三株产CTX-M型超广谱β-内酰胺酶大肠埃希菌的研究  

Study on three strains of Escherichia coli producing extended-spectrum β-lactamases of CTX-M type

作　　者：殷俊[1] 程君[1] 孙震[1] 张晓妮[1] 高帆[1] 朱玉林[1] 叶英[1] 李家斌[1] 

机构地区：[1]安徽医科大学第一附属医院感染病科,合肥市230022

出　　处：《中华实验和临床感染病杂志（电子版）》2010年第2期1-5,共5页Chinese Journal of Experimental and Clinical Infectious Diseases(Electronic Edition)

基　　金：国家自然科学基金项目(No.30972631)

摘　　要：目的探讨3株产CTX-M型超广谱β-内酰胺酶(extended-spectrum β-lactamases,ESBLs)的大肠埃希菌分子生物学特征的改变及其与该突变位点的关系。方法 3株产CTX-M型ESBLs的大肠埃希菌于2005年收集自安徽省内3家医院。克隆这3株细菌的突变基因后进行序列分析,测定野生株、转移接合子与克隆子的最低抑菌浓度,提取突变基因所编码的蛋白质,进行等电聚焦电泳和酶动力学检测,并使用脉冲场凝胶电泳分析这3株细菌的同源性。结果 PCR和序列分析结果显示,与CTX-M-14相比,这3株大肠埃希菌的编码产物各有2处氨基酸突变,1株为第77位丙氨酸(Ala)突变为缬氨酸(Val)、第167位脯氨酸(Pro)突变为亮氨酸(Leu)(等电点为9.1);另2株均为第77位Ala突变为Val、第272位丝氨酸(Ser)突变为精氨酸(Arg)(等电点为9.3)。最低抑菌浓度和酶动力学检测结果表明,这两种酶均可高度水解头孢噻肟。脉冲场电泳结果提示这3株细菌无同源性。结论本研究发现了2种新的分离自大肠埃希菌的CTX-M酶,探讨突变位点与其生物学性状改变的关系,为遏制耐药基因及研究新的抗菌药物作用靶点提供依据,具体机制需进一步深入研究。Objective To describe the molecular characteristics of two novel extended-spectrum β-lactamases（ESBLs）of CTX-M type.Methods Three strains of Escherichia coli producing ESBLs of CTX-M type were collected from three hospitals of Anhui Province in 2005.Minimal inhibitory concentrations were detected by agar dilution method.Molecular characterization was carried out through polymerase chain reaction,single-strand conformation polymorphism,automated sequencing,clone and kinetic parameters and isoelectric focusing.Genetic relatedness was determined by pulsed-field gel electrophoresis.Results PCR and DNA sequencing revealed that two novel ESBLs,one with deduced amino acid sequence that different from CTX-M-14 by the substitutions Ala77→Val and Pro167→Leu（with pI 9.1）;the other by the substitutions Ala77→Val and Ser 272→Arg（with pI 9.3）.Both had potent hydrolytic activity against cefotaxime.Enzymatic studies revealed that the two novel ESBLs had no homology.Conclusions The study found two novel ESBLs of CTX-M type from Escherichia coli,exploring the mutation and the biological characteristics in order to curb the spread of resistance genes and provide new antibacterial drug targets,and revealing the specific mechanisms need further study.

关 键 词：大肠埃希菌 超广谱Β-内酰胺酶 耐药性 

分 类 号：R446.5[医药卫生—诊断学]