美洲型与欧洲型PRRSV双重RT-PCR检测方法的建立  

Development of a duplex RT-PCR assay for the American and European genotypes of porcine reproductive and respiratory syndrome virus

作　　者：高雅[1] 武欣欣 蒋小玲 贺晓霜 肖兴 周双海[1] GAO Ya;WU Xinxin;JIANG Xiaoling;HE Xiaoshuang;XIAO Xing;ZHOU Shuanghai(College of Animal Science and Technology,Beijing University of Agriculture,Beijing 102206,China;Hengyang Municipal Center for Animal Husbandry and Fisheries Affairs,Hengyang 421001,China)

机构地区：[1]北京农学院动物科学技术学院,北京102206 [2]衡阳市畜牧水产事务中心,湖南衡阳421001

出　　处：《北京农学院学报》2021年第4期76-80,共5页Journal of Beijing University of Agriculture

基　　金：北京高等学校高水平人才交叉培养实培计划-科研训练计划深化项目(PXM2020_014207_000009)。

摘　　要：【目的】为了建立一种具有高敏感性的能够区分猪繁殖与呼吸综合征病毒(PRRSV)美洲型与欧洲型毒株的快速检测方法。【方法】从引物浓度与退火温度先对美洲型与欧洲型PRRSV的单项RT-PCR方法分别进行条件筛选与优化,之后对其双重RT-PCR方法进行条件筛选与优化,再进行特异性与敏感性试验和临床样品检测。【结果】建立的美洲型与欧洲型PRRSV双重RT-PCR检测方法不能检出猪瘟病毒与猪流行性腹泻病毒等当前流行较广的猪源RNA病毒,对美洲型与欧洲型PRRSV的检测灵敏度分别是66,40 copies/μL。此方法对2020年5个猪场的200份血清样品的检测结果显示,美洲型与欧洲型PRRSV的个体阳性率分别为44.0%和0,与单项RT-PCR方法检测结果完全一致。【结论】建立了一种高敏感性的美洲型与欧洲型PRRSV双重RT-PCR检测方法。[Objective]This study aimed to develop a rapid detection method with high sensitivity for distinguishing the American and European genotype strains of porcine reproductive and respiratory syndrome virus(PRRSV).[Methods]The primer concentration and annealing temperature conditions of the single RT-PCR assay for the American and European genotypes of PRRSV was screened and optimized respectively,and then the conditions of their duplex RT-PCR assay was screened and optimized.Consequently the specificity and sensitivity test and clinical sample detection were carried out.[Results]For this generated duplex RT-PCR assay for the American and European genotypes of PRRSV,current epidemic swine RNA viruses such as classical swine fever virus and porcine epidemic diarrhea virus could not be detected,the detection sensitivity for the American and European genotypes of PRRSV was 66,40 copies/μL respectively.The detection results of 200 serum samples from 5 pig farms in 2020 with the duplex RT-PCR assay showed that the positive rates of the American and European genotypes of PRRSV were 44.0%and 0%respectively,which were consistent with the results of their single RT-PCR assay.[Conclusion]A duplex RT-PCR assay with high sensitivity for the American and European genotypes of PRRSV was developed.

关 键 词：猪繁殖与呼吸综合征病毒 美洲型 欧洲型 双重RT-PCR 高敏感性 

分 类 号：S852[农业科学—基础兽医学]