一例碳青霉烯类抗菌药物耐药的肺炎克雷伯菌耐药机制研究

目的研究1株碳青霉烯类抗菌药物耐药的肺炎克雷伯菌A1500的耐药机制。方法采用浓度梯度法（E-test）测定细菌的抗菌药物最低抑菌浓度（MIC）,采用接合实验、聚合酶链反应（PCR）、质粒抽提、探针杂交印迹试验（Southern blot）、等电聚焦电泳及耐药基因克隆测序分析细菌的耐药机制。结果接合实验、质粒电泳和Southern blot结果显示耐药基因位于一个50 kb的可转移质粒上;等电聚焦电泳显示3条β-内酰胺酶条带,等电点（PI）分别为9.0、6.7（KPC-2）、5.4。PCR产物克隆分析比对为blaKPC-2型。结论碳青霉烯类抗菌药物耐药的肺炎克雷伯菌A1500携带由质粒介导的KPC-2酶。

Investigation of the resistant mechanism of a carbapenem-resistant Klebsiella pneumoniae

Objective To investigate the resistant mechanism of a carbapenem-resistant Klebsiella pneumoniae A1500. Methods The minimal inhibitory concentrations（ MIC ） of the antimicrobial agents were determined by E-test, conjugation, polymerase chain reaction（PCR） ,plasmid extraction,Southern blot,isoelectric focusing electrophoresis, cloning and sequencing were carried out for analyzing the resistant mechanism. Results Conjugation, plasmid electrophoresis and Southern blot demonstrated that the gene was encoded on an approximately 50 kb conjugative plasmid. Isoelectric focusing of the strain studies demonstrated three β-laetamases with isoionic point （PI）9.2,6.7 （KPC-2） and 5.4. The gene was blaKPC-2 through blasting. Conchlsions The strain of carbapenem-resistant Klebsiella pneumoniae A1500 produces a plasmid-mediated carbapenemase KPC-2.