摘要
为了确诊一例疑似SVA感染的临床病例,本试验将从广西地区某猪场母猪和肥育猪鼻及蹄部采集的出现典型水泡性病变的猪水泡液进行无菌处理后接种PK-15细胞,连续传代培养,通过细胞病变、RT-PCR扩增、基因序列测定、TCID_(50)和间接免疫荧光等方法对分离得到的病毒进行鉴定。结果显示:该病毒株在PK-15细胞上生长良好,可见明显细胞病变;接种PK-15细胞进行TCID_(50)测定,病毒滴度达到10^(-6.75)/mL;间接免疫荧光试验结果显示,接种第4代细胞培养物的BHK-21细胞出现特异的绿色荧光;扩增和测定了VP1基因序列后进行同源性及遗传进化分析,发现该毒株与Senecavirus病毒属中的病毒成员同源性最高,与之前2017年报道的广东分离株SVA-CH-GDYS02-2017和SVA-CH-GDLZ01-2017位于较近的进化分支内。以上结果表明,已成功从样品中分离到了SVA,且该毒株可以在PK-15和BHK-21细胞上进行增殖。本研究成功分离到一株SVA,为后续塞内卡病毒的诊断和疫苗研制提供试验材料。
To confirm a SVA infection case on a pig farm in Guangxi province,monolayers of porcine kidney cell(PK-15)were inoculated with the clinical samples that were RT-PCR positive.The virus isolated from PK-15 cells was identified by CPE formation,RT-PCR,sequencing and indirect immunofluorescent assay(IFA).The result showed that the strain grew well and caused obvious CPE in PK-15 cells.The virus harvest was titrated on PK-15 cells and the virus titer(TCID_(50))reached to 10^(-6.75)/mL.Moreover,specific green fluorescence was observed in inoculated BHK-21 cell at the fourth passage as showed in IFA.Homology and genetic evolution analysis based on the VP1 gene sequence indicated that the strain belonged to Seneca virus and located in a relatively close evolutionary branch with Guangdong isolates SVA-CH-GDYS02-2017 and SVA-CH-GDLZ01-2017 that were reported in 2017.The availability of the SVA strain provided materials for the diagnosis and vaccine development for Seneca virus disease.
作者
牛晨霞
王豪
农作荣
全东群
曾悦
任同伟
王玉旭
王景隆
刘畅
陈樱
欧阳康
黄伟坚
韦祖樟
NIU Chenxia;WANG Hao;NONG Zuorong;QUAN Dongqun;ZENG Yue;REN Tongwei;WANG Yuxu;WANG Jinglong;LIU Chang;CHEN Ying;OUYang Kang;HUANG Weijian;WEI Zuzhang(Laboratory of Animal infectious Diseases and molecular Immunology,College of Animal Science and Technology,Guangxi University,Nanning 530005,China)
出处
《中国动物传染病学报》
CAS
北大核心
2022年第2期20-26,共7页
Chinese Journal of Animal Infectious Diseases
基金
国家自然科学基金(31972666)。
关键词
塞内卡病毒
水疱性疾病
分离鉴定
序列分析
Seneca Valley virus
vesicular disease
isolation and identification
sequence analysis
