产新德里金属β-内酰胺酶大肠埃希菌耐碳青霉烯类药物相关机制研究和分子分型

Carbapenem-resistance mechanisms and molecular typing of Escherichia coli producing New Delhi metalβ-lactamase

目的研究某三甲医院临床分离的产新德里金属β-内酰胺酶(NDM)大肠埃希菌的耐药机制和分子流行病学特征。方法在某三甲医院收集的耐碳青霉烯类大肠埃希菌中利用聚合酶链反应(PCR)筛查是否携带NDM基因,通过测序确定NDM型别并进一步检测其它碳青霉烯酶相关基因和广谱β-内酰胺酶类耐药基因。通过载体构建和表达研究NDM-1、NDM-6及NDM-9对亚胺培南的水解能力。采用多位点序列分型(MLST)对产NDM大肠埃希菌进行分子分型。结果共筛选出8株携带NDM大肠埃希菌,其中2株为NDM-9型,6株为NDM-6型。4株大肠埃希菌同时携带NDM-6、TEM和CTX-M-1基因,1株同时携带NDM-6、TEM和CTX-M-9基因。携带NDM-1、NDM-6与NDM-9重组菌对亚胺培南均耐药,最小抑菌浓度(MIC)分别为32μg/mL、128μg/mL和64μg/mL。产NDM大肠埃希菌MLST分型结果为ST226、ST648和ST1284型各1株,ST101型5株,此5株均来源于神经内科。结论首次发现同时携带NDM-6、TEM和CTX-M-1基因和携带NDM-6、TEM和CTX-M-9基因的大肠埃希菌,NDM-6、NDM-9对亚胺培南的耐药性比NDM-1强,应引起重视。MLST分型结果提示某三甲医院可能存在耐碳青霉烯类大肠埃希菌的院内局部传播。

The aim of this study was to investigate the resistance mechanisms and molecular epidemiological features of Escherichia coli strains producing New Delhi metalβ-lactamase(NDM)collected in a third class A hospital.NDM genes were screened with PCR in carbapenem-resistant Escherichia coli strains.Types of NDM genes were identified by sequencing.Other carbapenem-resistance related genes and extended-spectrum-β-lactamase genes were detected by PCR for NDM gene positive strains.An expression system was used to evaluate the imipenem-hydrolysis ability of NDM-1,NDM-6 and NDM-9.Molecular typing of Escherichia coli producing NDM was performed with the multilocus sequence typing(MLST)method.Eight strains of Escherichia coli producing NDM were identified,among which six were NDM-6 strains,and the remainder were NDM-9 strains.Four strains of Escherichia coli were found to simultaneously carry NDM-6,TEM and CTX-M-1 genes,and one strain was found to simultaneously carry NDM-6,TEM and CTX-M-9 genes.Recombinant Escherichia coli strains expressing NDM-1,NDM-6 and NDM-9 genes were resistant to imipenem,with minimum inhibitory concentrations of 32μg/mL,128μg/mL and 64μg/mL,respectively.MLST revealed four distinct sequence types(STs),among which five strains of ST101 were collected from the neurological unit,and the remaining three strains were ST226,ST648 and ST1284,respectively.This study reports the first identification of Escherichia coli simultaneously carrying NDM-6,TEM and CTX-M-1 genes,or NDM-6,TEM and CTX-M-9 genes.Notably,NDM-6 was associated with the highest imipenem hydrolysis ability,and was followed by NDM-9 and NDM-1.The MLST results indicated that a nosocomial infection of carbapenem-resistant Escherichia coli occurred locally in a third class A hospital.