两株田间重组猪繁殖与呼吸综合征病毒的分离鉴定、进化分析及其致病性研究  

Isolation,Identification and Evolution Analysis of Two Recombinant PRRSV Strains in Fields and Their Pathogenicity to Piglets

作　　者：王新港[1] 王傲杰 周峰[1] 崔丹丹[1] 常洪涛[1] 陈陆[1] 王川庆[1] WANG Xingang;WANG Aojie;ZHOU Feng;CUI Dandan;CHANG Hongtao;CHEN Lu;WANG Chuanqing(College of Animal Husbandry and Veterinary Science, Henan Agricultural University, Zhengzhou 450002, Chin)

机构地区：[1]河南农业大学牧医工程学院

出　　处：《病毒学报》2018年第3期362-371,共10页Chinese Journal of Virology

基　　金：河南省高校科技创新团队与支持计划资助项目(项目号:14IRTSHN015),题目:动物疫病防控新技术~~

摘　　要：为了解2型猪繁殖与呼吸综合征病毒（PRRSV2）田间毒株重组特征,本研究对两株分离物（HENXX-8、HENJY-2）进行全基因测序、进化分析及毒力测定,并对其全基因序列进行重组分析。结果显示,两个毒株均属于PRRSV 2,两者与其代表株VR-2332的核苷酸相似性分别为85.6%、85.7%;与高致病性PRRSV（HPPRRSV）毒株JXA1、TJ和WUH4株分别为85.7%-85.8%、85.4%-85.5%;与经典毒株CH-1a、HB-2（sh）分别为84.7%-85.7%、84.5%-85.7%;而与所有NADC30类毒株均在90.0%以上。全基因、ORF5及Nsp2序列进化分析结果均显示两个分离物与国内报道的类NADC30毒株遗传距离较近,同处于一个分支。全基因组重组分析结果表明,两个分离毒株存在明显重组现象,且重组模式均以类NADC30毒株为骨架病毒,与HP-PRRSV毒株发生重组。但重组对象不同：HENJY-2是由类NADC30毒株与TJ株发生重组;而HENXX-8则是由类NADC30毒株与WUH4、TJ株发生3毒株间重组。由于重组部位序列缺乏特异性分子标志,因此无法确定与类NADC30发生重组的是HP-PRRSV还是其减毒的疫苗毒株。两株分离物的重组部位与早期分离毒株HENANHEB、JL580等有所不同,均未涉及到ORF5基因,而是集中在Nsp1-Nsp2以及ORF2a-ORF3区域,主要发生在病毒基因组的靠5#端（30-7 000bp）和3#端（11 000-13 000bp）处。对部分重要基因的核苷酸相似性分析结果显示,两个分离株的ORF2a、ORF3基因与HP-PRRSV毒株相似性最高,表明重组病毒的这两个基因可能来自HPPRRSV毒株。致病性试验结果表明,参考毒株HENXC-4的毒力略高于分离毒株HENXX-8,主要表现在体温升高、日均增重降低和肺部病变上。但两个分离物均未引起发病猪死亡。以上结果表明,目前PRRSV2在田间的基因重组事件存在随机性,提示不同毒株在田间的存在会加剧PRRSV重组事件的发生和流行、毒株类型更加复杂,从而增加了临床防控难度。因此,慎重使用活疫苗�In order to understand the recombination characteristics of field porcine reproductive and respiratory syndrome virus（PRRSV）,the whole genome of PRRSV isolates（HENXX-8 and HENJY-2）was sequenced and analyzed for genetic evolution,virulence and recombination.The result showed that HENXX-8 and HENJY-2 belonged to the North American genotype（species PRRSV2）,and shared 85.6% and85.7% nucleotide identity with the reference strain VR-2332,respectively.Compared to the Chinese strains,HENXX-8 and HENJY-2 shared 85.7%-85.8% and 85.4%-85.5% nucleotide identities with the highly pathogenic PRRSV（HP-PRRSV）strains JXA1,TJ and WUH4,and 84.7%-85.7% and84.5%-85.7% with the Chinese classical PRRSV strains CH-1 aand HB-2（sh）,respectively,while90.0% with all reference NADC30-like strains.Complete genomic sequence alignment and phylogenetic analysis revealed that HENXX-8 and HENJY-2 are most closely related to the NADC30-like strains recently reported in China and they were in the same subgroup.Recombination analysis indicated that the two isolates had a similar recombination model with NADC30-like strain as skeleton virus recombined with ORF2 a and ORF3 genes of HP-PRRSV.The major difference between two recombinants is that HENJY-8 was recombined by two strains NADC30-like and TJ,while HENXX-8 was recombined by three strains NADC30-like,WHU4 and TJ.Since the sequence at the recombination site lacking specific molecular markers,so it was difficult to distinguish whether ORF2 aor ORF3 genes in two recombinants were from HP-PRRSV or its attenuated live vaccine strain.The recombination sites of the two isolates were different from those of the earlier isolates HENAN-HEB and JL580,etc,and were focused in the Nsp1-Nsp2 and ORF2 a-ORF3 regions mainly occurred in the 5′（30-7,000 bp）and 3′（11,000-13,000 bp）of the viral genome.Based on the nucleotide similarity analysis of some important genes ORF2 aand ORF3 of PRRSV,the two isolates had the highest similarity with HP-PRRSV,which indicated that the recombinant

关 键 词：猪繁殖与呼吸综合征病毒(PRRSV) 病毒分离鉴定 NADC30 HP-PRRSV 全序列 遗传进化分析 重组分析 致病性 

分 类 号：S852.65[农业科学—基础兽医学]