鹅星状病毒、鹅细小病毒双重PCR检测方法的建立及初步应用

Establishment and Primary Application of a Duplex PCR Methodfor Detection of Goose Astrovirus and Goose Parvovirus

为快速鉴别诊断鹅星状病毒(GoAstV)和鹅细小病毒(GPV),根据GoAstV和GPV基因序列保守区域设计了两对特异性引物,通过PCR扩增目的片段并构建重组质粒,建立了一种可同时检测这两种病毒的PCR诊断方法。该方法能够特异性扩增出两种病毒的相应片段,而对鹅副粘病毒(GPMV)、鹅流感病毒(GAIV)、鹅腺病毒(GADV)、鹅圆环病毒(GoCV)均无扩增,对重组质粒的最低检测拷贝数分别为1.25×10^(3) copies和1.25×10^(5) copies。应用该方法对50份鹅临床脏器样品进行检测,结果检测出GoAstV 35份、GPV 2份,GoAstV和GPV混合感染2份。结果表明,研究建立的双重PCR检测方法快速、特异、敏感,可用于GoAstV和GPV的流行病学调查和疾病监测。

In order to quickly detect goose astrovirus(GoAstV)and goose parvovirus(GPV)simultaneously,a duplex PCR method was established by designing two pairs of specific primers targeting the conserved sequence of GoAstV and GPV,amplifying the target fragment and constructing recombinant plasmid.The results showed that the target fragments of these two viruses were amplified,but no amplification of goose paramyxovirus(GPMV),goose influenza virus(GAIV),goose adenovirus(GADV),goose circovirus(GoCV).The minimum detection copy number for recombinant plasmid of GoAstV and GPV were 1.25×10^(3) and 1.25×10^(5),respectively.A total number of 50 fecal samples from goose were tested by the established duplex PCR,of which 35 samples were GoAstV positive,2 samples were GPV positive,and 2 samples were GoAstV and GPV positive.The above results indicate that the duplex PCR method established in this study is rapid,specific and sensitive,and can be used for epidemiological investigation and disease surveillance of the above two viruses.