A型塞内卡谷病毒SDMY-CH-2021的分离鉴定及全基因组序列分析

Isolation and identification of Senecavirus A SDMY-CH-2021 and whole genome sequence analysis

为了查明山东省某养猪场水疱性传染病的病原体,试验采集了该养猪场猪只的水疱皮病料,并采用RT-PCR、病毒分离纯化、基因测序及分子生物学分析等方法对样品进行了病毒分离、鉴定和全基因组序列测定。结果表明:确诊该养猪场猪只发生水疱性传染病的病因为A型塞内卡谷病毒(SVA)感染,同时排除了猪口蹄疫病毒和猪水疱病病毒混合感染的情况,并成功分离到一株SVA,命名为SDMY-CH-2021株。该分离株在BHK-21细胞中盲传3次可引起明显的致细胞病变效应,噬斑纯化3代测定其TCID_(50)为1×10^(-8.2)/100μL;基因全长为7388 bp,开放阅读框大小为6546 bp,编码2181个氨基酸。与参考毒株相比,该分离株与广州分离株CH-01-2015和武汉分离株HB-CH-2016亲缘关系较近;VP1蛋白氨基酸序列与美国首个分离株SVV-001的VP1蛋白氨基酸序列相似性最低,为92.5%;VP1蛋白存在9个B细胞线性抗原表位,相较于参考毒株VP1蛋白的氨基酸序列存在多处突变及缺失。说明分离株SDMY-CH-2021发生了部分基因变异。

In order to identify the pathogen of vesicular infectious disease in a pig farm in Shandong Province,in the test,the blister skin samples of pigs in the farm were collected.The virus was isolated and identified by RT-PCR,virus isolation and purification,gene sequencing and molecular biological analysis,and the whole genome sequence was determined.The results showed that the pigs in the farm were infected with vesicular infectious disease because of SVA.At the same time,the co-infection of porcine Foot-and-mouth disease virus and Swine vesicular disease virus was ruled out,and a SVA strain was successfully isolated and was named SDMY-CH-2021.The blind transmission of the strain in BHK-21 cells for 3 generations caused obvious cytopathic effect.The virus titer of the 3rd generation of plaque purification was 1×10^(-8.2)TCID_(50)/100μL.The total length of the isolate was 7388 bp;the open reading frame was 6546 bp,and it encoded 2181 amino acids.Compared with the reference strains,this strain was in a relatively close evolutionary branch with Guangzhou isolate CH-01-2015 and Wuhan isolate HB-CH-2016.The amino acid sequence of VP1 protein of this isolate was the least similar to that of SVV-001,the first isolate in the United States,which was 92.5%.The VP1 protein of this strain had 9 B-cell linear epitopes,and there were many mutations and deletions in the amino acid sequence of VP1 protein compared with the reference strains.These results indicated that SDMY-CH-2021 had some genetic variation.