耐碳青霉烯铜绿假单胞菌金属β-内酰胺酶和外膜蛋白机制研究

Study on metallo-β-lactamase and outer membrane protein OprD2 of carbapenem-resistant pseudomonas aeruginosa

目的探讨铜绿假单胞菌金属酶肛内酰胺酶和外膜蛋白OprD2表达与碳青霉烯类抗生素耐药之间的关系。方法采用微量肉汤稀释法测定多利培南、美罗培南、亚胺培南对铜绿假单胞菌的最低抑菌浓度；应用亚胺培南-乙二胺四乙酸（EDTA）双纸片增效法对耐碳青霉烯菌株进行金属酶表型检测；采用聚合酶链反应（PCR）检测耐碳青霉烯菌株的blaIMP和blaVIM型金属酶基因和外膜孔道蛋白OprD2基因表达。结果应用亚胺培南-EDTA双纸片增效试验检测30株耐碳青霉烯铜绿假单胞菌金属酶表型，结果均为阴性，PCR未扩增出blaIMP、blaVIM型基因。25株耐碳青霉烯铜绿假单胞菌OprD2基因表达缺失。25株表达缺失菌株全部对亚胺培南耐药，仍有7株对美罗培南、13株对多利培南敏感。结论产金属酶不是我院铜绿假单胞菌对碳青霉烯类抗生素耐药的主要机制，外膜蛋白OprD2表达缺失是铜绿假单胞菌对亚胺培南耐药的主要机制，对美罗培南和多利培南的耐药还需要其他机制共同作用。

Objective To study the relation between metallo-β-1actamase and outer membrane protein OprD2 with carbapenem-resistance in pseudomonas aeruginosa. Methods MIC of doripenem, meropenem, imipenem against pseudomonas aeruginosa were determined by broth dilution method. Metallo-β-enzymes phenotypic screening of the carbapenem-resistant strains was examined by IMP-EDTA combined disk test. Polymerase chain reaction （PCR） confirmed the blaVIM, blaIMP and OprD2 gene. Results The results of IMP-EDTA combined disk test showed that no strains was positive. There were no blaIMP and blaVIM-type MBLs detected in the 30 carbapenem-resistant pseudomonas aeruginosa by PCR. 25 carbapenem-resistant PA were determined as lack of OprD2 gene. All of the 25 PA loss of OprD2 were resistant to imipenem,but 7 PA loss of OprD2 were sensitive to meropenem and 13 were sensitive to doripenem. Conclusions MBLs were not the main mechanism of carbapenem resistance for pseudomonas aeruginosa in our hospital. Loss of OprD2 was the main mechanism of imipenem-resistant pseudomonas aeruginosa, but there were other mechanisms in doripenem, meropenem-resistant pseudomonas aeruginosa.