鹅细小病毒PCR检测方法的建立及初步应用

根据GenBank中鹅细小病毒(GPV)的7株全基因序列,在相对保守的NS区域设计了一对引物,以GPV-98E株鹅胚尿囊液的核酸提取物为模板,经优化反应条件,建立了特异性检测GPV的PCR方法。敏感性试验结果显示,该方法对GPV核酸的最小检出量为4.194 pg,尿囊液的最小检出量为3.09个ELD50;特异性试验结果显示,采用该方法检测GPV能够扩增出与预期大小相符的476 bp的特异性片段,而对作为对照的鸭瘟病毒(DPV)、禽流感病毒(AIV)、鹅副粘病毒(GPMV)的核酸均未扩增出任何条带。在临床检测中,对52份雏鹅的心、肝、肠等组织进行PCR扩增,能够检测到相应的特异性病毒核酸片段,表明建立的鹅细小病毒PCR检测方法具有敏感性高、特异性强的特点,具有潜在的临床应用价值。

Development and preliminary application of PCR assay for detection of goose parvovirus

One pair of primers was designed in relatively conservative NS region according to all the sequences of seven goose parvovirus strains in Genbank.Nucleic acids of GPV-98E strain which was extracted from allantoic fluid of goose embryo serve as template and a PCR assay was developed based on optimization of reaction conditions.The sensitivity of the PCR reached 4.194 pg for the GPV nucleic acids and 3.09 ELD50 for allantoic fluid of goose embryo respectively.Using the PCR assay,the specific fragment of 476 bp was amplified from DNA sample of the GPV only,which agreed with expectation,but not amplified from duck plague virus,avian influenza virus and goose paramyxovirus.All the specific fragments were amplified by the assay from 52 samples of goslings infected clinical with GPV including heart,liver and intestine,indicating the PCR assay developed for GPV was sensitive and specific,which had potential clinical value.