北美H7亚型禽流感病毒SYBR Green Ⅰ实时荧光定量PCR检测方法的建立  

SYBR Green Ⅰ real-time PCR assay for detection of North American H7 subtype avian influenza virus

作　　者：王晨曦[1] 张谞霄 蒲娟[1] 孙洪磊[1] WANG Chen-xi;ZHANG Xu-xiao;PU Juan;SUN Hong-lei(College of Veterinary Medicine,China Agricultural University,Beijing 100193,China)

机构地区：[1]中国农业大学动物医学院,北京海淀100193

出　　处：《中国兽医杂志》2018年第4期32-34,I0001,共4页Chinese Journal of Veterinary Medicine

基　　金："十二五"国家科技支撑项目"新发动物流感病原分子演化规律和检测技术"(2013BAD12B01)

摘　　要：H7N9亚型禽流感病毒(AIV)对家禽养殖业和公共卫生的持续威胁提示我们应加强对H7亚型流感病毒的监测。根据血凝素(HA)基因的分子遗传进化特点,H7亚型AIV分为欧亚和北美两个进化分支,我国自然分离到的H7亚型AIV均属于欧亚分支,但是目前我国尚未检测到北美H7亚型AIV。本研究通过分析NCBI数据库中H7亚型AIV HA基因序列,设计特异性引物,建立了针对北美H7亚型AIV HA基因的实时荧光定量PCR检测方法。该方法仅针对北美分支H7亚型AIV HA基因出现特异性扩增曲线,不能检出欧亚分支H7亚型AIV、其他亚型(H3、H5和H9)流感病毒以及新城疫病毒和鸡传染性支气管炎病毒;检测方法的灵敏度可达53 Copies/μL。该检测方法的建立为监测北美H7亚型AIV提供了良好的技术支撑,可用于外来AIV的监测。The emergence of H7 N9 avian influenza virus（ AIV） in human and chicken highlights the importance of surveillance for H7 subtype AIV. Based on the phylogeny of hemagglutinin（ HA） gene,and H7 subtype AIVs are divided into two clusters,the North American-like and Eurasian-like. All the H7 viruses isolated in China belong to the Eurasian-like,while North American H7 AIVs have not been found in China. In this study,a pair of specific oligo nucleotide primers was designed based on HA sequences available from NCBI database,and SYBR Green Ⅰ real-time PCR assay was established for the detection of North American H7 subtype AIV. The amplification by the real-time PCR assay was positive for North American H7 AIV but negative for Eurasian H7 subtype AIVs（ H7 N2 and H7 N9）,other subtypes（ H3,H5 and H9） of influenza viruses,Newcastle disease virus（ NDV） and infectious bronchitis virus（ IBV）. The sensitivity was 53 copies/μL. This assay will provide a useful detection method for North American H7 subtype AIV in the surveillance of exotic diseases.

关 键 词：H7亚型禽流感病毒 北美分支 HA 荧光定量PCR 

分 类 号：S852.655[农业科学—基础兽医学]