ORIGINAL_ARTICLE Pulsed-field gel electrophoresis used for typing of extended-spectrum-β-lactamases- producing Escherichia coli Isolated from infant ҆ s respiratory and digestive system Escherichia coli infections are becoming increasingly difficult to treat because of emerging antimicrobial resistance, mostly to expanded-spectrum cephalosporins, due to the production of extended-spectrum β-lactamases (ESBLs).Despite extensive studies of ESBL- producing E.coli in adult patients, there is a lack of information about the epidemiology and spread of ESBL organisms in pediatric population. The aim of this study was to examine the gastrointestinal tract as an endogenous reservoir for the respiratory tract colonization with ESBL- E. coli in children, hospitalized because of the severity of the respiratory illness. The study group consists of 40 children with ESBL-producing E. coli strains isolated from the sputum and from the rectal samples. A control group of 15 E. coli isolated from rectal swabs of healthy children were included in the analysis. The comparison of the strains was done by using antimicrobial susceptibility patterns of the stains, and pulsed field gel electrophoresis was performed for molecular typing, using XbaI digestion. 90% of the compared pairs of strains in the study group were with identical antimicrobial susceptibility patterns and indistinguishable in 79.2% by the obtained PFGE – profiles.33.3% (5/15) of confirmed E. coli strains from the control group were found to be ESBL – producers. Resulting band profiles of all isolates demonstrated presence of 12 pulsotypes, with 100% similarity within the pulsotypes. Although, some isolates obtained from different patients were genetically indistinguishable, these strains were not hospital acquired, as none of the patients satisfied the criteria for hospital acquired pneumonia, and there was a lack of an obvious transmission chain. All ESBL –E. coli isolated from sputum in clinical cases were obtained from patients under the age of one. According to the resistance profile of the compared pairs and the PFGE comparison of all isolates, it can be concluded that the gastrointestinal tract is the main reservoir of ESBL-E. coli. Small age in infants is a risk factor for translocation of bacteria, enabling the colonization of the respiratory tract. https://macvetrev.mk/Files/Article/2020/10.2478/macvetrev-2018-0016/macvetrev-2018-0016.pdf 2018-10-15T09:00:00 133 141 10.2478/macvetrev-2018-0016 ESBL-producing Escherichia coli resistance profile GUT colonization PFGE- typing Gorica Popova gorica.popova@yahoo.com false 1 Department of Clinical Microbiology, Institute for Respiratory Diseases in Children, Skopje, Republic of Macedonia LEAD_AUTHOR Dean Jankuloski false 2 Department of Molecular Microbiology, Faculty of Veterinary Medicine, Food Institute,University Ss Cyril and Methodius in Skopje, Republic of Macedonia AUTHOR Benjamin Felix false 3 French Agency for Food, Environmental and Occupational Health and Safety, Laboratory for Food Safety, Paris, France AUTHOR Katerina Boskovska false 4 Pediatric Department, Institute for Respiratory Diseases in Children, Skopje, Republic of Macedonia AUTHOR Biljana Stojanovska - Dimzovska false 5 Department of Microbiology, Faculty of Veterinary Medicine, Food Institute, University Ss