大肠埃希菌和肺炎克雷伯菌外排泵转录水平与耐药的关系

Relationship between the level of efflux pump in Escherichia coli and Klebsiella pneumonia and drug resistance

目的调查细菌耐药性与其主动外排泵系统的作用关系,并探讨大肠埃希菌外排泵调控基因对其外排泵结构基因表达水平的影响,分析其耐药机制。方法收集大肠埃希菌和肺炎克雷伯菌共59株,根据头孢他啶耐药情况分为实验组(耐药)与对照组(敏感),多重聚合酶链反应(PCR)扩增外排泵调控基因acrR并测序,实时定量逆转录(RT)-PCR检测外排泵及调控基因marA的相对转表达水平,三维试验检测AmpC酶。结果实验组大肠埃希菌46.7%(7株)acrAB基因转录水平增高,肺炎克雷伯菌中51.9%(14株)acrAkp基因转录水平增高,敏感组外排泵基因均未见增高。实验组大肠埃希菌中,3株acrR发现突变,9株marA基因转录水平增高。59株菌株中39株(66.1%)AmpC阳性。结论主动外排泵系统是细菌耐药机制之一,常与其他耐药机制共同存在。acrR突变与marA表达增高均可引起大肠埃希菌acrAB表达增加。

Objective To investigate the relationship between bacterial resistance and its active role of efflux pump systems,explore the influence between the regulatory genes of efflux pump in Escherichia coli and the expression level of its structure gene,and analyze the resistant mechanism.Methods 59 strains of Escherichia coli and Klebsiella pneumonia were collected,and the strains were classified into two groups as experimental group and control group according to the resistance of ceftazidime.The multiplex polymerase chain reaction（PCR） was used for the amplification and seguencing of the regulatory gene acrR,and the real time（RT）-PCR was used for the detection of the expression level of efflux pump and the regulatory gene marA.The three-dimension test was used for the detection of AmpC beta-lactamase.Results 7 strains of Escherichia coli（46.7%） had high acrAB transcription level in experimental group,and 14 strains of Klebsiella pneumonia（51.9%）had high acrAkp transcription level in experimental group.Among the Escherichia coli in experimental group,3 isolates had mutation of acrR gene,and 9 isolates had high marA gene transcription level.AmpC beta-lactamase was detected in 39 of 59（66.1%） isolates.Conclusions The efflux pump system is one of the mechanisms of bacterial resistance,which often enhances the extent to resistance with other resistance mechanisms.The acrR mutation and increased expression of marA of Escherichia coli can cause the increasing of acrAB expression.