鸭细小病毒04Nb株的分离鉴定与rep基因测序与分析  

Isolation and Identification of Duck Parvovirus 04Nb Strain and Sequence Analysis of rep Gene

作　　者：张彦鹏[1] 李静[1] 寇铮[1] 陈绳亮[1] 范兆军[1] 张忠[1] 李天宪[1] 

机构地区：[1]中国科学院武汉病毒研究所病毒学国家重点实验室,湖北武汉430071

出　　处：《中国病毒学》2006年第2期173-177,共5页Virologica Sinica

基　　金：国家863计划项目(2005AA219070)

摘　　要：采集浙江宁波地区以腹泻、呼吸困难为主要症状的病鸭肝组织,接种正常鸭胚尿囊腔增殖病毒。雏鸭感染试验显示发病症状及病理变化明显,死亡率为75%。电镜下可见纯化病毒直径约20nm左右的球形病毒粒子。免疫琼脂扩散实验结果显示与鸭细小病毒(duckparvovirusDPV)标准株阳性血清有明显沉淀线。经SDS-PAGE呈现3条结构蛋白带,与DPV标准株一致;参照GenBankDPV非结构蛋白基因序列设计引物,PCR扩增反应获得目的条带,克隆测序后,与DPV代表株序列同源性达98%。根据上述实验结果,确定引起本次鸭场疫病的病原为DPV。为进一步研究该分离株rep基因的序列特征,对其rep基因克隆测序,与GenBank中两株DPV、两株鹅细小病毒(GPV)进行序列比对,结果显示rep基因核苷酸序列与DPV参考毒株同源性为98%以上,与GPV同源性为80%左右。A severe contagious disease broke out in several duckeries in Ningbo, Zhejiang province with the symptoms of diarrhea and grasping. The liver tissues from the dead ducks were collected and inoculated into the allantoic cavity of the 11-day-old embryos. Through purification by sucrose density gradient centrifugation, a diameter of 20nm virions with typical duck parvovirus （DPV） characters was observed by electron microscopy. In the test of agar-gel immunodiffusion, the reaction between the stuff from dead embryos and positive serum was obvious. Through SDS-PAGE, three main segments were visible which coincided with DPV. In animal test, similar symptoms were also observed. Using the primers designed according to DPV genome, we acquired an anticipative 600bp DNA fragment, which was cloned into E.coli and sequenced. The sequence shared 98% identity with the representative DPV strain. From all above results, the pathologic agent was diagnosed as Duck Parvovirus. To acquire more information of the rep gene, we sequenced and aligned it with 4 other sequences of DPV or goose parvovirus （GPV） from GenBank. The result indicated the homology of rep gene was about 98% with DPV and 81% with GPV.

关 键 词：鸭细小病毒 分离鉴定 rep基因 同源性 

分 类 号：S852.65[农业科学—基础兽医学]