Cyril and Methodius in Skopje AUTHOR Velibor Tasic false 6 Nephrology Department, University Children, s Hospital, Skopje, Republic of Macedonia AUTHOR Katerina Blagoevska false 7 Department of Molecular Microbiology, Faculty of Veterinary Medicine, Food Institute,University Ss Cyril and Methodius in Skopje, Republic of Macedonia AUTHOR Akil, I., Yilmaz, O., Kuruepe, S., Deqwrli, K., Kavukcu, S. (2006). Influence of oral intake of Saccharomyces boulardii on Escherichia coli in enteric flora. Pediatr Nephrol. 21, 807-810. PMid:16703374 1 10.1007/s00467-006-0088-4 Backhed, F., Ley, R.E., Sonnenburg, J.L., Peterson, D.A., Gordon, J.I. (2005). Host bacterial mutualism in the human intestine. Science 307, 1915-1920. PMid:15790844 2 10.1126/science.1104816 Qin, J., Li, R., Raes, J., Arumugam, M., Burgdorf, K.S., Manichanh, C., et al. (2010). A human gut microbial gene catalogue established by metagenomic sequencing. Nature 464 (7285):59-65. PMid:20203603 PMCid:PMC3779803 3 10.1038/nature08821 Penders, J., Thijs, C., Vink, C., Stelma, F.F., Snijders, B., Kummeling, I., van den Brandt, P.A., Stobberingh, E.E. (2006). Factors influencing the composition of the intestinal microbiota in early infancy. Pediatrics 118 (2):511-521. PMid:16882802 4 10.1542/peds.2005-2824 Jernberg, C., Lofmark, S., Edlund, C., Jansson, J.K. (2010). Long –term ecological impacts of antibiotic administration on the human intestinal microbiota. Microbiology 156, 3216-3223. PMid:20705661 5 10.1099/mic.0.040618-0 de la Cochetiere, M.F., Durand, T., Lepage, P., Bourreille, A., Galmiche, J.P., Dore, J. (2005). Resilience of the dominant human fecal microbiota upon short-course antibiotic challenge. J Clin Microbiol 43, 5588-5592. PMid:16272491 PMCid:PMC12≊7 6 10.1128/JCM.43.11.5588-5592.2005 Perez-Cobas, A.E., Gosalbes, M.J., Friedrichs, A., Knecht, H., Artacho, A., Eismann, K., et al. (2013). Gut microbiota disturbance during antibiotic therapy:a multi-omic approach. Gut 62, 1591-1601. PMid:23236009 PMCid:PMC3812899 7 10.1136/gutjnl-2012-303184 Paterson, L.D., Bonomo, A.R. (2005). Extended spectrum beta lactamases:a clinical update. Clin Microbiol Rev. 18(4):657-686. PMid:16223952 PMCid:PMC1265908 8 10.1128/CMR.18.4.657-686.2005 Bush, K., Jacoby, G.A. (2010). Update functional classification sheme of β–lactamases. Antimicrob Agents Chemother. 54(3):969-976. PMid:19995920 PMCid:PMC2825993 9 10.1128/AAC.01009-09 Philippon, A., Labia, R., Jacoby, G. (1989). Extended-spectrum β-lactamases.Antimicrob Agents Chemother. 33, 1131–1136. 10 10.1128/AAC.33.8.1131 Srivastava, A., Singhal, N., Goel, M., Virdi, J.S., Kumar, M. (2014). CBMAR:a comprehensive β-lactamase molecular annotation resource. Database (Oxford). 2014:bau111 PMid:25475113 PMCid:PMC4255060 11 10.1093/database/bau111 Bonnet, R. (2004). Growing group of extended spectrum:the CTX-M enzymes. Antimicrob Agent Chemother. 48, 1-14 PMCid:PMC310187 12 10.1128/AAC.48.1.1-14.2004 Lewis, J.S., Herrera, M., Wickes, B., Patterson, J.E., Jorgensen, J.H. (2007). First report of the emergence of CTX-M-type extended-spectrum β-lactamases (ESBLs) as the predominant ESBL isolated in a U.S. Health Care System. Antimicrob Agents Chemother. 51(11):4015-4021. PMid:17724160 PMCid:PMC2151438 13 10.1128/AAC.00576-07 Alobwede, I., Mzali, F.H., Livermore, D.M., Hentige, J., Todd, N., Hawkey, P.M. (2003). CTX-M extended-spectrum beta-lactamases arrives in UK. J Antimicrob Chemother. 