施马伦贝格病毒实时荧光RT-RAA快检方法的建立

Development of a Real-time RT-RAA Assay for Rapid Detection of Schmallenberg Virus

旨在建立一种快速检测施马伦贝格病毒(SBV)的方法。本研究对于SBV S基因,设计特异性引物和探针,通过优化反应条件,建立SBV反转录重组酶介导核酸恒温扩增(RT-RAA)检测方法。结果显示,该方法在38℃条件下,8 min内即可特异性检出SBV,与口蹄疫病毒(FMDV)、小反刍兽疫病毒(PPRV)、非洲马瘟病毒(AHSV)、塞内卡谷病毒(SVV)、水疱性口炎病毒(VSV)无交叉反应,其敏感性为103拷贝/反应,且批内和批间重复性变异系数均小于5%。对模拟阳性和阴性绵羊样品的检出率为100%,与SBV的RT-qPCR检测结果一致。本研究成功建立了一种快速、敏感、特异的SBV实时荧光RT-RAA方法,为SBV防控提供新技术手段。

The aim of this study was to establish a rapid detection method of Schmallenberg virus(SBV).A real-time reverse transcription recombinase acid amplification(RT-RAA)was developed with specific primers and probes based on of SBV S gene.The results showed that,through optimizing experimental conditions,the assay could complete detect SBV specifically within 8 minutes at 38℃.which can There was no cross-react with foot-and-mouth disease virus(FMDV),small pestedes petites ruminants disease virus(PPRV),African horse distemper virus(AHSV),Seneca valley virus(SVV),or vesicular stomatitis virus(VSV).The detection of limit(LOD)was 103 copies/reaction.At the same time,the co-efficient of variations in intra-and inter-assay were both less than 5%.The detection rate for the spinked positive samples and negative sheep samples was 100%,which was consistent with SBV RT-qPCR results.This study successfully established a fast,sensitive,and specific real-time RT-RAA assay for detection of SBV,which provided a new technical method for prevention and control of SBV infection.