Supplementary issue:02 - 119 - Analysis of two different outbreaks in PICU of a tertiary care centre in two different time zones Fatima Khan1, Uzma Tayyaba1, Asfia Sultan1, Anees Akhtar1, Shariq Ahmed1 1Department of Microbiology, JN Medical College, AMU, Aligarh Correspondence: Dr. Asfia Sultan Assistant Professor, Department of Microbiology, JN Medical College, Faculty of Medicine, Aligarh Muslim University, Aligarh-202002, U.P., India ORCID ID: 0000-0003-0343-9513 E-mail ID: drasfia@gmail.com ABSTRACT Klebsiella pneumoniae is one of the common cause of nosocomial outbreak. With the emergence of multi-drug resistance, clinical management has become a serious challenge. The present study is a comparative analysis of changing trends of antimicrobial resistance (AMR) between two outbreaks of Klebsiella pneumoniae in PICU occurring over a gap of 8 years. Materials & Methods: 1054 Blood culture samples received from paediatric patients over a period of 6 months were tested by BacT/Alert-3D & VITEK-2 system. Survey of environmental samples was also conducted in Paediatric ICU. All the findings were then analysed with the data from another outbreak of MDR Klebsiella pneumoniae in PICU reported from the same institute in the year 2015. Result: From 154 (14.6%) positive paediatric blood samples, Gram-negative bacilli was most commonly isolated (75/154, 48.7%), followed by Gram-positive cocci (53, 34.4%). Amongst gram-negative isolates Klebsiella pneumoniae was commonest (40/75, 53.3%), followed by Pseudomonas speices (17.3%), Escherichia coli (16%), Burkholderia cepacia-complex (6.7%), and Acinetobacter baumanii-complex (4%). Out of 40 Klebsiella pneumoniae, 90% were extensively-drug resistant (XDR), i.e resistant to cephalosporins, carbapenems, aminoglycosides, fluoroquinolone, Co-trimaxazole as well as Tigecyclin. 45% of these patients were admitted in the PICU. However, previous outbreak reported 90 Klebsiella pneumoniae (43.3% of all isolates) over a period of 4 month, out of which 45.5% were MDR & 18.8% were ESBL producers. However, mailto:drasfia@gmail.com Supplementary issue:02 - 120 - Carbapenems were effective in majority of them (98.9%), which is a contrast to our present finding (97.5% resistance). Moreover, multiple MDR strains of Klebsiella species (7/16 isolates, 43.7%) were obtained from PICU samples in previous outbreak, with antibiogram similar to that of patient samples. Similarly PICU sampling in the current outbreak also yielded XDR Klebsiella pneumoniae from medicine table, with similar profile that is seen in patients sample. Conclusion: On analysis of two outbreaks from same hospital in two different time zones, we observed changing trend in resistance pattern, as Carbapenems were effective in previously isolated MDR strains, but now XDR stains are isolated which are resistant to carbapenems as well. Urgent and effective measures are needed to restrain emergence and transmission of XDR strains within the hospital. Stringent infection control practices should be advocated and strictly followed. KEYWORDS: Outbreak, Klebsiella pneumoniae, PICU, MDR, XDR Introduction Bloodstream infections (BSI) are common cause of febrile illness and is associated with high morbidity and mortality worldwide. The bacterial pathogen Klebsiella pneumoniae is a major cause of nosocomial infections, especially in immunocompromised individuals (1). Increasing antimicrobial resistance among these notorious bacteria, particularly with emergence of carbapenem resistance seen in recent years (2), and their worldwide dissemination (3), possess serious threat to public health worldwide. Because infections caused by these Carbapenem- resistant Klebsiella pneumoniae (CRKP) are difficult to treat (4), and are associated with high mortality (5). CRKP strains produces a variety of Cabapenamases enzymes and/or Extended- spectrum Beta-lactamases (ESBLs) along with porin loss and efflux pump overexpression, leading to emergence of multi-drug resistant (MDR), extensively-drug resistant (XDR) and pan-drug resistant (PDR) bacteria. (6) Another aggravating factor is the tendency of Klebsiella pneumoniae to cause outbreaks in healthcare facilities, particularly in ICUs (7–9). Moreover, Klebsiella pneumoniae has propensity to silently colonize patients and hospital personnel, without causing any sign of infections. These carriers function as reservoirs and help in its dissemination, making control of outbreak a difficult challenge (9). Furthermore, because of extensive use of antibiotics in hospital setting, multiple Supplementary issue:02 - 121 - drug resistance have selective advantage in perseverance of such nosocomial bacteria in hospitalised patients as well as in hospital environment (7). To apply productive infection control measures, it is crucial to know relevant environmental contamination and in-hospital transmission routes. Therefore the present study was done to describe changing pattern of antimicrobial resistance (AMR) between two outbreaks of Klebsiella pneumoniae associated with bacteriemia in paediatric patients reported from the same institute occurring over a gap of 8 years (10). Materials & Methods: The study was conducted in a tertiary care centre in two different time zone. Present data was taken from July 2022 to December 2022. Blood samples for culture were collected from paediatric patients taking all sterile precautions. Blood culture positive samples identified by BacT/ALERT 3D (bioMérieux) automated system were plated on 5% sheep Blood agar, MacConkey agar and Chocolate agar. Bacterial identification as well as antimicrobial sensitivity of positive growth was done by VITEK-2 (bioMérieux) automated system. Survey of environmental samples was also conducted in Paediatric ICU. Environmental samples were taken from various sites including patient bed, door handle, medicine table, medicine basket, Ventilator knob, suction pipe, warmer and disinfectant bottle. Air culture was done by settle plate method. Samples were inoculated on brain heart infusion broth, from which subculture was done next day on 5% sheep blood agar and MacConkey agar. Bacterial identification was done by conventional methods and confirmed by VITEK-2. All the findings of present outbreak was then compared & analysed with another outbreak of MDR Klebsiella pneumoniae reported in paediatric blood culture isolates from the same institute 8 years back in the year 2015. (10) Result & Discussion: A total of 1834 blood samples were received in the enteric lab over the period of 6 months, out of which 1054 (57.5%) were from paediatric patients. Out of these 1054 paediatric blood samples, Supplementary issue:02 - 122 - 154 (14.6%) were identified positive. Gram negative bacilli was most commonly isolated (75/154, 48.7%), followed by Gram positive cocci (53, 34.4%), then yeast like fungi (24, 16.9%). Amongst gram negative bacilli, maximum isolates were of Klebsiella pneumoniae (40/75, 53.3%), followed by Pseudomonas speices (13/75, 17.3%), Escherichia coli (12, 16%), Burkholderia cepacian complex (5, 6.7%), Acinetobacter baumanii complex (3, 4%), Citrobacter species (1, 1.3%), and Enterobacter cloacae complex (1, 1.3%) as depicted in Figure1. Figure1 Out of 40 paediatric samples from which Klebsiella was isolated 27 (67.5%) were of infants, which includes 17 neonates (42.5%). 45% of these patients were admitted in the PICU, and 36.8% of patients were intubated and were on mechanical ventilation. Antimicrobial susceptibility profile by VITEK-2 system showed 97.5% (n=39) of total 40 Klebsiella pneumoniae were resistant to beta lactams antibiotics (namely Ampicillin, Amoxicillin/clavulanic acid, Piperacillin/Tazobactum, Cefuroxime, Ceftriaxone, Supplementary issue:02 - 123 - Cefoperazone/Sulbactum, Cefepime, Ertapenem, Imipenem, Meropenem), Tigecyclin, as well as aminoglycosides (Amikacin & Gentamicin). Ciprofloxacin resistance was seen in 95% (n=38), and Co-trimaxazole resistance in 90% (n=36). Thus, 90% of the strains were Extensively drug resistant (XDR), which is defined as non-susceptible to at least one agent in all but two or fewer antimicrobial categories (6). Another outbreak of MDR Klebsiella pneumoniae was reported 8 years back in the year 2015 in paediatric blood culture isolates from the same institute (10). In the previous outbreak 90 Klebsiella pneumoniae (43.3% of all isolates) were isolated over a period of 4 month. Out of which 71.1% were from PICU. Antimicrobial susceptibility testing showed resistance rates of 62.3% to amikacin, 84.5% to gentamycin, 83.4% to ceftriaxone, 85.6% to cefoperazone, 66.7% to cefoperazone+sulbactum, 68.9% to piperacillin+tazobactum and 53.4% to levofloxacin (10). 