临床分离耐喹诺酮阴沟肠杆菌gyrA基因突变的研究

目的：检测临床分离耐喹诺酮阴沟肠杆菌DNA促旋酶gyrA基因突变情况。方法：收集临床分离耐喹诺酮阴沟肠杆菌30株及敏感株10株，测定其对萘啶酸、环丙沙星、氧氟沙星的最低抑菌浓度(MIC)；用聚合酶链反应(PCR)扩增gyrA基因部分片断，对PCR产物进行限制性片断长度多态性(PCR—RFLP)及单链构象多态性(PCR—SSCP)分析，检测gyrA突变情况。结果：30株耐喹诺酮菌株都检测到gyrA基因突变，PCR—RFLP均显示两条不同于敏感株的电泳带，其PCR—SSCP则检测到4种不同于敏感菌的迁徙率条带；而10株敏感菌均未检测到gyrA基因突变。结论：阴沟肠杆菌对喹诺酮类药物耐药性与gyrA基因突变有关，gyrA基因第83位氨基酸密码子突变可能为其耐药的主要原因。

Study on the mutation of gyrA gene in quinolone-resistant clinical isolates of Enterobacter cloacae

Objective: To study the mutation of gyrA gene in quinolone-resistant clinical isolates of Enterobacter cloacae.Methods:Using age the micro broth diluent to determine the MICs of three quinolones of 30 quinolone-resistant and 10 quinolone-susceptible isolates,the gyrA gene in these isolates was amplified with polymerase chain reaction(PCR),then by restrictive fragments length polymorphism(RFLP) and single strand conformation polymorphism(SSCP) to detect the mutation of gyrA gene.Results:All 30 quinolone-resistant isolates were detected the mutation of gyrA gene,the results of PCR-RFLP of them revealed two stripes while the RFLP-SSCP revealed four kinds of stripe different from 10 quinolone-susceptible isolates.No mutation in gyrA gene of 10 quinolone-susceptible isolates and ATCC13047 was detected by PCR-RFLP and PCR-SSCP.Conclusions:Mutation of gyrA gene in clinical isolates of Enterobacter cloacae implicated in resistance to quinolones,the mutation of DNA gyrase at the 83rd amino acid may be play a principle role in the resistant to quinolones of Enterobacter cloacae isolates.