福建农业学报2024，Vol.39Issue(11)：1242-1247,6.DOI:10.19303/j.issn.1008-0384.2024.11.005
赤羽病病毒、牛疱疹病毒1型和牛病毒性腹泻病毒多重RT-PCR检测方法的建立
A Multiplex RT-PCR Assay for Simultaneous Detecting Akabane Virus,Bovine Herpesvirus 1,and Bovine Viral Diarrhea Virus
摘要
Abstract
[Objective]A multiplex RT-PCR assay for rapid and simultaneous detection of akabane virus(AKAV),bovine herpesvirus 1(BoHV-1),and bovine viral diarrhea virus(BVDV)was developed.[Method]Three pairs of primers were designed for the conservative regions of S genes of AKAV,gH of BoHV-1,and 3'UTR of BVDV for establishing an RT-PCR assay capable of detecting the three viruses simultaneously under optimized reaction conditions.[Result]The assay detected positive only on AKAV,BoHV-1,and BVDV but negative on bovine parvovirus(BPV),bovine respiratory syncytial virus(BRSV),bovine parainfluenza virus 3(BPIV-3),foot-and-mouth disease virus(FMDV),and bovine ephemeral fever virus(BEFV).The minimum detection thresholds of the assay were 1.0×103 copies·μL-1 on the recombinant plasmids of the three target viruses with high intra-and inter-batch reproducibility.On 143 whole blood/tissue specimens of cattle with reproductive disorders that included cases with mixed infections,it delivered the positivity rates on AKAV,BoHV-1,and BVDV of 2.80%(4/143),21.68%(31/143),and 38.46%(55/143)as well as the conformity rates with the local standards for AKAV,BoHV-1,and BVDV of 99.3%,98.6%,and 97.2%with the Kappa coefficients of 0.885,0.960,and 0.942 respectively.[Conclusion]The newly established multiplex RT-PCR assay displayed high specificity,sensitivity,and reproducibility in detecting AKAV,BoHV-1,and BVDV simultaneously with a considerably reduced operation cost.关键词
赤羽病病毒/牛疱疹病毒1型/牛病毒性腹泻病毒/多重RT-PCR/繁殖障碍Key words
arkaban virus/bovine herpesvirus 1/bovine viral diarrhea virus/multiplex RT-PCR/reproductive disorders分类
农业科技引用本文复制引用
张帆帆,韦家禛,李杰茂,谭美芳,胡利珍,曾艳兵,龚小齐,杨群,徐俊..赤羽病病毒、牛疱疹病毒1型和牛病毒性腹泻病毒多重RT-PCR检测方法的建立[J].福建农业学报,2024,39(11):1242-1247,6.基金项目
江西省重点研发计划项目(20201BBF61008) （20201BBF61008）
江西省现代农业科研协同创新专项(JXXTCX201702、JXXTCX202405) （JXXTCX201702、JXXTCX202405）
