MexAB-OprM主动外排系统在全耐药铜绿假单胞菌耐药中的作用及机制  

The Contribution and Mechanism of MexAB-OprM Active Efflux System in Pan-resistant Pseudomonas Aeruginosa

作　　者：邓芳[1,2] 徐元宏[3] 沈继龙[1] 

机构地区：[1]安徽医科大学病原生物学教研室,合肥230001 [2]亳州市人民医院检验科 [3]安徽医科大学第一附属医院检验科

出　　处：《临床输血与检验》2010年第1期6-10,共5页Journal of Clinical Transfusion and Laboratory Medicine

基　　金：安徽省自然科学基金(No.09020103002)资助

摘　　要：目的探讨MexAB-OprM主动外排系统(外排泵)在全耐药铜绿假单胞菌耐药机制中的作用及其过度表达的机制。方法联合L-酚丙氨基-L-氨基酰-β-萘胺(MC-207,110)和环丙沙星(CIP),测定26株全耐药铜绿假单胞菌的MIC;用RT-PCR法扩增MexAB-OprM内膜转运蛋白MexB的结构基因mexB和内参照基因16SrDNA片段,根据mexB与16SrDNA扩增产物的电泳扫描比值,分析mexBmRNA的表达水平,判断MexAB-OprM的表达情况;用PCR法扩增MexAB-OprM调控基因mexR并对其产物测序,用Blast软件在GenBank中与已知序列比对。结果联合MC-207,110后有20株(76.9%)CIP的MIC值下降4倍及以上;26株全耐药铜绿假单胞菌中有22株MexAB-OprM高表达,其中有15株mexR基因发生变异,13株发生点突变,出现氨基酸替换,2株发生插入突变,导致氨基酸插入;点突变位点是336G-A、603G-A、660G-A、584C-G、653T-A,后两者导致氨基酸替换103Ala-Gly、126Val-Glu,插入突变是在470和474核苷酸间插入碱基GGC,导致66和68氨基酸间插入Ala。结论MexAB-OprM主动外排系统的高表达对铜绿假单胞菌的耐药性起重要作用,调控基因mexR的突变是其高表达的原因之一。Objective To investigate the effect of MexAB-OprM active efflux system in pan-resistant Pseudomonas aeruginosa （PRPA） and mechanisms of overexpression of MexAB-OprM. Methods To observe the inhibition of efflux pump inhibitors （EPIs） MC-207,110. The levels of mexB mRNA of wild strain （PA01） and 26 strains of PRPA were determined by RT-PCR,the 16SrRNA gene was also amplified as control,the levels of relative mexB mRNA were analyzed according to the ratio of electrophoresls scan results of their PCR products and the 16SrDNA＇s. mexR was amplified with PCR and PCR products were analyzed with DNA sequencing. Results The excretion of CIP can be inhibited by MC-207, 110 in 20 strains （76.9%） of PRPA. The levels of mexB mRNA in 22 strains of PRPA were significantly higher than that of PA01. Among 22 strains overexpressing MexAB-OprM, 15 carried mutations in mexR. The point mutations were 336G-A, 603G-A, 660G-A, 584C-G, 653T-A,the two latters resulted in the substitution of amino acids, which were 103Ala-Gly, 126Val-Glu respectively. The insertion mutation （insertion of GGC between 470 and 474） resulted in insertion of Gly between 66 and 68. Conclusions The overexpression of MexAB-OprM active efflux system are associated with resistance of pseudomonas aeruginosa. The mutations of mexR gene may be one of the reasons for high level expression of MexAB-O-prM.

关 键 词：铜绿假单胞菌 主动外排 MexAB—OprM RT—PCR 

分 类 号：R378.99[医药卫生—病原生物学] R392.11[医药卫生—基础医学]