凡纳滨对虾肠道上皮细胞微胞子虫PCR检测方法的建立与应用Development and Application of a PCR Detection Assay of Microsporidium Enterocytozoon hepatopenaei in Pacific White Leg Shrimp Litopenaeus vannamei
雷燕,肖洋,张会军,王娟,张文文,卢刚,王雪鹏
摘要(Abstract)：
根据Gen Bank中对虾肠道上皮细胞微胞子虫(Enterocytozoon hepatopenaei,EHP)的基因保守序列,设计一对特异性引物,通过优化PCR扩增条件,建立快速检测凡纳滨对虾(Litopenaeus vannamei)EHP的PCR方法。用该方法对EHP阳性虾进行PCR扩增,得到与实验设计相符的330 bp特异性扩增条带,而对白斑综合症病毒(WSSV)、桃拉病毒(TSV)、传染性皮下及造血器官坏死病毒(IHHNV)、对虾杆状病毒(BP)、"棉花虾"微孢子虫、黏孢子虫的阳性虾,以及健康虾的扩增结果为阴性。测序比对发现,PCR产物序列与Gen Bank EHP基因序列的同源性为99.8%,表明该PCR方法检测结果准确。敏感性试验表明,该方法最低可检测出约100 fg的EHP质粒DNA。用该PCR方法检测广东、广西、江苏、山东、海南等地的755份临床样品,共检出阳性样品21份。该PCR方法可用于凡纳滨对虾EHP的快速检测。
关键词(KeyWords)： 凡纳滨对虾;肠道上皮细胞微胞子虫;PCR检测;临床应用
基金项目(Foundation): 山东省现代农业产业技术体系(SDAIT-19);; 泰安市科技发展计划项目(201440774);; 浙江省近岸水域生物资源开发与保护重点实验室开放基金项目(J2013006)
作者(Author): 雷燕,肖洋,张会军,王娟,张文文,卢刚,王雪鹏
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