一株猪源1型呼肠孤病毒的分离及鉴定

Isolation and identification of a strain of porcine reovirus serotype 1 in China

为了研究猪呼肠孤病毒的分子生物学特性,通过在细胞培养液中添加胰酶的方法,从仔猪腹泻粪样中分离、纯化并鉴定了1株能在Vero细胞上稳定产生细胞病变,以细胞颗粒增多、肿胀、漂落等细胞病变为主要特征的猪源1型呼肠孤病毒SHR-A株。根据呼肠孤病毒的理化特性,对病毒进行初步纯化,然后进行电镜观察、核酸提取、RNA电泳、RT-PCR、各病毒基因扩增和克隆测序等常规病毒学鉴定。SHR-A株S1基因的同源性分析和核苷酸系统发育进化树分析表明,该病毒与哺乳动物呼肠孤病毒血清1型的同源性较高,而与血清2型和3型的同源性较低,因此初步判断该SHR-A株为血清1型,为国内首次报道从猪体内分离到血清1型的呼肠孤病毒。进一步分析发现,SHR-A的S1基因全长为1 465bp,其3′-UTR为3型,其余部分则为1型,说明其S1基因是1型和3型的嵌合体,此结果表明,哺乳动物呼肠孤病毒作为RNA病毒,基因重组可能是其除基因突变和基因重排以外的第3种病毒变异进化的重要方式。

In order to study the molecular characteristics of porcine reovirus,a strain of reovirus,SHR-A,was isolated from Vero cell line by the method of adding trypsin into the culture DMEM from piglets＇ diarrhea fecal sample.The stable cytopathic effect caused by the virus including cell granule increasing,cellular swelling and shedding were observed.SHR-A strain was purified based on reovirus physical and chemical properties,and identified by conventional virological methods.Sequence analysis showed that the strain belonged to reovirus serotype 1.S1 gene（1 465 bp in full-length） of SHR-A showed higher similarity to that of mammalian orthoreovirus（MRV） serotype 1 than that of other serotype viruses.This is the first isolate of reovirus serotype 1 from porcine herds in China.3′-UTR of SHR-A S1 genome segment shared higher homology to that of serotype 3 than to serotype 1,indicating that the S1 gene was a chimera of serotype 1 and serotype 3.The results provided evidence that MRV as RNA virus might have undertaken genomic recombination in the process of viral evolution,which is the third process beside genomic reassortment and genomic point mutation.