51. 470-471. PMid:12562729 14 10.1093/jac/dkg096 Mendonça, N., Ferreira, E., Louro, D., ARSIP Participants, Caniça, M. (2009). Molecular epidemiology and antimicrobial susceptibility of extended- and broad-spectrum beta-lactamase-producing Klebsiella pneumoniae isolated in Portugal. Int J Antimicrob Agents. 34(1):29-37. PMid:19272757 15 10.1016/j.ijantimicag.2008.11.014 Kiratisin, P., Apisarnthanarak, A., Laesripa, C., Saifon, P. (2008). Molecular characterization and epidemiology of extended-spectrum-beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae isolates causing health care-associated infection in Thailand, where the CTX-M family is endemic. Antimicrob Agents Chemother. 52(8):2818-2824. PMid:18505851 PMCid:PMC2493136 16 10.1128/AAC.00171-08 The European Committee on Antimicrobial Susceptibility Testing. Breakpoint tables for interpretation of MICs and zone diameters. Version 4.0, 2014. http://www.eucast.org. 17 The EUCAST subcommittee for detection of resistance mechanisms and specific resistance of clinical and/or epidemiological importance. EUCAST guidelines for detection of resistance mechanisms and specific resistances of clinical and/or epidemiological importance.Version 1.0, 2013. http://www.eucast.org. 18 Ribot, E.M., Fair, M.A., Gautom, R., Cameron, D.N., Hunter, S.B., Swaminathan, B., Barrett, T.J. (2006). Standardization of pulsed-field gel electrophoresis protocols for the subtyping of Escherichia coli O157:H7, Salmonella, and Shigella for PulseNet. Foodborne Pathog Dis. 3(1):59-67. PMid:16602980 19 10.1089/fpd.2006.3.59 Caprioli, A., Maugliani, A., Michelacci, V., Morabito, S. (2014). Molecular typing of Verocytotoxin - producing E. coli (VTEC) strains isolated from food, feed and animals :state of play and standard operating procedures for pulsed field gel electrophoresis (PFGE) typing, profiles interpretation and curation. EFSA journal EN704, 55. 20 Barrett, T.J., Gerner-Smidt, P., Swaminathan, B. (2006). Interpretation of pulsed-field gel electrophoresis patterns in foodborne disease investigations and surveillance. Foodborne Pathog Dis. 3(1):20-31. PMid:16602976 21 10.1089/fpd.2006.3.20 Peters, T.M., Maguire, C., Threlfall, E.J., Fisher, I.S., Gill, N., Gatto, A.J. (2003). The Salm-gene project - a European collaboration for DNA fingerprinting for food-related salmonellosis. Euro Surveill. 8, 46-50. PMid:12631975 22 10.2807/esm.08.02.00401-en Winokur, P.L., Cantón, R., Casellas, J.M., Legakis, N. (2001). Variations in the prevalence of strains expressing an extended-spectrum β-lactamase phenotype and characterization of isolates from Europe, the Americas, and the Western Pacific region. Clin Infect Dis. 32, 94–103. PMid:11320450 23 10.1086/320182 Pitout, J.D., Hanson, N.D., Church, D.L., Laupland, K.B. (2004). Population-based laboratory surveillance for Escherichia coli-producing extended-spectrum β-lactamases:importance of community isolates withblaCTX-M genes. Clin Infect Dis. 38, 1736–1741. PMid:15227620 24 10.1086/421094 Ben-Ami, R., Schwaber, M.J., Navon-Venezia, S., Schwartz, D., Giladi, M., Chmelnitsky, I., Leavitt, A., Carmeli, Y. (2006). Influx of extended-spectrum β-lactamase-producing Enterobacteriaceae into the hospital. Clin Infect Dis. 42, 925–934. PMid:16511754 25 10.1086/500936 Chandramohan, L., Revell, P.A. (2012). Prevalence and molecular characterization of extended-spectrum-β-lactamase-producing enterobacteriaceae in a pediatric patient population. Antimicrob Agents Chemother. 