石鲽鱼鱼苗中一种弹状病毒的分离与鉴定  

The isolation and characterization of a rhabdovirus from stone flounder,Kareius bicoloratus

作　　者：孙颖杰[1,2] 江育林[3] 刘荭[3] 高隆英[3] 史秀杰 何俊强[3] 王哲[1] 

机构地区：[1]吉林大学畜牧兽医学院,吉林长春130062 [2]辽宁出入境检验检疫局,辽宁大连116001 [3]深圳出入境检验检疫局,广东深圳518001

出　　处：《中国兽医学报》2009年第3期277-282,共6页Chinese Journal of Veterinary Science

摘　　要：从养殖石鲽鱼(Kareius bicoloratus)鱼苗中分离到1株弹状病毒(080113株)。用研磨离心除菌后的组织匀浆液,接种11种鱼类细胞系。结果在鲤上皮瘤细胞(EPC)和肥头鲤细胞系(FHM)等8种细胞中均出现明显的细胞病变(Cytopathic effect,CPE),在鲤上皮瘤细胞(EPC)中的滴度最高。对该毒株的理化性质进行了测定,结果显示080113株对温度和pH敏感;对5′-碘-2′脱氧尿密啶(I UDR)不敏感;对氯仿敏感,提示是一种有囊膜的RNA病毒。制备电镜超薄切片,观察到在细胞质中有大小为(80～118)nm×(28～55)nm、形态为典型的弹状的病毒粒子。SDS-PAGE电泳分析该毒株的蛋白质图谱,结果表明该毒株有5条主要的蛋白带,相对分子质量大小分别为200 000、56 000、42 000、29 000和24 000。用传染性造血器官坏死病毒(I HNV)、病毒性出血性败血症病毒(VHSV)和牙鲆弹状病毒(HRV)等水生动物弹状病毒特异性引物对该毒株进行逆转录聚合酶链式反应,结果用HRV的引物能扩增出阳性片段。对该片段进行测序分析,发现与HRV的参考序列有99%以上的相似性。因此,初步鉴定该毒株为牙鲆弹状病毒。A strain of rhabdovirus （strain 080113） was isolated from the larvae of cultured stone flounder Kareius bisoloratus. The obvious cytopathic effect （CPE） was observed in 8 cell lines inoculated with tissue homogenates,with the highest titer in Epithelioma papulossum of carp cell line （EPC）. The characterization of strain 080113 was studied. The strain was sensitive to temperature, pH3 and pH10, chloroform and not sensitive to 5-iodo-2-deoxyuridine （IUDR）, which suggested that the strain was RNA virus with envelope. Typical bullet-shape particles, with the size of （80 118） nm × （28 55） nm,were observed in the cytoplasm of EPC cells infected by strain 080113. Five protein bands were observed by SDS-PAGE,with the molecular weight of 200 000,56 000,42 000,29 000 and 24 000, respectively. Reverse transcription polymerase chain reaction （RT-PCR） was tested with the specific primers of infectious hematopoietic necrosis virus （IHNV）,viral haemorrhagic septicemia virus （VHSV）,hirame rhabdovirus （HRV） and other aquatic animal rhabdovirus. The fragment was amplified with the primers of HRV and sequenced. More than 990/00 homology was shared with the reference strain of HRV. It was deduced that the strain 080113 is HRV. The etiology and the pathogenicity will be carried in the future.

关 键 词：石鲽鱼 牙鲆弹状病毒 分离 鉴定 

分 类 号：S852.65[农业科学—基础兽医学]