猪盖他病毒RT-PCR检测方法的建立及其应用

Establishment and application of RT-PCR method for detection of Getah virus

参考GenBank中猪盖他病毒(Getah virus,GETV)的全基因组序列,针对NSP3保守区域设计1对扩增片段为810bp的引物,建立了一种检测GETV的RT-PCR方法,其检测灵敏度为102.25 TCID50/mL,检测PRV、PCV2、PRRSV、CSFV、JEV、PEDV、TGEV和PoRV均为阴性,特异性和可重复性均良好。应用此方法首次从79份猪临床病料中检出GETV阳性4份(5.06%);从6个厂家6个种类的63批次猪用活疫苗中共检出来源于一个厂家同一种类的3个批次GETV阳性活疫苗(批次阳性率4.76%)。结果表明,该方法具有快速、灵敏、特异等优点,可分别用于临床发病猪群和疫苗污染GETV的检测,为本病的快速诊断和确保疫苗质量提供了有效手段。

According to the complete genome sequence of GETV published in GenBank,apair of primers for the conserved region of NSP3was designed for amplifying the 810bp fragment.A RTPCR assay for detection of Getah virus was established with the sensitivity of 102.25 TCID50/mL,highly specific to GETV detection,while all amplifications were negative for PRV,PCV2,PRRSV,CSFV,JEV,PEDV,TGEV and PoRV.Moreover it possessed a good repeatability at the same time.Four positive cases(5.06%)of GETV were identified from 79porcine clinical samples using this method for the first time in China.Three batches(4.76%)of one kind of live vaccines from the same manufacture were detected as positive to GETV from all 63batches of 6kind pig live vaccines from 6manufacturers.The results showed that the method developed was rapid,sensitive,specific for GETV detection for sick swines and vaccine contamination and could benefit rapid diagnosis of the disease and the quality of the vaccine.