guarantee the quality, efficacy, and safety of medicines, 35, 36 the new partnership for africa's development has found that there is either limited or declining government funding for mras in the east african community partner states. 37 many have looked to who for this leadership, given its successful implementation of the public health treaty on tobacco control. however, some argue that the u.n. system, including the who, is poorly suited to be in a leadership role because of sparse technical expertise in products, manufacturing, and quality systems. u.n. agencies are beholden to member states and cannot regulate or enforce anything easily, especially, in india and china. in this regard, who could serve the role of a partner rather than a leader. the recently revitalized rapid alert program at who has begun to "track and trace" poor quality drugs as reported voluntarily by member countries. 38 rapid alert notices are published periodically by who indicating the fake drug type, lot number, quantity of product, and place detected. strong action by countries can stem the tide as shown in rwanda 39 and cambodia, 40 although unique situations and major multi-sectorial engagements exist in these countries. one solution is creation of regional harmonization networks, addressing some elements of drug registration tied to regional economic communities; the african medicines registration harmonization initiative is one example of such a network. the u.n. office on drugs and crime (unodc) has also made recent attempts at facilitating international cooperation against falsified medicines. one proposal has been a trilateral coalition of the unodc, who, and interpol. 17 still, active and transparent support from the fda, drug companies, individual countries, and other partners will be needed; the fda may be the most qualified as a leadership organization based on their technical expertise and global influence. mechanisms for training technical staff, regulating products, improving manufacturing practices, and stopping criminal production are needed to assure a good supply of medicines. given that the problem of substandard and falsified medicines should be approached primarily from a public health and equity perspective, it is important that the negotiations on the way forward be led by the ministries of health along with the ministers of finance and trade, while respecting legitimate intellectual property rights. could and should who be the lead organization in curbing the spreading epidemic of falsified pharmaceuticals? who's ability to take more assertive action is strengthened by the revised international health regulations. who's director general can convene emergency committees in response to public health emergencies as has been done recently for the influenza a (h1n1) pandemic in 2009, the middle east respiratory syndrome (mers-cov) in 2013, the polio crises in 2014, and the ebola epidemic in 2014-2015. illicit drug trafficking is an emergency. the drug quality and security act, signed into law by president obama in 2013, outlines steps to build an electronic system to identify and trace certain prescription drugs in the united states. the results of this system are awaited. finally, the millennium development goals (mdgs), under revision, should include measurable objectives for good quality drugs. this will encourage national establishment of baseline status and achievable targets, particularly for essential drugs. establishment of mdg targets and sustainable development goals (sdgs) will help greatly to solve the poor quality drug epidemic by application of available technology and good pharmaceutical vigilance and governance. 41 one incentive that would transform the current system is applying a "universal quality standard" to drug products. for example, if india allows a substandard manufacturer to sell products in africa, the fda could ban import of products from india. although difficult to develop and implement, a combination of incentives and penalties driven at the political and economic levels is needed. the major urgent needs to improve the quality of the world's medicines fall into three main areas: 1) research to develop and compare the most accurate and affordable tools to identify high-quality drugs at point of sale and deployment of the best methods; 2) an international convention and national legislation to facilitate production and use of high-quality drugs and protect all countries from the criminal and the negligent who make, distribute, and sell life-threatening products; 3) designation of a highly qualified, well-supported international organization, possibly the fda or who, that will establish standards, training programs, drug quality surveillance, and authoritative guidance for countries in cooperation with national medical regulatory agencies, pharmaceutical companies, and international agencies, all of which have an urgent interest and investment in ensuring that patients throughout the world have access to good quality medicines. the organization would also advocate strongly for including targets for achieving good quality medicines in the mdgs and sdgs including certification of pharmaceutical products entering countries that request such services, and participate in global disease control and elimination programs. poorquality antimalarial drugs in southeast asia and sub-saharan africa substandard and counterfeit medicines: a systematic review of the literature the global counterfeit drug trade: patient safety and public health risks substandard/spurious/ falsely-labeled/falsified/counterfeit medical products analysis of counterfeit cialis tablets using raman microscopy and multivariate curve resolution confirmed glyburide poisoning from ingestion of "street valium characterization and identification of suspected counterfeit miltefosine capsules empirical analysis of counterfeit drug penetration in global legitimate medicine supply chains: a descriptive assessment internetordered viagra (slidenafil citrate) is rarely genuine a dictionary of epidemiology new definition for "substandard medicines how to achieve international action on falsified and substandard medicines impact of poorquality medicines in the 'developing' world united states pharmacopeia ten global health organizations united in a worldwide campaign to protect patients from fakes countering the problem of falsified and substandard drugs improving global health governance to combat counterfeit drugs: a proposal for a unodc-who-interpol trilateral mechanism guidelines for field surveys of the quality of medicines: a proposal recommendations on the content of a survey protocol for surveys of the quality of medicines integration of novel low-cost colorometric and visual fluorescent techniques for rapid identification of falsified artemether-lumefantrine and other drugs in resource poor areas survey of the quality of selected antimalarial medicines circulating in madagascar, senegal, and uganda the global pandemic of falsified medicines: laboratory and field innovations policies and perspectives evaluation of a new handheld instrument for the detection of counterfeit artesunate by visual florescence comparison fda facts: fda's counterfeit detection device cd-3 india aims to clock usd 15.5 bn pharma exports fy13 import of human drugs and human drug components global health and the law follow the money: how the billions of dollars that flow from smokers in poor nations to companies in rich nations greatly exceed funding for global tobacco control and what might be done about it counterfeiting, a global spread, a global threat counterfeit bevacizumab and endophthalmitis progress report for state and federal regulators medicines counterfeiting is a complex problem: a review of key challenges across the supply chain fda takes action against thousands of illegal internet pharmacies. food and drug administration news release effective medicines regulation: ensuring safety, efficacy and quality availability of drug regulatory and quality assurance elements in member states of the who african region the african medicines regulatory harmonization initiative: rationale and benefits international medical products anti-counterfeiting taskforce (impact) combating substandard and falsified medicines: a view from rwanda quality of antimalarials at the epicenter of antimalarial drug resistance: results from an overt and mystery client survey in cambodia why the mdgs need good governance in pharmaceutical systems to promote global health this is an open-access article distributed under the terms of the creative commons attribution license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-309900-4nln90jn authors: doornekamp, laura; stegers-jager, karen m.; vlek, odette m.; klop, tanja; goeijenbier, marco; van gorp, eric c. m. title: experience with a multinational, secondary school education module with a focus on prevention of virus infections date: 2017-07-12 journal: am j trop med hyg doi: 10.4269/ajtmh.16-0661 sha: doc_id: 309900 cord_uid: 4nln90jn worldwide, virus infections are responsible for many diseases in terms of morbidity and mortality. vaccinations and therapies are only available for relatively few virus infections and not always where they are needed. however, knowledge of transmission routes can prevent virus infection. in the context of this study, we measured the effects of a secondary school education module, named viruskenner, on knowledge, attitude, and risk behavior as these relate to virus infections. a nonrandomized intervention study was conducted between april and august 2015 to assess the effect of this 2-month education module on knowledge, attitude, and behavior of 684 secondary school students in the netherlands, suriname, and indonesia. for the netherlands, a control group of a further 184 students was added. factor analysis was performed on questions pertaining to attitude and behavior. comparative analyses between preand posttest per country were done using multiple linear regression, independent sample t-tests, and one-way analysis of variance. these showed a significant increase in knowledge about virus infections and the prevention of infectious diseases among the dutch and surinamese groups, whereas a trend of increased knowledge was evident among the indonesian participants. the dutch control group showed an overall decrease in knowledge. regression analyses showed that there was a significant interaction effect between participation and time on knowledge, attitude, and awareness and behavior and risk infection. attitudes improved significantly in the intervention group. pearson correlation coefficients between knowledge, attitude, and behavior were found to be positive. viruses are responsible for almost half of all emerging infections worldwide and are among the most emerging pathogens. [1] [2] [3] most virus infections are not treatable with antivirals and neither are they preventable with vaccines. therefore, education plays a key role in raising awareness for infectious diseases and preventing the spread of virus infections. 4 a population that is aware of the different ways a virus can be transmitted and does know how to embed effective preventive methods in daily life can protect themselves against virus infections. this principle is based on the knowledge, attitude, behavior (kab) model, showing that increased knowledge can change people's attitudes and lead to behavioral change. [5] [6] [7] a foundation for health-related attitudes and behavior is laid in early stages of life. following many theories, the likelihood of changing attitude is high in adolescence. 8 also, the world health organization (who) states with their health promoting school framework that schools are a good environment to start promoting health. 9 five years ago, a consortium of scientists and teachers developed a school-based education module named viruskenner, regarding virus infections. this module aimed to teach students how to prevent virus infections. the module started with one secondary school in the netherlands and evolved to a project with eight different secondary schools in the netherlands. in 2014, the first surinamese school joined and in 2015 the first indonesian school. these two countries were already involved with the organizing institute by an international collaboration on emerging infectious disease population studies, facilitating easy communication and logistics. the viruskenner module was extensively evaluated by independent researchers in the early years of the project. the conclusions of these evaluations led to improvement of the module and the questionnaires used. for example, in 2012, the concept of students being coached by an infectious disease expert was introduced. when becoming an international education module, it was interesting to see the impact of viruskenner in different countries on knowledge, attitudes, and behavior as they relate to virus infections, and find out which educational factors play a role in these changes. educational programs that address infectious diseases are quite common, although most education is focused on a specific infection or a group of infections, particularly human immunodeficiency virus (hiv) and other sexually transmitted infections. a recent systematic review and meta-analysis evaluated 64 school-based sex education programs in middleand low-income countries. most of these programs (55 of 64) focused on comprehensive sex education, with the remaining nine focusing on sexual abstinence. about half of the studies (33) were included in the meta-analysis and showed an overall positive effect on hiv-related knowledge, condom use, the initiation of sexual intercourse, the number of sexual partners, and self-efficacy. 10 although hiv is among the virus infections that place the highest burden on society, it is not the only virus that significantly impacts global health. besides hiv, lower respiratory tract infections (e.g., influenza), and diarrheal diseases (like norovirus) also belong to the 10 leading contributors to the global burden of disease. 11 furthermore, arthropodborne diseases (like dengue) have a very high incidence. 12 remarkably, virus infections other than hiv are less frequently addressed in education modules. for example, only a few trials were carried out to measure the impact of an educational intervention for viral hepatitis, human papillomavirus (hpv), dengue, and influenza. [13] [14] [15] [16] [17] most of the educational interventions that were analyzed showed positive results in improving knowledge and attitude pertaining to the subject of the intervention. given the success of education programs about hiv, education modules about other virus infections that have global impact might also work. we developed an education module that focuses on multiple viruses with different transmission routes and all with global impact: hiv, dengue, hantavirus, chikungunya, middle east respiratory syndrome (mers) coronavirus, hpv, norovirus, viral hepatitis, measles, and influenza. we studied the efficacy and success of the education module in three countries, netherlands, suriname, and indonesia, each differing in culture, circulating viruses, and infection pressure. the education module aims to effectively increase knowledge, attitudes, and behavior regarding several virus infections in each of these different circumstances. participants and setting. schools in three countries participated in this nonrandomized intervention study: four in the netherlands, a high-income country in europe; one in suriname, an upper middle-income country in latin-america; and one in indonesia, a lower middle income country in southeast asia. 18 the effect of the education module was measured per country, by comparing the results of a pre-and posttest. the situations per country, for example, culture and school system, were too different to make a fair comparison between countries. however, the target group for the education module is the same in each country and the concept of the module and measurements were as comparable as possible. secondary schools in the netherlands, suriname, and indonesia had been invited to apply to participate in the education module with their 10th grade students (generally 14 or 15 years of age). all schools were well-known public schools for students with an above-average socioeconomic status. the school in suriname that participated had about 840 students and was located in paramaribo, the capital and largest city of the country in inhabitants. the indonesian school that participated was located in surabaya, the second largest city in the country. this was a senior high school (grade [10] [11] [12] and had about 1,200 students. the four schools in the netherlands ranged in number of students from 1,600 to 2,400 and were from different regions but all in the dutch urban agglomeration, including one school from amsterdam, the capital city of the netherlands. the 10th grade is the final stage of the junior high school in the netherlands, which means that all students have, until then, followed the same subjects and have expressed their interest in the choice for a special curriculum. for example, a beta scientific curriculum, which includes the following subjects: biology, physics, chemistry, and mathematics. in the netherlands, the schools that were invited to participate were all schools that offer students an option for technasium, which is an elective course for students interested in beta scientific subjects. 19 the participating students had all chosen this special curriculum with additional technical courses. information about the module was disseminated via the project website and the technasium network coordinator. a control group for the dutch intervention group was selected at one of the participating dutch schools. thus, although they had not opted for the technasium curriculum, they do have a similar background and social environment. school curricula are defined differently in each country. in the netherlands, students choose a profile and we defined "nature and science" and "nature and health" as scientific profiles. in suriname, students can choose biology, and we defined this as a scientific profile. indonesian students can choose between a social profile (ilmu pengetahuan sosial [ips]) and a science profile (ilmu pengetahuan alam [ipa]). we defined ipa as a scientific profile. in both suriname and indonesia, schools that matched most closely, in terms of grade and education level, with the dutch intervention group were invited to participate in the study. of the dutch schools, two were preuniversity education level (known in the netherlands as "vwo") and two were mixed preuniversity education level and advanced general secondary education (known in the netherlands as "havo"). the surinamese participants were from one vwo school, which is comparable with the dutch vwo education level. these surinamese participants can therefore be seen as preuniversity education level. 20 the indonesian participants were from one sekolah menengah atas (sma) (high school), which is comparable with havo in the netherlands and internationally known as advanced general secondary education. 21 the dutch control group consisted of students with preuniversity education level and advanced general secondary education level. design of the intervention. the viruskenner education module is based on the "learning-by-doing" principle. students are challenged to create a prevention tool for a specific virus infection. by involving students in real-life science-based problems and stimulating active learning (searching for information, test possible solutions and present their idea) a high impact can be achieved. [22] [23] [24] in each country, the 2-month module started with a national opening day, during which all participants of that country were introduced to the field of infectious diseases and viruses by means of four short lectures from experts in the field of virology, public health, and infectious diseases. an optimal learning effect can be reached by bringing students in contact with experts. 25 so, in all countries one or two dutch experts from the department of viroscience in the erasmus medical center in rotterdam were assigned to a class to coach them during the project. the students were supposed to work in groups of four to six students in competition with the other groups. 22, 26 each group worked on one of the viruses of the subject list including hiv, dengue, hantavirus, chikungunya, mers coronavirus, hpv, norovirus, viral hepatitis, measles, and influenza. students developed a prevention tool to disseminate this knowledge among their peers and, in doing so, help prevent virus infections that impact local or global health. during the three national final days (one in each country), the best groups per class, selected by the teachers and coach during a school final, presented their results and final product to their peer students and a jury. this independent jury was selected per country and based on proven expertise in virology, communication strategy, and/or overall creativity. in each country, the jury chose two winners: the most informative presented prevention tool and the most creative prevention tool. the study was conducted between april and august 2015. a pretest was performed 1 or 2 days before the start of the module to assess their basic knowledge, attitude, and behavior; a posttest 5-7 days after its completion to let the information settle in their memory and give the students some time to evaluate their attitude and behavior a few days after the final day. other measurement instruments were used to get additional information ( figure 1 ). during the intervention, students could use the modules' website (www.viruskenner.nl) and other supportive resources, like a youtube channel and a facebook page (all in dutch and english and available for all participating countries), to find more information on the project and on virus infections and to disseminate information about their prevention tools. [27] [28] [29] instruments. the effect of the education module was measured by the kab model. given that there was no validated instrument to assess knowledge, attitude, and behavior regarding several viral infections, we used a self-designed questionnaire (supplemental table 1 ). the questionnaire was based on 5 years' experience with the education module. a team comprising two senior virologists, a communication scientist, and an education expert developed the questionnaire, which was refined after a pilot with a group of 60 students from a school similar to the participating schools. the questionnaires used for the dutch and surinamese schools were in dutch. the questionnaires for the indonesian students were first translated into english by a dutch researcher and then into indonesian by a native speaker of the language. the pre-and posttest questionnaires addressed five areas: 1) sociodemographic factors, 2) stigma and fear, 3) attitude and behavior, 4) knowledge on viruses and infectious diseases in general, and 5) the opportunity to write down questions or comments about the questionnaire or module. the posttest had an additional category-6) perceptions of the project. a principal component analysis (pca) with varimax rotation was performed for the attitude and behavioral questions on the results of the pretest questionnaires, as suggested in literature. 30 varimax was the preferred rotation because this results in a small number of factors per variable and a small number of variables per factor. this is the most popular type of rotation because it makes the interpretation of the data more reliable and easier. 31 one of the behavior items ("i do not use a condom when i have sexual intercourse") was excluded because of more than 10% missing values. the remaining missing values were randomly spread over the sample population. the sample size was big enough to delete these cases list wise. the kaiser-meyer-olkin (kmo) value is a statistic that measures how much two random variables correlate. a kmo value greater than 0.8 represents a small partial correlation which makes a factor analysis more useful. in this study, the kmo value was 0.849, which means there were relatively compact patterns of correlations and the factor analysis would provide reliable components. 32 the number of extracted factors was based on the objective and interpretability criteria mentioned in schönrock-adema and others: 1) the screen test, 2) eigenvalues > 1.5, 3) > 5% of the variance explained by all factors, and 4) interpretability. however, the criterion of eigenvalue > 1.5 led to only two components, which was not interpretable. therefore, we set the norm of an eigenvalue back to greater than one (kaiser's criterion). 33, 34 the pca with varimax rotation finally resulted in four components. the reliability per component was calculated by cronbach's alpha (table 1 ). internal consistency for the components "attitude and awareness" and "behavior and life science" was above 0.7 and therefore acceptable. the components "attitude and risk infections" and "behavior and risk infection" should be interpreted with caution, because of the diversity of the constructs. 35 an additional instrument to measure knowledge was a live multiple-choice quiz, which was implemented at the end of the final day. in the netherlands and suriname, portable electronic devices (keypad and software from interactive voting system ® ) were used by the students to answer 40 knowledge questions. in indonesia, these portable electronic devices were not available, so the knowledge quiz was done by voting with colored papers; therefore, recording these results was not possible. to obtain more information about factors that influenced the impact of the education module, teachers of all four participating schools in the netherlands were interviewed when they had completed the education module. the aim of this additional qualitative component was to determine possible confounders which might have influenced the difference in outcomes between the pre-and posttests and to find out whether the teachers noticed increased knowledge or improved attitude and/or behavior among their students. although the teacher interviews were carried out in the netherlands only, the module was evaluated in each country. in suriname and indonesia, the project was evaluated with the local organizing teams but not per individual teacher. in the indonesian and surinamese culture, hierarchy is strong and extensive evaluation uncommon. therefore, the teachers preferred a general evaluation with the head of the school. however, we do feel these interviews were less helpful because the heads of the schools were not closely involved in the project. the dutch teacher interviews were semistructured and took about 30 minutes each. questions that were asked included "how was the contact with the coaches?" and "what have the students learned during the project?" teachers were interviewed in their classrooms after the classes had filled out the posttest questionnaire. finally, user data from the website and social media were analyzed after the completion of the module to find out which supportive resources were most popular during the education module (supplemental figure 1) . outcomes. the primary outcomes in this study were knowledge, attitude, and behavior and stigma and fear. stigma and fear, attitude, behavior were measured on a 5point likert scale. all these outcomes ranged from 1 (strongly disagree) to 5 (strongly agree). stigma and fear were measured with two questions and mean values were calculated. the outcome "stigma" was used in this study to describe a negative thought regarding people with a hiv infection. the stigma was expected to be high before the module started. by gaining knowledge, the stigma could be decreased. the outcome "fear" in this study aims to measure how afraid people are to get infected in case of a large outbreak; at the time of this study, ebola was the best example. the attitude and behavior questions were subdivided into four components by the factor analysis and the mean score per component was calculated. (table 1) . unstandardized coefficients (b) as outcomes of the regression analysis showed which factors contribute significantly to these four attitude and behavior components and to knowledge. the outcome knowledge represented the student's knowledge regarding infectious diseases in general and the viruses in specific that were included in the education module. knowledge was measured in the questionnaire by means of the responses to 32 questions, which had to be answered with "true" or "false." each correct answer resulted in one point, an incorrect answer in zero points. the mean percentage of all knowledge questions that were answered correctly was calculated per group and ranged from 0% to 100%. knowledge outcomes from the quiz were calculated in percentages. as a secondary endpoint, the perceptions of the students about their participation in the project were evaluated. ten statements measured if students enjoyed working on the project and if they thought the project was informative. this was scored on a 5-point likert scale and ranged from 1 (strongly disagree) to 5 (strongly agree). data management and analysis. the questionnaires were read by the open source optical mark recognition program sdaps (benjamin berg, karlsruhe). correct reading was checked manually by two different persons. all data were imported into one database and analyzed with ibm spss version 21. all questionnaires in which less than 90% of the knowledge questions had been answered were excluded. cases that showed a variance equal to zero in the likert scale questions were excluded for analysis on these outcomes. items with more than 10% missing values were deleted. in all analyses, p < 0.05 was considered significant. descriptive analyses were performed to calculate the frequencies of students' characteristics and pearson's χ 2 test was used to identify significant differences between the characteristics of the groups in the pre-and posttest. correlations between knowledge, attitude, and behavior were calculated for all students in the intervention group, with a pearson's coefficient. the average knowledge per country in the pre-and posttest situations was compared by an independent sample t-test. effect sizes (es) were calculated with cohen's d. effect sizes greater than or equal to 0.30 were considered medium, and those greater than or equal to 0.50 as large. 36 multiple linear regression analysis was used to find factors that influenced the knowledge, attitude, and behavior outcomes. time point (pre-and posttest) and participation (intervention and control group) and the interaction between these two variables were added as independent variables, as well as gender, age, education level, school, and country. tolerance values were computed to assess multicollinearity. values below 0.2 were viewed as potentially problematic. 30 stigma and fear were compared between pre-and posttest with a one-way analysis of variance (anova). the sample size allowed us to calculate differences between the components of the factor analysis in pre-and posttest per country with a one-way anova test. perceptions of the project were measured only after the module had finished. means and standard deviation were summarized per country. the data set is provided in the supplementary materials. ethics. the study was carried out in accordance with the declaration of helsinki. according to dutch law, this study was exempt from medical ethical approval requirements. the technasium network in the netherlands approved this study to be performed at the dutch technasium schools and informed the students and parents. in suriname and indonesia, the headmasters of the schools approved conducting the viruskenner module and evaluations at their schools and informed the students and their parents. participation was voluntary and anonymity was guaranteed. participants and setting. in 2015, a total of 684 (of 738) secondary school students participated in the viruskenner education module. two of the participating schools in the netherlands, dropped out (54 of 260 dutch students, representing 20.7% of them) because of noncompletion of the module and evaluation program. one school dropped out because the teacher got sick after the kick-off and the other school could not attend the final day because it clashed with another school activity that day (figure 2 ). in suriname, 158 students participated and there was no dropout. this was also the case in indonesia, where all 320 students completed the education module. response rates for the netherlands were 95.6% of participants for the pretest and 70.9% for the posttest. in suriname, these percentages were 90.5 and 96.8%, respectively, and for indonesia 97.2 and 99.1%, respectively. the control group had a response rate of 100% for the pretest and 73.4% for the posttest. table 2 presents the pre-and posttest characteristics of the module participants from all three countries and the control group. in all groups, except the surinamese group, the age category in the posttest was significantly higher than in the pretest. however, for gender and education, the characteristics did not show any significant differences between the pre-and posttest per country. in indonesia, the preference for science was significantly higher in the posttest than in the pretest. on average, in the netherlands, most of the participating students were male, whereas in indonesia, they were mostly female. generally speaking, in the netherlands, the students from both the control group and the intervention group were significantly younger than average. the pretest showed that on average more students attended preuniversity education in the netherlands, suriname, and in the control group. in indonesia, all students attended advanced general secondary education. in the posttest, the percentage of preuniversity education students in the dutch intervention group rose to 65.3%. in the control group, this percentage decreased to 64.1%. in both the pre-and posttest, the amount of participants from the intervention group in the netherlands and suriname that chose scientific profiles was not significantly different from the average. the control group consisted of less students with scientific profiles than average and indonesia had more students with science-related profiles. correlations between knowledge score and attitude and behavior. pearson's coefficients showed a positive and significant correlation between the knowledge scores and all four components regarding attitude and behavior (table 3) . knowledge was most strongly correlated with attitude and awareness (r = 0.20). students who scored higher on attitude and awareness also scored higher on behavior regarding risk of infection (r = 0.47) and behavior regarding life sciences (r = 0.51). knowledge. during the project, the answer to one of the 32 knowledge questions changed, due to the mers epidemic in south korea. because of the confusion surrounding this question, we decided to exclude it from the analysis. analyses per country showed differences in achieved knowledge ( figure 3) , with mean knowledge increasing in all three participating countries. for suriname and the netherlands, this increase was significant (p < 0.001). the overall effect size (cohen's d) for all intervention groups was 0.43, which represents a medium effect. at 0.77, the effect size for suriname was the highest. the effect size for the netherlands was 0.52, which also represents a large the given percentages have been calculated from the number of students for which data is available for that variable. the percentages have been rounded off to one decimal place. pearson chisquare was used to calculate differences per country and the control group between pre-and post-test. t = 0 represents the pre-test and t = 1 represents the post-test after 10 weeks. education represents the level of education, in which havo stands for advanced general secondary education and vwo stands for pre-university education level. science represents the interest of the students, measured by their (preferred) choice of curriculum. figure 3. the impact of the viruskenner on students' knowledge. the knowledge of the participating and nonparticipating students per country before and after the intervention is represented by the mean percentage of the true/false questions in the questionnaire that were answered correctly. the blue line represents the netherlands, without the control group. the orange line represents all intervention groups, so from the netherlands, suriname, and indonesia. * p < 0.05; ** p < 0.01; *** p < 0.001. this figure appears in color at www.ajtmh.org. the correlation coefficients shown have been calculated from all values in the intervention groups at both the pre-and posttest. * p < 0.05 ** p < 0.01 *** p < 0.001. effect. 