45.5% were reported as MDR. Fortunately, Imipinem was resistant in only 1.1% isolates, which is contrast to our present study, where carbapenems are resistant in 97.5% of Klebsiella isolates. Figure 2 depicts increasing trend of resistance pattern from 2015 outbreak to the present outbreak, especially to Carbapenems. Alarming rise in resistance rates of Carbapenems seen over 8 years, is a cause of concern as this leaves us with only few available treatment options (11). Supplementary issue:02 - 124 - Figure2 For successful containment of an outbreak it is very important to understand how transmission is occurring. To identify the source of contamination within the PICU environmental samples were taken. Air culture by settle plate method grew Bacillus and Coagulase negative Staphylococcus species. No growth was seen in sample from patient bed side and disinfectant bottle. Different bacteria isolated from different samples were Bacillus, Coagulase negative Staphylococcus species (CONS), Acinetobacter baumanii complex, Klebsiella pneumoniae, Pseudomonas stutzeri, and Enterococcus species (Table). Apart from variety of different bacteria isolated from different sites, XDR Klebsiella pneumoniae was isolated from medicine table, with similar antimicrobial resistance profile that is seen with patients samples, which could be source of contamination. Strict infection control practices were advocated to stop any further cross contamination to the patients. Table 1 S.No. Site Organism isolated Susceptibility profile A m ik a c in A z tr e o n a m G e n ta m ic in C e ft ri a x o n e C e fo p e ra z o n e + S u lb a c tu m C e fe p im e Im ip e n e m M e ro p e n e m C ip ro fl o x a c in C o tr im a x a z o l e T ig e c y c li n M in o c y c li n 1. Air Bacillus - - - - - - - - - - - - CONS - - - - - - - - - - - - 2. Medicine Table Pseudomonas stutzeri S R S - - R - S - S - S Klebsiella pneumoniae R - R R R R R R R R R - 3. Medicine Basket Pseudomonas species S S R - - S - R - R - R Bacillus - - - - - - - - - - - - 4. Syringe tray CONS - - - - - - - - - - - - Enterococcus species - - - - - - - - - - - - 5. Ventilator knob Acinetobacter baumanii complex - - R R R R R R R R R - 6. Suction pipe Klebsiella pneumoniae S - S R S R S S R S R - 7. Warmer Acinetobacter baumanii complex - - S R R R R R R R I - 8. Door handle Pseudomonas species S S R - - R - S - S - S Supplementary issue:02 - 125 - Bacillus - - - - - - - - - - - - Similarly in the previous outbreak of 2015, multiple MDR strains of Klebsiella species (7/16 isolates, 43.7%) were obtained from environmental sampling, with antibiogram similar to that of patient samples (10). Most of the environmental MDR strains were still susceptible to Imipenem, which is contradictory to our present study where carbapenems are ineffective. Presence of such XDR strains in healthcare environment is unsettling, since cross contamination of these organisms may cause grave danger to patients. It warrants the need to create improved cleaning protocols and ensuring strict adherence to infection control practices within the PICU. Our hospital is a tertiary referral centre as well as a teaching hospital. Environment such as ours, with enormous burden of patients, unrestrained movement of medical as well as paramedical students inside wards for teachings and rounds and duty exchange within nursing staff, lead to reduced compliance with infection prevention practices and facilitate transmission of multidrug resistant organisms (MDRO), from either hospital environment or hands of healthcare staff to the patients (12). Poor hand hygiene practices among staff is the most plausible reason. Strict infection prevention practices among healthcare staff have been described as an effective tool to combat MDRO transmission within the healthcare system (13). In recent years Klebsiella pneumoniae have been identified as major cause of nosocomial outbreaks