56(9):4765-4770. PMid:22733062 PMCid:PMC3421901 26 10.1128/AAC.00666-12 Mitchella, D.J., Mc Clurea, B.G., Tubmanb, T.R.J. (2001). Simultaneous monitoring of gastric and oesophageal pH reveals limitations of conventional oesophageal pH monitoring in milk fed infants. Arch Dis Child. 84, 273-276. PMCid:PMC1718697 27 10.1136/adc.84.3.273 Orozco-Levi, M., Torres, A., Ferrer, M., Piera, C., El-Ebiary, M., de la Bellacasa, J.P., Rodriguez-Roisin, R. (1995). Semirecumbent position protects from pulmonary aspiration but not completely from gastroesophageal re- flux in mechanically ventilated patients. Am J Respir Crit Care Med. 152, 1387–1390. PMid:7551400 28 10.1164/ajrccm.152.4.7551400 Davis, K.J., Johannigman, J.A., Campbell, R.S., Marraccini, A., Luchette, F.A., Frame, S.B., Branson, R.D. (2001). The acute effects of body position strategies and respiratory therapy in paralyzed patients with acute lung injury. Crit Care. 5, 81–87. PMid:11299066 PMCid:PMC30713 29 10.1186/cc991 Drakulovic, M.B., Torres, A., Bauer, T.T., Nicolas, J.M., Nogue, S., Ferrer, M. (1999). Supine body position as a risk factor for nosocomial pneumonia in mechanically ventilated patients:a randomised trial. Lancet 354, 1851–1858. 30 10.1016/S0140-6736(98)12251-1 Pingleton, S.K., Hinthorn, D.R., Liu, C. (1986). Enteral nutrition in patients receiving mechanical ventilation:multiple sources of tracheal colonization include the stomach. Am J Med. 80, 827–832. 31 10.1016/0002-9343(86)90623-6 Tablan, O.C., Anderson, L.J., Besser, R., Bridges, C., Hajjeh, R., Healthcare Infection Control Practices Advisory Committee, Centers for Disease Control and Prevention. (2004). Guidelines for preventing health-care–associated pneumonia, 2003:recommendations of the CDC and the Healthcare Infection Control Practices Advisory Committee. MMWR Recomm Rep. 53(RR-3):1–36. PMid:15048056 32 Prosperi, M., Veras, N., Azarian, T., Rathore, M., Nolan, D., Rand, K., Cook, R.L., Johnson, J., Morris, J.G., Salemil, M. (2013). Molecular epidemiology of community-associated methicillin-resistant Staphylococcus aureus in the genomic era:a cross-sectional study. Sci Rep. 3, 1902 PMid:23712667 PMCid:PMC3664956 33 10.1038/srep01902 Tschudin-Sutter, S., Frei, R., Dangel, M., Strauden A., Widmer, A.T. (2012). Rate of Transmission of extended-spectrum beta-lactamase–producing enterobacteriaceae without contact isolation. Clin Infect Dis. 55 (11):1505-1511. PMid:22955436 34 10.1093/cid/cis770 Jain, R., Kralovic, S.M., Evans, M.E., Ambrose, M., Simbartl, L.A., Obrosky, D.S., Render, M.L., Freyberg, R.W., Jarnigan, J.A., Muder, R.R., Miller, L.J., Roselle, G.A. (2011). Veterans affairs initiative to prevent methicillin-resistant Staphylococcus aureus infections. N Engl J Med. 364:1419-30. PMid:2148∼ 35 10.1056/NEJMoa1007474 Ostrowsky, B.E., Trick, W.E., Sohn, A.H., Quirk, S.B., Holt, S., Carson, L.A., Hill, B.C., Arduino, M.J., Kuehnert, M.J., Jarvis, W.R. (2001). Control of vancomycin-resistant enterococcus in health care facilities in a region. N Engl J Med. 344:1427-1433. PMid:11346807 36 10.1056/NEJM200105103441903