2013年中国典型对虾养殖区白斑综合征病毒流行株高变异区序列的分析比较  

Variation sequence comparison of shrimp WSSV from different parts of China in 2013

作　　者：孙新颖[1,2] 万晓媛[1] 刘庆慧[1,3] 黄倢[1,3] 

机构地区：[1]农业部海洋渔业可持续发展重点实验室,中国水产科学研究院黄海水产研究所,山东青岛266071 [2]上海海洋大学,上海201306 [3]青岛海洋科学与技术国家实验室,山东青岛266071

出　　处：《中国水产科学》2016年第3期693-703,共11页Journal of Fishery Sciences of China

基　　金：国家重点基础研究发展计划项目(2012CB114401);山东省“泰山学者”建设工程专项经费;农业部科研杰出人才和创新团队专项

摘　　要：为了解中国不同地区白斑综合征病毒的流行变异情况,本研究取用2013年3—12月从7个省市发病地区采集到的64份WSSV阳性样本,以特异的引物扩增目的片段,通过测序分析不同样本的缺失及变异差异。结果显示,在开放阅读框ORF14/15的扩增中,分别有6530 bp、6533 bp和5138 bp的片段缺失,而在ORF23/24扩增中有12070bp大片段的缺失,不同地区样本中未能成功扩增ORF75,而ORF94的重复单元数目分别为0、3、4、12不等,ORF125的重复单元数目为0、7。SNP分析表明,含有3个重复单元的ORF94在48位的碱基为T、T、T,重复单元数为4的在48位的碱基为T、T、T、T,重复单元数为12的在48位的碱基分别为T及11个A。而ORF125所有重复单元数为7的情况在8、18、25、66、69位置的碱基均为G、G、G、G、A,在9、50、53、61位的碱基也普遍出现了变异。结果表明,WSSV在中国不同地区存在一定程度的变异,其在序列中的缺失、重复单元数目以及SNP的差异较为明显。White spot syndrome virus （WSSV） is highly pathogenic to penaeid shrimp and has caused significant economic losses in the aquaculture industry worldwide. With the constant expansion of the areas and scale of prawn fann ing, variations between different strains of WSSV have emerged. The differences between isolates may cause variations in the virulence of WSS. Deletions in the ORF14/15 and ORF23/24 variable regions have been used as molecular markers to study the epidemiology of the virus. The repeat units of ORF75, ORF94, and ORF125 vary in different isolates and the numbers of repeat units in these three genomic regions are a useful marker for epidemiological studies and for genotyping strains of WSSV. However, each repeat unit may contain single-nucleotide polymorphisms （SNPs） that are important for differentiating haplotypes with the same numbers of repeat units. The results of epidemiological studies may differ if the variability provided by these SNPs is not considered and only the number of repeat units is taken into account. To understand the variations in WSSV ORF14/15 and ORF23/24 and the variable number of tandem repeats （VNTR） and SNPs located in ORF75, ORF94, and ORF125 in different regions of China, we investigated 64 samples that were WSSV-positive on PCR, collected in a disease outbreak area between March and December 2013. The tested samples were from Hainan, Guangdong, Guangxi, Zhejiang, Shandong, Hebei, and Tianjin. Using specific primers, WSSV-positive samples were genotyped with PCR and the amplified fragments were ligated to a T-vector and used to transform Topl0 cells. The positive clones were selected and sequenced. After sequencing, the fragments deleted from ORF14/15 and ORF23/24 in different samples were compared with the sequences of Th-96-II and TW respectively, and the three WSSV VNTR regions and SNPs in ORF75, ORF94, and ORF125 from different samples were analyzed with the DNAMAN software. The results showed that there were five samples which had the products of ORF 14/1

关 键 词：WSSV ORF 缺失 变异 PCR 

分 类 号：S945[农业科学—水产养殖]