猪流行性腹泻病毒云南流行毒株分离与鉴定

Isolation and Identification of Porcine Epidemic Diarrhea Virus in Yunnan

为有效控制和预防猪流行性腹泻(PED),分离获得猪流行性腹泻病毒(PEDV)云南地方流行毒株,并选取部分经RT-PCR鉴定为阳性的样品,处理后接种在Vero细胞中进行分离培养,细胞培养物盲传至15代后可观察到典型的细胞病变(CPE),对细胞培养物通过RT-qPCR方法检测和间接免疫荧光(IFA)鉴定,并对分离株S基因全序列进行克隆和遗传进化分析。结果表明:获得的细胞培养物均为PEDV阳性,病毒半数细胞感染量(TCID_(50))为10^(-4.85)/0.1mL,将其命名为YNDH-2017毒株。该分离株S基因与参考毒株的同源性为92.8%～98.2%,和国内近年来PEDV流行毒株亲缘关系较近,存在相同的突变位点,而与经典毒株CV777株在两个大的分支上,亲缘关系较远。试验分离获得的YNDH-2017株属于当前国内流行毒株。

The isolated PEDV strain identified as a positive sample by RT-PCR is inoculated into Vero cells for separation culture and the typical cytopathic effect (CPE) of cell cultures is observed after 15 th generation culture. The cell cultures are detected and identified by RT-qPCR and indirect immunofluorescence assay (IFA) and total sequence and genetic evolution of S gene of the isolated strain are cloned and analyzed respectively to effectively control and prevent porcine epidemic diarrhea (PED). Result: The obtained cell culture with PEDV-positive and 10 -4.85 /0.1 mL of TCID 50 is named as YNDH-2017 strain. The S gene homology between YNDH-2017 strain and reference strains reaches 92.8%- 98.2%. YNDH-2017 strain has a near relationship to PEDV epidemic strains for the past few years in China and there are the same mutation sites between YNDH-2017 strain and other PEDV epidemic strains. YNDH-2017 strain has a distant relationship to typical CV777 strain. The isolated YNDH-2017 strain belongs to current epidemic strain in China.