鳗鲡疱疹病毒的分离与鉴定

为获知鳗鲡"脱粘败血病"与病毒的关系,实验用蔗糖密度梯度离心的方法从发病的欧洲鳗鲡内脏器官组织匀浆液中纯化了病毒粒子,负染后利用电镜观察;进一步用EO细胞对病毒进行了分离、培养,并对感染病毒的细胞进行超薄切片,电镜观察;然后,提取病毒DNA,利用鳗鲡疱疹病毒的PCR检测方法对其进行了鉴定。结果显示,接种匀浆上清液的EO细胞出现细胞融合的病变效应;分离病毒的病毒粒子具囊膜,大小约为200 nm;从感染病毒的细胞上清液DNA中扩增出特异性条带,序列测定与比对分析表明,该序列与鳗鲡疱疹病毒欧洲株(AngHV-1)的序列完全一致。研究表明,利用EO细胞分离了一株鳗鲡病毒,经形态观察和DNA分析,确认该病毒为鳗鲡疱疹病毒,命名为AngHV-FJ。该研究为深入开展鳗鲡疱疹病毒的致病机制及鳗鲡"脱粘败血病"的防控研究奠定了重要基础。

Isolation and identification of a herpesvirus from cultured European eels Anguilla anguilla in China

To clarify whether the "mucus sloughing and hemorrhagic septicemia disease" of cultured European eels,Anguilla anguilla,is related with virus,the internal organs of the diseased European eel were extracted and homogenized,and virions were purified by sucrose density gradient centrifugation;then EO cells were incubated with the supernatant of the tissue homogenate,and the cytopathic effects of cell fusion appeared.In addition,the virus infected cells were thin sectioned,also the negatively stained purified virions were observed by electron microscope to photograph the virus morphology.It showed that the virus was enveloped with size in about 200 nm.Furthermore,DNA were extracted from the supernatant of the infected cells,and used for Anguillid herpesvirus(AngHV)PCR detection,and then the amplified fragment was sequenced and alignment analyzed.The results showed that it was exactly the same sequence with the AngHV European strain,AngHV-1.Therefore,the observation by electron microscope and DNA polymerase gene sequencing confirmed that the isolated virus is an AngHV strain,named AngHV-FJ,which is the first cell isolation in China mainland.The research has laid an important foundation for the study of the pathogenic mechanism of AngHV-FJ and the prevention and control of eels "mucus sloughing and hemorrhagic septicemia disease".