耐碳青霉烯类大肠埃希菌临床分布、耐药特征及携带mcr 基因分析

目的　分析临床耐碳青霉烯类大肠埃希菌（carbapenem-resistance Escherichia coli , CREC）分布及耐药特征，检测其碳青霉烯酶合成基因及质粒介导的多黏菌素耐药（mobile colistin resistance, mcr）基因携带情况，为多重耐药菌防控提供参考。方法　收集2020 年7 月～ 2021 年6 月期间安徽医科大学第二附属医院住院患者送检的各类临床标本分离非重复CREC 菌株，微量肉汤稀释法进行药敏试验。聚合酶链反应（polymerase chain reaction, PCR）扩增及测序技术检测碳青霉烯酶基因及mcr 系列基因。多黏菌素诱导试验检测mcr-9 阳性菌株的诱导耐药特征，脉冲凝胶电泳（pulsedfieldgel electrophoresis, PFGE）分析其同源性。结果　共收集CREC 菌株33 株，主要分离自尿液标本（30.3%）、痰液标本（24.2%）、脓液/ 分泌物标本（18.2%）及血液标本（15.2%），菌株呈现多重耐药表型。PCR 检测结果显示，23 株CREC（69.7%）携带blaNDM 基因，8 株CREC（24.2%）携带blaKPC 基因。有3 株（9.1%）blaNDM 阳性CREC 菌株同时携带mcr-9 基因，PFGE 分析显示3 株细菌间无同源性。上述mcr-9 阳性菌株经过1/2 倍最低抑菌浓度（minimuminhibitory concentration, MIC）多黏菌素B 药物诱导6h 后，对多黏菌素B 的MIC 值均升高。结论　临床CREC 呈现多重耐药，主要碳青霉烯酶基因为blaNDM 及blaKPC。部分菌株同时携带blaNDM 及mcr-9 基因，且呈现多黏菌素诱导耐药特征。实验室应加强监测，防控其流行播散。

Clinical Distribution,Antimicrobial Agent Resistance and mcr Genes Analysis of Carbapenem-resistant Escherichia coli

Objective　To analyze the clinical distribution, antimicrobial agent resistance rates of carbapenem resistance Escherichia coli (CREC), and detect the carbapenemse synthesis gene and the carrier of plasmid mediated mobile colistin resistance (MCR) gene, so as to provide reference for the prevention and control of multi-drug resistant bacteria. Methods　Form July 2020 to June 2021, Non-repetitive CREC strains isolated from various clinical specimens submitted by inpatients in the Second Hospital of Anhui Medical University were collected. The antimicrobial susceptibility testing was detected by microbroth dilution method. Carbapenemase encoding and mcr genes were analyzed by PCR amplification and sequencing. The polymyxin B induction assays of mcr-9-positive-isolates were performed, and the homology of mcr-9-positive-isolates was analyzed by pulsed-field gel electrophoresis (PFGE). Results　A total of 33 CREC isolates were collected, which mainly isolated from urine (30.3%), sputum (24.2%), pus/secretion (18.2%) and blood (15.2%), presenting multiple drug resistance phenotypes. 69.7% of the isolates (n=23) harboured blaNDM, while 24.2% of them(n=8) harboured blaKPC. Three CREC isolates (9.1%) carried both blaNDM and mcr-9 genes, and PFGE analysis showed no homology among the three strains. After pretreatment with polymyxins B [1/2×minimum inhibitory concentration (MIC)], the MIC values of mcr-9-positive-strains to polymyxins B were increased. Conclusion　The clinical CREC isolates performed multiple drug resistance phenotype, mainly harboured blaNDM and blaKPC carbapenemase encoding genes. CREC isolates co-harboring blaNDM and mcr-9 were detected, and the polymyxin-induced drug resistance phenotype were detected in all this strains. Laboratory monitoring should be strengthened to prevent and control its spread.