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Antimicrobial resistance (AMR) is a major health threat for public and animal health in the twenty-first century. In Ecuador, antibiotics have been used by the poultry industry for decades resulting in the presence of multi-drug resistant (MDR) bacteria in the poultry meat production chain, with the consequent risk for public health. This study evaluated the prevalence of ESBL/AmpC and mcr genes in third-generation cephalosporin-resistant Escherichia coli (3GC-R E. coli) isolated from broiler farms (animal component), broiler carcasses (food component), and human enteritis (human component) in Quito-Ecuador. Samples were collected weekly from November 2017 to November 2018. For the animal, food, and human components, 133, 335, and 302 samples were analyzed, respectively. Profiles of antimicrobial resistance were analyzed by an automated microdilution system. Resistance genes were studied by PCR and Sanger sequencing. From all samples, 122 (91.7%), 258 (77%), and 146 (48.3%) samples were positive for 3GC-R E. coli in the animal, food, and human components, respectively. Most of the isolates (472/526, 89.7%) presented MDR phenotypes. The ESBL blaCTX-M-55, blaCTX-M-3, blaCTX-M-15, blaCTX-M-65, blaCTX-M-27, and blaCTX-M-14 were the most prevalent ESBL genes while blaCMY-2 was the only AmpC detected gene. The mcr-1 gene was found in 20 (16.4%), 26 (10.1%), and 3 (2.1%) of isolates from animal, food, and human components, respectively. The implication of poultry products in the prevalence of ESBL/AmpC and mcr genes in 3GC-R must be considered in the surveillance of antimicrobial resistance.
Keywords: AmpC beta-lactamases, broiler farms, broiler carcasses, E. coli, extended-spectrum beta-lactamase (ESBL), human, mcr-1.
1. IACG. No time to wait: Securing the Future from Drug-Resistant Infections. WHO, editor (2019). Available online at: https://www.who.int/docs/ default-source/documents/no-time-to-wait-securing-the-future-fromdrug-resistant-infections-en.pdf?sfvrsn=5b424d7_6
2. O’Neill J. Tackling Drug-Resistant Infections Globally: Final Report and Recommendations. The Review on Antimicrobial Resistance. (2016). Available online at: https://amr-review.org
3. Van Boeckel TP, Brower C, Gilbert M, Grenfell BT, Levin SA, Robinson TP, et al. Global trends in antimicrobial use in food animals. Proc Natl Acad Sci USA. (2015) 112:5649–54. doi: 10.1073/pnas.1503141112
4. OECD/FAO. Agricultural Outlook 2016–2025. (2016). Available online at: https://doi.org/10.1787/agr_outlook-2016-10-en
5. Vinueza-Burgos C, Wautier M, Martiny D, Cisneros M, Van Damme I, De Zutter L. Prevalence, antimicrobial resistance and genetic diversity of Campylobacter coli and Campylobacter jejuni in Ecuadorian broilers at slaughter age. Poult Sci. (2017) 96:2366–74. doi: 10.3382/ps/pew487
6. Conave. Estadísticas Del Sect Avícola. (2019). Available online at: https://www. conave.org/informacion-sector-avicola-publico/
