key: cord-274477-jkjyh4cs authors: zhang, zhiruo; liu, shelan; xiang, mi; li, shijian; zhao, dahai; huang, chaolin; chen, saijuan title: protecting healthcare personnel from 2019-ncov infection risks: lessons and suggestions date: 2020-03-23 journal: front med doi: 10.1007/s11684-020-0765-x sha: doc_id: 274477 cord_uid: jkjyh4cs the outbreak of a novel coronavirus disease (covid-19, caused by the 2019-ncov infection) in december 2019 is one of the most severe public health emergencies since the founding of people’s republic of china in 1949. healthcare personnel (hcp) nationwide are facing heavy workloads and high risk of infection, especially those who care for patients at the epicenter of the outbreak, hubei province. sadly, as of february 20, 2020, over two thousand covid-19 cases are confirmed among hcp from 476 hospitals nationwide, with nearly 90% of them from hubei province. based on literature search and interviews with some hcp working at wuhan, capital city of hubei, we have summarized some of the effective measures taken to reduce infection among hcp, and also made suggestions for improving occupational safety during an infectious disease outbreak. the experience and lessons learned should be a valuable asset for international health community to contain the ongoing covid-19 epidemic around the world. the local outbreak of novel coronavirus epidemic (covid-19, caused by the 2019-ncov infection) in wuhan, hubei province of china, last december has rapidly spread nationwide with a cumulative number of 80 026 laboratory-confirmed cases and 2912 deaths as of march 1, 2020 [1] . as the epicenter of the outbreak, hubei province has been hit hardest, accounting for over 83% of all the confirmed cases in china [1] . more than 40 000 healthcare personnel (hcp) from around the country rushed to hubei, working around the clock with local hcp in the public health intervention, diagnosis and treatment, patient care, field investigation, samples testing and sterilization [2] . sadly, as of february 20, 2020, 2055 laboratory-confirmed cases were reported among hcp from 476 hospitals across china. the majority of hcp cases (88%) were reported from hubei [2] (table 1) . so far, no super spreader has been identified among the hcp infections. such a high number of infections among hcp in this ongoing epidemic is very alarming, testifying the magnitude of the epidemic, the lack of understanding of the novel virus, and the need for improvement in the medical system. little knowledge about the novel virus led to a surge of hcp infection in the first month after the outbreak. in addition, the sudden tide of patients in january 2020 added unprecedented stress on the municipal healthcare system of wuhan. healthcare staff from different medical departments in the city had to hastily take actions in a short period of time. many did not have the required experience, knowledge, and preparedness for dealing with infectious diseases, such as proper sequence of use, replacement, and disposal of personal protective equipment (ppe). some healthcare workers had to work ten or more hours daily with no break time due to a high volume of patients and severe staff shortage. stress and extreme fatigue could further challenge the immune system and increase the susceptibility to 2019-ncov among hcp (fig. 1) . the rapid expansion of the epidemic also caused a severe shortage of critical ppe. as many mass production of medical supplies was halted when workers left for family reunification at the chinese new year (january 25 , 2020), the healthcare system was completely caught off guard, and quickly exhausted the emergency ppe reserves. some hospitals in hubei had to reduce the replacement of critical ppe. the grim shortage of protective equipment greatly increased the risk of hcp infections at the onset of the outbreak, exacerbating the spread of diseases among visitors, staff, and patients. a healthy and effective team of hcp is crucial to successfully preventing the on-going epidemic from further expansion. the high infection toll underscores the necessity and urgency of protecting hcp from covid-19. it is laudable that the central government has attached great importance to protecting the health of hcp and has taken a number of immediate actions [3] , such as improved guidance on the proper use of ppe, strengthened logistics and medical supplies, and enhanced disinfection at the hotels where hcp stay. in addition, an emergency surveillance system is now in place to monitor all exposed hcp, contributing to prompt detection, effective triage, and isolation of infected hcp. a special medical expert group is making every effort to diagnose and treat medical staff of suspected and confirmed infection. in addition, a special health and life insurance fund is set up for all hcp working in the frontline at both national and provincial levels [4] . all of these are conducive to ensuring the confidence and efficiency of the hcp, but more needs to be done for further protection of their occupational health in the long run. the covid-19 outbreak calls for the establishment of exposure risk assessment and management system in healthcare settings. public health schools should be given more support within medical education system and all medical students should receive regular theoretical courses and practical training of public health, including the knowledge and skills to deal with large scale health emergencies, such as covid-19. standard guidelines and procedures should be in place to detect infectious diseases at an early stage, to timely announce the pathogens, transmission paths, diagnosis, and treatment among hcp. furthermore, improvement in professional development as essential part of continuous medical education at all public health and medical institutions is another critical step toward reducing infection rate among healthcare workers. hcp, regardless of the disciplines they are in, should have routine emergency drills for infectious diseases, receive professional development periodically in the protection from occupational hazards. especially, medical staff related to the handling of infectious diseases should be well trained to properly use ppes, and continuing education certificate can be mandatory for key hcp or staff members in all medical institutions. in addition, easy access to mental health service for hcp should be available throughout their career journey, especially during the time of crisis when they need for anxiety and stress relief. with the stabilizing of epidemics and measures taken by the decision-makers, the shortage of ppe in china was significantly attenuated by mid-february [5] . yet, the covid-19 outbreak alerts us that a carefully planned stockpile of ppe and other essentials is key to effective infectious disease preparedness and to the optimal function of hcp [6] . an epidemic can affect a broad population, hence the availability and appropriate use of ppe, such as n95 respirators, face masks, gowns, and gloves, are crucial to protecting the health of hcp [7] . while it is very difficult to predict an outbreak of widespread epidemic, all the healthcare facilities should stockpile a certain amount of critical ppe to ensure an adequate supply at the onset. in addition, it is also important to set up a centralized and coordinated network of emergent ppe supply among central and local governments, healthcare facilities, and medical equipment, so as to meet the demand for consumable and durable supplies when a wide spread epidemic lasts long. the ratio of confirmed hcp infection to patients is on the decrease, from 5.72% (january 11-20, 2020) to 2.68% (february 1-11, 2020), thanks to the increased supply of critical ppe, enhanced vigilance, and accumulated experienced among hcp [8] . it is hard to predict when and where an epidemic, like the covid-19, will occur in the world as disease knows no national boundaries. therefore, the compelling lessons learned in the fight against coronavirus in china must be remembered so that better public health emergency response preparedness mechanisms can be established not only in china but all over the world. national health commission of the people's republic of china. the latest situation of novel coronavirus pneumonia who. report of the who-china joint mission on coronavirus disease national health commission of the people's republic of china. notification for further strengthening the protection of health personnel during 2019-ncov outbreak the state council. press conference of the joint prevention and control mechanism of the state council information office of hubei provincial people's government. press conference on prevention and control of new coronavirus infected pneumonia protecting health-care workers from subclinical coronavirus infection association between 2019-ncov transmission and n95 respirator use novel coronavirus pneumonia emergency response epidemiology team. the epidemiological characteristics of an outbreak of 2019 novel coronavirus diseases (covid-19) in china we would like to extend sincere gratitude to mengyun luo, yue fang, kun qian, xueyuan li, jiawei xu, and jiahui li who collected data. zhiruo zhang, shelan liu, mi xiang, shijian li, dahai zhao, chaolin huang, and saijuan chen declare no conflicts of interest. this manuscript is a commentary and does not involve a research protocol requiring approval by the relevant institutional review board or ethics committee. key: cord-344480-6tcush4w authors: zhou, guangbiao; chen, saijuan; chen, zhu title: back to the spring of wuhan: facts and hope of covid-19 outbreak date: 2020-03-14 journal: front med doi: 10.1007/s11684-020-0758-9 sha: doc_id: 344480 cord_uid: 6tcush4w nan since december 2019, an atypical pneumonia has been spreading from wuhan, a beautiful city located at the center of china (fig. 1) , to the whole country. this disease originated from the huanan seafood wholesale market that was closed on january 1, 2020. very rapidly, a novel coronavirus was isolated and named first the 2019 novel coronavirus (2019-ncov) [1] and subsequently severe acute respiratory syndrome coronavirus 2 (sars-cov-2) [2] , and is suggested to be named as human coronavirus 2019 (hcov-19) [3] . meanwhile, the disease is chronologically called pneumonia of unknown origin, novel coronavirus pneumonia (ncp), and coronavirus disease-2019 (covid-19) (fig. 2 ). superspreading events of this virus have also taken place on the diamond princess cruise off the coast of yokohama, japan [4] . the chinese central [5] and local governments [6] have been endeavoring unprecedented efforts to constrain the outbreak, and more than 30 000 medical professionals especially doctors and nurses outside wuhan have been joining the local ones to handle this emergency. in early december 2019, the first case was reported; by december 31, 104 cases were diagnosed [7] . the disease took 30 days to spread to all the 34 provinces/regions (including hong kong, macao, and taiwan) of china. the number of confirmed cases of covid-19 rose to 11 791 in chinese mainland as of january 31, including 259 deaths. the confirmed cases increased rapidly, and by february 21, 75 567 confirmed cases, 5206 suspected cases, and 2239 deaths, have been reported in china. the disease also spread to 26 countries, with 1151 confirmed cases and 8 deaths reported by february 21 [8] . by february 21, severe cases in wuhan, other regions of china, and the whole country were 9628 (21.2%), 2005 (6.6%), and 11633 (15.4%), respectively ( table 1 ). the reproduction number (r 0 ) is between 2.2 and 4.8 [9, 10] . the commonest symptoms of the covid-19 include fever, dry cough, fatigue, sputum, and shortness of breath, with a small proportion of patients (889/72 314, 1.2%) as asymptomatic and probably spreaders [7] . cytokine storm that is induced by virus particles, is associated with disease severity [11, 12] . a first histological examination of covid-19 patient showed diffuse alveolar damage with cellular fibromyxoid exudates, desquamation of pneumocytes and hyaline membrane formation, pulmonary edema with hyaline membrane formation, interstitial mononuclear inflammatory infiltrates, and multinucleated syncytial cells with atypical enlarged pneumocytes in the lungs [13] . these results shed insights into the pathogenesis of covid-19 and may help design therapeutic strategies against the disease. a previously unknown betacoronavirus was discovered from bronchoalveolar-lavage fluid samples of the patients, which is the seventh member of the family of coronaviruses that infect humans [1, 14] . the virus was most closely related to a group of sars-like coronaviruses (genus betacoronavirus, subgenus sarbecovirus) previously sampled from bats in china [14, 15] , and a virus with 99% sequence homology to the receptor-binding domain (rbd) and 90% to the sars-cov-2 genome was also found in malayan pangolins in southern china [16] . however, higher viral loads were detected in the nose than in the throat of symptomatic and asymptomatic covid-19, a pattern resembling that of influenza but not sars-cov [17] . human angiotensin-converting enzyme 2 (ace2) has been shown to be the putative receptor for the virus to enter into host cells [15, 18] , and biophysical and structural evidence shows that the sars-cov-2 spike glycoprotein binds ace2 with high affinity [19] . ace2 can also bind spike protein through association with b0at1 [20] . ace2 locates on the organs such as lung, heart, esophagus, kidney, bladder and ileum, and in particular on the cell types such as type ii alveolar cells, myocardial cells, proximal tubule cells of kidney, ileum and esophagus epithelial cells, and bladder urothelial cells, providing clues for further investigating the pathogenesis covid-19 [21] . so far, neither drug nor vaccine has been approved to treat the novel covid-19. while supportive treatment regimens including oxygen therapy are widely used, antivirus (oseltamivir) and anti-hiv (lopinavir/ritonavir) drugs are also applied in treating covid-19. some emerging therapeutics are being tested in clinical trials. for example, virally targeted agents, approved nucleoside analogs (favipiravir and ribavirin) and experimental nucleoside analogs (remdesivir and galidesivir), may have potentials against sars-cov-2 [22] . remdesivir has been shown to be able to block virus infection [23] and exert therapeutic efficacy in the first covid-19 case in the united states [24] . two phase iii trials have been initiated to evaluate remdesivir in covid-19. anti-malaria drug chloroquine shows activity in blocking sars-cov-2 infection [23] and is being evaluated in an open-label trial [22] . convalescent patient plasma that contain anti-sars-cov-2 antibody holds promise to beat this disease [25] . ace2 and spike protein represent two novel therapeutic targets for the disease [26] . an ongoing study using tocilizumab, a specific monoclonal antibody antagonist of interleukin-6 receptor (il-6r) which proved to be effective in alleviating cytokine release syndrome, showed preliminary positive effects [27] . traditional chinese medicine (tcm) plays an active role in fighting infectious disease, exemplified by ancient formula maxingshigan-yinqiaosan in the treatment of h1n1 influenza [28] . in combination with western medicine [29] or used alone, tcm is widely applied or being evaluated in clinical trials to treat covid-19. two and a half months have witnessed the great efforts that china has been endeavoring to control the outbreak of the covid-19. these include lockdown of wuhan and related cities, control of population mobility, and input of huge resource to the regions. china races against the clock to build virus hospitals, including huoshenshan hospital with 1000 beds built in 9 days and leishenshan hospital with 1500 beds completed in 15 days. more than 15 fangcang hospitals (with large open space and necessary anti-infection conditions) have been built with a capacity of more than 20 000 beds for the isolation and treatment of the patients. the updated genome sequence of sars-cov-2 has been shared to the public, clinical trials are undergoing, and scientists from china and overseas are working together to combat this public health emergency. with great contributions of our domestic selfless medical professionals and supports from public health scientists around the world [30] , by using smart but firm public health measures preventing spreadout of the virus and more specific combinatorial therapeutic strategies, such as the anti-viral convalescent plasma and drugs against cytokine storm for severe cases, as well as effective tcm drugs and technologies, we believe that the mankind will win this battle, and wuhan will be back to the spring. guangbiao zhou, saijuan chen, and zhu chen declare no conflict of interests. this manuscript does not involve a research protocol requiring approval by the relevant institutional review board or ethics committee. china novel coronavirus investigating and research team. a novel coronavirus from patients with pneumonia in china severe acute respiratory syndromerelated coronavirus: the species and its viruses -a statement of the coronavirus study group a distinct name is needed for the new coronavirus epidemiological research priorities for public health control of the ongoing global novel coronavirus (2019-ncov) outbreak a novel coronavirus outbreak of global health concern china's local governments are combating covid-19 with unprecedented responses -from a wenzhou governance perspective novel coronavirus pneumonia emergency response epidemiology team. the epidemiological characteristics of an outbreak of 2019 novel coronavirus diseases (covid-19) in china databased analysis, modelling and forecasting of the novel coronavirus (2019-ncov) outbreak early transmission dynamics in wuhan, china, of novel coronavirus-infected pneumonia clinical features of patients infected with 2019 novel coronavirus in wuhan, china epidemiological and clinical characteristics of 99 cases of 2019 novel coronavirus pneumonia in wuhan, china: a descriptive study pathological findings of covid-19 associated with acute respiratory distress syndrome genomic characterisation and epidemiology of 2019 novel coronavirus: implications for virus origins and receptor binding a pneumonia outbreak associated with a new coronavirus of probable bat origin identification of 2019-ncov related coronaviruses in malayan pangolins in southern china sars-cov-2 viral load in upper respiratory specimens of infected patients functional assessment of cell entry and receptor usage for lineage b β-coronaviruses, including 2019-ncov cryo-em structure of the 2019-ncov spike in the prefusion conformation structure of dimeric fulllength human ace2 in complex with b 0 at1 the single-cell rna-seq data analysis on the receptor ace2 expression reveals the potential risk of different human organs vulnerable to wuhan 2019-ncov infection therapeutic options for the 2019 novel coronavirus (2019-ncov) remdesivir and chloroquine effectively inhibit the recently emerged novel coronavirus (2019-ncov) in vitro washington state 2019-ncov case investigation team. first case of 2019 novel coronavirus in the united states 32 recovered patients donate plasma to others with coronavirus infection therapeutic strategies in an outbreak scenario to treat the novel coronavirus originating in wuhan, china. (version 2; peer review: 2 approved scientists target major cause of critically ill virus deaths. china daily oseltamivir compared with the chinese traditional therapy maxingshigan-yinqiaosan in the treatment of h1n1 influenza: a randomized trial combination of western medicine and chinese patent medicine in treating a family case of covid-19 in wuhan statement in support of the scientists, public health professionals, and medical professionals of china combatting covid-19 key: cord-308673-dkmpayk4 authors: ni, li; zhou, ling; zhou, min; zhao, jianping; wang, dao wen title: combination of western medicine and chinese traditional patent medicine in treating a family case of covid-19 in wuhan date: 2020-03-13 journal: front med doi: 10.1007/s11684-020-0757-x sha: doc_id: 308673 cord_uid: dkmpayk4 in december 2019, an outbreak of novel coronavirus (2019-ncov) occurred in wuhan, hubei province, china. by february 14, 2020, it has led to 66 492 confirmed patients in china and high mortality up to ∼2.96% (1123/37 914) in wuhan. here we report the first family case of coronavirus disease 2019 (covid-19) confirmed in wuhan and treated using the combination of western medicine and chinese traditional patent medicine shuanghuanglian oral liquid (shl). this report describes the identification, diagnosis, clinical course, and management of three cases from a family, suggests the expected therapeutic effects of shl on covid-19, and warrants further clinical trials. since december 31, 2019, a cluster of patients with pneumonia of unknown cause has been reported in wuhan, china. this special pneumonia was associated with a novel coronavirus, 2019-ncov, named initially by the world health organization (who) in january 2020 [1] . an outbreak of 2019-ncov pneumonia (officially and internationally named as covid-19 on february 11, 2020 by the who) occurred and spread to the entire china and multiple countries worldwide [2] . by february 14, the number of patients confirmed with covid-19 reached 66 492, 8969 suspected patients, and 1523 deaths were identified in china [3] . by february 14, the number of confirmed patients and deaths were 37 914 and 1123, respectively, and the mortality of covid-19 was 2.96% in wuhan city, hubei province, which is considered as the origin of first cluster of patients [3] . the confusing and difficult thing for physicians is that they do not have specific drugs to either treat or prevent the aggravation and serious complications of covid-19 for such patients [4] . thus, chinese herbs attract our attention, and investigations for different clinical trials are in progress. the present report described a family case, including three cases who received western medicine and chinese traditional patent medicine shuanghuanglian oral liquid (shl) treatments and achieved rapid recovery. the family case includes parents and a daughter. the family lives in an apartment 3 km from the huanan seafood wholesale market in wuhan, hubei province. these cases are a typical familial cluster where all of them had covid-19 [5] . all the patients were informed about the clinical trial (chictr2000029605) and signed informed consent. case 1 is a 51-year-old female. on january 17, 2020, the patient presented a feeling of general malaise and coldness. on january 19, she had fever with body temperature of 37.3°c and experienced diarrhea and vomiting. she started to receive intravenous injection of cefotaxime in community clinic and took oral jinyebaidu granules (another chinese traditional patent medicine) and oseltamivir (75 mg, twice a day) for 4 days. she had persistent severe fever with body temperature from 37.6°c to 38.3°c for four days. on january 23, her chest computed tomography (ct) scan showed multiple patchy ground glass opacity and consolidation shadow in bilateral lung and subpleural regions (fig. 1a) . she was highly suspected with covid-19. oral moxifloxacin and arbidol were prescribed, and she continued to take jinyebaidu granules and oseltamivir. on january 24, her body temperature reached 39.9°c and experienced severe fatigue, diarrhea, and breathlessness despite continuous treatments of the above drugs. she felt better after intravenous injection of immunoglobin (ivig, 5 g per day) and dexamethasone (5 mg, once to twice a day). however, the patient experienced recurrent fever (body temperature of 39°c) and breathlessness, and her blood oxygen saturation fluctuated from 90% to 95% at the night of january 27. on january 28, the second chest ct scan indicated that her pneumonia aggravated for the past 5 days (fig. 1b) . a nasopharyngeal swab specimen was obtained and sent for detection of 2019-ncov. although the patient's 2019-ncov test was negative (table 1) , she was diagnosed of covid-19 in accordance with her symptoms and chest ct display. on the same day, she was confined into an isolation ward and started to take oral shl (twice a day, 20 ml once). on the next day, shl administration increased to three times a day (20 ml once) without using any other drugs. from january 29 to 31, the patient's symptoms resolved with body temperature decreasing from 37.3°c to 36.5°c and without vomiting and diarrhea. the patient gradually felt strong except for slight cough. after february 6, the patient's symptoms disappeared, and her third chest ct scan (fig. 1c ) showed significant absorption of bilateral ground glass opacity compared with the previous ones. the association of her symptoms with treatments is shown in fig. 2 , and the clinical laboratory results are shown in table 2 . cases 2 and 3 became simultaneously ill 9 days after they had close contacts with case 1 [5] . case 2 is a 27-year-old female nurse at the clinical trial center of division of cardiology of tongji hospital, wuhan, daughter of case 1, who took care of her mother. on january 26, the patient presented mild weakness, diarrhea, and low fever, and she started to take oral jinyebaidu granules, oseltamivir, moxifloxacin, and arbidol. on the next day, all her symptoms aggravated with body temperature reaching to 38.3°c, frequent vomiting, and diarrhea for 5 times a day. at the night of january 27, her body temperature reached 39.5°c with chest tightness and shortness of breath. although she took nonsteroidal anti-inflammatory drug (loxoprofen), her fever remained high (39°c). on january 28, her chest ct scan showed consolidation shadow in the left lung, and her 2019-ncov test showed negative using nasopharyngeal swab specimen. considering her contact history with her mother, she was also confined into an isolation ward and started to take shl 20 ml once for three times a day without taking other drugs. her body temperature ranged from 37.5°c to 38.5°c during january 29 to 31 and decreased from 37.5°c to 36.5°c on february 1. all other symptoms resolved on february 2 with recovered appetite and spirit. two repeated 2019-ncov tests were negative (table 1) . on february 6, her second chest ct scan showed the absorption of the left lung shadow (fig. 3) . after her disease symptoms disappeared, the oral dose of shl reduced to 10 ml once for three times a day. the patient's clinical laboratory results are shown in table 3 . case 3 is a 53-year-old male, husband of case 1 and father of case 2, who presented mild diarrhea, vomiting, and fever on january 26. on january 28, the patient was diagnosed of covid-19 with positive 2019-ncov test using nasopharyngeal swab specimen (table 1 ) and chest ct scan showing patchy ground glass opacity in the right lower lung subpleural fields (fig. 4) . he had no fever, cough, and breathlessness. thus, the patient started to isolate himself at home and took shl (20 ml once, three times a day), moxifloxacin, and arbidol on january 28. since february 2, all his symptoms resolved with exception of light nausea, and he continued taking shl with other drugs. on february 4, he felt slightly weak. on february 7, the patient fully recovered, and the repeated 2019-ncov rna test was negative (table 1 ). this report described the first typical family case of covid-19 treated using the chinese traditional patent medicine shl because of poor response to other treatments. the three patients were a close family, and the mother was the first victim with typical symptoms of severe viral pneumonia and confirmed as covid-19 in accordance with the symptoms and chest ct scan. cases 2 and 3 had close contact history with case 1, and 9 days later they became ill with positive diagnosis of covid-19. all their symptoms resolved after using the chinese traditional patent medicine shl and rapidly recovered without obvious adverse effects when the patients showed no response and their symptoms continued to aggravate after other treatments, including ivig (5 g per day) and dexamethasone, antibiotics, and antivirus compounds. shl, a chinese traditional patent medicine containing extracts of three chinese herbs, namely, honeysuckle, forsythia, and scutellaria baicalensis, is usually used to treat cold, sore throat, and cough with fever. shl has been used in clinical practice for a long time because of its affordable cost and no serious adverse reaction. recent news from preliminary study findings indicated that shl can inhibit 2019-ncov (http://www.cas.cn/yw/202001/ t20200131_4733137.shtml, accessed on january 31, 2020). considering that no specific drugs are recommended to treat covid-19, we started our clinical trial (chictr2000029605) to investigate whether shl can treat this disease and the family case is a part of the clinical trial. these cases suggest that shl might be effective for covid-19 although subsequent clinical trials are needed. in this family case report, we described two patients who had poor response to other treatments but responded well to shl therapy. case 3 showed positive therapeutic effect although he simultaneously received arbidol. early treatments may contribute to patients' outcome and several errors, including taking antibiotics, and combination of two antivirus drugs should be avoided. this report suggests that shl treatment might be effective for covid-19 and warrants subsequent clinical trials to obtain sufficient evidence for clinical recommendation. china novel coronavirus investigating and research team. a novel coronavirus from patients with pneumonia in china early transmission dynamics in wuhan, china, of novel coronavirus-infected pneumonia national health commission of the people's republic of china pillai sk; washington state 2019-ncov case investigation team. first case of 2019 novel coronavirus in the united states a familial cluster of pneumonia associated with the 2019 novel coronavirus indicating person-to-person transmission: a study of a family cluster li ni, ling zhou, min zhou, jianping zhao, and dao wen wang declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article. informed consent was obtained from all the patients in which their identifying information are included in this article. other ethical board approval is not applicable in this case report. key: cord-312361-ermbijfq authors: gong, fanghua; xiong, yong; xiao, jian; lin, li; liu, xiaodong; wang, dezhong; li, xiaokun title: china’s local governments are combating covid-19 with unprecedented responses — from a wenzhou governance perspective date: 2020-03-12 journal: front med doi: 10.1007/s11684-020-0755-z sha: doc_id: 312361 cord_uid: ermbijfq the covid-19 caused by a novel strain of coronavirus has been spreading rapidly since its onset in wuhan, the capital city of central china’s hubei province, in december 2019. it is highly communicable through human-to-human transmission. china has been making unprecedented efforts in treating the confirmed cases, identifying and isolating their close contacts and suspected cases to control the source of infection and cut the route of transmission. china’s devotion in handling this epidemic has effectively and efficiently curbed communication domestically and across the border. representative measures adopted by wenzhou, the worst hit city out of hubei province, are examined to elucidate those massive undertakings with the aim of enhancing international understanding and building global rapport in fighting this evolving epidemic situation. a novel strain of coronavirus has caused a pneumonia outbreak in the central chinese city of wuhan since december 2019 and has been quickly spreading to other parts of china. sporadic imported cases are also identified in other countries and regions of the world because of cross-border mobility of travelers originating or who had contact with people from wuhan. named as 2019 novel coronavirus (2019-ncov) initially by the world health organization (who) in january 2020, it was caused by a previously unknown pathogen and has escalated into an unprecedented outbreak. the pneumonia caused by this viral strain was named "novel coronavirus pneumonia" by china's national health commission on february 7, 2020 [1] and later officially named internationally as covid-19 on february 11, 2020 by the who [2] . as commented by the who director-general dr. tedros adhanom ghebreyesus, the outbreak has been met by an unprecedented response [3] from the chinese government and grassroot communities. a total of 44 730 confirmed and 16 067 suspected covid-19 cases have been reported in china with 4742 cured and 1114 deaths by midnight of february 11, 2020 [4] and an additional 441 confirmed cases overseas with 1 death in the philippines as of 17:30 on february 12, 2020 [5] . the trends in confirmed cases, suspected cases, and deaths that resulted from this disease are shown in figs. 1-3, respectively. the world is in a race against time to identify the root cause and find a vaccine for this new virus, and china is taking the lead in this combat as the originating and worst hit country. the world may be well informed of the amazing speed of the chinese in constructing and manning two purposebuilt hospitals in wuhan, the epicenter of the virus outbreak. the two dedicated hospitals, huoshenshan (fire god mountain) hospital with 1000 beds and leishenshan (thunder god mountain) hospital with 1600 beds, only took 9 and 14 days to complete from scratch, respectively, and were staffed with medics from hospitals affiliated to the people's liberation army (pla) joint logistic support force and medical universities of the army, navy, and air force of the pla [6] . the two makeshift hospitals are playing major roles in improving the cure rate and decreasing the death rate of infected patients and help china and the world win the combat of prevention against and control of this growing epidemic situation. although this event is extremely impressive, the strategy and measures taken by wenzhou merit special attention and even commendation. wenzhou, a city 900 km away from wuhan and well-known for its private economy and entrepreneurial spirits of its people, is again catching the whole nation's attention as the worst hit place second only to hubei province by covid-19. statistics show that almost 180 thousand wenzhou people were in wuhan for business or study in 2019, and over 78.8 thousand of them returned to wenzhou for the traditional chinese spring festival at a time when the virus was announced and the city of wuhan was placed into full lockdown. consequently, the effective control of the source of the virus and efficient prevention of internal communication became a paramount challenge for the city. in response to the chinese central government's mandate, wenzhou's municipal government convened emergency meetings (mostly through video conferences to avoid mobility and contact) in the last weeks to deliberate on coping policies, preventive measures, and implementation plans with objective investigation and across-the-board examination of the rapidly worsening epidemic reality. on january 29, the municipal government convened a video conference pledging to curb the deterioration of the epidemic situation and reach the inflection point by making concerted efforts and working vigorously for 10 days [7] until the lantern festival, a day when the traditional chinese spring festival officially ends. in the evening of january 31, the municipal government rolled out "twenty-five emergency measures" that cover the control measures for human mobility, traffic, and daily necessity outlets and the prevention and control measures for post-holiday work and production resumption. those measures include: postponing resumption of production for various enterprises until february 17 except for those closely related to public utilities, everyday life supply, epidemic prevention and control, or concerning national interest and people's livelihood; postponing resumption of work for public institutions until february 9 except for those involved in epidemic prevention and control; postponing resumption of school until march 1 for all levels and categories of educational institutions and organizations; temporary suspension of all city bus and metro lines; strict checking arrangement for all private houses and public places; requirement of wearing protective face masks when one has to go out; closure of badly affected villages, communities, and residential areas; cancellation of all public fairs and large gatherings and closing of parks, libraries, museums, cinemas, swimming pools, large department stores, and commercial complexes; closing of central air-conditioning at densely populated areas; reward for reporting to municipal authorities of hiding patients or suspected individuals secretly returning from hubei. since january 24, nine circulars had been issued by wenzhou municipal leading group for covid-19 prevention and control, which was headed by the mayor. the latest circular issued on february 9 announced measures with even elevated rigidness to curb possible covid-19 transmission at famer markets and supermarkets. measures from previous circulars include: body temperature monitoring, daily sterilization, and improving ventilation at high-density public places such as airport, train and bus stations, passenger piers, and expressway entrances and exits (circular 1, january 24); 24/7 whistleblowing hotline for reporting to municipal epidemic prevention and control office of recent returnees from hubei province who were not quarantined for medical observation (circular 2, january 25); strengthening of management and control of 14-day home or concentrated quarantine (at designated hotels by arranging government purchase of services with medical staff and resources deployed) for source of risks including all returnees from hubei province, their close contacts, and patients with fever, with one-to-one follow-up and tracking scheme in place (circular 3, january 27) (see fig. 4 for total number of close contacts subsequently traced); heightened control of household mobility by allowing only one family member to go out for daily necessity shopping every other day (circular 6, february 1); and temporarily closing 46 expressway entrances and exits within its administration, keeping only 9 with one for each district or township under its jurisdiction (circular 7, february 2). updated measures followed the above policies considering the development of the actual circumstances. the battery of rigorous measures had been appropriate given the gravity of the situation and proved timely and conducive to the effective prevention and control of this epidemic in wenzhou. there had been an evident decrease in newly confirmed cases (illustrated in fig. 5 in dotted line) . thus, the total number of confirmed cases is leveling out recently. wenzhou people are known far and wide as people with entrepreneurship, determination, and collective spirits. with this conjuncture, wenzhou is demonstrating outstanding organizing and coping capacities in the face of unexpected crisis and showing the rest of the country that a war of defense against covid-19 had been waged. furthermore, the city is on the right track to containment and recovery. similar mass mobilization and harsh measures are under way in severely affected areas in china to ensure the health and safety of people, both chinese and internationals, and the effective prevention and control of this epidemic for the shared well-being of mankind. like any other unexpected outbreak, covid-19 caused fear, anxiety, and panic locally, nationally, and globally. the good news is that china's response on how to combat and stop the spread of the coronavirus had been undoubtedly swift and amazing as who director-general dr. tedros adhanom ghebreyesus noted: "i will praise china again and again because its actions actually helped in reducing the spread of the novel coronavirus to other countries … in many ways, china is actually setting a new standard for outbreak response. it's not an exaggeration." [3] "the nation is currently experiencing a peak period of confirmed cases. with prevention and control measures in place, the number of future infections will be significantly reduced," said li lanjuan, a renowned chinese epidemiologist and academician of the chinese academy of engineering. as the virus is highly contagious, the entire world is looking to china's prevention and control efforts. wenzhou has been taking timely, rigorous, and systematic measures in fighting the spread of this epidemic within its jurisdiction. the unprecedented responses prove effective and efficient in curbing the source of infection and cutting the route of transmission and are setting new standards for other cities to overcome the challenge and difficulties together. chinese president xi jinping has stressed releasing information in a timely manner and deepening international cooperation to fight this crisis. when meeting with who director-general dr. ghebreyesus, xi stressed that china is ready to work with the who and the international community to safeguard regional and global public health security [8] . in a deeply interwoven world, the only way we will defeat this outbreak is for all countries to work together in a spirit of solidarity and cooperation. this is the time for facts, not fear. this is the time for science, not rumors. we, as a global community with a shared future, are all in this combat together. together, we must and we will win this fight. fanghua gong, yong xiong, jian xiao, li lin, xiaodong liu, dezhong wang, and xiaokun li declare no conflicts of interest. this article does not involve a research protocol requiring approval by the relevant institutional review board or ethics committee. national health commission of the people's republic of china. notice by national health commission on the provisional designation of novel coronavirus pneumonia world health organization. novel coronavirus (2019-ncov) situation report -22. 