摘要
鸭甲型肝炎病毒（Duck hepatitis A virus,DHAV）是引起雏鸭病毒性肝炎的主要病原之一。本研究将2013-2015年分离的5株DHAV进行了鸭胚传代复苏,发现JS2013株的鸭胚适应性最好,可导致鸭胚在接毒后2-5d出现有规律的死亡。应用血凝试验和PCR方法进行了鸭坦布苏病毒（DTMUV）、鸭瘟病毒（DPV）和鸭圆环病毒（Du CV）等的检测,结果均为阴性。将DHAV JS2013株继续在鸭胚传代并测定其病毒滴度,发现其在5至8代之间病毒滴度稳定提升,第8代病毒的ELD50为10-7.5/0.1m L,但8至20代没有出现有规律的病毒滴度提升。为了揭示DHAV JS2013株的基因组特征,应用RT-PCR方法分9段扩增了病毒基因片段,通过序列拼接获得了全基因组序列,Gen Bank登录号为KP721458。同源比对发现,与DHAV JS2013株基因同源性最高的毒株属于基因I型DHAV。将其编码多聚蛋白的序列与同源性最高的10株已登录序列的DHAV毒株进行比对,发现存在8处氨基酸位点变异。该研究探明了DHAV JS2013株在鸭胚的增殖特性及其基因特征,为研制新型鸭肝炎病毒灭活疫苗奠定基础。
Duck hepatitis A virus（ DHAV） is one of the main pathogens that causes duck viral hepatitis. In this study,five strains of DHAV isolated between 2013 and 2015 were recovered by passaging in duck embryos. Among the five strains,the inoculation of DHAV JS2013 led to the most regular death of duck embryo during 2 to 5 days post inoculation,indicating the best adaptation to duck embryo.Then,other duck virus such as DTMUV,DPV and Du CV were detected by hemagglutination test and PCR method,each receiving the negative results. DHAV strain JS2013 was then continuously passaged in duck embryos,and it was found that the virus titer stably increased from the fifth to eighth generation. The eighth generation of DHAV JS2013 was determined with the ELD50 of 10- 7. 5/0. 1m L. However,from the8 th to the 20 th generation,no further increase in viral titer was observed. To reveal genomic features of DHAV JS2013,nine fragments covering the whole genome were amplified by RT- PCR and then the complete genome sequence was obtained with the Gen Bank accession number KP721458. Sequence analysis indicated that DHV JS2013 strain belongs to genotype I of DHAV. By analyzing the amino sequences among JS2013 and the top 10 DHAV strains with higher homology in genome,eight sporadic amino acid mutations were identified. In conclusion,the multiplication characteristics of DHAV JS2013 in duck embryos and its genetic features were discovered in the present study,which paved the way for the development of inactive duck viral hepatitis vaccine with DHAV JS2013 strain.
出处
《畜牧与兽医》
北大核心
2016年第9期38-44,共7页
Animal Husbandry & Veterinary Medicine
