猪Deltacoronavirus RT-PCR检测方法的建立及其应用

Development and application of a RT-PCR assay for detection of porcine Deltacoronavirus

猪Deltacoronavirus（PDCoV）是2014年在美国检测出的一种新型猪冠状病毒，能够引起感染猪出现水样腹泻、呕吐和脱水等症状。为及时评估该病毒在我国猪群中的感染和流行情况，本研究根据GenBank中登录的PDCoV核苷酸序列设计并合成一对特异性引物，在我国首次建立了PDCoV的RT．PCR检测方法。研究结果表明，所建立的RT．PCR检测方法仅对PDCoV核酸具有特异性扩增，对其它猪主要病毒核酸的扩增均呈阴性，具有较强的特异性；对PDCoV最低检测限为4．05×103拷贝／μL，敏感性较高；采用该方法对2014年～2015年我国华东地区猪场的526份临床样品进行检测，结果显示PDCoV总阳性率为4．75％（25／526），其中腹泻样品中阳性率为25．76％（17／66），表明我国猪群中存在PDCoV感染。本研究建立的RT—PCR方法可以作为临床检测PDCoV的一种有效手段。

Porcine Deltacoronavirus （PDCoV） is a new coronavirus that has been identified in United States since 2014, which caused watery diarrhea, vomiting, and dehydration. To investigate the prevalence of PDCoV in China, a RT-PCR assay was developed for detection of PDCoV with a pair of primers designed according to PDCoV sequence available in GenBank. The test results showed that the RT-PCR assay was specific for PDCoV detection, but had no cross-reaction with closely related coronavirus or other animal viruses, and the detection limit of the assay was approximately 4.05×103 copies/μL. In addition, PDCoV infection in pigs was detected in 25 out of 526 （4.75%） clinical samples （25.76% for samples from diarrhea pigs） collected from 2014 to 2015 from pig farms in Eastern China. In this study, we present the first evidence of PDCoV infection in mainland of China using the RT-PCR assays. The RT-PCR assay could be applied as a diagnostic technique for the detection of PDCoV.