碳青霉烯类抗生素诱导耐药铜绿假单胞菌外膜蛋白D2表达与编码基因多点突变  

Expression of outer membrane protein D2 and its gene mutation of coding region in drug resistant Pseudomonas aeruginosa induced by carbopenem antibiotics

作　　者：叶晓光[1] 余名华[1] 

机构地区：[1]广州医学院第二附属医院感染科,510260

出　　处：《中华生物医学工程杂志》2008年第4期269-273,共5页Chinese Journal of Biomedical Engineering

摘　　要：目的探讨对碳青霉烯类抗生素耐药的铜绿假单胞菌编码外膜蛋白D2（OprD2）基因变异与OprD2表达的关系。方法由临床分离敏感铜绿假单胞菌，使用不同浓度亚胺培南与美罗培南含药琼脂平板法筛选耐药菌株。应用十二烷基磺酸钠-聚丙烯酰胺凝胶电泳（SDS—PAGE）、凝胶成像系统分析OprD2。应用聚合酶链反应及测序分析OprD2基因编码区。结果人工诱导能产生出与临床分离株一样的对碳青霉烯类抗生素耐药的菌株。在SDS—PAGE图谱上亚胺培南与美罗培南耐药株与同源敏感株相比均有OprD2的减少。测序结果显示在多株耐药菌中，OprD2基因编码区3个基因位点同时出现碱基突变：在＋84位点产生突变，胞嘧啶突变为胸腺嘧啶（C→T）；在＋401位点突变（C→A），造成第134位氨基酸苏氨酸突变为天冬氨酸（Th→Asp）；在＋959位点突变（A→G），造成第320位氨基酸赖氨酸突变为精氨酸（Lys→Arg）。结论同时出现3个或更多的碱基点突变可能导致铜绿假单胞菌OprD2缺失或表达减少，可能是对碳青霉烯类抗生素产生耐药性的原因。Objective To investigate the expression of outer membrane protein D2 （OprD2） and its gene mutation of coding region in drug resistant Pseudomonas aeruginosa （ PA ） induced by carbopenem antibiotics. Methods The resistant mutant strains were selected from the clinically isolated sensitive strains of PA by passage on Muller-Hinton agar containing different concentrations of meropenem or imipenem. OprD2 was analyzed by gel imaging system after SDS-PAGE. The coding region of OprD2 gene was detected by polymerase chain reaction and sequencing analysis. Results The carbopenems-resistant strains of PA by artificial induction were similar to those clinically isolated strains. The expression of OprD2 in carbopenemsresistant strains was significantly lower than that in the homologous sensitive strains in SDS-PAGE atlas. The nucleotide sequence showed three gene mutations of the OprD2 coding region in carbopenems-resistant strains of PA at the same time： at position ＋ 84 （C→T） ; at position ＋401 （C→A）, Tbr134→Asp; at position ＋ 959 （A→G）, Lys320→Arg. Conclusion The low expression of OprD2 is due to three or more gene mutations at the same time in the coding region of OprD2 gene in carbopenems-resistant strains of PA.

关 键 词：假单胞菌 铜绿 碳青霉烯类 外膜蛋白172 抗药性 突变 亚胺培南 美罗培南 

分 类 号：R446.5[医药卫生—诊断学]