猪Delta冠状病毒的流行病学调查及M基因序列分析

Epidemiological investigation and M gene sequence analysis of Porcine Deltacoronavirus

为了解猪Delta冠状病毒(Porcine Deltacoronavirus,PDCoV)在发生腹泻疫情猪群中的感染情况,根据PDCoV M基因的保守序列设计了一对RT-PCR检测引物,建立了猪Delta冠状病毒的RT-PCR的检测方法,其最低核酸检测限量为3.92×10^3 copies/μL,特异性和灵敏度均较好。利用该方法对2015—2017年上海周边猪场的518份猪腹泻粪便样品进行检测,检测出25份PDCoV阳性样品,感染阳性率达4.8%。在所检出的阳性样本中,PDCoV与猪嵴病毒(porcine kobuvirus,PKV)和猪星状病毒(porcine astrovirus,PAstV)的混合感染率较高,分别为40%和48%;PDCoV与猪流行性腹泻病毒(porcine epidemic diarrhea virus,PEDV)混合感染率为8.0%,未检测到PDCoV与猪传染性胃肠炎病毒(transmissible gastroenteritis virus,TEGV)混合感染的样本。随机选取10份PDCoV阳性样本,对扩增的M基因片段同源性进行分析,结果,10份样品中核苷酸同源性达95.0%—99.6%,与GenBank登录的PDCoV毒株的同源性为96.1%—100%,说明目前流行的PDCoV毒株的遗传差异不大,该研究为了解PDCoV流行情况提供了参考依据。

To investigate the epidemiology of porcine deltacoronavirus(PDCoV)in domestic area,a pair of primers for amplifying M gene of PDCoV was designed and used for establishing PCR detection method.Results showed that the primer was in good specificity with the amplification fragment of 281 bp.The minimum nucleic acid detection limit was 3.92×1O^3 copies/μL which confirmed the good sensitivity.Around Shanghai area,518 fecal samples collected in 2015—2017 from the pig herds with diarrhea symptom in clinic were detected using this method.Twenty-five samples were PDCoV positive with the infection rate of 4.8%.Furthermore,co-infection of PDCoV with PKV or PAstV were commonly seen with the co-infection rates of 40%and 48%respectively.Coinfection of PDCoV and PEDV was 8.0%.No co-infection of PDCoV and TGEV was observed.Ten PDCoV positive samples were further selected for M gene nucleic acid amplification and sequence analysis.Their nucleotides identities were among 95.0%—99.6%.And the identity with other PDCoV genomes from GenBank were 96.1%—100%,which meaned the PDCoV was conserved till now.The study could provide the basis for epidemiological survilliance of PDCoV.