7. WHO. Critically Important Antimicrobials for Human Medicine: 5th Revision. Geneva: WHO (2017).
8. WHO. Global Priority List of Antibiotic-Resistant Batceria to Guide Research, Discovery, and Development of New Antibiotics. Geneva: WHO (2017). Available online at: https://www.who.int/news-room/detail/27-02-2017- who-publishes-list-of-bacteria-for-which-new-antibiotics-are-urgentlyneeded
9. Ortega-Paredes D, Barba P, Zurita J. Colistin-resistant Escherichia coli clinical isolate harbouring the mcr-1 gene in Ecuador. Epidemiol Infect. (2016) 144:2967–70. doi: 10.1017/S0950268816001369
10. Ortega-Paredes D, Haro M, Leoro-Garzón P, Barba P, Loaiza K, Mora F, et al. Multidrug-resistant Escherichia coli isolated from canine faeces in a public park in Quito, Ecuador. J Glob Antimicrob Resist. (2019) 18:263– 8. doi: 10.1016/j.jgar.2019.04.002
11. Yamamoto Y, Calvopina M, Izurieta R, Villacres I, Kawahara R, Sasaki M, et al. Colistin-resistant Escherichia coli with mcr genes in the livestock of rural small-scale farms in Ecuador. BMC Res Notes. (2019) 12:1– 5. doi: 10.1186/s13104-019-4144-0
12. Overdevest I, Willemsen I, Rijnsburger M, Eustace A, Xu L, Hawkey P, et al. Extended-spectrum β-lactamase genes of Escherichia coli in chicken meat and humans, The Netherlands. Emerg Infect Dis. (2011) 17:1216– 22. doi: 10.3201/eid1707.110209
13. Kluytmans JAJW, Overdevest ITMA, Willemsen I, Kluytmans-van den Bergh MFQ, van der Zwaluw K, Heck M, et al. Extended-spectrum β-lactamaseproducing Escherichia coli from retail chicken meat and humans: comparison of strains, plasmids, resistance genes, and virulence factors. Clin Infect Dis. (2013) 56:478–87. doi: 10.1093/cid/cis929
14. Moser KA, Zhang L, Spicknall I, Braykov NP, Levy K, Marrs CF, et al. The role of mobile genetic elements in the spread of antimicrobial-resistant Escherichia coli from chickens to humans in small-scale production poultry operations in rural Ecuador. Am J Epidemiol. (2018) 187:558–67. doi: 10.1093/aje/kwx286
15. Pacholewicz E, Liakopoulos A, Swart A, Gortemaker B, Dierikx C, Havelaar A, et al. Reduction of extended-spectrum-β-lactamase- and AmpC-β-lactamase-producing Escherichia coli through processing in two broiler chicken slaughterhouses. Int J Food Microbiol. (2015) 215:57– 63. doi: 10.1016/j.ijfoodmicro.2015.08.010
16. Mughini-Gras L, Dorado-García A, van Duijkeren E, van den Bunt G, Dierikx CM, Bonten MJM, et al. Attributable sources of community-acquired carriage of Escherichia coli containing β-lactam antibiotic resistance genes: a population-based modelling study. Lancet Planet Heal. (2019) 3:e357– 69. doi: 10.1016/S2542-5196(19)30130-5
17. Vinueza-Burgos C, Ortega-Paredes D, Narvaéz C, De Zutter L, Zurita J. Characterization of cefotaxime resistant Escherichia coli isolated from broiler farms in Ecuador. PLoS ONE. (2019) 14:e207567. doi: 10.1371/journal.pone.0207567
18. Bej AK, Dicesare JL, Haff L, Atlas RM. Detection of Escherichia coli and Shigella spp. in water by using the polymerase chain reaction and gene probes for UID. Appl Environ Microbiol. (1991) 57:1013–7.
19. CLSI. Antimicrobial Susceptibility Testing Standards. CLSI Supplement for Global Application.(2020). Available online at: http://em100.edaptivedocs.net/ GetDoc.aspx?doc=CLSIM100~ED30:2020&scope$=$user
20. Rebelo AR, Bortolaia V, Kjeldgaard JS, Pedersen SK, Leekitcharoenphon P, Hansen IM, et al. Multiplex PCR for detection of plasmidmediated colistin resistance determinants, mcr-1, mcr-2, mcr-3, mcr-4 and mcr-5 for surveillance purposes. Eurosurveillance. (2018) 23:1–11. doi: 10.2807/1560-7917.ES.2018.23.6.17-00672
