H13亚型禽流感病毒实时荧光定量PCR检测方法的建立及其应用Establishment and Application of a Real-time Fluorescence Quantitative PCR Assay for Detection of H13-Subtype Avian Influenza Virus
朱彤;商瑞;程凯慧;于志君;吴家强;
摘要(Abstract):
H13亚型禽流感病毒以往只在海鸥、鸭等水禽中被发现,2018年,我们实验室报道了蛋鸡感染H13亚型禽流感病毒的证据,表明H13亚型禽流感病毒存在从水禽中溢出感染陆禽的风险,因而有必要加强对H13亚型禽流感病毒的监测。实时荧光定量PCR由于具有特异性强、灵敏度高、检测时间短等优点,在病原监测工作中得到了广泛应用。本研究以H13亚型禽流感病毒的HA基因作为靶基因,设计了一对特异性荧光定量PCR检测引物,通过特异性试验、敏感性试验等,建立了H13亚型禽流感病毒实时荧光定量PCR检测方法,并应用建立的检测方法对海鸥粪便样品进行核酸检测。实验结果发现,该检测方法的灵敏度为1.60×100拷贝/μL;该检测方法对H1、H3、H5、H6、H7和H9等6个亚型的禽流感病毒无交叉反应;该检测方法的敏感性是常规PCR的10倍;海鸥粪便样品中H13亚型禽流感病毒的核酸检测阳性率为18.8%。我们建立的H13亚型禽流感病毒实时荧光定量PCR检测方法具有灵敏度高、特异性强等特点,该检测方法可用于大量临床样品的快速检测,为H13亚型禽流感病毒监测工作的顺利开展提供了技术支撑。
关键词(KeyWords): H13亚型禽流感病毒;荧光定量PCR;检测;HA基因
基金项目(Foundation): 山东省重大科技创新工程(项目号:2020CXGC010801),题目:非洲猪瘟防控关键技术研究与用;; 山东省农业重大应用技术创新项目(项目号:SD2019XM007),题目:畜禽主要疫病混合感染防控新模式的建立和应用;; 家禽所学科团队建设(项目号:CXGC2018E11),题目:家禽研究所学科团队建设;; 山东省农业科学院高层次人才及创新团队引进计划;; 济南市自主培养创新团队项目(项目号:2019GXRC025),题目:畜禽重要疫病防控及健康养殖技术研究与推广;; 山东省农业科学院农业科技创新工程(项目号:CXGC2021A12),题目:主要畜禽重要疫病防控关键技术与产品创制~~
作者(Authors): 朱彤;商瑞;程凯慧;于志君;吴家强;
DOI: 10.13242/j.cnki.bingduxuebao.004027
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