2020 world health organization. who director-general's statement on ihr emergency committee on novel coronavirus national health commission of the people's republic of china real-time update: latest on covid-19 makeshift hospital in wuhan receives coronavirusinfected patients wenzhou municipal health commission. fight ten days for the inflection point! wenzhou sounds the bugle call to beat epidemic spread commanding china's fight against novel coronavirus outbreak number of confirmed covid-19 cases in wenzhou from key: cord-310331-29srzbuk authors: xu, jiuyang; chen, yijun; chen, hao; cao, bin title: 2019 novel coronavirus outbreak: a quiz or final exam? date: 2020-03-20 journal: front med doi: 10.1007/s11684-020-0753-1 sha: doc_id: 310331 cord_uid: 29srzbuk the 2019 novel coronavirus (2019-ncov) is an emerging pathogen and is threatening the global health. strikingly, more than 28 000 cases and 550 deaths have been reported within two months from disease emergence. armed with experience from previous epidemics in the last two decades, clinicians, scientists, officials, and citizens in china are all contributing to the prevention of further 2019-ncov transmission. efficient preliminary work has enabled us to understand the basic characteristics of 2019-ncov, but there are still many unanswered questions. it is too early now to judge our performance in this outbreak. continuous and strengthened efforts should be made not only during the epidemic, but also afterwards in order to prepare for any incoming challenges. the 2019 novel coronavirus (2019-ncov) outbreak is currently bringing challenges to china and the whole world. first reported in december 2019, it has caused over 28 000 cases in around 2 months, among whom at least 3800 have severe illness and more than 550 deaths have been reported (data as of february 5, 2020) [1] . on january 30, world health organization has announced it as a public health emergency of international concern [2] . since direct human transmission and asymptomatic infection have been revealed for 2019-ncov [3] , health authorities in china are facing an enormous challenge on disease management and control. 2019-ncov is the third highly pathogenic coronavirus emerging in the 21st century, also one of the epidemics or pandemics since the millennium (fig. 1) . human coronavirus had been overlooked for a long time before the severe acute respiratory syndrome coronavirus (sars-cov) outbreak in 2002, leaving 8096 infections with a mortality rate of nearly 10% [4] . fortunately, public health measures, including isolation and quarantine, finally put an end to the sars pandemic in summer 2003. in 2009, the influenza a/h1n1 swept over 200 countries and regions, leaving with more than 18 449 deaths [5] . subsequently, middle east respiratory syndrome coronavirus (mers-cov), another coronavirus, has posed a continuous threat to global health since 2012. considered as a zoonotic virus, mers-cov infection is reported occasionally and sometimes in cluster, such as the outbreak in south korea where one infected traveler back from the middle east led to 186 infections and 36 deaths in 2015 [6] . by november 2019, mers-cov has infected 2494 individuals and caused 858 deaths [7] . in addition, ebola virus outbreak in 2014 and zika virus outbreak in 2015 both contributed dramatic damage to the affected countries. public responses to 2019-ncov outbreak and previous experience experience from previous epidemics has trained us to perform more positively in the incoming threat. for example, the disastrous outbreak of sars in 2003 promoted the reformation of chinese center for disease control and prevention (ccdc), driving the establishment of a better disease surveillance network. as a result, relatively quick response and actions were made during the mers outbreak in south korea in 2015, avoiding further spread in china. realizing the severity of sars-cov as a respiratory virus, the detection of pneumonia of unknown origin in 2019-ncov outbreak enabled relatively early alarm from health authorities and raised awareness for medical staff to equip personal protection methods when dealing with suspected cases. however, material reserve (i.e., gloves, protective suits) for health worker is far from enough with the outbreak evolving, indicating the insufficiency of financial support and emergency practice. the rapid progress in science nowadays has also prepared us for the recent 2019-ncov outbreak. first, modern biological techniques developed in the last decade have greatly promoted the speed of pathogen detection. the whole-genome sequencing and isolation of 2019-ncov strains were accomplished in early january 2020, within two weeks from identifying the outbreak. in 2003, the same process took 4 months [8] . second, the more advanced hospital facilities in china, including advanced life support devices (i.e., extracorporeal membrane oxygenation), have improved our ability to manage severe cases, although the equipment is still in shortage in the central disease area. the outbreaks of sars and mers informed us about the possibility of highly pathogenic coronavirus, but there are still many unanswered questions. although basic research has generated several promising candidates [9, 10] , we are still not armed with vaccines or specific antiviral drugs against coronaviruses 17 years after sars and 7 years after mers outbreak. the novel coronavirus indeed hit this weakness. as of january 31, the number of confirmed cases for 2019-ncov has surpassed that of sars-cov and the number is still climbing. besides lack of specific antiviral treatments and deficiency in the understanding of novel virus, many other factors have also contributed to the wider transmission of 2019-ncov, such as increased population density coming with the increased urbanization, easier longdistance travel resulting from improved public transportation, as well as the surge of travelers during the spring festival travel rush. while the more convenient lifestyle enhances the transmission of 2019-ncov, it also facilitates personal prevention after the outbreak. for example, the more advanced personal terminal equipment such as smartphones facilitates information sharing and real-time epidemic update for disease control. in addition, onlineordered service including take-out food, online shopping, and fresh product delivery make self-isolation in urban areas relatively possible. we are glad to see that multiple public health measures were taken to prevent the disease transmission. 2019-ncov was identified as category b infectious disease and managed as class a on january 20 according to the law of the people's republic of china on prevention and treatment of infectious diseases. on january 23, wuhan city, the initial epidemic focus, suspended all public transport to prevent further transmission during the spring festival holiday (starting from january 25), which was then simulated by several other neighboring cities. at the same time, chinese community-based governments also adapt receptive measures to propagate the threat brought by 2019-ncov and recommend personal hygiene in the communities. with the popularity of smartphones, these propagations arrive at almost every family to partially slow down the spread of the disease. as of february 3, a total of 47.0 billion rmb has been applied to disease control and prevention [11] . the authorities as well as the scientific community in china have been working diligently to battle against the outbreak. after the local authority was informed of the cluster of pneumonia of unknown origin associated with huanan seafood market on december 31, 2019, an expert team of clinicians, epidemiologists, scientists, and officials was dispatched to wuhan, and suspected cases were soon transferred to a designated hospital (jinyintan hospital). several groups have independently isolated a novel pathogen from the lower respiratory tract of the patients in early january 2020. zhu et al. identified the pathogen as a novel coronavirus (2019-ncov) by unbiased sequencing and obtained the transmission electron microscopy image [12] . this finding was supported by data from other groups and genomic studies showed that 2019-ncov was closest to a sars-like coronavirus in bats, and that the receptorbinding domain (rbd) was close to that of sars-cov, suggesting ace2 as the possible receptor [13] [14] [15] [16] . however, more studies are needed to fulfill koch's postulate for 2019-ncov and identify its cellular receptor. hunting for the animal source of 2019-ncov is also under way. apart from the basic science findings, clinicians have worked efficiently to provide first-hand experience with 2019-ncov patients. chan et al. reported a case of a family cluster of 2019-ncov infection, five of whom had travel history to wuhan but one did not, providing the first evidence that 2019-ncov was able to transmit from person-to-person [3] . an epidemiology study conducted by li et al. also confirmed cluster cases indicating early human transmission [17] . huang et al. gathered clinical features of the first cohort of 41 laboratory-confirmed cases and found that fever, dry cough, and dyspnea were common symptoms for the new disease [18] . chen et al. extended the description by gathering another 99 cases and adding details on combined bacterial and fungal coinfections [19] . experience from the front line helped to form a national guideline for the management of the novel coronavirus induced pneumonia patients (now fifth edition) [20] . this is not the end of clinical research on 2019-ncov. we are expecting good news from chinese physicians and scientists. in this era of we media nowadays, there are more challenges than ever in preventing and controlling infectious diseases. on one hand, social media promotes rapid information flow, facilitating efficient information exchange and enabling timely decision-making. on the other hand, it also allows rumor, misunderstanding, and fear to quickly turn around in the vast population. it should be noted that the words spoken by specialists are easily spread in public, with a high possibility of misunderstanding the jargons and following irrational behaviors. although the academic journal is still the best place for scientists to share and exchange viewpoints, it is not the only place, especially for civilians who do not have an academic background but are eager to know more. popular science readings are equally important, or at least as a supplement to the academic articles, in sharing scientific knowledge. there will be less fear when armed with knowledge. the 2019-ncov outbreak is an examination for the whole world, and for everyone. at the current stage, it is still too early to judge our actions in this outbreak. we have accomplished many, and more efforts are needed to better understand the novel virus, with various questions on its source, animal reservoirs, transmission and evolution pattern, pathogenesis, as well as appropriate treatment. 2019-ncov is only a quiz but not the final examination, with novel threats around the corner. therefore, a continuous and strengthened emphasis should be put on potential pathogens, such as coronaviruses, that might infect humans and cause epidemic. financial support, research work, and personal awareness of crisis are not supposed to wane with the departure of a sudden or "temporary" threat. we do need to be prepared, rather than be surprised. jiuyang xu, yijun chen, hao chen, and bin cao declare no conflicts of interest. this manuscript is a commentary and does not involve a research protocol requiring approval by the relevant institutional review board or ethics committee. china national health commission. the latest information for 2019 novel coronavirus infected pneumonia by the-second-meeting-of-the-international-health-regulations-(2005)-emergency-committee-regarding-the-outbreak-of-novel a familial cluster of pneumonia associated with the 2019 novel coronavirus indicating person-to-person transmission: a study of a family cluster world health organization. summary of probable sars cases with onset of illness from 1 world health organization. pandemic (h1n1) 2009-update 112 middle east respiratory syndrome coronavirus (mers-cov) -republic of korea middle east respiratory syndrome coronavirus (mers-cov) the genome sequence of the sars-associated coronavirus coronaviruses--drug discovery and therapeutic options antibodies and vaccines against middle east respiratory syndrome coronavirus ministry of finance of the people's republic of china. the government at all levels continued to increase input and allocated a total of 47 billion yuan for epidemic prevention and control china novel coronavirus investigating and research team. a novel coronavirus from patients with pneumonia in china genomic characterization of the 2019 novel human-pathogenic coronavirus isolated from a patient with atypical pneumonia after visiting wuhan identification of a novel bat-origin coronavirus causing severe pneumonia in human: a descriptive study genomic characterisation and epidemiology of 2019 novel coronavirus: implications for virus origins and receptor binding a pneumonia outbreak associated with a new coronavirus of probable bat origin early transmission dynamics in wuhan, china, of novel coronavirus-infected pneumonia clinical features of patients infected with 2019 novel coronavirus in wuhan epidemiological and clinical characteristics of 99 cases of 2019 novel coronavirus pneumonia in wuhan, china: a descriptive study china national health commission. the issuance of the diagnosis and treatment programme for pneumonia of novel coronavirus infections key: cord-347414-t88654wo authors: peng, mian; liu, xueyan; li, jinxiu; ren, di; liu, yongfeng; meng, xi; lyu, yansi; chen, ronglin; yu, baojun; zhong, weixiong title: successful management of seven cases of critical covid-19 with early noninvasive-invasive sequential ventilation algorithm and bundle pharmacotherapy date: 2020-08-06 journal: front med doi: 10.1007/s11684-020-0796-3 sha: doc_id: 347414 cord_uid: t88654wo we report the clinical and laboratory findings and successful management of seven patients with critical coronavirus disease 2019 (covid-19) requiring mechanical ventilation (mv). the patients were diagnosed based on epidemiological history, clinical manifestations, and nucleic acid testing. upon diagnosis with covid-19 of critical severity, the patients were admitted to the intensive care unit, where they received early noninvasive–invasive sequential ventilation, early prone positioning, and bundle pharmacotherapy regimen, which consists of antiviral, anti-inflammation, immune-enhancing, and complication-prophylaxis medicines. the patients presented fever (n = 7, 100%), dry cough (n = 3, 42.9%), weakness (n = 2, 28.6%), chest tightness (n = 1, 14.3%), and/or muscle pain (n = 1, 14.3%). all patients had normal or lower than normal white blood cell count/lymphocyte count, and chest computed tomography scans showed bilateral patchy shadows or ground glass opacity in the lungs. nucleic acid testing confirmed covid-19 in all seven patients. the median mv duration and intensive care unit stay were 9.9 days (interquartile range, 6.5–14.6 days; range, 5–17 days) and 12.9 days (interquartile range, 9.7–17.6 days; range, 7–19 days), respectively. all seven patients were extubated, weaned off mv, transferred to the common ward, and discharged as of the writing of this report. thus, we concluded that good outcomes for patients with critical covid-19 can be achieved with early noninvasive–invasive sequential ventilation and bundle pharmacotherapy. the transmission of pathogens from animals to humans and resultant zoonotic infectious diseases have been associated with several major global outbreaks and constitute global public health burden. generally, coronaviruses (covs) are enveloped rna viruses that can cause respiratory and intestinal infections in humans and other animals. human infections with covs had been associated with mild upper respiratory disease before the 2003 outbreak of severe acute respiratory syndrome (sars) [1] . however, the discovery that sars is caused by a new cov, sars-cov, redefined our understanding of covs [2] . the potential pathogenicity of covs was further elucidated by the 2012 middle east respiratory syndrome (mers) outbreak, which is attributed to infection with yet another new cov, mers-cov. mers, which presents as acute pneumonia and occasional renal failure in severe cases, had an alarming 30% mortality rate in the 2012 outbreak [3] . the severe morbidities of cov diseases over the past two decades have shown that covs represent a serious infectious disease concern. in december of 2019, a cluster of patients with pneumonia of unknown cause was linked to transmission at a seafood wholesale market in wuhan, china. a previously unknown β-cov was discovered in samples from affected patients. the 2019 novel cov causing the ongoing pneumonia pandemic was identified to be of the same genus as the sars-cov and the seventh cov known to infect humans following 229e, nl63, oc43, hku1, mers-cov, and sars-cov [4, 5] . this causative virus was named as severe acute respiratory syndrome coronavirus 2 (sars-cov-2). the relevant infectious disease was named as coronavirus disease 2019 (covid-19) by the world health organization [6] . the cases of covid-19 can be classified as mild, moderate, severe, or critical based on clinical symptoms and radiologic images [7] . a total of 87 137 confirmed cases globally, including 79 968 confirmed cases in china and 7169 outside china, with 2977 deaths (3.4%) had been reported by the world health organization as of march 1, 2020 [8] . sars-cov-2 exhibits rapid transmission and causes atypical clinical symptoms, including fever, cough, myalgia, weakness, and dyspnea, as well as clinical signs recognizable in chest images that may be helpful for the early detection of emergent covid-19 [9] . patients with covid-19 who become critically ill and require mechanical ventilation (mv) have a particularly high mortality risk. therefore, an effective standard treatment regimen for covid-19 that can limit disease severity should be established to quell public panic. here, we report the successful management of seven critically ill patients diagnosed with covid-19 who suffered acute respiratory failure. we collected the clinical and laboratory data of the seven critically ill patients admitted to the intensive care unit (icu) of the third people's hospital of shenzhen. respiratory specimens (nasopharyngeal swabs, sputum, or bronchoalveolar lavage) from each patient were tested for sars-cov-2 with real-time reverse-transcriptase polymerase chain reaction testing. the patients who presented the clinical symptoms of covid-19 sought care in our hospital from january 20 to february 6, 2020. suspected diagnoses of covid-19 were made based on clinical symptoms and then confirmed based on white blood cell count (wcc)/lymphocyte count, chest computed tomography (ct) images, and nucleic acid test results. the patients were treated in the icu for covid-19. at present, these seven critically ill patients who suffered respiratory failure and required mv were treated successfully and discharged. the seven critically ill patients (four women; 57.1%), who are the subjects of this case series, had a median age of 56 years (interquartile range (iqr), 41-65 years; range, 36-71 years). the group included four patients (57.1%) with body mass index > 25, three patients with preexisting chronic diseases (42.9%), and six patients with wuhan contact history (85.7%). all seven patients had fever (100%), three had dry cough (42.9%), two experienced weakness (28.6%), one had chest tightness (14.3%), and one reported muscle pain (14.3%). the demographic and clinical characteristics of these patients are summarized in table 1 . laboratory tests (results reported in table 2 ) showed that all seven patients (100%) had normal or lower than normal wccs and lymphocyte counts, as well as bilateral patchy shadows or ground glass opacity in the lungs visible in chest ct scans (fig. 1 ). all the patients (100%) were treated with the following bundle pharmacotherapy regimen: lopinavir/ritonavir tablets (500 mg every 12 h) plus α-interferon (5.0 mu atomized inhalation every 12 h) as an antiviral treatment, thymosin α1 (1.6 mg/day subcutaneous injection) to enhance immunity, methylprednisolone (40 mg/day for the first 3 days) to reduce pulmonary exudation, traditional chinese medicine xuebijing (100-ml injection every 12 h) to attenuate inflammation, and low-molecular-weight heparin (4.0 kiu subcutaneous injection each day) to prevent deep vein thrombosis. the drugs (lopinavir/ ritonavir and α-interferon) were stopped when two consecutive negative respiratory sample nucleic acid tests with a sampling interval of ≥ 1 day were achieved. the patients received noninvasive ventilation (niv) with 60% oxygen absorption concentration (fio 2 ) and inspired/expiratory positive airway pressure (ipap/epap) levels of 8-12 and 6 cmh 2 o, respectively. however, the patients' conditions were not relieved in 2 h (pao 2 /fio 2 ratio reported in table 2 ). subsequently, all the patients were advanced to invasive mv with prone positioning (12-14 h/day) in the first 3-5 days. the median mv duration was 9.9 days (iqr, 6.5-14.6 days; range, 5-17 days). the mv start and end dates and duration for each patient, as well as icu admission and transfer-out dates and duration of icu stay, are reported in table 3 . none of the patients received extracorporeal membrane oxygenation. all seven patients were extubated and weaned off their ventilators by february 25, 2020 and exhibited clinically smooth breathing without fever or any other covid-19 symptoms. additionally, chest ct scans showed absorption of the aforementioned lung changes in all cases (fig. 2) . the patients were thus transferred from the icu to the common ward. the patients had a median icu stay of 12.9 days (iqr, 9.7-17.6 days; range, 7-19 days; table 3 ) and were discharged as of the writing of this report. thus, sars-cov-2 and covid-19 has rapidly spread [10, 11] . patients infected with sars-cov-2 have since been found in china, other countries in asia, the middle east, europe, and north america [12] . thus, an urgent international need for the establishment of effective therapeutic strategies has emerged to achieve the rapid control of sars-cov-2 infection and covid-19. according to china's national guidelines for the diagnosis of covid-19 (summarized in fig. 3) [7], these seven cases were diagnosed with covid-19 because of the positive nucleic acid affirmation of sars-cov-2 in their respiratory samples ( table 2) . covid-19 can be classified as mild, moderate, severe, or critical. mild covid-19 is characterized by mild clinical symptoms without signs of pneumonia on imaging. moderate covid-19 is characterized by fever, respiratory symptoms, and imaging manifestations of pneumonia. moderate-type covid-19 is elevated to severe-type if the patient shows respiratory distress (defined by a respiratory rate of ≥ 30 breaths/min, a resting oxygen saturation rate of ≤ 93%, or a partial arterial oxygen pressure (pao 2 )/ fio 2 ≤ 300 mmhg). severe covid-19 is considered to have progressed to critical covid-19 if the patient develops respiratory failure requiring mv, sepsis, or combined with organ failure requiring icu monitoring and care. the seven cases of covid-19 presented in this paper exhibited respiratory failure requiring invasive mv and thus were classified as critical-type covid-19. our department is focused on the early diagnosis and severity typing of covid-19. our management plan upon confirmation of a covid-19 diagnosis includes the immediate treatment of symptoms and the delivery of a comprehensive bundle pharmacotherapy. monitoring and supportive treatment is usually sufficient in patients with mild or moderate covid-19 to ensure good outcome. we provide close monitoring and bundle pharmacotherapy, as well as supplemental oxygen therapy, in patients with severe covid-19 to avoid the progression of covid-19 to critical level, which can be fatal. therapy plans may be individualized depending on the particular characteristics of individual patients ( table 3) . the patients with critical covid-19 were weaned off mv successfully, transferred to the common ward, and subsequently discharged. our experience with the concentrated treatment of this patient population from shenzhen, china has yielded three noteworthy lessons explained below. first, we found that the early determination of covid-19 severity is of utmost importance. missed diagnosis and misdiagnosis caused by atypical symptomology results in the loss of opportunity for early intervention [9] . patients with the following factors are at risk of severe or critical covid-19 [13, 14] : age > 60 years; body mass index > 25; underlying chronic disease; low lymphocyte count ( < 0.95 â 10 9 /l); and nasopharyngeal swab viral nucleic acid value < 30 or positive results with blood, stool, urine, or tear specimen. we believe that the most important among these factors may be lymphopenia. three among the seven patients described in this report were over 60 years old with a chronic underlying disease and lymphocyte counts < 0.95 â 10 9 /l, which indicate a less than robust immunocompetence. patients with lymphopenia tend to have the most serious symptoms, especially severe hypoxemia, and are prone to developing sepsis or multiple organ failure. second, early sequential noninvasive-invasive respiratory support and early prone position are effective measures for improving outcomes. respiratory support is a very important technique that can save patients with severe covid-19 who suffer from respiratory failure by improving oxygenation to reduce mortality. therefore, appropriate respiratory support technique is the key point related to its success or failure [15] . early respiratory support can facilitate disease recovery and improve prognosis [16] . we have come to favor a sequential noninvasive-invasive ventilation algorithm (summarized in fig. 4 fig. 4) . we restricted the duration of niv to ≤ 2 h because respiratory failure cannot be relieved with niv and the patients will continue to incur further oxygen debt if they have massive lung consolidation. mv with sedation and analgesia allows patients with critical covid-19 to rest while giving time for our comprehensive pharmacotherapy regimen to exert therapeutic efficacy in the lungs. the patients were rested in prone position to ensure that the diaphragm is in a favorable position to enable its movement while reducing the pressure gradient in the chest cavity. this position increases the functional residual capacity of the lungs, which improves oxygenation by improving the ventilationto-blood flow ratio and enhancing secretion drainage and fluid movement in the lungs. third, the administration of a bundle pharmacotherapy can support recovery from covid-19. we recommend a bundle therapy (see case report text for dosage information) with three principal components: (1) antiviral (lopinavir/ ritonavir plus α-interferon) and anti-inflammation drugs (methylprednisolone and xuebijing), (2) immune system support (thymosin α1), and (3) prevention/treatment of complications (low-molecular-weight heparin). all seven patients were treated with this bundle pharmacotherapy while receiving respiratory support and recovered well. all of them have been discharged as of the writing of this report. our discharge criteria for patients recovering from covid-19 are: normal body temperature for ≥ 3 days; alleviation of respiratory symptoms; majority resolution of radiological lung changes; and two consecutive negative respiratory sample nucleic acid tests with a sampling interval of ≥ 1 day [7] . among the seven critically ill patients whose cases are described in this report, five patients had mv durations of 5-9 days, and two patients required extended mv durations (16 and 17 days) because of ventilator-acquired pneumonia infections, which caused sepsis in one case (see table 3 for bacteria identified from sputum cultures and individualized treatments). consistent with the findings, we found that complications due to secondary infection are rare if the period of disease requiring mv is within a week or so. conversely, patients requiring 2 weeks or more of mv become exposed to greater risk of acquiring negative bacilli, positive cocci, and fungal infections, which make the patients at increased risk of systemic infection, sepsis, multiple-organ dysfunction, and death. thus, the strict control of hospital-acquired infections is of utmost importance in patients requiring mv to improve their likelihood of achieving rescue success. we aim to facilitate such success in part by enhancing patient immunity as discussed above. this report has a notable limitation in that only seven cases of successfully treated critical ill patients with covid-19 were included. more cases are needed to further confirm the effectiveness of our recommended treatment regimen. we hope that our treatment protocol will be applied to patients more broadly to collect data on its efficacy from broader populations. good treatment outcomes can be achieved for critically ill patients with covid-19 with early sequential noninvasive-invasive ventilation and bundle pharmacotherapy, which comprises antiviral and anti-inflammation medicines, immune system support, and prophylaxis for vascular complications. genetic and molecular biological analysis of protein-protein interactions in coronavirus assembly identification of a novel coronavirus in patients with severe acute respiratory syndrome isolation of a novel coronavirus from a man with pneumonia in saudi arabia tan w; china novel coronavirus investigating and research team. a novel coronavirus from patients with pneumonia in china a pneumonia outbreak associated with a new coronavirus of probable bat origin coronavirus disease 2019 (covid-19): what we know interpretation of guidelines for the diagnosis and treatment of novel coronavirus (2019-ncov) infection by the national health commission (trial version 5) the origin, transmission and clinical therapies on coronavirus disease 2019 (covid-19) outbreak-an update on the status early detection and disease assessment of patients with novel coronavirus pneumonia an emerging coronavirus causing pneumonia outbreak in wuhan, china: calling for developing therapeutic and prophylactic strategies a familial cluster of pneumonia associated with the 2019 novel coronavirus indicating person-to-person transmission: a study of a family cluster potential for global spread of a novel coronavirus from china expert panel of critical care medicine for covid-19 pneumonia in shenzhen. the shenzhen 2020 guidelines for the diagnosis and treatment of severe/critical covid-19 host susceptibility to severe covid-19 and establishment of a host risk score: findings of 487 cases outside wuhan respiratory support for severe 2019-ncov pneumonia suffering from acute respiratory failure: time and strategy clinical characteristics of novel coronavirus cases in tertiary hospitals in hubei province we acknowledge the writing guidance provided by prof. kunmei ji and the training camp for medical research held by shenzhen medical association and huada. we are grateful to the physicians and nurses of the third people's hospital of shenzhen who participated in the clinical examinations and sample collection. mian peng, xueyan liu, jinxiu li, di ren, yongfeng liu, xi meng, yansi lyu, ronglin chen, baojun yu, and weixiong zhong declare no conflict of interest. written informed consent was obtained from all the patients. key: cord-338258-bdyalfcd authors: chen, enfu; wang, fenjuan; lv, huakun; zhang, yanjun; ding, hua; liu, shelan; cai, jian; xie, li; xu, xiaoping; chai, chengliang; mao, haiyan; sun, jimin; lin, junfen; yu, zhao; li, lianhong; chen, zhiping; xia, shichang title: the first avian influenza a (h7n9) viral infection in humans in zhejiang province, china: a death report date: 2013-06-10 journal: front med doi: 10.1007/s11684-013-0275-1 sha: doc_id: 338258 cord_uid: bdyalfcd this study reports the first death caused by a novel avian influenza a (h7n9) virus in zhejiang province, china. the patient had chronic hepatitis b and history of exposure to poultry. the patient initially complained of diarrhea and influenza-like symptoms on march 7 and 14 respectively. the disease progressed to severe pneumonia, sustained hypoxia, and coagulation abnormalities. the patient died on march 27 because of respiratory failure, multiple organ failure, and disseminated intravascular coagulation without oseltamivir treatment. this h7n9 virus from zhejiang is highly similar to isolates obtained from shanghai, jiangsu, anhui, etc. analysis of hemagglutinin, neuramidinase, and matrix genes indicated that the isolates share the same avian origin, have low virulence, and are sensitive to oseltamivir, but are resistant to adamantine. only the isolate that caused the fatality exhibited substitution of q226i in the ha gene, which indicates a potentially enhanced human affinity. the secondary transmission rate was 1.6% (2/125). only two health workers presented with influenza-like symptoms, and they subsequently tested negative for h7n9 rna. in conclusion, underlying disease, late diagnosis, and untimely antiviral treatment are possible high-risk factors for infections and death caused by the lowpathogenicity avian influenza a (h7n9). person-to-person transmission of the h7n9 virus was not detected among close contacts, but such transmission should be investigated in the future. expanding and enhancing surveillance will help in the early discovery and diagnosis of suspected cases, which will reduce the number of severe cases and deaths. an emerging infectious disease that presents with acute high fever, cough, and pink frothy sputum with leukocytopenia was identified in china, from shanghai, beijing, jiangsu, zhejiang, and anhui since february 2013 [1, 2] . some of the cases developed severe pneumonia and acute respiratory distress syndrome (ards) [3] , subsequently requiring intensive care and mechanical ventilation. about 24.43% (32/131) of the patients died of respiratory failure and multiple organ failure (mof). in march 2013, epidemiologic, clinical, and pathologic investigations were performed that subsequently identified the etiology as a novel re-assortment avian influenza a (h7n9) virus distinct from the previous circulating human influenza a viruses [2, 4] . it was the first reported case of a human infected with a low-pathogenicity h7n9 virus that had a fatal outcome. as of may 16, 2013 , 131 laboratory-confirmed cases, including 32 deaths, have been reported in china since the first patient was identified on march 25, 2013 [5] . zhejiang province is one of the areas involved, and it is adjacent to shanghai, jiangsu, and anhui. by may 16, 2013, 46 confirmed cases were reported, including 10 deaths, after the first h7n9 case was diagnosed on april 1, 2013 [5] . sporadic cases were found daily in more areas because of the expanding and enhanced surveillance of influenza/avian influenza, pneumonia, and severe acute respiratory infection since the first h7n9 case was confirmed. in the current paper, we describe the discovery and epidemiological, clinical, and virologic characterization of the first fatal case of infection by the novel h7n9 virus in zhejiang province. this report is important for assessing and controlling similar cases, and would provide meaningful insights into the early and rapid diagnosis, treatment, prevention, and control of human influenza a (h7n9) viral infections. the case and contact definitions were set based on the diagnosis and treatment guidelines (second edition) established by the chinese ministry of health. a confirmed h7n9 case was defined as a suspected case with respiratory specimens positive for h7n9 virus based on isolation of the h7n9 virus or on positive real-time reverse transcription polymerase chain reaction (rrt-pcr) assays for the h7n9 virus. patients with pneumonia-complicated respiratory failure or other organ failure was defined as severe cases [6] . contacts were defined as (1) medical staff or relatives of the patient who did not take protective measures upon the diagnosis and treatment