鸡源大肠杆菌强毒株耐药基因的定位及耐药质粒消除

摘要

To preliminarily locate the resistant gene with multi-resistance in pathogenic E. coli clinically isolated from chicken and provide reasonable evidence to therapeutic treatment. The E. coli strain was isolated from liver and heart of died chicken examined. The drug resistant plasmid, extracted with plasmid extraction kit, was purified after transferred to gene engineering strain JM109. The anti-drug gene was preliminarily located through electrophoresis and drug sensitivity examination. The weeping function of the extracted material from Artemsia argyi to the resistant plasmid was tested. The results indicated that the E.coli strain with strong virulence, isolated from chicken and identified successfully, waa multi-resistance to drugs, and only sensitive to fluorine spectinomycin and streptomycin. The preliminary location of anti-drugs gene, which resisted to cipro-floxacin, penicillin, ofloxacin, norfloxacin, lincomycin, and cotrimoxazole, was on the certain drug resistant plasmid. The characteristics of resistance to drug could be delivered to the gene engineering strain E. coli, in which the drug resistant plasmid extracted was transferred to. Boiled leaves solution can eliminate 60% the bacteria resistance plasmid. Susceptibility test results showed that the bacteria eliminated resistant plasmids restored sensitivity to ciprofloxacin, penicillin, ofloxacin, norfloxacin, lincomycin and cotrimoxazole. Therefore, the anti-drug genes were located on plasmid and chromosome of the isolated strain. The extracted material from leaves had strong weeping function, and could be used as clinical therapeutic medicine.%本试验对临床分离的多重耐药鸡源致病性大肠杆菌强毒株的耐药基因进行初步定位,为临床选择合适的治疗策略提供理论依据.从送检病死鸡的肝脏、心脏中分离鉴定致病菌,质粒提取试剂盒提取分离菌的耐药质粒,转化入基因工程菌JM109,通过质粒纯化、电泳和药敏试验对耐药基因进行了初步定位.并用艾叶水煮液对该菌株进行体外耐药质粒消除试验.结果分离鉴定到1株强毒力鸡源大肠杆菌,该菌呈多重耐药性,且仅对氟奇霉素和链霉素敏感；由质粒转化和药敏试验结果可初步将耐环丙沙星、青霉素、氧氟沙星、氟哌酸、林可霉素和复方新诺明的基因定位于耐药质粒上,并可随质粒的转移而使转化菌获得耐药性；用艾叶水煮液可使该菌的耐药质粒消除率达60％；质粒消除菌的药敏试验结果表明,消除耐药质粒的细菌恢复了对环丙沙星、青霉素、氧氟沙星、氟哌酸、林可霉素和复方新诺明的敏感性.本研究结果表明,分离菌的耐药基因分别位于质粒和染色体上,艾叶对耐药质粒有较强的消除作用,可作为临床治疗用药.