36 for example, in the netherlands, the percentage of correct answers on the statement "dengue is a virus infection that is transmitted by a tiger mosquito" raised from 71% correct in the pretest to 90% correct in the posttest (the correct answer is true). the score for "if someone is infected with hiv this person has aids" raised from 76% to 83% (the correct answer is false because acquired immunodeficiency syndrome is a syndrome in which the immune system is suppressed and opportunistic infections can cause illness, which can be prevented in hiv infected individuals by taking antiretrovirals).although in some other questions the percentage of correct answers differs only one percentage point between the pre-and posttest. the mean percentage of correct answers on a few questions declined. the mean total knowledge in the control group decreased significantly (p = 0.032). in the multiple regression analyses, the variable participation (control group or intervention group) contributed significantly (p < 0.001; b = 0.078) to the knowledge outcome. the variable time point (pre-or posttest), however, did not. most information about the impact of the module on knowledge is given by the interaction between participation and time point, which was significant (p < 0.001; b = 0.053). other variables that contributed significantly to knowledge were gender, age > 16 years, and the school ( table 4 ). the mean tolerance of all variables in the regression analyses is 0.3. although this suggests that there is some multicollinearity between predictors, this value is no reason for concern. 30 the data from the knowledge quiz showed that the netherlands had a mean score of 70.7%, with suriname scoring 59.6%. stigma and fear. the first question regarding stigma and fear was "i don't want to mix with people who have hiv" and the second one was "i am afraid that i will get infected by ebola." generally speaking, the module participants' answers did not change significantly. however, the results per country showed a significant decrease in suriname on both questions (p = 0.009; effect size [es] = 0.31 and p = 0.001; es = 0.42, respectively); other countries showed no significant differences in separate analyses, neither did the control group. figure 4 shows the changes in the four components regarding attitude and behavior. in the intervention group (all countries combined), attitude and awareness increased significantly (p = 0.028; es = 0.12) and so did attitude and risk infection (p < 0.001; es = 0.30). behavior and risk infection increased, but with the chosen p value of 0.05, the increase was on the borderline of significance (p = 0.062; es = 0.11). behavior and life sciences also increased with borderline significance (p = 0.060; es = 0.10). in the control group, attitude and awareness and behavior and risk infection decreased significantly (p < 0.001; es = 0.67 and p < 0.001; es = 0.51, respectively). attitude and risk infection and behavior and life sciences both showed a slight, but nonsignificant increase. although attitude, and even behavior, in the intervention group seemed to increase, in the subanalysis per country, we only found a significant increase in attitude and risk infection for the netherlands and suriname (figure 4) . the multiple regression analysis showed that as main effects, participation and time point both contributed significantly to attitude and awareness. we also found significant interaction between participation and time point for this outcome (p < 0.001; b = 0.430). the independent variables gender, school 2, education level, and countries also contributed significantly to attitude and awareness. for the attitude and risk infection outcome, only surinamese students had higher scores (p < 0.05; b = 0.213). for behavior and risk infection, the main variable participation was not significant. but time point was, and it had a negative effect (p < 0.001; b = _ 0.288). the interaction between these two resulted in a significantly positive effect (p < 0.001; b = 0.310). being older, the school and the country also contributed significantly, to behavior and risk infection. behavior and life sciences were influenced by participation in the module; however, time point had no significant effect and the interaction was not significant either. countries and education level, however, did contribute significantly to behavior and life sciences ( table 4) . appreciation of the project. generally speaking, the students enjoyed participating in the education module and said that it taught them a lot about infectious diseases. the score on the statement "i enjoyed working on the project viruskenner" was measured on a scale from 1 (totally disagree) to 5 (totally agree). the mean score in the netherlands was 3.2, in suriname 4.37, and in indonesia 3.93. in total, 90% of all students that participated gave a score of 3 or higher. supplemental table 1 in the supplementary data reports how they answered the other evaluation questions. teacher interviews. the first and second school participated in the project for 5 and 4 years, respectively, but the head teacher of the first school was involved for the first time. the project was completely new for the third and fourth schools. the first school allowed the most time for students to work on the project, 6 hours a week for 8 weeks. the fourth school allowed 5 hours a week for 8 weeks. the second and third schools allowed 5 and 4 hours a week, respectively, for 6 weeks in each case. in the first three schools, the students had no other lessons about viruses during the project period; only in school four did the biology teacher pay some extra attention to them. none of the teachers let the students prepare for the kick-off, but the teacher in the fourth school told them to read the manual. in the first and fourth schools the students themselves decided on the composition of the collaboration groups and chose their subject of preference. students in the second school also decided their group composition, but straws were drawn to allocate the subjects. the teacher in the third school divided the students and subjects over the groups randomly. all teachers reported that contact with their coaches during the project was good, although it has to be said that there were some communication problems with the teachers in the second school. and while school number three's teacher said that the contact with the coach was very helpful and amicable, he added that the students got to learn more about the world of scientific research, and that this aspect might have been emphasized even more. another remark was made by a teacher in the first school, who said that the website should be promoted for learning purposes more frequently and contact with the coaches could be more intensive. all teachers responded positively to the question: "what do you think the students learned from the module?" the teacher of the first school said he thought students are now more aware of the worldwide impact of infectious diseases. he even remarked that during the break on the final day he noticed that more students washed their hands after going to the bathroom. the teacher in the second school insisted that students are now more focused on viruses in the news, such as ebola or mers, and that there is a gap between knowing and doing. finally, a teacher in the fourth school concluded that during the completion of the posttest questionnaires he got the distinct impression that the students learned a lot. user data online resources. although the education was mainly face to face, online supportive resources were available to increase the educational impact. the graph in s1 shows the use of the website and social media in time. 27 after adjusting for age, sex, education level, school, and country, viruskenner proved to be an effective education module for increasing the knowledge of young people in the netherlands and suriname of virus infections, according to this nonrandomized intervention study. with all limitations of this study design taken in mind, we describe a positive correlation in knowledge, attitude, and behavior in the participating secondary school students. participation had a positive effect on attitude and awareness. this effect was higher among females and students who had attained a higher level of education. knowledge, behavior, and risk infection were higher in female and older students (16+). and while the attitude components increased in the figure 4 . the impact of the education module on students' attitude and behavior. the graphs illustrate the changes per country in the four different components of the attitude and behavior questions that were answered on a 5-point likert scale. a higher score represents a more positive attitude or healthier behavior. panel a shows the score for the attitude and aswareness component, panel b for attitude and risk infection, panel c for behavior and risk infection and panel d for behavior and life sciences. the blue line represents the netherlands, without the control group. the orange line represents all intervention groups, so from the netherlands, suriname, and indonesia. * p < 0.05; ** p < 0.01; *** p < 0.001. this figure appears in color at www.ajtmh.org. intervention group, the behavior components only showed an increasing trend. there was no significant effect of participation on attitude and risk infection, but there was on behavior and risk infection. this might be explained by the positive effect in the control group for attitude and risk infection but negative for behavior and risk infection. it might be due to there being less motivation in the control group to fill in the questionnaires. the education module had less impact on students' knowledge in indonesia. the somewhat limited impact on indonesian participants could be explained by their lower level of involvement. 37 all students in the netherlands and suriname developed a prevention tool and prepared a presentation. however, the evaluations found that in indonesia only a selection of the students did. additionally, in suriname (and partly in the netherlands), family members were invited to attend the final day. in indonesia, this was not possible due to the limited space. the engagement of families could well have had a positive effect. another striking fact in indonesia was the relatively high scores for attitude and awareness and behavior, in both pre-and posttests. the same was true for suriname, which might point to cultural differences with the netherlands. collectivistic countries, like suriname and indonesia, tend to give more socially desirable answers to questionnaires than individualistic countries like the netherlands. 38 overall, most students of all countries enjoyed working on the project. although most outcomes in the intervention group showed a positive trend or change, in the control group knowledge, attitude, and awareness and behavior and risk infection decreased. these students did not differ significantly in gender, education level, or profile between pre-and posttest. the decreased outcomes might be explained by reduced motivation in doing the same test twice. to our knowledge, this is the first study to evaluate an education module on several viruses in several continents. the heterogeneity of the study population increases the external validity of the study. comparing the results of the same education module in different countries gave insights in the importance of educational factors on the impact. in each country, the pre-and posttests were compared. however, a limitation of the study is that only the netherlands had a control group. the control group consisted of more students that had chosen a nonscience curriculum than the intervention group. however, there was no significant difference in knowledge score between the nonscience and science students in the control group. although science students scored higher on attitude and awareness and behavior and life science questions. although the time that schools spend on the project differed, no direct relationship was found between the hours spent and the results achieved. making the results translatable to schools that could participate to the module and would spend at least 4 hours per week during 6 weeks. due to logistics, randomization of schools was not possible. for a maximum effect, it is important to embed the project in the curriculum, so schools were chosen by a curriculum in which it would fit, as it is in the netherlands with technasium. multiple participating schools per country would have added value as it would have enabled us to perform a proper multilevel analysis, instead of a multiple linear regression analysis. furthermore, it might be good to measure any balancing measure, for example, the mean grades, to determine whether there are any unanticipated harms to scores on other subjects in school, due to the time the students spent on the project. although the project is embedded in the curriculum, the harms to other subject would be minimized. the questionnaires were composed with accuracy in dutch (the national language in the netherlands as well as in suriname). the ones that were used in indonesia were translated to bahasa indonesia without back translation. self-reported questionnaires are useful to measure knowledge changes. however, self-reported attitude and behavior have to be interpreted with caution. the effect was little and could even be due to overestimation. the effect of participation in the module on knowledge, however, was large in two of three countries. measuring a long-term effect in these countries as well would be of additional value. a clear effect on knowledge, but a negligible or nonexistent effect on attitude and behavior is common in educational research. several studies pertaining to hiv or sex education show that knowledge increased after participation in an education module. 10, 39, 40 we only found a few studies in which peer education did not increase knowledge. 41, 42 the literature about stigma, awareness, and attitude is inconclusive. some studies conclude that awareness can be increased or that attitudes can be changed, whereas others conclude that the effects on these is limited. [41] [42] [43] [44] [45] [46] [47] [48] [49] [50] [51] according to the available literature, behavior is the most difficult part to measure and improve. most studies about virus education evaluate hiv prevention programs. condom use or the intention to use condoms are measured most frequently. in self-reported questionnaires these outcomes improved significantly in some studies, which is promising. however, other studies did not find a significant improvement. 10, 41, 45, 47, 52 we found two studies, both conducted in africa, that tested behavioral change based on the prevalence of virus infections. in them, participants' blood samples were tested for hiv and herpes simplex virus antibodies, before and 3-8 years after an education module or compared with a control group. however, no significant differences were evident in infection rates. 53, 54 so what factors play a role in making an educational intervention effective in changing behavior? we found some studies that based their intervention on the health promoting school framework of the who proved to be successful in changing health-related behavior. 55 important elements of this framework which were applied in these studies were implementation of the intervention in the school curriculum, involvement of the school environment in the project, and involvement of family and society in the intervention. 55, 56 the viruskenner education module was implemented in the technasium curriculum in the netherlands, but was not part of the curriculum in suriname and indonesia. the family and society were involved in the project, particularly in the netherlands and suriname. however, stronger involvement of the school environment and ethos in prevention of infectious diseases might increase the impact of the intervention on attitude and behavior in all countries. this might be reached by additional interventions like handwashing posters in the sanitary facilities or selling machines for mosquito nets in the schoolyard, for example. knowledge that is not translated into behavior change would not make a difference in numbers of virus infections. so adjustments to the viruskenner module are needed to have a greater impact on attitude and behavior. active learning has the best chance of being successful if every individual student participates. students' families have to be closer involved and a sharper focus on infection prevention in school environments is needed. increasing knowledge is a great first step, because it correlates with attitude and behavior. however, significant improvements in attitude and behavior must be reached to have a possible impact on infection rates. therefore, further exploration of contributing elements of education modules that reached behavioral changes would be very useful. global trends in emerging infectious diseases diseases of humans and their domestic mammals: pathogen characteristics, host range and the risk of emergence population biology of emerging and reemerging pathogens the spread of awareness and its impact on epidemic outbreaks health impacts of education: a review matching the message to the process: the relative ordering of knowledge, attitudes, and practices in behavior change research the knowledge, attitudes, and behaviors approach how to evaluate performance and learning in complex environments development of attitude strength over the life cycle: surge and decline the world health organization's health promoting schools framework: a cochrane systematic review and meta-analysis school based sex education and hiv prevention in lowand middle-income countries: a systematic review and metaanalysis dengue and severe dengue evaluation of the hepatitis b prevention education programme in poland an intervention study for viral hepatitis. peer-led health education among high school students the effect of school-based cervical cancer education on perceptions towards human papillomavirus vaccination among hong kong chinese adolescent girls an educational programme on dengue fever prevention and control for females in jeddah high schools effects of an influenza prevention program using non-pharmaceutical prevention measures to improve the knowledge, attitudes and practices of elementary school students in nakhon phanom province countries and economies stichting technasium the surinamese education system described and compared with the dutch system the indonesian education system described and compared with the dutch system learning effects of an international group competition project where's the evidence that active learning works? a conceptual framework for organizing active learning experiences in biology instruction doing science at the elbows of experts: issues related to the science apprenticeship camp is peer interaction necessary for optimal active learning? viruskenner: knowledge is the antidote viruskenner facebook page discovering statistics with spss factor rotations in factor analysis. encyclopedia of social sciences research methods an index of factorial simplicity necessary steps in factor analysis: enhancing validation studies of educational instruments. the pheem applied to clerks as an example the application of electronic computers to factor analysis the handbook of psychological testing a visitor's guide to effect sizes: statistical significance versus practical (clinical) importance of research findings does active learning work? a review of the research social desirability in crosscultural research evaluation of a school-based hiv prevention intervention among yemeni adolescents effects of a rapid peer-based hiv/aids educational intervention on knowledge and attitudes of high school students in a high-income arab country effectiveness of a peer-led hiv prevention intervention in secondary schools in rwanda: results from a non-randomized controlled trial an evaluation of a peerbased hiv/aids education program as implemented in a suburban high school setting effectiveness of school-based education on hiv/aids knowledge, attitude, and behavior among secondary school students in wuhan characteristics of effective interventions in improving young people's sexual health: a review of reviews effectiveness of a school hiv/aids prevention program for spanish adolescents evaluation of hiv/aids secondary school peer education in rural nigeria longitudinal study of a school based hiv/aids early prevention program for mexican adolescents effectiveness of an hiv prevention program for secondary school students in mongolia the effects of a social-cognitive method based education on knowledge and attitudes intentions with respect to hiv transmition among school learners in maragheh acquired immunodeficiency syndrome educational program: effects on adolescents' knowledge and attitudes a systematic review of school-based sexual health interventions to prevent sti/hiv in sub-saharan africa effects of the culturallysensitive comprehensive sex education programme among thai secondary school students long-term biological and behavioural impact of an adolescent sexual health intervention in tanzania: follow-up survey of the community-based mema kwa vijana trial the regai dzive shiri project: results of a randomized trial of an hiv prevention intervention for youth the who health promoting school framework for improving the health and well-being of students and their academic achievement reducing obesity via a school-based interdisciplinary intervention among youth: planet health acknowledgments: we would like to thank the european union for the assignment of the erasmus+ grant (ka2-cooperation for innovation and the exchange of good practices-capacity building in the field of youth), the viroscience department, and cirion foundation for sponsoring the project. we would also like to thank erik sickmann, georgina arron, and wilco zwennis for their contributions in the planning and management of the module. furthermore, we would like to thank benjamin berg, who developed the sdaps software to process the questionnaires and henri starmans for processing the scans of the questionnaires. finally, we would, of course, like to thank all participating schools and students for their invaluable input and time. this is an open-access article distributed under the terms of the creative commons attribution license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-344503-gw5d721n authors: yousefzadegan, sedigheh; rezaei, nima title: case report: death due to covid-19 in three brothers date: 2020-04-10 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0240 sha: doc_id: 344503 cord_uid: gw5d721n we report fatal cases of novel coronavirus disease (covid-19) in three brothers in iran. an increased susceptibility to specific pathogens has been reported for a number of genetic defects. considering the fact that most of them who are affected by covid-19 recover, deaths in three brothers who lived separately and had no known underlying disease suggest genetic predisposition to covid-19 in some individuals. abstract. we report fatal cases of novel coronavirus disease in three brothers in iran. an increased susceptibility to specific pathogens has been reported for a number of genetic defects. considering the fact that most of them who are affected by covid-19 recover, deaths in three brothers who lived separately and had no known underlying disease suggest genetic predisposition to covid-19 in some individuals. novel coronavirus disease (covid-19), caused by infection with severe acute respiratory syndrome coronavirus-2 (sars-cov-2), was identified in late 2019 in wuhan, china. it has rapidly spread around the world, and was named a pandemic by the who on march 11, 2020. 1,2 in iran, the first covid-19 case was diagnosed on february 19, 2020; unfortunately, the ministry of health reports increasing numbers of cases and deaths from covid-19. 3 interestingly, covid-19 seems to be strangely and tragically selective. only some infected people become sick. although most critically ill covid-19 patients are either elderly or have underlying medical problems such as cardiovascular disease, hypertension, diabetes mellitus, or cancer, some previously healthy and even relatively young individuals have died from covid-19. sequencing of patients' genomes for dna variations associated with severe illness may help to explain this mystery. 4 herein, we present the cases of three brothers, all of whom died from covid-19 with a relatively similar pattern after less than 2 weeks of illness. the brothers, aged 54-66 years, lived in different locations in tehran, iran, and were previously healthy, without histories of underlying diseases, including hypertension, diabetes mellitus, cardiac or hepatic disease, or malignancy. patient 1, aged 60 years, worked in a travel agency. he referred to an outpatient clinic with fever, without any indication for admission. a dry cough started on the following day, followed by dyspnea 1 day later. he was then hospitalized with progression of dyspnea and suspicion for covid-19. he was admitted to the intensive care unit (icu) and died after 3 days with respiratory failure. a sars-cov-2 reverse transcriptase-polymerase chain reaction (pcr) test was positive; his spouse tested negative. patient 2, aged 54 years, worked in the same travel agency as his brother, case 1, with close contact with his brother beginning a week before the onset of illness. he started with fever 2 days after his brother. he was admitted promptly to another hospital. he developed cough and dyspnea 1 day later. the national treatment protocol for covid-19 (hydroxychloroquine plus oseltamivir) was started. his condition improved by the fifth day of admission, and he was discharged from the hospital with o 2 saturation of 87%. at home, despite good care, his condition deteriorated, and he died on the ninth day of illness. a sars-cov-2 reverse transcriptase-pcr test was negative, but the diagnosis of covid-19 was highly suspected. sars-cov-2 tests for his spouse and their two daughters were negative. patient 3, aged 66 years, had only short contact for less than an hour with case 1 on his second day of illness. patient 3 developed fever and cough 1 week later. he was admitted to a hospital with suspicion of covid-19. he progressed to severe dyspnea 1 day later and, subsequently, was transferred to the icu. he died on the next day with acute respiratory distress syndrome. a sars-cov-2 test was positive; tests were negative for his spouse and their two children. the results of laboratory tests for the patients are presented in table 1 . chest computed tomography scans are in figure 1 . there is some evidence for a role for the immune response and inflammation in the pathogenesis of covid-19. 5 an increased susceptibility to specific pathogens has been reported for a number of genetic defects. 6 there have been reports of deaths from covid-19 in multiple members of a family; for example, four family members died of covid-19 in wuhan, china, the epicenter of the outbreak, after they went into self-quarantine. 7 considering the fact that most of them who are affected by covid-19 recover, death in three brothers who lived separately is of interest. of note, the patients' spouses and children were not shown to be affected by covid-19, with negative tests for sars-cov-2. therefore, the three brothers who experienced fatal covid-19 may have had a particular predisposition to infection or severe illness. it can be hypothesized that there might be a genetic predisposition to covid-19 and/or to severe illness from covid-19 that renders some individuals without any underlying disease at particular risk of severe or fatal illness. who director-general's opening remarks at the media briefing on covid-19-11 iranian ministry of health and medical education covid-19 how sick will the coronavirus make you? the answer may be in your genes immune-epidemiological parameters of the novel coronavirus -a perspective primary immunodeficiency diseases: definition, diagnosis, and management four deaths in one family show danger of wuhan's home quarantine policy all scans show diffuse ground glass opacities (ggos), with peripheral distribution, consistent with novel coronavirus disease (covid-19) lung involvement. patients 1 and 2: crazy paving appearance, suggestive of late covid-19. patient 3: diffuse widespread ggos and bilateral diffuse opacification due to acute respiratory distress syndrome key: cord-345554-x9q6pqgw authors: kay, alexander w.; ness, tara e.; martinez, leonardo; mandalakas, anna m. title: it ain’t over till it's over: the triple threat of covid-19, tb, and hiv date: 2020-08-31 journal: am j trop med hyg doi: 10.4269/ajtmh.20-1089 sha: doc_id: 345554 cord_uid: x9q6pqgw it ain't over till it's over: the triple threat of covid-19, tb, and hiv. as the sars-cov-2 outbreak rapidly spread to pandemic proportions, the global tuberculosis (tb) and hiv care communities voiced increasing concerns. can we contain the inevitable disruption of routine tb and hiv services and the resultant detrimental impacts to patient care and disease control? tuberculosis alone kills 4,000 people each day and 1.5 million people each year. the impact of tb is greatest in settings with high burdens of both tb and hiv, where a third of people living with hiv die from tb. in 2018, an estimated 10 million people developed tb, nearly half a million drugresistant tb. although an estimated 58 million lives were saved through tb diagnosis and treatment between 2000 and 2018, 1 the current sars-cov-2 pandemic threatens to reverse these gains by overwhelming the healthcare system, disrupting patient access to care, and spurring reallocation of resources from tb and hiv services. furthermore, in already stressed settings with high burdens of tb and hiv infection, the risk and impact of triple infection with tb, hiv, and sars-cov-2 will likely be greatest. in this issue of the ajtmh, two articles describe four patients with triple infection with drug-sensitive mycobacterium tuberculosis, hiv, and sars-cov-2. 2,3 farais et al. describe two adult male patients in brazil, both with poor antiretroviral treatment (art) adherence and viremia. one individual had known active tb, but did not complete treatment, and the other had tb diagnosed upon presentation with covid-19. although these patients ultimately recovered, the authors note the challenge of delaying arts because of tb treatment. rivas et al. describe another two adult males, both with uncontrolled viremia from hiv with low cd4 counts. one had tb diagnosed during hospitalization, and the other was admitted during the continuation phase of tb treatment. whereas art initiation was delayed because of tb treatment, management of covid infection did not alter the overall treatment course for tb or hiv; however, none of the patients were treated with remdesivir, a drug increasingly used in the care of patients with severe sars-cov-2, which has a significant interaction with the tb drug rifampicin. the authors of these case reports effectively describe the preliminary and often conflicting available data on the association between covid-19 and clinical outcomes in patients with hiv and tb coinfection. the diversity of conclusions drawn in the literature to date, from those suggesting that hiv may have a protective effect, 2,3 to a negligible effect, 4-6 and to a deleterious effect, 7 may reflect the heterogeneity of the global hiv epidemic itself. emerging data from south africa, a country with a generalized hiv epidemic, suggest that hiv is associated with an increased risk of covid-19 mortality. even in this setting, however, the impact of hiv on covid-19 mortality is dwarfed by the impacts of noncommunicable diseases such as diabetes mellitus and hypertension, which are increased among people living with hiv. 7 similarly, the same risk factors that predispose individuals to tb, such as advanced age, kidney disease, diabetes, and male gender, also confer risk for severe covid-19 disease. 