of suspected or confirmed cases, or those who took care of the patient; (2) persons who lived with or were in close contact with the suspected or confirmed cases for at least 1 week; (3) the investigators who were also considered close contacts [7] . we conducted a retrospective descriptive study of the clinical, laboratory, and radiographic characteristics of a confirmed case of avian influenza a (h7n9) viral infection during the study period (april 5, 2013 to april 10, 2013). all available medical records were reviewed by 10 clinicians, using a standardized data collection tool. epidemiologists and local public health doctors interviewed family members and medical staff using a standard questionnaire. pharyngeal swabs from the patient and from 46 of the 125 contacts were submitted to zhejiang cdc and china cdc for testing for h7n9 viral rna via rrt-pcr assays. avian influenza a (h7n9) virus rna extraction was performed following the instructions of the kit (qiagen company), and 200 μl of template was dissolved in doubledistilled water, aliquoted into four samples (50 μl each), and stored at -80°c. rrt-pcr was used to detect the ha, na, and m genes of seasonal influenza viruses (h1, h3, and b), h5n1, h7n9, severe acute respiratory syndrome coronavirus (sars-cov), and novel coronavirus. their specific primer and probe sets were provided by the china cdc. a pharyngeal swab specimen positive for h7n9 viral rna was further cultured in mdck cells maintained in hank's balanced solution at 37°c for 7 days. the cell cultures were observed for 2 weeks. the subtypes were identified using h7n9 ha and na type-specific primers when the cells exhibited the characteristic cytopathic effect (cpe). the subtypes were determined via a hemagglutination inhibition test (hai) assays using type-specific reference antibodies. the nucleotide sequences of the amplified products were directly determined via dideoxy sequencing using an abi prism bigdye terminator cycle sequencing kit. the ha, m, and na segments were analyzed and compared using mega 5.05 and dnastar. phylogenetic trees were constructed via the neighbor-joining method to estimate the viral gene relationships with selected influenza a virus strains obtained from the gisaid database (http://platform.gisaid.org/epi3/ frontend#575253) and genbank (http://www.ncbi.nlm.nih. gov/nuccore/?term = h7n9) [2] . on march 7, 2013, a 39-year-old male cook with chronic hepatitis b who works in suzhou, jiangsu province and lives in hangzhou, zhejiang province, china, reported cough and diarrhea. he consulted an outpatient clinic in hangzhou on march 9 and he was admitted to the second hospital of jiande in hangzhou on march 18, 2013 for the first time. on admission, he had fever (39.5°c), cough, and expectoration of bloody sputum, and his chest radiograph showed shadows in the right lower lobes. blood tests showed a white cell count of 2.33 â 10 9 /l and his c-reactive protein (crp) was 43.7 mg/l. on march 20, 2013 he was transferred to the first people's hospital in xiaoshan in hangzhou to hospitalization for the second time, because the cefodizime + levofloxacin treatment was ineffective. his second chest radiograph showed diffuse bilateral consolidation with right pleural effusion. he was diagnosed with "acute pneumonia." his condition continued to deteriorate and he developed multiple organ failure (mof) on march 24 despite oxygen therapy, broad-spectrum antibiotics (ceftriaxone + tazobactam), and corticosteroid (urbason) treatment. consequently, he was intubated and ventilated on the same day. on march 27, he died of ards, disseminated intravascular coagulation (dic), and mof. the swab samples collected on march 24, 2013 and the rna tested positive for influenza a but excluding seasonal influenza a/h1, h3, h5n1, 2009 pandemic h1n1 and influenza b, sars-cov, and hcov-erasmus medical center under rrt-pcr assays. he was confirmed as an avian influenza a (h7n9) virus case by analyzing eight segments, performed in china cdc on april 1, 2013. the delay in seeking medical care resulted in the patient presenting with severe acute lower respiratory symptoms with several complications upon hospital admission. thus, he was diagnosed with avian influenza a (h7n9) 5 days after he died and he did not receive antiviral treatment during the clinical course of the disease. in early march 2013, he worked as a cook in suzhou, jiangsu province and bought meat and poultry every morning from the local live poultry market 2 miles away, and then prepared lunch and dinner for about 30 company staff. the patient had no travel history before onset on march 7. according to the epidemiologic investigation, the patient had 125 close contacts, including 13 relatives, 45 colleagues, 57 doctors and nurses, and 10 patients exposed within the same ward. among these contacts, one doctor and one nurse developed cough one day after the patient was diagnosed and treated. both of the symptomatic contacts were negative for h7n9 avian influenza rna under rrt-pcr assay. the symptoms resolved spontaneously within 2 days. a total of 98 serum and 46 throat swabs samples from close contacts were tested using rrt-pcr on april 2, and none tested positive for h7n9. the demographic and epidemiologic characteristics of the patient are summarized in table 1 . clinical diagnosis and treatment the initial symptoms reported on march 7 and march 14 were diarrhea and cough respectively, which worsened, and was accompanied by fever (39.5°c) and hemoptysis on march 17. then patient developed shortness of breath, weakness, poor appetite, cyanosis, coma, and anuria because of progression to severe pneumonia with pleural effusion, and mof. he had died of ards and mof. the laboratory tests show that the patient's white blood cell count decreased to 1.6 â 10 9 /l during the early stages, which slowly increased to normal levels after treatment, and reached abnormally increased levels in the later stages. the neutrophil count exhibited a similar trend as the white cell count. the lymphocyte and mononuclear cell counts were consistently lower than normal, but the red cell count remained stable at normal levels. see fig. 1 . d-dimer remained abnormally high and the platelet counts remain low. activated partial thromboplastin time (aptt), prothrombin time, and thrombin time were initially normal levels, but became prolonged in the later stage. see fig. 2 . fig. 3 . liver and heart function were damaged, as indicated by the increased levels of lactate dehydrogenase (ldh), creatine kinase (ck), creatine kinase mb (ck-mb), alanine aminotransferase (alt), aspartate aminotransferase (ast), direct bilirubin (db), and total bilirubin (tb). however, kidney function was unchanged, as indicated by normal blood urea nitrogen (bun) and cr (creatinine). blood electrolytes such as na + and k + were normal. crp was persistently increased. these clinical findings are summarized in table 2 . the patient (temperature, sao 2 ) continued to deteriorate and eventually died on march 27, 2013 despite the administration of broad-spectrum antibiotics such as piperacillin sodium and tazobactam sodium, imipenem and cilastatin sodium, moxifloxacin hydrochloride and methylprednisolone sodium succinate (ranging from 40 mg to 240 mg) with intubation and continuous ventilation. see fig. 5 . the virus was confirmed using fluorescence quantitative pcr for detecting nucleic acids. n30d and t215a) . the m2 protein was found to exit s31n substitution, indicating resistance to adamantanes (amantadine and rimantadine). see table 3 . the patient swab was inoculated into mdck cell cultures. the typical cpes of h7n9 were observed under an inverted microscope: infected cells began to swell, became rounded, and developed larger intercellular spaces. the cells ruptured within 24 h after infection (fig. 9 ). the icid50 of the isolated virus was between 10 -4 and 10 -5 . the presence and subtype was further confirmed as h7n9 via an hai test and the specific primers used to amplify the h7n9 rna. influenza a (h7) virus outbreaks have occurred among avian populations in most european countries. the predominant subtypes circulating in poultry population were h7n1, h7n3, h7n4, h7n7, h7n8, and h7n9 [8] . human infections with h7 influenza viruses (h7n2, h7n3, and h7n7) were reported in the netherlands, italy, canada, and so on [9] [10] [11] . most of these infections occurred in association with poultry outbreaks. the infections mainly resulted in conjunctivitis, mild upper respiratory symptoms, and moderate illness resulting in hospitalization (lower respiratory tract disease), except for one death [12] [13] [14] . only 100 h7 cases have been confirmed worldwide from 1996 to 2012. the 2013 h7n9 virus reported in china is an avian influenza virus that is the first bird flu subtype (h7n9) found in humans. the novel virus is very different from other h7n9 viruses previously found in birds [1, 2] . according to the updated data from 131 cases and 32 deaths, the novel influenza a (h7n9) virus circulating in the urban and rural areas of beijing and shanghai, as well as anhui, jiangsu, zhejiang, etc., differs from the avian h5n1 reported in rural areas [15] . the main high-risk population is similar to that of h5n1, which consists of individuals with frequent contact with poultry, those with suppressed immune function, and the elderly with underlying diseases. the initial epidemiology suggests that the confirmed h7n9 cases were isolated, without sustained transmission among people, although two family clusters have been reported, which differs from the limited person-to-person transmission in h5n1 [16] . the source of infection and the mode of transmission are currently unknown. although some of the confirmed cases were exposed to birds and animals or to outdoor environments, the cases are not associated with disease outbreaks among animals or with direct exposure to animals. about 65% of the confirmed cases had histories of avian exposure, which indicates that contact with live poultry and excreta is a high-risk factor, similar to the h5n1 virus infection in china [17, 18] . the confirmed h7n9 cases developed critical and fatal illness, which indicates that the h7n9 virus is more virulent in humans than other h7 viruses and the 2009 h1n1 virus [19] . the current h7n9 case fatality rate is similar to that for reported h5n1 viral infections [20] . the first death was reported in zhejiang province, caused by a highly homologous avian h7n9 virus circulating in other areas of china. phylogenetic analysis showed that this novel virus originated from avian populations. the ha gene sequence from the patient indicated that the virus may be better adapted to infecting mammals than other avian influenza viruses because of the presence of q226i in the zhejiang strain and q226l in the shanghai 01 strain; the ha protein potentially enhanced the ability to bind to mammalian-like receptors with sialic acid moieties linked to galactose via α-2,6 linkages [21, 22] . cleavage site had only one r, which indicates its low pathogenicity, similar to other chinese strains. the na sequence data indicate antiviral resistance to adamantanes and susceptibility to neuraminidase inhibitors. no differences in active sites of mp gene, glycosylation sites, and na stalk deletion were observed between the zhejiang strain and other chinese strains. additional analyses are needed to understand its significance and the effect of single mutations. no outbreaks were identified in zhejiang avian population before the onset of the illness. the source of infection was thought to be exposure to poultry. the virus needs to be tested further to support the exposure in the live poultry market. although the source of infection and the mode of transmission have not yet been determined, control measures such as closing live poultry markets should be implemented. although secondary cases developed (the nurse and doctor) during diagnosis and treatment, the attack rate was only 1.6% (2/125) and no close contacts tested positive for h7n9, exclusion of possible person-to-person transmission cannot be considered [13] . the main clinical features of the first patient in zhejiang province were similar to those reported in other areas of china. they share features such as acute onset and rapid development to severe and sustained hypoxia accompanied by respiratory failure, ards, pleural effusion, and mof. despite the administration broad-spectrum antibacterial agents, cortisone, and mechanical ventilation, his condition : an isolate from the first confirmed h7n9 case in zhejiang; : isolates from the confirmed h7n9 patients from anhui and shanghai in china; : isolates from human h7 cases; : influenza vaccine strains in 2012. deteriorated, resulting in death. considering the patient had chronic hepatitis b, the case significantly differs from other cases in china. the patient presented with hemoptysis, sustained severe liver function, serious coagulation dysfunction, and dic. considering the diagnosis was confirmed 5 days after the patient died, he did not receive oral oseltamivir, which contributed greatly to the fatal outcome. in summary, we report the first human death caused by human avian influenza (h7n9) virus in zhejiang province. the patient was exposed to poultry before he initially : an isolate from the first confirmed h7n9 case in zhejiang; : isolates from the confirmed h7n9 patients from anhui and shanghai in china; : isolates from human cases . : an isolate from the first confirmed h7n9 case in zhejiang; : isolates from the confirmed h7n9 patients in anhui and shanghai, china. the main laboratory findings were increasing ast, alt, ldh, pt, aptt, cpk, and crp, but decreasing sao 2 % with disease progression. the major complications were ards, mof, and dic. the patient did not receive any antiviral treatment, although he received antibiotics, glucocorticoid hormone therapy, and invasive mechanical ventilation. the advanced infection control was achieved by testing all contacts, identifying the source of the virus and suspected cases, surveillance in zhejiang province, and sterilization. considering h7n9 viral infections have not occurred in humans before, persons of all ages might be susceptible. we are uncertain whether the influenza a (h7n9) virus could cause a pandemic. enhanced and expanded surveillance to ensure immediate discovery, diagnosis, and treatment of suspected cases will reduce the number of severe h7n9 cases and deaths. information on the emergent h7n9 virus is currently limited; therefore, studies on the source of the virus, transmission models, serologic investigations, vaccines, and drug development are necessary. the sampling activities were approved by the chinese medical ethical committee and the national health and family planning commission approved the study. this study was supported by the program for zhejiang leading team of science and technology innovation (grant no. 2011r50021) and for zhejiang provincial health bureau (2013kya043 and 2012kya161). none of the funders had any role in the study design, the collection, analysis, and interpretation of data, the writing of the article, and the decision to submit it for publication. the researchers confirm their independence from funders and sponsors. global concerns regarding novel influenza a (h7n9) virus infections human infection with a novel avian-origin influenza a (h7n9) virus world health organization. interim who surveillance recommendations for human infection with avian influenza a (h7n9) virus avian influenza a (h7n9) virus number of confirmed human cases of avian influenza a (h7n9) reported to who diagnostic and treatment protocol for human infections with avian the mdck cells exhibited the characteristic cytopathic effect (cpe) when inoculated with the patient's swabs. the cpe presented as cell swelling and becoming rounded, with increased intercellular spaces; the cells ruptured within 24 h after infection ( â 100). (a) control; (b) cpe; yellow arrow, cpe site. influenza a (h7n9) prevention and control protocol for human infections with avian influenza a (h7n9) avian influenza in north america novel avian influenza h7n3 strain outbreak outbreak of low pathogenicity h7n3 avian influenza in uk, including associated case of human conjunctivitis influenza virus a (h10n7) in chickens and poultry abattoir workers avian influenza a virus (h7n7) associated with human conjunctivitis and a fatal case of acute respiratory distress syndrome preliminary report: epidemiology of the avian influenza a (h7n9) outbreak in china human illness from avian influenza h7n3, british columbia seroprevalence of avian influenza a/h5n1 among poultry farmers in rural indonesia probable limited person-toperson transmission of highly pathogenic avian influenza a (h5n1) virus in china case-control study of risk factors for avian influenza a (h5n1) disease, hong kong human influenza a (h5n1) cases, urban areas of people's republic of china pandemic characteristics and controlling experiences of influenza h1n1 virus 1 year after the inception in hangzhou world health organization. cumulative number of confirmed human cases for avian influenza a (h5n1) reported to who host-range barrier of influenza a viruses receptor binding and membrane fusion in virus entry: the influenza hemagglutinin key: cord-330198-pwkxgbxk authors: cai, xiaofang; jiang, hanlan; zhang, simin; xia, shengying; du, wenhui; ma, yaoling; yu, tao; li, wenbin title: clinical manifestations and pathogen characteristics in children admitted for suspected covid-19 date: 2020-10-27 journal: front med doi: 10.1007/s11684-020-0820-7 sha: doc_id: 330198 cord_uid: pwkxgbxk coronavirus disease 2019 (covid-19), which is caused by severe acute respiratory syndrome coronavirus-2 (sars-cov-2), has spread around the world. however, approaches to distinguish covid-19 from pneumonia caused by other pathogens have not yet been reported. we retrospectively analyzed the clinical data of 97 children with probable covid-19. a total of 13 (13.4%) patients were confirmed positive for sars-cov-2 infection by nucleic acid rt-pcr testing, and 41 (42.3%) patients were found to be infected with other pathogens. notably, no pathogen was detected in 43 (44.3%) patients. among all patients, 25 (25.8%) had familial cluster exposure history, and 52 (53.6%) had one or more coexisting conditions. fifteen (15.5%) patients were admitted or transferred to the picu. in the 11 confirmed covid-19 cases, 5 (45.5%) and 7 (63.6%) were positive for igm and igg against sars-cov-2, respectively. in 22 patients with suspected covid-19, 1 (4.5%) was positive for igg but negative for igm. the most frequently detected pathogen was mycoplasma pneumonia (29, 29.9%). one patient with confirmed covid-19 died. our results strongly indicated that the detection of asymptomatic covid-19 or coexisting conditions must be strengthened in pediatric patients. these cases may be difficult to diagnose as covid-19 unless etiologic analysis is conducted. a serologic test can be a useful adjunctive diagnostic tool in cases where sars-cov-2 infection is highly suspected but the nucleic acid test is negative. electronic supplementary material: supplementary material is available in the online version of this article at 10.1007/s11684-020-0820-7 and is accessible for authorized users. coronavirus disease 2019 (covid19) , which is caused by severe acute respiratory syndrome coronavirus 2 (sars-cov-2), has spread around the world, constituting a public health emergency of international concern [1] . sars-cov-2 is the seventh coronavirus that has been identified to date that is known to cause human disease. compared with sars and middle east respiratory syndrome (mers) viruses that were responsible for pandemics in 2003 and 2012, respectively, sars-cov-2 is associated with a lower mortality rate but a higher transmission speed [2] [3] [4] [5] . most people, including children, are susceptible to sars-cov-2. however, data on pediatric patients with covid-19 are lacking. similar to sars and mers, most children infected with covid-19 have mild clinical presentation unless they have underlying comorbidities or coexisting conditions [6] . as the largest children's hospital in central china, wuhan children's hospital affiliated to tongji medical college of huazhong university of science and technology, was the first hospital assigned by the chinese government to treat children with covid-19 in hubei province, china. during the outbreak, only the fever clinic and emergency department remained open to tend to outpatients, all of whom were required to undergo two preexaminations and triage. all febrile or suspected covid-19 cases were referred to the fever clinic, and the others-including critically ill children-were received by the emergency department after pediatric 5-level triage. the majority of children with confirmed covid-19 were admitted to the hospital through the fever clinic, whereas some with probable covid-19 were admitted from the emergency department. we retrospectively analyzed the clinical data of these children admitted from the emergency department to characterize thoroughly the features of covid-19 that can be evaluated to distinguish this novel disease from pneumonia caused by other pathogens in pediatric patients. the study was approved by the institutional ethics board of wuhan children's hospital affiliated to tongji medical college of huazhong university of science and technology. oral consent was obtained from the legal guardians in this study. we retrospectively recruited a total of 97 children (excluding newborn cases) with probable covid-19 who were admitted by the emergency department of wuhan children's hospital from january 30, 2020 to march 15, 2020 . the clinical diagnostic criteria for covid-19 were based on chinese experts' consensus statement [7] . the patients' medical records were analyzed by the research team at the emergency department of wuhan children's hospital. epidemiological, clinical, laboratory, and radiological characteristics, as well as treatment and outcome data, were obtained by two trained staff using standardized data collection forms from electronic medical records. after admission, all 97 cases in the study underwent testing for a common set of pathogens, including influenza, parainfluenza virus, respiratory syncytial virus, adenovirus, enterovirus, mycoplasma, chlamydia, and legionella. routine bacterial and fungal examinations were also performed as necessary. a confirmed covid-19 case was defined as positive for sars-cov-2 nucleic acid in nasopharyngeal swab and/or anal swabs by real-time rt-pcr according to who guidance. clinical specimens were collected from patients with probable covid-19. real-time rt-pcr was performed using a nucleic acid test kit (bgi biotechnology co., ltd., wuhan, china) according to the manufacturer's instructions. serological test for detection of specific igm and igg antibodies against sars-cov-2 in the early phase of the covid-19 pandemic, no suitable assay for sars-cov-2 specific igm and igg detection was available. some children with probable covid-19 underwent serological testing in the hospital starting on march 5, 2020. serum samples were analyzed for igm and igg against sars-cov-2 by using chemiluminescence test kits (shenzhen yhlo biotech co., ltd., shenzhen, china) following the manufacturer's instructions. treatment, release from isolation, and follow-up no effective antiviral treatments for covid-19 have been developed to date [4, 7, 8] . we chose interferon α-2b aerosol inhalation, ribavirin intravenous infusion, oseltamivir, arbidol, and traditional chinese medicine oral administration as antiviral therapies for some patients following the recommendations of the pediatrics society of chinese medical association [7, 9] . appropriate oxygen support (nasal cannula and noninvasive or mechanical ventilation) was administered to patients according to the severity of hypoxaemia. glucocorticoid and immunoglobulin were used in severe cases. if the patient's body temperature (if febrile) returned to normal for more than 3 days, respiratory symptoms remarkably improved, pulmonary images clearly showed that the focus was evidently absorbed, and the nucleic acid test results were negative two consecutive times (the sampling interval was at least 1 day), then the patient was released from isolation or discharged. constant health monitoring and medical observation for 14 days at the patient's home was recommended. after 2 weeks post discharge, two trained staff followed up the patients via telephone. continuous variables were expressed as median and interquartile range (iqr) values and compared with mann-whitney u test or kruskal-wallis test. categorical variables were expressed as number (%) and compared with χ 2 test or fisher's exact test. a two-sided α value < 0.05 was considered statistically significant. data were analyzed using spss version 20.0 software (spss inc., chicago, il, usa), unless otherwise indicated. owing to the outbreak of covid-19, wuhan was put under lockdown starting on january 23, 2020, and nearly all citizens were required to remain at home. children with probable covid-19 were admitted to wuhan children's hospital starting on january 30, 2020. the majority of these admissions occurred between january 30 and march 15. during this period, out of the 4312 outpatients received by the emergency department, 97 (2.25%) with probable covid-19 were admitted to the hospital. out of the 97 patients, 13 (13.4%) cases were confirmed (classified as children with confirmed covid19) , and 41 cases were found to be infected with other pathogens (classified as children with non-covid-19). however, no pathogen was detected in 43 cases (classified as children with suspected covid-19) (fig. 1 ). at the same time, between january 30 and march 15, a total of 773 children with probable covid-19 were admitted to the hospital, of which 245 were confirmed as covid-19 cases, that is, 12.5% (97/ 773) probable covid-19 and 5.3% (13/245) confirmed covid-19 cases were admitted from the emergency department. all 97 patients lived in wuhan within 14 days before the onset of symptoms; 25 (25.8%) children had documented exposure to one or more adult family members with confirmed or suspected covid-19. compared with the 5 (11.6%) patients with suspected covid-19 and 10 (24.4%) patients with non-covid-19 causes, the exposure incidence of the 10 (76.9%) confirmed covid-19 cases was significantly higher (p < 0.01) ( table 1 ). out of the 97 patients, 52 (53.6%) had one or more comorbidities or coexisting conditions, including 6 (46.2%) confirmed covid-19 cases, 27 (62.8%) suspected covid-19 cases, and 19 (46.3%) non-covid-19 cases. epilepsy and encephalopathy (11, 11 .3%), acute abdomen (10, 10.3%), and immunodeficiency (8, 8 .2%) were the most common coexisting conditions (table 1) . the median time from symptom onset to hospital admission for the 97 patients 19 cases (7 (16.3%) and 1 (2.3%), respectively) and the non-covid-19 cases (1 (2.4%) and 3 (7.3%), respectively) (p = 0.013 and 0.040, respectively). the incidence of fever was higher in non-covid-19 cases (31, 75.6%) than that in confirmed covid-19 cases (7, 53.8%) and suspected covid-19 cases (19, 44.2%) (p = 0.013). a total of 19 patients with shortness of breath received oxygen, of which 4 (30.8%) were confirmed covid-19 cases, 6 (14.0%) were suspected covid-19 cases, and 9 (22.0%) were non-covid-19 cases. out of the 97 patients, 15 (15.2%) were admitted or transferred to pediatric intensive care unit because of organ dysfunction (table 1) . white blood cell (wbc) count was normal in 59 (60.8%) out of the 97 patients; 4 (30.8%) of the 13 confirmed covid-19 cases, 6 (14.0%) of the 43 suspected covid-19 cases, and 7 (17.1%) of the 41 non-covid-19 cases showed leucopenia (wbc count < 5.5 â 10 9 /l). most patients had slightly elevated c-reactive protein (5.00 mg/l; iqr, 0.75-23.00 mg/l) (54 out of 97 cases (55.7%)) and procalcitonin (0.12 ng/ml; iqr, 0.07-0.42 ng/ml) (69 out of 84 cases (82.1%)) ( table 2) . out of the 97 patients, alanine aminotransferase and aspartate aminotransferase were elevated in 12 (12.4%) and 35 (36.1%) cases, respectively. out of 91 patients, increased lactate dehydrogenase, creatine kinase (ck), and ck isoenzyme levels were observed in 23 (25.3%), 18 (19.8%), and 62 (68.1%) cases, respectively. out of 95 patients, blood urea nitrogen and creatinine were elevated in 7 (7.4%) and 3 (3.2%) cases, respectively ( table 2) . out of 42 patients, plasma concentrations of interleukin (il)-6 and il-10 increased in 16 (38.1%) and 20 (47.6%) cases (mostly severe), respectively. however, il-2, il-4, and tumor necrosis factor (tnf)-α levels were normal. no differences were found in il-6 and il-10 levels among confirmed, suspected, and non-covid-19 cases (p > 0.05). out of 36 patients, counts of cd4 + t cells, cd8 + t cells, and cd16 + cd56 + natural killer (nk) cells were below normal in 5 (14.3%), 10 (28.6%), and 13 (37.1%) cases, respectively ( table 2 ). counts of helper t cells (cd4 + ), suppressor t cells (cd8 + ), and nk cells (cd16 + cd56 + ) were reduced mainly in critically ill patients. no differences were observed in the numbers of cd4 + t cells, cd8 + t cells, and cd16 + cd56 + nk cells among confirmed, suspected, and non-covid-19 cases (p > 0.05). all 97 patients underwent rt-pcr testing for sars-cov-2 nucleic acid upon admission. the median number of tests was 3 (range, 2-5) in patients with confirmed covid-19, 2 (range, 1-5) in those with suspected covid-19, and 2 (range, 1-4) in those with non-covid-19 causes. out of the 97 patients, 53 (54.6%) patients underwent serum anti-sars-cov-2 igm and igg testing starting on march 5, 2020, including 11 (84.6%) confirmed covid-19 cases, 22 (51.2%) suspected covid-19 cases, and 20 (48.8%) non-covid-19 cases. out of the 11 confirmed covid-19 cases, 5 (45.5%) and 7 (63.6%) were positive for igm and igg, respectively. out of the 22 suspected covid-19 cases, 1 (4.5%) was positive for igg but negative for igm (table 3 , fig. 1 (table s1) . upon admission, chest imaging abnormalities were observed in all patients. typical findings in chest computed tomography (ct) images were bilateral distribution of patchy shadows or ground-glass opacity, and/or infiltrating shadows in the confirmed covid-19 cases (fig. s1) . the most commonly used antiviral therapy was interferon α-2b aerosol inhalation, which was administered to 13 (100%) confirmed covid-19 cases, 29 (67.4%) suspected covid-19 cases, and 27 (65.9%) non-covid-19 cases. four (30.8%) confirmed covid-19 cases, 2 (4.7%) suspected covid-19 cases, and 3 non-covid-19 cases received intravenous ribavirin. oseltamivir was orally administered to 3 (23.1%) confirmed covid-19 cases, 2 (4.7%) suspected covid-19 cases, and 6 (14.6%) non-covid-19 cases. three (3.5%) non-covid-19 cases were given oral arbidol. azithromycin was the most commonly used antibiotic (for mycoplasma infection) in 27 to date, 12 (92.3%) out of the 13 confirmed covid-19 cases and all suspected covid-19 cases and non-covid-19 cases have been cured and discharged. one (7.7%) confirmed covid-19 case had intussusceptions and multiorgan failure and eventually died [6] . at 2 weeks after discharge, all patients were followed up by telephone. only one patient was readmitted because of diarrhea, but nucleic acid retesting by rt-pcr remained negative. the other patients were in good condition with no symptoms related to covid-19. the most striking feature of this cohort was that the early symptoms in these children with probable covid-19 were apparently nonspecific or covered by other diseases. although we strictly adhered to two pre-examinations and triage for all outpatients, most of the probable and confirmed covid-19 cases were admitted from the fever clinic, and 12.5% (97/773) of the probable covid-19 cases (including 5.3% (13/245) of confirmed covid-19 cases) with atypical symptoms or critical illness were admitted from the emergency department. the initial symptoms of these patients did not always include fever or cough. some pediatric patients were brought to the emergency department because of digestive tract symptoms (vomiting, diarrhea, and abdominal pain), nervous system symptoms (convulsion), or even trauma. only when chest ct showed signs of pneumonia, they were hospitalized as probable or confirmed covid-19 cases. therefore, this approach was different from that of lu et al. [6] . owing to the parents' fear that their children were infected with sars-cov-2, the median time from symptom onset to hospital admission was shorter for confirmed covid-19 cases (2.0 days) than that for suspected covid-19 cases (3.0 days) and non-covid-19 cases (4.0 days) (p < 0.05). however, because of the mandatory isolation period, the median length of hospital stay was longer for confirmed covid-19 cases (15.0 days) than that for suspected covid-19 cases (10.0 days) and non-covid-19 cases (9.0 days) (p = 0.08). the clinical symptoms of covid-19 in children differ from those in adults [10] . most children with covid-19 had mild symptoms or were asymptomatic. in our study, the main symptoms in confirmed covid-19 cases during hospitalization were fever (53.5%), diarrhea (38.5%), vomiting (30.8%), shortness of breath (30.8%), cough (23.1%), and abdominal pain (23.1%). compared with suspected covid-19 and non-covid-19 cases, the incidence of gastrointestinal symptoms (diarrhea and abdominal pain) was higher in confirmed covid-19 cases (p < 0.05) ( table 1) . sars-cov-2 infection in children is mainly the result of family aggregation [11, 12] . our study showed that 10 (76.9%) out of the 13 confirmed covid-19 cases had documented exposure to one or more other adult family members with confirmed or suspected covid-19. thus, aggregative onset is an important feature in pediatric cases, and it also underscores the highly contagious nature of the virus [13] . children with covid-19 who are asymptomatic or have coexisting conditions can be misdiagnosed and may become potential vectors for virus transmission. similar to adult patients with covid-19, most children have normal or only slightly reduced leukocyte count without evident organ dysfunction [14, 15] . severely ill adult patients often have coexisting medical conditions [14, 16] . although severe cases are less frequent among children, they can progress to death when they have coexisting conditions [6] . we analyzed cytokine (il-2, il-4, il-6, il-10, and tnf-α) levels and cellular immune functions (cd3 + t cells, cd4 + t cells, cd8 + t cells, and cd16 + cd56 + nk cells) in some patients and found that the increase in il-6 and il-10 levels and the decrease in numbers of helper t cells (cd4 + ), suppressor t cells (cd8 + ), and nk cells (cd16 + cd56 + ) mainly occurred in severe cases with no difference between confirmed, suspected, and non-covid-19 cases (p > 0.05) ( table 2 ). this result suggested that the virus causes direct damage to organs and that further impairment of organ function related to cytokine storm induced by the viral invasion in critical cases is not unique to sars-cov-2 infection because it was also found in children that were severely infected with other pathogens. as with sars and mers, children with sars-cov-2 infection have low morbidity [17] [18] [19] [20] [21] . the reason for this phenomenon is unknown, but it has been attributed to the fact that children have a more active innate immune response, healthier respiratory tracts, and fewer underlying disorders than adults. a more vigorous immune response in adults may also explain the detrimental immune response in acute respiratory distress syndrome [22] . sars-cov-2, a newly emerged virus, binds with high affinity to human angiotensin converting enzyme 2 (ace2) and employs it as an entry receptor to invade target cells [23] . the ace2 protein is abundantly expressed in lung alveolar epithelial cells and enterocytes of the small intestines and is present in arterial and venous endothelial cells and arterial smooth muscle cells of all organs [24, 25] . therefore, sars-cov-2 may infect patients not only through the respiratory tract in the form of air droplets but also through the digestive tract by contact or fecal-oral transmission. we previously described chest imaging features in five covid-19 cases with nonrespiratory symptoms as the first manifestation in children [26] . in the early phase of the disease, chest x-rays showed no abnormalities, potentially leading to missed diagnosis. as in adults, chest ct images in children with cocid-19 revealed the presence of viral pneumonia, with early lesions frequently detected in the lung periphery and showing ground-glass opacity and/or infiltrating shadows. with disease progression, the lesions increased in number and expanded to involve multiple lung lobes. in the serious cases, the ct images showed diffuse consolidation in the lungs, as well as so-called white lung and air bronchogram. rt-pcr was one of the first laboratory diagnostic methods to be established in this pandemic and can yield results within 2 h. in our study, 13 out of the 97 children were confirmed to be positive for sars-cov-2 infection by this method. owing to the sudden outbreak of covid-19, detection kits varying in sensitivity and specificity were used for testing, and cases that were highly suspected of sars-cov-2 infection were sometimes found to be negative although they could not be completely ruled out [27] . in fact, nucleic acid testing of nasal and pharyngeal swabs has a detection time window [28] . dynamic monitoring of sars-cov-2 nucleic acid profiles by rt-pcr showed that the positive rate of oral swabs gradually decreases, whereas that of anal swabs increases with the duration of hospitalization [12] . at the same time, serum igm positivity increased from 50% (8/16) upon admission to 81% (13/16) after 5 days of hospitalization, whereas igg positivity rate increased from 81% (13/16) to 100% (16/ 16) [29] . therefore, to improve the reliability of diagnosis and avoid false-negative results, different biological specimens (nasal and pharyngeal swabs, anal swab, and blood) should be used for nucleic acid testing, and serum igm and igg levels should be monitored. serological tests can serve as an important supplementary tool for covid-19 diagnosis. therefore, serologic testing must be conducted in children who are highly suspected of having covid-19 but who have tested negative via rt-pcr. notably, in the 97 children with probable covid-19 who were initially admitted from the emergency department, the most frequently detected pathogen was m. pneumonia in 27 (65.9%) out of the 41 non-covid cases and 2 (15.4%) out of the 13 confirmed covid-19 cases. additionally, out of the 41 non-covid-19 cases, we detected other viral infections in 15 cases, bacterial infections in 6 cases, and fungal infection in 1 case. four cases had coinfection with two pathogens, and two cases had coinfection with three pathogens. therefore, clinical manifestations, laboratory test results, and findings from chest imaging of children with covid-19 can be nonspecific, and these cases cannot be definitively diagnosed as covid-19 unless etiological analysis is performed. in