一例PPRV阳性羊鼻拭子样本N、P、M、F、H基因的序列测定和遗传进化分析

Sequencing and phylogenetic analysis of N, P, M, F, H genes of PPRV in a positive nasal swab sample detected in goat

为对采集自昆明某大型牲畜交易市场的一例PPRV阳性鼻拭子样本进行追根溯源,设计引物采用RT-PCR方法进行PPRV病毒基因组关键基因N、P、M、F、H核苷酸序列扩增,并进行序列测定和遗传进化分析。结果显示:N、P、M、F、H基因均有预期片段扩增,其读码框(ORF)长度分别为1 578、1 531、1 008、1 641及1 830 nt;N、P、M、F及H构建的遗传进化树均显示所测毒株为基因Ⅳ型,同2013年在新疆伊犁山羊中检测到的毒株(KM091959)同源性最高,N、P、M、F、H核苷酸同源性分别为99.2%、99.0%、99.4%、99.6%和99.3%,推断氨基酸同源性分别为98.7%、98.4%、100.0%、99.8%和99.3%。研究表明,检测到的毒株仍为2013~2014年我国PPRV流行期的基因Ⅳ型毒株,变异较小;所测毒株的N、P、M、F、H 5个关键基因中,M基因最保守、其次是F/H基因,而N、P基因略有变异。

In order to trace the source of a PPRV positive nasal swab sample collected in a large livestock market in Kunming, N, P, M, F, H genes of PPRV virus genome were amplified through RT-PCR method with designed primers, and nucleotide sequence and phylogenetic analysis were carried out. The results showed that N, P, M, F, H had expected fragment amplification. The ORF length of N, P, M, F and H genes were 1 578, 1 531, 1 008, 1 641 and 1 830 nt, respectively. The phylogenetic trees constructed based on N, P, M, F and H all showed that the tested strain belonged to genotype Ⅳ, which had the highest homology with the strain(KM091959) detected in goat, Xinjiang Yili, 2013. Nucleotide homology of N, P, M, F, H was 99.2%, 99.0%, 99.4%, 99.6% and 99.3%, respectively, and deduced amino acid homology was 98.7%, 98.4%, 100.0%, 99.8% and 99.3% respectively. We concluded that the virus strain detected in the study was closest with those pandemic during 2013~2014 nation-wide outbreak and variation was minimal. Among the five key genes including N, P, M, F, H, M gene was the most conservative, followed by F and H genes, while N and P genes were slightly genetically variant.