对虾传染性皮下及造血组织坏死病毒(IHHNV)PCR检测方法的建立  

Development of a polymerase chain reaction (PCR) for the detection of infectious hypodermal and hematopoietic necrosis virus

作　　者：杨冰[1] 宋晓玲[1] 黄倢[1] 史成银[1] 刘莉[1] 刘庆慧[1] 宋微波[2] 

机构地区：[1]中国水产科学研究院黄海水产研究所,青岛266071 [2]中国海洋大学海水养殖教育部重点实验室,青岛266003

出　　处：《海洋水产研究》2005年第2期1-5,共5页Marine Fisheries Research

基　　金：国家重点基础研究项目(G1999012002);人事部留学回国人员科技活动资助项目;山东省出入境检验检疫局项目(SK200228 2)共同资助

摘　　要：对虾传染性皮下及造血组织坏死病毒(IHHNV)病是国际兽疫局(OIE)划定的甲壳类其他重要疾病之一,它分布较广,危害严重,对世界对虾养殖业发展影响重大。本文根据Genbank登录的IHHNV基因序列(AF218266),设计了1对特异性引物,从纯化的IHHNV DNA和感染IHHNV凡纳滨对虾(Litopenaeus vannamei)组织DNA中成功地扩增出产物大小为703bp的DNA片段,该对引物对IHHNV DNA的检测灵敏度为19. 85fg(8. 83×103 病毒拷贝),与健康对虾组织DNA、对虾白斑综合征病毒(WSSV) DNA及对虾肝胰腺细小病毒(HPV) DNA无交叉反应。本方法可快速、灵敏、特异地检测出对虾感染和携带IHHNV状况,为对虾健康养殖、无特定病原(SPF)种群选育及流行病调查提供了有效的检测手段。Infectious hypodermal and hematopoietic necrosis virus (IHHNV) is one of the pathogens leading to other significant diseases of crustaceans pointed by the OIE (Office international desépizooties) manual. The PCR primers for the detection of IHHNV were designed from a region of the IHHNV genomic sequence (GenBank AF218266) that encodes for structural protein. Purified IHHNV DNA and nucleic acids extracted from tissue samples of clinically healthy shrimp Litopenaeus vannamei yielded an evident amplification product,showing the expected mobility of a 703bp DNA fragment, thereby confirming the specificity of our primers. The detection sensitivity was 19.85fg testing IHHNV DNA. IHHNV-free shrimp tissue DNA, WSSV DNA and HPV DNA showed no cross-reactions. An effective diagnostic tool is thus provided for detecting IHHNV infections, diagnosing epidemic diseases and breeding specific pathogen free (SPF) shrimp.

关 键 词：传染性皮下及造血组织坏死病毒(IHHNV) 聚合酶链式反应(PCR) 检测 

分 类 号：S945.46[农业科学—水产养殖]