particular, coinfection with another pathogen-especially m. pneumonia-could mask the symptoms of the primary infection. our study had some limitations. first, it was a retrospective, single-center study with a small sample of patients admitted by the emergency department. standardized data from a larger prospective cohort study could provide more information on the features of covid-19 in children. second, we only conducted serologic testing in a subset of our patients and found just one case that was positive for sars-cov-2 specific igg in suspected covid-19 cases. detection of nucleic acid in different biological specimens of suspected covid-19 cases and dynamic monitoring of serum antibodies could reveal additional cases and thus reduce the risk of missed diagnosis. finally, according to the standards of experts' consensus statement [7] , suspected cases may be considered as confirmed based on supportive evidence from nucleic acid or serologic testing. however, serologic testing was not initiated until march 5, 2020 and was only performed in some children (25 cases) . to ensure the consistent application of diagnostic criteria to all patients, we used nucleic acid test results as the basis for diagnosis, and this approach may have resulted in some cases being overlooked. to address this potential omission, we are currently following up these children and advising them to undergo serologic testing. in summary, our findings highlighted the characteristics of covid-19 in pediatric patients admitted from the emergency department, which differ from those admitted from the fever clinic [6] . in children with similar clinical manifestations and even chest ct images, distinguishing covid-19 from pneumonia caused by other pathogens (e.g., m. pneumoniae) before etiologic analysis (nucleic acid and serological testing) is impossible. moreover, serologic testing can serve as an important adjunctive method for covid-19 diagnosis, especially when the patient is highly suspected of sars-cov-2 infection but is found to be negative by nucleic acid testing. with the ethical standards of the responsible committee on human experimentation (institutional and national) and with the helsinki declaration of 1975, as revised in 2000 (5) . oral consent was obtained from the legal guardians in this study. supplementary material is available in the online version of this article at https://doi.org/ 10.1007/s11684-020-0820-7 and is accessible for authorized users. coronavirus disease 2019 (covid-19): a clinical update 2019 novel coronavirus outbreak: a quiz or final exam unique epidemiological and clinical features of the emerging 2019 novel coronavirus pneumonia (covid-19) implicate special control measures severe acute respiratory syndrome coronavirus 2 (sars-cov-2) and coronavirus disease-2019 (covid-19): the epidemic and the challenges protecting healthcare personnel from 2019-ncov infection risks: lessons and suggestions chinese pediatric novel coronavirus study team. sars-cov-2 infection in children chinese medical association; chinese medical doctor association committee on respirology pediatrics; china medicine education association committee on pediatrics; chinese research hospital association committee on pediatrics; chinese non-government medical institutions association committee on pediatrics; china association of traditional chinese medicine, committee on children's health and medicine research global pediatric pulmonology alliance. diagnosis, treatment and prevention of 2019 novel coronavirus infection in children: experts' consensus statement washington state 2019-ncov case investigation team. first case of 2019 novel coronavirus in the united states recommendations for the diagnosis, prevention and control of the 2019 novel coronavirus infection on children (first interim edition) covid-19 patients' clinical characteristics, discharge rate, and fatality rate of meta-analysis novel coronavirus infection in hospitalized infants under 1 year of age in china 2019-novel coronavirus infection in a threemonth-old baby sars-cov-2 infection in children: transmission dynamics and clinical characteristics clinical characteristics of 138 hospitalized patients with 2019 novel coronavirus-infected pneumonia in wuhan, china dysregulation of immune response in patients with covid-19 in wuhan, china first case of neonate infected with novel coronavirus pneumonia in china first case of severe childhood novel coronavirus pneumonia in china middle east respiratory syndrome coronavirus disease in children other members of the hospital for sick children sars investigation team. children hospitalized with severe acute respiratory syndrome-related illness in toronto 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 are children less susceptible to covid-19? structure, function, and antigenicity of the sars-cov-2 spike glycoprotein tissue distribution of ace2 protein, the functional receptor for sars coronavirus. a first step in understanding sars pathogenesis pulmonary angiotensin-converting enzyme 2 (ace2) and inflammatory lung disease clinical characteristics of 5 covid-19 cases with non-respiratory symptoms as the first manifestation in children improved molecular diagnosis of covid-19 by the novel, highly sensitive and specific rdrp/hel real-time reverse transcription-polymerase chain reaction assay validated in vitro and with clinical specimens positive rate of rt-pcr detection of sars-cov-2 infection in 4880 cases from one hospital in molecular and serological investigation of 2019-ncov infected patients: implication of multiple shedding routes we acknowledge all healthcare workers involved in the diagnosis and treatment of patients in wuhan children's hospital. this study was supported by the national natural science foundation of china (no. 81741099). key: cord-307995-8q7efrqk authors: al-tawfiq, jaffar a.; omrani, ali s.; memish, ziad a. title: middle east respiratory syndrome coronavirus: current situation and travel-associated concerns date: 2016-05-04 journal: front med doi: 10.1007/s11684-016-0446-y sha: doc_id: 307995 cord_uid: 8q7efrqk the emergence of middle east respiratory syndrome coronavirus (mers-cov) in 2012 brought back memories of the occurrence of severe acute respiratory syndrome coronavirus (sars-cov) in 2002. more than 1500 mers-cov cases were recorded in 42 months with a case fatality rate (cfr) of 40%. meanwhile, 8000 cases of sars-cov were confirmed in six months with a cfr of 10%. the clinical presentation of mers-cov ranges from mild and non-specific presentation to progressive and severe pneumonia. no predictive signs or symptoms exist to differentiate mers-cov from community-acquired pneumonia in hospitalized patients. an apparent heterogeneity was observed in transmission. most mers-cov cases were secondary to large outbreaks in healthcare settings. these cases were secondary to community-acquired cases, which may also cause family outbreaks. travel-associated mers infection remains low. however, the virus exhibited a clear tendency to cause large outbreaks outside the arabian peninsula as exemplified by the outbreak in the republic of korea. in this review, we summarize the current knowledge about mers-cov and highlight travel-related issues. the emergence of middle east respiratory syndrome coronavirus (mers-cov) in 2012 brought back memories of the occurrence of severe acute respiratory syndrome coronavirus (sars-cov) in 2002 [1, 2] . as of march 17, 2016 , 1715 mers-cov cases were confirmed with a case fatality rate (cfr) of approximately 40%, whereas 8000 sars-cov cases were recorded in six months with a 10% cfr [1] [2] [3] . an upsurge in the number of cases was observed in march-may 2014 because of the outbreak in large healthcare facilities in jeddah, kingdom of saudi arabia (ksa) [4, 5] . more recently, a larger outbreak occurred in riyadh, ksa and in multiple hospitals in south korea [6] [7] [8] [9] . wolfe et al. [10] described the five stages of pathogen evolution that lead to diseases confined to humans. in stage 1, the pathogen is confined to an animal host. humans become infected by animals only in stage 2. stage 3 is limited human-to-human transmission. in stage 4, long outbreaks with numerous cycles of human-to-human transmission occur. in stage 5, the pathogen exclusively infects humans [10] . mers-cov has not yet reached stage 5. the interest and concern of the public and the health community in emerging infectious diseases stem from the ability of the emerging pathogens to cause pandemics with high fatality rates and the associated economic effects on affected countries. for example, sars-cov caused $30-50 billions of economic losses in mainland and hong kong of china, singapore, and canada, and the pandemic influenza h1n1 in 2009 resulted in $45-55 billion losses worldwide [11] . in this review, we focus on the important aspects of the recently emerged mers-cov, its effects on humans, and the transmission patterns of the disease based on available scientific data. year-old male from bisha [12, 13] . the patient was hospitalized with community-acquired pneumonia; the disease rapidly progressed, resulting in acute renal failure, respiratory failure, and death [13] . a summary of the major events in the development of the mers-cov epidemic is shown in fig.1 . since the first case of mers-cov, a detailed investigation was conducted to determine the environmental and animal source of the virus. extensive contact investigation was carried out on the first case that originated from bisha but was eventually hospitalized in a private hospital in jeddah [12, 14] . the contact investigation started in the patient's hometown of bisha in southern ksa. the investigation included his immediate household contacts (3 wives, 10 sons, 11 daughters, 12 grandchildren, and 1 house maid), as well as the community and healthcare facility in bisha, which included 2 shepherds who took care of his five camels and 14 healthcare workers (hcws) (11 nurses and three physicians) [14] . in total, 53 contacts in bisha were screened and all of them tested negative for mers-cov by polymerase chain reaction (pcr) [14] . an extensive investigation of the 48 out of 56 hcw who had significant contact with the same patient during his 10-day stay in the private hospital in jeddah tested negative [15] . during the october 2012 investigation on the source of the virus, a team representing three agencies, namely, saudi ministry of health, center for infection and immunity of columbia university, and ecohealth alliance, interviewed the family of the index case-patient from bisha. the team also collected samples from bats within 12 km from his home, as well as an abandoned date palm orchard, the area within 1 km from his place of employment, a hardware store that fronted his garden, and a date palm orchard. although none of his family members nor employees recalled seeing bats, the investigating team observed the roosting bats and guano in abandoned wells and ruins within 12 km of his home and insectivorous bats at dusk in the garden behind his store. a sample from a taphozous perforatus bat (egyptian tomb bat) captured in bisha showed 100% identity to the human β-cov 2c emc/2012 cloned from the index case-patient [16] . the largest data set on the contact investigation revealed that the percentage of positive cases was 2.5%, 1.12%, 3.03%, and 2.09% among hospital patients, hcw contacts, family and contacts, and overall [17] . with the expansion of testing to identify the full disease spectrum and the inclusion by the world health organization (who) of the positive cases by serology in july 2014 [18] , the cfr gradually decreased from 60% to 40% as more asymptomatic and mildly symptomatic cases were included [19] . a nationwide serosurvey for mers-cov conducted in the ksa between december 2012 and december 2013 suggested that around 45 000 infected individuals were not aware of their infection, which confirmed an extremely high incidence of asymptomatic to mildly symptomatic disease [20] . a good example of a family cluster of mers-cov infections was published in the new england journal of medicine [21] . once the second case in that cluster was identified, an epidemiologic investigation identified the index case who was the father of the second case who was admitted earlier with congestive heart failure and community-acquired pneumonia. subsequently, the third and fourth cases where recognized [21] . in that cluster, 28 persons lived in the same extended household, including nine children ( < 14 years of age) [21] . aside from the four patients, no other family members had major respiratory symptoms and none of the 124 hcws who managed the index case before the mers-cov diagnosis exhibited symptoms [21] . a second family cluster was also described by omrani et al. [22] . three patients lived in one large house in urban riyadh. none of the other family contacts had positive pcr tests. community outbreak was also described [23] . the first patient infected his cousin, and each of the patients infected their parents. genome analysis showed multiple introduction of the virus and determined three distinct genotypes, which confirm very low patient-to-patient transmission [23] [24] [25] [26] . a number of healthcare-associated outbreaks were reported previously [14, 19, 27, 28] . a cluster of acute respiratory illnesses was reported in the intensive care unit (icu) in zarqa, jordan in april 2012 [29] . thirteen hcw cases were detected (intensivists and icu nurses) with two mortalities. an initial investigation revealed no etiology. after the announcement of the first case of mers-cov in september 2012 in ksa, samples from these patients were retested for mers-cov and confirmed to be positive in two patients by pcr and eight contacts by serology [29, 30] . thus, this case demonstrated mers-cov as a healthcare outbreak. the second large outbreak occurred in al-hasa, ksa in april 2013 [26] . a total of 23 confirmed cases and 11 probable cases were recorded. a detailed transmission map was drawn based on the best available epidemiological data linking these patients epidemiologically [26] . subsequent genotyping showed multiple introduction of the virus leading to the outbreak rather than a single introduction [25] . four of the cases did not match the transmission, which indicates that the disease was not likely transmitted between the cases but had a different genome, thus indicating multiple community introductions [25] . the outbreak was controlled with simple infection control measures in three weeks. mers-cov was shown to have minor clades, and the most recent common ancestor of mers-cov was introduced into humans at the end of 2010 [31] . mers-cov is closely related to pipistrellus bat cov hku5 (pi-batcov hku5) in bats from hong kong and these bats diverged from a common ancestor several centuries ago [32] . in 2014, one of the largest healthcare-associated outbreaks of mers-cov infection took place between february 17 and april 26 in jeddah, ksa [4] . a total of 128 mers-cov patients were treated in 14 hospitals [4, 5] . the number of cases in each hospital varied from 2 to 45 cases [4, 5] . in all the cases, 33% were primary cases and 60% (including 39 hcw) were healthcare-associated infections [33] . the rapid increase in the cases was attributed to more sensitive case detection, active case determination, and contact tracing with changes in testing algorithms [4] . a breakdown in infection control measures was observed with no change in the virus [4] . a near full genome sequence of the three viruses from the early phase of the jeddah outbreak showed a highly similar virus with no genome insertions or deletions [4] . the genetic marker influencing transmissibility was 100% identical to known mers-cov genomes [4] . in 2015, the largest outbreak outside the middle east took place in south korea [6] . the index case was a 68year-old male who visited bahrain, ksa, united arab emirates, and qatar [6] . he developed symptoms on may 11, 2015 and visited multiple hospitals in south korea [6] . he caused an outbreak involving five health care facilities and 63 cases [7] , with 34 cases in hospital b, 18 cases in hospital d, 5 cases in hospital e, and three cases in hospital f [7] . as of june 19, 2015 the outbreak in the republic of korea involved 72 health care facilities, and six facilities exhibited nosocomial transmission [8] . the total number of cases as of july 21, 2015 was 186 cases with 36 deaths [34, 35] . based on epidemiological data monitoring over the last three years, the potential seasonality of mers-cov from march to may and from september to november was observed. in april and may 2014, the number of cases increased [4] . one of the reasons for this increase in the number of cases is the parallel surge in mers-cov tests in jeddah [4] . this increase is also facilitated by an intensified intra-hospital and inter-hospital transmission of mers-cov with no change in the virus genetic composition or ability to cause disease [4, 27] . thus, seasonality is difficult to establish because sporadic cases were documented with amplifications mainly occurring during nosocomial outbreaks. available data to date show that mers-cov behaves differently in various conditions and in different population of patients. isolated sporadic cases, small family clusters, as well as large healthcare-acquired infections and clusters, were recorded. most cases present with respiratory symptoms and about one-third had gastrointestinal symptoms (table 1) [26, [36] [37] [38] [39] [40] . early symptoms are mild and non-specific, which last several days prior to progressing to severe pneumonia. no predictive signs or symptoms exist to differentiate mers-cov from community-acquired pneumonia in hospitalized patients [38] . an apparent heterogeneity in transmission was observed. the severity of the disease is usually seen in primary or index cases, immune-compromised individuals, and people with underlying comorbidities. mild or asymptomatic disease usually occurs in secondary cases and was initially thought to infect the young and previously healthy individuals. however, mortalities and severe cases were observed among primary cases and young individuals [10] . however, person-to-person transmission as a definite route of transmission is still unclear. the median incubation period was 5.2 days (95% ci, 1.9 to 14.7), and the serial interval was 7.6 days (95% ci, 2.5 to 23.1) [26] . cfr is directly related to the number of comorbidities, the increasing detection of asymptomatic to mildly symptomatic cases over the last three years [39] . however, mortalities were reported among healthy individuals. the median time to hospitalization was 4 days, icu admission was 5 days, mechanical ventilation was 7 days, and death was 11.5 days [26, 41] . mers-cov pcr was standardized, and it works extremely well with lower respiratory samples in experienced laboratories. confirmatory testing in national or regional reference laboratories with experience and load of samples will avoid reporting false positive cases. on november 5, 2013, a spanish case from hajj was initially tested positive for mers-cov but was eventually sent to an outside reference laboratory for confirmatory testing; all tests were negative [42] . if mers-cov infection is suspected and initial testing is negative, repeat testing is recommended and lower respiratory tract samples would yield higher positivity [43] . in july 2014, a key change in mers-cov case definition is the inclusion of a confirmed case based on serology [44] . serologic mers-cov confirmation requires sero-conversion in two samples taken at least 14 days apart by a screening (elisa, ifa) and a neutralization assay [44] . since the emergence of mers-cov and till october 2015, 1715 cases were reported in different countries [45] . the cases included 1403 cases in the middle east, 15 cases in europe, 191 in asia, and 7cases in other countries [45] . the who international health regulations (ihr) emergency committee convened a mers-cov emergency committee meeting on multiple occasions; after extensive deliberations and reviews of available data, the diseases weakness did not fulfill the ihr requirements to be defined as a public health emergency of international concern (pheic) and mainly sustained human-to-human transmission [46, 47] (fig.2) . using a mathematical model, the risk of mers-cov was estimated to be one to seven cases per hajj and three to ten umrah pilgrims per year [48] . in another model, 6.2 pilgrims were estimated to develop mers-cov symptoms during the hajj, and 4.0 foreign pilgrims will be infected but return home before developing symptoms [49] . travelrelated mers-cov occurred infrequently among pilgrims performing the umrah [50] . however, millions of pilgrims who performed the annual hajj did not exhibit mers-cov symptoms [51] . a cross sectional study of 839 african hajj pilgrims returning to ghana, west africa in 2013 showed that none of the pilgrims was positive for mers-cov [52] . a cohort of french pilgrims exhibited no mers-cov infection [53] . no mers-cov was detected by pcr among 5235 adult pilgrims from 22 countries [54] . although the risk of travel-associated mers-cov remains low, the potential for healthcareassociated infections in relation with an imported mers-cov is a real concern. this event took place in the republic of korea [6] [7] [8] [9] . thus, all hcws should be vigilant to the importation of mers from returning travelers and healthcare organizations should implement a strategy to screen, isolate, and diagnose these patients. serologic testing allowed the detection of eight of the 124 contacts in the jordan cluster [30] . the positive results were confirmed in six of nine outbreak members, one in 26 household contacts, and one in 89 hcws [30] . interest-ingly, one hcw who tested positive did not recall having respiratory symptoms at the time of the outbreak [30] . a few published serology studies did not present any background on mers-cov. in one study, eight out of 356 abattoir workers and blood donors had weak positive tests by ifa, and none of these individuals tested positive by nt in jeddah and makkah in 2012 [55] . in a second study, none of 268 children with respiratory tract disease and blood donors showed neutralizing antibodies in dammam, ksa in 2010-2012 [55] . an initial study of 268 samples that were tested for mers-cov antibodies in the eastern saudi arabia revealed no positive samples [55] . a serologic evaluation of 280 household contacts of 26 index patients showed 12 probable cases of secondary transmission (4%; 95% confidence interval, two to seven) [56] . a large-scale study of more than 10 000 samples demonstrated that the overall seroprevalence in ksa was 0.15% and detected an increase of 17-and 26-fold of antibody detection rate among camel shepherds and abattoir workers compared to that in the general population [20] . the full genome sequences from mers patients with known dates and locations can help answer these questions: how fast does the virus change? when did the virus begin circulating in its current form? is the virus adapting to humans? can the geographical patterns help locate an animal source? what are the transmission patterns? a previous study showed that sequence success is as function of viral load, which is inversely proportional to the threshold cycle value (ct) in real-time pcr assays [25] . ct values below 33 are associated with good sequencing success rates. thus, ct values are sufficient predictors of the success that is independent on sample type or source [24, 25] . mers-cov was found to be stable in the environment. the virus can survive on plastic and steel for up to 48 h at lower temperatures and humidity; once temperature and humidity increase, the virus becomes less viable [57] . the virus is viable at 20°c and 40% humidity for 48 hours, at 30°c and 30% humidity for 24 hours and at 30°c and 80% humidity for 8 hours only [57] . in another study, increasing the temperature from 25°c to 65°c adversely affects viral infectivity [58] . the who advocates contact and droplet precautions with airborne isolation in hospital settings when dealing with an aerosol-generating procedure [56] ; the united states centers for disease control and prevention (cdc) and the european centre for disease prevention and control (ecdc) call for airborne infection isolation precautions [59] [60] [61] . multiple hospital outbreaks of mers-cov infection in ksa were controlled effectively using infection control precautions recommended by the who and the saudi ministry of health [4, 5, 26, 36] . there are no proven therapeutic agents for the treatment of mers-cov patients. existing antiviral agents can be repurposed against mers-cov [62, 63] . interferon and ribavirin were suggested to be possible therapeutic options based on the sars data [63] . these two agents were used to treat five patients with mers-cov infection [64] . the median time to therapy was 19 days, and no improvement was observed [64] . ribavirin and interferon were used in 20 patients with mers-cov at a median of three days [65] . the 14-day survival was 70% (14 of 20 patients) in the treatment group vs. 29% (7 of 24 of patients) in a historical group (p = 0.004) with no survival improvement at 28 days (30% vs. 17%; p = 0.054) [63] . in an observational study, interferon-α2a with ribavirin and interferon-β1a with ribavirin showed similar results in treating mers-cov [40] . the potential repurposed drugs for mers therapy include ribavirin with or without interferon, hiv protease inhibitors (lopinavir and nelfinavir), cyclophilin inhibitors (cyclosporin and alisporivir), chloroquine, mycophenolic acid, and nitazoxanide [66] . the use of interferon-α2b and ribavirin decreased viral replication in the rhesus macaques model within 8 h of mers-cov infection [67] . in vitro, ribavirin inhibits mers-cov; however, the required doses are extremely high to obtain in vivo [68, 69] . in vitro, nelfinavir and lopinavir achieved inhibitory concentrations against mers-cov [70] . in a primate model, the mortality rate at 36 h post-inoculation was 67% in untreated versus 0%-33% in the lopinavir-or ritonavirtreated and interferon-β1b-treated animals [71] ; the combination was used in another case [72] . to understand the mers-cov disease further, resolving the issues related to the specific host and the specific transmission mode and determining the factors that increase transmission within healthcare environments are crucial. the viral kinetics of mers-cov within different body compartments is another aspect that needs further examination. the optimal therapeutic options and strategies to predict the occurrence and severity of the disease require further analysis. jaffar a. al-tawfiq, ali s. omrani, and ziad a. memish declare that they have no conflict of interest. this manuscript is a review article and does not involve a research protocol requiring approval by the relevant institutional review board or ethics committee. travel implications of emerging coronaviruses: sars and mers-cov coronaviruses: severe acute respiratory syndrome coronavirus and middle east respiratory syndrome coronavirus in travelers middle east respiratory syndrome coronavirus (mers-cov) -kuwait an observational, laboratory-based study of outbreaks of middle east respiratory syndrome coronavirus in jeddah and riyadh mers-cov outbreak in jeddah-a link to health care facilities middle east respiratory syndrome coronavirus (mers-cov) -republic of korea south korea coronavirus mers case list-including imported and exported cases middle east respiratory syndrome 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infections: new drug development and therapeutic options treatment with interferon-α2b and ribavirin improves outcome in mers-covinfected rhesus macaques broad-spectrum antivirals for the emerging middle east respiratory syndrome coronavirus inhibition of novel coronavirus replication by a combination of interferon-α2b and ribavirin broad-spectrum antivirals for the emerging middle east respiratory syndrome coronavirus treatment with lopinavir/ritonavir or interferon-β1b improves outcome of mers-cov infection in a nonhuman primate model of common marmoset combination therapy with lopinavir/ritonavir, ribavirin and interferon-α for middle east respiratory syndrome: a case report key: cord-253452-feb75ei4 authors: yang, xingyue; liu, yali; hou, aijuan; yang, yang; tian, xin; he, liyun title: systematic review for geo-authentic lonicerae japonicae flos date: 2017-04-19 journal: front med doi: 10.1007/s11684-017-0504-0 sha: doc_id: 253452 cord_uid: feb75ei4 in traditional chinese medicine, lonicerae japonicae flos is commonly used as anti-inflammatory, antiviral, and antipyretic herbal medicine, and geo-authentic herbs are believed to present the highest quality among all samples from different regions. to discuss the current situation and trend of geo-authentic lonicerae japonicae flos, we searched chinese biomedicine literature database, chinese journal full-text database, chinese scientific journal full-text database, cochrane central register of controlled trials, wanfang, and pubmed. we investigated all studies up to november 2015 pertaining to quality assessment, discrimination, pharmacological effects, planting or processing, or ecological system of geo-authentic lonicerae japonicae flos. sixty-five studies mainly discussing about chemical fingerprint, component analysis, planting and processing, discrimination between varieties, ecological system, pharmacological effects, and safety were systematically reviewed. by analyzing these studies, we found that the key points of geo-authentic lonicerae japonicae flos research were quality and application. further studies should focus on improving the quality by selecting the more superior of all varieties and evaluating clinical effectiveness. lonicerae japonicae flos, also known as jin yin hua, is a dried flower of lonicera japonica thunb., which originates from east asia. in traditional chinese medicine (tcm), it is commonly used as anti-inflammatory, antiviral, and antipyretic herbal medicine for some diseases, such as acute tonsillitis, pneumonia, dysentery, and arthritis in clinic [1, 2] . according to statistics, lonicerae japonicae flos is one of the most frequently used herbs in the treatment and precaution of severe acute respiratory syndromes (sars) and influenza a virus a (h1n1), which are serious viral diseases [3] . lonicerae japonicae flos is also an economic plant with high value and is widely recognized as tea, food, and ornamental plant in asia and europe because of its unique aroma and beautiful flowers [4] . lonicerae japonicae flos is widely planted in china, but its quality is influenced by many factors, such as the product area, planting and processing, and ecological system. research showed that these herbs are different in qualities and morphological characteristics from different regions [5] . in tcm, geo-authentic herbs, which are planted in specific areas, are believed to present the highest quality among all samples from different regions, and they have been considered with appropriate and classic planting and processing; they have highly effective pharmaceutical use and contain the highest contents of active constituents [6] . currently, with the standardization of tcm, the quality of chinese herbs, including the planting and processing, contents of active constituents, and injurious ingredients, is becoming more and more important to control. therefore, the geo-authentic herb is becoming the hot research point. as for lonicerae japonicae flos, it has been confirmed that henan and shandong provinces in china are geoauthentic product areas [7] . the research about geoauthentic lonicerae japonicae flos is growing in recent years, but no systematic review exists. thus, in this article, we searched the studies about geo-authentic lonicerae japonicae flos to discuss the current situation and trend, to find out the difference between geo-authentic and non-authentic product, and to assess the quality of geo-authentic lonicerae japonicae flos. we planned to discuss the studies about geo-authentic lonicerae japonicae flos. the studies should meet the inclusion criteria of geo-authentic lonicerae japonicae flos (cultivated in shandong or henan province). additionally, reviews, news reports, or studies discussing other varieties, such as lonicera hypolauca miq, lonicera confusa dc, lonicera dasystyla rehd, or the main study objects that do not focus on geo-authentic lonicerae japonicae flos, were excluded. the following sources were up to november 2015. we searched four chinese databases, including chinese biomedicine literature database (cbm), chinese journal full-text database, chinese scientific journal full-text database, and wanfang database. the keywords were "jinyinhua," "shuanghua," "rendonghua," "erhua," "yinhua," or "rendong." moreover, we refined the titles or abstracts of these studies with the word "daodi" or "didao." we also searched the english databases, including pubmed, cochrane central register of controlled trials, and embase database. the main keywords were "lonicera japonica," "lonicerae japonicae flos," "jinyinhua," "shuanghua," "rendonghua," "erhua," "yinhua," or "rendong." we continued to refine these studies through the geo-authentic habitats. each database was used independently. for example, we searched cbm using "jinyinhua," "shuanghua," "rendonghua," "erhua," "yinhua," or "rendong" and "daodi" or "didao" in title/abstract/keyword/mesh. finally, we obtained 75 articles from this database. two reviewers assessed whether the studies should be accepted or excluded. both reviewers independently assessed studies, and disagreement was resolved by discussion or through the third reviewer. we made a data extraction table based on the detailed situations of studies regarding geo-authentic lonicerae japonicae flos. the table included the basic information of articles and the main information about geo-authentic lonicerae japonicae flos, such as chemical fingerprint, component, discrimination, pharmacological effects, planting, and processing. we accomplished the data extraction table after screening these articles. we performed a descriptive analysis as it was not possible to undertake a meta-analysis due to the heterogeneity of these studies. we classified all these studies into different parts according to the main discussing point, and in each part, we clearly listed the main point. we initially searched 363 potentially relevant articles. a total of 235 articles were excluded based on the headings and abstracts. of the 128 articles remained, 61 articles were excluded according to the inclusion and exclusion criteria. in this work, 67 articles were supposed to be reviewed; however, we could not access the full text of two articles. hence, we analyzed 65 articles in our review. data are shown in fig. 1 . only 1 article was published in english, whereas 64 articles were published in chinese. all of these studies were classified into 6 parts, based on the main idea of articles, mainly discussing the following points: chemical fingerprint, component analysis, planting and processing, ecological system, pharmacological effects and safety, and discrimination between different varieties. we also analyzed the years that these studies were published. only one study was published before 2000, and most of the studies regarding lonicerae japonicae flos were published between 2001 to 2015. data are presented in table 1 . from table 1 , we could see that there were 18 studies using chemical fingerprint to analyze the quality of lonicerae japonicae flos. among the 18 studies, hplc was the most commonly used method, accounting for 88%. many other methods were also used, such as gas chromatography, polyacylamide gel electrophoresis, infrared spectrum, rapid resolution liquid chromatography. most of these articles used these methods to analyze the content of main components, including chlorogenic acid and galuteolin; these were also used to form the standard chemical fingerprint. the details are shown in table 2 . a total of 12 studies mainly discussed the component of geo-authentic lonicerae japonicae flos. we put emphasis on the main component, chlorogenic acid, and galuteolin. out of the 12 studies analyzed, 10 showed chlorogenic acid content, and only 2 of them showed galuteolin. other components, such as volatile oil, heavy mental, and total ash, were also accessed in some studies. the details are shown in table 3 . twelve studies discussed the planting and processing of geo-authentic lonicerae japonicae flos. two studies [37, 38] investigated the germplasm resources from the main producing areas of lonicerae japonicae flos in china. through the investigation, they analyzed the existent problems about producing areas and germplasm resources, including aimless introduction of new strain, lack of strain selection technology, and unreasonable management. they also provided some suggestions and methods, such as setting standards, assessment, and screening of good strain. four studies [39] [40] [41] [42] discussed the processing methods of geo-authentic lonicerae japonicae flos. wang et al.'s research [39] indicated that different plucking time and processing methods made a great effect on the quality of lonicerae japonicae flos through measuring the content of chlorogenic acid. to make sure of the high effectiveness, the more appropriate time to pluck lonicerae japonicae y indicates the study included in the current research on this point; hplc, high-performance liquid chromatography; gc, gas chromatography; page, polyacylamide gel electrophoresis; tlc, thin-layer chromatography; ir, infrared spectrum; hnmr, nuclear magnetic resonance spectra; rapd, random amplified polymorphic dna; hs-gc-ms, headspace solid-gas chromatography-mass spectrometry; issr, inter-simple sequence repeat; rrlc, rapid resolution liquid chromatography. flos was at the beginning of may when the flowers start to bloom and then dried in the sun or placed in a stove after drip washing. additionally, other research [43] [44] [45] [46] suggested that the plucking time and processing methods, influenced by the natural and man-made factors, determined the quality and production of lonicerae japonicae flos; they also discussed the standard operation procedure of planting, including the technical requirements for the production suitability, growth, field management, diseases, and pest control, picking and processing, packing and storing, transporting, and quality monitoring of lonicerae japonicae flos. another two articles [47, 48] reported the hybridization and induction of allotetraploid of lonicerae japonicae flos and culturing in vitro to satisfy the need of modern market and make a foundation to generalize new varieties and rapidly propagate in vitro through biotechnology and industrial production. five studies mainly discussed the morphological characteristics discrimination of geo-authentic lonicerae japonicae flos. we could see from these studies that they observed the leaves and buds, such as exterior of petal and corolla, pollen grain seen under the