养殖环境中耐氟喹诺酮类药物大肠杆菌gyrA基因的检测与序列分析  

Detection and Analysis of gyrAin Fluoroquinolone-Resistant Escherichia coli Isolates Collected from Different Breeding Environments

作　　者：曹立亭[1] 周晏丞[1] 仲崇华 陈鑫[1] 马跃[1] 

机构地区：[1]西南大学(荣昌校区)动物医学系,重庆荣昌402460 [2]农标普瑞纳(重庆)饲料有限公司,重庆荣昌402460

出　　处：《西南大学学报（自然科学版）》2014年第9期9-14,共6页Journal of Southwest University(Natural Science Edition)

基　　金：西南大学博士基金资助项目(09BSR04);西南大学本科毕业论文基金资助项目

摘　　要：该文探讨从养殖场动物、环境、饲养员等分离的大肠杆菌耐氟喹诺酮类药物的靶位基因gyrA突变特征.对养殖环境中分离的8株大肠杆菌进行临床常用氟喹诺酮类药物的耐药性分析、PCR扩增gyrA基因并测序,并采用DNASTAR软件对gyrA基因的氟喹诺酮耐药决定区进行生物信息学分析.结果表明:养殖场环境中分离的菌株编号为1,6和8的3株大肠杆菌对环丙沙星、左氧氟沙星、诺氟沙星敏感,菌株2,3,4,5和7的5株大肠杆菌对环丙沙星、左氧氟沙星、诺氟沙星耐药;氨基酸序列分析发现,敏感菌株氨基酸位点未发生突变或83位发生单突变;耐药菌株的突变发生在83位Ser→Leu替代和87位Asp→Asn替代,且发生氨基酸双替代的菌株均为高度耐药菌株.表明不同来源的耐氟喹诺酮类药物的大肠杆菌gyrA基因的氨基酸变异具有多种类型,该研究将更好地诠释养殖场大肠杆菌的氟喹诺酮类耐药机制.The mutations of the fluoroquinolone-resistant gyrA gene in 8 Escherichia coli isolates collected from human feces ,animal feces and the environment were investigated .The resistant phenotypes of 8 E.coli isolates to commonly used fluoroquinolones were determined by the K-B method with the standard strain ATCC25922 .Chromosome DNAs from all isolates of E. coli were extracted .The mutations in the fluoroquinolone resistance-determining regions of the gyrA gene were analyzed by PCR and sequencing . The results indicated that isolates of strain 1 ,strain 6 and strain 8 were sensitive to all the selected fluoroquinolones （ciprofloxacin ,Lomefloxacin and norfloxacin ） ,strain 2 ,strain 3 ,strain 4 ,strain 5 and strain 7 were multidrug-resistant isolates .Sequence analysis showed 2 mutations in the amino acids coded by fluoroquinolone resistance-determining regions in gyrA gene of all multidrug resistant isolates ,which resulted in 83Ser→Leu and 87Asp→ Asn .The strains in which both amino acid mutational sites existed were highly resistant .For the sensitive isolates ,there were no amino acid mutational sites or was only one amino acid mutational site in 83 Ser→Leu .A variety of amino acid changes in gyrA gene were detected in fluoroquinolone-resistant E. coli isolates isolated from different sources .These mutations may give a satisfactory explanation of the gene mutation mechanism of resistance to fluoroquinolones in E. coli .

关 键 词：大肠杆菌 氟喹诺酮类药物 耐药性 GYRA基因 

分 类 号：S853[农业科学—临床兽医学]