大肠埃希氏菌和沙门菌氨基糖苷耐药基因PCR检测方法的建立  

Development of PCR method for detection of aminoglycoside resistance genes of Escherichia coli and Salmonella

作　　者：马孟根[1] 王红宁[2] 余勇[3] 李成忠[2] 张东[3] 羊云飞[2] 邢坤[3] 刘世贵[1] 

机构地区：[1]四川大学生命科学学院,四川成都610064 [2]四川农业大学动物科技学院,四川雅安625014 [3]四川省动物防疫监督总站,四川成都610041

出　　处：《中国兽医科技》2005年第4期277-283,共7页Chinese Journal of Veterinary Science and Technology

基　　金：国家"十五"重大科技攻关项目(2002BA514A 12 5)

摘　　要：采用平板稀释法,选用氨基糖苷类药物链霉素、庆大霉素、卡那霉素和新霉素对22 株猪、鸡大肠埃希氏菌和30株猪、鸡沙门菌进行了药敏试验。结果显示,在4种药物中大肠埃希氏菌对卡那霉素的耐药率最高,达81.8%,而沙门菌则对链霉素的耐药率最高,达60%,两类细菌都对新霉素的耐药率最低。设计了5对引物,对耐药基因aph(3′) Ⅱ、aadA1、aadA2、aadB和aac(3) Ⅰa进行了扩增,获得的基因片段与预期的大小一致;序列分析表明,扩增产物与GenBank中的相应序列有很高的同源性(>99.8%)。采用建立的PCR技术,对22株猪、鸡大肠埃希氏菌和30 株猪、鸡沙门菌的耐药基因进行了检测,证实耐药基因普遍存在,在大肠埃希氏菌中aadA1 基因的检出率最高(12/22),在沙门菌中aph(3′) Ⅱ基因的检出率最高(18/30),在所有的菌株中都未能检测出aac(3) Ⅰa基因。The streptomycin, gentamycin, kanamycin, and neomycin resistance indexes of 22 Escherichia coli (isolates) and 30 Salmonella isolates from swine and chickens were detected. The results showed that the kanamycin resistance rate of 22 E.coli isolates was 81.8%, the streptomycin resistance rate of 30 Salmonella isolates was 60.0%, and the neomycin resistance rates of 22 E.coli and 30 Salmonella isolates was the lowest among the 4 antibiotics. The antibiotic resistance genes aph(3′)-Ⅱ, aadA1, aadA2, aadB and aac(3)-Ⅰa were amplified with the 5 designed pairs of primers. The sequencing showed the 4 genes shared over 99.8% of homology with that from GenBank. The antibiotic resistance genes were detected in the 22 E.coli isolates and the 30 Salmonella isolates by the developed PCR. The results showed that the antibiotic resistance genes were commonly present in the above-mentioned isolates, and the frequency of aadA1 gene was the highest in the E.coli isolates and the frequency of aph(3′)-Ⅱ gene was the highest in the Salmonella isolates, whereas aac(3)-Ⅰa gene was not found in all the isolates.

关 键 词：大肠埃希氏菌 沙门菌 耐药基因 氨基糖苷类药物 

分 类 号：S852.612[农业科学—基础兽医学] Q503[农业科学—兽医学]