microscope, the skin hair of buds, and the length or width or diameter or number of the leaves and buds. the details are shown in table 4 . three studies discussed the heredity of lonicerae japonicae flos using different methods. to study the formation of geoherbalism of lonicerae japonicae flos on the gene level, li et al. [54] extracted genomic dnas from different populations, amplified, and sequenced the 5s-rrna gene spacer region. they found that the sequences were different between genuine and ungenuine drugs and could be identified by sequencing. sun et al. [55] and han et al. [56] used the intersimple sequence repeat (issr) to analyze genetic polymorphism of lonicerae japonicae flos, providing evidence of molecular biology for the genetic resources, genetic relationship, and planting. the results showed that there were obvious genetic differences among regions and plant sources. in the research of kang [21] , except the hplc fingerprint, he also discussed the genetic diversity using issr, revealing that geographical factors had a significant effect on the variety of lonicerae japonicae flos. others hplc, page, and rapd were used to compare the content of main available components, isozyme, and gene sequence to offer scientific basis for the discrimination of different varieties of lonicerae japonicae flos. the amino acid content and extracorporeal bacteriostatic effectiveness were also compared [57] . li et al. [58] used electron probe method to analyze the element weight in the cell of two different species to identify the quality of lonicerae japonicae flos and distinguish the geo-authentic and non-authentic species. the results showed that the method of electron probe was effective in appraising and distinguishing the quality of lonicerae japonicae flos because it could be used to distinguish the content of elements in the cell of different lonicerae japonicae flos, which was controlled by genetic factor. wang et al. [59] used restriction fragment length polymorphism analysis to confirm geographical authenticity. the results showed that the cleavage rate of pcr products by econ i correlated remarkably with the geographical origin of lonicerae japonicae flos. therefore, this method could also be used to classify geoauthentic lonicerae japonicae flos. wang [60] sought for further research on shandong provincial lonicerae japonicae flos cultivars, and three cultivars, jizhuahua, maohua, and sijihua, were studied on original, macroscopical, and microscopical identification, hplc fingerprint atlas, and rapd fingerprint atlas. the results showed that these methods could distinguish the characteristic identification of three cultivars. five articles discussed the ecological system, including the geological background and soil characteristics. detailed points, such as longitude, latitude, altitude, geology, soil type, vegetation regionalization, climatic elements (annual average temperature, rainfall, sunshine), soil texture, trace element, ph, organic material, and base saturation, were mentioned. some studies analyzed the component and economic benefits of geo-authentic lonicerae japonicae flos to investigate the effect of ecological system. the details are shown in table 5 . two of these studies were randomized clinical trials [66, 67] , investigated the clinical effectiveness for gum inflammation and oral ulcer of honeysuckle from geo-authentic product area and sichuan province, and the results showed that the effects of authentic production were better than that from sichuan province in the treatment of mouth ulcers and gum inflammation. however, the problems were that these two studies did not clearly express the randomized and blinded methods, inclusive criteria, and efficacy assessment standards. thus, the qualities of these two studies were low. furthermore, li et al. [68] found that the scavenging capacities on free radicals, o -2 , $oh, and h 2 o 2 varied from locality and species and were highly relative to the contents of phenolic acids. as the same species, the scavenging capacities of the samples from henan and shandong provinces were stronger than those from jiangsu province. lei [69] found that antibacterial effect of lonicerae japonicae flos from shandong province (ji) was lower than that from henan province (mi). lonicerae japonicae flos had certain antipyretic effect to fevered rat caused by fresh cerevisiae. on the other hand, he also discovered that l. macranthides hand-mazz's ld50 was higher than lonicerae japonicae flos, which meant that l. macranthides hand-mazz's had less toxicity than lonicerae japonicae flos. results of the study could also be confirmed in other articles [70, 71] . lonicerae japonicae flos is widely used clinically, and the application of geo-authentic herbs has always been paid attention by clinical experts. many experts focus on geoauthentic lonicerae japonicae flos because of its higher quality and better clinical effect than non-authentic product. the chinese herbs are used to prevent and treat diseases; thus, the quality of herbs directly influences the clinical effectiveness and even sometimes affects the patients' lives to some extent. it is an important factor to the inheritance and development of tcm. thus, most of the studies on geo-authentic lonicerae japonicae flos focus on its quality. among the articles we searched, 18 studies focused on the chemical fingerprint explored the methods to access the quality of geo-authentic lonicerae japonicae flos. additionally, combined with the time these studies published, it indicates that chemical fingerprint is becoming a hot topic in recent years. it will provide significant scientific basis for assessment and control of the quality of geo-authentic lonicerae japonicae flos to make a standard chemical fingerprint. the chemical fingerprint of chinese herbs is a method used in modern technology under given conditions to analyze the chemical and biological information and expressed in image mode. chemical fingerprint not only emphasizes the uniqueness of certain herbs but also reflects the similarity of different varieties. it has great advantages with respect to monitoring of the quality of chinese herbs, and some developed countries have regarded it as standard of quality control. at present, the commonly used methods are hplc, ir, gc, rrlc, rapd, and hnmr. among these methods, hplc has the most advantages, such as high separation efficiency, high sensitivity, fast analysis speed, and load liquid velocity, which is the main method used in the component analysis and fingerprint creation of geo-authentic lonicerae japonicae flos. these studies show that it is reasonable to distinguish geo-authentic and non-authentic lonicerae japonicae flos. analyzing all of these studies, the chemical fingerprint is the most often used method to qualitatively analyze the component content, focusing on the content of chlorogenic acid and galuteolin, which accords with the component analysis method in chinese materia medica grand dictionary and quantitatively establish the standard chemical fingerprint. one reason for the high quality of geo-authentic lonicerae japonicae flos is that its active component is high and stable. the active component is also the basis of quality standard of chinese herbs. chinese materia medica grand dictionary explains the content of chlorogenic acid, galuteolin, and noxious substances. in the research that focuses mainly on the component analysis, they pointed at the content of chlorogenic acid, which was also the main analyzing component in chemical fingerprint, ecological system, and other research. chlorogenic acid is the main active component that confirmed as an anti-inflammatory agent. flavonoid compounds, including galuteolin, are antioxidants that can remove free radicals of ultra oxygen ions in the human body, increase immunity, and has anti-aging effect. in chinese materia medica grand dictionary, it also refines the content of noxious substances, such as heavy mental and residual insecticides. the high content of these noxious substances is also an important reason limiting international popularization and application of chinese herbs. however, few studies involved the assessment of these noxious substances. thus, further studies should improve this aspect. the content of water, total ash, and volatile oil are also seldom involved, so the studies about geo-authentic lonicerae japonicae flos should also stress this point. the planting and processing of lonicerae japonicae flos is also an important factor influencing its quality. the geo-authentic lonicerae japonicae flos with appropriate planting and processing has higher quality, but the quality and yield is unstable. therefore, the major task is to form the scientific management and normalized planting and processing to improve the yield and effectiveness and diminish drug toxicity. from the studies we have searched, we can see that many studies have begun to explore the standardization of planting and processing of geo-authentic lonicerae japonicae flos. the research indicates that the best picking time is sanqing, erbai, and dabai stage, just before the beginning of summer. these herbs are directly dried in the sun or placed in a stove, which is a better processing method compared with other methods. this method will increase the yield and the content of the active component. ecological system environment, such as geology, geomorphology, soil types, and climate factors, is another important factor that influences the herbal quality. in tcm, henan and shandong provinces are recognized as the authentic product areas. analysis of the ecological system environment of these two provinces showed that they both are located in the 34â°-35â°n, 113â°-117â°e; the altitude height is 100-800 m; the soil types and geology are similar; the climate factors, such as annual sunshine, annual rainfall, and climate type, are almost the same. the accumulation of active components is closely related with ecological conditions, and the content of geo-authentic lonicerae japonicae flos is highly above non-authentic product. by comparing the geo-authentic and nonauthentic lonicerae japonicae flos, we found that it is sunshine rather than soil style that may be the critical factor that affects the active components. the geo-authentic lonicerae japonicae flos, through long-term natural and artificial selection, has made obvious variations and formed different varieties in authentic product areas. the different varieties show diversity in morphological characteristics and heredity. the most common varieties are damaohua, xiaomaohua, dajizhua, and xiaojizhua. first, the varieties of the geo-authentic lonicerae japonicae flos should be discriminated. different organs are differently influenced by the environment. the research shows that the color, morphology of buds and leaves, density, and length of non-glandular hair outside the buds are the discrimination point between different varieties. second, different varieties have different heredity features, which can be discriminated through technologies. from the inclusive research, we can see that the issr is the most commonly used method recently. issr combines rapd and ssr labeling technique and has advantages such as simple, rapid, and efficient operation, high genetic diversity, and good repeatability. issr can provide more genome dna information and is often used in germplasm resource discrimination, evolutionary and genetic relationship analysis, and genetic diversity research. other technologies, such as hplc, electron probe, rapd fingerprint, and restriction fragment length polymorphism analysis, are also used. the aim of the discrimination of morphological characteristics and heredity is to select the superior quality of geo-authentic lonicerae japonicae flos; however, until now, no study reports the mode of selection for superior quality and the method to improve the quality. thus, this point should be strengthened. the clinical effectiveness of geo-authentic lonicerae japonicae flos is superior than that of non-authentic in tcm theory. however, scientific data to support it are lacking as we can see little evidence from the studies we searched. the qualities of these two clinical trials are too low, and the studies about its anti-inflammation and antipyretic pharmacologic action are still with animals rather than clinical application. thus, we should make further investigation to explore the clinical effectiveness of geo-authentic lonicerae japonicae flos. for example, high-quality clinical trials can be conducted to compare the clinical effectiveness between geo-authentic lonicerae japonicae flos and non-authentic ones. furthermore, geoauthentic herbs that target illness are also important to control quality or develop chinese herb extracts. briefly, research shows that geo-authentic lonicerae japonicae flos is mainly characterized by chemical fingerprint, component analysis, planting and processing, discrimination between varieties, ecological system, pharmacological effects, and safety and focuses on the quality assessment technology. the factors involve yield and quality. the key point of geo-authentic lonicerae japonicae flos research is the quality and application. however, we are facing many problems such as identifying noxious substances of lonicerae japonicae flos, selecting the most superior of all varieties, and evaluating clinical effectiveness. therefore, further studies should focus on resolving these problems and focus on the application and further development of geo-authentic lonicerae japonicae flos. separation of chlorogenic acid from honeysuckle crude extracts by macroporous resins lonicera japonica thunb.: ethnopharmacology, phytochemistry and pharmacology of an important traditional chinese medicine research and comprehensive utilization of honeysuckle analysis of chlorogenic acids in beverages prepared from chinese health foods and investigation, in vitro, of effects on glucose absorption in cultured caco-2 cells rapid quantitative analysis of adulterant lonicera species in preparations of lonicerae japonicae flos the characteristics of geoauthentic herbs comparative study on quality of flos lonicerae between geoauthentic and non-authentic producing areas finger print studies on flos lonicerae and its preparation study of the fingerprints atlas on multi-elements and multi-data of flos lonicera japonica thunb study of flos lonicerae in local herbs and other areas by ftir spectrum geo-authentic medicinal in shandong-flos lonicera's quality inspect. food science (shi pin ke xue) studies in pharmacognosy and hplc fingerprint chromatogram of the chinese herbal medicine flos lonicerae study on hplc fingerprint chromatogram of shandong trueborn flos lonicerae establishment fingerprint of the flower of lonicera japonica thunb. and salvia miltiorrhiza bge with gc and hplc study on the key technology of quality control and quality evaluation system of flos lonicerae application and study of hplc fingerprint of lonicera japonica for identification of geo-authenticity study on hplc fingerprint chromatogram of shandong trueborn flos lonicerae. the first honeysuckle peak bbs application study on the modern quality control of flos lonicerae from shandong province the key technology of quality control of flos lonicerae. the first honeysuckle peak bbs lonicera japonica thunb. germplasm resources evaluation and honeysuckle genuine chemical fingerprint study on the genetic diversity by issr maker and the hplc fingerprint of jizhao strain of lonicera japonica chemical fingerprint chromatogram of henan trueborn flos lonicerae study on hplc fingerprint of lonicera japonica thunb. from henan genuine study on rrlc fingerprint of flos lonicerae from shandong genuine producing areas. communication conference about development of crude drug quality control technology and resources and utilization study of lonicerae japonicae flos from genuine producing areas of shandong on rapid resolution liquid chromatography fingerprint research on different phenological phase of total dynamic accumulation of chlorogenic acid in flos lonicerae comparing about different producing area of flos lonicerae naphtha using gc-ms analysis determination of chlorogenic acid in 5 species and one variety of flos lonicerae by hplc study oil variation of chlorogenie acid in flos lonicerae with phenoiogieal phases and localities comparative study on quality of flos lonicerae between geo-authentic and non-authentic producing areas. the seminar of standardized cultivation research determination of the organochlorine insecticide residues in lonicera japonica thunb. by gas chromatography the quality control of flos lonicerae planted in henan province determination of acid ingredients and content variations in flos lonicerae at different time by hplc-esi-ms. chin pharm (zhongguo yao fang) study on the correlation between endophytic fungus with metabolites in lonicera japonica thunb active ingregients of shandong famous region drug extraction isolation and identification principal component analysis and cluster analysis of inorganic elements in lonicerae japonical flos from different areas the investigation of the honeysuckle germplasm resources in shandong investigation on the germplasm resources for main producing area of flos lonicerae japonicae in china the quality evaluation of the different collection periods and processing technologies of flos lonicerae in henan processing and storage of honeysuckle harvesting and modern processing of honeysuckle investigation and analysis about processing method of some geo-authentic herbs in henan. herb processing association of china association of the standardization of planting of shu and mi flowers (sop) the standardization of planting of flos lonicerae. spec econ anim plant standard operating procedure for raw materials of reduning injection-lonicerae japonicae flos. chin tradit herbal drugs study on effects of soil and water and fertilizer on the quality of honeysuckle hybridization and induction of allotetraploid of lonicera japonica research on medicinal herbs of yu honeysuckle in vitro culture technology studies on the floral morphology of flos lonicerae studies on the morphology of flowering bud of lonicera japaoncia thunb. and their singificnace in division of germplasm comparative study on botanical morphological character of germplasm resources of lonicera japonica thunb comparison of leaf shape characters in lonicera japonica cultivation varieties study on characteristics of non-glandular hairs of cultivated lonicera japonica preliminary attempt to identify geoherbalism of flos lonicerae by sequence divergence of 5s-rrna gene spacer region. chin tradit herbal drugs issr analysis on genetic diversity of germplasm resource of lonicerae flos issr-based genetic diversity of lonicera japonica in authetic region varieties and quality evaluation of shandong honeysuckle cultivars damao and jizhua flower application of electron probe method on quality identification of flos lonicerae discrimination of lonicera japonica thunb. from different geographical origins using restriction fragment length polymorphism analysis the identified research of three cultivars of flos lonicerae japonicae of the famous-region chinese materia medica in shandong province research on the relationship between genuine and the environmental factors of honeysuckle. the 5th natural medicinal material resources seminar analyses on the trace elements of soils in geo-authentic and non-authentic production areas of flos lonicerae analysis on the geologic background and physicochemical properties of cultivated soil of flos lonicerae in the geo-authentic and non-authentic producing areas the correlation between quality of flos lonicerae and ecosystem ecological adaptation of five different germplasm of honeysuckle in the real traditional production areas of henan the influence of varieties of flos lonicerae for clinical application clinical efficacy influence of different varieties of honeysuckle on gum inflammation and oral ulcer scavenging capacities on radicals of flos lonicerae by chemiluminescence comparative study on the pharmacological action and safety between lonicera macrathodes hands mazz. and the certified flos lonicerae comparative experiments on antipyretic effect between lonicera macrathodes hands mazz. and the certified flos lonicerae comparative experiments on safety between lonicera macrathodes hands mazz. and the certified flos lonicerae this study was supported by a grant from program for state key laboratory breeding base of dao-di herbs (no. z0310-3). xingyue yang, yali liu, aijuan hou, yang yang, xin tian, and liyun he declare that they have no conflict of interest. this manuscript is a systematic review article and does not involve a research protocol requiring approval by the relevant institutional review board or ethics committee. key: cord-287222-wojyisu0 authors: zhou, min; zhang, xinxin; qu, jieming title: coronavirus disease 2019 (covid-19): a clinical update date: 2020-04-02 journal: front med doi: 10.1007/s11684-020-0767-8 sha: doc_id: 287222 cord_uid: wojyisu0 coronavirus disease 2019 (covid-19) caused by severe acute respiratory syndrome coronavirus-2 (sars-cov-2) has posed a significant threat to global health. it caused a total of 80 868 confirmed cases and 3101 deaths in chinese mainland until march 8, 2020. this novel virus spread mainly through respiratory droplets and close contact. as disease progressed, a series of complications tend to develop, especially in critically ill patients. pathological findings showed representative features of acute respiratory distress syndrome and involvement of multiple organs. apart from supportive care, no specific treatment has been established for covid-19. the efficacy of some promising antivirals, convalescent plasma transfusion, and tocilizumab needs to be investigated by ongoing clinical trials. currently, coronavirus disease 2019 (covid-19) poses a significant threat to global health. world health organization (who) has declared this outbreak as a "public health emergency of international concern" on january 31, 2020. within the first two months of the outbreak, the epidemic spread rapidly around the country and the world. as of march 8, 2020, a total of 80 868 confirmed cases and 3101 deaths had been reported in chinese mainland by national health commission of china, and 90 other countries are affected. covid-19 as an emerging disease, has unique biological characteristics, clinical symptoms, and imaging manifestations, though considerable progress has been made on the clinical management. this article will summarize the epidemiological, etiological, clinical, pathological, and radiological characteristics of covid-19 and review the latest advancements in the treatment. epidemic curves reflect that this epidemic may be a mixed outbreak pattern, with early cases suggestive of a continuous common source, potentially at huanan seafood wholesale market (hswm), and later cases suggestive of a propagated source as the virus began to be transmitted from person to person [1] . a retrospective analysis on the first 425 patients with confirmed covid-19 showed that during the early stages of this outbreak, the basic reproduction number r 0 was estimated to be 2.2 [2] . another modeling study estimated that the r 0 for covid-19 was 2.68 [3] . considering the strict prevention and control measures implemented by the chinese government, a phase-adjusted estimation of epidemic dynamics assumed that the effective reproduction number r 0 was 3.1 at the early phase of the epidemic, and could be gradually decreased [4] . of the first 99 laboratory-confirmed patients, 49 (49%) had been exposed to hswm, which was reported to be the possible initial source of severe acute respiratory syndrome coronavirus-2 (sars-cov-2) [5] . a shenzhen family cluster without exposure history to wuhan markets or wild animals also proved the possibility of person-to-person transmission [6] . another family cluster of patients provided evidence that asymptomatic carriers may also be potential sources of sars-cov-2 infection [7] . evidence has recently been obtained to suggest transmission along a chain of 4 generations [8] . sars-cov-2 spread mainly through respiratory droplets or close contact. while in the later stage of infection, the virus is also detectable in anal swabs, suggesting the possibility of oral-fecal route transmission [9] . significant environmental contamination by patients carrying sars-cov-2 through respiratory droplets and fecal shedding suggests that the environment serves as a potential medium of transmission and supports the requirement for strict adherence to environmental and hand hygiene [10] . currently, there is no clear evidence of infection caused by vertical transmission or aerosol transmission. sars-cov-2 is the causative pathogen of covid-19, identified as the seventh type of coronavirus to infect humans [11] . six other kinds of coronaviruses are known to cause human disease, including severe acute respiratory syndrome coronavirus (sars-cov) and middle east respiratory syndrome coronavirus (mers-cov) with high mortality rate [12] . according to the genome characteristics, coronavirus is separated into four genera: α-cov, β-cov, γ-cov, and δ-cov [12] . deep sequencing revealed that this novel coronavirus isolated from lower respiratory tract samples of patient with covid-19 belongs to β-cov [11] . coronavirus has the appearance of crown under electron microscopy. they are enveloped viruses with a singlestrand, positive-sense rna genome, which is the largest known genome for an rna virus [13] . all coronaviruses share the same genome organization and expression pattern, with two large overlapping reading frames (orf1a/b) which encode 16 nonstructural proteins, followed by orfs for four major structural proteins: spike (s), envelope (e), membrane (m), and nucleocapsid (n) [13] . the sars-cov-2 protein also contains eight accessory proteins [14] . spike protein plays an essential role in binding to receptors and is critical for determining host tropism and transmission capacity. it is functionally divided into s1 domain and s2 domain, responsible for receptor binding and cell membrane fusion respectively. the receptor binding domain (rbd) of β-cov is commonly located in the c-terminal domain of s1 [15] . a team analyzed the cryogenic electron microscopy (cryo-em) structure of the sars-cov-2 spike protein and found that it has 10 to 20-fold higher binding affinity to human angiotensin-converting enzyme 2 (ace2) than sars-cov does [16] . phylogenetic analysis of the evolution history showed that sars-cov-2 shared a closer sequence homology toward the genomes of sars-cov than to that of mers-cov [17] . sars-cov-2 is highly similar to a bat coronavirus ratg13, with an overall genome sequence identity of 96.2% [18] , indicating that bat, which was discovered to be the natural reservoir host of various sars-related coronaviruses [19] , may also be the original host of sars-cov-2. the intermediate host in the process of transmission remains uncertain. clinical manifestation covid-19 has an incubation period of 1-14 days, mostly ranging from 3 to 7 days [20] . the most common symptoms in mild to moderate patients are fever, fatigue, and dry cough, followed by other symptoms including headache, nasal congestion, sore throat, myalgia, and arthralgia [5, [21] [22] [23] . a minority of patients had gastrointestinal symptoms, such as nausea, vomiting, and diarrhea, especially in children. in the study of 1099 covid-19 patients, 43.8% cases presented fever at onset of illness and the percentage further increased to 88.7% during following hospitalization [24] . notably, fever may occasionally be absent from elderly persons or immunocompromised ones. a part of patients may progress to shortness of breath, usually in the second week of the illness, and might be accompanied by or progress to hypoxemia [25, 26] . for patients presenting tachypnea, chest indrawing, or inability to feed or drink, severe pneumonia should be considered. in 10% to 20% of severe patients, the respiratory injury will inevitably develop into acute respiratory distress syndrome (ards) during 8-14 days of the illness, defined as partial pressure of oxygen (pao 2 ) to fraction of inspired oxygen (fio 2 ) ratio lower than 300 mmhg, as well as resultant non-cardiogenic pulmonary edema and mechanical ventilation [24, 25, 27] . ards, as the main cause of respiratory failure, is associated with high morbidity and mortality. risk factors for developing into severe to critical cases include advanced age, underlying comorbidities such as hypertension, diabetes, cardiovascular disease, and cerebrovascular disease [23, 25, 26] . as disease progresses, a series of complications tend to occur, especially in critically ill patients admitted to icu, including shock, sepsis, acute cardiac injury, acute kidney injury, and even multi-organ dysfunction [23, 24, 26] . patients may manifest altered mental status, low oxygen saturation, reduced urine output, weak pulse, cold extremities, low blood pressure, and mottled skin. besides, patients with acute cardiac injury would present tachycardia or bradycardia. critically ill ones may also suffer acidosis and increased lactate [23] [24] [25] . current studies reported the peak value of temperature in non-survivors of covid-19 was significantly higher than that in survivals during hospitalization [23, 24] . thus, patients presenting hyperthermia and chill should exclude the possibilities of co-infection with bacteria or other pathogens. attentions should be paid to prevent hospital-acquired pneumonia (hap) in critical cases and ventilator-associated pneumonia (vap) in those receiving mechanical ventilation. coagulopathy and thrombocytopenia are also common complications for covid-19 infection, which increase the risk of hemorrhage and thrombosis. mottled skin, petechial or purpuric rash, appearance of black stool or hematuresis could be found in some cases. patients with the syndrome of persistent hypoxemia, chest pain, pre-syncope or syncope, and hemoptysis should be suspected of having pulmonary thromboembolism (pte) [28] . the manifestation of limb pain, swelling, erythema, and dilated superficial veins should be suspected of deep vein thrombosis (dvt). nearly 20% of patients had abnormal coagulation function, and most of severe and critical patients presented coagulation disorders and had the tendency to develop into disseminated intravascular coagulation (dic) [5, 25, 26] . in the early stage of infection, the involved lung lobe presented obvious alveolar edema, proteinaceous exudates, and reactive pneumocyte hyperplasia, accompanied by mild inflammatory infiltration [29] . on gross examination, the whole lung showed bronzing surface and diffuse congestive appearance, with partly hemorrhagic necrosis, as same as the cut surfaces. on histological examination, the typical manifestations were extensive proteinaceous and serous exudation in the alveolar, hyaline membrane formation, and inflammatory infiltration with multinucleated syncytial cells. type ii alveolar epithelial cells showed extensive hyperplasia, and some presented necrosis and desquamation. viral inclusions could be identified in epithelium and macrophage. besides, alveolar septal vessels manifested congestion with alveolar edema. the infiltration of monocytes and lymphocytes in alveolar cavity and microthrombosis were prominent. some parts showed alveolar exudate organization and pulmonary interstitial fibrosis. with a fraction of desquamation of mucosal epithelium, bronchi were covered by mucus even mucus plug [20, 30, 31] . in addition, other organs also suffered pathological damage to some extent [20] . the atrophic spleen showed significantly reduced lymphocytes, focal hemorrhage and necrosis, and macrophage hyperplasia. with degeneration and necrosis of cardiomyocytes, a small number of monocytes, lymphocytes, and/or neutrophils were infiltrated in the myocardial interstitium. protein exudation was seen in renal glomerulus and within hyaline cast, and renal tubular epithelium degenerated and desquamated. besides, hepatocytes degeneration, necrosis, and inflammatory infiltration also occurred. the brain presented congestion, edema, and degeneration of some neurons. meanwhile, microthrombosis could be found in multiple organs. radiological images play an important role in the diagnosis and providing guidance for treatment. guan et al. found that 86.2% of patients manifested abnormalities in chest ct images, of whom more than 75% had bilateral lung involvement, mainly with peripheral and diffused distribution [24] . patients of different severity presented significant different lesions on chest ct (fig. 1) . mild patients manifested unilateral and focal ground-glass opacity (ggo) which gradually develops to bilateral or multilobular lesions. as the disease progressed further, ggos evolved to consolidation lesions, presenting mixpattern or pure consolidation, with the latter being more common in critically ill patients admitted to icu [5, 25, 32] . consistent with the interstitial involvement in viral pneumonia, zhao et al. suggested that 48.5% of ct images manifested reticular patterns, and 28.7% presented interlobular septal thickening [33] . unlike influenza pneumonia, which usually exhibited unilateral ggo and significant solid nodules, only 6% of covid-19 patients had solid nodules [32, 34, 35] . moreover, other lesions included adjacent pleura thickening, vascular enlargement, bronchial wall thickening, traction bronchiectasis, air bronchogram, pericardial effusion, etc. [32, 33, 36] . follow-up of ct scan could help to monitor disease changes and evaluate therapeutic effects [32] . some dynamic images fluctuate repeatedly, and showed coexistence of absorption of primary lesions and emergence of new ones. during disease deterioration, increased number of or enlarged lesions could be observed in radiological imaging, and part of them even developed into a "white lung" with diffusely involved lung [37] . a majority of covid-19 patients showed normal leucocyte count, and nearly one third had leucopenia [21, 24] . lymphocytopenia, as one of the most typical laboratory abnormalities, was present in 83.2% of patients, with an even higher proportion in severe ones [24, 26] . in addition, previous studies also revealed that increased ddimer level and prolonged prothrombin time were also common features of covid-19, especially for severe patients [24] [25] [26] . meanwhile, sars-cov-2 might damage liver and myocardium to some extent, showing elevated levels of aminotransferase, creatine kinase, and myoglobin with diverse degrees, as well as increased troponin in critical patients [5, 23, 25] . a few patients had renal dysfunction, presenting increased serum creatinine or blood urea nitrogen [5] . as for infection-related serum biomarkers, our studies have reported that most of patients had increased concentration of c-reactive protein, interleukin-6 (il-6), and erythrocyte sedimentation rate [5] . likewise, huang et al. observed similar phenomenon and proposed that icu patients might suffer severe cytokine storms, with a overproduction of il-7, il-10, gcsf, ip10, mcp1, mip1a, and tnf-α, etc. [25] . multi-drug resistant acinetobacter baumannii and klebsiella pneumoniae have been isolated in covid-19 patients [5, 23] . other identified microorganisms included pseudomonas aeruginosa, aspergillus flavus, aspergillus fumigatus, candida albicans, and candida glabrata [5, 23] . laboratory confirmed covid-19 patients had positive results on real-time reverse transcriptase polymerase chain reaction (rt-pcr) of nasal and pharyngeal swab, sputum, blood, faeces, and urine specimens [25] . the collected clinical specimens need to be transported to designated laboratories promptly, and extracted for rna correctly, followed by rt-pcr detection with primers and probes of appropriate sequences [25] . the value of cycle threshold (ct) was the criterion to determine the detection result, with less than 37 being defined as negative, above 40 as positive and a medium load (37) (38) (39) (40) calling for confirmation by retesting [2] . the detection of sars-cov-2 specific igm and igg antibodies can also be used for diagnosis [20] . covid-19 infection could be determined with one of the following criteria: positive specific igm, the transformation of specific igg from negative to positive, a 4-fold increase in igg titer during recovery period compared with the result of acute phase. although antibody detection was simple, rapid, and inexpensive, it is still not widely used due to inherent limitations, for example, false-negativity resulted from the existence of window period, noncomparable sensitivity and specificity with pt-pcr, absence of exclusion criteria making it a diagnosis tool only. these is no specific antiviral treatment which has been proven to be effective for covid-19. combinations of over three antivirals are not suggested. current treatment options are mainly based on previous experience showing clinical benefits in treating influenza, ebola, mers, sars, and other viral infections. it is reported that most of covid-19 patients received antiviral therapy in china [5, 21, 25] , and here we will introduce some commonly used drugs. ribavirin is representative of nucleoside analogs. the combination of ribavirin and recombinant interferon, a broad spectrum antiviral agent, showed augmentation effect in inhibiting mers-cov replication and reduced doses of both ribavirin and interferon [38] . however, most of clinical experiences in mers patients come from limited case reports and observational studies, making the quality of evidence for ribavirin and interferon treatment efficacy very low [39] . it is recommended to administer ribavirin by intravenous infusion in combination with inhaled interferon-α or oral lopinavir/ritonavir in the 5th version guideline on covid-19 diagnosis and treatment issued by chinese national health commission [20] . notably, ribavirin is not suggested by military medical team coming to hubei [40] and interferon-α inhalation is worried to increase the risk of virus-containing aerosol production and airway stimulation. lopinavir/ritonavir is a combination of a protease inhibitor and a booster used for the treatment of human immunodeficiency virus (hiv) infection. currently, randomized controlled trials for the efficacy of a combination of lopinavir/ritonavir with interferon-α in mild to moderate patients (chictr2000029387) and severe to critical patients with covid-19 (chictr2000029308) are in progress. remdesivir, a novel nucleotide analog rna polymerase inhibitor, is considered as the most promising antiviral drug for the treatment of covid-19. it showed broad spectrum antiviral activities, from inhibition of human and zoonotic coronavirus (including sars-cov-2 [41] as well ebola virus) in vitro, to prophylactic and therapeutic effects in animal model of mers-cov and sars-cov infection [42, 43] . the first covid-19 patient identified in the united states was given remdesivir without obvious adverse reactions. two trials on efficacy of remdesivir have been