8 a south african cohort demonstrated a 1.5-to 2-fold increased risk of covid-19 mortality associated with prior or current tb disease, respectively. however, it is difficult to separate the impact of covid-19 from the increased risk of tb itself, and other case series have not found a similar association. 9 these cases, and the limited literature to date, highlight the importance of better establishing the roles that hiv-and tb-specific risk factors play in covid-19 outcomes, either independently or together. although comorbidities associated with hiv and tb may primarily drive covid-19 disease severity in these populations, the disruption to health systems caused by the covid-19 pandemic is likely to have a greater impact on the lives of people with hiv infection and tb than is coinfection with covid-19. 10 preliminary data from south korea, china, and nigeria, all locations that have seen substantial reductions in tb over the past decade, suggest that the covid-19 pandemic has severely disrupted tb control services. in south korea, tb notifications decreased by 24% in 2020 compared with prior years. 11 similarly, in nigeria, there were 35% and 34% decreases in the number of presumptive and active tb notifications from january to may 2020, respectively. 12 mathematical models predict that by 2024, deaths due to hiv and tb will increase by 10% and 20%, respectively, compared with levels predicted without a covid-19 pandemic. 13 there is, however, a potential opportunity to harness efforts to control covid-19 in a way that also mitigates impacts on tb and hiv control. 14 this may include coordinating contact-tracing efforts, which are critical to controlling outbreaks of both covid-19 and tb. importantly, novel technology that has been developed for covid-19 contact tracing could be an important tool to increase efficiency of contact tracing of tb patients. 15, 16 in addition, community health workers newly designated to screen for covid-19 may provide a workforce with experience and training compatible with hiv testing and tb screening services. the covid-19 pandemic has highlighted the need for all countries to invest in universal health coverage and to ensure equitable distribution of not only covid-19 treatment and vaccines but also tb and hiv infection treatments. covid-19 has already depleted and disrupted the global supply chain for all drugs, including tb and hiv medicines, and personal protective equipment (e.g., n95 respirators) used in a variety of illness settings. focusing solely on covid-19, especially in these low-resource settings, will damage the very fragile gains we have made in tb and hiv control. an approach that does not consider tb and hiv control will inevitably be less * address correspondence to anna m. mandalakas, baylor college of medicine and texas children's hospital, 1102 bates st., houston, tx 77030-3411. e-mail: anna.mandalakas@bcm.edu effective, and will stand as a missed opportunity to rise to the challenge of ending tb and hiv infection. world health organization why aren't people living with hiv at higher risk for developing severe coronavirus disease 2019 (covid-19)? could hiv infection alter the clinical course of sars-cov-2 infection? when less is better clinical characteristics and outcomes in people living with hiv hospitalized for covid-19 clinical features and outcomes of hiv patients with coronavirus disease 2019 covid-19 id team, 2020. description of covid-19 in hiv-infected individuals: a single-centre, prospective cohort hiv and risk of covid-19 death: a population cohort study from the western cape province factors associated with covid-19-related death using opensafely tuberculosis, covid-19 and migrants: preliminary analysis of deaths occurring in 69 patients from two cohorts tackling two pandemics: a plea on world tuberculosis day effect of covid-19 on tuberculosis notification impact of covid-19 on tb care: experiences of a treatment centre in nigeria potential impact of the covid-19 pandemic on hiv, tuberculosis, and malaria in low-income and middleincome countries: a modelling study how covid-19 could benefit tuberculosis and hiv services in south africa child contact management in high tuberculosis burden countries: a mixed-methods systematic review digital tools for covid-19 contact tracing key: cord-342790-70vn1nrm authors: rosenthal, philip j.; bausch, daniel g.; goraleski, karen a.; hill, david r.; jacobson, julie a.; john, chandy c.; breman, joel g. title: keep politics out of funding decisions for medical research and public health date: 2020-07-22 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0850 sha: doc_id: 342790 cord_uid: 70vn1nrm keep politics out of funding decisions for medical research and public health. authors are members of the american society of tropical medicine and hygiene's executive committee the american society of tropical medicine and hygiene is the largest international scientific organization of experts dedicated to reducing the worldwide burden of tropical infectious diseases and improving global health. as leaders of the society, we are compelled to speak up for the integrity of science, and specifically for research funding and public health decisions based on merit, with policies rooted in data. recently, key decisions have lacked this basis, and science is under attack. on april 24, 2020, the nih abruptly canceled funding for an ongoing program studying coronaviruses that had been funded since 2014. this decision was highly unusual for several reasons. first, the nih almost never cancels grants after funding is approved based on rigorous peer review, unless there is financial or scientific misconduct. second, it is remarkable that, of all grants to cancel in the midst of the covid-19 pandemic, it is a grant to study coronaviruses. third, it has been substantiated that the decision to cancel the grant came not from the nih but from the highest levels of the u.s. government. there is little doubt that a productive research program, indeed a program directly relevant to our attempts to control the greatest respiratory virus pandemic of the last century, was canceled because of political pressure. canceling a major coronavirus research program at exactly the time when more, not less, research in this area is needed is deeply disturbing. the research program that was canceled was led by the ecohealth alliance, a u.s.-based global nonprofit organization that conducts research on five continents in biosurveillance, deforestation, wildlife conservation, and, most relevant to this conversation, zoonotic diseases and pandemic prevention. this research, which included collaboration with other international institutions, focused on coronavirus spread from bats to other species, including humans. the research results contributed to studies to prevent the spread of and to develop drugs and vaccines against sars-cov-2, the cause of covid-19. 1 ecohealth alliance research has helped us to understand how coronaviruses jumped from bats to humans to cause the current pandemic; the canceling of this program without scientific basis seems a flagrant example of "shooting the messenger." the reason for cutting the coronavirus research project is quite transparent. the trump administration reportedly learned about the project, including its links to the wuhan institute of virology, from a reporter on april 17. soon thereafter, it was announced that the program would be cut. the department of health and human services justified the cut with the claim that "the grantee was not in compliance with nih's grant policy," but the director of the nih institute that funded the study later stated in congressional testimony that "it was canceled because the nih was told to cancel it." clearly, the actual reason for cancellation was political, apparently based on an attempt to assign blame for the covid-19 pandemic to china, and consistent with claims that the pandemic was the result of a laboratory accident or a deliberate attempt to initiate a viral outbreak in wuhan. these claims are unsubstantiated and readily refuted by available molecular data. 1 on another front, on may 29, the white house announced that the united states will terminate its relationship with the who; this was confirmed on july 6 when the united states sent an official withdrawal letter to the united nations. the who was founded in 1948 to coordinate international health policy, establish global disease surveillance networks, and lead disease control and eradication programs. it is supported by and under the control of its 194 member states. although burdened through its history by limited control over international health policy and bureaucratic constraints, overall, the who has been remarkably successful and is an essential global institution. the who played a pivotal role in the eradication of smallpox, it has led successful efforts to control and eliminate many other infectious diseases, and it is the key international agency to lead efforts to address international outbreak and epidemic diseases, from influenza to hiv/aids, multidrug-resistant tuberculosis, polio, onchocerciasis, guinea worm, ebola, zika, and now covid-19. the reason for cutting ties with the who seems as clear as that for the nih grant cut described previously. with a failing response to the covid-19 pandemic at home and a pending election, the u.s. administration acted to deflect criticism to the who. although initial international responses to the emerging pandemic may have been slower than optimal, the who played a major role in rapid dissemination of information regarding virus isolation and characterization within a few weeks of recognition of the outbreak, oversaw the rapid development of reliable tests for the infection, and has regularly written and updated evidence-based guidance for control of the pandemic. who coordination and strong national programs and leadership are helping to curb covid-19 in many countries. as with cutting funding for coronavirus research during the pandemic, cutting funding for the who is profoundly troubling. at this time of deep human tragedy and economic disaster due to the covid-19 pandemic and of political upheaval in many countries, the headlines change quickly, and politically motivated mandates of great importance can fail to generate the attention that they deserve. we have therefore highlighted two recent decisions by the white house that are profoundly misguided. indeed, they seem the worst possible choices as we grapple with an overwhelming pandemic. but these choices can be reversed. we urge president trump to 1) reinstate and work with congress to increase funding for essential research on coronaviruses to help us develop new tools and strategies to address current and future pandemics, and 2) continue the country's long-standing relationship with the who, offering full financial support and cooperation for this essential international body. more broadly, we implore our elected officials at every level of government to keep politics out of decisions regarding medical research and public health. the health of americans and of the population of our planet must not be a bargaining chip used to seek political gains or deflect accountability. rather, health is an indisputable human right. as we fight the worst respiratory pandemic of the last century, it is foolhardy to limit support for the best research and the best public health institutions that are working to stem the pandemic. the origin of covid-19 and why it matters key: cord-345067-kummh0g7 authors: nachega, jean b.; leisegang, rory; kallay, oscar; mills, edward j.; zumla, alimuddin; lester, richard t. title: mobile health technology for enhancing the covid-19 response in africa: a potential game changer? date: 2020-05-29 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0506 sha: doc_id: 345067 cord_uid: kummh0g7 mobile health technology for enhancing the covid-19 response in africa: a potential game changer? the who africa region is experiencing an increase in the number of novel covid-19 cases. as of may 20, 2020, 63,521 cases with 1,796 deaths (2.8% case fatality) have been reported from 45 countries. 1 although these numbers are small compared with those in united states or europe, the who recently estimated that up to 190,000 people could die of covid-19 in africa if the pandemic is not controlled. 2 these projections are threatening the already overstretched health services in africa, where governments have been implementing mitigating strategies to flatten epidemic curves at manageable levels. these include education, personal hygiene practices, social distancing, travel bans, and partial or total lockdowns. 3 however, as lockdowns and social distancing measures are currently being lifted in stages by most african countries, governments will need to ensure that public health infrastructure and needed resources are put in place for community surveillance to identify cases and clusters of new infections through active case finding, large-scale testing, and contact tracing. cost-efficient testing strategies with rapid turnaround and community-based contact-tracing approaches are cornerstones for containment during epidemics. to do so at scale and over the anticipated prolonged course of this pandemic, african countries will need to capitalize on digital health innovations. [4] [5] [6] the global system for mobile communication association reports that 50% of africans own mobile phones and that 39% are internet-connected, numbers which are rapidly increasing, and approach 80% access when phone-sharing is considered. 7 mobile phone technology (mhealth) platforms are effective in improving service delivery and outcomes for many health conditions in africa and globally, including hiv infection, tuberculosis, and chronic noncommunicable diseases. [4] [5] [6] [7] in the context of covid-19, mhealth solutions offer opportunities to directly support public education, case management, and contact tracing, and to perhaps even provide geolocation and exposure notification. 7, 8 with the support of global mobile technology companies and small and medium enterprises within africa, mhealth offers opportunities ranging from text messaging to mobile apps to mitigate the spread of covid-19. the use of mobile phones reduces the need for physical contact, exchange of materials, and movement by health workers, and thus maximizes safety. several ongoing digital and mobile initiatives related to covid-19 have been identified across africa ( figure 1 ). district health information software 2 is an open-source, web-based health management information system platform already used by 67 low-and middle-income countries. district health information software 2 has a covid-19-specific application package that several african countries are using for field data collection. 9 in rwanda and uganda, the weltel virtual care system serves as a real-time remote monitoring platform. covid-19 cases and contacts in home isolation receive semi-automated daily text message check-ins via sms for 2 weeks using an open language format, allowing self-reporting of new symptoms or issues. responses are viewed by health officials on a dashboard, and patients are triaged much faster than would be the case with traditional field outreach or telephone calls, saving critical human resource capacity. novel natural language processing computing tools promise to reveal insights into the issues that patients face during home quarantine. the provision of monitoring packaged with interactive support helps people undertake home isolation/quarantine most effectively. 10 in ghana, a short ussd code (*920*222#) dialed on mobile phones allows residents to respond electronically to questions about their symptoms, who they have been in contact with, and their travel history. the opine health assistant compiles the results into maps and graphs to make it easier to understand, monitor, and share. 11 in senegal, sms services are used to broadcast good hygiene practices to rural communities to disrupt the spread of covid-19. 12 in south africa, community screening, referral for testing, and communication of results of using an mhealth platform are being rapidly expanded to more than 28,000 trained community health workers. 13 mobile phones and apps also support livelihoods and enable remote access to critical services such as education and food. in kenya, transaction fees for using m-pesa, a cashless, mobile money platform with 20 million users, have been waived to provide a safe method by which to transfer funds within community settings. in south africa, mobile data costs of accessing some teaching and learning websites have been waived by major cellphone providers to ensure that primary and secondary school and university students can continue to access learning materials. globally, mobile counseling, support hotline, and social media platforms are assisting with public health information as well as mental health counseling, food relief, domestic violence concerns, and other support. government and private alignment within these platforms should be encouraged, as oversight by public health agencies will ensure accurate content. in conclusion, there appears to be a limited window of opportunity in which to contain the spread of covid-19 in africa and keep economies afloat. there is a significant body of innovation and evidence to inform mhealth best practices that have emerged from africa over the past decade. 14-16 mhealth may be a game changer if it is introduced swiftly and widely in this pandemic. to succeed, barriers to access to and use of mobile phones and the latest technologies need to be defined, and there must be cooperation among all stakeholders to enable rapid deployment and scale-up of promising or evidencebased solutions. if mhealth is rigorously implemented, scaledup, and evaluated through implementation science, then africa will reap the benefits of this technology for the remainder of the covid-19 crisis and be better positioned for future pandemics and for improving all aspects of public health. publication charges for this article were waived due to the ongoing pandemic of covid-19. coronavirus disease (covid-2019) situation reports new who estimates: up to 190000 people could die of covid-19 in africa if not controlled the late arrival of covid-19 in africa -mitigating pan-continental spread effects of a mobile phone short message service on antiretroviral treatment adherence in kenya (weltel kenya1): a randomised trial sub-saharan africa-the new breeding ground for global digital health africa: opportunities for growth mobile phone penetration through sub-saharan africa covid-19 surveillance digital data package get virtual care for your covid-19 response institute of ict professionals ghana. 2020. how ussd code is breaking grounds in ghana -part covid-19: our response in senegal south africa is hunting down coronavirus with thousands of health workers mobile phone-delivered reminders and incentives to improve childhood immunisation coverage and timeliness in kenya (m-simu): a cluster randomised controlled trial a hybrid mobile approach for populationwide hiv testing in rural east africa: an observational study controlling ebola through mhealth strategies covid-19 and mobile health technology in africa key: cord-314921-u74jut63 authors: levine, shankar; dhakal, guru prasad; penjor, tshering; chuki, pem; namgyal, kesang; tshokey,; watts, melanie title: case report: the first case of covid-19 in bhutan date: 2020-04-20 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0259 sha: doc_id: 314921 cord_uid: u74jut63 the initial cases of novel coronavirus disease-19 (covid-19) in a country are of utmost importance given their impact on healthcare providers, the country’s preparedness response, and the initial molding of the public perception toward this pandemic. in bhutan, the index case was a 76-year-old immunocompromised man who had traveled from the united states and entered bhutan as a tourist. he presented initially with vague gastrointerestinal symptoms and later a cough. his atypical presentation led to a delay in diagnosis, but ultimately he was isolated and tested. on confirming the diagnosis of covid-19, the patient was isolated in a separate hospital with a dedicated medical care team. all contacts were traced and quarantined. the patient’s respiratory status deteriorated despite broad-spectrum antivirals, antibiotics, and intensive supportive care. he required intubation and was given a trial of intravenous immunoglobulin to modulate his likely aberrant immune response. subsequently, the patient’s clinical status improved, and after 8 days of hospitalization, he was transferred out of the country, where he recovered. this was a learning experience for the treating medical staff, the government, and the people of bhutan. as novel coronavirus disease (covid-19) rapidly spreads across the globe; the initial cases in a country and their outcomes may have a tremendous impact. in a small, relatively isolated country such as bhutan, this impact may be felt not only by the patients and their families but also by the country's population, as the case and the country's response are keenly followed. this report describes the first patient diagnosed and treated for covid-19 in bhutan. the index case was a 76-year-old american man, with a history of hypertension, hyperlipidemia, and neuropathy for which he was on medications; surgical history was notable for a splenectomy due to mantle cell lymphoma. the patient traveled from the united states through airports in london, mumbai, and kolkata, before arriving in jorhat, assam, india. halfway through a week-long cruise on the brahmaputra river in assam, he started feeling ill. following the cruise, he flew to bhutan, and over the first 2 days in country, he sought medical care for intermittent symptoms including bloating, loss of appetite, diarrhea, and fatigue. on his fourth day in bhutan, he presented to the emergency department in thimphu, where he was noted to be afebrile but hypoxic, with an oxygen saturation of 78% on room air, and he endorsed 4 days of cough. based on preparedness screening protocols, the patient was clinically screened for covid-19, and the decision was made to isolate and test the patient. initial chest x-ray revealed mild bilateral patchy infiltrates, and he was started on oseltamivir, ceftriaxone, and doxycycline, and was kept in respiratory isolation on supplemental oxygen. the patient's swab reverse transcriptase-polymerase chain reaction test for covid-19 was confirmed positive at midnight, and by morning, the ministry of health had traced approximately 90 contacts from the healthcare sector and hotel and restaurant staff; based on their degree of exposure, the patient contacts were instructed to either home quarantine or present to a quarantine facility. on royal command, a recently constructed ophthalmology hospital building was rapidly converted into a covid-19 treatment facility including separate equipment, medications, and staffing. a dedicated care team was assembled comprising an emergency critical care physician, two internal medicine physicians, and nurses. over the first days of hospitalization, the patient's oxygen requirement gradually worsened; his white blood cell (wbc) count and c-reactive protein (crp) continued to increase. point-of-care ultrasound revealed worsening b lines ( figure 1 ). despite the addition of lopinavir and ritonavir and switching antibiotics to meropenem and vancomycin, he developed and maintained a low-grade fever. computed tomography of the chest revealed diffuse ground-glass opacities consistent with acute respiratory distress syndrome (ards) (figure 2 ). on the fifth day after diagnosis, the patient's oxygen requirement and work of breathing dramatically worsened, and he was intubated and placed on a ventilator for respiratory support. having reviewed the data of intravenous immunoglobulin (ivig) in the setting of ards, 1,2 a decision was made to give a 3-day course of 0.5 g/kg (40 g daily). the dose was largely based on the limited quantity of available ivig. the patient received his first dose of ivig on the sixth day after diagnosis and underwent prone positioning for ards. 3 by the next morning, his oxygen requirement had improved, and 48 hours after the first dose of ivig, the patient's wbc count and crp had decreased (figure 3) . after 8 days of treatment and significant clinical improvement in bhutan, the patient was evacuated to his home country. his clinical status continued to improve, and he was extubated 7 days after evacuation. fifteen days after his initial positive covid-19 test and hospitalization, he no longer required oxygen therapy and rehabilitative therapy was initiated. as the covid-19 pandemic continues to spread to new countries, some aspects pertaining to the patient's presentation, interventions to provide patient care, and the country's measures of containment in response to this first case in bhutan may be of broad interest. the patient's initial presenting symptoms were gastrointestinal; he subsequently developed a cough. although early studies suggested that gastrointestinal symptoms were uncommon with covid-19, subsequent studies have suggested that such symptoms are more common, noted in 10% and 18.6% in two studies. [4] [5] [6] the patient's lack of appetite prompted him to seek medical care. in a recent study, 78.6% of covid-19 patients suffered from a lack of appetite at presentation. 6 the patient's presentation did not fall into the case definition for covid-19 that bhutan was using at that time, which was limited to fever and respiratory symptoms. this case is a reminder of atypical presentations and the need for allowing medical providers' clinical discretion in management and for regular updating of case definitions during a new disease outbreak. as the patient's clinical status worsened, it seemed possible that this may have been due to an aberrant immune response, which is described in patients with viral infections, leading to ards. 2, 7 prior case reports of ivig successfully treating ards secondary to viral illnesses have been documented. 1,2 after receiving ivig, the patient had dramatic improvement of oxygenation and a downward trend in his inflammatory markers. although the efficacy of ivig, timing, dosage, and patient selection all clearly require further study, this case suggests that a trial of ivig for the treatment of covid-19 is warranted. the public health strategies initiated by bhutan's ministry of health and government are beyond the scope of this case report, but the immediate steps pertaining to the patient's contacts and medical staff caring for the patient are of interest. some relevant lessons from singapore and hong kong, which were able to curtail the rapid spread of the virus fairly effectively, include the need for the following: 1) appropriate personal protective equipment for medical staff in contact with a suspected patient, 2) disinfecting surfaces, 3) treating patients in a separate ward or hospital with a separate team, and 4) contact tracing and quarantine. 8, 9 in singapore, contact tracing accounted for the primary detection of approximately half of the country's first 100 covid-19 patients. 8 despite the patient's atypical presentation, bhutan's preparedness played a role in limiting exposure and making the initial diagnosis. the triage nurse, trained to screen patients for recent travel, alerted the emergency physician, who then took the patient to a predesignated isolation area for suspected cases. after the patient tested positive, a decision was rapidly made to designate a separate hospital building as a dedicated covid-19 hospital. separate staff including doctors, nurses, ambulance drivers, and cleaners were designated for this facility. these staff lived in a separate facility for the duration of treating the patient, after which they were put in a 14-day quarantine and monitored for symptoms. all of the patient's contacts were traced and quarantined. those with exposures deemed high risk were tested initially and at the end of their 14 days of quarantine. although this is not common practice globally, it led to the diagnosis of covid-19 infection in the patient's partner, who was asymptomatic. no other initial contacts or medical staff tested positive by the end of their 14day quarantine. to prevent the import of cases and subsequent local transmission from occurring, bhutan rapidly initiated travel restrictions barring the entry of all nonnationals and instituted a mandatory quarantine in designated hotels for all persons entering bhutan. to date, there have been five cases of covid-19; the three additional cases were diagnosed among repatriated quarantined bhutanese students who were studying abroad. there has been no evidence of local community spread. mails: sailevine@gmail.com, gurudhakal@hotmail.com, tsheringp@ jdwnrh.gov.bt, pchuki@jdwnrh.gov.bt, dkesangnam@gmail.com, tshokey@ jdwnrh a case of acute respiratory distress syndrome associated with novel h1n1 treated with intravenous immunoglobulin pneumonia, acute respiratory distress syndrome, and early immune-modulator therapy prone positioning in severe acute respiratory distress syndrome epidemiological and clinical characteristics of 99 cases of 2019 novel coronavirus pneumonia in wuhan, china: a descriptive study clinical characteristics of 138 hospitalized patients with 2019 novel coronavirus-infected pneumonia in wuhan, china. jama e201585 clinical characteristics of covid-19 patients with digestive symptoms in hubei, china: a descriptive, cross-sectional, multicenter study immune responses in covid-19 and potential vaccines: lessons learned from sars and mers epidemic evaluation of the effectiveness of surveillance and containment measures for the first 100 patients with covid-19 in singapore keeping the coronavirus from infecting health-care workers. the new yorker graph demonstrating white blood cell and c-reactive protein in relation to days of illness and timing of treatments given acknowledgments: we would like to acknowledge the leadership and rapid implementation of necessary public health measures by lyonpo dechen wangmo, minister of health. publication charges for this article were waived due to the ongoing pandemic of covid-19. this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-312861-6ayidmj3 authors: rayner, craig r.; dron, louis; park, jay j. h.; decloedt, eric h.; cotton, mark f.; niranjan, vis; smith, patrick f.; dodds, michael g.; brown, fran; reis, gilmar; wesche, david; mills, edward j. title: accelerating clinical evaluation of repurposed combination therapies for covid-19 date: 2020-08-21 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0995 sha: doc_id: 312861 cord_uid: 6ayidmj3 as the global covid-19 pandemic continues, unabated and clinical trials demonstrate limited effective pharmaceutical interventions, there is a pressing need to accelerate treatment evaluations. among options for accelerated development is the evaluation of drug combinations in the absence of prior monotherapy data. this approach is appealing for a number of reasons. first, combining two or more drugs with related or complementary therapeutic effects permits a multipronged approach addressing the variable pathways of the disease. second, if an individual component of a combination offers a therapeutic effect, then in the absence of antagonism, a trial of combination therapy should still detect individual efficacy. third, this strategy is time saving. rather than taking a stepwise approach to evaluating monotherapies, this strategy begins with testing all relevant therapeutic options. finally, given the severity of the current pandemic and the absence of treatment options, the likelihood of detecting a treatment effect with combination therapy maintains scientific enthusiasm for evaluating repurposed treatments. antiviral combination selection can be facilitated by insights regarding sars-cov-2 pathophysiology and cell cycle dynamics, supported by infectious disease and clinical pharmacology expert advice. we describe a clinical evaluation strategy using adaptive combination platform trials to rapidly test combination therapies to treat covid-19. abstract. as the global covid-19 pandemic continues, unabated and clinical trials demonstrate limited effective pharmaceutical interventions, there is a pressing need to accelerate treatment evaluations. among options for accelerated development is the evaluation of drug combinations in the absence of prior monotherapy data. this approach is appealing for a number of reasons. first, combining two or more drugs with related or complementary therapeutic effects permits a multipronged approach addressing the variable pathways of the disease. second, if an individual component of a combination offers a therapeutic effect, then in the absence of antagonism, a trial of combination therapy should still detect individual efficacy. third, this strategy is time saving. rather than taking a stepwise approach to evaluating monotherapies, this strategy begins with testing all relevant therapeutic options. finally, given the severity of the current pandemic and the absence of treatment options, the likelihood of detecting a treatment effect with combination therapy maintains scientific enthusiasm for evaluating repurposed treatments. antiviral combination selection can be facilitated by insights regarding sars-cov-2 pathophysiology and cell cycle dynamics, supported by infectious disease and clinical pharmacology expert advice. we describe a clinical evaluation strategy using adaptive combination platform trials to rapidly test combination therapies to treat covid-19. for novel covid-19, there has been extensive focus on repurposing previously approved drugs against sars-cov-2. 