蓝狐细小病毒分离鉴定及VP2基因遗传进化分析
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摘要
为了鉴定从蓝狐粪便中分离的病毒是否为细小病毒,试验采用病毒培养、电镜观察、中和试验、间接免疫荧光试验和分子生物学试验对其进行鉴定。结果表明:分离毒株在F81细胞中培养96 h后出现细胞病变(CPE);电镜观察可见直径为20 nm左右的病毒粒子;该病毒能被水貂肠炎细小病毒(MEV)阳性血清中和;PCR检测可扩增得到大小为570 bp的特异性条带,并证明分离毒株为细小病毒,将其命名为BFPV-HeB05/16;VP2全基因遗传进化与氨基酸序列分析显示,分离毒株BFPV-HeB05/16与MEV和BFPV处在同一分支上,具有较高的同源性,其关键氨基酸位点(80,300,426,564,568位)与BFPV和MEV保持一致。
To identify the virus isolated from Alopes lagopus feces,the virus was detected by cell culture, electron microscopy, neutralization test, indirect immunofluorescent assay and molecular biology. The results indicated that the isolate virus showed cytopathic effect after 96 h in F81 cells. The virus particles with a diameter of about 20 nm were observed by electron microscope. The isolate virus could be neutralized by MEV positive serum. PCR assay could be amplified to obtain specific bands with size of 570 bp, and it is proved that the isolated strain was parvovirus, which was named as BFPV-HeB05/16. The analysis of VP2 genetic evolution and amino acid sequence suggested that BFPV-HeB05/16 was on the same branch with MEV and BFPV,and had high homology, and its key amino acid sites(80,300,426,564,568) were consistent with BFPV and MEV.
To identify the virus isolated from Alopes lagopus feces,the virus was detected by cell culture, electron microscopy, neutralization test, indirect immunofluorescent assay and molecular biology. The results indicated that the isolate virus showed cytopathic effect after 96 h in F81 cells. The virus particles with a diameter of about 20 nm were observed by electron microscope. The isolate virus could be neutralized by MEV positive serum. PCR assay could be amplified to obtain specific bands with size of 570 bp, and it is proved that the isolated strain was parvovirus, which was named as BFPV-HeB05/16. The analysis of VP2 genetic evolution and amino acid sequence suggested that BFPV-HeB05/16 was on the same branch with MEV and BFPV,and had high homology, and its key amino acid sites(80,300,426,564,568) were consistent with BFPV and MEV.
引文
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[4] 成大荣, 焦库华, 李俊宝. 蓝狐细小病毒性肠炎的诊断[J]. 中国兽医杂志, 1999, 25(8):22.
[5] CHEN X Y, XIE Z J, ZHAO Z P, et al. Genetic diversity of parvovirus isolates from dogs and wild animals in China[J]. J Wildl Dis, 2011, 47(4): 1036-1039.
[6] 张海玲, 闫喜军, 柴秀丽, 等. 水貂肠炎细小病毒 PCR 检测方法的建立和应用[J]. 特产研究, 2007, 29(2): 1-3.
[7] 闫喜军,张海玲, 柴秀丽, 等. 水貂肠炎细小病毒结构蛋白 VP2基因的克隆和序列分析[J]. 特产研究, 2007, 29(4): 1-3.
[8] TRUYEN U.Evolution of canine parvovirus-a need for new vaccines[J]. Vet Microbiol, 2006, 117(1): 9-13.
[9] TSAO J, PARRISH C R. The three-dimensional structure of Canine parvovirus and its functional implications[J]. Science, 1991, 251(5000):1456-1464.
[10] 张传鹏, 张航, 高全新, 等. 犬细小病毒 SH15 株的分离鉴定与全基因组分析[J]. 中国兽医科学, 2017, 47(2): 165-171.
[2] VEIJALAINEN P. Characterization of biological and antigenic properties of raccoon dog and blue fox parvovirus: a monoclonal antibody study[J]. Vet Microbiol,1988,16(3):219-230.
[3] MIZAK B. Isolation of Polish strains of blue fox parvovirus[J].J Neurochem,1994,38(2):98-104.
[4] 成大荣, 焦库华, 李俊宝. 蓝狐细小病毒性肠炎的诊断[J]. 中国兽医杂志, 1999, 25(8):22.
[5] CHEN X Y, XIE Z J, ZHAO Z P, et al. Genetic diversity of parvovirus isolates from dogs and wild animals in China[J]. J Wildl Dis, 2011, 47(4): 1036-1039.
[6] 张海玲, 闫喜军, 柴秀丽, 等. 水貂肠炎细小病毒 PCR 检测方法的建立和应用[J]. 特产研究, 2007, 29(2): 1-3.
[7] 闫喜军,张海玲, 柴秀丽, 等. 水貂肠炎细小病毒结构蛋白 VP2基因的克隆和序列分析[J]. 特产研究, 2007, 29(4): 1-3.
[8] TRUYEN U.Evolution of canine parvovirus-a need for new vaccines[J]. Vet Microbiol, 2006, 117(1): 9-13.
[9] TSAO J, PARRISH C R. The three-dimensional structure of Canine parvovirus and its functional implications[J]. Science, 1991, 251(5000):1456-1464.
[10] 张传鹏, 张航, 高全新, 等. 犬细小病毒 SH15 株的分离鉴定与全基因组分析[J]. 中国兽医科学, 2017, 47(2): 165-171.