launched in china among mild to moderate patients (nct04252664) and severe to critical patients (nct04257656) infected with sars-cov-2. neuraminidase inhibitors (nais), such as oral oseltamivir and intravenous peramivir, showed substantial clinical improvement in treating influenza patients [44] . oseltamivir was widely used for suspected and confirmed covid-19 patients in china [26] , however, there is no exact evidence that supports its application. a research team from zhejiang university reported that abidol has the potential to inhibit sars-cov-2, which was previously used for influenza. there is a multicenter, randomized, and controlled trial (chictr2000029573) to evaluate the efficacy of abidol and lopinavir/litonavir, either alone or in combination with a new type of interferon, novaferon. according to current who interim guidance on covid-19 management [27] , corticosteroids were not recommended as routine therapy unless indicated for another reason, because possible harms and higher risk of mortality attributed to corticosteroids therapy have been identified by studies on other coronaviruses and influenza. an epidemiological study conducted in wuhan observed a larger percentage of patients receiving corticosteroids in icu groups when compared with non-icu groups (6 (46%) vs. 3 (11%); p = 0.013), while we still cannot determine the effects of corticosteroids due to the limited sample size [25] . according to the latest guidelines issued by national health commission of china (version 7) [20] and the interim guidance of who [27] , when sars-cov-2 infection is suspected, corticosteroids should be recommended to use with caution. new coronavirus infection diagnosis and treatment scheme (trial version) published by military support hubei medical team also put forward that for mild to moderate covid-19 patients, corticosteroids should not be given principally and highdose corticosteroid pulse therapy was not recommended. only patients presenting ongoing deterioration in oxygenation index, or rapid progression of radiological findings, or excessive activation of immune responses, will be considered to use short-term corticosteroid therapy within 10 days of illness onset. seven designated hospitals in zhejiang province gave patients corticosteroids when they showed increased resting respiratory rate ( > 30 breaths/ minute), drop in oxygen saturation ( < 93%) on room air, or multi-lobular progression ( > 50%) on imaging within 48 h [21] . timely and appropriate use of corticosteroids combined with ventilator support should be considered for severe patients to prevent progression to ards [30] . the pharmacologic use of corticosteroids in covid-19 treatment should vary with severity [20, 40] . for severe cases, it is suggested to start at a dose of 40 to 80 mg/day methylprednisolone and slowly taper over 7 to 10 days, and some suggested for a shorter period of 3 to 5 days. for critically ill cases, a starting dose of 80 to 160 mg/day methylprednisolone, following a slow withdrawal within 7 to 10 days is considered. it is widely recognized that many patients, especially critically ill patients were susceptible to secondary infections. patients receiving corticosteroids had increased risks of developing hap due to the immunosuppression effects, and those who received mechanical ventilation were susceptible to vap. the latest guidelines issued by national health commission of china for the diagnosis and treatment of covid-19 infection (version 7) [20] advise against inappropriate and unnecessary use of antimicrobial therapy, especially combination of broadspectrum antibiotics. if the sputum or blood specimens showed a clear evidence of etiology or the pct levels increased, administration of antimicrobial agents should be considered. as shown in a study of 99 patients with covid-19, acinetobacter baumannii, klebsiella pneumoniae, and aspergillus flavus were simultaneously cultured in one patient. meanwhile, one case of fungal infection was attributed to candida glabrata and three cases of fungal infection were caused by candida albicans [5] . when selecting antimicrobial agents for initial empiric treatment, in addition to the local epidemiological data of hap/vap pathogens, imaging features of pulmonary lesions should also be taken into account [45] . as for fungal infections, voriconazole is recommended for the treatment of aspergillus infections, while fluconazole is more suitable for candida spp. infections. when patients are suspected with pneumocystis pneumonia, sulfamethoxazole and caspofungin should be promptly administrated [45] . in clinical practice, nearly 20% of patients with covid-19 are found to have abnormal coagulation function, and almost all severely and critically ill patients presented coagulation disorders [5, 25, 26] . in view of no relevant experience for reference, anticoagulation should be given with great caution in patients with dic though microthrombosis was observed in lung, liver, and other organs by autopsy. when patients exhibit a bleeding tendency or when surgical treatment is needed, platelet transfusion or administration of fresh-frozen plasma is recommended to correct coagulopathies analogs [46] . low molecular weight heparin (lmwh) can be used for drug prevention. as for subjects with clinical manifestations, clinicians need to be alert to the occurrence of pte, initiate the diagnostic procedures, and develop corresponding treatment strategies based on risk stratification. considering the risk of disease transmission and the false positive results caused by the presence of lung lesions, the diagnosis of pte by pulmonary ventilation-perfusion imaging is not recommended. if the critically ill patients cannot take examination due to specific conditions and the infectivity of covid-19, it is recommended to perform anticoagulant therapy for patients without contraindications. if the condition is lifethreatening and bedside echocardiography indicates new onset of right ventricular volume overload or pulmonary hypertension, thrombolytic therapy or other cardiopulmonary support treatments, such as extracorporeal membrane oxygenation (ecmo) can be initiated with the patient's full informed consent. for mild to moderate patients with hypoxemia, nasal catheters and masks and even high-flow nasal cannula oxygen therapy (hfnc) are advised. while for severe and critical patients with respiratory distress, hfnc, noninvasive mechanical ventilation (niv) or invasive mechanical ventilation, and even ecmo should be considered. hfnc can provide accurate oxygen concentration and a certain positive airway pressure to promote alveolar expansion to improve oxygenation and respiratory distress [47] . however, according to expert consensus on the use of hfnc for covid-19, patients with cardiac arrest, weak spontaneous breathing, pao 2 /fio 2 < 100 mmhg, paco 2 > 45 mmhg and ph < 7.25 and upper airway obstruction are contraindicated. for severe patients with respiratory distress or hypoxemia that cannot be alleviated after standard oxygen therapy, niv can also be considered with close surveillance [24, 26] . dangers et al. considered that changes in dyspnea could be used as a variable to predict the failure of noninvasive ventilation [48] . if the patient continuously deteriorates or the respiratory rate cannot be improved after a short time (about 1-2 h), timely tracheal intubation and invasive ventilation are required [49] . notably, patients with hemodynamic instability, multiple organ failure or abnormal mental status should not receive noninvasive ventilation. lung protection ventilation strategies (small tidal volume, limited plateau pressure, and permissive hypercapnia) are suggested to be adopted in invasive mechanical ventilation to reduce ventilator-related lung injury [50] . compared with niv, invasive mechanical ventilation can more effectively improve the pulmonary ventilation function and respiratory mechanics of patients with acute respiratory failure. it can effectively increase the sao 2 level and is more conducive to lower the plasma bnp level [51] . however, invasive mechanical ventilation requires tracheotomy, or oral/nasal tracheal intubation to establish an artificial airway, which is very likely to cause damage to patients, such as mediastinal emphysema, ventilatorrelated lung injury, and other related complications, such as reduced swallowing function, gastresophageal reflux, infections, etc. what's more, invasive mechanical ventilation also increases the risk of secondary infections transmitted by aerosol particles [52] . for critical patients, crrt can support organ function, reduce cytokine storms and maintain internal environment stability [53] . three clinical studies showed that the incidence of aki in patients with covid-19 was 3% to 7%, and 7% to 9.0% were treated with crrt. in icu, the rate of crrt application was 5.6% to 23.0% and reached as high as 66.7% to 100% in patients with aki [5, 26, 54] . crrt is recommended for patients who exhibit aki indications (hyperkalemia, acidosis, pulmonary edema, severe sodium ion disorders) or patients with ckd who have not undergone hemodialysis. during septic shock, crrt can effectively remove inflammatory mediators and significantly improve hemodynamics. when ards appears in combination with multiple organ dysfunction syndrome (mods), early crrt is recommended [55] . crrt combined with the treatment of ecmo may remove cytokines, reduce the activity of macrophages and monocytes, and better preserve lung parenchyma. some studies reported that early convalescent plasma treatment for influenza and sars-cov infection is associated with decreased viral load and reduction in mortality [56] , however, the studies were heterogeneous and of low quality. the who deemed convalescent plasma transfusion as the most promising therapy for mers-cov infection, while the efficacy remained inconclusive, with a lack of adequate clinical trials [56] [57] [58] . since the virological and clinical characteristics among sars, mers, and covid-19 were comparable [59] , convalescent plasma could have immunotherapeutic potential in covid-19 treatment and further investigations are needed to prove its safety and efficacy. one possible explanation for the efficacy of convalescent plasma therapy is that the neutralizing antibodies from convalescent plasma might suppress viremia [60] , so understanding the antibody response during the course of sars-cov-2 infection could provide strong empirical support for the application of convalescent plasma therapy. a study reported that on day 5 after treatment, an increase of viral antibodies can be seen in nearly all patients, igm positive rate increased to 81%, whereas igg positive rate increased to 100%, which was considered as a transition from earlier to later period of infection [9] . preliminary study has showed that patients who have recovered from covid-19 with a high neutralizing antibody titer and could provide a valuable source of the convalescent plasma. plasma transfusion may cause adverse effects, so convalescent plasma therapy is recommended as a last resort to improve the survival rate of severe patients with covid-19. the optimal dose and treatment time point, as well as the therapeutic indications of convalescent blood products in covid-19 remain uncertain, which need to be further investigated in randomized clinical studies. tocilizumab is a humanized igg1k monoclonal antibody which can specifically bind soluble or membrane-type il-6 receptors (sil-6r and mil-6r), and has been widely used in the treatment of autoimmune diseases such as rheumatoid arthritis [61] , adult-onset still's disease [62] , and large vessel vasculitis [63] . for covid-19 infection, clinical studies have shown that serum levels of inflammatory mediators in severe patients are significantly higher than those in common patients [25] . excessive immune responses can trigger cytokine storms and cause damage to multiple target organs. recent guidelines also point that a progressive rise in il-6 may be a clinical warning indicator for the deterioration of covid-19. a domestic research team found that tocilizumab can block the signaling pathways of two key inflammatory factors, il-6 and gm-csf, and reduce the inflammatory response. a multicenter, randomized, controlled clinical study has been coducted to evaluate the efficacy and safety of tocilizumab in the treatment of moderate patients at high risk to develop into severe and critical patients (registration number: chictr2000029765). for patients with elevated il-6 levels, the efficacy of tocilizumab can be expected. in this review, we gave an overview of epidemiological, etiological, clinical, pathological, and imaging characteristics of covid-19 and introduced the latest advancements in the treatment. this novel virus spread mainly through respiratory droplets and close personal contact. a series of complications tend to develop during disease progression, especially in critically ill patients. pathological studies of autopsy showed typical presentations of acute respiratory distress syndrome and involvement of multiple organs. apart from supportive care, no specific treatment has been established for covid-19. the efficacy of some promising antivirals, convalescent plasma transfusion, and tocilizumab needs to be further validated by ongoing clinical trials. 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 early transmission dynamics in wuhan, china, of novel coronavirus-infected pneumonia nowcasting and forecasting the potential domestic and international spread of the 2019-ncov outbreak originating in wuhan, china: a modelling study phase-adjusted estimation of the number of coronavirus disease epidemiological and clinical characteristics of 99 cases of 2019 novel coronavirus pneumonia in wuhan, china: a descriptive study a familial cluster of pneumonia associated with the 2019 novel coronavirus indicating person-to-person 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patients with 2019 novel coronavirus-infected pneumonia in wuhan clinical management of severe acute respiratory infection when novel coronavirus (ncov) infection is suspected: interim guidance esc guidelines for the diagnosis and management of acute pulmonary embolism developed in collaboration with the pulmonary pathology of early phase 2019 novel coronavirus (covid-19) pneumonia in two patients with lung cancer pathological findings of covid-19 associated with acute respiratory distress syndrome clinical pathology of critical patient with novel coronavirus pneumonia (covid-19) radiological findings from 81 patients with covid-19 pneumonia in wuhan, china: a descriptive study relation between chest ct findings and clinical conditions of coronavirus disease (covid-19) pneumonia: a multicenter study radiographic and ct features of viral pneumonia infectious pulmonary nodules in immunocompromised patients: usefulness of computed tomography in predicting their etiology imaging and clinical features of patients with 2019 novel coronavirus sars-cov-2 novel coronavirus pneumonia (covid-19) ct distribution and sign features. chin j tuberc respir dis (zhonghua jie he he hu xi za zhi) inhibition of novel β coronavirus replication by a combination of interferon-α2b and ribavirin a systematic review of therapeutic agents for the treatment of the middle east respiratory syndrome coronavirus (mers-cov) diagnosis and treatment of disease 2019 novel coronavirus infection suitable for military support hubei medical team remdesivir and chloroquine effectively inhibit the recently emerged novel coronavirus (2019-ncov) in vitro broad-spectrum antiviral gs-5734 inhibits both epidemic and zoonotic coronaviruses prophylactic and therapeutic remdesivir (gs-5734) treatment in the rhesus macaque model of mers-cov infection influenza virus-related critical illness: prevention, diagnosis, treatment chinese guidelines for the diagnosis and treatment of hospital-acquired pneumonia and ventilator-associated pneumonia in adults the japanese clinical practice guidelines for management of sepsis and septic shock feasibility of high-flow nasal cannula oxygen therapy for acute respiratory failure in patients with hematologic malignancies: a retrospective single-center study research network in mechanical ventilation) and the groupe de recherche en réanimation respiratoire en onco-hématologie (grrroh); list of contributors who included study patients: angers university hospital, angers, france. dyspnoea in patients receiving noninvasive ventilation for acute respiratory failure: prevalence, risk factors and prognostic impact: a prospective observational study european society of intensive care medicine, and society of critical care medicine. an official american thoracic society/european society of intensive care medicine/society of critical care medicine clinical practice guideline: mechanical ventilation in adult patients with acute respiratory distress syndrome diagnosis and treatment in acute respiratory distress syndrome-reply effect of invasive and non-invasive positive pressure ventilation on plasma brain natriuretic peptide in patients with chronic obstructive pulmonary disease and severe respiratory failure severe acute respiratory syndrome (sars): lessons learnt in hong kong coronavirus epidemic: preparing for extracorporeal organ support in intensive care kidney impairment is associated with in-hospital death of covid-19 patients cytokine reduction in the setting of an ards-associated inflammatory response with multiple organ failure convalescent plasma study group. 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 sars: systematic review of treatment effects current treatment options and the role of peptides as potential therapeutic components for middle east respiratory syndrome (mers): a review emerging threats from zoonotic coronavirusesfrom sars and mers to 2019-ncov convalescent plasma as a potential therapy for covid-19 tocilizumab discontinuation after attaining remission in patients with rheumatoid arthritis who were treated with tocilizumab alone or in combination with methotrexate: results from a prospective randomised controlled study (the second year of the surprise study) tocilizumab in patients with adultonset still's disease refractory to glucocorticoid treatment: a randomised, double-blind, placebo-controlled phase iii trial trial of tocilizumab in giant-cell arteritis this work was funded in part by a grant from innovative research team of high-level local universities in shanghai. min zhou, xinxin zhang, and jieming qu declare that they have no conflict of interest. this manuscript is a review article that does not involve a research protocol requiring approval by the relevant institutional review board or ethics committee. key: cord-331228-wbd0s4fo authors: shehata, mahmoud m.; gomaa, mokhtar r.; ali, mohamed a.; kayali, ghazi title: middle east respiratory syndrome coronavirus: a comprehensive review date: 2016-01-20 journal: front med doi: 10.1007/s11684-016-0430-6 sha: doc_id: 331228 cord_uid: wbd0s4fo the middle east respiratory syndrome coronavirus was first identified in 2012 and has since then remained uncontrolled. cases have been mostly reported in the middle east, however travel-associated cases and outbreaks have also occurred. nosocomial and zoonotic transmission of the virus appear to be the most important routes. the infection is severe and highly fatal thus necessitating rapid and efficacious interventions. here, we performed a comprehensive review of published literature and summarized the epidemiology of the virus. in addition, we summarized the virological aspects of the infection and reviewed the animal models used as well as vaccination and antiviral tested against it. coronaviruses (cov) became known to cause human disease in the twentieth century. hcov-229e and hcov-oc43 were discovered in the 1960s and shown to cause respiratory infections in humans [1, 2] . with the emergence of sars-cov in 2003 [3] , two other human coronaviruses were discovered, hcov nl63, hcov hku1 [4] . in 2012, a new type of coronavirus was detected as the cause of severe respiratory illness in humans. the first case was a 60-year-old male from saudi arabia admitted to hospital with acute respiratory illness leading to pneumonia and acute renal failure. the virus initially named as human corona virus-emc [5] , is currently known as the middle east respiratory syndrome coronavirus (mers-cov) [6] . classification and nomenclature of mers-cov phylogenetically, mers-cov is a lineage c β coronavirus (β-cov) and is closely related to bat coronaviruses hku4 and hku5. the rooted phylogenetic analysis showed that mers-cov had an amino acid sequence identity less than 90% to all other known covs [7] . the virus initially named by many different working groups as novel coronavirus, human coronavirus emc, human b coronavirus 2c emc, human b coronavirus 2c england-qatar, human b coronavirus 2c jordan-n3, and b coronavirus england 1, which represented the places where the first complete viral genome was sequenced (erasmus medical center, rotterdam, the netherlands) or where the first laboratory-confirmed cases were identified or managed (jordan, qatar, and england) was later named as mers-cov by the coronaviruses study groups of ictv [5, 6, 8] . mers-cov is an enveloped virus with a positive sense rna genome. coronavirus genomes range between 25 to 32 kb in size. the complete sequence of hcov-emc-2012 resulted in 30 119 nucleotides sequence [7] . coronavirus genomes are polycistronic with large replicase open reading frames orf1a and orf1b which are subsequently cleaved into 15 or 16 nonstructural proteins (nsps). the region downstream of orf1b encode smaller genes including the spike (s), envelope (e), membrane (m), and nucleocapsid (n) structural protein [9] [10] [11] . the functional receptor for mers-cov is the dipeptidyl peptidase 4 (dpp4) which is present on human nonciliated bronchial epithelial cells surfaces [12] . the dpp4 protein displays high amino acid sequence conservation across different species, including the sequence that was obtained from bat cells. cell lines susceptibility studies showed that mers-cov infected several human cell lines, including histiocytes as well as respiratory, kidney, intestinal, and liver cells [13] . the range of tissue tropism in vitro was broader than that for any other known human coronavirus [14] . mers-cov can also infect nonhuman primate, porcine, bat, civet, rabbit, and horse cell lines all possessing the dpp4 receptor [15] . the replication cycle of mers-cov consists of numerous steps as illustrated by lu et al. [30] . the mers-cov s protein is a class i fusion protein composed of two subunits: the amino n-terminal receptor binding s1 and carboxyl c-terminal membrane fusion s2 subunits. the s1/s2 junction is a protease cleavage site which is responsible for membrane fusion activation, virus entry, and syncytium formation. the s1 c domain contains the receptor binding domain (rbd), and an n domain [13] . neutralizing monoclonal antibodies against the rbd may inhibit virus entry into cells and receptor-dependent syncytium formation in cell culture, hence vaccines containing the rbd induced high levels of neutralizing antibodies in mice and rabbits [16] [17] [18] . dpp4 is the cell key functional receptor for the mers-cov s protein [19] . mers-cov is the first cov that has been identified to use dpp4 as a receptor [19, 20] . dpp4 has important roles in glucose metabolism, t cell activation, chemotaxis modulation, cell adhesion, and apoptosis [19, 21] . the s2 subunit contains five domains: a fusion peptide, the heptad repeat 1 (hr1) and hr2 domains, a transmembrane domain, and a cytoplasmic domain, which form the stalk region of s protein that facilitates fusion of the viral and cell membranes [22, 23] . the binding of the s1 subunit to the cellular receptor triggers conformational changes in the s2 subunit, which inserts its fusion peptide into the target cell membrane to form a six-helix bundle fusion core between the hr1 and hr2 domains that approximates the viral and cell membranes for fusion. mers-cov utilizes many pathways for membrane fusion depending on available host proteases, such as transmembrane protease serine protease 2 (tmprss2), trypsin, chymotrypsin, elastase, thermolysin, endoproteinase lys-c, and human airway trypsin-like protease. proteases cleave the s protein into the s1 and s2 subunits to activate the mers-cov s protein for endosome-independent host cell entry at the plasma membrane [24] [25] [26] . in addition to the pervious fusion proteases furin has been identified recently to play an essential role in the mers-cov s protein cleavage activation into their biologically active forms [27, 28] . after cell entry, the virion particle disassembles to release the nucleocapsid and viral rna into the cytoplasm for expression of viral polyproteins pp1a and pp1ab. doublemembrane vesicles and convoluted membranes are formed by the attachment of the hydrophobic domains of the mers-cov replication machinery to the limiting membrane of auto-phagosomes [29] . the viral polyproteins pp1a and pp1ab are cleaved by papain-like protease and 3c-like protease into nsp1 to nsp16 [7, 30, 31] . these nonstructural proteins form the replication-transcription complex, which regulates transcription and viral protein expression [29] . after the production of abundant viral rna and structural and accessory proteins, the n protein binds to the genomic rna in the cytoplasm to form the helical nucleocapsid (viral core). the viral core is enveloped by budding through intracellular membranes between the endoplasmic reticulum and golgi apparatus [32] . the s, e, and m proteins are transported to the budding virion, where the nucleocapsid probably interacts with m protein to generate the basic structure and complexes with the s and e proteins to induce viral budding and release from the golgi apparatus [33] . mers-cov replication cycle is completed by releasing the progeny virions through the cell membrane via exocytosis pathway. mice mers-cov strain hcov-emc/2012 was inoculated to three different mouse strains (immunocompetent balb/c mice, 129s6/svev and innate immune-deficient 129/ stat1 -/mice) intranasally. no significant weight loss was observed and infectious virus could not be detected in the lungs. only moderate pathological lesions were observed in the lungs. hence no viral replication was observed in these strains of mice [34] . zhao et al. developed a mouse model transduced with a recombinant adenovirus vector expressing hdpp4 (ad5-hdpp4) in lung tissue. inoculation of mers-cov in these mice resulted in mers-cov replication but without mortality. young mice cleared from mers-cov in 6-8 days and old mice in 10-14 days. perivascular and peribronchial lymphoid infiltration was observed, with progression to an interstitial pneumonia postinfection [35] . in another study, transgenic mice expressing hdpp4 were susceptible to mers-cov infection. infectious virus was isolated from lung and brain tissue and weight loss was observed [36] . pascal et al. developed humanized transgenic mouse. no mortality or clinical signs was observed but interstitial pneumonia and significant lung disease were observed histopathologically, suggesting that humanized dpp4 mouse is a model for mers-cov infection in which pathological changes resembles mers-cov infection in humans [37, 38] . the rhesus macaque was the first animal model used for mers-cov infection as it possessed dpp4 receptor [38, 39] . in infected animals, an increase in respiratory rates, body temperature, cough and reduced appetite was observed with mild to moderate severity. infectious virus isolated only from the lower respiratory tract. viral rna was detected in the conjunctiva, nasal mucosa, tonsils, pharynx, trachea, bronchus and lungs. mild to marked interstitial pneumonia with dark red lesions appeared in lungs. seroconversion of neutralizing antibodies began at 7 dpi and increased in titer with time. the development of a transient pneumonia, rapid replication, and tropism of mers-cov for the lower respiratory tract resembled the severity of the disease observed in humans [38, 40, 41] . similarly, the common marmoset was shown to possess the dpp4 receptor [42] . radiographic imaging showed mild to severe bilateral interstitial infiltration and extensive bronchointertitial pneumonia in infected animals. infectious virus was detected in lower and upper respiratory tract tissue and viral rna was detected in nasal mucosa, oropharyngeal swabs, blood, conjunctiva, lymph nodes, gastrointestinal tract, kidney, heart, adrenal gland, liver, spleen, brain and lungs [42] . inoculation of syrian hamsters and ferrets with mers-cov did not result in infection [12, 43] . rabbits may be used as a model to study pathogenesis, transmission, and disease control strategies of mers-cov in vivo as they seroconvert and shed virus after inoculation [44] . in september 2012, a novel coronavirus infection was noted in promed mail [45] . the virus was isolated from the sputum of a 60-year-old saudi male, who was admitted to a hospital with pneumonia and acute kidney injury in june 2012. a few days later, another report appeared describing an almost identical virus detected in a patient in qatar with acute respiratory syndrome and acute kidney injury. the patient had a recent travel history to saudi arabia and then traveled to uk for further medical care [5, 46, 47] . two cases from jordan (april 2012) were retrospectively diagnosed as mers patients. since that time, more than 1542 cases of mers-cov infection have been reported including 544 deaths [48] . the actual number of cases could be higher than those reported [49] . an outbreak of more than 180 confirmed cases including 36 deaths occurred in south korea in may and june 2015. the median age of korean cases were 55 years (range: 16 to 87 years), 60% were men, and 14% were health care professionals. the index case was a 68-year-old male who had recently traveled to several countries in the arabian peninsula [50] . disease control and prevention (cdc), and the ministry of health of saudi arabia (mohsa) as asymptomatic, mild, severely symptomatic, or mortal. cases may be classified into suspected, probable, and confirmed [52, 53] . any person with laboratory confirmation of infection with mers-cov irrespective of clinical signs and symptoms is considered as a confirmed case. who criteria for laboratory confirmation require detection of viral rna or acute and convalescent serology. the presence of nucleic acid can be confirmed by positive results from at least two sequence-specific rrt-pcrs or a single sequence-specific rrt-pcr test and direct sequencing from a separate genomic target [54] . a case confirmation by serological methods requires demonstration of seroconversion in two samples collected at least 14 days apart using at least one screening assay (enzyme-linked immunoassay, immunofluorescence assay) and a neutralization assay. a probable case is defined by the following criteria, a febrile acute respiratory illness as pneumonia or acute respiratory distress syndrome, direct contact with a confirmed mers-cov case and unavailability of mers-cov testing or results being inconclusive for a single inadequate specimen. any person who developed a fever and pneumonia or acute respiratory distress syndrome with a history of travel to countries in or near the arabian peninsula within 14 days before symptom onset or was in contact with a traveler from this region who developed a febrile respiratory illness is considered as a mers-cov suspected case. the who, cdc, and mohsa recommended laboratory diagnostics for mers-cov infection [6, 9, 51, 55, 56] . mers-cov cases must be confirmed by at least two positive qrt-pcr tests on two different specific genomic regions or single positive qrt-pcr with a sequence of another positive genome fragment [57] . the who algorithm for testing mers-cov relies on qrt-pcr and sequencing [58] . available real-time tests include an assay targeting the rna upstream of the e gene (upe) as a highly sensitive screening assay and three confirmatory assays targeting open reading frames (orf 1a and 1b) and/ or n gene. the orf 1a assay is of equal sensitivity to the upe assay. the orf 1b assay is less sensitive but is useful for confirmation. these assays are specific for mers-cov and have not shown cross-reactivity with other respiratory human coronaviruses. for sequencing, two target genes, the rna-dependent rna polymerase (rdrp, present in orf 1b) and n genes are enough to confirm the existence of mers-cov rna in the samples of a patient [57] . several serologic assays including immunofluorescence assays, protein microarray assay, enzyme-linked immunosorbent assay (elisa) have been developed for the detection of mers-cov antibodies [57, [59] [60] [61] . any positive test by one of these assays should be confirmed with a neutralization assay. single serological result may be valuable for definition of probable case and should be followed by further testing for confirmation of mers-cov infection [62] [63] [64] . incubation period of mers-cov infections was studied by assiri et al. in 2013. the median incubation period was 5.2 days (95% ci 1.9-14.7 days) [65] . in another report from france of a secondary case, a patient who shared a room with an infected patient, the incubation period was estimated at 9 to 12 days [66] . in the recent outbreak in south korea during may/june 2015, the median incubation period was 6.3 days [67] . who and cdc recommended that individuals that returned from the arabian peninsula and other affected countries must be evaluated for mers-cov infections up to at least 14 days [68] . clinical features of mers-cov infections range from asymptomatic cases to mildly ill, severe pneumonia, acute respiratory distress syndrome, septic shock and mortal with multi-organ failure (table 1) [64, 65] . many other clinical features such as gastrointestinal symptoms (anorexia, nausea, vomiting, abdominal pain, diarrhea), pericarditis, and disseminated intravascular coagulation were reported [65, 69, 70] . specific clinical conditions (comorbidities) were apparently proportionate with high severity of mers-cov infections. a study by assiri et al. in saudi arabia showed that of a total of 47 patients with mers-cov infection in 2013, 45 (96%) had underlying clinical conditions, including diabetes mellitus (68%), hypertension (34%), chronic cardiac disease (28%), and chronic kidney disease (49%) [65] . this high rate of comorbidities must be interpreted with some caution, since diabetes mellitus is common in saudi arabia, and because approximately half of those 47 were part of an outbreak in a hemodialysis unit, where rates of comorbidities might be high due to chronic kidney disease [65, 71] . in another study, being on dialysis, diabetes mellitus, and age > 50 years was associated with mortality [72] . in this study, testing positive for mers-cov in a plasma sample was a predictor of severe outcome [72] . younger adults and children appeared to be less susceptible to mers-cov infection. only one study described mers-cov infection in children [73] . all of those children were discovered during contact investigations of older patients. only 2 of 11 children developed symptoms of mers-cov infection. these two children had underlying conditions (cystic fibrosis and down syndrome). the other 9 children were asymptomatic. there are few reports of mers-cov infections in pregnant women. a five-month pregnant female developed vaginal bleeding and abdominal pain after one week, then delivered a stillborn infant [74] . another case in the united arab emirates was near term phase, she gave birth to an apparently healthy baby, and died after delivery [52] . mild and asymptomatic mers-cov infections have been reported, a majority of whom were identified among the contacts of patients [62, 75, 76] . in a report from mohsa, more than 3000 contacts of patients were screened using qrt-pcr and seven healthcare workers with mers-cov infection were identified, two of whom were asymptomatic and five of whom had mild upper respiratory tract symptoms [75] . epidemiological and virological studies were conducted in attempts to determine person to person transmission of mers-cov. they studied case clustering in household and hospital outbreaks in the uk, tunisia, italy, and in healthcare facilities in saudi arabia, france, iran, and lately in south korea. those studies provided strong evidence that human-to-human transmission occurs [70, [77] [78] [79] [80] . the number of contacts infected by individuals with confirmed infections, however, appears to be limited [62] , except the outbreak of south korea in may/ june 2015, where most cases were secondary and some cases were tertiary infections [67, 81] . secondary cases often were milder or symptomless [62] . possible modes of transmission include droplet and close contact transmission, air borne transmission, and fomite transmission [82] . the majority of all laboratory-confirmed secondary cases have been associated with healthcare settings [82] . the majority of cases of jeddah, saudi arabia hospital outbreak during the spring of 2014 were acquired through human-to-human transmission due to systematic weaknesses in infection control [76] . secondary transmission rates were assessed within households and the transmission rate was around 4%, suggesting that the actual number of infection is greater than reported [62] . during the outbreak in south korea during may/june 2015, 25 secondary infections were associated with the index case, who was hospitalized from may 15 to may 17 and 11 were tertiary [83] . the median incubation period was six days for secondary cases and six days for tertiary cases. this outbreak also clearly demonstrated roles of "superspreaders," who may be responsible for a high proportion of cases [83] . for instance, a single patient infected more than 70 other people while being treated in the emergency room of a hospital in south korea for three days, 27-29 may 2015. transmissibility and epidemic potential studies of mers-cov revealed that the reproduction number (r 0 ) of patients infected with mers-cov ranged between 0.6 to 0.69 [84, 85] . the finding of an r 0 < 1 suggests that mers-cov does not yet have pandemic potential. other study suggested that r 0 values might reach to 0.8 to 1.3 in the absence of infection control [49] . shedding periods of mers-cov in humans was reported to be long as viruses were detected in lower respiratory samples of symptomatic patients for more than two weeks [86] . at instances, prolonged shedding for 6 weeks was detected in an asymptomatic healthcare worker. these findings raise concerns that asymptomatic persons could transmit infection to others in a silent manner [87] . the majority of cases have occurred in saudi arabia and united arab emirates [88] [89] [90] . many cases have also been reported outside the arabian peninsula in north africa, europe, asia, and north america as shown in table 2 . almost all cases reported outside the arabian peninsula had a travel history to it. the first cluster was in october/november 2012 in four men of the same family