美罗培南耐药铜绿假单胞菌多药外排泵研究

目的 研究美罗培南耐药铜绿假单胞菌的多药外排泵机制.方法 采用琼脂稀释法测定美罗培南对141株铜绿假单胞菌的最低抑菌浓度(MIC),同时观察加入泵抑制剂MC207110后的美罗培南对141株铜绿假单胞菌MIC变化,检测多药外排泵机制.实时定量PCR检测外排泵mRNA水平的表达.PCR扩增外排泵编码基因及调控基因并测序分...

Study on the multi-drug efflux pump of carbapenem-resistant Pseudomonas aerug- inosa

Objective To study the multi-drug efflux pump of meropenem-resistant Pseudornonas aeruginosa. Methods The minimum inhibitory concentrations （MICs） of meropenem against 141 strains of P. aeruginosa were determined by agar dilution method with or without the presence of efflux pump inhibitor MC207110 （20 mg/L）. The expression level of efflux pumps mRNA were analyzed by real-time PCR. The coding genes and regulatory genes of efflux pumps were amplified and analyzed by PCR and DNA sequencing. Results The results of efflux pump phenotype testing showed that MIC values of meropenem de- creased to 1/4 to 1/512 for 69. 1% of the strains tested. The mRNA expression level of 4 efflux pump genes increased in 20 strains of meropenem-resistant P. aeruginosa, primarily MexEF-OprN （10 strains）, followed by MexCD-oprJ （5）, MexAB- OprM （4）, and MexXY-OprM （3）. The mRNA expression of both MexA and MexC genes increased in 2 of these strains. Sequencing results of the 4 efflux pump genes showed that MexAB-OprM overexpression was due to MexR mutation in 3 strains, and nalC mutation in 1 strain, nMD mutation was not identified. All the 5 strains with MexCD-OprJ overexpression showed nfxB mutation. All the 10 strains with MexEF-OprN overexpression had mexT mutation. All the 3 strains with MexXY-OprM overexpression showed mexZ gene mutation. The coding genes mexB, mexD, mexF, mexY of efflux pump did not mutate in the 20 strains. Conclusions Efflux pump may be one of the main mechanisms of P. aeruginosa resistance to meropenem. The overexpression of the efflux pump system is primarily due to the mutation of corresponding regulatory genes