犬细小病毒广西流行株的分离鉴定及生物学特性分析  

Isolation and identification of canine parvovirus isolated from Guangxi and analysis of it's biological characteristics

作　　者：刘金凤 廖文军 陈凤莲 白安斌 钟华[2] 凌丹 吴健敏 LIU Jin-feng LIAO Wen-jun CHEN Feng-lian BAI An-bin ZHONG Hua LING Dan WU Jian-min(Guangxi Key Laboratory of Veterinary Biotechnology, Guangxi Veterinary Research Institute, Nanning 530001, China College of Marine and Bioteehnology, Guangxi University For Nationalities, Nanning 530001, China Wuming Animal Disease Prevention and Control Center, Nanning 530001, China)

机构地区：[1]广西壮族自治区兽医研究所广西兽医生物技术重点实验室,广西南宁530001 [2]广西民族大学海洋与生物技术学院,广西南宁530006 [3]武鸣县动物疫病预防控制中心,广西南宁530199

出　　处：《中国兽医科学》2017年第7期877-883,共7页Chinese Veterinary Science

基　　金：国家重点研发计划重点专项(2016YFD0501000)

摘　　要：为了解当前犬细小病毒广西地方毒株的生物学特性,本研究收集广西各地疑似犬细小病毒(CPV)感染病犬的粪便通过接种F81细胞进行病毒的分离培养,并通过血凝试验、抗性分析、同源性分析和攻毒试验等对分离毒株的生物学特性及致病力进行研究。结果显示,接种3代后,细胞出现明显的病变(CPE),血凝效价为1∶512,第5代病毒的TCID_(50)为1×10^(-3.6)/0.1 m L,能抗热、酸、氯仿和乙醚;电镜观察到CPV为直径约25 nm圆形、无囊膜的结构,应用CPV特异性引物检测到长约1 419 bp和1 254 bp的特异性条带,提示成功分离到一株犬细小病毒毒株;测序分析结果表明,VP2基因与周边国家地区及我国各地方的参考毒株、流行株核苷酸同源性为98.9%~99.8%;系统进化发生分析表明,广西分离株的基因型为CPV-2a亚型,与泰国参考毒株VT148亲缘关系最近;与其它CPV-2a亚型相比,分离株的第267位、305位、324位和440位氨基酸位点发生了变异;动物回归试验结果表明,犬细小病毒广西分离株属于强毒株。本研究成功分离到犬细小病毒广西地方流行株,并对其主要衣壳蛋白VP2基因进行遗传变异分析,为更好的预防和控制犬细小病毒提供依据。In order to understand the biological characteristics of Guangxi local canine parvovirus（CPV）strain,in this study,feces were collected from dogs suspected of CPV infection in Guangxi province and were inoculated with FK81 cells to isolate CPV.Research was also done on the biological characteristics and pathogenicity of the isolate with hemagglutination test,resiitance analysis,homology analysis and challenge test.The results showed that the typical CPE was spotted and the HA blood coagulation rate was 1 ∶ 512 after the cells were passaged to the third generation.The virus TCID50 of the fifth generation virus was 1×10^-3.6/0.1 mL,and the strains was anti-heat,anti-acid and insensitive to chloroform and aether.Round and non-enveloped particles about 25 nm in diameter were observed by electron microscope.1 419 bp and 1 254 bp bands were detected by specific primers against CPV.The results suggested that a canine parvovirus strain was successfully isolated.This strain,according to VP2 gene homology analysis,shared from 98.9% to 99.8% homology with reference strains reported in neighboring countries and our country.Phylogenetic analysis showed that the genotype of this isolated strain was CPV-2a subtype,and had the closest relationship with Thailand VT148 reference strain.267 th,305th,324 th and 440 th amino acid of this strain were detected to have mutated,completely different from other isolates of CPV-2a subtype.Animal regression test results also suggested that this isolated CPV strain belonged to virulent strain.To sum up,in this research, an epidemic strain of canine parvovirus in Guangxi was successfully isolated,and the genetic variation of its major capsid protein VP2 gene was analysed.The results will provide basic information for the better prevention and control of canine parvovirus in the future.

关 键 词：犬细小病毒 分离 生物学特性 

分 类 号：S852.659.2[农业科学—基础兽医学]