1 repurposing is attractive, as it allows the use of existing information on human pharmacology and clinical safety to enable faster clinical trial development and rollout. 2 in addition, repurposing drugs offers the potential to rapidly and efficiently scale effective treatments, in contrast to newer therapies that require upscaling drug manufacturing and supply chain pathways. to illustrate, a recent large-scale compound repurposing effort identified more than 20 antivirals that should be further investigated for application for covid-19. 3 antiviral agents for covid-19 can be categorized by two broad mechanisms of action: 1) those targeting viral proteins or nucleic acids related to infection of host cells, viral production via hijacking cellular machinery, or release from host cells and circulation of virions; and 2) those targeting essential host functions for viral replication including agents such as interferon that boost the cellular immune response to infection. each category is complex, and of the armament of potential repurposed drugs for covid-19, multitudes of mechanisms and pathways are in consideration. selectively combining agents to complement each other by variable mechanisms of action across these categories may yield effective treatments for select phases of sars-cov-2 infection. analogous to the evolution of antiretroviral therapies for hiv infection, it is important to formulate and test combination therapy regimens. instead of sequentially testing monotherapies that will likely have modest clinical effects on their own, additive or synergistic effects can potentially be gained by combining antiviral drugs exploiting pharmacology throughout the spectrum of covid-19 illness. 4 there are more than 750 clinical trials evaluating repurposed therapies for covid-19. 1 these trials cover all antiviral mechanisms noted earlier and applications from prophylaxis to treatment to reduction in the sequelae of the host inflammatory response. however, to date, only dexamethasone and remdesivir have been identified as effective therapies for severely ill patients. 5, 6 this low success rate might be due to the fact that the majority of covid-19 clinical trials (87%) are evaluating repurposed drugs as monotherapy. 1 combination therapies for covid-19 represent an attractive approach to drug development. among the first trials published of effective interventions for covid-19 was a combination of interferon beta-1b, lopinavir-ritonavir (lpv/r), and ribavirin for hospitalized patients. 7 in comparison to the control group that received lpv/r monotherapy, the triple combination arm showed faster viral clearance and alleviation of symptoms, and shorter hospital stays. 7 although lpv/r monotherapy was no better than placebo alone in another hospitalized trial (recovery), 8 it is unclear whether both interferon beta-1b and ribavirin or triple combination therapy drove clinical benefits for hospitalized patients. under normal circumstances, development of combination treatment strategies entails a stepwise evaluation process whereby first the individual components of a potential combination regimen are tested for clinical efficacy in isolation or as individual arms within a trial evaluating both single and combination regimens. this strategy allows sequential compilation of evidence for each drug before studying combinations, as outlined by the u.s. food and drug administration in describing the so-called combination rule. 9 this process is based on reducing exposure to ineffective or toxic drugs among participants until individual components have demonstrated treatment effects. although this process is both scientifically rigorous and safe, it is slow, and may delay identification of unexpected synergistic effects, resulting in failure to evaluate potentially potent treatment cocktails. as the global pandemic continues, unabated and therapeutic options evaluated to date demonstrate limited effectiveness, and there is a pressing need to accelerate treatment evaluations. among the options for accelerated development is the evaluation of combination strategies in the absence of prior monotherapy data. this approach is appealing for a number of reasons. first, combining two or more drugs with complementary antiviral or therapeutic effects permits a multipronged approach addressing the variable pathways of the disease. second, if an individual component of a combination strategy offers a therapeutic effect, then the clinical trial should still detect treatment effects unless antagonism between components is present. third, this strategy saves time, as it accelerates evaluation of prioritized combination regimens much earlier than would occur with a stepwise approach evaluating monotherapies before combinations. finally, given the severity of the current pandemic, and the current absence of verified treatment options, the likelihood of detecting a treatment effect with combination therapy maintains scientific and patient enthusiasm for repurposed treatments. despite these advantages, there has been limited research activity investigating combination therapy for covid-19, in clear contrast to other disease areas. among the 1971 registered clinical trials evaluating all therapeutic interventions (i.e., not only previously approved treatments) for covid-19, 1 only 258 (13.1%) trials are evaluating combination therapies. of these, 120 trials are taking a stepwise approach, and 138 trials are evaluating combination strategies without associated individual monotherapies. determining whether these trials are implementing a forward (evaluating monotherapy before combination therapy) or backward stepwise approach is not possible from clinical trial registries. among the many scientific lessons of the covid-19 pandemic has been the widespread embrace of adaptive clinical trials, particularly platform trials in which multiple interventions are compared simultaneously against a common control arm, and in which interventions may be added over time. 10, 11 figure 1. a platform trial for combination therapy. here, in this example, there are several interim analyses planned for the platform trial testing combination therapies using a backward stepwise approach. at the first interim analysis, combination arm 1 is dropped for futility followed by combination arm 2 dropped at the second interim analysis. at the third interim analysis, combination arm 3 shows superiority over placebo (pbo), and, thereafter, individual monotherapies are added and evaluated after. although this concept is not new, it has historically been controversial and limited to industry-run clinical trials, especially in oncology. in adaptive clinical trial designs, prespecified modifications are permitted with decision rules based on accumulated interim data. 12 adaptive designs have been controversial because interim data are often assessed multiple times, leading to a fear of inflated type i error rates. however, statistical measures can be implemented to control the type i error rate at the usual 5%. important examples of adaptive platform trials directed toward covid-19 include the recovery, 5 solidarity, 13 and remap-cap 14 trials. adaptive platform trials provide capacity in combination trials to add or remove ineffective treatment arms, allowing for sequential comparisons and a perpetual design wherein treatments can be prioritized or de-prioritized as evidence accumulates. we propose a platform trial to evaluate combination therapies in the absence of monotherapy evaluations (figure 1 ). if the combination is ineffective, then there is reduced incentive to evaluate the monotherapies. if the combination demonstrates efficacy, then monotherapy arms can be studied to determine which components of the combination therapy are efficacious. our proposed approach is not without limitations. evaluating combinations assumes that individual components are nonantagonistic and likely either additive or synergistic. there are important examples of drug therapies that exhibit antagonism. 15, 16 thus, combination strategies may miss efficacy of individual components undermined by antagonism. furthermore, drug interactions may result in adverse events and toxicity not observed with monotherapy. therefore, it is critical to engage clinical pharmacologists and infectious disease experts in guiding the selection and refinement of regimens to ensure that insights on sars-cov-2 pathophysiology, cell cycle dynamics, mechanisms of action, drug-drug interactions, safety signals, and clinical utility are appropriately incorporated in the selection of combinations for study. ultimately, we argue that the best approach to quickly establish efficacy of potential therapies for covid-19 is through studying combination therapies, ideally with demonstrated in vitro and in vivo activities and strong preclinical properties, in an adaptive framework to maximize the probability of clinical efficacy. this strategy challenges the traditional drug development dogma, but we believe that it will accelerate the development of repurposed drugs as combination therapies to treat covid-19. a realtime dashboard of clinical trials for covid-19 rapid repurposing of drugs for covid-19 discovery of sars-cov-2 antiviral drugs through large-scale compound repurposing. nature (epub ahead of print modelinformed drug repurposing: viral kinetic modelling to prioritize rational drug combinations for covid-19 dexamethasone in hospitalized patients with covid-19-preliminary report remdesivir for the treatment of covid-19-preliminary report triple combination of interferon beta-1b, lopinavir-ritonavir, and ribavirin in the treatment of patients admitted to hospital with covid-19: an open-label, randomised, phase 2 trial no clinical benefit from use of lopinavir-ritonavir in hospitalised covid-19 patients studied in recovery guidance for industry: codevelopment of two or more new investigational drugs for use in combination adaptive platform trials: definition, design, conduct and reporting considerations an overview of platform trials with a checklist for clinical readers adaptive designs for clinical trials clinical trial for covid-19 treatments the randomized embedded multifactorial adaptive platform for community-acquired pneumonia (remap-cap) study: rationale and design dexamethasone inhibits paclitaxel-induced cytotoxic activity through retinoblastoma protein dephosphorylation in non-small cell lung cancer cells in vivo antagonism with zidovudine plus stavudine combination therapy this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-316712-1ngcwdln authors: laxminarayan, ramanan; jameel, shahid; sarkar, swarup title: india’s battle against covid-19: progress and challenges date: 2020-08-24 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0992 sha: doc_id: 316712 cord_uid: 1ngcwdln india's battle against covid-19: progress and challenges. india was fortunate not to be among the first countries hit by covid-19. the first reported case of infection with the sars-cov-2, the virus that causes covid-19, in india was reported on january 30, 2020 in an indian student evacuated from wuhan, and the first death was reported on march 12, 2020. although it is possible that the sars-cov-2 was circulating in india earlier, the first known secondary transmission of the virus within india was reported only in early march. even when the case load was low, india was predicted to be at high risk for covid-19 for a number of reasons. a dense population, especially in urban settings, could exacerbate the spread of sars-cov-2. the indian population has high rates of uncontrolled hypertension 1 and diabetes, 2 both of which have been shown to be risk factors for severe covid-19 and mortality. although children were found less likely to be infected in china 3 and other countries, it was unclear how children in india, who have high levels of stunting and early exposure to infections, would fare against sars-cov-2. 4 on the plus side, it was hypothesized that india's relatively young population-65% of india's population is younger than 35 years, and only 6% is older than 65 years-would be spared high mortality from a disease that targets the elderly. in comparison, 22% of the population of italy and 10% of that of china are older than 65 years. how did india respond and how has it fared? the indian government responded to covid-19 with temperature screening of incoming passengers on flights from east asia. large public events, including one to greet u.s. president donald trump, were held as late as february 24, although disease transmission was well underway in europe at that time. on march 24, a national lockdown was announced with 4 hours' notice. 5 although a state-level or more localized lockdown would have been preferable, the precise locations of disease hotspots were unknown because of low levels of testing. 6 the sudden lockdown imposed a significant burden on the urban poor and migrants, who found themselves both out of work and with no means to return to their villages. at the time of the lockdown, india had officially counted about 500 cases of covid-19 and 10 deaths because of the contagion. the lockdown represented a law-and-order solution to a problem for which india was poorly equipped from a public health standpoint. at that time, india had approximately 1.9 million hospital beds, 95,000 intensive care unit (icu) beds, and 48,000 ventilators, against a need of 270,000 icu beds under an optimistic scenario of covid-19 burden. 7 the lockdown was essential to buy time to prepare for the eventual flood of cases. model-based estimates 8 produced in march 2020 had indicated that a national lockdown could reduce the number of infections at the peak of the pandemic-expected in early may-by 70-80%, depending on the degree of public compliance with physical distancing. these projections estimated that, in the absence of any intervention, india could expect to see 5-20% of its urban population infected in a first wave. 8, 9 at an overall mortality rate of 1%, the death toll could climb to hundreds of thousands, if not higher. by all accounts, the national lockdown was tightly enforced and has been described as one of the harshest in the world. 10 since late june, the lockdown has been lifted in stages and has transitioned to state-level lockdowns that have been largely reactive to local caseloads at any given time. some form of restrictions on movement exists in most states. however, universal adherence to masking and social distancing has been difficult to enforce, and compliance has varied across states and districts. physical distancing is not practicable in many lowincome communities where there is significant crowding both within households and in public spaces. as of august, schools, colleges, movie theaters, places of worship, and most other places of mass gathering remain closed. the future outlook for reopening is unclear at this time. caseloads and deaths are now increasing at the fastest rate of any large country, and india currently records more than 800 covid-19 deaths every day (figures 1 and 2 ). as of august 3, 2020, india was third in the world in the number of reported sars-cov-2 infections (2,456,785) and fourth in the number of reported covid-19 deaths (48,117). 11 both infections and deaths are likely underestimated because of the low levels of testing in india. mortality rates (based on reported cases and deaths) appear to be low in india, as they are in most countries in the region, perhaps indicative of both limited testing and other unexplained factors. india was slow to provide testing despite significant capacity for reverse transcription-polymerase chain reaction (rt-pcr) testing in both public and private laboratories. testing in the early days of the epidemic was limited to a few public laboratories. private laboratories, which typically provide the bulk of pathology services, were not allowed to test at all. the restriction was not only ostensibly to maintain quality but presumably also to control information. testing for covid-19 in india continues to rank among the lowest in the world on a per-capita basis. testing rose from 3,000 tests per day on march 24, 2020 (approximately 2 per million population) to more than 700,000 per day (500 per million) in august (figure 3 ), although much of this increase was due to the introduction of rapid antigen tests that have far lower sensitivity than rt-pcr. 12 at the current time, india has conducted approximately 18,000 tests per million population, a rate that is a third that of south africa, about 60% that of nepal, and among the lowest of any large country. testing rates have been highly variable across states. daily testing in delhi, the capital city, was around 670 per million at the end of july, comparable to that in the united states, but rates elsewhere were much lower. as of august 4, the state of bihar was conducting just 20,000 tests per day, for a population of 105 million (190 per million). the low level of testing nationally has likely led to a large number of infections being missed. as a consequence, reported infections in india are likely indicative of only a small proportion of the total infections. according to a serological study based on 21,387 samples conducted by india's national cdc from june 27 to july 10, 23% of the population of delhi had been infected. 13 the number of seropositives-roughly 4.7 million-was approximately 40 times greater than the reported cumulative number of infections in delhi, indicating that a large number of infections were likely missed by the testing process. another unpublished seroprevalence study conducted in mumbai in early july indicated that 57% of those living in slums had antibodies to sars-cov-2, compared with 16% of those residing in other parts of the city. 14 unpublished data from the indian council of medical research indicated that 10 million people (0.7% of the indian population) had been infected through early may, at a time when fewer than 50,000 cumulative infections had been reported. 15 similar undercounts by a factor of 40-200 have been uncovered by comparing seroprevalence data with reported infection data. an important caveat is that the seroprevalence data have yet been published in the peer-reviewed literature. despite the size of the covid-19 problem, there has been limited published epidemiological or clinical research emerging from india. preliminary data from comprehensive surveillance in the indian states of tamil nadu and andhra pradesh (home to ∼128 million people) from a cohort of 4,206 index cases and 64,031 contacts found that the risk of transmission from an index case to a close contact ranged from 2.6% in the community to 9.0% in the household; these results did not differ with respect to the age of the index case. 16 this finding indicated an important role for children and young adults in transmission, with a third of infected individuals younger than 30 years. early analysis from this contact-tracing dataset, which represents one of the largest prospective studies of infections among exposed individuals to date, found that superspreading events were the rule, rather than the exception. prospective follow-up testing of exposures revealed that 83% of infected individuals did not infect any of their contacts, whereas 5% accounted for 80% of observed new infections. unlike in high-income countries, where deaths are mostly in the age-group older than 65 years, covid-19-related deaths were concentrated at ages 50-64 years, and the incidence of reported cases did not increase with older age. strikingly, these differences cannot be fully accounted for by differences in population age distributions. contrary to the long hospital stays reported in high-income settings, 17 the median time to death was 5 days following admission. the study also found substantial reductions in the reproduction number associated with the implementation of india's countrywide lockdown. india is a large country, and it does not face a single homogenous epidemic. currently, 80% of cases are reported from < 10% of its districts. 18 the epidemic is in different stages in different parts of the country, but the response has been driven by a national, overarching centralized strategy instead of being locally owned. although opportunities for containment of infections are limited, given the tremendous economic and human cost of lockdowns, a number of measures could help reduce the mortality rate and facilitate a quicker exit from the pandemic. 1. an important aspect of covid-19 management is averting deaths. the current national guidelines do not prioritize high-risk individuals for early testing, and this is a missed opportunity for averting deaths in vulnerable populations of the elderly and those with comorbidities. 2. reporting of deaths is incomplete, and because many individuals die without a covid-19 test, the number of reported deaths is likely an underestimate of the true numbers. identification of deaths offers an opportunity to learn about the disease and, thereby, prevent future cases and deaths. a formal system of mortality surveillance, specifically to measure the additional mortality attributable to covid-19, needs to be put in place. 19 3. the epidemic response should be data driven and locally owned. more granular data and greater openness to data sharing and coordination would enable surveillance data to be used for management decisions, including planning regarding personal protective equipment, medicines, supplies, and, most importantly, icu capacity and healthcare personnel. this would provide a clear picture of the impact of covid-19 to the public and could encourage greater compliance with personal protection and distancing. 4. nongovernmental organizations and civil society have been largely missing from the response to the pandemic and should be involved in helping mitigate the continued effects of the lockdown and enabling access to health care. should be updated rapidly, consistent with global research findings, and communicated clearly to clinicians. despite national guidelines, there is confusion about how best to care for patients at home with asymptomatic infection, in hospital with mild-to-moderate disease, with serious disease requiring high flow oxygen, and with severe disease requiring mechanical ventilation. 6. india is now in a season during which other diseases including dengue, chikungunya, malaria, and seasonal influenza have symptoms that are similar to those of covid-19. as these diseases are likely to have overlapping spread in the country, a clinical and testing strategy to enable distinction between the diseases is needed. 20 the covid-19 pandemic is an opportunity to invest in the public health infrastructure of india, an area of systemic neglect over the past few decades. in the short-to-medium term, developing protocols for clinical trials to investigate candidate vaccines, drugs, and monoclonal antibodies against sars-cov-2 infection will be critical to ensure optimal preventive and therapeutic management of the disease, particularly to protect those at high risk of death. 21 in the long term, a blueprint should be developed to empower and strengthen india's national and state level mechanisms for public health research, surveillance, and policy activities. as was the case in other countries, india's pandemic preparedness plan was largely abandoned in the face of a real pandemic. the response to covid-19 has been driven by political priorities rather than by public health and epidemiological expertise. given the country's size and its large global diaspora, india's battle with covid-19 will play a large role in the fate of the pandemic. as the world's largest vaccine producer, india will likely be a major supplier of vaccines against covid-19, if and when they are approved. the country's largest vaccine manufacturers are gearing up to produce covid-19 vaccines at scales that have not been attempted before. if india's vaccine industry is successful, then it will help ensure that these vaccines will be available not only to those who can pay for them but also to the hundreds of millions of impoverished people in india and in other low-and middle-income countries who need a vaccine. india stands at a critical juncture. although covid-19 is exacting a significant health and economic impact on the country, it offers an opportunity to rethink india's approach to public health. if done correctly, the legacy of covid-19 could be a much needed public investment in health, a wellequipped workforce to respond to future pandemics, and system capacity for surveillance, contact tracing, research, disease modeling, and response. emerging trends in hypertension epidemiology in india government survey found 11.8% prevalence of diabetes in india epidemiological characteristics of 2143 pediatric patients with 2019 coronavirus disease in china risks to children under-five in india from covid-19. medrxiv coronavirus in india: modi orders total lockdown of 21 days. the new york times state-wise shutdowns may be only way to prepare for spike in covid-19jopinion. india news, hindustan times statewise estimates of current hospital beds, intensive care unit (icu) beds and ventilators in india: are we prepared for a surge in covid-19 hospitalizations? medrxiv covid-19 in india: potential impact of the lockdown and other longer-term policies. center for disease dynamics prudent public health intervention strategies to control the coronavirus disease 2019 transmission in india: a mathematical modelbased approach oxford covid-19 government response tracker coronavirus cases coronavirus: india is turning to faster tests to meet targets sero-prevalence study conducted by national center for disease control ncdc, mohfw the hindu, 2020. coronavirusj57% of mumbai slum population has developed antibodies: study. the hindu. available at just 0.73% of people exposed, but india still susceptible to covid epidemiology and transmission dynamics of covid-19 in two indian states. medrxiv incidence, clinical outcomes, and transmission dynamics of severe coronavirus disease 2019 in california and washington: prospective cohort study the indian council of medical research (icmr), 2020. icmr press statement tracking covid-19 mortality in india, where deaths aren't registered properly. the wire science covid-19 and dengue fever: a dangerous combination for the health system in brazil the pandemic pipeline this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-331109-a8e7r80d authors: ibrahim, yassmin s.; karuppasamy, gowri; parambil, jessiya v.; alsoub, hussam; al-shokri, shaikha d. title: case report: paralytic ileus: a potential extrapulmonary manifestation of severe covid-19 date: 2020-08-31 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0894 sha: doc_id: 331109 cord_uid: a8e7r80d the covid-19 pandemic has recently spread worldwide, presenting primarily in the form of pneumonia or other respiratory disease. in addition, gastrointestinal manifestations have increasingly been reported as one of the extrapulmonary features of the virus. we report two cases of sars-cov-2 infection complicated by paralytic ileus. the first patient was a 33-year-old man who was hospitalized with severe covid-19 pneumonia requiring ventilator support and intensive care. he developed large bowel dilatation and perforation of the mid-transverse colon, and underwent laparotomy and colonic resection. histopathology of the resected bowel specimen showed acute inflammation, necrosis, and hemorrhage, supporting a role for covid-19–induced micro-thrombosis leading to perforation. the second patient was a 33-year-old man who had severe covid-19 pneumonia, renal failure, and acute pancreatitis. his hospital course was complicated with paralytic ileus, and he improved with conservative management. both cases were observed to have elevated liver transaminases, which is consistent with other studies. several authors have postulated that the angiotensin-converting enzyme 2 receptors, the host receptors for covid-19, that are present on enterocytes in both the small and large bowel might mediate viral entry and resultant inflammation. this is a potential mechanism of paralytic ileus in cases of severe covid-19 infection. recognizing paralytic ileus as a possible complication necessitates timely diagnosis and management. covid-19, the disease caused by sars-cov-2, was initially identified as a cluster of pneumonia cases in wuhan, a city in hubei, china, toward the end of 2019. 1 several extrapulmonary features have been reported for this disease. studies have estimated that 30-40% of cases are asymptomatic, irrespective of the viral load. 2, 3 gastrointestinal symptoms have been reported to occur in 17.6% of infected patients. 4 the most common symptoms documented are loss of appetite, followed by diarrhea, nausea/vomiting, and abdominal pain. 4 abnormal liver biochemical tests and fecal shedding of the virus have also been observed. [4] [5] [6] however, there is scarcity of knowledge in the implication of this infection on gastrointestinal complications. we describe two cases of severe covid-19 pneumonia who developed paralytic ileus during their disease course, which may represent one of the luminal manifestations of severe sars-cov-2 infection. the first case was a 33-year-old man who presented with a 2-day history of fever, cough, and sore throat. initial laboratory investigations were normal apart from elevated inflammatory markers. chest x-ray showed haziness along the mid and lower lung zones, leading to suspicion of viral pneumonia, and his nasopharyngeal swab reverse transcriptase-pcr (rt-pcr) tested positive for sars-cov-2. two days after admission, he developed tachypnea and desaturation with the progression of infiltrates on chest imaging. he was intubated, received vasopressor support, and admitted to intensive care. the patient received covid-19 treatment (ceftriaxone, azithromycin, hydroxychloroquine, methylprednisolone, and a dose of tocilizumab) guided by the local protocol. the patient was extubated after 4 days but was re-intubated because of increased oxygen requirements. examination revealed distended and tense abdomen with epigastric tenderness, exaggerated bowel sounds, and empty rectum. he was noted to have abnormal liver enzymes, alanine aminotransferase (alt) was 1,589 u/l, and aspartate aminotransferase (ast) was 1,856 u/l. abdominal x-ray showed bilateral free air under the diaphragm and distended bowel loops ( figure 1a and b). computed tomography (ct) scan of the abdomen showed large bowel dilatation from the cecum down to the rectum with no stricture or transitional area of obstruction, and patent mesenteric vessels ( figure 2a ). surgical evaluation was sought, and emergency laparotomy was performed. the operative findings were pyogenic membranes and purulent fluid in the peritoneal cavity and a 2-mm perforation in the anterior aspect of the mid-transverse colon with three serosal tears of 3-4 cm distal to the perforation; the rest of the bowel was healthy. he underwent primary repair of the perforated colon and lavage. his condition improved, and he was gradually introduced to enteral feeding. liver enzymes improved over the next week. however, 10 days later, the patient again developed respiratory distress, and repeat ct abdomen showed significant free peritoneal air with fluid contrast layering at the rectovesical pouch, raising suspicion of extra-luminal bowel contrast leak. no bowel dilatation was found. urgent laparotomy revealed a localized leak of the repaired perforated transverse colon; transverse loop colostomy was performed. the patient's hospital course was also complicated by renal failure, which was attributed to acute tubular necrosis secondary to infection and bowel perforation. he remains hospitalized in intensive care at the time of writing this report. the second case was a 33-year-old man who was brought to the hospital in severe respiratory distress. he had abdominal pain; examination revealed epigastric and right hypochondrial tenderness with guarding. his laboratory evaluation showed leukocytosis, elevated inflammatory markers, and impaired renal function with blood urea of 55.78 mmol/l, creatinine 1,891 μmol/l, and bicarbonate 1.9 mmol/l. other significant investigations included lipase of > 1,200 u/l, amylase 390 u/l, alt 187 u/l, and ast 125 u/l. he was noted to have deranged electrolytes, with potassium of 2.2 mmol/l and calcium 0.96 mmol/l. his nasopharyngeal rt-pcr was positive for sars-cov-2. bilateral pneumonic patches were observed on the chest x-ray ( figure 1c ). he was intubated and urgent hemodialysis was initiated, and he was admitted to the intensive care unit as a case of severe covid-19 pneumonia, renal failure, and acute pancreatitis. ultrasound abdomen revealed healthy gallbladder but noted dilated fluid-filled bowel loops in the left lower abdomen. he was extubated on day 5 of admission. two days after extubation, the patient developed bilious vomiting and abdominal discomfort. he did not have bowel movements for the preceding 4 days but was passing flatus. on examination, he had a distended abdomen with no guarding, but bowel sounds were sluggish. abdominal x-ray showing diffuse dilatation of the small and large bowel loops ( figure 1d ). computed tomography scan of the abdomen showed diffusely distended large bowel, from the anus to the cecum, filled with fluid, and air-fluid levels extending to involve the small bowel proximally up to the third part of the duodenum. there was no transitional zone of narrowing, mass, or bowel wall thickening. minimal peripancreatic fat stranding was noted with no fluid collection. both kidneys were small in size, and an incidental finding of soft tissue lesion at the right upper pole was noted ( figure 2b ). he was evaluated by the surgery team, who advised conservative management with a nasogastric tube insertion and correction of electrolytes. the patient was kept nil per oral, and total parenteral nutrition was initiated. he received prokinetic agents and intravenous potassium replacement to maintain target serum potassium of 4 mmol/l. the patient's condition improved over the next week, and he was able to tolerate an oral diet. a repeat ct scan performed for further evaluation of renal mass showed improvement in the significant bowel dilatation and resolution of small bowel dilatation. the patient is currently on renal replacement therapy three times a week through a permanent subclavian tunneled catheter. gastrointestinal involvement has been increasingly reported in association with the sars-cov-2 infection. a multicenter cohort study of hospitalized u.s. adults found that two-thirds of patients with covid-19 had at least one gastrointestinal symptom. 