in riyadh, saudi arabia, two of whom died [75] . the second cluster was reported in jordan in april 2012 involving 10 healthcare workers exposed to fatal patients. in addition, seven surviving hospital contacts seroconverted suggesting that they had mers-cov infection [91] . the third cluster was reported in uk during january/ february 2013. an english resident had a travel history to saudi arabia and pakistan in january, developed a severe respiratory illness, and tested positive for both mers-cov and h1n1 influenza a, and died in march 2013 after infecting several contacts [92] . a cluster of 43 cases of mers-cov was reported in al-hasa in saudi arabia during april 2013. all those cases were directly linked to human to human contact in the same hospital. there were only two confirmed cases of healthcare workers, and three family members were detected by a survey of over 200 household contacts that visited this hospital [77] . in france, may 2013, an infection of mers-cov was reported in a patient who recently traveled to the united arab emirates. a second case who shared the hospital room with the first case tested positive. the first patient died and the second patient was critically ill. a survey of 100 healthcare workers found no other infections with mers-cov, despite the lack of use of personal protective equipment [70] . a surge in mers-cov cases was reported in saudi arabia and the united arab emirates during march and april 2014 [55, 76] . the majority of cases were associated with hospital-based outbreaks jeddah, riyadh, tabuk, and madinah in saudi arabia as well as in al ain, and abu dhabi in united arab emirates. cases included several healthcare workers, visitors, patients, and ambulance staff. person to person transmission was confirmed in > 75% of cases. the majority of infected health care workers developed mild symptomatic or asymptomatic infection, but about 15% had severe illness or died [93] . the recent outbreak of south korea occurred in may 2015. the index case was a man who had recently traveled to bahrain, the united arab emirates, saudi arabia, and qatar [55] . as of late july 2015, > 180 secondary cases were reported including 36 death and many cases had been reported among household and hospital contacts [55, 67] . in china, one case occurred in a man who traveled to china from korea following exposure to two relatives with mers-cov infection [55] . in spite of reporting of mers-cov infections throughout the year, some evidence on disease seasonality occurred. the first identified cases of mers-cov infection were reported in april and june 2012 [5, 46, 47] . a high increase in cases was reported in april and may 2013 followed by a surge in case reporting in april and may 2014. increase in case reporting in march to may 2013 were attributed to infection from infected young camels [94, 95] sources and modes of transmission of mers-cov are still unclear. initially, a bat origin of mers-cov was suggested based on the relation of genome sequences between mers-cov and bat coronaviruses [96] . cell tropism studies showed that both bat coronavirus hku4 and mers-cov shared the same cell type receptors, dpp4 [4, 7, 75] . mers-cov grows readily in several bat-derived cell lines [14] . there is no evidence for direct or indirect transmission of mers-cov from bats to humans. virological studies performed in europe, africa, and asia, including the middle east, have shown that coronavirus rna sequences are found frequently in bat feces. some of the sequences were closely related to mers-cov sequences [97] [98] [99] . in a survey from saudi arabia, 823 fecal and rectal samples were tested by pcr for mers-cov, many coronaviruses sequences were detected [97] . most of the detected sequences were unrelated to mers-cov, but one sequence of 190 nucleotide in the rna-dependent rna polymerase (rdrp) gene had a 100% identity with a mers-cov. this sequence was detected from feces of a taphozous perforatus bat captured near the home of the index saudi patient. uncommon contact of humans with bats indicates that bats are not the intermediate host of mers-cov transmission but may be the reservoir of the virus [100] . dromedary camels (camelus dromedarus) appear to be the source of mers-cov. other animals like sheep, goats, and cows tested negative to anti-mers-cov antibodies. camel sera from oman, canary islands, and egypt were positive for anti-mers-cov antibodies in about 100%, 14%, and > 90% of the samples respectively [63, 101, 102] . retrospective studies on archived human sera showed no evidence of infection with mers-cov before 2012 [103] , but anti-mers-cov antibodies were detected in archived camel sera in saudi arabia in 1993 [95] , and united arab emirates in 2003 [104] , indicating circulation of mers-cov in camels for many years. bactrian camels in mongolia tested negative for mers-cov antibodies [105] . serologic studies from around the middle east suggested that camels are one of the sources of mers-cov as > 90% of adult camels tested positive and had high titers of antibodies. seropositivity was different in juvenile camels and was usually lower than in adults. these results suggested that mers-cov infections in camels occurred in young ages followed by frequent boosting [94, 95, 102, 104] . camels in other parts of the world, far from the middle east like in europe, australia, and the americas do not have mers-cov antibodies and have no evidence of infection [106] . table 3 summarizes camel serologic studies. in a study aimed to evaluate virus infectivity and shedding in camels, three adult dromedary camels were inoculated with mers-cov intratracheally, intranasally, and conjunctivally. those camels shed large quantities of virus from the upper respiratory tract and infectious virus was detected in nasal secretions for 7 days post-inoculation and viral rna for up to 35 days post-inoculation [113] . human infections with mers-cov were linked to camels. the first evidence was a study in saudi arabia in which the mers-cov full genome sequences of isolates from a man with fatal infection and from one of his camels were identical. this patient had a direct contact with his deceased camels some days before the onset of symptoms. these results suggested that mers-cov can infect dromedary camels and can be transmitted from them to humans by direct close contact [86] . in other studies, phylogenetic analyses of camel and human isolates of the mers-cov genome demonstrated that the viruses were highly identical or in some cases were similar to each other [107, 108, 114] . seroepidemiological studies shown low prevalence of mers-cov antibodies in humans in saudi arabia [103, 115] . a survey of 10 009 individuals representative of the general population of saudi arabia resulted in 15 seropositive subjects (0.15%), however, seropositivity increased 15-23 folds in camel-exposed individuals [78] . in a separate report, 7 of 87 camel shepherds and 140 slaughterhouse workers (3.1%) tested positive for mers-cov antibodies [103] . an overview of mers-cov transmission routes is illustrated in fig.2 . the development of an effective vaccine is critical for prevention of a mers-cov pandemic. some investigators have indicated that the rbd protein of mers-cov s protein is a good candidate antigen as a subunit vaccine. various rbd fragments showed the highest dpp4 binding affinity and induced the highest-titer of igg ab and neutralizing ab in mice and rabbits [17, 18, [116] [117] [118] [119] [120] . a robust neutralizing antibody response was elicited in balb/c mice against mers-cov after immunization with purified full s protein nanoparticles produced in sf9 cells infected with specific recombinant baculovirus containing the s gene [121] or a recombinant human adenoviral vectors (rad5 or rad41) containing the s or s1 genes [122, 123] . vaccinia ankara was encoded with full s protein and inoculated to balb/c mice that developed high levels of neutralizing antibodies and had induction of humoral and cell-mediated immunity [124, 125] . another study using ad5-hdpp4-transduced balb/c mice immunized with venezuelan equine encephalitis virus replicon particles containing s protein elucidated a reduction of viral titers to nearly undetectable levels and increased neutralizing antibodies [35] . recently, wang et al. developed two candidate vaccines, a subunit (full s and s1 protein fraction) and a dna vaccine (full s and s1 gene in a mammalian vrc8400 vector). the vaccine containing the full s dna and s1 protein was the most efficacious in mice and rhesus macaques [126] . using antibodies to deter mers-cov infection appears to have some promise. transfer of sera containing anti-mers-cov-s protein to or seropositive camel sera to ad5-hdpp4-transduced mice accelerated virus clearance, inhibited virus attachment, and reduced weight loss [35, 37, 127] . recently, corti et al. successfully isolated monoclonal antibodies from serum obtained from a mers-cov survivor after 200 days of infection [128] . transduced ad5-hdpp4 balb/c mice were immunized with 15 mg/kg of the mab and showed decreased lung [105] viral titers, no weight loss, and decreased peribronchial lymphoid infiltration [128] . no approved antivirals for use against mers-cov infection are yet available. the first approach performed when a new unknown virus like mers-cov emerges is testing drugs used as antiviral for similar viruses [29, 129, 130] . type i interferons and ribavirine combination exhibited acceptable results in cell culture and rhesus macaques by decreasing the host inflammatory response, replication of virus, and improved clinical outcome [129, 131, 132] . a human cohort study in saudi arabia showed that treatment with combination of ribavirin and interferon-α2b to 5 did not improve clinical outcomes but this may have been due to late treatment or due to the immunocompromised state of the patients [133] . in a retrospective study of 20 mers-cov infected patients treated with ribavirin and interferon α-2a, results showed 14-day and 28-day survival was improved by 70% and 28% in the treated group as compared to an untreated group [134] . the second approach is screening of approved drugs with known safety profiles and transcriptional signatures in different cell lines. several drugs, including antiparasitics, neurotransmitters, antibacterials, inhibitors of clathrinmediated endocytosis estrogen receptor, lipid or sterol metabolism, protein processing, and dna synthesis or repair were tested on culture cells [119, [135] [136] [137] [138] [139] . lopinavir-ritonavir combined with pegylated interferon and ribavirin therapy showed improved outcomes in infected marmosets [140] . the third approach involves in vitro inhibition of s protein to block virus entry into host cells using designed antiviral peptides targeting the hr2 domain of the s2 subunit of the mers-cov and preventing the interaction between the hr1 and hr2 domains required for the formation of the heterologous six-helix bundle in viral fusion core formation [22, 23] . other drugs that act as inhibitors for viral proteases and helicase to suppress mers-cov infection were tested [141] [142] [143] [144] [145] . other investigators studied inhibition of mers-cov infection by competitive inhibition of dpp4 cell receptor using compounds such as sitagliptin, vildagliptin, and saxagliptin [19, 146] . more than three years have passed since the first detection of mers-cov human infection and the virus, uncontrolled, continues to cause major outbreaks in the middle east. the recent outbreak in korea demonstrated that a single index case can lead to 185 more infections in a short period of time, hence raising questions about the accuracy of the number of cases being reported in the middle east. furthermore, the korean outbreak confirmed the high fatality rate of mers-cov infection as being true rather than overestimated in case only the more severe cases are detected. in all, public health, veterinary health, and research efforts need to be consolidated in order to answer the following high priority questions: -what is the true extent of human infection with mers-cov? -what antivirals and vaccines are to be used in humans? -what infection control measures are needed in healthcare settings to prevent nosocomial outbreaks? -what measures need to be in place in order to prevent zoonotic infections from camels? -is it possible to control the virus in the camel population and if so, how? -are there other animal species involved in the mers-cov transmission cycle? a new virus isolated from the human respiratory tract recovery in tracheal organ cultures of novel viruses from patients with respiratory disease coronavirus as a possible cause of severe acute respiratory syndrome discovery of seven novel mammalian and avian coronaviruses in the genus deltacoronavirus supports bat coronaviruses as the gene source of alphacoronavirus and betacoronavirus and avian coronaviruses as the gene source of gammacoronavirus and deltacoronavirus isolation of a novel coronavirus from a man with pneumonia in saudi arabia middle east 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important differences with sars coronavirus antiviral potential of erk/mapk and pi3k/akt/mtor signaling modulation for middle east respiratory syndrome coronavirus infection as identified by temporal kinome analysis testing of middle east respiratory syndrome coronavirus replication inhibitors for the ability to block viral entry screening of an fda-approved compound library identifies four small-molecule inhibitors of middle east respiratory syndrome coronavirus replication in cell culture interferon-β and mycophenolic acid are potent inhibitors of middle east respiratory syndrome coronavirus in cell-based assays treatment with lopinavir/ritonavir or interferon-β1b improves outcome of mers-cov infection in a nonhuman primate model of common marmoset prediction and biochemical analysis of putative cleavage sites of the 3c-like protease of middle east respiratory syndrome coronavirus assessing activity and inhibition of middle east respiratory syndrome coronavirus papain-like and 3c-like proteases using luciferase-based biosensors evaluation of ssya10-001 as a replication inhibitor of severe acute respiratory syndrome, mouse hepatitis, and middle east respiratory syndrome coronaviruses thiopurine analogs and mycophenolic acid synergistically inhibit the papain-like protease of middle east respiratory syndrome coronavirus the newly emerged sars-like coronavirus hcov-emc also has an "achilles' heel": current effective inhibitor targeting a 3c-like protease inhibition of middle east respiratory syndrome coronavirus infection by anti-cd26 monoclonal antibody mahmoud m. shehata, mokhtar r. gomaa, mohamed a. ali, and ghazi kayali declare that they have no conflict of interest. this manuscript is a review article and does not involve a research protocol requiring approval by the relevant institutional review board or ethics committee. key: cord-006788-kz8i4a6h authors: li, xiaokun title: the fgf metabolic axis date: 2019-09-07 journal: front med doi: 10.1007/s11684-019-0711-y sha: doc_id: 6788 cord_uid: kz8i4a6h members of the fibroblast growth factor (fgf) family play pleiotropic roles in cellular and metabolic homeostasis. during evolution, the ancestor fgf expands into multiple members by acquiring divergent structural elements that enable functional divergence and specification. heparan sulfate-binding fgfs, which play critical roles in embryonic development and adult tissue remodeling homeostasis, adapt to an autocrine/paracrine mode of action to promote cell proliferation and population growth. by contrast, fgf19, 21, and 23 coevolve through losing binding affinity for extracellular matrix heparan sulfate while acquiring affinity for transmembrane α-klotho (kl) or β-kl as a coreceptor, thereby adapting to an endocrine mode of action to drive interorgan crosstalk that regulates a broad spectrum of metabolic homeostasis. fgf19 metabolic axis from the ileum to liver negatively controls diurnal bile acid biosynthesis. fgf21 metabolic axes play multifaceted roles in controlling the homeostasis of lipid, glucose, and energy metabolism. fgf23 axes from the bone to kidney and parathyroid regulate metabolic homeostasis of phosphate, calcium, vitamin d, and parathyroid hormone that are important for bone health and systemic mineral balance. the significant divergence in structural elements and multiple functional specifications of fgf19, 21, and 23 in cellular and organismal metabolism instead of cell proliferation and growth sufficiently necessitate a new unified and specific term for these three endocrine fgfs. thus, the term “fgf metabolic axis,” which distinguishes the unique pathways and functions of endocrine fgfs from other autocrine/paracrine mitogenic fgfs, is coined. fibroblast growth factors (fgfs) are pleiotropic signal molecules for all types of cell and tissue systems in metazoans [1] [2] [3] . fgfs share a conserved core structure of β-trefoil fold consisting of 12-stranded β-sheets arranged in three similar lobes around a central axis, of which six strands form an antiparallel β-barrel [4, 5] . except for the four fgf-homologous intracrine factors that are functionally reminiscent of the ancestor fgf, the fgfs can be classified into mitogenic and metabolic fgfs, which overtly regulate cellular proliferation and substrate/energy metabolism, respectively, on the basis of their distinct functions and endpoint biological effects [6, 7] . both fgf classes signal through the same types of transmembrane receptor tyrosine kinases, that is, the fgf receptors (fgfrs) 1 to 4 with multiple splicing variants [8] . however, in physiology, these two types of regulatory activities driven by the two fgf classes appear to be spatially and temporally segregated. at a physiological level, mitogenic fgfs appear to be incapable of traveling far to other tissues, including metabolic tissues, to promote cellular metabolism because of local trapping after secretion that is mediated by high affinity binding to the extracellular matrix heparan sulfate (hs). on the other hand, metabolic fgfs circulate but are inactive for nonmetabolic tissues or cells that often undergo active tissue remodeling via the renewed cycles of cell proliferation and population growth because of the lack of critical transmembrane accessory coreceptors. this divergence necessitates a distinction of the metabolic axis that is a term as we call hereafter, which the metabolic fgfs drive, from the mitogenic axis that the mitogenic fgfs drive. the metabolic axis still shares the major aspects of structural coevolution [9,10] while gaining unique structural and functional divergence with the mitogenic axis within each subfamily (table 1) , as our recent structural studies have revealed [2, 5, 11] . from the evolutionary standpoint, although the two axes largely parallel and drive differential effects via divergent intracellular mechanisms, they aim for a common goal of promoting the survival and homeostasis of each cell/tissue system and the organism as a whole (fig. 1) , as we have summarized in a previous review [7] . the classic fgf family consists of 17 structurally related polypeptides, which are secreted and act as extracellular signaling molecules, in humans [1, 3, 7, 12] . for the most part of fgf history beginning in the late 1970s [13, 14] , fgfs are known as short-range mitogens in a wide variety of cell types in the developing ectoderm, mesoderm, and endoderm. fgfs elicit a chemoattractant activity to promote cell migration and tissue remodeling and antiapoptotic effects to promote cell survival. fgf1 and 2 are the prototypes that are initially isolated based on potent mitogenic activity toward fibroblasts or fibroblastlike cells [13, 14] . it was found early that the mitogenic fgfs bind tightly to the local extracellular matrix hs chains, do not circulate, and accordingly act in a paracrine or autocrine mode. this heparin/hs binding property renders their potent activity temporarily contained but timely released locally upon injury or demand of tissue remodeling [15] . these mitogenic fgfs include 14 members (table 1) , which strongly promote genomic dna synthesis and subsequent cell division and population growth [12, 16, 17] . therefore, mitogenic fgfs play critical roles in the development of multiple tissues/organs [18] [19] [20] . they initiate the mitogenic axis by binding to the ig-like ectodomains of their cognate transmembrane fgfrs in complex with hs motifs on diverse target cells and tissues in the first step [1, 21, 22] . the subsequent activation of the intracellular kinase domains of fgfrs results in downstream signal relay primarily through the pi3k-akt, ras-mapk, and plcg-pkc pathways [23] [24] [25] , as we have summarized previously [3] . these hs and fgfr dependent activities driven by the mitogenic fgf axes contribute not only to the regulation of virtually all aspects of development and organogenesis but also to many natural processes of active post-developmental tissue repair, remodeling, and homeostasis [26] . among mitogenic fgfs, fgf7, which is also known as keratinocyte growth factor (kgf), has the highest specificity for receptor isotypes [12, 22, 27] . fgf7 only activates the iiib-type isoform of fgfr2. given that fgf7 is produced in mesenchyme cells, while fgfr2iiib resides on the epithelial or keratinocyte cells, fgf7 forms a unidirectional paracrine communication axis with fgfr2 from mesenchyme to epithelium compartment within a tissue or organ. on the other hand, epithelial cells secrete specific fgfs (e.g., fgf1 or 9), which then acts on mesenchymal cells that harbor fgfr1iiic within two compartmental tissues. therefore, these fgf1 and fgf7 driven mutual cell communication axes are poised to drive tissue remodeling and maintain tissue homeostasis [28] . the prolonged or abnormal activation of the fgfr-hs binary complexes by mitogenic fgf axes contributes to an array of cell/tissue-specific developmental diseases and multiple cancer types [3, 29] (see a brief summary in table 1 ). the proliferation-and survival-promoting fig. 1 scheme of fgf metabolic axis evolution. the fgf family originates from a common fgf13-like ancestor molecule in early metazoans that bifurcates into the so-called intracrine fgf-homologous factor (fhf) subgroup (black arrow), including fgf11, 12, 13, and 14 (not shown), and fgf4-like molecule, which continues to bifurcate into two major functional subgroups with diverging structural and functional specifications. the socalled mitogenic fgf subgroups, including the fgf5, 8, 9, and 10 subfamilies (red arrows, table 1 ), bind extracellular matrix heparan sulfate and drive autocrine/paracrine mitogenic signal axes to promote cell proliferation and population growth. by contrast, the endocrine fgf subgroup members (green arrow, table 2 ), including fgf19, 21, and 23, drive metabolic signal axes that elicit broad-spectrum functions in regulating the metabolic homeostasis of bile acid, lipids, glucose, energy, and minerals without direct proliferation-promoting activity. however, both the fgf mitogenic and fgf metabolic axes are designed to promote cell and organismal survival in the vertebrates (orange arrows and blue-colored font). activities of the diverse mitogenic fgf axes have been a major focus of utilities as regenerative and repair agents in a range of medical settings [30] [31] [32] [33] [34] . in the past, we demonstrated the benefits of the application of mitogenic fgfs to tissue damage complications of diabetes mellitus, including diabetic cardiomyopathy, nephropathy, and neuropathy [35] [36] [37] , as well as to wound healing and spinal cord injury repair [38] [39] [40] . on the other hand, the mitogenic fgf mediated cell miscommunications have also been on the menu for developing inhibitors to be used in cancer therapy [29, 41, 42] . it should be pointed out that, although at a physiological level mitogenic fgfs are not evolutionarily designed to circulate and target distal tissues or organs for an endocrine effect, at pharmacological or supraphysiological levels, mitogenic fgfs exert certain regulatory activities beyond promoting cell proliferation and growth possibly due to their accumulation sufficiently to achieve an effect in distal metabolic tissues, where a cognate fgfr isotype is expressed. it was shown in the early 1990s that a bolus intravenous injection of fgf1 or 2 could target vascular endothelium to decrease arterial blood pressure [43] . fgf16 is expressed in classical brown fat depots during the later stages of embryonic development, and recombinant fgf16 is a mitogen for adipocytes [44] . mice overexpressing fgf16 delivered by adeno-associated virus display dramatic weight loss and uncoupling protein-1 (ucp1) upregulation in inguinal white adipose tissue (wat), which is a common site for emergent active brown adipose tissue (bat). these effects are likely a combined result of reduced food and water intake and abnormal feces replete with lipid and bile acid due to the brain, liver, and intestinal actions of overexpressed fgf16 [45] . mice deficient in fgf1 exhibit insignificant phenotypes under standard dietary conditions; however, under a chronic high-fat diet, these mice develop an aggressive diabetic phenotype coupled with aberrant adipose phenotypes, including multiple histopathologies in the adipose vasculature network, accentuated inflammatory response, aberrant adipocyte size distribution and expansion, and ectopic expression of pancreatic lipases [46] . in particular, we show by structure-based mutagenesis that fgf1 can be designed to have full metabolic activity of wild-type fgf1 but with reduced proliferative potential both in vitro and in vivo [47] . these studies underscore the important role of fgf1 in maintaining local adipose tissue homeostasis, which upon significant tissue perturbations impinges on the metabolic functions that subsequently affect the systemic metabolic state. thus, the metabolic effects of several mitogenic fgf axes may be due to either a local function in maintaining cellular homeostasis that is closely associated with local metabolic state at a physiological concentration or an induced metabolic response to a supraphysiological concentration from circulation in the metabolic tissues or organs where fgfr resides. however, at pharmacological levels, few mitogenic fgfs may also be designed to elicit systemic metabolic effects. in contrast to mitogenic fgfs, the metabolic fgf subfamily contains only three members, namely, fgf19 (mouse fgf15), 21, and 23 [2, 7, [48] [49] [50] [51] . however, the metabolic axes of these three fgfs regulate a wide range of metabolic pathways, resulting in tissue and organismal metabolic homeostasis of bile acids, lipid, glucose, energy, and minerals. although the metabolic fgf axes do not overtly promote dna synthesis, thereby leading to cell proliferation [12, 52, 53] , both metabolic and mitogenic fgf axes appear to enhance cell survival and promote an optimal state of homeostasis in the target tissues and organisms [7] . based on current knowledge, the metabolic fgfs appear to originate from a common fgf13-like ancestor molecule as mitogenic fgfs and then bifurcate in early evolution through an fgf4-like molecule from all other mitogenic members by acquiring unique structural and mechanistic properties [5, 10, 11, 54] , thereby leading to specific activities in modulating metabolic states in specific cell and tissue types [2] . instead of acting locally, metabolic fgfs take a hormonal or endocrine route of action by traveling through circulation from the originating tissue to other peripheral tissues/organs. this endocrine action can be attributed to the loss of the structurally conserved hsbinding domain characteristic of the mitogenic fgfs [5] . both the expression and target tissues of the metabolic fgfs are relatively limited to the metabolically active endocrine organs, such as liver, intestine, adipose tissue, pancreas, muscle, bone, kidney, heart, parathyroid, and specific neurons in specific regions of the central nervous system (cns) [55, 56] . in expression tissues, metabolic fgf genes are subject to direct transcriptional control by several major metabolite-responsive nuclear receptors, including farnesoid x receptor (fxr), peroxisome proliferator-activated receptor α (ppara) and γ (pparg), carbohydrate-response element-binding protein (ch-rebp), sterol regulatory element-binding protein-1c (srebp1c), retinoic acid-related orphan receptor α (rora), liver x receptor β (lxrb), vitamin d receptor (vdr) [48] [49] [50] [57] [58] [59] [60] [61] [62] [63] [64] [65] , and stress-sensing transcription factors, such as atf4 [66] , depending on the location of specific nutrition/energy-sensing cells in specific tissues. in target tissues, the biological effects of the metabolic fgf axes are still mediated by fgfrs but in a different binary complex with a new transmembrane nonkinase accessory coreceptor, the α-klotho (kl) or β-kl (klb) [5, 11, 55, 67] ( table 2) , to which mitogenic fgfs do not bind. structurally, metabolic fgfs coevolve with coreceptor kl/klb but also acquire new structural elements that direct specific contact interactions with kl/klb and fgfrs, thereby leading to a tethered basic triad complex and subsequent activation of intracellular kinase domains of fgfrs [5, 11] . the c-terminus of the metabolic fgfs mimics the interaction mode of a sugar chain that docks into the pseudo-glycolytic pocket of kl/klb while interacting with fgfr ectodomains through the domains that are conserved across the fgf family [5, 68] . meanwhile, the interacting kl/klb protrudes an "arm" from the membrane-proximal glycosidase domain griping onto the fgfr ectodomain. although the fgfrs, in particular fgfr1, are broadly expressed, the highly restricted expression of kl/klb and metabolic fgfs, and the new structural elements and mutual interaction modes, set the tone for tissue-specific functions of the metabolic fgf axes ( table 2 ). the different intracellular molecular constituents in different cells types, which are tailored to perform specific biological functions, may be also an important limiting factor. for instance, the adult adipocytes are not poised in a normal context to increase population by direct proliferation due to the loss of several key proliferation-controlling pathways, thereby partly accounting for the inability of the activated fgfr1 by fgf21 to promote adipocyte proliferation. overall, metabolic fgfs appear to be inducible stress factors in response to organismal metabolic perturbations [7,69] and signal distal peripheral tissues through the fgfr-kl/klb complex to control due metabolic pathways. in this sense, the metabolic fgf acts as a key to ignite the fgf-fgfr-klb/klb triad complex, which functions similarly as an engine with an axis to drive effects in a tissue-specific manner, thereby leading to beneficial effects that offset the initial adverse metabolic changes and prevent metaflammation and tissue damage not only in the fgf-producing tissues but also systemically [2, 7] (table 2) . consequently, both the analogs of endocrine fgfs and the agonists of fgf-kl/ klb have been actively pursued clinically for the prevention and treatment of a wide range of metabolic diseases and comorbidities [2, [70] [71] [72] [73] [74] [75] . fgf19 is the prime controller of diurnal bile acid flux, and the fgf19-driven metabolic axis is a temporal interorgan crosstalk from the ileum to the liver in response to the increase in the postprandial serum and transintestinal flux of bile acids [2, 49] (fig. 2) . this axis serves to control the enterohepatic and systemic levels of bile acids negatively, which facilitate the uptake and absorption of dietary lipids after a meal but are toxic as biodetergent if the flux is prolonged at increased levels. the ileal initiation of the fgf19 signal is under the transcriptional control of fxr, which is stimulated by the reabsorbed enterocyte bile acids as a natural ligand that is originally released from gallbladder and mixed with food traveling down from the duodenum to jejunum and ileum. this enterocytederived fgf19 activates the remote fgfr4-klb complex [67] residing across the membrane of hepatocytes in the liver, resulting in a major feedback termination of the transcription of the rate-limiting enzymes cyp7a1 and cyp8b1 in the bile acid biosynthesis pathways [49, 76] . therefore, the fgf19 axis triggers the shut-off of hepatic biosynthesis of new bile acids from cholesterol and the refilling of gallbladder approximately 2 h after the peak of serum bile acids is reached. in experimental animals, fgf19 overexpression or administration elicits other metabolic effects [77, 78] . excessive fgf19 promotes lipolysis, metabolic rate, and energy expenditure and reduces body weight, serum glucose, and lipids. the fgfr1-klb complex on adipose tissues, including wat and bat, was suggested in a large part to mediate these metabolic effects [79] (fig. 2) . however, the direct metabolic roles of bile acid fluctuation and bile acid-activated fxr and tgr5 cannot be excluded. although there is no evidence for any genetic mutation of fgf19 gene that may be involved in human metabolic diseases, its reduced synthesis and blood levels are suggestive of a causative factor of chronic bile acid diarrhea [80, 81] and certain metabolic disorders, such as metabolic syndrome, nonalcoholic fatty liver disease (nafld), and insulin resistance. experimentally, the fig. 2 fgf19 metabolic axis. the major fgf19 metabolic axis drives a temporal interorgan crosstalk from the ileum to the liver in response to the increase in postprandial serum and transintestinal flux of bile acids to discontinue the biosynthesis of new bile acids after sufficient food digestion, thereby preventing the prolonged exposure of tissues to potential bile acid toxicity. pharmacological fgf19 may also initiate multiple signal axes to drive effects on multiple tissues/organs, such as promoting (green arrow) energy expenditure in white and brown adipose tissues, increasing muscle mass and insulin sensitivity, and preventing (red long-tailed "t" sign) systemic hyperglycemia and hyperlipidemia. faa: free fatty acids. neutralization of fgf19 by specific anti-fgf19 antibodies causes severe diarrhea in monkeys accompanied by the increases in bile acid synthesis, serum and fecal total bile acids, specific bile acid transporters, and liver toxicity [82] . in obese patients who undergo roux-en-y gastric bypass bariatric surgery, fgf19 increases to normal values, which at least partially underlie the benefits of this approach [83] . on the other hand, high fgf19 expression levels are found in the livers of patients with extrahepatic cholestasis [84, 85] , suggesting fgf19 as a therapeutic target for this disease. recently, the fgf19 axis was shown to elicit hypertrophic and protective effects on the skeletal muscle presumably through a klb-fgfr4-dependent mechanism by increasing myofiber size in the soleus, muscle mass, and grip strength [86] . pharmacological fgf19 ameliorates skeletal muscle atrophy and prevents muscle wasting in mice with glucocorticoid treatment, obesity, or sarcopenia. these results highlight a potential treatment strategy for muscle wasting induced by glucocorticoid treatment, obesity, aging, and cachexia. however, whether the same treatment will have a similar adverse effect on the liver still has to be determined because muscle-specific transgenic mice developed prominent hepatocellular carcinoma (hcc) [87] . despite the tumorigenic concern, the fgf19 analog ngm282 was tested in patients with nonalcoholic steatohepatitis (nash). it markedly reduced liver fat content but with significant side effects [70] . in a phase 2 trial in patients with type 2 diabetes and chronic idiopathic constipation, ngm282 significantly improved bowel function by accelerating gastric emptying and colonic transit [81] . furthermore, ngm282 was further tested in mouse models and human patients with cholestasis and primary biliary cholangitis, showing efficacy in significantly reducing bile acid levels and improving hepatic inflammatory injury and fibrosis [84, 88, 89] . fgf21 is a prime lipid catabolic factor that regulates energy balance. however, the physiological roles and pharmacological effects of fgf21-driven metabolic axes are multifaceted [2, 7, 90] (fig. 3) . fgf21 was discovered as a driver of glucose uptake in adipocytes and a pparαdependent hepatic starvation hormone [48, 50, 51] . in mice, fgf21 levels are induced when calories are restricted or when glucose is low to allow fats to be burned for energy supply. the increasing levels of fgf21 drive diverse aspects of the adaptive starvation response, including stimulation of hepatic fatty acid oxidative for ketone body production during prolonged fasting and starvation. whether this action of fgf21 is autocrine/paracrine in the liver or endocrine in adipose tissues through adipose lipolysis and fatty acid oxidation is a matter of debate. the liver is a major contributor to the circulating fgf21 levels, which is associated with hepatic fat content and adiposity but inversely associated with serum glucose levels [91] [92] [93] . the hepatic expression of fgf21 is responsive not only to starvation but also to a broad spectrum of cellular, metabolic, or pathological changes in the liver as well as systemic metabolic perturbations [7, 69, 94] . as fgf21 is incapable of activating fgfr4-klb complex [67] , which is predominant in the liver that expresses fgfr1-klb with lower levels, hepatic fgf21 acts mainly as an endocrine factor to drive the metabolic pathways in peripheral tissues, including wat, bat, muscle, heart, kidney, and cns that express high levels of fgfr1/2/3-klb, leading to the correction of metabolic derangements and amelioration of metaflammation and stress damage (fig. 3) [7,94] . although the liver is unlikely a major direct target of fgf21, the effects of fgf21 on the liver are prominent. in addition to its role as a regulator of integrated hepatic metabolism in multiple aspects [48, 50, [95] [96] [97] [98] , including fatty acid oxidation, ketogenesis, gluconeogenesis, and macronutrient preference, fgf21 counteracts hepatic pathologies in response to a number of nutritional and chemical insults, including ketogenic diet, high fat diet, high fructose diet, methionine and choline deficient diet, ethanol-supplemented diet, and diethylnitrosamine [99] [100] [101] [102] [103] . under a chronic obesogenic diet, fgf21-deficient mice developed a spectrum of progressive fatty liver disease, including simple hepatosteatosis to nash, fibrosis, and hcc, which is the most lethal complication of this disorder. these findings highlight the role of fgf21 metabolic axis as a defensive barrier for the deleterious stress damage caused by metabolic disorders in the liver [104] . current clinical trials with fgf21 analogs show promising efficacy against nafld, nash, and fibrosis without noticeable adverse side effects [73] . acting on wat and bat, the fgf21 axis drives an array of catabolic effects, including insulin-independent glucose uptake, lipid droplet expansion