产AmpC酶铜绿假单胞菌对亚胺培南耐药机制的研究  

Mechanism of imipenem resistance in AmpC β-lactamases-producing Pseudomonas aeruginosa

作　　者：李小燕[1] 吴爱武[1] 

机构地区：[1]广州医学院医学检验系,广州510182

出　　处：《中国感染与化疗杂志》2008年第6期423-428,共6页Chinese Journal of Infection and Chemotherapy

基　　金：广州市医药卫生科技立项资助项目(项目编号:2005-YB-134)

摘　　要：目的研究产AmpC酶的铜绿假单胞菌对亚胺培南耐药的分子机制。方法2003—2007年从临床标本中分离到铜绿假单胞菌220株,采用三维试验筛选产β内酰胺酶(包括AmpC酶)的铜绿假单胞菌,应用多重PCR检测产AmpC酶的铜绿假单胞菌质粒携带的AmpC耐药基因,应用荧光定量RT-PCR的方法检测染色体AmpC酶基因和oprD2基因的表达情况。结果共检出40株产β内酰胺酶的菌株,其中产AmpC酶、ESBLs、金属酶(MBL)和未知酶型铜绿假单胞菌的构成比分别是62.5%(25/40)、20%(8/40)、7.5%(3/40)和10%(4/40)。25株产AmpC酶菌株均有不同程度AmpC酶基因表达量升高,其中,仅1株细菌携带DHA质粒型AmpC酶基因,12株菌oprD2基因表达量减低,这些菌株均对亚胺培南耐药,13株菌oprD2基因表达量正常,其中仅5株菌对亚胺培南耐药;7株菌(占产AmpC酶菌株的28%,不产金属酶)的酶粗提物能微弱水解IPM,这7株菌AmpC酶基因的表达量均有明显升高,其中5株菌同时伴有oprD2基因表达量降低。结论铜绿假单胞菌产生的AmpC酶可微弱水解亚胺培南,且其水解亚胺培南可能与AmpC酶产生量有一定关系;产AmpC酶铜绿假单胞菌对亚胺培南耐药的原因可能是细菌AmpC酶表达量增加和oprD2表达量减低两者共同作用的结果。Objective To investigate the molecular mechanism of imipenem （IPM） resistance in AmpC β-lactamases-producing P. aeruginosa. Methods Two hundred and twenty strains of P. aeruginosa were isolated from inpatients between 2003 and 2007. Three-dimensional test was used to identify the P. aeruginosa producing β-lactamases （including AmpC β-lactamases）. PCR and Multiplex PCR were performed to detect oprD2 genes and plasmid-mediated AmpC β-lactamase genes, respectively. The expression of chromosomal AmpC β-lactamases and oprD2 genes in P. aeruginosa were analyzed by real-time reverse transcriptase PCR. Results Production of β-lactamases was found in 40 isolates. AmpC β-lactamases, extended spectrum β-lactamases （ESBLs）, metallo-β-lactamases （MBL） and an unknown type of β-lactamases were identified in 62.5% （25/40）, 20% （8/ 40）, 7.5% （3/40） and 10%（4/40） of these isolates, respectively. Of the 25 isolates expressing increased AmpC β-lactamases, the plasmid mediated AmpC β-lactamase gene DHA was identified in only 1 isolate. Diminished expression of oprD2 was found in 12 isolates. These strains were resistant to imipenem. The other 13 isolates had normal oprD2 expression. Five of these 13 strains were resistant to imipenem. The AmpC lactamase crude extract （not MBL） from 7 isolates could weakly hydrolyze imipenem. Five of these 7 isolates also showed lower expression of oprD2 gene. Conclusions Some AmpC β-lactamases produced by P. aeruginosa can weakly hydrolyze imipenem. This hydrolytic activity is dependent on the expression level of AmpC β-lactamases. The mechanism of imipenem resistance in P. aeruginosa is a result of the interplay between diminished production of oprD2 and increased activity of AmpC β-lactamase.

关 键 词：铜绿假单胞菌 AMPC酶 OprD2蛋白 荧光定量反转录聚合酶链反应 

分 类 号：R516[医药卫生—内科学]