牛诺如病毒、牛星状病毒和牛环曲病毒多重PCR检测方法的建立及应用  

Establishment and application of a multiple PCR for detection of BNoV,BAstV and BToV

作　　者：师志海[1,2] 徐照学 兰亚莉[1] 张彬 孟红丽[1] 王亚州[1] 金磊[1] 王文佳[3] SHI Zhi-hai;XU Zhao-xue;LAN Ya-li;ZHANG Bin;MENG Hong-li;WANG Ya-zhou;JIN Lei;WANG Wen-jia(Institute of Animal Husbandry and Veterinary Medicine,Henan A-cademy of Agricultural Sciences,Zhengzhou 450002,China;Henan Key Laboratory of Farm Animal Breeding and Nutritional Regulation,Zhengzhou 450002,China;College of Veterinary Medicine and Pharmaceutical Engineering,Henan University of Animal Husbandry and Econo-my,Zhengzhou 450046,China)

机构地区：[1]河南省农业科学院畜牧兽医研究所,河南郑州450002 [2]河南省畜禽繁育与营养调控重点实验室,河南郑州450002 [3]河南牧业经济学院动物医药学院,河南郑州450046

出　　处：《中国兽医学报》2020年第10期1924-1928,共5页Chinese Journal of Veterinary Science

基　　金：河南省重点研发与推广专项(科技攻关)资助项目(192102110074);“十三五”国家重点研发计划资助项目(2018YFD051700);国家现代农业肉牛牦牛产业技术体系资助项目(CARS-37)。

摘　　要：为建立能同时检测牛诺如病毒(BNoV)、牛星状病毒(BAstV)和牛环曲病毒(BToV)的方法,根据BNoV、BAstV和BToV基因组的保守区域设计3对特异性引物,预期特异性扩增BNoV、BAstV和BToV的片段大小分别为542,376,698 bp,建立了BNoV、BAstV和BToV的多重PCR检测方法。结果显示,该方法能特异性扩增BNoV、BAstV和BToV的目的条带,且未扩增出其他犊牛腹泻病原;BNoV、BAstV和BToV的最低检测限分别为1.34×10^4,1.06×10^4,1.03×10^4拷贝/μL。对采集自河南省的221份犊牛腹泻样品的检测结果显示,BNoV、BAstV和BToV的检出率分别为9.96%(22/221),18.10%(40/221)和19.00%(42/221)。三者与单一PCR检测结果相一致。结果表明,本试验建立的多重PCR方法可用于BNoV、BAstV和BToV的检测和流行病学调查。To establish a PCR method for simultaneous detection of bovine norovirus(BNoV),bovine astrovirus(BAstV) and bovine torovirus(BToV),three pairs of specific primers were designed based on the conserved regions of BNoV,BAstV and BToV genomes.The specific amplification of BNoV,BAstV and BToV were 542,376 and 698 bp,respectively.The method had a sensitivity of 1.34×10^4 copies/μL for BNoV,1.06×10^4 copies/μL for BAstV and 1.03×10^4 copies/μL for BToV,as well as a good specificity,which was only positive for BNoV,BAstV and BToV,but negative for other calf diarrhea virus pathogens.Furthermore,this multiple PCR method was used to test 221 clinical samples that collected from the diarrheic calves in Henan Province.The positive rate was 9.96%(22/221),18.10%(40/221) and 19.00%(42/221) for BNoV,BAstV and BToV,which is consistent with single PCR results.The multiple PCR method can be used for the rapid detection and epidemiological investigation of BNoV,BAstV and BToV.

关 键 词：牛诺如病毒 牛星状病毒 牛环曲病毒 犊牛腹泻 多重PCR 

分 类 号：S852.65[农业科学—基础兽医学]