泛耐药铜绿假单胞菌耐药机制研究

目的 研究铜绿假单胞菌(PA)泛耐药(PDR)机制.方法 琼脂稀释法测定14种抗菌药物对泛耐药铜绿假单胞菌(PDR-PA)的最低抑菌浓度(MIC).肠杆菌基因间重复一致序列-聚合酶链反应(ERIC-PCR)分析耐药菌株同源性.PCR扩增检测超广谱B内酰胺酶(ESBL)、碳青霉烯酶、质粒介导AmpC酶等分析B.内酰胺类的耐药机制;PCR扩增检测16种氨基糖苷钝化酶基因分析氨基糖苷类耐药机制;PCR扩增检测qnr基因、DNA旋转酶基因gyrA、gyrB和拓扑异构酶Ⅳ基因parC和parE,分析氟喹诺酮类耐药机制;PCR扩增oprD2编码基因及序列分析和MC207110检测外排泵分析膜屏障机制引起的碳青霉烯类耐药机制.结果 19株PDR-PA的ERIC-PCR分型结果显示具有5个型别,其中以A和B型为主,分别有6株和7株.17株检出VEB-3型ESBL基因,其中1株同时检出OXA-10型ESBL基因;未检测出质粒AmpC酶和碳青霉烯酶基因.19株均检出ant(3")I氨基糖苷钝化酶基因,其中18株同时捡出ant(3)Ⅱ酶基因;19株发生gyrA突变,其中14株同时发生了parC突变,未检测出qnr基因.19株oprD2编码基因均发生小片段缺失.16株显示具外排泵机制.结论 提示产生VEB-3型ESBL、aac(3)Ⅱ和ant(3")I氨基糖苷钝化酶、DNA旋转酶gyrA和拓扑异构酶ParC基因发生突变,以及OprD2蛋白缺失和外排泵高表达是本组PDR-PA泛耐药的重要原因.

Mechanisms of pandrug-resistance of Pseudomonas aerugionosa

Objective To explore the mechanisms of pandrug-resistance(PDR)of Pseudomonas aerugiorLosa(PA).Methods Nineteen strains of PA were coilected from Huashan Hospital,Shanghai.Agar dilution method was used to detect the levels of minimum inhibitory concentration(MIC)of 14 antimicmbial drugs to the PA strains.Strain homology was investigated by entembacterial repetitive intergenic consensus-polymerase chain reaction(ERIC-PCR).To analyze the B-lactam resistant mechanisms,genes of extended-spectnnn B-lactamases(ESBLs),carbapenemase,and plasmid-mediated AmpC were amplified and analyzed by PCR and DNA sequencing.To analyze the aminoglyeoside resistant mechanisms.16 aminoglycoside modifying enzyme were screened by PCR.PCR and DNA sequencing were used to amplify and analyze the genes of DNA gyrase genes gyrA and gyrB,topoisomerase Ⅳ genes parc and pare.and qnr gene for the fluoroquinolone resistance mechanisms,to amplify and analyze study the oprD2 coding genes and inhibitor MC207110 were used to detect efflux pump for the carbapenem resistance mechanisms.Results Five types were identified in the 19 PDR-PA by ERIC-PCR,mainly type A(n=6)and type B(n=7).17 of the 19 PDR-PA strains produced VEB-3 type ESBL,1 strain of which also preduced OXA-10 type ESBL simultaneously.Both of the carbapenemase and plasmid-mediated AmpC were negative.All of the 19 PDR-PA strains produced aminoglycoside modifying enzyme,yielding ant(3")I and 18 strains of which produced aac(3)Ⅱ simultaneously.All 19 PDR-PA strains carried gyrA mutations,14 of which carried parC mutation simultaneously.qnr gene w85 negative.OprD2 coding gene sequencing analysis revealed that small fragment missing occurred to all oprD2 genes of the 19 strains.16strains showed emux pump mechanism.Conclusion The resistance mechanism of PDR-PA to cephalespofins,B-lactam/B-Lactamsse inhibitor,carbapenem,fluoroquinolones,and aminoglycosides are due to production of VEB-3-ESBL,aac(3)H,and ant(3")I aminoglycoside modifying enzyme,mutations of DNA gyrase gyrA and topoisomerase parC gene,OprD2 protein deficiencies,and emux pump overexpression.