湖南省禽白血病病毒分子流行病学调查及env基因序列分析

Molecular epidemiology and env gene sequence analysis of avian leukosis virus from chicken and pheasant in Hunan Province

为了解禽白血病病毒(ALV)在湖南省各禽类养殖场的感染情况,于2018年1月至2019年1月采集湖南省不同地区21个养殖场的156份组织与血清样品。分别设计针对ALV 5′非翻译区(5′UTR)的检测引物和env基因近全长扩增引物,通过RT-PCR首先用检测引物对样品进行检测,然后扩增阳性样品ALV env基因全长并测序分析。结果显示,19个养殖场检测出E亚群ALV(ALV-E),其env基因与GenBank中ALV-E的核苷酸序列同源性为99.38%~99.90%;1个养殖场检测出ALV-J,其env基因推导的氨基酸序列与GenBank中ALV-J的氨基酸序列同源性最高为92.63%,并且在该场同一样品中同时检出ALV-E和ALV-J;1个野鸡养殖场检测出ALV-F,其env基因推导的氨基酸序列与已发表的ALV-F的氨基酸同源性为88.69%;同时检测出2株新型的ALV,其env基因推导的氨基酸序列与已发表的A、B、C、D、E、F、J和K亚群的氨基酸同源性为44.68%~58.42%。结果表明,ALV-E在湖南省禽养殖场普遍存在,并首次在我国养殖野鸡中检测出ALV-F,而且在野鸡中发现了很可能是ALV新亚群的2株病毒。本试验为ALV在湖南禽类养殖场的流行情况研究提供了依据。

In order to investigate the infection situation of avian leukosis virus(ALV) in poultry farms from Hunan Province,we collected 156 tissue/sera samples from 21 farms in various regions of Hunan Province from January 2018 to January 2019.The samples were first checked by RT-PCR with detection primers specific for a fragment of ALV 5’ untranslated region(UTR),and then the positive samples were reamplified by primers for near full env gene amplification.The PCR products of env gene were cloned and sequenced.The results of the sequence analysis indicated that the subgroup ALV-E was detected in 19 farms,with high env nucleotide sequence identities of 99.38%-99.90% to the known ALV-E sequences from GenBank;a strain classified as ALV-J was detected in one farm,with the highest identity of 92.63% to the ALV-J sequences from GenBank,and moreover ALV-E was also detected in the same sample.Additionally, from a pheasant farm,a strain belonging to ALV-F was identified with amino acid(aa) similarity of 88.69% of ENV to the only ALV-F sequence in GenBank,and two novel env sequences were also identified,with low aa identities of 44.68%-58.42% to the sequences of the known subgroups of A to F,J and K.This study suggested the ALV-E is common in poultry farms of Hunan Province,and the ALV-F strain was first identified in China,and two novel ALV strains may be classified as new ALV subgroup.The present investigation provided for the first time the preliminary epidemiologic data of the ALV subgroups in Hunan Province.