in the ICU (8,9). Patients admitted in the Intensive critical units of hospitals are always at a risk of acquiring nosocomial infections, because of their critical condition, low immunity and prolonged hospitalisation (14). Furthermore, infants and newborns admitted in PICU are at greater risk due to their immature immune system, low birth-weight and regular usage of antimicrobials and invasive devices (15). Numerous studies have described Klebsiella pneumoniae as most common etiological agent of outbreak within ICU (16,17) and Neonatal ICUs (NICU) (18,19). Similar to our report, nosocomial outbreak of XDR Klebsiella pneumoniae have been reported worldwide, like report of nosocomial BSI by XDR Klebsiella pneumoniae in a teaching hospital in China by Wenzi B et al (20), and by Gasper et al in Brazil in 2022 (21) and so on. XDR isolates show high resistance to beta-lactams antibiotics (including carbapenems and beta-lactam/ beta-lactamase inhibitors combination), Supplementary issue:02 - 126 - aminoglycosides, fluoroquinolones, tigecycline and/or polymyxins, and may lead to emergence of pan drug-resistant (PDR) isolates. (22,23). The rising prevalence and worldwide dissemination of these XDR isolates, especially in healthcare system, is a major public health threat, as there are very few effective therapeutic options left (24) and are associated with high mortality (25). Conclusions Our study describes an outbreak of XDR Klebsiella pneumoniae associated with bacteriemia in paediatric patients. On comparative analysis of two outbreaks from same hospital in two different time zones, we observed changing trend in resistance pattern seen in nosocomial pathogen. In the present outbreak XDR stains are isolated which are resistant to carbapenems as well, which were effective in previously isolated MDR strains. This makes us wonder, from here, where do we go next? Urgent and effective measures are needed to restrain emergence and transmission of XDR strains within the hospital. Vigorous infection control practices should be advocated and strictly followed. Also, high mortality associated with BSIs caused by XDR Klebsiella pneumoniae should alert clinicians to institute effective and rational antibiotic policy. References: 1. Wyres KL, Holt KE. Klebsiella pneumoniae Population Genomics and Antimicrobial- Resistant Clones. Trends Microbiol. 2016 Dec;24(12):944–56. 2. Campos AC, Albiero J, Ecker AB, Kuroda CM, Meirelles LEF, Polato A, et al. Outbreak of Klebsiella pneumoniae carbapenemase–producing K pneumoniae: A systematic review. American Journal of Infection Control. 2016 Nov 1;44(11):1374–80. 3. Lee CR, Lee JH, Park KS, Kim YB, Jeong BC, Lee SH. Global Dissemination of Carbapenemase-Producing Klebsiella pneumoniae: Epidemiology, Genetic Context, Treatment Options, and Detection Methods. Front Microbiol. 2016 Jun 13;7:895. 4. Paczosa MK, Mecsas J. Klebsiella pneumoniae: Going on the Offense with a Strong Defense. Microbiol Mol Biol Rev. 2016 Sep;80(3):629–61. Supplementary issue:02 - 127 - 5. Munoz-Price LS, Poirel L, Bonomo RA, Schwaber MJ, Daikos GL, Cormican M, et al. Clinical epidemiology of the global expansion of Klebsiella pneumoniae carbapenemases. Lancet Infect Dis. 2013 Sep;13(9):785–96. 6. Magiorakos AP, Srinivasan A, Carey RB, Carmeli Y, Falagas ME, Giske CG, et al. Multidrug-resistant, extensively drug-resistant and pandrug-resistant bacteria: an international expert proposal for interim standard definitions for acquired resistance. Clinical Microbiology and Infection. 2012 Mar 1;18(3):268–81. 7. Peña C, Pujol M, Ardanuy C, Ricart A, Pallares R, Liñares J, et al. Epidemiology and Successful Control of a Large Outbreak Due to Klebsiella pneumoniae Producing ExtendedSpectrum β-Lactamases. Antimicrob Agents Chemother. 1998 Jan;42(1):53–8. 8. Jarvis WR, Munn VP, Highsmith AK, Culver DH, Hughes JM. The epidemiology of nosocomial infections caused by Klebsiella pneumoniae. Infect Control. 1985 Feb;6(2):68–74. 9. Asensio A, Oliver A, González-Diego P, Baquero F, Pérez-Díaz JC, Ros P, et al. Outbreak of a multiresistant Klebsiella pneumoniae strain in an intensive care unit: antibiotic use as risk factor for colonization and infection. Clin Infect Dis. 2000 Jan;30(1):55–60. 