贵州省地方品种鸡禽白血病病毒流行株的亚型鉴定及其分子特征分析

摘要

为了解贵州省9个地方品种鸡群中禽白血病病毒(ALV)流行株亚型与分子特征,本研究2016年从该省8个地区9个地方品种鸡群共采集380份鸡蛋样品.采集卵白用ELISA试剂盒检测ALV-P27抗原,阳性卵白接种CEF和DF1细胞分离病毒,培养7d后检测细胞培养液中ALV-P27抗原,对感染细胞cDNA以4对ALV特异性引物进行PCR鉴定与分型.并选取9个鸡群的部分ALV分离株进行gp85基因的克隆测序与序列分析.结果表明,9个鸡群的卵白样品ALV-P27抗原阳性率为0～36％;阳性卵白的CEF和DF1细胞培养上清P27抗原检测结果相同,阳性率为26.8％(15/56),并鉴定出其亚型全部为J亚型(ALV-J).已测序的21株ALV-J gp85基因序列与国内外ALV-J参考株同源性为86.6％～100％;进化树分析显示,21株ALV-J分离株在不同的分支,其进化关系和来源之间未发现明显规律.本研究表明,贵州省9个地方品种鸡群有6个存在ALV-J感染.其中,3个当地培育的品种鸡ALV感染率为0;而来自3个地区的绿壳蛋鸡ALV-P27抗原阳性率普遍较高(26.7％～36.0％).本研究补充了我国禽白血病流行病学的数据并为该病的防制提供了调查依据.%To investigate the subtype of prevalent avian leukosis virus (ALV) and characterize the molecular of ALV in the loacal chicken flocks from Guizhou province,a total of 380 egg samples were collected from nine local chicken flocks in eight areas of Guizhou in 2016.ALV-P27 was detected by ELISA kit from the albumen,and the CEF and DF1 cells were inoculated with the positive albumen to isolate the virus.The gp85 genes of some ALV isolates from nine chicken flocks were cloned and sequenced.The result showed that the positive rates of ALV-P27 in nine chicken flocks were from 0 to 36％;All the ALV isolates were identified to be J subtypes (ALV-J).Sequencing analysis revealed that the gp85 gene of the 21 isolates of ALV-J shared 86.6％ to 100％ identity with reference strains,and the 21 isolates of ALV-J were classified in different branches in the evolutionary tree.This study showed that ALV-J was detected from six of nine local chicken flocks in Guizhou,and the infection rates was 0 from three local chicken breeds.However,the positive rates of ALV-P27 was generally higher (from 26.7％ to 36.0％) in Lvkedan chickens in three areas.These results provide epidemiological data for the ALV prevention and control.