7 a review of 29 studies noted that 12% of patients with sars-cov-2 infection had gastrointestinal symptoms, including diarrhea, nausea, and vomiting. 8 the clinical significance of this presentation was demonstrated in a review by mao et al., 9 who reported that patients who presented with gastrointestinal system involvement had delayed diagnosis and also tended to have a poorer disease course. it has been proposed that the angiotensin-converting enzyme 2 (ace2) receptor plays a central role in the mechanism of gastrointestinal tract involvement in covid-19. the ace-2 receptor is the functional host receptor for the sars-cov-2 virus. 10, 11 although ace2 is highly expressed in alveolar cells in the lungs, ace2 receptors are also abundant in the gastrointestinal tract, especially in the small and large intestines. 12, 13 the gastrointestinal symptoms that appear early during covid-19 suggest that the small bowel may be an important entry site for the virus. 14 zhang et al. 15 postulated that ace2 expression on small intestinal enterocytes may mediate the invasion of the virus and activation of gastrointestinal inflammation. therefore, this could be a potential mechanism of paralytic ileus in severe covid-19 cases. there is a growing body of evidence describing large bowel involvement in covid-19 infection. carvalho et al. 16 described a case of sars-cov-2 gastrointestinal infection causing acute hemorrhagic colitis with endoscopic findings of coloproctopathy but normal histologic findings. sattar et al. 17 reported three cases of covid-19 with colonic findings, including colitis and air in the bowel wall. however, there are few reports of paralytic ileus in patients with covid-19. kaafarani et al. 18 described a series of 141 critically ill patients with covid-19, where half of the patients developed hypomotility-related complications of variable severity. only two patients were diagnosed with a colonic paralytic ileus (ogilvie's syndrome). similarly, the patient in case 1 had extensive large bowel dilatation and mid-transverse colon perforation. histopathology of the resected bowel specimen showed fat necrosis, acute inflammation, reactive fibroblastic proliferation, and hemorrhage. however, the mesenteric vessels were patent on imaging, suggesting that the etiology could be sars-cov-2-induced micro-thrombosis. it has been hypothesized that sars-cov-2 causes vascular endothelial cell inflammation, leading to impaired microcirculatory function in different vascular beds. 19 moreover, bhayana et al. 20 reported bowel wall abnormalities in 31% of abdominal ct scan images in covid-19 patients, including pneumatosis and portal venous gas. the patients were found to have bowel infarction due to ischemic enteritis with patchy necrosis and fibrin thrombi in the arterioles. our second case had diffuse distension of both small bowel and large bowel but responded well to conservative management. the development of ileus, in this case, may also be explained by electrolyte derangement, as the patient had hypokalemia and hypocalcemia. however, sars-cov-2 may cause ileus in an independent mechanism, as alluded previously. our cases were both observed to have elevated liver transaminases. in a systematic review, abnormal liver enzyme levels were noted in 15-20% of patients. 11 studies by mao et al. 9 and guan et al. 21 found that patients with severe covid-19 had higher rates of abnormal liver function. 12, 21 case 2 developed severe acute pancreatitis of unclear etiology, with elevated lipase and peripancreatic fat stranding on imaging. a few similar cases have been reported, but there is uncertainty about its pathogenesis. 22, 23 theories include acute inflammation associated with the expression of ace2 in the pancreas and heightened systemic inflammatory response from cytokine storm syndrome leading to multi-organ dysfunction. 24 a review by bourgonje et al. 25 also points out that it is the relative proportion of ace to ace2 receptors that is responsible for the resultant pro-inflammatory and pro-fibrotic symptomatology, and it has been reported that these may be affected by gender, with males having a relatively higher proportion of ace to ace2, thus favoring inflammation and colitis. both our patients were male, supporting this theory. further studies are needed to elaborate on the causative relationship between sars-cov-2 and the gastrointestinal manifestations of covid-19. in conclusion, we report paralytic small and large bowel ileus as a complication of covid-19. furthermore, we explore a potential mechanism of ileus and explore the management strategies. the added value of the present case report is the detailed histopathological evidence supporting a role for covid-19-induced micro-thrombosis, thereby compromising microcirculatory function and resultant colonic bowel dilatation and perforation in the first patient. recognizing paralytic ileus as a possible complication necessitates timely diagnosis and management. covid-19 and thrombotic or thromboembolic disease: implications for prevention, antithrombotic therapy, and follow-up: jacc state-of-the-art review prevalence of asymptomatic sars-cov-2 infection: a narrative review sars-cov-2 viral load in upper respiratory specimens of infected patients gastrointestinal manifestations of sars-cov-2 infection and virus load in fecal samples from a hong kong cohort: systematic review and meta-analysis liver injury in covid-19: management and challenges evidence for gastrointestinal infection of sars-cov-2þ prevalence and characteristics of gastrointestinal symptoms in patients with sars-cov-2 infection in the united states: a multicenter cohort study prevalence of gastrointestinal symptoms and fecal viral shedding in patients with coronavirus disease 2019: a systematic review and meta-analysis manifestations and prognosis of gastrointestinal and liver involvement in patients with covid-19: a systematic review and meta-analysis sars-cov-2 and the pathophysiology of coronavirus disease 2019 (covid-19) assessing ace2 expression patterns in lung tissues in the pathogenesis of covid-19 quantitative mrna expression profiling of ace 2, a novel homologue of angiotensin converting enzyme expression of the sars-cov-2 cell receptor gene ace2 in a wide variety of human tissues covid-19, coronavirus, sars-cov-2 and the small bowel specific ace2 expression in small intestinal enterocytes may cause gastrointestinal symptoms and injury after 2019-ncov infection sars-cov-2 gastrointestinal infection causing hemorrhagic colitis: implications for detection and transmission of covid-19 disease three cases of covid-19 di sease with colonic manifestations gastrointestinal complications in critically ill patients with covid-19 endothelial cell infection and endotheliitis in covid-19 abdominal imaging findings in covid-19: preliminary observations. radiology (epub ahead of print clinical characteristics of coronavirus disease 2019 in china covid-19 presenting as acute pancreatitis coronavirus disease-19 (covid-19) associated with severe acute pancreatitis: case report on three family members sars-cov-2 associated acute pancreatitis: cause, consequence or epiphenomenon sars-cov-2 and the pathophysiology of coronavirus disease 2019 (covid-19) this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-348192-ibohbjfb authors: odih, erkison e.; afolayan, ayorinde o.; akintayo, ifeoluwa; okeke, iruka n. title: could water and sanitation shortfalls exacerbate sars-cov-2 transmission risks? date: 2020-06-09 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0462 sha: doc_id: 348192 cord_uid: ibohbjfb sars-cov-2, the etiologic agent of covid-19, is shed in stool. sars coronaviruses have been detected in wastewater during outbreaks in china, europe, and the united states. in this perspective, we outline the risk fecal shedding poses at locations without safely managed sanitation, as in most of nigeria where we work. we believe that feco-oral transmission could occur if community transmission becomes high and sustained in densely populated cities without proper sanitation in nigeria and many other african and asian settings. in the absence of basic sanitation, or where existing sanitation is not safely managed, groundwater, which is often drawn up from wells and boreholes for drinking and household use, can become contaminated with enteric bacteria and viruses from fecal matter. endemic and epidemic transmission of multiple feco-oral pathogens via this route continues to be documented in areas without safely managed sanitation, and, therefore, the risk of sars-cov-2 transmission needs to be evaluated, tracked, and forestalled in such settings. we suggest that fecal matter from treatment facilities and recovered patients should be carefully and properly disposed. furthermore, environmental surveillance of sars-cov-2 in wastewater and accumulated human waste, as well as efforts to mitigate the virus’ entry into unprotected household water sources, should be a priority part of the covid-19 response in settings without safely managed sanitation for the duration of the pandemic. every effort must be deployed to limit continued spread of the etiologic agent of the covid-19 pandemic. mounting evidence shows that sars-cov-2 is amplified in the gastrointestinal tracts of infected people, excreted in stool, and detectable in wastewater at high levels [1] [2] [3] (rimoldi et al., 2020 ; medrxiv preprint doi: https://doi.org/10.1101/2020. 05.01.20086009). we are concerned that, in areas without safely managed sanitation, drinking and household water supplies could become contaminated with the virus. this potential risk of feco-oral transmission is highest in densely populated urban centers. sars-cov-2 has largely been transmitted via respiratory droplets and fomites from infected persons to the respiratory systems of susceptible individuals. 4 however, the virus replicates in gut enterocytes 1 and is detected in stool from patients with severe or mild covid-19, as well as from presymptomatic and asymptomatic individuals. 2, 3 recovered covid-19 patients may continue to shed virus for as long as 42 days after symptoms have ceased, even after they test negative by conventional respiratory tests. 3, 5, 6 considerable concern has been expressed in the literature that the feco-oral transmission potential for sars-cov-2 places endoscopists, caregivers of diapered children who shed the virus, 7 and fecal transplant recipients 8 at high risk of contracting the infection. for intestinal sars-cov-2 to transmit via fecal matter, it would have to be viable when shed and persist in the environment until a count greater than an oral infective dose is ingested by a susceptible individual. the cycle could potentially be shortcut by direct dissemination of fecal matter inadvertently from person-to-person or by pests like flies and cockroaches, or it could be broken if wastewater or domestic water treatment inactivates sars-cov-2. because wastewater treatment eradicates sars-cov-2 and most of the worst affected countries have robust water purification systems, community feco-oral transmission has been less extensively discussed. feco-orally transmitted pathogens are endemic in nigeria, which is among the top five countries worldwide contributing to diarrhea-derived under-five mortality. 9 a principal reason why the burden from feco-oral pathogens is so high is that for most nigerians, sewage systems are nonfunctional, incompletely functional, or nonexistent. of six major northern nigeria cities conducting polio virus environmental surveillance, only abuja operates a sewage plant. 10 in lagos, there are multiple sewage treatment plants, but their performance is suboptimal, and they therefore pose a risk of enteric pathogen transmission to surrounding areas. 11, 12 sewage handling capacity has not grown in tandem with the explosive and continued growth of this megacity so that coverage does not extend to all residents. this situation prevails in many african urban centers and in some south asian settings: between 60% and 95% of african city dwellers are not connected to sewerage and instead use a range of autonomous solutions or resort to open defecation. 13 as a result, fecal matter can be deposited into the open environment, pour from toilets unconnected to sewerage into surface water, or be buried underground in soakaways and pits from where, if these receptacles are not adequately protected, it can seep into shallow wells used for irrigation, drinking, and household purposes. 10, [13] [14] [15] according to the who/unicef, 663 million people in africa and asia do not have access to safe water, and diarrheal disease is a major cause of illness and death in those populations. 16 in recent years, surveillance of household water at a number of african and asian locations has revealed frequently found indicators of recent fecal contamination, such as escherichia coli, or outright pathogens, including enteric viruses. in each case, links have been made to human or animal open defecation, proximal latrines, or improperly processed wastewater. [17] [18] [19] in those settings, even pathogens known not to persist or thrive in the environment are among those recovered from contaminated household water or causing outbreaks. 20, 21 although there are as yet no reports of transmission of sars-cov-2 via sewage or fecal matter in settings without safely managed sanitation, or recovery from household water, these examples demonstrate that feco-oral transmission by endemic pathogenic organisms is commonplace in these settings. there is now convincing evidence, from countries with adequate sanitation, that sars-cov-2 is present in feces, around toilets, and in wastewater. 2, 22, 23 both sars-cov and sars-cov-2 nucleic acid have been detected in wastewater during outbreaks. 22, 24, 25 replicable virus is less commonly reported, although it is less commonly sought, and has been found. in laboratory studies, wang et al. 25 found that sars-cov, the agent of the 2003 sars epidemic, remains viable in water for 14 days at 4°c but for only 2 days at 20°c, suggesting that survival in tropical regions may be inadequate to sustain viability in stored water. however, coronaviruses are protected by organic matter, 26 and this will greatly affect their survival under real-world conditions. sars-cov-2 rna has been detected in substantial concentrations in wastewater and downstream water bodies. 27 rimoldi et al. reported that sars-cov-2 is susceptible to wastewater treatment and that viral infectiveness in wastewater is negligible; viral rna was amplified in untreated but not treated wastewater. 28 zhang and coworkers 29 in another preprint, however, found the china cdc-recommended sodium hypochlorite treatment of wastewater to be ineffective for the removal of sars-cov-2 rna. these studies await peer review, and further investigation is needed to clarify risks. as noted by lodder and de roda husman, 22 early finding of a covid-19 case in the united states with no known exposure to an infected case suggests that a form other than human-tohuman respiratory transmission of covid-19 may be possible. additional evidence comes from a recently published systematic review from wuhan, which spotlighted a small number of patients with diarrhea but no respiratory symptoms. 30 however, most patients in this pandemic who could have had the opportunity to be infected feco-orally to date have also been exposed to respiratory droplets or fomites; thus, the magnitude of the risk is challenging to gauge. fecooral transmission nonetheless remains a valid, if untested, hypothesis. 1, 2, 22, 26, [30] [31] [32] [33] our review of the evidence suggests that the risk of this mode of transmission in communities without basic sanitation may be high. unfortunately, countries without effective sanitation and water purification are also those least likely to have the wherewithal to detect live virus in environmental samples (detection of viral nucleic acid does not infer infectivity) and therefore measure this risk. 10, 34 as at the time of writing, most african and asian cities without basic or safely managed sanitation had reported relatively few covid-19 cases. however, case numbers are increasing, and, as they rise, the viral load in untreated fecal waste pools could escalate. this is particularly true of urban settings experiencing rapid rises in case numbers such as lagos and kano, two nigerian megacities. as at may 23, 2020, the nigerian centre for disease control had confirmed 3,357 and 883 cases in these states, respectively, together representing 56.3% of the number of cases nationally. because of occupancy pressures on isolation facilities, most nigerian cities have erected makeshift isolation and treatment facilities for patients who have tested positive, to supplement the few facilities that were available at the start of the pandemic. these new facilities often lie outside hospitals that manage their own wastewater. the same pressures on treatment facilities mean that sars-cov-2-infected persons in nigeria are discharged as soon as two consecutive respiratory swabs test negative: symptom free but likely shedding the virus when they return to their communities. other factors could additionally combine to alter the risk of feco-oral sars-cov-2 transmission. in kathmandu, nepal, where salmonella enterica typhi and paratyphi have been shown to leach into the municipal water system, breaches occur more heavily during the rains. 35 we note that african countries on the upswing of their covid-19 epidemics are just beginning the rainy season. rainy season sewage overflows can overwhelm even properly managed wastewater plants, leading to heightened enteric virus transmission. 36 on the other hand, it is possible that only very high counts of sars-cov-2 would yield orally infectious doses. thus, feco-oral transmission may only occur when the epidemic reaches an as yet unknown threshold. either way, those at risk within those settings are poor urban communities and informal settlements, which have the worst sanitation options and access to health care. individuals could become infected even if they were able to implement physical distancing recommendations, which themselves are a challenge, and need to be decisively protected from fecal sars-cov-2. 37 possible options for halting feco-oral sars-cov-2 transmission are disinfection of known open defecation sites, intensifying handwashing messages, encouraging boiling or chemical treatment of household water, and explicitly treating waste from isolation and treatment facilities. safer sewage management should be instituted or reinstituted, as priority where possible. these include ensuring that standard operating procedures are followed, and there are no interruptions in sewage decontamination, as well as quality assurance to ensure that decontamination goals are met. 11 stalled or slowed sanitation projects should be expedited, and new ones could be explored. individuals who have to work in or close to wastewater handling facilities, particularly those operating suboptimally, should be informed of their risk and provided with protection where feasible. in those situations, as research from the 2003 sars outbreak demonstrated, aerosolized virus poses a risk for respiratory transmission in addition to any feco-oral risk that may exist. 38, 39 on the positive side, fecal shedding of sars-cov-2 can be exploited for community surveillance of wastewater or human waste using similar methods that would be required for risk evaluation. 22, 40 enteric pathogen, polio, and antimicrobial resistance environmental surveillance could be leveraged, where these have been initiated, 10, 41, 42 but sites with no access to sewerage, typically not used for surveillance, must also be included. in high-risk settings, waste and wastewaterbased epidemiology could help balance sampling biases inherent in case-and contact-tracing-based human testing for covid-19 and consequently predict prevalence. 42, 43 it would also preemptively identify epidemic foci and ascertain the exact risk of community transmission via the fecal-oral route. indeed, it could represent a dedicated strategy to protect the poor and marginalized in whom outbreaks in this pandemic have typically been detected with significant lags. 44 although sanitation shortfalls risk sars-cov-2 feco-oral transmission much is focused on the current emergency, the potential risk from feco-oral sars-cov transmission should motivate and even initiate concrete steps toward lasting wastewater and sewage systems wherever possible. this would leave a post-covid-19 development legacy that could impact disease transmission, extend the value of other disease control strategies, 45 and improve the quality of life in the long term. sars-cov-2 productively infects human gut enterocytes the presence of sars-cov-2 rna in the feces of covid-19 patients characteristics of pediatric sars-cov-2 infection and potential evidence for persistent fecal viral shedding who, 2020. modes of transmission of virus causing covid-19: implications for ipc precaution recommendations: scientific brief prolonged presence of sars-cov-2 viral rna in faecal samples asymptomatic sars-cov-2 infected case with viral detection positive in stool but negative in nasopharyngeal samples lasts for 42 days faecal-oral transmission of sars-cov-2: practical implications screening faecal microbiota transplant donors for sars-cov-2 by molecular testing of stool is the safest way forward estimates of the global, regional, and national morbidity, mortality, and aetiologies of diarrhoea in 195 countries: a systematic analysis for the global burden of disease study contribution of environmental surveillance toward interruption of poliovirus transmission in nigeria assessment of wastewater discharge impact from a sewage treatment plant on lagoon water isolation and identification of enteroviruses from sewage and sewage-contaminated water in assessment of domestic wastewater management practices in the communal district i of maradi city, niger republic waste of a nation: garbage and growth in india we need to talk about crapping progress on sanitation and drinking water: 2015 update and mdg assessment safe drinking water and waterborne outbreaks bacterial contamination of drinking water sources in rural villages of mohale basin, lesotho: exposures through neighbourhood sanitation and hygiene practices the burden and characteristics of enteric fever at a healthcare facility in a densely populated area of kathmandu the dengue virus in nepal: gaps in diagnosis and surveillance a large and persistent outbreak of typhoid fever caused by consuming contaminated water and street-vended beverages sars-cov-2 in wastewater: potential health risk, but also data source aerodynamic analysis of sars-cov-2 in two wuhan hospitals coronavirus in water environments: occurrence, persistence and concentration methods -a scoping review concentration and detection of sars coronavirus in sewage from xiao tang shan hospital and the 309th hospital survival of coronaviruses in water and wastewater how sewage could reveal true scale of coronavirus outbreak presence and vitality of sars-cov-2 virus in wastewaters and rivers potential spreading risks and disinfection challenges of medical wastewater by the presence of severe acute respiratory syndrome coronavirus 2 (sars-cov-2) viral rna in septic tanks of fangcang hospital review article: gastrointestinal features in covid-19 and the possibility of faecal transmission covid-19 faecal-oral transmission: are we asking the right questions? enteric involvement of coronaviruses: is faecal-oral transmission of sars-cov-2 possible? alert for sars-cov-2 infection caused by fecal aerosols in rural areas in china divining without seeds: the case for strengthening laboratory medicine in africa the ecological dynamics of fecal contamination and salmonella typhi and salmonella paratyphi a in municipal kathmandu drinking water the impact of combined sewage overflows on the viral contamination of receiving waters slum health: arresting covid-19 and improving well-being in urban informal settlements evidence of airborne transmission of the severe acute respiratory syndrome virus environmental transmission of sars at amoy gardens leveraging africa's preparedness towards the next phase of the covid-19 pandemic global monitoring of antimicrobial resistance based on metagenomics analyses of urban sewage pathogen surveillance in the informal settlement, kibera, kenya, using a metagenomics approach letter to the editor: wastewater-based epidemiology can overcome representativeness and stigma issues related to covid-19 the moment to see the poor impact of rotavirus vaccination varies by level of access to piped water and sewerage: an analysis of childhood clinic visits for diarrhea in peru sanitation shortfalls risk sars-cov-2 feco-oral transmission key: cord-355850-jgcjscjq authors: elhadi, muhammed; msherghi, ahmed; alkeelani, mohammed; zorgani, abdulaziz; zaid, ahmed; alsuyihili, ali; buzreg, anis; ahmed, hazim; elhadi, ahmed; khaled, ala; boughididah, tariq; khel, samer; abdelkabir, mohammed; gaffaz, rawanda; bahroun, sumayyah; alhashimi, ayiman; biala, marwa; abulmida, siraj; elharb, abdelmunam; abukhashem, mohamed; elgzairi, moutaz; alghanai, esra; khaled, taha; boushi, esra; ben saleim, najah; mughrabi, hamad; alnafati, nafati; alwarfalli, moaz; elmabrouk, amna; alhaddad, sarah; madi, farah; madi, malack; elkhfeefi, fatima; ismaeil, mohamed; faraag, belal; badi, majdi; al-agile, ayman; eisay, mohamed; ahmid, jalal; elmabrouk, ola; bin alshiteewi, fatimah; alameen, hind; bikhayr, hala; aleiyan, tahani; almiqlash, bushray; subhi, malak; fadel, mawada; yahya, hana; alkot, safeya; alhadi, abdulmueti; abdullah, abraar; atewa, abdulrahman; amshai, ala title: assessment of healthcare workers’ levels of preparedness and awareness regarding covid-19 infection in low-resource settings date: 2020-06-18 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0330 sha: doc_id: 355850 cord_uid: jgcjscjq covid-19, caused by the sars-cov-2 virus, is spreading rapidly worldwide, with devastating consequences for patients, healthcare workers, health systems, and economies. as it reaches lowand middle-income countries, the pandemic puts healthcare workers at high risk and challenges the abilities of healthcare systems to respond to the crisis. this study measured levels of knowledge and preparedness regarding covid-19 among physicians and nurses. a cross-sectional survey was conducted among healthcare workers in libya between february 26 and march 10, 2020. we obtained 1,572 valid responses of a possible 2,000 (78.6%) participants from 21 hospitals, of which 65.1% were from physicians and 34.9% from nurses. the majority of participants (70%) used social media as a source of information. a total of 47.3% of doctors and 54.7% of nurses received adequate training on how to effectively use personal protective equipment. low confidence in managing suspected covid-19 patients was reported by 83.8% of participants. furthermore, 43.2% of healthcare workers were aware of proper hand hygiene techniques. less than 7% of participants received training on how to manage covid-19 cases, whereas 20.6% of doctors and 26.3% of nurses felt that they were personally prepared for the outbreak. awareness and preparedness for the pandemic were low among frontline workers during the study. therefore, an effective educational training program should be implemented to ensure maintenance of appropriate practices during the covid-19 pandemic. in late 2019, a novel covid-19 was reported to cause severe viral pneumonia in wuhan, china. it has since spread worldwide, resulting in a pandemic that has now infected more than 1.3 million people, causing more than 80,000 deaths globally. 1, 2 in february 2020, the who named the condition covid-19, which stands for coronavirus disease 2019. 3 the severity of symptomatic infections ranges from mild to critical. approximately 80% of patients have mild symptoms, whereas less than 20% experience severe symptoms such as dyspnea and shock; respiratory failure occurs in less than 5% of patients. [4] [5] [6] [7] elderly patients and/or patients with comorbidities, such as cardiovascular diseases, respiratory diseases, hypertension, diabetes mellitus, and chronic kidney diseases, are at a higher risk for severe illness. they have a higher risk of mortality than younger, or otherwise healthier, individuals. 8, 9 previous reports have confirmed that hospitalized patients had a mean age of 49-55 years. 6, 10 in an earlier report provided by the united states, regarding covid-19 patients treated between february 12, 2020 and march 2020, about 67% of those hospitalized were older than 45 years, which is similar to a prior report from china. 11, 12 acute respiratory distress syndrome (ards) is a significant complication for covid-19 patients; an estimated 20-41% of patients develop ards following a covid-19 infection and require mechanical ventilation. 5, 13 this characteristic of the disease can significantly increase the existing burden on healthcare facilities, and it requires extra resources and appropriate management. covid-19 poses a higher risk for physicians and nurses who work in critical care, emergency medicine, infectious diseases, and pulmonary medicine departments. personal protective equipment (ppe), proper handwashing, and hand hygiene are critical in decreasing the transmission and risk of infection of covid-19 in hospitals. therefore, adequate training, knowledge, and resources are necessary to prevent hospital-acquired infections due to crosscontamination to other patients who receive care in these departments. [14] [15] [16] as of april 12, 2020, the libya national center for disease control (ncdc) reported 524 suspected cases of covid-19, with 25 confirmed diagnoses. 17 however, these numbers raise a question regarding the local authorities' ability to perform adequate testing for covid-19, as the libya ncdc can perform an average of 30 tests per day. furthermore, given the current civil war, limited financial resources, and a shortage of machines and materials, libya is more vulnerable to the covid-19 pandemic. in 2017, the who released a report on the healthcare system in libya, revealing that approximately 20% of hospitals were shut down. in addition, several major hospitals only operate at 80% capacity or less. the report states that chronic respiratory disease readiness is less than 29%, whereas the readiness of emergency services is less than 47% of the target. furthermore, readiness for emergency health services is less than 18% in terms of training and 40% for diagnosis abilities. the report indicates that libya does not have a rapid response team or the ability to diagnose and respond to alerts in an organized manner. 