奶牛乳房炎大肠杆菌整合子与耐药表型的相关性研究

Correlation between Antimicrobial Resistance and Integrons of Escherichia coli Strains Isolated from Bovine Mastitis Milk Samples

为研究自乳房炎奶牛分离的大肠杆菌整合子携带情况与其耐药表型之间的相关性,在获得菌株对20种抗菌药物耐药谱的基础上,设计Ⅰ、Ⅱ、Ⅲ类整合子简并引物扩增菌株的整合酶基因,并结合内切酶限制性片段长度多态性分析,调查菌株携带的各类整合子,然后进一步扩增菌株整合子的基因盒插入区,测序分析其携带的基因盒。结果发现,80株自奶牛乳房炎病例分离的大肠杆菌中有37株检出Ⅰ类整合子(检出率46.25%),未检出Ⅱ类和Ⅲ类整合子。Ⅰ类整合子阳性菌中31株扩增出3种不同大小的基因盒插入区片段,插入基因盒主要是二氢叶酸还原酶基因(dfr)和氨基糖苷类抗生素耐药基因(aadA),最主要的基因盒排列为dfrA17-aadA5。分析表明,Ⅰ类整合子集中分布在耐受7种以上抗菌药物的菌株中,与菌株的多重耐药性高度相关,而菌株Ⅰ类整合子携带的耐药基因盒与其耐药谱缺乏对应关系。

This study investigated the correlation between antimicrobial resistance of Escherichia coli strains isolated from bovine mastitis milk samples and the integrons. After acquirement of the antimicrobial resistance patterns, integrase gene was amplified by PCR using degenerate primers. The integrons were confirmed by restriction fragment length polymorphism （RFLP） analysis of positive PCR products, then integrated gene cassettes were examined. The incidence of class I integron in the 80 strains of E. coli was 46.25% （37/80）. All integrons were revealed as class I integron. Neither class Ⅱ nor class Ⅲ integron was detected. Thirty-one strains carried the integrated gene cassttes were detected in thirty-seven integron-positive srtains. The class Ⅰ integron was prevalent in strains which were resistant to more than seven antimicrobials. Class Ⅰ integrons harbored dfr and aadA gene cassettes confered resistance to trimethoprim and streptomycin, respectively. The order of main inserted gene cassettes was dfrA17-aadA5. There was a good correlation between integron carried and the multi-resistance, but the antimicrobial resistance patterns did not consist with the gene cassettes.