猪库布病毒RT-PCR检测方法的建立及湖北省流行病学初步调查  

Porcine kobuvirus RT-PCR detection assay establishment and application in primary epidemiology investigation in Hubei Province

作　　者：胡军勇[1] 汤细彪[2] 胡睿铭[2] 刘望宏[1] 倪德斌[1] 吴斌[2,3] 

机构地区：[1]华中农业大学动物科技学院,武汉430070 [2]华中农业大学动物传染病诊断中心,武汉430070 [3]华中农业大学农业微生物学国家重点实验室,武汉430070

出　　处：《华中农业大学学报》2012年第4期485-489,共5页Journal of Huazhong Agricultural University

基　　金：湖北省研究与开发计划项目(2010BBB008)

摘　　要：根据猪库布病毒的3 D基因设计1对引物,建立了两步法RT-PCR检测方法,使用该方法分别对CSFV、PRRSV、JEV、SIV、PEDV、TGEV、GARV阳性模板及包含猪肠道病毒3 D基因和口蹄疫病毒3 D基因的重组质粒进行PCR检测,结果从以上9种常见猪病病原的阳性模板中均不能扩增出323bp大小的PCR产物,说明该检测方法的特异性很好,能够用作临床样品的检测。敏感性试验显示,本试验建立的检测方法能够检测到的模板最低质量浓度为180fg/mL。应用该方法对湖北省各大猪场进行了临床病料检测,在采集的165份病料中有118份样品检测为猪库布病毒阳性。在4个发生腹泻疫情的规模化猪场进行了分群抽样调查,结果显示,猪库布病毒在猪群中集中分布在发生腹泻疫情的猪群,说明猪库布病毒和现阶段的腹泻疫情有紧密的联系。One pair of primers was designed based on the 3D gene sequences of porcine kobuvirus, and the two-step RT-PCR assay was established in this study. The positive templates of CSFV, PRRSV, JEV, SIV, PEDV, TGEV, GARV and recombinant plasmids （pMD18T-PEV3D, pMD18T-FMDV3D） which contain either FMDV＇s or PEV＇s 3D gene were used to test the specificity of the porcine kobuvirus RT-PCR assay we established and the assay showed excellent specificity. Sensitivity test showed that the minimal concentration of positive template the assay can detect is 180 fg/mL. The epidemiology of porcine kobuvirus in Hubei Province was then investigated by applying this assay. Among 165 samples of pigs with diarrhea, 118 samples were kobuvirus positive. To investigate the distribution of porcine kobuvirus in different age groups in pig farm with diarrhea, the samples of different age groups from 4 different pig farms were collected. The results clearly indicated that porcine kobuvirus were centralized distributed in the pig group with diarrhea, which suggested that positive relationship exists between diarrhea disease outbreak and porcine kobuvirus.

关 键 词：猪库布病毒 RT-PCR 检测方法 腹泻 流行病学调查 

分 类 号：S852.651[农业科学—基础兽医学]