18 this raises concerns about the local healthcare authorities' preparedness and capacity to provide an adequate response to covid-19 infection. therefore, critical preparedness, readiness, and knowledge regarding covid-19 are needed for physicians and nurses on the front line. 16, 19 few researchers have addressed the overall issues of preparedness of healthcare systems for covid-19, especially in developing countries, where resources and facilities are limited. considerable concerns have been raised regarding countries' preparedness for covid-19 and their ability to maintain control. this study aimed to determine the preparedness and knowledge of libyan healthcare workers regarding covid-19, and to develop and validate a measurement tool for estimating healthcare preparedness on a global scale. study design. a cross-sectional survey study was conducted in 21 hospitals in libya between february 26, 2020 and march 10, 2020. setting and participants. the study was conducted among physicians and nurses working in the emergency department, intensive care units (icus), and respiratory and infectious disease departments, all of whom may expect to encounter covid-19 patients. the study methodology was explained to the participants, and they were asked to provide consent before participating in the study. doctors and nurses working in other departments or private clinics were excluded from the study. a total of 2,000 targeted participants were given a paper-based questionnaire at their workplace. instruments. the self-administered and anonymous questionnaire was developed by the authors of the study and was validated in a pilot study of 30 participants, who did not participate in the final analysis. some of the questions were based on a framework similar to that of previous studies on infectious disease outbreaks. [20] [21] [22] the structured questionnaire was validated to address questions about healthcare workers' level of preparedness and knowledge of covid-19 (supplemental files i-ii). the first part of the survey was designed to collect data regarding the background characteristics of participants (age, gender, department, years of experience, information sources, and previous experience with outbreaks). the second part comprised seven items intended to evaluate the knowledge and awareness of physicians and nurses regarding covid-19 infection. the third part comprised 11 items to assess the overall preparedness in terms of managing cases of covid-19 infection. the questionnaire evaluates information sources, training experience with covid-19, diagnosis and management of covid-19 patients, use of ppe, safety precautions, isolation procedures, measures to prevent infection, and reporting procedures. to assess a given participant's knowledge, each correct answer was given a score of 1, and an incorrect answer was given a zero. scores were summed for a total score of 7, ranging from 0 to 7 for knowledge. the preparedness ranges from 0 to 11. those who scored ³ 5 on the knowledge scale were deemed to have a high level of knowledge, whereas those who scored ³ 8 on the preparedness scale had adequate preparedness. the questionnaire was developed in english for doctors and was translated into arabic for nurses using a forwardbackward translation by two qualified, independent linguistic translators at the university of tripoli. each translator independently performed a forward translation of the original questionnaire. backward translation from arabic to english was carried out by another linguistic translator. three researchers then reviewed the translated versions of the questionnaire for discrepancies. pilot tests of both versions were carried out, and cronbach's alpha was used to measure the internal consistency reliability of both versions. participants were recruited in relative proportion to the estimated number of doctors and nurses working in these departments in libya. we assumed that 70% of participants had basic knowledge and preparedness regarding covid-19, and the estimated sample size was calculated with 80% power at a 95% confidence limit, with a design effect equal to 2, and using 20 clusters. statistical analysis. data entry and analyses were performed using spss (ibm spss statistics for windows, version 25.0; ibm corp., armonk, ny). descriptive statistics were used to describe the study variables. frequency, percentage, and mean scores were used to report the descriptive analysis. the chi-square test was used to assess the association difference between the groups. statistical significance was considered for p < 0.05. ethical approval. ethical approval for this study was obtained from the bioethics committee at biotechnology research center in libya [reference number: bec-btrc-109.3-2020]. all participants provided consent before participating in the study. we obtained 1,572 valid responses out of a possible 2,000 (78.6%) participants from 21 hospitals in libya. among the 1,572 total healthcare personnel, 65.1% were physicians and 34.9% were nurses. table 1 summarizes the characteristics of the study population. figure 1 provides the distribution of participants based on departments and profession. the mean age of the population (±sd) was 35.5 ± 7.3 years for doctors and 27.8 ± 5.4 years for nurses, with women making up 64.6% of all respondents. approximately half of the participants had low confidence regarding their ability to manage covid-19 patients. most participants (70% of both doctors and nurses) named social media as a source of information for covid-19, which was statistically significant, whereas fewer than 4% viewed training courses as such. doctors had an average of 7.8 ± 7.6 years of healthcare experience, whereas nurses had an average of 5.8 ± 4.1 years. all participants worked in general public hospitals: 30.7% were in icus, 58.8% in the emergency department, 3.2% in the infectious disease department, and 7.4% in respiratory departments. a total of 47.3% of doctors and 54.7% of nurses did not receive adequate training in the use of ppe. meanwhile, 16.2% of doctors and 13.3% of nurses indicated confidence in their ability to handle suspected covid-19 patients, versus 83.8% of doctors and 86.7% of nurses who did not covid-19 level of preparedness and awareness indicate confidence in this regard. about 45% of doctors and 37% of nurses indicated that they were not prepared to handle cases of infection transmitted at the hospital level. the measurements of knowledge, awareness, and preparedness are presented in table 2 . the reliability of the knowledge and preparedness questionnaires was determined using cronbach's alpha, which revealed scores of 0.72 and 0.68 for the knowledge and preparedness scales, respectively. the results showed that less than half of all participants knew the symptoms of covid-19 infection (44.4% of doctors and 31.7% of nurses). similar results were obtained for case identification. however, less than 70% of the participants were aware of the mode by which covid-19 infection was transmitted. in addition, 52.7% of doctors and 45.3% of nurses knew about ppe. interestingly, only 43.2% of doctors and nurses were aware of proper handwashing and hand hygiene techniques to prevent the transmission of covid-19. in terms of preparedness, less than 7% of both doctors and nurses had taken courses, or training, on covid-19. about 18% of participants reported the presence of an isolation room and the availability of a known protocol for isolation. however, only about 13% of all healthcare workers felt that hospitals were prepared for the covid-19 outbreak. meanwhile, 20.6% of doctors and 26.3% of nurses perceived themselves as personally prepared for covid-19. the majority (65.8%) also reported that they were not prepared to take precautions to prevent aerosol transmission via covid-19 patients. the majority (73.5%) reported an inadequate level of knowledge on covid-19. there was no significant association between hospital department type and level of participant preparedness (p = 0.319) or knowledge (p = 0.166). tables 3 and 4 show the level of preparedness and knowledge among healthcare workers by department. a significant number of healthcare workers expressed low levels of awareness and preparedness regarding covid-19. this raises a concern regarding the ability of the libyan healthcare system and its healthcare workers to combat covid-19 infection. despite these concerns, along with the poor local healthcare infrastructure in libya, healthcare workers continue to work during covid-19, risking their lives to save their patients. meanwhile, no official courses or training programs are available, and healthcare workers have to purchase ppe themselves, as they are not provided by the hospitals in adequate amounts. our study provides considerable insights into the necessity of immediate and determined efforts focused on training programs and providing an adequate supply of ppe to alleviate these challenges during the covid-19 pandemic. inadequate knowledge is a risk factor for disease transmission, as it can lead to low levels of care. our study demonstrated that only 6.7% of doctors and 5.5% of nurses had participated in training courses. our study also reported that about 70% of participants received information from social media, which is lower than previously reported (91.1%). furthermore, our study indicated the lowest rate of knowledge compared with previous studies, where we found that the overall rate of respondents providing correct answers on the knowledge questionnaire was a mere 26.5%, compared with a previous study reporting that 90% of healthcare workers provided correct answers. 23 another recent study demonstrated that 89% of healthcare workers had sufficient knowledge on covid-19. 24 another primary concern emerging from this study is that only 21.2% of doctors and 23.9% of nurses knew the criteria for evaluating persons under investigation for covid-19 infection. in addition, only about 25% of doctors knew how to report potential covid-19 cases, which could prompt an unexpected increase in undiagnosed cases, thereby increasing the burden of infection within the community. moreover, approximately 18% of participants were unaware of isolation room specifications and processes for potential covid-19 patients, which could increase the risk of infection within hospitals. interestingly, only 44.2% of doctors and 33.3% of nurses were aware of the proper handwashing and hand hygiene techniques and disinfectants. in addition, about 45.1% of doctors and 37% of nurses were not prepared to manage a case with signs and symptoms of covid-19 infection. this could highlight the risk of cross-contamination within hospitals and could lead to a higher rate of hospital-acquired infections. training and safety precautions, focused on the direct decontamination of contact points among healthcare workers, are needed to prevent the spread of infection. 25, 26 the majority of healthcare workers (77.4%) felt personally unprepared to address covid-19 infection. a total of 50.1% of participants were uneducated about ppe, whereas about 54.3% were not trained to use it. in addition, ppe is very limited, and hospital workers reported that they independently purchase their own ppe because of the inadequate supply provided by libyan hospitals. furthermore, about 35.1% of 27 our study found that doctors and nurses were buying ppe themselves, prompting questions around the hospitals' inabilities to provide this essential equipment. only a small proportion (13.4%) of participants perceived their hospital as prepared for the outbreak. more training and education are needed on the triage and isolation of suspected cases, as approximately 25% of participants are not prepared, or trained, to conduct these protocols. limitations. although the study provided a large-scale sample from 21 healthcare centers, and used a sample size that was sufficiently large enough to allow the adequate assessment of healthcare workers' knowledge and preparedness regarding covid-19 infection, some limitations should still be clarified. the study was conducted in a single country with low resource levels and a lower number of detected covid-19 infections than other countries, which may have affected the results. future multinational studies, using more extensive and varied populations, are needed to validate these findings. in conclusion, our study has illuminated the current level of knowledge and awareness of covid-19 among doctors and nurses, with special consideration for those working in departments responsible for caring for covid-19 patients. we focused on healthcare workers who may come into direct contact with covid-19 patients, and are thus expected to have adequate knowledge and preparedness. by contrast, other studies have focused on more general populations of healthcare workers. 23, 24, 28, 29 this study provides an overview of healthcare workers' preparedness regarding the current pandemic. the respondents had a lower level of preparedness, which highlights the importance of education and training programs for healthcare workers, to control and prevent infection from covid-19. however, the absence of an organized and effective governmental plan, along with a poor healthcare infrastructure, renders developing countries vulnerable. moreover, educational initiatives, along with more tangible forms of support, such as the provision of ppe, should be carried out to help developing countries improve their abilities to control and prevent covid-19 infection. e-mails: muhammed.elhadi.uot@gmail.com, ahmedmsherghi@gmail.com, albshrimohamed@gmail.com e-mails: tariq.bugadeda@gmail.com, moutazashour@ yahoo.com, fatma.mousa91@gmail.com, mohamed.alfssi@gmail.com, belaladel94@gmail.com, and majdibadi3202@gmail.com. mohammed abdelkabir, faculty of medicine who director-general's opening remarks at the mission briefing on covid-19 -9 available at: https:// www.who.int/dg/speeches/detail/who-director-general-sopening-remarks-at-the-media-briefing-on-covid director-general's remarks at the media briefing on 2019-ncov on 11 a familial cluster of pneumonia associated with the 2019 novel coronavirus indicating person-to-person transmission: a study of a family cluster clinical characteristics of 138 hospitalized patients with 2019 novel coronavirus-infected pneumonia in wuhan, china epidemiological and clinical characteristics of 99 cases of 2019 novel coronavirus pneumonia in wuhan, china: a descriptive study persons evaluated for 2019 novel coronavirus -united states clinical course and risk factors for mortality of adult inpatients with covid-19 in wuhan, china: a retrospective cohort study characteristics of and important lessons from the coronavirus disease 2019 (covid-19) outbreak in china: summary of a report of 72,314 cases from the chinese center for disease control and prevention clinical features of patients infected with 2019 novel coronavirus in wuhan severe outcomes among patients with coronavirus disease 2019 (covid-19) -united states di napoli r, 2020. features, evaluation and treatment coronavirus risk factors associated with acute respiratory distress syndrome and death in patients with coronavirus disease 2019 pneumonia in wuhan, china infection prevention in the operating room anesthesia work area covid-19 in turkey: an urgent need for the implementation of preparedness and response strategies critical preparedness, readiness and response actions for covid-19 covid-19 status in april 10 health emergencies and humanitarian response update health-care workers' perspectives on preparedness of health-care facilities for outbreak of communicable diseases in nigeria: a qualitative study knowledge and attitude of healthcare workers about middle east respiratory syndrome in multispecialty hospitals of qassim, saudi arabia middle east respiratory syndrome-related knowledge, preventive behaviours and risk perception among nursing students during outbreak health-care workers' perspectives on preparedness of health-care facilities for outbreak of communicable diseases in nigeria: a qualitative study knowledge, attitudes, and practices towards covid-19 among chinese residents during the rapid rise period of the covid-19 outbreak: a quick online cross-sectional survey knowledge, attitude and practice regarding covid-19 among healthcare workers in henan how to train the health personnel for protecting themselves from novel coronavirus (covid-19) infection during their patient or suspected case care knowledge and attitudes of medical staff in chinese psychiatric hospitals regarding covid-19 covid-19: protecting health-care workers knowledge and attitude toward covid-19 among healthcare workers at district 2 hospital, ho chi minh city is pakistan prepared for the covid-19 epidemic? a questionnaire-based survey covid-19 level of preparedness and awareness acknowledgments: we would like to thank all of the staff who voluntarily participated in this study. publication charges for this article were waived due to the ongoing pandemic of covid-19. this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-315695-9sg7s1a1 authors: brett-major, david m.; schnaubelt, elizabeth r.; creager, hannah m.; lowe, abigail; cieslak, theodore j.; dahlke, jacob m.; johnson, daniel w.; fey, paul d.; hansen, keith f.; hewlett, angela l.; gordon, bruce g.; kalil, andre c.; khan, ali s.; kortepeter, mark g.; kratochvil, christopher j.; larson, luann; levy, deborah a.; linder, james; medcalf, sharon j.; rupp, mark e.; schwedhelm, michelle m.; sullivan, james; vasa, angela m.; wadman, michael c.; lookadoo, rachel e.; lowe, john-martin j.; lawler, james v.; broadhurst, m. jana title: advanced preparation makes research in emergencies and isolation care possible: the case of novel coronavirus disease (covid-19) date: 2020-03-30 journal: am j trop med hyg doi: 10.4269/ajtmh.20-0205 sha: doc_id: 315695 cord_uid: 9sg7s1a1 the optimal time to initiate research on emergencies is before they occur. however, timely initiation of high-quality research may launch during an emergency under the right conditions. these include an appropriate context, clarity in scientific aims, preexisting resources, strong operational and research structures that are facile, and good governance. here, nebraskan rapid research efforts early during the 2020 coronavirus disease pandemic, while participating in the first use of u.s. federal quarantine in 50 years, are described from these aspects, as the global experience with this severe emerging infection grew apace. the experience has lessons in purpose, structure, function, and performance of research in any emergency, when facing any threat. the university of nebraska medical center and its clinical partner nebraska medicine (unmc/nm) were confronted with a unique set of circumstances at the start of the u.s. experience with novel coronavirus disease (covid-19) that highlighted core lessons regarding research in emergencies that might be applied in any location, and to any disease. ultimately, unmc/nm conducted a prospective, observational cohort study beginning with covid-19-infected persons in isolation care. the rapidly traveled road to this study had many curves. the university of nebraska medical center and its clinical partner nebraska medicine are accustomed to responding to public health emergencies. it cared for patients with ebola virus disease from west africa; received persons exposed to other high consequence pathogens; established and maintained the nebraska biocontainment unit; with partners emory university, bellevue hospital, and the cdc, led the national ebola training and education center (netec); launched the national quarantine unit funded by the health and human services (hhs) assistant secretary of preparedness and response office; and established the global center for health security to coordinate its other national and international health emergency initiatives. the biocontainment unit was established in the aftermath of outbreaks of severe acute respiratory syndrome coronavirus (sars-cov) and avian influenza a in the early 2000s, getting its first use in the 2014-2016 west african ebola virus disease epidemic. the unit has critical care capabilities. the quarantine unit has airborne precaution capabilities but was designed to accommodate groups of individuals who are not ill, a need suggested by returned healthcare workers following occupational exposures in the west african epidemic. even for an institution with experience in both timely research and management of patients with highly communicable diseases, the conditions under which coronavirus disease (covid-19) was introduced to the united states and the pervasive challenges of patientcentered research in emergencies complicated considerations ( figure 1 ). nebraska's first guests related to covid-19 were 57 men, women, and children evacuated from wuhan, china, and in quarantine. they arrived on federally chartered aircraft and passed through carefully arranged screening checks manned by cdc officials. then, uniformed public health service personnel from other agencies within hhs and managed by the assistant secretary for preparedness and response office provided support. security was present, including u.s. marshals. everyone-including those who had placed a large nebraska welcome banner at the airport, deposited gift baskets in dormitory rooms, or wielded thermometers-wanted these persons to arrive well, stay well, and feel welcome. these individuals were glad to be back in the united states. they did not want others to become infected with covid-19 if, in fact, they proved to be ill. nonetheless, they were constrained by schedule, location, and physical and human barriers until their departure from quarantine. on the verge of the departure of the evacuees from china, unmc/nm received a mix of 11 isolated (infected) and two quarantined (not known to be infected) individuals evacuated by federal authorities from a cruise ship in japan. the evacuees from wuhan were among the first in the united states to be placed under quarantine by the federal government under new authorities established in the 2017 revision of the code of federal regulations 1,2 (box 1). federal quarantine orders-the first such use in over 50 yearspresented contextual challenges. in general, u.s. quarantine stations exist at major points of entry, such as at large international airports, where small numbers of sentinel cases of an emerging disease are thought to be most likely to be first encountered in the united states. 3 in late 2019, unmc/nm opened the first national quarantine unit. this twenty-bed unit is designed to host larger numbers of quarantined persons than existing, smaller quarantine stations. it is colocated with a national training resource for public health emergency personnel, in close proximity to the nebraska biocontainment unit, to enable more advanced care if needed. 4 this large group of 57 persons, however, were managed by federal authorities at camp ashland-a nebraska army national guard base outside of omaha-with some logistics support from unmc/nm. by the time that the quarantined persons from wuhan arrived, experts had already considered the possibility that sars-cov-2 might shed before symptoms, facilitating its ability to achieve sustained human-to-human transmission. 5 for this reason, unmc/nm initially sought to test asymptomatic individuals to inform their case management and how they were housed. however, a consensus regarding the advisability of testing could not be reached with authorities because of concerns regarding their personal autonomy (whether the quarantined persons understood the implications of testing and could make a choice freely) and uncertainty about what to do about isolated negative test results. testing was not pursued. in the end, none of the quarantined evacuees from wuhan demonstrated clinical evidence of covid-19. 6 the question of scope of presymptomatic shedding remained unanswered. toward the end of that quarantine, on 17 february, unmc/ nm received a group from a cruise ship in japan comprised mostly of covid-19-infected persons. 7 the infected individuals were under federal isolation orders as opposed to quarantine; they were known to be infected. whether simply being observed in the setting of few or no symptoms, or more ill and in need of hospital level care, they were housed at unmc/nm. by that time, asia had accumulated many cases, and a literature base was developing. 8, 9 nonetheless, cases in the united states remained few, and availability of information and specimens from affected areas in asia that were relevant to medical countermeasure development was limited. this prompted unmc/nm to launch its own research initiative for the prospective assessment of patients. it did so against a backdrop of initial hesitancy because of complex issues of patient autonomy under federal orders, interagency jurisdiction challenges as different governmental actors exercised their perceived obligations for oversight, and known larger patient populations in other countries that might make local research less important. once the decision to initiate research was made, one of the immediate questions was on what? unmc/nm participated in the national institute of allergy and infectious diseases (niaid) studies of drug therapy against ebola virus disease, and this collaboration continued in support of an adaptive randomized controlled trial with the antiviral drug remdesivir. 10, 11 as the first institution to initiate this trial for covid-19 patients in the united states, unmc/ nm assisted expansion of the trial to additional sites via its rapid response central institutional review board (irb) mechanism for the netec special pathogens research network. finding an effective drug, however, is not the only purpose of doing research in emergencies. early in response efforts, a critical questions and ethics committee was formed. pulling from a multidisciplinary base, its purpose was providing a space for leadership and others to air questions, concerns, and challenges that might represent an obstruction to effective risk management-a space to reflect amidst an otherwise operationally fast-paced environment. fielded questions were sometimes narrow and sometimes broad. they often highlighted uncertainty about the disease itself, which limited the ability to make evidence-based decisions. this process facilitated stakeholders coming together to start pursuing answers (box 2). the committee also undertook a survey of research associated with the response and started to link risk management challenges with sources of information that might assist decision-making. several needs were evident as research planning discussions ensued (box 3). importantly, these discussions led to a broad picture of how a platform for research might be applied, and a prospective, observational cohort study design was selected. fortunately, the international severe acute respiratory and emerging infection consortium (isaric) and the world health organization (who) had been working on a protocol for just such a prospective, observational cohort study for several years. 12 known as the clinical characterization protocol for severe emerging infections, it represents a longitudinal effort to generate and keep updated an internationally harmonized protocol for the evaluation of emerging infections. 13 the existence of a well-developed protocol with case report form, informed consent documents, and other supporting materiel had immediate advantages. from a science management perspective, the most striking aspect was that the well-documented evolution of the protocol simplified local scientific review requirements. moreover, it was easier to edit than to initiate writing. unmc/nm changes to documents reflected technical preferences, differences in local law or institutional requirements, or using the documents in a referral academic center rather than a resource-limited setting. overall, the isaric materials saved at least several days in the process and provided important guideposts. unmc/nm have several unique features in its irb. the irb has technical breadth, a dedicated pool of community representatives, and a process for rapid review. in addition, the university has invested in this office so that when called upon for rapid reviews, there are sufficient highly committed staff to participate in management and oversight of the process, as well as consultation with petitioning investigators. just as importantly, the irb has experience with reviews in emergencies and related exercises. it also has worked through how to facilitate cooperative research through its central irb mechanism for the special pathogens research network, comprising 10 academic centers that serve as regional referral isolation care hospitals. 14 the regulatory process reflects a general posture toward discovery in parallel with clinical care shared across its network partners. 15 operational efficiency such as that provided by the central irb was impactful in ensuring the window of opportunity was not lost. a curious, structural aspect of research preparedness that became clear while assisting other sites considering adoption of the unmc/nm prospective, observational cohort study was the importance of routine access. unmc/nm and other referral location personnel regularly access isolation care spaces in training and response activities, as well as participate in community coordination in the management of patients who may have an infection with a high consequence pathogen. consequently, the primary pool of investigators box 1 a revised quarantine law's first use on january 31, 2020, the cdc issued a federal mandatory quarantine order for 195 americans evacuated out of wuhan, china, on january 29. 18 effective february 2, health and human services declared a mandatory quarantine for any u.s. citizens or permanent residents returning to the united states who had returned from the hubei province of china in the previous 14 days. in addition, all u.s. citizens and permanent residents returning from mainland china were required to undergo two weeks of self-monitoring. this marked the first time in over 50 years that mandatory federal quarantine had been invoked under the cdc's jurisdiction. by contrast, federal isolation orders have been comparatively common. isolation differs from quarantine in that isolation requires infection with a quarantinable, communicable disease, not just exposure. between 2005 and 2016, the cdc issued 12 federal isolation orders, 19 relying mainly on port-of-entry screening. historically, state and local health departments have executed most quarantine orders. in 2019, to decrease the spread of measles in california, los angeles county did so for more than 200 individuals at two college campuses. 20 the quarantine measure met little resistance. other state orders have not been so well received. in 2014, new jersey issued a quarantine order for a nurse returning to the united states after caring for ebola patients in sierra leone. afterward, the nurse filed suit in federal court, stating that new jersey had violated her constitutional rights to liberty and due process. she later dropped her suit, settling in favor of changes to the state's quarantine regulations, including provisions for the right to counsel, notice of hearings, visitor rights, and the right to privacy. 21 the case suggests where some points of friction may arise as the cdc continues its covid-19 orders. coronavirus disease-19 quarantines also were the first to test recently updated regulations. in 1967, quarantine authority shifted to the cdc for cases involving ports of entry, with interstate quarantine added to the cdc's jurisdiction in 2000. related regulations have had several updates, most recently in 2017, with a stated focus on individuals' due process rights. some anticipated issues include mandatory reassessment of quarantine cases, social distancing practices, compensation for lost wages, and payment for the care and treatment of quarantined individuals. under the current regulations, any federal isolation or quarantine order must be reassessed within 72 hours of issuance of the order, seemingly impractical in light of large numbers of related cases, if conducted individually. it may be impossible to house each person alone, despite the consequences for housemates if the person is infected. in addition, regulations do not expressly direct payment for the care and treatment of individuals subject to a federal quarantine. these costs may include diagnostic testing. the regulations allow that the director of the cdc may authorize payment for such care and treatment, but that payment is in the cdc's sole discretion. this language leaves matters of payment open to interpretation and negotiation, which may be a hindrance to real-time decision-making. many of these issues relate to differences between small-scale quarantines and the additional challenge of larger scale events, as relevant for covid-19. as this health emergency evolves, ambiguous guidelines, combined with the unprecedented nature and scale of this quarantine, could impact the operational response. the cdc is in a unique situation to take precedent-setting action, establishing new standards for how federal quarantine should occur in the united states for many years to come. needed at the bedside and in the laboratory are readily able to undertake practices and follow procedures within containment areas, including institutional biosafety committeeappropriate laboratory spaces. consequently, when an emergency such as the covid-19 pandemic occurs, the work is feasible. in each emergency, some structures preexist, some must be applied anew, and priorities must be set. 16 for covid-19 with a remdesivir drug trial from niaid on site and its potential to impact care generally, unmc/nm tiered offers of enrollment to its patients, first screening for the drug trial before considering other research on a given patient. in just over a week from conception, in the beginnings of delivering isolation care, seven participants with covid-19 infections were enrolled in a prospective, observational cohort study for severe emerging infections. the study rapidly accumulated both prospectively collected and residual clinical specimens. in contrast to accumulated experiences in asia and some other affected areas, the cohort was small. nonetheless, it captured high-quality specimens coupled with data of value to researchers and product developers in the midst of a new emerging infectious disease. the unmc/nm prospective observational cohort study incorporated a tissue bank, allowing the later use of study specimens. in accordance with regulatory requirements, it has a governance structure. a priorities steering committee was established immediately, including some members of the investigator group and other stakeholders. a formal request process for use of data and specimens was instituted, and a request tracker quickly filled with governmental, academic, and industry requests that were as varied as they were rapid. the committee adopted a long view for use of the tissue bank, recognizing the need to balance exigent with future possibilities for use. it recognized the importance of transparency, cooperative work meeting aims not achievable by smaller groups or individuals, and the need to facilitate meaningful innovation that might not be served by other initiatives. in emergencies, however, long views may not be popular, requests are not always rational, disclosures and realistic assessments are not often available on the actual utility of an experiment in the context of the emergency at hand, and respect for autonomy and appreciation of the intent of a gift of data and sample by a patient are not always appreciated. however, that investigators believe in their work and seek to advance discovery is important for both patients and science. being arbiters of limited resources in this context means that everyone must make compromises. unmc/nm launched the prospective, observational cohort study without external funding. cohort studies often are box 2 critical questions and ethics vignette the university of nebraska medical center and its clinical partner nebraska medicine established a critical questions and ethics committee immediately before experiencing its first covid-19 patients. this allowed decision-makers and implementers alike a space in which to air concerns based on unanswered questions or perceived operational or organizational risks. the committee was advisory in nature. one such question asked how best to prioritize n95 respirators that were anticipated to be in short supply. the conversation revolved around fit-testing requirements. at a center like unmc/nm, several hundred respirators are consumed each year in quantitative fit testing for staff who have newly arrived, or for required periodic testing. logistical, ethical, legal, and operational considerations included finding the right balance between the need for appropriate fit-especially if at high risk of sars-cov-2 exposure, differences in regulatory intent for fit testing and a more rigorous standard applied by the university, and preconceived notions of need, practice, and requirements. several small program adjustments were thought to have promise. these were reevaluating nondestructive or qualitative fit testing, using a survey to enable a longer interval before retesting, and prioritizing new employees and areas with higher risk for encounters with ill patients. important research avenues emerged, and this process highlighted the need for interdisciplinary approaches. environmental hygiene, logistics, and implementation science aims all arose from the conversation in ways that might not otherwise have emerged. decision-related knowledge needs relevant to the prospective, observational cohort study described in this article have included viral shedding dynamics, clinical course relevant to resource demand, and the horizon of available medical countermeasures and their development. â�¢ risk identification and characterization of the disease in patients â�¢ hypotheses generation with a potential to impact patient-and community-centered outcomes â�¢ continual patient population assessment so that work to test hypotheses is best designed and fundamental processes are well framed and practiced â�¢ flexibility to interact with clinical care and public health teams when the study could provide meaningful information, particularly in real time or near real time, including coordination with environmental sampling and testing â�¢ potential to explore data, specimens, and the results of analysis over time, to include the potential for cooperative work with partners across stakeholder groups â�¢ flexibility to adjust the schedule of events when exigencies such as when infection prevention and control posture or immediate patient interests require changes â�¢ durable rather than fleeting investment of time and other resources, so that all are ready when new health emergencies present challenging to fund. research dollars tend not to align with durable, multi-threat capabilities. 