王方洲;孙普;张婧;李平花;马雪青;赵志荀;李凤娟;李国秀;李冬;王健;袁红;曾巧英;刘在新;卢曾军;

• 1:中国农业科学院兰州兽医研究所家畜疫病病原生物学国家重点实验室国家口蹄疫参考实验室
• 2:甘肃农业大学

摘要(Abstract)：

美洲型猪繁殖与呼吸综合征病毒（Porcine reproductive and respiratory syndrome virus,PRRSV）在我国出现了多种基因亚型毒株混合流行和重组的复杂变化，其引起的猪繁殖与呼吸综合征（PorcineReproductiveand Respiratory Syndrome,PRRS）已成为危害程度仅次于非洲猪瘟的重要疫病。为建立一种快速而敏感性高的美洲型PRRSV检测方法，本研究分析了国内美洲型全基因组序列，利用重组酶聚合酶扩增技术（Recombinase Polymerase Amplification,RPA），设计合成特异性引物和探针，优化反应体系和条件，建立美洲型PRRSV实时荧光RPA检测方法，通过特异性、敏感性、重复性试验及田间样品的比对检测，评价该方法的检测能力，并测定和分析了田间阳性样品的ORF5基因序列。结果显示，该方法在42℃、20 min内即可完成检测，最低检测美洲型PRRSV载量为50拷贝/μL，且与欧洲型PRRSV、口蹄疫病毒、猪瘟病毒、塞内卡病毒、非洲猪瘟病毒、猪伪狂犬病毒、猪圆环病毒、猪流行性腹泻病毒无交叉反应，特异性好；检测50份田间样品该方法的阳性检出率为34%，与实时荧光RT-PCR检测结果的符合率为100%;ORF5序列分析显示检出阳性样品中NADC30-Like毒株占比为88%。结果表明，本研究建立的美洲型PRRSV实时荧光RPA检测方法特异性强、敏感性高，满足PRRSV的快速诊断和流行病学调查。

关键词(KeyWords)： PRRSV;美洲型;RPA;应用;ORF5基因

Abstract:

Keywords:

基金项目(Foundation): 甘肃省农业农村厅科技项目（项目号：GSKL-2020-7-2）,题目：甘肃省生猪健康养殖疫病防控及生物安全技术集成试验示范项目;; 中国农业科学院兰州兽医研究所创新工程揭榜挂帅项目（项目号：CAAS-ASTIP-JBGS-20210601）,题目：猪繁殖与呼吸综合征表位缺失标记疫苗的研制~~

作者(Authors): 王方洲;孙普;张婧;李平花;马雪青;赵志荀;李凤娟;李国秀;李冬;王健;袁红;曾巧英;刘在新;卢曾军;

DOI: 10.13242/j.cnki.bingduxuebao.004263

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