基于N基因的小反刍兽疫病毒贵州流行株分子特征与分群研究

Molecular Characteristics and Cluster of PPRV Guizhou Strains Based on N Gene

为探讨小反刍兽疫病毒(peste des petits ruminants virus,PPRV)贵州流行株N基因分子特征和分群,试验设计了1对特异性引物,应用RT-PCR技术对小反刍兽疫(peste des petits ruminants,PPR)临床样本进行N基因扩增,克隆至pMD19-T载体,对阳性重组质粒进行测序,应用DANStar软件对测序序列和参考序列进行核苷酸同源性、氨基酸同源性、变异位点及系统进化树分析。结果显示:PPRV贵州流行株N基因扩增长度为1 578bp,其相互间核苷酸、氨基酸同源性分别为99.6%~100.0%及99.2%~100.0%,与国内参考株N基因的核苷酸序列(97.7%~99.9%)及氨基酸序列(98.3%~100.0%)同源性较国外参考株(88.5%~97.7%和92.2%~98.5%)高;PPRV贵州流行株N基因编码的氨基酸同疫苗株Nigeria 75-1相比存在26个位点突变,但没有氨基酸的缺失或增加;基于N基因系统进化分析显示,PPRV贵州流行株同国内参考株处于同一个进化分支,但与国外参考株处于不同进化分支;其属于病毒进化的Ⅳ基因群,与国内参考株处于同一系统分群,但与疫苗株Nigeria 75-1(Ⅰ基因群)处于不同基因群。

In order to investigate the N gene molecular characteristics and clustering of the peste des petits ruminants virus（PPRV）of Guizhou province,the primers of PPRV N gene were designed and the target gene was amplified from clinical samples and then was cloned to pMD19-T vector.The positive recombinant plasmids were sequenced and nucleotide homologoy,amino acids homology,mutation sites and phylogenetic tree analysis of N gene of Guizhou PPRV strains and reference strains were performed by DANStar software.The results showed that the length of amplified N gene was 1 578 bp and the homology of nucleotide and amino acid sequences among Guizhou PPRV strains were 99.6%to 100.0%and 99.2%to 100.0%,respectively.The homolo-gy of nucleotide（97.7%to 99.9%）and amino acids（98.3%to 100.0%）compared with domestic reference strains were higher than those of other countries（88.5% to 97.7% and 92.2% to98.5%）.Compared with the vaccine strain Nigeria 75-1,there were 26 multiple sites in Ngene of Guizhou PPRV strains while there was no amino acid loss or increase.The results of phylogenetic analysis showed that Guizhou PPRV strains were in the same evolutionary branch with the domestic reference strains,while the foreign reference strains in a different evolutionary branch.Guizhou PPRV strains belonged toⅣ gene group and vaccine strain Nigeria 75-1 belonged to Ⅰgene group.