17 such studies may be supported in part through sub-study funding, for instance, to test a particular device and assay in the laboratory on samples from cohort members. without new funding solutions, the development of valuable cohorts may not be possible. the university of nebraska medical center and its clinical partner nebraska medicine quickly established a prospective, observational cohort study for severe emerging infections during the 2020 covid-19 emergency, while supporting national quarantine and isolation care activities and launching an niaid randomized, controlled drug trial. this was possible thanks to preexisting resources from the international community and durable partners, as well as structures that support research review and execution with intrinsic aspects that allow flexibility. studies in emergencies must be designed in ways mindful of the context in which they start, and yet have a long view. as in all science, aims must be clear, mechanisms for governance present, and opportunities for reflection and input encouraged. despite challenges and sometimes a lack of external funding support, research is a worthwhile undertaking to advance understanding and seek risk management solutions. 20-fb, unmc/nm clinical characterization protocol for severe emerging infections. this study is currently operated on institutional funding only, although hhs support of quarantine and isolation efforts is received. the niaid rct for remdesivir also is running at unmc/nm with irb approval and extramural support. views expressed are those of the authors and not necessarily those of the state of nebraska, the u.s. government, or any of their agencies e-mails: david vice chancellor for research office code of federal regulations 71.33, persons: isolation and surveillance. washington, dc: us government quarantine and liability in the context of ebola quarantine stations 2020 global center for health security capabilities 2020 report of the who-china joint mission on coronavirus disease 2019 (covid-19) rescued americans depart camp ashland daily coronavirus update. omaha, ne: nebraska medicine early transmission dynamics in wuhan, china, of novel coronavirus-infected pneumonia characteristics of and important lessons from the coronavirus disease 2019 (covid-19) outbreak in china: summary of a report of 72314 cases from the chinese center for disease control and prevention a randomized, controlled trial of zmapp for ebola virus infection nih, 2020. nih clinical trial of remdesivir to treat covid-19 begins. bethesda, md: nih. available at open source clinical science for emerging infections the special pathogens research network: enabling research readiness a novel approach to infectious disease preparedness: incorporating investigational therapeutics and research objectives into full-scale exercises catching chances: the movement to be on the ground and research ready before an outbreak consequences of pathogen lists: why some diseases may continue to plague us secretary azar delivers remarks on declaration of public health emergency for 2019 novel coronavirus control of communicable diseases final rule a. and california state-los angeles order quarantines amid measles outbreak. the new york times the cdc's communicable disease regulations: striking the balance between public health and individual rights acknowledgments: we thank our patients; our clinical and administrative staff; emergency medical services; public health officials at county, state, and federal levels; and many others for their important contributions to daily work and care of delivered patients, without which this research would be impossible. unmc/nm nursing is superb and important to application of study procedures. shahnaz benner conducted phlebotomy in isolation care, and morgan shradar conducted swab sampling. teresa hartman and sara donovan assisted in developing the critical questions and ethics committee this is an open-access article distributed under the terms of the creative commons attribution (cc-by) license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-321514-knyw023l authors: bénet, thomas; picot, valentina sanchez; awasthi, shally; pandey, nitin; bavdekar, ashish; kawade, anand; robinson, annick; rakoto-andrianarivelo, mala; sylla, maryam; diallo, souleymane; russomando, graciela; basualdo, wilma; komurian-pradel, florence; endtz, hubert; vanhems, philippe; paranhos-baccalà, gláucia title: severity of pneumonia in under 5-year-old children from developing countries: a multicenter, prospective, observational study date: 2017-07-12 journal: am j trop med hyg doi: 10.4269/ajtmh.16-0733 sha: doc_id: 321514 cord_uid: knyw023l pneumonia is the leading cause of death in children. the objectives were to evaluate the microbiological agents linked with hypoxemia in hospitalized children with pneumonia from developing countries, to identify predictors of hypoxemia, and to characterize factors associated with in-hospital mortality. a multicenter, observational study was conducted in five hospitals, from india (lucknow, vadu), madagascar (antananarivo), mali (bamako), and paraguay (san lorenzo). children aged 2–60 months with radiologically confirmed pneumonia were enrolled prospectively. respiratory and whole blood specimens were collected, identifying viruses and bacteria by real-time multiplex polymerase chain reaction (pcr). microbiological agents linked with hypoxemia at admission (oxygen saturation < 90%) were analyzed by multivariate logistic regression, and factors associated with 14-day in-hospital mortality were assessed by bivariate cox regression. overall, 405 pneumonia cases (3,338 hospitalization days) were analyzed; 13 patients died within 14 days of hospitalization. hypoxemia prevalence was 17.3%. detection of human metapneumovirus (hmpv) and respiratory syncytial virus (rsv) in respiratory samples was independently associated with increased risk of hypoxemia (adjusted odds ratio [aor] = 2.4, 95% confidence interval [95% ci] = 1.0–5.8 and aor = 2.5, 95% ci = 1.1–5.3, respectively). lower chest indrawing and cyanosis were predictive of hypoxemia (positive likelihood ratios = 2.3 and 2.4, respectively). predictors of death were streptococcus pneumoniae detection by blood pcr (crude hazard ratio [chr] = 4.6, 95% ci = 1.5–14.0), procalcitonin ≥ 50 ng/ml (chr = 22.4, 95% ci = 7.3–68.5) and hypoxemia (chr = 4.8, 95% ci = 1.6–14.4). these findings were consistent on bivariate analysis. hmpv and rsv in respiratory samples were linked with hypoxemia, and s. pneumoniae in blood was associated with increased risk of death among hospitalized children with pneumonia in developing countries. despite reduced mortality rates in recent years, pneumonia is the foremost cause of death from infectious diseases in under 5-year-old children worldwide, accounting for 15% of total deaths, mostly in developing countries. 1 hypoxemia, frequently associated with pneumonia and a marker of disease severity, leads to 3-to 4-fold increased risk of death in children with pneumonia. 2 a cochrane review 3 has reported that systematic hypoxemia screening with pulse oximetry and appropriate oxygen supply are effective in preventing death from pneumonia in children. a large simulation study estimated that systematic pulse oximetry may globally prevent almost 150,000 deaths from pneumonia annually. 4 however, the microbiological agents linked with hypoxemic pneumonia are poorly recognized. the identification of such etiological agents would serve to better target preventive (i.e., vaccination) and curative measures (i.e., antibiotics and antiviral drugs), reducing the global burden of hypoxemia and pneumonia. because of its high incidence and related mortality, particular attention must be paid to hypoxemic pneumonia in developing coun-tries. pulse oximetry is still rarely available in health-care settings of developing countries. 5 in the absence of oximeter, hypoxemia can be detected by several clinical signs or symptoms, including cyanosis and increased respiratory rate. however, none is sensitive and specific enough to reliably detect hypoxemia. [6] [7] [8] most studies of hypoxemia in children with pneumonia were performed in one country and rarely investigated the relationship between hypoxemia and microbiological results, so it would be useful to reassess them in a more recent multicontinental investigation. 2 the risk factors of death among children with pneumonia in developing countries have already been identified, 9, 10 but rarely regarding the relationship between microbiological findings and mortality. assessment of clinical, para-clinical, and microbiological predictors of death would be useful to prioritize public health campaigns. identification of microbiological agents associated with death and/or hypoxemia would be useful to better focus therapeutic measures. indeed, hypoxemic pneumonia can be treated with oxygen in conjunction with other measures, whereas non-hypoxemic pneumonia with poor vital prognosis might need other regimens, such as antibiotics/antivirals or intensive care. the objectives of the present study are to assess the microbiological agents linked to hypoxemia in hospitalized children with pneumonia in developing countries, to identify clinical and para-clinical predictors of hypoxemia and to pinpoint factors associated with death within 2 weeks after admission. study sites and design. findings from a prospective, hospital-based, multicenter, longitudinal study, conducted at five sites in four countries located on three continents, were analyzed: lucknow and vadu in india, antananarivo in madagascar, bamako in mali, and san lorenzo in paraguay. the participating sites are members of the gabriel (global approach to biological research, infectious diseases and epidemics in low-income countries) network established by fondation mérieux. 11 the study protocol and sites are described elsewhere. 12 pneumonia cases from the original case-control study were followed up during hospital stay and constituted the analyzed cohort. the study population comprised children aged between 2 and 60 months, complying with protocol definitions and inclusion criteria. eligible patients were identified by study clinicians at each participating site. all consecutive patients hospitalized in pediatric departments, who were eligible for study entry, were enrolled during each season (dry and rainy) for at least a 1-year period. the study aimed at obtaining an equal number of individuals in each season at each study site. incident cases were defined as hospitalized children aged 2-60 months, with clinical features of pneumonia, as described latter, radiological confirmation of pneumonia on chest x-ray as per world health organization (who) guidelines, 13 and informed consent statement signed by the children's parents or legal guardian. wheezing at auscultation was initially an exclusion criterion, but was amended because it slowed the inclusion process. thus, it was finally decided to include children presenting pneumonia with or without "wheezing." the present study selectively comprised sites with better quality data on oxygen saturation (so 2 ) at admission, mortality among pneumonia cases, and documented recording of patient follow-up during hospitalization. main characteristics were compared by site, and multivariate analysis accounted for heterogeneity of sites regarding observed or non-observed potential confounding factors. patients with missing data on follow-up, so 2 measurement, and vital status at discharge were excluded (n = 59). excluded patients did not differ from those analyzed for gender and weight-for-height z score, but were older (p = 0.03). definition of pneumonia. pneumonia cases were defined by the following criteria: data sources and quality control. data quality was monitored and evaluated by each site and by the emerging pathogens laboratory (lyon, france) for pooled data analysis. demographic characteristics, underlying diseases, medical history, clinical examination at enrollment, therapeutics, vaccinations, and outcomes were recorded prospectively for each patient on a standardized paper form. each potential error was discussed with local investigators, and a final ruling was applied. the principal investigator at each site was informed about quality assessments and was involved in their resolution. hypoxemia was defined as so 2 < 90%, according to who recommendations. 15 so 2 was measured at hospital admission, before the administration of oxygen or other therapeutics. vital status was recorded until patient discharge. biological samples. samples were collected in the first 48 hours of patient hospitalization. nasal swabs/aspirates, whole blood, and pleural effusions (in case of pleurisy) were sampled from all patients. urine was collected at patient admission to ascertain history of antibiotic use. biological samples were taken before the in-hospital administration of antibiotics. whole blood allowed complete blood count and culture, with real-time multiplex polymerase chain reaction (rt-pcr) assay for the identification of staphylococcus aureus, streptococcus pneumoniae, and haemophilus influenzae type b. c reactive protein (crp) and procalcitonin (pct) were quantified in serum. respiratory specimens permitted the identification of viruses and bacteria by rt streptococcus pneumoniae-positive specimens were serotyped by multiplex rt-pcr that detects 29 different serotypes. a centralized, blinded pcr respiratory quality control panel was provided to all sites to ensure procedure validation on-site before specimens were processed locally. statistical methods. qualitative variables were described as numbers and percentages with comparison by χ 2 test, if appropriate, or fischer's exact test. quantitative variables were reported as median and interquartile range (iqr) and compared by the mann-whitney u test or kruskal-wallis one-way analysis of variance. positive likelihood (lr+) and negative likelihood (lr _ ) ratios of various clinical signs and symptoms were calculated to detect hypoxemic cases with the following formulae: lr+ = sensitivity/(1 _ specificity) and lr _ = (1 _ sensitivity)/specificity. it has been underlined that lr calculation is useful to improve diagnostic accuracy. 16 lr could thus express the proportion of hypoxemic children who presented a particular sign or symptom divided by the proportion of non-hypoxemic children with the same result. microbiological findings from different sample sites associated with hypoxemia were assessed by logistic regression modeling. multivariate analysis was performed after univariate analysis, with forced adjustment on patient age, time per quarter, and study center. microorganisms with p < 0.15 values on univariate analysis were initially entered in the multivariate model. thus, backward stepwise deletion was applied until all p values were < 0.05. models were compared by wald testing. factors associated with in-hospital mortality were assessed with kaplan-meier curves and compared by log-rank test. follow-up was censored at 14 days after admission or discharge, if duration of hospitalization was less than 14 days. the characteristics of patients deceased within 2 weeks (n = 13) were compared with non-deceased patients (n = 392). univariate and bivariate proportional hazard cox regression analyses were undertaken. no multivariate cox model was fitted owing to the limited number of events. bivariate analyses expressed the effect of one major risk factor from univariate analysis adjusted on one other possible confounder (age category, human immunodeficiency virus [hiv] seropositivity, time per quarter, or weightfor-height z score). all tests were two tailed, and p < 0.05 was considered significant. statistical analysis was conducted with stata version 13.0 (statacorp., college station, tx). ethics. the study protocol, informed consent statement, clinical research form, amendments, and all other study documents were submitted to and approved by the institutional research ethics committee of each site. population description. overall, 405 children with pneumonia, accounting for 3,338 hospitalization days, were included. among them, 235 (58.0%) were male. median age was 14 months (iqr = 8-27 months). ninety-six (23.7%) patients came from lucknow, india, 71 (17.5%) were from vadu, india, 26 (6.4%) were from antananarivo, madagascar, 114 (28.1%) from bamako, mali, and 98 (24.2%) from san lorenzo, paraguay. the study periods at each site were patients differed between sites according to median weight-for-height z score (p = 0.001) and age category (p = 0.007), but did not differ between sites according to hiv seropositivity (p = 0.33) and mortality (p = 0.37). seventy patients were hypoxemic at admission. global prevalence of hypoxemia was 17.3% (95% confidence interval [ci] = 13.9-21.3%). median so 2 was 95% (iqr = 92-97%), without differences between countries (from 95% [iqr = 87-96%] in mali to 96% [iqr = 89-98%] in vadu, india, p = 0.14). table 1 compares the characteristics of hypoxemic and nonhypoxemic patients. hiv prevalence was 1.0% (n = 3). median weight-for-height z score was _ 1.1 (iqr = _ 2.4; +0.1). median length of hospital stay was 7 days (iqr = 4-10 days). median crp level at admission was 24 mg/l (iqr = 6-90 mg/l), median white blood cell count was 12,600 × 10 9 cells/l (iqr = 1,000-25,000 × 10 9 cells/l), and median neutrophil proportion was 47% (iqr = 28-64%). median pct level at admission was 1.9 ng/ml (iqr = 0.3-8.1 ng/ml), with mean of 15.6 ng/ml (minimum: 0.05, maximum: 585.5 ng/ml). overall, 70.5% tested positive for urinary antibiotics at admission. among the 402 (99.3%) patients given antibiotics during hospitalization for a median duration of 7 days (iqr = 5-10 days), 298 (74.5%) received monotherapy, with some also getting multiple antibiotic lines. the main drugs were ceftriaxone (n = 135, 26.0%), amoxicillin (n = 120, 23.1%), ampicillin (n = 51, 9.8%), amoxicillin/sulbactam (n = 36, 6.9%), amoxicillin/clavulanic acid (n = 35, 6.7%), oxacillin (n = 35, 6.7%), and vancomycin (n = 22, 4.2%). microbiological agents associated with hypoxemia in children with pneumonia. hypoxemic (n = 70) and nonhypoxemic (n = 335) pneumonia cases did not differ by median number of bacteria (1 versus 1, respectively, p = 0.85) or viruses detected (1 versus 1, respectively, p = 0.72) in nasal swabs/aspirates. infection types (bacterial/viral/mixed) did not differ in hypoxemic and non-hypoxemic patients (p = 0.96). table 2 reports the microbiological agents linked with hypoxemia. univariate analysis disclosed that hmpv and rsv detection in nasal samples was associated with increased risk of hypoxemia (p = 0.04 and 0.008, respectively). after adjustment on age, center, and calendar time, microorganisms independently associated with heightened risk of hypoxemia were hmpv (adjusted odds ratio [aor] = 2.5, 95% ci = 1.1-5.3) and rsv (aor = 2.4, 95% ci = 1.0-5.8). median so 2 was lower in rsv-and hmpv-positive patients than in -negative patients but not different between rsv-and hmpv-positive patients ( figure 1 ). pneumococcus serotypes 6ab and 10a from respiratory samples were more frequent in hypoxemic than in non-hypoxemic patients (22.9% versus 9.5%, respectively, p = 0.002; 7.1% versus 1.5%, respectively, p = 0.006). the distribution of other serotypes was not significantly different in hypoxemic and non-hypoxemic children (supplemental figure 1) . clinical and para-clinical presentation in children with hypoxemic pneumonia. hypoxemic patients differed from non-hypoxemic patients according to age (p = 0.02), history of common cold/pharyngitis (p < 0.001), receipt of one dose of pentavalent vaccine (p < 0.001), breathing rate (p = 0.009), blood pressure (p < 0.01), chest indrawing (p = 0.02), cyanosis (p = 0.003), conjunctivitis (p = 0.01), rasping (p = 0.03), radiological presentation (p = 0.009), mean white blood cell count (p < 0.001), and pct (p < 0.001) at admission ( table 1) . lr+ of lower chest indrawing was 2.3 (95% ci = 1.1-4.9), and lr-was 0.9 (95% ci = 0.8-0.95). lr+ of cyanosis was 2.4 (95% ci = 1.4-4.1) and lr _ was 0.7 (95% ci = 0.5-0.99). other signs and symptoms were less predictive of hypoxemia (data not shown). hypoxemic patients differed from non-hypoxemic patients in mean white blood cell count (p = 0.001) and pct at admission (p = 0.03) but not regarding crp level. factors associated with death. fourteen (3.5%) patients died during hospital stay. among them, 13 died within 14 days after hospital admission. the mortality rate was 8.6% in hypoxemic and 2.4% in non-hypoxemic patients (p = 0.01). eight deceased patients were not hypoxemic at admission. among them, the causes of death were multiple organ dysfunction syndrome (n = 4), acute respiratory distress syndrome with septic shock (n = 2), severe pneumonia in hiv (n = 1), and cardiac arrest (n = 1). table 3 reports the microbiological, clinical, and paraclinical findings associated with death on univariate cox analysis. streptococcus pneumoniae detection by blood pcr, hypoxemia, and pct ³ 50 ng/ml at admission were associated with increased risk of death (log-rank test: p = 0.003, p = 0.002, and p < 0.0001, respectively, figure 2a -c). other characteristics or microorganisms, including s. pneumoniae in respiratory samples, were not significantly associated with death ( supplemental tables 1 and 2) . no figure 2 ). in addition, s. pneumoniae detection by blood pcr was associated with increased risk of death (aor = 4.0, 95% ci = 1.3-12.4), independently of hypoxemia at admission (aor = 4.3, 95% ci = 1.4-12.8). one of the objectives of this study was to assess microbiological agents and other predictors of hypoxemia and death in under 5-year-old hospitalized children with pneumonia from developing countries. we observed that two viruses, namely rsv and hmpv, detected in respiratory samples by pcr, were independently associated with increased risk of hypoxemia, while no bacterial agent was significantly linked with it. on the other hand, s. pneumoniae detection by blood pcr was associated with a higher rate of in-hospital mortality in the study population independently of hypoxemia at admission. several predictors of hypoxemic pneumonia were identified. however, none had high likelihood ratio. elevated pct concentration and hypoxemia were straightforward predictors of death in children with pneumonia. the contribution of hypoxemia to the risk of death was independent of pneumococcus detection by blood pcr. several studies have investigated factors associated with hypoxemia, particularly clinical predictors. [6] [7] [8] 17 however, few of them have researched the links between microbiological findings and severity in different developing countries, with a standardized protocol. we observed that two viruses were associated with increased risk of hypoxemia. viral pneumonia induced diffuse, bilateral, pulmonary damage, 18 compared with bacterial pneumonia, with more frequent, well-systematized alveolar localization. 19 this is probably the reason why we noted that the two viral etiological agents evoking pneumonia were associated with hypoxemia. rsv is the leading cause of viral pneumonia in children, 20 frequently in association with severe disease. 21 we observed that it might be also a major cause of hypoxemic pneumonia. on the other hand, rsv detection was not related to increased mortality, but it is estimated that 66,000-199,000 children could die of rsv-associated pneumonia worldwide every year. our study's power was probably too limited to demonstrate such associations. however, with prevalence of exposure in non-hypoxemic patients ranging from 10% to 50%, with bilateral tests and α < 0.05, study power was ³ 80% to detect or ³ 2.5. similar results have been reported recently in a study from botswana, 22 where researchers noted that rsv pneumonia in children induced more complications and longer duration of hospitalization, but mortality was lower in comparison to other agents. hmpv is recognized as a frequent etiological agent of pneumonia, causing severe disease. 23 here, we determined that detection of this virus was associated with hypoxemic pneumonia in children. these findings might be related to the fact that these two viruses in respiratory samples might be the etiological agents of pneumonia. another hypothesis is that they might be associated with co-infections. however, we did not discern any relationship between infection type (bacterial, viral, or coinfection) and the risk of hypoxemia. the main clinical predictors of hypoxemic pneumonia were lower chest indrawing and cyanosis with lr+ ratios between 2 and 3. if these signs are present, hypoxemia must be suspected in the absence of pulse oximetry, and oxygen therapy should be initiated promptly. nevertheless, we did not discern that one sign had a high lr+ ratio, confirming that pulse oximetry is important for initial evaluation of pneumonia severity and should be implemented more widely in developing countries. 4 lower chest indrawing has been reported to be predictive of hypoxemia in children from nigeria, 8 while breathing rate ³ 60 cycles/minute has been encountered with hypoxemia in children from papua new guinea. 24 other predictors are less known. the designation and validation of a simple, robust score predicting hypoxemia might be useful in resource-limited settings. streptococcus pneumoniae is recognized as the main etiological agent of severe pneumonia and death from pneumonia in children. 25 the diagnosis of pneumococcus pneumonia is, however, difficult at the individual level, because of the low sensitivity of blood culture, particularly in case of previous exposure to antibiotics. in addition, the clinical meaning of pneumococcus detection in nasal samples by molecular testing is difficult to interpret because of the high prevalence of s. pneumoniae respiratory carriage in asymptomatic children. 26 we did not observe associations between pneumococcus detection in nasal samples and disease severity, although pneumococcus-positive blood pcr was linked with greater mortality. molecular s. pneumoniae detection by blood pcr is thus helpful in identifying bacterial pneumonia cases with the poorest prognosis who might need intensive therapies. interpretation of molecular methods, such as pcr, to identify the etiology of pneumonia in children is, however, challenging. respiratory viruses can be identified in asymptomatic children, and secondary bacterial infections in the lungs can easily be missed by these methods. in addition, identification of s. pneumoniae in blood by highly sensitive pcr may detect children with nasopharyngeal colonization only, which could lead to potential misclassification bias, particularly when using results of nasopharyngeal specimens to determine the etiology of pneumonia in children. we must be cautious with results interpretation. moreover, pct was the major biomarker associated with in-hospital death in our cohort. several studies previously found an association between pct and the risk of death or bacterial disease. [27] [28] [29] [30] however, such linkage has rarely been seen in pneumoniainfected children living in developing countries. the main strength of the present study is the prospective data collection on a standardized form at different sites, with advanced molecular diagnosis in all cases, which reinforces internal validity. its main limitations include paucity of information on exposures before hospital admission (i.e., breastfeeding, food intake, or vitamin supplementation). in addition, microbiological diagnosis of pneumonia is difficult because sensitive and specific tests are not routinely available in practice. thus, based on respiratory samples, we were unable to differentiate colonization from infection, particularly by s. pneumoniae. 31 however, analysis of the relationship between results from different samples and severity was contributive: we did not find associations between nasal colonization by pneumococcus and disease severity, hypoxemia, or death. finally, selection bias might have occurred because of patient recruitment in hospital with inclusion of more severe cases or patients with easier access to care. however, we did not discern significant heterogeneity regarding so 2 of infants at admission and inhospital mortality, which suggests that the results might be generalizable to different settings. we must acknowledge that 41% of the study population was enrolled in two sites from india, which might limit external validity. this proportion is, however, in accordance with estimates of global pneumonia incidence and related mortality: india might have accounted for almost 30% of the total number of severe pneumonia cases in children and 40% of the number of deaths worldwide. 