我国28家医院中铜绿假单胞菌对碳青霉烯类抗生素的耐药性及同源性分析

Resistance of the carbapenems and homology analysis of Pseudomonas aeruginosa in 28 hospitals in China

目的调查中国16个城市28家医院铜绿假单胞菌碳青霉烯类抗生素耐药情况、产金属酶情况及同源性。方法收集中国16个城市28家医院2006年7月到2007年7月临床分离的我国医院内病原菌耐药监测（NPRS）所保留的铜绿假单胞菌菌株654株。采用琼脂稀释法测定亚胺培南、美罗培南的MIC，脉冲场凝胶电泳（PFGE）分析碳青霉烯类耐药菌株同源性；Etest法进行金属酶表型筛选；PCR扩增及克隆测序分析金属酶基因型，实时荧光定量逆转录PCR（realtime RT-PCR）检测金属酶的转录水平。结果共筛选出275株碳青霉烯类抗生素不敏感株，其中259株为耐药株，亚胺培南和美罗培南的耐药率分别为37．5％（245／654）和28．0％（183／654）。275株碳青霉烯类抗生素不敏感菌株中金属酶表型阳性率6．55％（18／275），金属酶基因型阳性率8．73％（24／275）。6株金属酶基因型阳性而表型阴性菌株的金属酶相对表达量显著低于18株金属酶表型阳性菌株（Z=-2．335，P〈0．05）。259株碳青霉烯类抗生素耐药菌株PFGE分属89个型，24株金属酶基因型阳性菌株来自6个城市，分为9个PFGE克隆型。结论我国铜绿假单胞菌对碳青霉烯类抗生素耐药率高；未发现铜绿假单胞菌大范围单克隆株流行。产金属酶不是铜绿假单胞菌对碳青霉烯类抗生素耐药的主要机制。

Objective To investigate the resistance of carbapenems, the production of metallo- beta-lactamases（MBLs） and homology analysis of Pseudomonas aeruginosa in China. Methods A total of 654 Pseudomonas aeruginosa isolates were collected from 28 hospitals in 16 different regions during the period from July 2006 to July 2007 as part of the Nosocomial Pathogen Resistance Surveillance （NPRS）. MICs of imipenem and meropenem were determined by agar dilution. PFGE was used to analyze the molecular epidemiology of carbapenem-resistant strains. MBLs were detected by Etest and PCR among carbapenem-non-sensitive strains. The transcription levels of MBLs were determined by real-time reverse transcriptase （ RT ）-PCR. Results Of the 645 Pseudomonas aeruginosa strains, 245 （ 37.5% ） were resistant to imipenem and 183 （28.0%） were resistant to meropenem. A total of 275 carbapenem-nonsusceptible strains and 259 carbapenem-resistant strains were chosen for further study. Among the carbapenem-nonsusceptible strains, 8.73% （24/275） of the strains carrried MBLs genes by PCR, but only 6. 55% （18/275） of the strains were detected as MBLs-producers by MBL Etest. The MBLs relative expression levels of the 6 MBL genotype-positive but phenotype-negative strains were significantly lower than those of the other 18 MBL genotype- and phenotype-positive strains （P 〈 0. 05 ）. The 259 carbapenem-resistant Pseudomonas aeruginosa strains were divided into 89 clones by PFGE. The 24 MBL genotype-positive strains collected from 6 cities were divided into 9 clones by PFGE. Conclusions The carbapenems resistance of Pseudomonas aeruginosa is severe in China. Clonal dissemination of Pseudomonas aeruginosa has not been detected throughout China. MBLs are not the major carbapenems resistance mechanism in Pseudomonas aeruginosa.