三重PCR方法对猪、鸡源细菌中四环素类药物耐药基因的检测  

Detection of tetracycline resistant genes in bacterial by using a triple-PCR method

作　　者：夏青青[1] 王红宁[1] 张安云[1] 

机构地区：[1]四川大学生命科学学院动物疫病防控与食品安全四川省重点实验室,生物资源与生态环境教育部重点实验室,"985工程"西南资源环境与灾害防治科技创新平台,成都610064

出　　处：《四川大学学报（自然科学版）》2013年第1期171-176,共6页Journal of Sichuan University(Natural Science Edition)

基　　金：农业部蛋鸡现代产业技术体系项目(2009CYTX01);四川省科技支撑计划项目(2011NZ0073-1;10KJT-02;2012NZ0037)

摘　　要：针对GenBank上已公开的四环素类耐药基因tetA、tetC、tetM序列进行比对分析,设计特异性扩增引物,建立三重PCR法快速检测方法.结果表明,单重PCR和三重PCR的符合率为100%.应用本检测方法和传统药敏试验方法(药敏纸片法)对412猪、鸡源细菌的四环素耐药基因和表型同时进行检测.PCR检测实验结果显示,在412株待检细菌中,tetA、tetC、tetM的检出率分别为47.57%,38.35%和34.95%.药敏实验结果表明,待测细菌对四环素的耐药率最高,为95%;对多西环素的耐药率次之,为92%;对米诺环素耐药率相对最低,为84%.比较两种方法,发现其检出结果的符合率为88%.说明两种方法具有较高的一致性.本研究建立了一种猪、鸡源细菌四环素类药物耐药基因三重PCR检测方法,并进行了初步应用.PCR specific primers were designed based on sequences published on Genbank. A triple-PCR was developed based on conventional PCR and reaction parameter we optimized to develop a detection method. To characterize the accuracy of this method, conventional PCR and triple-PCR were used to test same samples. Results indicated that the coincidence rate of these two methods were 100%. Triple- PCR and disk diffusion method were both performed to detect the genotype and phenotype of 412 bacte- rial isolated from swine and chicken. The result showed that bacterial resistance rate to tetracycline was 95%, to doxycycline was 92%, and to minocycline was 84%. The detection rate of tetA, tetC and tetM were 47.57%, 38.35%, 34.95%, respectively. The coincidence rate of two methods was 88%, which suggested high consistency.

关 键 词：四环素耐药性 tetA TETC tetM 三重PCR 

分 类 号：S855[农业科学—临床兽医学]