21. Zankari E, Hasman H, Cosentino S, Vestergaard M, Rasmussen S, Lund O, et al. Identification of acquired antimicrobial resistance genes. J Antimicrob Chemother. (2012) 67:2640–4. doi: 10.1093/jac/ dks261
22. Hoek AHAM Van, Veenman C, Florijn A, Huijbers PMC, Graat EAM, Greeff S De, et al. Longitudinal study of ESBL Escherichia coli carriage on an organic broiler farm. J Antimicrob Chemother. (2018) 73:3298– 304. doi: 10.1093/jac/dky362
23. Apostolakos I, Mughini-Gras L, Fasolato L, Piccirillo A. Assessing the occurrence and transfer dynamics of ESBL/pAmpC-producing Escherichia coli across the broiler production pyramid. PLoS ONE. (2019) 14:e217174. doi: 10.1371/journal.pone.0217174
24. González D, Gallagher E, Zúñiga T, Leiva J, Vitas AI. Prevalence and characterization of β-lactamase-producing Enterobacteriaceae in healthy human carriers. Int Microbiol. (2019) 23:171– 7. doi: 10.1007/s10123-019-00087-z
25. Bagus Wasito E, Shigemura K, Osawa K, Fardah A, Kanaida A, Raharjo D, et al. Antibiotic susceptibilities and genetic characteristics of extendedspectrum beta-lactamase-producing Escherichia coli isolates from stools of pediatric diarrhea patients in Surabaya, Indonesia. Jpn J Infect Dis. (2017) 70:378–82. doi: 10.7883/yoken.JJID.2016.234
26. Reyes JA, Melano R, Cárdenas PA, Trueba G. Mobile genetic elements associated with carbapenemase genes in South American enterobacterales. Brazilian J Infect Dis. (2020) 24:231–8. doi: 10.1016/j.bjid.2020.03.002
27. Zurita J, Alcocer I, Ortega-Paredes D, Barba P, Yauri F, Iñiguez D, et al. Carbapenem-hydrolysing β-lactamase KPC-2 in Klebsiella pneumoniae isolated in Ecuadorian hospitals. J Glob Antimicrob Resist. (2013) 1:229– 30. doi: 10.1016/j.jgar.2013.06.001
28. Soria-Segarra C, Soria-Segarra C, Catagua-González A, Gutiérrez-Fernández J. Carbapenemase producing Enterobacteriaceae in intensive care units in Ecuador: results from a multicenter study. J Infect Public Health. (2019) 13:80–8. doi: 10.1016/j.jiph.2019.06.013
29. Ortega-Paredes D, Barba P, Mena-López S, Espinel N, Crespo V, Zurita J. High quantities of multidrug-resistant Escherichia coli are present in the Machángara urban river in Quito, Ecuador. J Water Health. (2020) 18:67– 76. doi: 10.2166/wh.2019.195
30. Lartigue MF, Poirel L, Poyart C, Réglier-Poupet H, Nordmann P. Ertapenem resistance of Escherichia coli. Emerg Infect Dis. (2007) 13:315– 7. doi: 10.3201/eid1302.060747
31. Vinueza C. Salmonella and Campylobacter in broilers at slaughter age: a possible source for carcasses contamination in Ecuador (Ph.D. thesis), Ghent University, Ghent, Belgium (2017).
32. Vieira DC, Lima WG, de Paiva MC. Plasmid-mediated quinolone resistance (PMQR) among Enterobacteriales in Latin America: a systematic review. Mol Biol Rep. (2020) 47:1471–83. doi: 10.1007/s11033-019-05220-9
33. Correia S, Poeta P, Hébraud M, Capelo JL, Igrejas G. Mechanisms of quinolone action and resistance: where do we stand? J Med Microbiol. (2017) 66:551– 9. doi: 10.1099/jmm.0.000475
34. Farajzadehsheikh A, Veisi H, Shahin M, Getso M, Farahani A. Frequency of quinolone resistance genes among extended-spectrum?-lactamase (ESBL)- producing Escherichia coli strains isolated from urinary tract infections. Trop Med Health. (2019) 47:1–7. doi: 10.1186/s41182-019-0147-8
35. Ortega-Paredes D, Zurita J. Integrones, plataformas bacterianas de recombinación. Rev Ecuat Med Cienc Biol. (2013) XXXIV:167–85. doi: 10.26807/remcb.v34i1-2.242
36. Donado-godoy P, Byrne BA, León M, Castellanos R, Vanegas C, Coral A, et al. Prevalence, resistance patterns, and risk factors for antimicrobial resistance in bacteria from retail chicken meat in Colombia. J Food Prot. (2015) 78:751– 9. doi: 10.4315/0362-028X.JFP-14-349
