河南地区NADC30-like PRRSV毒株的增殖特性与遗传进化分析

Proliferation Characteristics and Phylogenetic Analysis of One NADC30-like PRRSV Strain Isolated in Henan

为研究河南地区猪繁殖与呼吸综合征病毒(PRRSV)的流行特点和遗传变异情况,采集河南省滑县地区猪繁殖与呼吸综合征(PRRS)发病猪场中的病料,经研磨、稀释、离心处理后将悬液上清接种于原代猪肺泡巨噬细胞,进行病毒分离培养,结果显示,PAM出现典型细胞病变效应,将得到的分离株命名为HNhx。应用PRRSV N蛋白的特异抗体进行间接免疫荧光(IFA)试验,结果表明,接种HNhx分离株的PAM出现特异性荧光,证明该分离株为PRRSV。应用RT-PCR技术对Nsp2区进行扩增分析,根据扩增目的产物大小推测HNhx分离株为NADC30-like毒株。遗传分析发现,与其他参考毒株相比,HNhx毒株的ORF3、ORF4和ORF5基因与NADC30毒株的同源性最高。基于ORF3、ORF4和ORF5序列构建进化树,系统进化分析的结果表明,HNhx归于NADC30-like亚群。

In order to investigate the epidemic characteristics and genetic diversity of PRRSV in Henan province,lungs from diseased pigs showing clinical signs suspected of PRRSV were collected from pig farm in Huaxian area of Henan province. The samples were homogenized,diluted and centrifuged and the obtained supernatants were inoculated into PAM to isolate PRRSV. PAM showed obvious cytopathic effect（CPE）,which revealed that PRRSV strain was isolated and was named HNhx. The result of indirect immunofluorescence（IFA） showed that the PAM infected with HNhx were positive for N protein of PRRSV.The isolated HNhx strain was identified as a NADC30-like PRRSV by RT-PCR using specific primers of NSP2. Genetics analysis showed that ORF3,ORF4 and ORF5 shared highest similarity with NADC30,respectively. Moreover,phylogenetic tree based on ORF3,ORF4 or ORF5 revealed that the PRRSV strainHNhx was clustered into NADC30-like subgroup.