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目的 研究铜绿假单胞菌遗传性耐药(gyrA基因突变和MexAB OprM主动外排)在生物被膜形成过程中对药物抗性的作用。方法 通过PCR技术鉴定gyrA基因突变株和MexAB OprM主动外排株,利用三亲杂交的方法将绿色荧光蛋白(GFP)基因转到铜绿假单胞菌药物敏感株中,测定gyrA基因突变株、MexAB OprM主动外排株以及带有绿色荧光蛋白的铜绿假单胞菌在特氟隆上产生生物被膜耐药的规律。结果 筛选出gyrA基因突变株和MexAB OprM主动外排株。通过三亲杂交成功构建了带有pGFPuv质粒的铜绿假单胞菌。将突变株和敏感株培养在最低杀菌浓度(MBC)的环丙沙星溶液中形成生物被膜使菌株的存活率均在50%以上。结论 比较遗传性耐药和生物被膜耐药的作用表明gyrA基因突变和主动外排在生物被膜耐药形成前期发挥主要作用,当生物被膜完全形成之后,铜绿假单胞菌的耐药性有可能是由生物被膜介导的。
Objective To study the effects of genetic resistance to P. aeruginosa (gyrA gene mutation and MexAB OprM active efflux) on drug resistance in biofilm formation. Methods The gyrA gene mutant and MexAB OprM active efflux were identified by PCR. The green fluorescent protein (GFP) gene was transferred to the Pseudomonas aeruginosa drug susceptible strain by triparental hybridization. The gyrA gene mutant, MexAB OprM active efflux plants and green fluorescent protein-containing Pseudomonas aeruginosa produce biofilm resistance on Teflon. Results The gyrA gene mutant and MexAB OprM active efflux were screened out. Pseudomonas aeruginosa with pGFPuv plasmid was successfully constructed by crossed parents. The mutant strains and sensitive strains were cultured in the minimum bactericidal concentration (MBC) of ciprofloxacin solution to form a biofilm so that the survival rate of the strains are above 50%. Conclusions The comparison of genetic resistance and drug resistance of biofilm indicates that gyrA gene mutation and active efflux play a major role in biopharmaceutical drug resistance formation. After the biofilm is completely formed, the drug resistance of Pseudomonas aeruginosa may be Is mediated by the biofilm.