豫南地区猪伪狂犬病病毒流行株的主要毒力基因遗传变异分析

为掌握豫南地区猪群中流行的伪狂犬病病毒（Pseudorabies virus，PRV）分子遗传特征，本研究利用PCR方法从PRV感染猪的组织中扩增其主要毒力基因gB、gE、gC、gD和TK，并进行核苷酸序列测定。利用MegAlign软件进行流行毒株的主要毒力基因与已发表的参考序列的相似性、进化树和关键位点氨基酸变异分析。测序结果表明，本研究成功从5份PRV感染猪组织中扩增出PRV的gB、gE、gC、gD和TK基因。氨基酸相似性和进化树分析结果表明，豫南地区的5株PRV流行株与国内流行毒株在G1群，与G1群国内流行毒株的gB、gE、gC和gD氨基酸相似性分别为98.5%~100%、97.2%~100%、97.5%~100%和98.3%~100%；与G2群亲缘关系较远（以Bartha、Becker、NiA3株为代表的欧美地区流行毒株），与G2群内毒株的gB、gE、gC和gD氨基酸相似性分别为96.4%~97.4%、94.6%~95.7%、92.7%~94.0%和96.3%~99.0%。关键氨基酸变异分析结果表明，与Bartha株（或Becker和NiA3株等）相比，5株流行毒株的gB氨基酸存在75-77位"PGL"的缺失，94位"G"的插入；gE氨基酸存在48和496位有"D"的插入，gC氨基酸存在57-63位"VSGTTGA"的插入和65-69位"SPEAG"突变为"ASTPA"，gD氨基酸存在278-281位"RP"或"RPRP"的插入。此外，流行株的gB、gE、gC、gD和TK氨基酸序列存在多个单位点的氨基酸突变。因此，豫南地区PRV流行株具有PRV变异毒株的分子遗传特征。上述结果证实，5株PRV流行毒株均为变异毒株，与疫苗毒株Bartha-K61株亲缘关系较远。

Genetic Variation Analysis of Major Virulence Genes of Pseudorabies Virus Epidemic Strains in Southern Henan Province

In order to investigate the molecular genetic characteristics of Pseudorabies virus (PRV) in Southern Henan province, the main PRV virulence genes gB, gE, gC, gD and TK from the tissues of PRV-infected pigs were amplified by PCR and sequenced.The similarity, phylogenetic tree and amino acid variation analysis of major virulence genes between epidemic strains and published reference sequences were performed by MegAlign software.Sequencing results showed that the PRV gB, gE, gC, gD and TK genes from 5 PRV-infected pig tissues were successfully amplified.Amino acid similarity analysis and phylogenetic tree results showed that the 5 PRV epidemic strains in Southern Henan province were in G1 group with the domestic epidemic strains, the amino acid similarity of gB, gE, gC and gD were 98.5%-100%, 97.2%-100%, 97.5%-100% and 98.3%-100%, respectively, relatively distantly related to G2 group (the Bartha strain, Becker strain, and NiA3 strain in Europe and America), the amino acid similarity of gB, gE, gC and gD were 96.4%-97.4%, 94.6%-95.7%, 92.7%-94.0% and 96.3%-99.0%, respectively.Compared with the Bartha-K61 strain (or Becker strain, NiA3 strain, etc.), there were amino acids deletion, insertion and mutation in the 5 epidemic strains, such as ‘PGL’ deletion at position 75-77 and ‘G’ insertion at position 94 in gB amino acids, the ‘D’ insertions at positions 48 and 496 in gE amino acid sequence, the ‘VSGTTGA’ insertions at positions 57-63 and the ‘SPEAG’ mutation to ‘ASTPA’ at positions 65-69 in the gC amino acid, the ‘RP’ or ‘RPRP’ insertion at positions 278-281 in gD amino acid.In addition, there were single amino acid mutations in gB, gE, gC, gD and TK amino acid, respectively.Therefore, the epidemic strains of PRV in Southern Henan province showed the molecular genetic characteristics of the variant strains.The above results confirmed that the 5 PRV epidemic strains were variant strains, which were relatively distantly related to the vaccine strain Bartha-K61.