inhibition, lipolysis, fatty acid oxidation, white adipocyte beigeing, and thermogenic dissipation of energy [79, 105, 106] . this route of action has been proposed as a major endocrine axis of fgf21 for insulin sensitization; lowering of systemic glucose, triacylglycerol, and ldl; fighting against obesity, diabetes, fatty liver diseases, hyperlipidemia, and associated comorbidities; and achieving metabolic health [2, 73, 74, 107] . some of these effects are likely mediated by adipokines, such as ccl11 and adiponectin, as shown in mice [108, 109] . in cold-induced nonshivering thermogenesis or exercise stress condition, bat also becomes a source of endocrine fgf21 in a β-adrenergicand campdependent manner, which in turn facilitates mitochondrial genesis, oxidative capacity, uncoupling, and heat generation, leading to adaptation to cold conditions and core body temperature maintenance [110] [111] [112] . in line with the beneficial effects of fgf21 on maintaining metabolic homeostasis during diverse adverse conditions, pharmacological fgf21 markedly extends the lifespan of mice by blunting the growth hormone/insulinlike growth factor-1 signaling pathway in the liver without reducing food intake or affecting longevity-associated markers of nad + metabolism, amp kinase, and mtor signaling pathways [113] . the thymus functions in producing new t cells for the immune system, but with age, it becomes fatty and loses the ability to produce a sufficient amount of new t cells, which is an important cause of increased risks of infections, obesity, diabetes, and certain cancer types, leading to reduced lifespan in the elderly people. the fgf21 level in thymic epithelial cells is several folds higher than that in the liver. the high level of fgf21 is proposed to protect thymus from the agerelated fatty degeneration and to increase the production of new t cells to bolster immune function, thereby lowering the incidence of diseases and promoting longevity [114] . the acinar cell compartment in the pancreas expresses the highest levels of fgf21 constitutively among tissues, but contributes little to the circulation [56, 115] . acinar cells appear to be both the dominant source and target (via fgfr1-klb complex) of pancreatic fgf21. the high levels of fgf21 is proposed to act as an exocrine pancreas secretagogue to stimulate pancreatic digestive enzyme secretion and pancreatic juice flow to the intestine, thereby relieving potential self-digestion caused proteostasis stress and protecting pancreas from pancreatitis, including but not limited to those caused by high-fat diet, pancreatic toxins, and alcoholism [116] . although islets express significantly lower amounts of fgf21, acinar cell derived or endocrine fgf21 helps protect against fatty pancreas, high-fat diet induced islet hyperplasia, and inflammatory damage [117] [118] [119] . demyelination in the cns can cause severe neurological deficits, such as multiple sclerosis and neurological dysfunction. pancreatic fgf21 acts on oligodendrocyte precursor cells to promote the remyelination process, leading to better recovery of neurological functions in mice [120] . the liver is the major organ of origin of endocrine fgf21 in response to a broad spectrum of stress conditions. the hepatic and pharmacological fgf21 drive multiple signal axes in multiple tissues/organs, resulting in multifaceted beneficiary metabolic effects, including promoting (green arrow) glucose, lipid, and energy homeostasis; offsetting metabolic derangements; and preventing (red long-tailed "t" sign) metaflammation, inflammatory tissue damage, and tissue-specific pathogenesis, including obesity, type 2 diabetes, fatty liver disease, metabolic syndrome, and associated comorbidities. faa: free fatty acids. black semicircular arrows indicate possibility of paracrine mode of fgf21 within local tissue environment. exposure to alcohol or sugar induces hepatic fgf21 through chrebp, which then acts on the hypothalamus reward pathway to suppress the desire for sugar and alcohol in favor of drinking water in mice depending on the β-adrenergic circuit [97, 98, 121, 122] . this finding may represent a new hydration pathway that is independent of the classical renin-angiotensin-aldosterone thirsty pathway in the kidney in response to nutritional stress, suggesting a previously underappreciated association of water intake to metabolism through the fgf21 metabolic axis. a human rs838133 allele in fgf21 is associated with higher alcohol and sugar intake and higher blood pressure and waist-hip ratio, with lower total body-fat percentage [123] . comparison of the genomes of more than 105 000 light and heavy social drinkers also identifies a variation in the rs11940694 locus of the klb gene in association with the aversion for alcohol [124] . neuronal cell stress reaction, such as those caused by disturbances in the mitochondria and endoplasmic reticulum (er), is an important factor in the development of neurodegenerative diseases. studies found that the integrated stress response induces neuronal fgf21, which presumably serves to attenuate stress and neural damage [125] . in addition to the liver, pancreas, and adipose tissues, cardiac muscle produces fgf21 in response to cardiac stress, cardio exercise, and endurance training [126, 127] , which then speeds up glucose uptake, lipid catabolism, and energy metabolism, and protects against cardiovascular stress damage, apoptosis, and heart dysfunctions, such as cardiac hypertrophy, myopathy, steatosis, ischemic infarction, and atherosclerosis [128] [129] [130] [131] [132] . through a multiorgan crosstalk, hepatic fgf21 drives the expression of angiotensin-converting enzyme 2 in adipocytes and renal cells, which hydrolyzes angiotensin ii to active vasodilator angiotensin-(1-7) in the renin-angiotensin system, thereby alleviating angiotensin ii-associated hypertension and reversing vascular damage [133] . skeletal muscle under the bouts of exercise or stress, such as mitochondrial myopathies, also induces fgf21 expression [134] [135] [136] . in turn, fgf21 acts on muscle and adipose tissue to reduce lipid load by increasing lipolysis, fatty acid utilization, energy expenditure, and insulin sensitivity, thereby preventing diet-induced obesity and insulin resistance [137] [138] [139] [140] . hepatic fgf21 acts on the paraventricular nucleus in the hypothalamus to drive the release of corticotropin-releasing factor, which then stimulates the involuntary sympathetic nerve activity. this leads to the activation of brown adipose tissue by upregulaing ucp1 and increases of glucose uptake, lipolysis, mitochondrial oxidation of fatty acids and glucose, body heat generation, and weight loss [141, 142] . the increase in corticotropin-releasing factor levels may also stimulate the pituitary gland to release adrenocorticotrophic hormone and subsequent corticoster-one production in adrenal cortex, leading to increased hepatic gluconeogenesis during prolonged fasting to prevent hypoglycemia [143] . hepatic fgf21 acts on the suprachiasmatic nucleus (scn) in the hypothalamus to suppress the vasopressin-kisspeptin and gonadotropinreleasing hormone signaling cascade, which then inhibit the proestrus surge in luteinizing hormone from anterior pituitary gland, thereby contributing to female infertility in response to nutritional challenge, such as prolonged starvation [144] . the scn action of fgf21 may also alter circadian behavior [145] . by increasing neuropeptide y levels and y1 receptor activation, the hypothalamus action of fgf21 may decrease locomotive activity, metabolic rate, and body temperature, leading to torpor under nutrition limitation [146] . fgf21 may also act on the hippocampus to decrease reactive oxygen species and inflammatory damage, thus decreasing brain cell damage and improving cognition [147, 148] . the endocrine fgf21 axes as well as the paracrine fgf21 axes within the local tissue compartments have been shown in many tissues and organs to counteract stress response and attenuate stress-ensued inflammation and inflammatory damage [7, 104, 117] . therefore, fgf21 is not only a stress-responsive or -induced factor but also an anti-stress and anti-inflammatory factor. the stress-offsetting effects, in particular the anti-inflammatory activities, can be attributed to the metabolic effects of fgf21 axes that prevent fatty degeneration, gluco-lipotoxicity, oxidative and er stress, and inflammatory and immune cell infiltration. these metabolic activities may be mediated in part through efficient and durable systemic and local glycemic and lipidemic control, improvement of insulin sensitivity, and promotion of lipid catabolism (lipolysis and fatty acid oxidation), adipose beigeing, and futile energy expenditure in adipose tissues, local adipocytes, and brain in both ucp1-dependent and adrenergic sympathetic nervous system-dependent mechanisms [79, 105, 106, 141, 149, 150] . as a result, fgf21 effectively reverses hepatic steatosis in obese mice and clinical obese patients [73, 105, 151] . furthermore, the pharmacological fgf21 analogs and fgfr1-klb agonists have been shown to directly improve the spectrum of adverse components of metabolic syndrome, including central obesity, insulin resistance, fasting hyperglycemia, dyslipidemia, systemic hypertension, and fatty liver, which are the major risk factors for cardiovascular disease (cvd), type 2 diabetes mellitus, chronic kidney disease (ckd), and all-cause mortality [2, 7, 73, 74, 107] . the fgf21 axes suppress atherosclerotic plaque by reducing hypercholesterolemia, oxidative stress, and smooth muscle cell proliferation via adiponectin-dependent and adiponectinindependent mechanisms [129] . fgf21-deficient mice developed significant islet hyperplasia and periductal lymphocytic inflammation upon chronic challenge of an obesogenic high-fat diet, indicating a protective role of fgf21 in compensatory islet hyperplasia and pancreatic inflammation associated with obesity [117, 152] . fgf21 directly suppresses triglyceride levels and lipid accumulation in kidney tissues, thereby reducing lipotoxicity, oxidative stress, inflammation, glomerular abnormalities, fibrotic renal injury in diabetic nephropathy, while deficiency of fgf21 aggravates these conditions [153, 154] , indicating a defensive role of fgf21 against kidney pathogenesis associated with obesity and diabetes. the anti-stress and anti-inflammatory effects of fgf21 may be also attributable to its direct action on nonmetabolic cells and non-metabolic activities. fgf21 directly inhibits cardiomyocyte apoptosis, oxidative stress, myocardial injury, thereby reducing the risk of pathological cardiac remodeling and dysfunction, cardiac hypertrophy, myocardial ischemia, and heart failure in ischemic heart tissue and diabetic cardiomyopathy [130, 155] . fgf21 protects the pancreas from caerulein-and larginine-induced pancreatitis, acinar cell injury, and fibrosis in mice [118, 119, 156] . fgf21 acts directly on renal mesangial cells to reduce glucose reabsorption and prevent hyperglycemia-induced fibrogenesis in db/db mice [157, 158] . interestingly, recent evidence supports that fgf21 can directly act on inflammatory and immune cells to attenuate inflammation and inflammatory damage. fgf21 activates thp-1-derived macrophages to promote cholesterol efflux, oxidized low-density lipoprotein (oxldl) uptake, and foam cell formation and inhibits macrophage inflammatory capacity through the nrf2 pathway [156, 159, 160] . adipose tissue is an endocrine organ and plays an active role in the inflammation in obesity that can favor cvd and ckd progression by inducing a chronic and low-grade inflammation via secreted proinflammatory adipokines and cytokines. studies in diet-induced obesity and pancreatitis models indicate that fgf21 promotes anti-inflammatory macrophage polarization in adipose depots and pancreas, wat browning, and insulin sensitivity, thereby effectively preventing adipose tissues from adapting proinflammatory profiles and the pancreas from inflammatory fibrosis [109, 156, 159, 161] . interestingly, fgf21 was found highly expressed in neutrophils and monocytes among circulating leukocytes and stimulates phagocytosis, glucose uptake, and reactive oxygen species production in a nadph oxidase-dependent manner in the neutrophil-like hl-60 and monocytic thp-1 cells [162] [163] [164] . in the type ii collagen-induced arthritis mouse model, fgf21 acts on the spleen to reduce inflammatory il-17, tnf-α, il-1β, il-6, il-8, and mmp3 and the number of splenic th17 cells, thereby alleviating arthritis severity [165] . these studies highlight the potential mediator role of fgf21 in innate immunity and inflammatory disorders. the direct impact of fgf21 on the function of inflammatory and immune cells and associated health consequences is yet to be validated. fgf23 is a key hormonal regulator of phosphate, vitamin d, and calcium metabolism, and its metabolic axes drive a complex interorgan crosstalk network for bone health and systemic mineral balance (fig. 4) [2, 61, [166] [167] [168] . osteoblastic cells in osseous tissue are the major source of fgf23 in response to elevated calcitriol, increased phosphate and calcium burdens, increased parathyroid hormone, iron and magnesium loss, and active bone remodeling in a vitamin d receptor dependent mechanism. acting on kidneys that express the fgfr1-kl complex, the fgf23 signal axis represses the expression of npt2a and npt2c, the sodium-phosphate cotransporters in the proximal tubule, thereby decreasing reabsorption and increasing secretion of phosphate in renal brush border membrane vesicles. another important function of this bone to kidney fgf23 signal axis is suppressing the expression of 25-hydroxyvitamin d3-1-α-hydroxylase and stimulating the expression of 1,25-dihydroxyvitamin d(3) 24-hydroxylase, thereby inhibiting the production of active calcitriol in renal proximal tubules, which subsequently inhibits the expression of npt2b and phosphate absorption in the apical brush border of small intestine. the bone fgf23 acts on the basolateral fgfr1-kl complex in the renal distal tubules to increase the intracellular transport of fully glycosylated trpv5 from the golgi apparatus to the plasma membrane, thereby stimulating calcium reabsorption in distal renal tubules and preventing calcium loss [169] . these fgf23-associated axes also directly increase the membrane abundance of the na + :clcotransporter ncc in distal renal tubules, and thus, increase sodium reabsorption, plasma volume, and blood pressure [170] . this change may be a new cause of high blood pressure and heart disease under the modern processed phosphaterich foods. bone fgf23 also acts on the parathyroid gland to inhibit the production and secretion of parathyroid hormone (fig. 4) [171] , which then reduces serum calcium through its effects on the bone, kidney, and intestine. high serum fgf23 levels in patients with ckd decrease calcitriol, thereby contributing to the development of secondary hyperparathyroidism, which has a crucial role in increasing the levels of fgf23 because the parathyroid hormone stimulates fgf23 expression. recent studies revealed the potential roles of the fgf23 axis in suppressing erythropoiesis in bone marrow. erythroid progenitor cells highly express fgf23 and fgfr-kl, suggesting that they are both a source and a target of fgf23. the loss of fgf23 or injection of an fgf23-blocking peptide in mice results in increased erythropoiesis, reduced erythroid cell apoptosis, and increased renal and bone marrow erythropoietin (epo) expression with increased circulating epo levels. on the other hand, the increased epo or acute blood loss increases fgf23 expression in the bone marrow with a concomitant increase in serum fgf23 [172, 173] . a recent study suggests that fgf23 is involved in the association between functional iron deficiency, increased epo levels, and death. the further elucidation of the role of the epo-fgf23 signaling axis in hereditary anemia and chronic hemolytic diseases, ckd, and mineralization disorders will add to the understanding of the pathophysiology of these diseases and life expectancy and will inform new treatment strategies for the diseases. current evidence indicates that fgf23 is more structurally unique than fgf19 and 21 [5] . fgf23 contains a conserved furin-sensitive 176 rhtr 179 cleavage site near the c-terminus, which inactivates the intact fgf23 upon cleavage, leading to signal attenuation. the biological importance of this activity control mechanism is demonstrated by point mutations (e.g., r176q, r179q, and r179w) of this site, which results in cleavage-resistant fgf23 and increased circulating levels of active fgf23, in autosomal dominant hypophosphatemic and vitamin ddeficient rickets characterized by renal phosphate wasting, hypophosphatemia, rickets, osteomalacia, leg deformities, short stature, bone pain, and dental abscesses [168, 174, 175] . fgf23 levels are increased and may play important roles in other hereditable and acquired phosphate wasting disorders, including x-linked dominant hypophosphatemic rickets, autosomal recessive hypophosphatemic rickets, hypophosphatemic rickets associated with mccune-albright syndrome/fibrous dysplasia of bone, and linear sebaceous nevus syndrome [176, 177] . the increased fgf23 levels are also found in acquired phosphate wasting disorders in some tumor types, such as the benign mesenchymal neoplasm phosphaturic mesenchymal tumor, causing tumor-induced osteomalacia, a paraneoplastic syndrome [168] . during post-translational modification, fgf23 is glycosylated at thr-178 in the cleavage site by galnt3, which facilitates its secretion and protects the protein from being broken down, suggesting a novel posttranslational regulatory model of fgf23 involving competing o-glycosylation and proteolytic processing to determine the level of secreted active fgf23 [178] . the importance of this glycosylation modification is demonstrated by inactivating galnt3 mutations that render fgf23 susceptible to fig. 4 fgf23 metabolic axis. the bone-derived fgf23 drives signal axes to promote (green arrows) the metabolic homeostasis of phosphate, vitamin d, and calcium through a complex interorgan crosstalk network for bone health and systemic mineral balance. the bone to the kidney axis of fgf23 is central to the metabolic roles of fgf23, which inhibits (red long-tailed "t" sign) the reabsorption of phosphate and the production of active calcitriol in renal proximal tubules while increasing the calcium and sodium reabsorption in renal distal tubules. the bone to parathyroid axis of fgf23 inhibits the production and secretion of parathyroid hormone that also plays critical roles in mineral and vitamin d balance. proteolysis [179, 180] , thereby reducing circulating intact hormone levels and leading to autosomal recessive familial tumoral calcinosis that manifests with hyperphosphatemic and massive calcium deposits in the skin and subcutaneous tissues throughout the body. consistently, at least seven mutations in the conserved backbone of fgf23, such as s71g, m96t, s129f, and f157l, destabilize the tertiary structure and render it susceptible to degradation, thereby resulting in autosomal recessive familial tumoral calcinosis with hyperphosphatemia [2, 5, [181] [182] [183] [184] . patients with ckd have increased serum levels of phosphate as well as fgf23, which lead to increased uptake of calcium by the kidneys, resulting in vascular calcification. this explains the cvd complications, such as cardiac hypertrophy and congestive heart failure, in patients with ckd [185, 186] . the inhibition of fgf23 or its axis could be a strategy to bring cvd and vascular calcification under control. the fgf23 level in patients with ckd can even indicate their life expectancy. the dysregulation of calcium levels can have an array of serious health consequences. chronic hypocalcemia can potentially lead to heart failure, nervous system and muscle disorders, and encephalopathy, while hypercalcemia can increase the risk of kidney stones, cause muscle weakness, and worsen psychological issues, such as dementia and depression. this may explain some current observations that people with high serum fgf23 can be at risk of dementia, and that mice lacking fgf23 exhibit defective learning and memory problems similar to those seen in kl-deficient mice [187, 188] . the three members of the metabolic fgfs, including fgf19, 21, and 23, share a conserved core structure of βtrefoil fold but diverge in functions from other mitogenic members of the fgf family during evolution (fig. 1) . these metabolic fgfs acquire specific structural elements that endow them with abilities to function via an endocrine mode and to bind new accessory receptors that have strict expression patterns in metabolic tissues. although metabolic fgfs still signal through the transmembrane fgfr tyrosine kinases as the mitogenic fgfs, these new properties divert their functions to metabolic regulation. as such, fgf19, 21, and 23 drive a wide range of diverse metabolic axes that function in maintaining the homeostasis of bile acids, glucose, lipids, energy, and minerals; offsetting detrimental metabolic derangements; and achieving optimal metabolic health without an overt effect on cell proliferation and population growth. in this sense, each of the metabolic axes of fgf19, 21, and 23 stands alone as a driver of specific metabolic effects with important physiological functions and pathological consequences. therefore, these axes together constitute the "fgf metabolic axis," which is a new term that we start to call hereafter, with broad-spectrum pathophysiological roles and consequences on the quality of survival (fig. 1) . i would like to acknowledge the long-term contributions of many members of my wenzhou fgf team to the fgf field research as i cited in the text that made the idea of "the fgf metabolic axis" possible. i thank dr. yongde luo for the expert assistance on the conceptual and practical aspects of the manuscript and dr. jin-san zhang and dr. jian xiao for their assistance. i apologize to those whose works have not been cited here due to the limited discussion scope on this evolving field. this work is supported by the national key r&d program of china (no. 2017yfa0506000, xiaokun li). xiaokun li declares no conflict of interests. this manuscript is a review article and does not involve a research protocol requiring approval by the relevant institutional review board or ethics committee. open access this article is licensed under a creative commons attribution 4.0 international license, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the creative commons licence, and indicate if changes were made. the images or other third party material in this article are included in the article's creative commons licence, unless indicated otherwise in a credit line to the material. if material is not included in the article's creative commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. to view a copy of this licence, visit https://creativecommons.org/ licenses/by/4.0/. the fgf family: biology, pathophysiology and therapy emerging structure-function paradigm of endocrine fgfs in metabolic diseases fibroblast growth factors, old kids on the new block threedimensional structure of human basic fibroblast growth factor comparative study of heparinpoloxamer hydrogel modified bfgf and afgf for in vivo wound healing efficiency heparin-based coacervate of fgf2 improves dermal regeneration by asserting a synergistic role with cell proliferation and endogenous facilitated vegf for cutaneous wound healing a thermosensitive heparin-poloxamer hydrogel bridges afgf to treat spinal cord injury therapeutics targeting fgf signaling network in human diseases anticancer molecules targeting fibroblast growth factor receptors hypotensive activity of fibroblast growth factor fibroblast growth factor-16 is a growth factor for embryonic brown adipocytes in vitro and in vivo analyses reveal profound effects of fibroblast growth factor 16 as a metabolic regulator a pparg-fgf1 axis is required for adaptive adipose remodelling and metabolic homeostasis uncoupling the mitogenic and metabolic functions of fgf1 by tuning fgf1-fgf receptor dimer stability maratos-flier e. hepatic fibroblast growth factor 21 is regulated by pparα and is a key mediator of hepatic lipid metabolism in ketotic states fibroblast growth factor 15 functions as an enterohepatic signal to regulate bile acid homeostasis endocrine regulation of the fasting response by pparα-mediated induction of fibroblast growth factor 21 fgf-21 as a novel metabolic regulator klotho co-receptors inhibit signaling by paracrine fgf8 subfamily ligands metabolic regulator βklotho interacts with fibroblast growth factor receptor 4 (fgfr4) to induce apoptosis and inhibit tumor cell proliferation evolutionary history and functional implications of protein domains and their combinations in eukaryotes tissue-specific expression of βklotho and fibroblast growth factor (fgf) receptor isoforms determines metabolic activity of fgf19 and fgf21 research resource: comprehensive expression atlas of the fibroblast growth factor system in adult mouse identification of a domain within peroxisome proliferator-activated receptor γ regulating expression of a group of genes containing fibroblast growth factor 21 that are selectively repressed by sirt1 in adipocytes glucose induces fgf21 mrna expression through chrebp activation in rat hepatocytes regulation of fgf21 expression and secretion by retinoic acid receptor-related orphan receptor α liver x receptor negatively regulates fibroblast growth factor 21 in the fatty liver induced by cholesterol-enriched diet vitamin d receptor in chondrocytes promotes osteoclastogenesis and regulates fgf23 production in osteoblasts 25-dihydroxyvitamin d3 upregulates fgf23 gene expression in bone: the final link in a renal-gastrointestinal-skeletal axis that controls phosphate transport the link between fibroblast growth factor 21 and sterol regulatory element binding protein 1c during lipogenesis in hepatocytes ablation of gp78 in liver improves hyperlipidemia and insulin resistance by inhibiting srebp to decrease lipid biosynthesis adipose fibroblast growth factor 21 is up-regulated by peroxisome proliferatoractivated receptor γ and altered metabolic states activating transcription factor 4-dependent induction of fgf21 during amino acid deprivation differential specificity of endocrine fgf19 and fgf21 to fgfr1 and fgfr4 in complex with klb structures of β-klotho reveal a 'zip code'-like mechanism for endocrine fgf signalling stressed liver and muscle call on adipocytes with fgf21 ngm282 for treatment of nonalcoholic steatohepatitis: a multicentre, randomised, double-blind, placebo-controlled, phase 2 trial effect of ngm282, an fgf19 analogue, in primary sclerosing cholangitis: a multicenter, randomized, double-blind, placebo-controlled phase ii trial ngm282 improves liver fibrosis and histology in 12 weeks in patients with nonalcoholic steatohepatitis bms-986036), a pegylated fibroblast growth factor 21 analogue, in patients with non-alcoholic steatohepatitis: a randomised, doubleblind, placebo-controlled, phase 2a trial a long-acting fgf21 molecule, pf-05231023, decreases body weight and improves lipid profile in non-human primates and type 2 diabetic subjects randomized trial of the anti-fgf23 antibody krn23 in x-linked hypophosphatemia elevated cholesterol metabolism and bile acid synthesis in mice lacking membrane tyrosine kinase receptor fgfr4 fibroblast growth factor 19 increases metabolic rate and reverses dietary and leptin-deficient diabetes transgenic mice expressing human fibroblast growth factor-19 display increased metabolic rate and decreased adiposity the breadth of fgf21's metabolic actions are governed by fgfr1 in adipose tissue a new mechanism for bile acid diarrhea: defective feedback inhibition of bile acid biosynthesis effects of ngm282, an fgf19 variant, on colonic transit and bowel function in functional constipation: a randomized phase 2 trial antibodymediated inhibition of fibroblast growth factor 19 results in increased bile acids synthesis and ileal malabsorption of bile acids in cynomolgus monkeys a role for fibroblast growth factor 19 and bile acids in diabetes remission after roux-en-y gastric bypass a nontumorigenic variant of fgf19 treats cholestatic liver diseases high expression of the bile salt-homeostatic hormone fibroblast growth factor 19 in the liver of patients with extrahepatic cholestasis fibroblast growth factor 19 regulates skeletal muscle mass and ameliorates muscle wasting in mice a mouse model of hepatocellular carcinoma: ectopic expression of fibroblast growth factor 19 in skeletal muscle of transgenic mice engineered fibroblast growth factor 19 reduces liver injury and resolves sclerosing cholangitis in mdr2-deficient mice ngm282 for treatment of patients with primary biliary cholangitis: a multicenter, randomized, double-blind, placebo-controlled trial fibroblast growth factor 21: a versatile regulator of metabolic homeostasis circulating levels of fgf-21 in obese youth: associations with liver fat content and markers of liver damage dynamic change of serum fgf21 levels in response to glucose challenge in human increased serum fgf21 levels in patients with nonalcoholic fatty liver disease a dozen years of discovery: insights into the physiology and pharmacology of fgf21 fgf21 is an endocrine signal of protein restriction a critical role for chrebp-mediated fgf21 secretion in hepatic fructose metabolism fgf21 mediates endocrine control of simple sugar intake and sweet taste preference by the liver fgf21 regulates sweet and alcohol preference fibroblast growth factor 21 limits lipotoxicity by promoting hepatic fatty acid activation in mice on methionine and cholinedeficient diets forced expression of hepatocyte-specific fibroblast growth factor 21 delays initiation of chemically induced hepatocarcinogenesis role of fibroblast growth factor 21 in the early stage of nash induced by methionine-and choline-deficient diet maratos-flier e. fibroblast growth factor 21 (fgf21) is robustly induced by ethanol and has a protective role in ethanol associated liver injury fibroblast growth factor 21 protects against acetaminopheninduced hepatotoxicity by potentiating peroxisome proliferatoractivated receptor coactivator protein-1α-mediated antioxidant capacity in mice maratos-flier e. deficiency of fibroblast growth factor 21 (fgf21) promotes hepatocellular carcinoma (hcc) in mice on a long term obesogenic diet fgf21-fgfr1 coordinates phospholipid homeostasis, lipid droplet function, and er stress in obesity treating diabetes and obesity with an fgf21-mimetic antibody activating the βklotho/fgfr1c receptor complex the effects of ly2405319, an fgf21 analog, in obese human subjects with type 2 diabetes adiponectin mediates the metabolic effects of fgf21 on glucose homeostasis and insulin sensitivity in mice the fgf21-ccl11 axis mediates beiging of white adipose tissues by coupling sympathetic nervous system to type 2 immunity irisin and fgf21 are cold-induced endocrine activators of brown fat function in humans thermogenic activation induces fgf21 expression and release in brown adipose tissue liver derived fgf21 maintains core body temperature during acute cold exposure the starvation hormone, fibroblast growth factor-21, extends lifespan in mice prolongevity hormone fgf21 protects against immune senescence by delaying age-related thymic involution fibroblast growth factor 21 is not required for the antidiabetic actions of the thiazoladinediones fgf21 is an exocrine pancreas secretagogue maratos-flier e. fibroblast growth factor 21 (fgf21) protects against high fat diet induced inflammation and islet hyperplasia in pancreas silencing of the fibroblast growth factor 21 gene is an underlying cause of acinar cell injury in mice lacking mist1 fibroblast growth factor 21 reduces the severity of cerulein-induced pancreatitis in mice peripherally derived fgf21 promotes remyelination in the central nervous system fgf21 is a sugar-induced hormone associated with sweet intake and preference in humans the hormone fgf21 stimulates water drinking in response to ketogenic diet and alcohol a common allele in fgf21 associated with sugar intake is associated with body shape, lower total body-fat percentage, and higher blood pressure neuronal mitochondrial dysfunction activates the integrated stress response to induce fibroblast growth factor 21 fibroblast growth factor 21 protects against cardiac hypertrophy in mice divergent effects of resistance and endurance exercise on plasma bile acids, fgf19, and fgf21 in humans fibroblast growth factor 21 is induced upon cardiac stress and alters cardiac lipid homeostasis fibroblast growth factor 21 prevents atherosclerosis by suppression of hepatic sterol regulatory element-binding protein-2 and induction of adiponectin in mice endocrine protection of ischemic myocardium by fgf21 from the liver and adipose tissue fibroblast growth factor-21 prevents diabetic cardiomyopathy via ampk-mediated antioxidation and lipid-lowering effects in the heart fibroblast growth factor 21 protects the heart from apoptosis in a diabetic mouse model via extracellular signal-regulated kinase 1/2-dependent signalling pathway fgf21 prevents angiotensin ii-induced hypertension and vascular dysfunction by activation of ace2/angiotensin-(1-7) axis in mice autophagy deficiency leads to protection from obesity and insulin resistance by inducing fgf21 as a mitokine fgf-21 as a biomarker for muscle-manifesting mitochondrial respiratory chain deficiencies: a diagnostic study exercise alleviates obesity-induced metabolic dysfunction via enhancing fgf21 sensitivity in adipose tissues opa1 deficiency promotes secretion of fgf21 from muscle that prevents obesity and insulin resistance acute exercise increases fibroblast growth factor 21 in metabolic organs and circulation fibroblast growth factor-21 protects human skeletal muscle myotubes from palmitate-induced insulin resistance by inhibiting stress kinase and nf-kb fgf21 is an akt-regulated myokine fgf21 acts centrally to induce sympathetic nerve activity, energy expenditure, and weight loss maratos-flier e. central fibroblast growth factor 21 browns white fat via sympathetic action in male mice fgf21 maintains glucose homeostasis by mediating the cross talk between liver and brain during prolonged fasting fgf21 contributes to neuroendocrine control of female reproduction fgf21 regulates metabolism and circadian behavior by acting on the nervous system role of pparα in the control of torpor through fgf21-npy pathway: from circadian clock to seasonal change in mammals fgf21 attenuates high-fat diet-induced cognitive impairment via metabolic regulation and anti-inflammation of obese mice fibroblast growth factor 21 protects mouse brain against d-galactose induced aging via suppression of oxidative stress response and advanced glycation end products formation fibroblast growth factor 21 action in the brain increases energy expenditure and insulin sensitivity in obese rats fgf21 promotes metabolic homeostasis via white adipose and leptin in mice fibroblast growth factor 21 reverses hepatic steatosis, increases energy expenditure, and improves insulin sensitivity in diet-induced obese mice loss of fibroblast growth factor 21 action induces insulin resistance, pancreatic islet hyperplasia and dysfunction in mice attenuation of hyperlipidemia-and diabetes-induced early-stage apoptosis and late-stage renal dysfunction via administration of fibroblast growth factor-21 is associated with suppression of renal inflammation fibroblast growth factor 21 improves insulin resistance and ameliorates renal injury in db/db mice liver-heart crosstalk controls il-22 activity in cardiac protection after myocardial infarction fibroblast growth factor 21 ameliorates pancreatic fibrogenesis via regulating polarization of macrophages fibroblast growth factor 21 ameliorates high glucose-induced fibrogenesis in mesangial cells through inhibiting stat5 signaling pathway fibroblast growth factor 21 regulates glucose metabolism in part by reducing renal glucose reabsorption fgf21 increases cholesterol efflux by upregulating abca1 through the erk1/2-pparg-lxrα pathway in thp1 macrophage-derived foam cells fibroblast growth factor 21 (fgf21) inhibits macrophage-mediated inflammation by activating nrf2 and suppressing the nf-kb signaling pathway fibroblast growth factor 21 increases insulin sensitivity through specific expansion of subcutaneous fat fibroblast growth factor 21 expressions in white blood cells and sera of patients with gestational diabetes mellitus during gestation and postpartum fgf-21 elevated il-10 production to correct lpsinduced inflammation a novel function for fibroblast growth factor 21: stimulation of nadph oxidase-dependent ros generation treatment of cia mice with fgf21 down-regulates th17-il-17 axis human fibroblast growth factor-23 mutants suppress na + -dependent phosphate co-transport activity and 1α,25-dihydroxyvitamin d3 production targeted ablation of fgf23 demonstrates an essential physiological role of fgf23 in phosphate and vitamin d metabolism cloning and characterization of fgf23 as a causative factor of tumor-induced osteomalacia fgf23 promotes renal calcium reabsorption through the trpv5 channel fgf23 regulates renal sodium handling and blood pressure the parathyroid is a target organ for fgf23 in rats erythropoietin induces bone marrow and plasma fibroblast growth factor 23 during acute kidney injury acute blood loss stimulates fibroblast growth factor 23 production autosomal dominant hypophosphataemic rickets is associated with mutations in fgf23 fgf-23 inhibits renal tubular phosphate transport and is a phex substrate fgf-23 in fibrous dysplasia of bone and its relationship to renal phosphate wasting elevated fibroblast growth factor-23 in hypophosphatemic linear nevus sebaceous syndrome polypeptide galnac-transferase t3 and familial tumoral calcinosis. secretion of fibroblast growth factor 23 requires o-glycosylation tumoral calcinosis presenting with eyelid calcifications due to novel missense mutations in the glycosyl transferase domain of the galnt3 gene the role of mutant udp-n-acetyl-α-d-galactosaminepolypeptide n-acetylgalactosaminyltransferase 3 in regulating serum intact fibroblast growth factor 23 and matrix extracellular phosphoglycoprotein in heritable tumoral calcinosis an fgf23 missense mutation causes familial tumoral calcinosis with hyperphosphatemia a novel homozygous missense mutation in fgf23 causes familial tumoral calcinosis associated with disseminated visceral calcification a novel mutation in fibroblast growth factor 23 gene as a cause of tumoral calcinosis a new missense mutation in fgf23 gene in a male with hyperostosis-hyperphosphatemia syndrome (hhs) fgf23 induces left ventricular hypertrophy fibroblast growth factor 23 and left ventricular hypertrophy in chronic kidney disease circulating fibroblast growth factor 23 levels and incident dementia: the framingham heart study impairment of spatial learning and memory in transgenic mice overexpressing human fibroblast growth factor-23 