2019年至2020年碳青霉烯类耐药大肠埃希菌的分子流行病学研究

Molecular epidemiology of carbapenem resistant Escherichia coli isolates from 2019-2020

目的对碳青霉烯类耐药大肠埃希菌的分子流行病学进行调查。方法通过微量肉汤稀释法测定中国细菌耐药监测研究(CARST)于2019年至2020年收集的大肠埃希菌的最低抑菌浓度;通过改良碳青霉烯灭活试验(mCIM)、EDTA改良碳青霉烯灭活试验(eCIM)检测菌株的碳青霉烯酶表型;通过聚合酶链反应和Sanger测序检测碳青霉烯酶相关耐药基因;通过接合实验验证碳青霉烯酶耐药基因所在质粒的水平传递性;通过多位点序列分型对碳青霉烯类耐药菌株进行同源性分析。结果36株对美罗培南或亚胺培南或厄他培南不敏感的菌株中22株只对厄他培南耐药。只对厄他培南耐药的22株大肠埃希菌碳青霉烯酶检测均为阴性,对美罗培南或亚胺培南耐药的14株大肠埃希菌碳青霉烯酶检测均为阳性,分别有8株携带bla_(NDM-5),3株携带bla_(NDM-1),1株携带bla_(NDM-7),1株携带bla_(NDM-16),1株碳青霉烯酶基因未测得。7株bla_(NDM-5)阳性菌株、3株bla_(NDM-1)阳性菌株、1株bla_(NDM-7)阳性菌株和1株bla_(NDM-16)阳性菌株可以通过接合实验使大肠埃希菌J53获得碳青霉烯抗性。14株碳青霉烯酶所致耐药的大肠埃希菌分属9个ST型,部分有关联。结论临床上只对厄他培南耐药但对亚胺培南和美罗培南敏感菌株的耐药性主要为非产碳青霉烯酶所致。而对亚胺培南或美罗培南耐药的14株菌mCIM和eCIM同时为阳性,提示临床分离株对亚胺培南或美罗培南耐药时,耐药机制较可能为产金属碳青霉烯酶。

Objective To investigate the molecular epidemiology of carbapenem resistant Escherichia coli isolates.Methods Escherichia coli isolates were collected by China Antimicrobial Resistance Surveillance Trial(CARST)from 2019-2020.Antibacterial susceptibility test was performed by microdilution broth method.Carbapenemase-producing Escherichia coli was detected by the modified carbapenem inactivation method(mCIM)and EDTA-carbapenem inactivation method(eCIM).Carbapenemase genes were detected by polymerase chain reaction(PCR)and Sanger sequence.Conjugation experiment was performed to verify the transferability of plasmids carrying carbapenemase genes.Homology analysis was performed by multilocus sequence typing(MLST).Result There were 36 Escherichia coli isolates resistant to any of the three carbapenem including imipenem,meropenem and ertapenem.14 carbapenemase producing strains were detected,in which,8 strains carried bla_(NDM-5),3 strains carried bla_(NDM-1),1 strain carried bla_(NDM-7),1 strain carried bla_(NDM-16),and 1 strain carried bla_(NDM-16).1 carbapenemase gene has not been detected and remains to be determined.12 carbapenemase gene positive strains succeed to transfer carbapenen resistance to Escherichia coli J53.14 strains of carbapenemase-producing Escherichia coli belonged to 9 ST types,with some correlation.Conclusion Clinical strains resistant to ertapenem but sensitive to imipenem and meropenem were mainly non-carbapenemase producing.14 strains were positive for mCIM and eCIM and were resistant to imipenem or meropenem,suggesting that when clinical isolates are resistant to imipenem or meropenem,the mechanism of resistance is more likely to be Metallo-carbapenemase production.