10. Khan F, Siddiqui N, Sultan A, Rizvi M, Aqbari S, Shukla I, et al. Klebsiella pneumoniae Outbreak in Paediatric Ward: Detection and Prevention. IntJCurrMicrobiolAppSci. 2015;(Special Issue-1):81–7. 11. Logan LK, Weinstein RA. The Epidemiology of Carbapenem-Resistant Enterobacteriaceae: The Impact and Evolution of a Global Menace. The Journal of Infectious Diseases. 2017 Feb 15;215(suppl_1):S28–36. 12. Casewell M, Phillips I. Hands as route of transmission for Klebsiella species. Br Med J. 1977 Nov 19;2(6098):1315–7. 13. Kim NH, Han WD, Song KH, Seo HK, Shin M jin, Kim TS, et al. Successful containment of carbapenem-resistant Enterobacteriaceae by strict contact precautions without active surveillance. Am J Infect Control. 2014 Dec;42(12):1270–3. 14. GiViTI Steering Committee, Bertolini G, Nattino G, Tascini C, Poole D, Viaggi B, et al. Mortality attributable to different Klebsiella susceptibility patterns and to the coverage of Supplementary issue:02 - 128 - empirical antibiotic therapy: a cohort study on patients admitted to the ICU with infection. Intensive Care Med. 2018 Oct;44(10):1709–19. 15. Ruiz E, Rojo-Bezares B, Sáenz Y, Olarte I, Esteban I, Rocha-Gracia R, et al. Outbreak caused by a multi-resistant Klebsiella pneumoniae strain of new sequence type ST341 carrying new genetic environments of aac(6′)-Ib-cr and qnrS1 genes in a neonatal intensive care unit in Spain. International Journal of Medical Microbiology. 2010 Nov 1;300(7):464–9. 16. Snitkin ES, Zelazny AM, Thomas PJ, Stock F, NISC Comparative Sequencing Program Group, Henderson DK, et al. Tracking a hospital outbreak of carbapenem-resistant Klebsiella pneumoniae with whole-genome sequencing. Sci Transl Med. 2012 Aug 22;4(148):148ra116. 17. Xu L, Sun X, Ma X. Systematic review and meta-analysis of mortality of patients infected with carbapenem-resistant Klebsiella pneumoniae. Ann Clin Microbiol Antimicrob. 2017 Mar 29;16(1):18. 18. Gastmeier P, Groneberg K, Weist K, Rüden H. A cluster of nosocomial Klebsiella pneumoniae bloodstream infections in a neonatal intensive care department: identification of transmission and intervention. American Journal of Infection Control. 2003 Nov 1;31(7):424–30. 19. Rastogi V, Nirwan PS, Jain S, Kapil A. Nosocomial outbreak of septicaemia in neonatal intensive care unit due to extended spectrum β-lactamase producing Klebsiella pneumoniae showing multiple mechanisms of drug resistance. Indian J Med Microbiol. 2010;28(4):380–4. 20. Bi W, Liu H, Dunstan RA, Li B, Torres VVL, Cao J, et al. Extensively Drug-Resistant Klebsiella pneumoniae Causing Nosocomial Bloodstream Infections in China: Molecular Investigation of Antibiotic Resistance Determinants, Informing Therapy, and Clinical Outcomes. Front Microbiol. 2017 Jun 30;8:1230. 21. Gaspar GG, Tamasco G, Abichabki N, Scaranello AFT, Auxiliadora-Martins M, Pocente R, et al. Nosocomial Outbreak of Extensively Drug-Resistant (Polymyxin B and Carbapenem) Klebsiella pneumoniae in a Collapsed University Hospital Due to COVID-19 Pandemic. Antibiotics (Basel). 2022 Jun 17;11(6):814. 22. Andrade LN, Vitali L, Gaspar GG, Bellissimo-Rodrigues F, Martinez R, Darini ALC. Expansion and evolution of a virulent, extensively drug-resistant (polymyxin B-resistant), QnrS1- Supplementary issue:02 - 129 - , CTX-M-2-, and KPC-2-producing Klebsiella pneumoniae ST11 international high-risk clone. J Clin Microbiol. 2014 Jul;52(7):2530–5. 23. Gaspar GG, Bellissimo-Rodrigues F, Andrade LN de, Darini AL, Martinez R. Induction and nosocomial dissemination of carbapenem and polymyxin-resistant Klebsiella pneumoniae. Rev Soc Bras Med Trop. 2015;48(4):483–7. 24. Lim TP, Cai Y, Hong Y, Chan ECY, Suranthran S, Teo JQM, et al. In vitro pharmacodynamics of various antibiotics in combination against extensively drug-resistant Klebsiella pneumoniae. Antimicrob Agents Chemother. 2015 May;59(5):2515–24. 25. Falagas ME, Tansarli GS, Karageorgopoulos DE, Vardakas KZ. Deaths Attributable to Carbapenem-Resistant Enterobacteriaceae Infections - Volume 20, Number 7—July 2014 - Emerging Infectious Diseases journal - CDC. [cited 2023 Feb 24]; Available from: https://wwwnc.cdc.gov/eid/article/20/7/12-1004_article