摘要
【目的】建立能同时检测新型鸭呼肠孤病毒(New-type duck reovirus,NDRV)、新型鹅细小病毒(Novel goose parvovirus,NGPV)和鸭坦布苏病毒(Duck Tembusu virus,DTMUV)的方法。【方法】根据NDRV、NGPV和DTMUV基因组的保守区域设计3对特异性引物,预期特异性扩增NDRV、NGPV和DTMUV的片段大小分别为594、467、328 bp,建立可同时检测NDRV、NGPV和DTMUV的多重PCR检测方法,对其进行特异性、敏感性及重复性检验,并在临床上进行初步应用。【结果】特异性检测结果表明,该多重PCR方法可同时扩增NDRV、NGPV、DTMUV的目的片段,且未扩增出其他鸭常见病原;敏感性结果显示NDRV、NGPV和DTMUV的检测下限分别为8.80×10^(4)、4.03×10^(4)、2.15×10^(4)copies·μL^(−1);重复性试验表明该方法具有良好的重复性。采用建立的多重PCR方法对167份临床样品进行检测结果显示,其检测结果与该3种病毒的常规单一PCR检测结果符合率为100%。【结论】建立的多重PCR检测方法具有特异性强、敏感性高、重复性好等优点,适用于临床样品中存在以上3种病原感染的检测和流行病学调查。
【Objective】A PCR method for simultaneous detection of new-type duck reovirus(NDRV),novel goose parvovirus(NGPV),and duck tembusu virus(DTMUV)was developed and tested for clinic applications.【Method】Separate sets of specific primers were designed based on the conserved regions of NDRV,NGPV,and DTMUV genomes.The target specific amplifications of NDRV,NGPV,and DTMUV were in the regions of 594 bp,467 bp,and 328 bp,respectively.The specificity,sensitivity,and repeatability of the assay method were determined prior to a trial determination on clinic specimens.【Result】The newly developed multiplex PCR methodology delivered highly repeatable results and showed a high specificity on the 3 viruses with negative results on other commonly found duck pathogens.The detection sensitivity on NDRV was 8.80×10^(4) copies·μL^(−1),4.03×10^(4)copies·μL^(−1) on NGPV,and 2.15×10^(4) copies·μL^(−1) on DTMUV.On 167 clinical samples a perfect 100%total coincidence rate between the new and conventional PCR methods were achieved in detecting the 3 viruses.【Conclusion】The newly established multiplex PCR assay was specific,sensitive,and repeatable in simultaneously detecting NDRV,NGPV,and DTMUV.It was considered adequate for clinic applications.
作者
李海琴
傅光华
康昭风
韦启鹏
谭美芳
黄江南
季华员
黄瑜
杨群
LI Haiqin;FU Guanghua;KANG Zhaofeng;WEI Qipeng;TAN Meifang;HUANG Jiangnan;JI Huayuan;HUANG Yu;YANG Qun(Institute of Animal Husbandry and Veterinary Medicine,Jiangxi Academy of Agricultural Sciences,Nanchang,Jiangxi 330200,China;Institute of Animal Husbandry and Veterinary Medicine,Fujian Academy of Agricultural Sciences,Fuzhou,Fujian 350013,China)
出处
《福建农业学报》
CAS
CSCD
北大核心
2022年第1期1-6,共6页
Fujian Journal of Agricultural Sciences
基金
江西省重点研发计划项目(20202BBFL63025、20192BBF60028、20161BBF60139)
江西现代农业科研协同创新专项(JXXTCX202106)
国家水禽产业技术体系建设专项(CARS-43-33)
江西省水禽产业技术体系项目(JXARS-09)
江西省“双千计划”人才项目(JXSQ2018102118)。