25 in conclusion, rsv and hmpv could be major causes of hypoxemia in children with severe pneumonia in developing countries, while s. pneumoniae detection by blood pcr is predictive of high risk of in-hospital mortality. viral etiology might be considered in hypoxemic patients, whereas in very severe pneumonia, which can lead to death, s. pneumoniae may be implicated as the primary cause, even in the absence of hypoxemia at admission. tachypnea and lower chest indrawing could be useful indicators of possible hypoxemia requiring oxygen therapy. pulse oximetry should be included for better diagnosis in developing countries. given these findings, preventive measures, such as increased vaccination coverage of children in developing countries, oxygen therapy of hypoxemic patients, and intensive cardiovascular support, even in non-hypoxemic patients, would reduce the burden of death by pneumonia in children. this protocol was developed on behalf of gabriel network members: http://gabriel. globe-network.org. we especially thank the following gabriel pneumonia experts: ron dagan from the pediatric infectious disease unit saha from the department of microbiology, bangladesh institute of child health, dhaka shishu hospital, bangladesh; and werner albrich from kantonspital aarau ag, bereich medizin, switzerland. we also thank ovid da silva for doing financial support: this study was funded by fondation mérieux and the gabriel network disclaimer: the corresponding author had full access to the data and has the final responsibility to submit the manuscript for publication e-mails: thomas. benet@chu-lyon.fr and philippe.vanhems@chu-lyon.fr. valentina sanchez picot, florence komurian-pradel, hubert endtz, and gláucia paranhos-baccalà, laboratoire des pathogènes emergents, fondation mérieux global, regional, and national causes of child mortality in 2000-13, with projections to inform post-2015 priorities: an updated systematic analysis the prevalence of hypoxaemia among ill children in developing countries: a systematic review oxygen therapy for lower respiratory tract infections in children between 3 months and 15 years of age evaluating the impact of pulse oximetry on childhood pneumonia mortality in resource-poor settings oxygen and pulse oximetry in childhood pneumonia: a survey of healthcare providers in resource-limited settings hypoxaemia in acute respiratory and non-respiratory illnesses in neonates and children in a developing country hypoxaemia in children with severe pneumonia in papua new guinea hypoxaemia in hospitalised under-five nigerian children with pneumonia who/unicef's child health epidemiology reference group (cherg), 2010. causes of deaths in children younger than 5 years in china clinical risk factors of death from pneumonia in children with severe acute malnutrition in an urban critical care ward of bangladesh enhancing research capacities in infectious diseases: the gabriel network, a joint approach to major local health issues in developing countries multicenter case-control study protocol of pneumonia etiology in children: global approach to biological research, infectious diseases and epidemics in low standardized interpretation of paediatric chest radiographs for the diagnosis of pneumonia in epidemiological studies case management of childhood pneumonia in developing countries guidelines for the management of common childhood illnesses. pocket book of hospital care for children refining clinical diagnosis with likelihood ratios epidemiology of hypoxaemia in children with acute lower respiratory infection viral pneumonia differentiation of bacterial and viral pneumonia in children global burden of acute lower respiratory infections due to respiratory syncytial virus in young pneumonia severity in children from developing countries children: a systematic review and meta-analysis viral etiology of severe pneumonia among kenyan infants and children association of respiratory viruses with outcomes of severe childhood pneumonia in botswana human metapneumovirus: review of an important respiratory pathogen hypoxaemia in children with severe pneumonia in papua new guinea global burden of childhood pneumonia and diarrhoea colonisation by streptococcus pneumoniae and staphylococcus aureus in healthy children procalcitonin is useful in identifying bacteraemia among children with pneumonia markers that predict serious bacterial infection in infants under 3 months of age presenting with fever of unknown origin procalcitonin in children admitted to hospital with community acquired pneumonia the diagnostic and prognostic accuracy of five markers of serious bacterial infection in malawian children with signs of severe infection streptococcus pneumoniae colonisation: the key to pneumococcal disease this is an open-access article distributed under the terms of the creative commons attribution license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. key: cord-328661-spxgox52 authors: yu, jianhai; li, xujuan; he, xiaoen; liu, xuling; zhong, zhicheng; xie, qian; zhu, li; jia, fengyun; mao, yingxue; chen, zongqiu; wen, ying; ma, danjuan; yu, linzhong; zhang, bao; zhao, wei; xiao, weiwei title: epidemiological and evolutionary analysis of dengue-1 virus detected in guangdong during 2014: recycling of old and formation of new lineages date: 2019-08-05 journal: am j trop med hyg doi: 10.4269/ajtmh.18-0951 sha: doc_id: 328661 cord_uid: spxgox52 the incidence of dengue is increasing in guangdong, china, with the largest outbreak to date in 2014. widespread awareness of epidemiological and molecular characteristics of the dengue virus (denv) is required. in 2014, we isolated the virus from patients and sequenced its genome. the sequences of denv isolated from guangdong and other countries screened since 2005 were studied to establish molecular evolutionary databases along with epidemiological data to explore its epidemiological, phylogenetic, and molecular characteristics. causes underlying the occurrence of the dengue epidemic included importation and localization of the virus. the number of indigenous cases significantly exceeded that of imported cases. dengue virus 1 is the most important serotype and caused the long-term epidemic locally. based on the data available since 2005, denv1 was divided into three genotypes (i, iv, and v). only genotypes i and v were detected in 2014. in 2014, an epidemic involving old lineages of denv1 genotype v occurred after 2 years of silence. the genotype was previously detected from 2009 to 2011. genotype i, which caused recent epidemics, demonstrated a continuation of new lineages, and a predictive pattern of molecular evolution since 2005 among the four lineages was present. the denv isolated from guangdong was closely related to those causing large-scale epidemics in neighboring countries, suggesting the possibility of its import from these countries. the lack of sufficient epidemiological data and evidence on the local mosquito-borne denv emphasizes the importance of studying the molecular evolutionary features and establishing a well-established phylogenetic tree for dengue prevention and control in guangdong. dengue virus (denv), a mosquito-borne flavivirus, is transmitted primarily by aedes aegypti and aedes albopictus, causing an acute infectious disease named dengue fever (df), 1 which gives rise to public health problems in tropical and subtropical regions worldwide, such as china, singapore, and brazil. [2] [3] [4] with the recent revision of the who dengue classification scheme, dengue patients are classified as having either dengue or severe dengue. the former refers to patients who recover without major complications, whereas the latter points to those who have any of the following conditions: plasma leakage resulting in shock, accumulation of serosal fluid sufficient to cause respiratory distress, or both; severe bleeding; and severe organ impairment. 5 before 1970, only nine countries had experienced severe dengue epidemics, but the disease is now endemic in more than 100 countries. 6 in recent decades, the spreading disease causes rapid upsurge in morbidity. one recent estimate indicates 390 million dengue infections per year, of which 96 million manifest clinically. 7, 8 hence, wasting a lot of health resources and causing the growing global burden of disease, df is regarded as the most widely distributed vector-borne disease with the highest morbidity and great harm. 5 the genome of denv is a single-stranded, positive-sense rna, and its single open reading frame encodes a polyprotein consisting of three structural proteins, which are as follows: the capsid, membrane-associated, and envelope (e) proteins. in addition, seven nonstructural proteins, ns1, ns2a, ns2b, ns3, ns4a, ns4b, and ns5, are also present in its structure. 9 four distinct denv serotypes (denv1, denv2, denv3, and denv4) have been identified, and the extensive diversity within denv1 enables it to recognize different genotypes, such as genotype i (southeast asia and east africa), genotype ii (thailand), genotype iii (malaysia), genotype iv (south pacific), and genotype v (america/africa). 10 dengue genotypes are phylogenetically distinct clusters of viruses, often associated with specific geographical regions, that are linked to epidemics of varying intensities and disease severity. 11, 12 phylogeny is a science that makes use of a set of relationships among groups of genes or organisms and reflects their evolutionary history. the maximum likelihood method is used to describe and analyze biological sequences. 13 alignment of the nucleic acid sequences of the denv followed by its phylogenetic analysis and subsequent generation of phylogenetic trees revealed information regarding its genetic evolution and epidemiology of the disease worldwide. [14] [15] [16] the e protein of the denv is responsible for its tropism and virulence. 17 the gene encoding the e protein has demonstrated its usefulness for decades for the phylogenetic reconstruction of the denv. thus, complete analysis of the coding region can help assign the correct denv genotype and infer the relationships within genotypes and lineages accurately. 18 therefore, the e protein provides adequate resolution to characterize genetic relationship and evolution of the denv. 19 guangdong province is located in the southern mainland of china and experiences tropical and subtropical monsoon climates with a hot and rainy environment that supports breeding of mosquitos, leading to epidemics and very high incidences of df. 20, 21 the first outbreak of dengue in the foshan city of guangdong province occurred in 1978, after which periodic infections and transmission of all four serotypes of dengue have occurred in the past 30 years. 22 the denv found in these areas may have different origins. there were two denv2 outbreaks in foshan in 1993 and 1998, respectively. phylogenetic analysis revealed that isolates from the 1998 epidemic and denv4 from the 1978 epidemic and the 1990 epidemic were closely related to those from the 1993 epidemic in thailand, 1973 epidemic in indonesia, and the 1984 epidemic in the philippines, respectively. since 1990, however, denv1 has been mainly isolated from the infected cases, and its continued existence in guangdong province indicated that endemic infectious agents of dengue may be circulating locally. sequence analysis of the viruses causing the epidemics at different time points revealed that the isolates were closely related to each other, implying that denv1 had probably circulated locally and caused the epidemics. 23 since 2009, all four serotypes were derived from autonomous patients from different outbreak localities in guangdong province. 24 in 2014, a total of 45,236 cases of dengue were reported, which exceeded the total number of cases reported over the previous 10 years. although three serotypes of the denv (denv1, denv2, and denv3) were identified, denv1 was found to be the major causative agent responsible for 98.72% of all 21,928 laboratory-confirmed cases diagnosed during this outbreak. 25 a recent study revealed that the detected sequences belonged to viruses of multiple origins, but the strain isolated in 2014 possibly originated from the isolates of 2013. 26 it can be reasonably speculated that the infectious agents of denv1 from the endemic, which were circulating locally, played a crucial role in causing the dengue epidemic in guangdong province. however, the data mentioned previously are from the studies based on the outbreaks of the particular year and have the limitation of space and time. as a result, comprehensive evaluation of the epidemiological situation and molecular evolution of the viral agents is of significance to warn against their risks and establish preventive and control measures for df. based on denv1 isolated from the outbreak in 2014, we systematically collected the e protein gene from 2005 to 2018 from genbank. with the epidemiological data since 2005 supplied by the guangdong provincial cdc, we studied phylogenetics, molecular characteristics, and epidemiology to strengthen the foundational research of denv1 for the prevention of large-scale dengue epidemics, providing preventive and control measures of df with important evidence. ethics statement. as this research involved human blood, the aims of our study were explained to all the dengue patients involved (all were adults) and all provided written informed consent. the collection methods of clinical samples and epidemiological data were reviewed and approved by the institutional ethics review board of southern medical university and were carried out in accordance with the approved guidelines. samples were selected randomly based on the laboratory diagnosis and clinical signs. sample collection and epidemiological data. dengue virus rna samples (n = 29) were obtained from the guangdong provincial maternal and child care service. all samples were extracted from patients suspected of dengue and were confirmed by reverse transcription-polymerase chain reaction (rt-pcr) using specific primers. the steps included an initial denaturation (94°c, 2 minutes); 35 cycles of denaturation added to make a comprehensive evaluation of the epidemiological situation and molecular evolution in the large 2014 dengue outbreak in guangdong. at the same time, e gene sequences of denv1 since 2005, comprising 187 sequences from guangdong and 1,886 from other countries, were downloaded from genbank. after excluding several sequences with uncertain epidemiological data, representative epidemic strains in every lineage were screened using phylogenetic methods. since then, a molecular evolution database for the denv1 e gene in guangdong and other countries has been established. based on representative strains of the e gene in lineages of the 2014 outbreak, as well as the molecular evolution database, we analyzed molecular characterization and possibility of local circulation for denv1 since 2005 in guangdong. three-dimensional (3d) structure prediction of denv1 protein e and molecular docking with 78-kda glucoseregulated protein (grp78). amino acid (aa) sequences of denv1 protein e were translated by using dnastar. protein 3d structures were simulated by discovery studio 2.5 (ds2.5) (http://accelrys.com/products/collaborativescience/biovia-discovery-studio/) 31 through the homology modeling method based on the modeler program. the optimal protein 3d model was selected by combining the probability density function and discrete optimized protein energy, and the reliability of the model was evaluated using the ramachandran plot and profile-3d. the optimal protein structure model was used to perform protein docking calculations with the grp78 protein (pdb number: 3ldp) using the zdock algorithm in ds2.5, and rdock was used to further optimize the docking configuration to minimize energy. finally, we analyzed aa sites in the binding interface of the optimal docking model. epidemiological findings. since 2005, epidemics of dengue in guangdong have been characterized by periodic outbreaks, the coexistence of importation and localization, and the significantly increased number of indigenous cases compared with that of imported cases ( figure 1a and c). the incidence peaked in 2006 and 2014. especially, during the large outbreak in 2014, a total of 45,236 cases of dengue were reported. in 2017, 1,650 cases were reported, with a continuous rising trend in comparison to 544 cases in 2016 ( figure 1a ). more seriously, as of november 4, 2018, a total of 2,835 cases were reported, an increase of 107.1% over the same period in 2017. among them, 257 cases were imported, which was 50.3% higher than those in the same period in 2017; 2,578 cases were local cases, which was 115.2% higher than those in the same period in 2017. based on the map of guangdong province, we diagrammed the distribution of the cumulative number of cases since 2005. this showed that guangzhou, foshan, zhongshan, and chaozhou were the main epidemic areas. the trends of the epidemics were anastomotic, with periods of fluctuation in guangdong province ( figure 2 ). after 2010, the epidemic trend featured the gradual coexistence of various serotypes, with up to four serotypes emerging in recent years. each denv serotype was identified. dengue virus 1 was the major cause of the outbreak of 2014, and it continued to be detected in guangdong province as the primary serotype, except in 2015. it is worth noting that the proportion of dengue 2 virus in the total cases also gradually increased ( figure 1d ). data of 56 samples were used to analyze the overall epidemiology. all patients who provided the samples had mild dengue and had no travel history (table 2) . generally, the collection date distribution was concentrated in september and october ( figure 1b ). the endemic areas were primarily located in guangzhou, followed by zhongshan and chaozhou, accounting for 85.72% of all samples. heyuan, huizhou, and shanwei were included in the collection areas ( figure 1e ). dengue virus 1 was the major causative agent of this outbreak. the serotype detected through type-specific primers (table 1 ) was denv1, which is similar to the results from guangdong cdc that of 323 df cases involved, 303 cases were due to denv1 infection ( figure 1d , table 2 ). phylogenetic analysis of denv1 outbreak in 2014. all 56 e genes were sequenced to construct phylogenetic trees. the dengue outbreak in 2014 involved asian genotype i and american/african genotype v. phylogenetic analysis showed that most of the sequences were clustered into a unified clade whose distribution differed in each city. among them, 47 e gene sequences were identified as genotype v with 98.9-100% similarity. they clustered into the same clade, which was closely related to the denv1 sequences in malaysia, singapore, and india and evolved into a lineage of genotype v. during 2013-2014, this lineage was involved in a co-epidemic in guangdong province, malaysia, and pakistan, with a continuous high-level local prevalence especially in singapore. significantly, this lineage was detected in six cities in our collection; 13 sequences from shanwei, chaozhou, and huizhou all belonged to genotype v. in addition, another nine sequences of the e gene were identified as genotype i, with two different lineages. lineage i involved eight sequences in our collection and other sequences in singapore and thailand in recent years, with a 99.6-100% similarity. only one sequence in guangzhou contributed to lineage ii, which had been involved in local epidemics in malaysia, singapore, and indonesia for many years ( figure 3 ). phylogenetic analysis of guangdong since 2005. we downloaded e gene sequences since 2005, comprising 187 from guangdong and 1886 from other countries. databases of e gene molecular evolution were created via a screening process based on epidemiologic and phylogenetic methods. among them, 81 strains in guangdong constituted the local database, whereas 403 strains in other countries made up the imported database. based on the evolutionary lineage of denv1 detected in 2014 in guangdong, seven sequences (p1023, p1078, p1258, p1277, p1278, p1306, and p1344) were added to the local we endeavored to clarify the relationship between guangdong and other countries concerning denv outbreaks. for this, we illustrated the origin of molecular evolution lineages and the potential of in situ evolution. representative strains from each lineage were added to the imported database for phylogenetic evolution analysis. in our collection, the molecular evolutionary lineage of the imported database proved to be highly consistent with the local database. the only distinctions were that the evolution time of lineages i and ii changed in genotype i, and denv1 in guangzhou, 2011, formed a clade alone, named clade a ( figure 5a ). genotype i, which has extensively circulated in guangdong and has been detected in each year of the epidemics, displayed some differences with epidemic countries surrounding china. after 2010, lineage i was not found in guangdong. lineage i was very homologous with lineages in singapore, thailand, and sri lanka in the same epidemic years. however, lineage i spread in neighboring countries, such as malaysia, myanmar, and new guinea, which contributed to cyclic epidemics ( figure 5b ). lineage ii in guangdong presented a more complicated epidemic situation in countries around china: for example, lineage ii extensively circulated in vietnam, singapore, thailand, malaysia, and cambodia, with vietnam and cambodia being hot spots. clade a only formed a cluster with denv1 in vietnam ( figure 5b ). recently, lineage iii in guangdong showed a complicated epidemic situation in surrounding countries as well. its sequence was related to sequences in thailand, laos, singapore, malaysia, and elsewhere. in addition, it was related to the co-epidemic in australia that occurred in 2013 and 2014 ( figure 5b ). there were few countries with lineage iv whose epidemic originated from the sequences in 2007. only malaysia, singapore, and especially indonesia shared clustering in denv1 with long-term cyclic epidemics ( figure 5b ). in addition, the occurrence of genotypes iv and v was inconsistent in different years. genotype iv sequences in 2007 shared extensive homology with those in indonesia, whereasthe strains in 2010 were related to those in the philippines with long-term cyclic epidemics ( figure 5c ). genotype v was related to the strains in india and maldives and was divided into clade 2009 and 2014 and clade 2010 and 2011; the sequences shared long-term coepidemics with india and singapore, respectively ( figure 5d ). protein conformational changes caused by mutation of e protein-specific aas between denv1 genotypes i and v. a total of eight uniform aa substitutions in the ectodomain of the e protein were mainly concentrated in domains i (five substitutions) (di) and iii (two substitutions) (diii) between genotypes i and v, whereas domain ii (dii) has only one. among them, two substitutions caused the protein's secondary structure to change from β-sheet to coil, e114 (i → l) in dii and e171 (t → s) in di, whereas only e37 (d → n), e155 (s → t), e161 (t → i), and e171 (t → s) of di were observed on the 3d conformation surface of the protein ( figure 6a and b, table 3 ). meanwhile, e161 (t → i) and e171 (t → s) of di were also found in the binding interface between the e protein of genotypes i and grp78, and other substitutions were not observed in either serotypes ( figure 6e ). in the docking model with the grp78 protein, all three domains of genotype i were involved, whereas genotype v had only di and diii. we found three identical docking sites of diii in the binding interface of the two genotypes: 331a-332p, 359t-360d, and 362e-363k ( figure 6c and d) . the distribution characteristic of denv1 in guangdong was determined from its long-term epidemic history. dengue virus 1 was first detected in zhongshan, guangdong province. since then, denv1 epidemics have occurred sporadically in specific regions over 2-3 years. 32 until 2006, denv1 was the leading serotype, which triggered massive outbreaks in guangzhou. 33 thereafter, denv was circulated continuously across multiple geographies in guangdong, and the isolated strains branched into several stable molecular evolutionary clades. 34, 35 since 2009, it has spread to different provinces of guangzhou, and imported cases were no longer primary. at indicated that although three denv serotypes (denv1, denv2, and denv3) were identified, denv1 was the major causative agent of this outbreak which had circulated continuously in multiple geographies. 41, 42 so far, the results mentioned were consistent with those of our study. the term "lineage" has been used to denote the viruses clustered in clades at a taxonomic level beneath the genotype. the appearance, change, and reappearance of specific lineages are closely linked to the transmission of those viruses. [43] [44] [45] here, 56 strains of viruses differing in their e genes were obtained, of which nine and 47 strains belonged to genotypes i and v, respectively. unlike the strains of genotype i which were linked to two lineages, each strain in genotype v branched into the same clade, forming a stable lineage. however, several lineages from different origins were responsible for the denv outbreak in 2014. moreover, strains belonging to genotype v were detected in six areas evaluated in this study. all 13 strains from shanwei, chaozhou, and huizhou gathered together also belonged to this genotype, illustrating that the same lineage was prevalent in multiple regions. coincidentally, some epidemiological data indicate that denv1 was restricted only to the local epidemics since 1997, 41 a phenomenon which was reported by guo et al. 24 overall, the aforementioned evidence strongly opposes the conventional belief that df in guangdong was completely imported. lee et al. used the term "in situ evolution" to characterize denv in singapore through phylogenetic analysis. results of the analysis explained the correlation between its genetic and evolutionary aspects, suggesting that denv had lurked locally and reappeared after some time. 46 rajarethinam et al. 47 described that the dengue in singapore from 2004 to 2016 demonstrated cyclic epidemic patterns dominated by serotypes 1 and 2. however, this in situ evolution, demonstrated by the step-ladder pattern of branching within each clade over time, has not been observed in guangdong since 2005. another possible transmission and evolution pattern of denv broke out in parts (switch of the lineage), followed by silence (change of the lineage), and was then prevalent on a large scale (continuation of the lineage), achieving continuous evolution of a new lineage and the silent circulation of old lineages. in 2006, when denv1 first became the leading serotype, it was due to a "switch" from the lineages in genotype i, which branched into lineages i-iii at the beginning. 48, 49 from 2007 to 2012, it was observed that the prevalence of denv was low and it frequently interchanged between the three lineages, leading to the occurrence of epidemics caused by denv1 belonging to each lineage. 50, 51 however, since 2012, the severity of dengue is increasing because of a continued epidemic caused by lineage iii. 52 in 2007, a switch in lineage iv, which was similar to the evolutionary process of lineage iii, was observed. the lineage of genotype iv was first formed in 2007, which reappeared in 2010, and was never detected in recent years. 53, 54 it is reported that genotype v had circulated continuously from 2009 to 2011, followed by an intermittent silence and further reappearance in 2014. this type of molecular evolution was similar to the silent circulation prevalent figure 3 . phylogenetic analysis of denv1 detected from the 2014 outbreak in guangdong. fifty-six envelope gene sequences were isolated in our study. these and 19 reference sequences from genbank were used to construct the phylogenetic tree. asian genotype i and american/african genotype v were involved in this outbreak. forty-seven sequences identified as genotype v clustered into the same clade, whereas genotype i was divided into lineages i and ii. thirteen sequences from shanwei (blue circles), chaozhou (green circles), and huizhou (violet circles) all belonged to genotype v. this figure appears in color at www.ajtmh.org. in indonesia, brazil, etc., 11, 55, 56 indicating that the same might happen in guangdong. castonguay-vanier et al. 57 also described the active circulation of denv in laos and the concurrent multiple introductions of new strains from neighboring countries, whereas moore et al. 58 claimed that the cocirculation of denv1-4 in png provides molecular evidence of its endemic transmission. meanwhile, compared with denv2, denv3, and denv4, denv1, which is the leading serotype in guangdong, rarely caused severe df. [59] [60] [61] in addition, because guangdong is the most densely populated province in southern china, which is surrounded by a large number of dengue-endemic countries and has a subtropical climate providing optimal environmental, social, and biological conditions for mosquito breeding and reproduction, a detailed evaluation of the current dengue epidemic is significant. nevertheless, there is still a lack of evidence on the local mosquito vector regarding its harboring of denv during epidemic and nonepidemic periods which can support the evidences of its molecular evolution revealed by the phylogenetic analysis. three genotypes (i, iv, and v) in denv1 from a variety of origins were identified. data obtained from the characterization of local or imported viruses with similarity in lineages found in guangdong demonstrated that these viruses, which were identified based on their molecular evolutionary analysis, were also present in many other countries, particularly those neighboring china. continuous occurrences of the epidemic in indonesia, malaysia, and singapore were consistent with the outbreaks in our country. the epidemiological data demonstrated that the early imported cases in guangdong had high correlation with those found in the neighboring countries, which indicated that these countries might be the source for the migration of denv1 to guangdong. 62, 63 however, specific switches in different genotypes were observed, which are as follows: myanmar, thailand, vietnam, and laos for genotype i; the philippines for genotype iv; and india and maldives for genotype v. the outbreaks of severe epidemics over the years in the neighboring countries and frequent mobility of their populations to china may be able to explain the pandemic pattern of the spread of denv, which began with its spread to guangdong province and then broke out in other parts of the country. a similar conclusion was presented in the study conducted by sun et al. 64 regarding the epidemiological characteristics and genetic diversity of denv in guangdong in 2014. nevertheless, since 2012, with an extremely high incidence of cases, denv spread more rapidly and affected a wider range of populations. the co-epidemic between other countries and guangdong and the dissemination of different genotypes and serotypes in multiple regions might not align totally with the theory that only the imported cases caused the epidemic. lee et al. 46 reported a theory which stated that multiple factors are involved in addition to the ones described in in situ evolution that can explain this phenomenon. however, we still need more evidence to prove the applicability of this hypothesis to the epidemic outbreaks in guangdong province. a prediction of the algorithm of the secondary structure of proteins suggests that it is closely related to the distribution of protein epitopes. the high chemical bond energy of the α-helix and β-sheet enables folding of the protein, making it difficult to bind to the antibody, whereas the β-turn and coil, because of their loose structure, are easily displayed on the surface as antigenic epitopes, facilitating binding of the antibody. [65] [66] [67] only two substitutions causing changes in the secondary structure from the β-sheet to coil were observed in our study, suggesting that its antigenicity was enhanced. however, e171 (t → s) located on the surface is more likely to be associated with protein function than e114 (i → l), which was located inside the protein. although di, the structure found in the central region of the e protein, was not significantly related to the protein function, in our study, majority of the aa substitutions were observed (5/8), and most of them were located on the surface of its 3d structure. even e161 (t → i) and e171 (t → s) were found in the binding interface between genotype i and grp78 but not genotype v, further suggesting that the substitution of e171 (t → s) may be playing a key role in the binding of the e protein of genotypes i and v to the receptor. moraes et al. 68 found that, with changes in the ph, the specific interaction between di and diii of the denv e protein is destroyed, resulting in its conformational change during entry into the cell, whereas nayak et al. 69 also observed the presence of a bundle structure consisting of four polar aa residues at the interface between di and diii, of which his-282 and his317 were unique to denv1, implying that the change of di conformation will also affect the realization of diii function. domain iii has an immunoglobulin-like structure and a functional region where denv binds to a cellular receptor. drumond et al. 70 predicted the structure of the denv1 e protein in brazil and observed that the substitution of e306 (s → f) can reduce its interaction with several residues (ser305, lys307, lys325, val380, val324, ile335, tyr326, and gly381), resulting in a change in the folding of the area, whereas genotypes i and v of denv1 in guangdong have a unified mutation in e324 (i → v). the residue qhg at position e314-e316 of the e protein of denv2 and 3 is highly conserved, and docking by the zdock method showed that it possibly interacts with the membrane receptor protein tim-1 71 , although we found the same region conserved at e316-e318. meanwhile, three conserved regions (331a-332p, 359t-360d, and 362e-363k) were concurrently marked in the binding interface of the two genotypes found in guangdong. chen et al. 72 localized the neutralizing determinants of the inhibitory mabs demonstrating strong effects to a sequence-unique epitope on diii of the denv1 e protein, centered near residues t346 and d360 (346tqngrlitanpivtd360) which were highly conserved among different genotypes of denv1 but different from those of the denv2, denv3, and denv4 serotypes and other flaviviruses. currently, we believe that vaccination and vector control are the fundamental measures to control df. however, research on vaccines and mosquito-control measures have not made significant breakthroughs. we need additional information to understand the epidemic situation of df and evolution of its virus in guangdong to develop effective preventive and control measures. epidemiological analysis reveals information on the cities and months during which df was substantially prevalent and helps to develop mosquito-surveillance and killing strategies. the phylogenetic tree revealed that denv1, which is the main serotype of the virus, has been prevalent in guangdong since a long time. the strains isolated from epidemic cases occurring during the same period are homologous, and genotype i has formed a stable evolutionary lineage in recent years. these results suggested that denv1 may be lurking and circulating in guangdong, although it cannot be stated with certainty. however, it is highly recommended that we detect the denv in local mosquito vectors urgently. at the same time, the phylogenetic tree of the input source suggests the possible countries and regions from which importation of df in guangdong can occur. this information is of great significance for the development of a plan to monitor the departure and entry of populations from regions with a high incidence of dengue. e-mails: chienhai@163.com, murong816@outlook. com, hexiaoen24@163.com, maiblume@163.com, xiyuxie89@126.com, zhuli89@126.com, 13078431254@163.com, milyx117@163.com, harpul@ sina.com, wenying@gdciq.gov.cn, and zhangb@smu.edu.cn. zhicheng zhong, and danjuan ma dengue fever: new paradigms for a changing epidemiology a local outbreak of dengue caused by an imported case in dongguan china dengue viruses in brazil guidelines for diagnosis, treatment, prevention and control world health organization refining the global spatial limits of dengue virus transmission by evidencebased consensus the global distribution and burden of dengue flavivirus genome organization, expression, and replication the origin, emergence and evolutionary genetics of dengue virus insights of the genetic diversity of denv-1 detected in brazil in 25 years: analysis of the envelope domain iii allows lineages characterization microevolution 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