37. Bartoloni A, Sennati S, Di Maggio T, Mantella A, Riccobono E, Strohmeyer M, et al. Antimicrobial susceptibility and emerging resistance determinants (blaCTX-M, rmtB, fosA3) in clinical isolates from urinary tract infections in the Bolivian Chaco. Int J Infect Dis. (2016) 43:1–6. doi: 10.1016/j.ijid.2015.12.008
38. Cyoia PS, Koga VL, Nishio EK, Houle S, Dozois CM, De Brito KCT, et al. Distribution of ExPEC virulence factors, bla CTX-M, fosA3, and mcr-1 in escherichia coliisolated from commercialized chicken carcasses. Front Microbiol. (2019) 10:3254. doi: 10.3389/fmicb.2018.03254
39. Gardiner BJ, Stewardson AJ, Abbott IJ, Peleg AY. Nitrofurantoin and fosfomycin for resistant urinary tract infections: old drugs for emerging problems. Aust Prescr. (2019) 42:14–9. doi: 10.18773/austprescr.2019.002
40. Sheu CC, Chang YT, Lin SY, Chen YH, Hsueh PR. Infections caused by carbapenem-resistant Enterobacteriaceae: an update on therapeutic options. Front Microbiol. (2019) 10:80. doi: 10.3389/fmicb.2019.00080
41. Ramirez MS, Tolmasky ME. Amikacin: uses, resistance, and prospects for inhibition. Molecules. (2017) 22:2267. doi: 10.3390/molecules22122267
42. Castellanos LR, Donado-Godoy P, León M, Clavijo V, Arevalo A, Bernal JF, et al. High heterogeneity of Escherichia coli sequence types harbouring ESBL/AmpC genes on IncI1 plasmids in the Colombian poultry chain. PLoS ONE. (2017) 12:e170777. doi: 10.1371/journal.pone.0170777
43. Vinueza-Burgos C, Cevallos M, Ron-Garrido L, Bertrand S, De Zutter L. Prevalence and diversity of Salmonella serotypes in ecuadorian broilers at slaughter age. PLoS ONE. (2016) 11:e159567. doi: 10.1371/journal.pone.0159567
44. Salazar GA, Guerrero-López R, Lalaleo L, Avilés-Esquivel D, Vinueza-Burgos C, Calero-Cáceres W. Presence and diversity of Salmonella isolated from layer farms in central Ecuador: [Version 2; peer review: 2 approved]. F1000Research. (2019) 8:1–12. doi: 10.12688/f1000research.18233.2
45. Zurita J, Ortega-Paredes D, Barba P. First description of Shigella sonnei Harboring bla(CTX-M-55) outside Asia. J Microbiol Biotechnol. (2016) 26:2224–7. doi: 10.4014/jmb.1605.05069
46. Bonnet R. Growing group of extended-spectrum B-lactamases: the CTX-M enzymes. Antimicrob Agents Chemother. (2004) 48:1–14. doi: 10.1128/AAC.48.1.1
47. Gharaibeh MH, Shatnawi SQ. An overview of colistin resistance, mobilized colistin resistance genes dissemination, global responses, and the alternatives to colistin: a review. Vet World. (2019) 12:1735–46. doi: 10.14202/vetworld.2019.1735-1746
48. Cantón R, Ruiz-Garbajosa P. Co-resistance: an opportunity for the bacteria and resistance genes. Curr Opin Pharmacol. (2011) 11:477– 85. doi: 10.1016/j.coph.2011.07.007
49. Ortega-Paredes D. High quantities of multidrug-resistant Escherichia coli are present in the Machángara urban river in Quito, Ecuador. J Water Health. (2020) 18:67–76. doi: 10.1348/000712610X493494
50. Lazarus B, Paterson DL, Mollinger JL RB. Do human extraintestinal Escherichia coli infections resistant to expanded-spectrum cephalosporins originate from food-producing animals? A systematic review. Clin Infect Dis. (2014) 1:439–52. doi: 10.1093/cid/ciu785
51. Saliu E-M, Vahjen W, Zentek J. Types and prevalence of extended– spectrum beta–lactamase producing Enterobacteriaceae in poultry. Anim Heal Res Rev. (2017) 18:46–57. doi: 10.1017/S14662523170 00020
52. Xiang R, Liu B, Zhang A, Lei C, Ye X, Yang Y, et al. Colocation of the polymyxin resistance gene mcr-1 and a variant of mcr-3 on a plasmid in an Escherichia coli isolate from a chicken farm. Antimicrob Agents Chemother. (2018) 62:e00501-18. doi: 10.1128/AAC.00 501-18