12株猪瘟病毒E2基因主要抗原区域的序列差异分析

用RTPCR扩增了12个不同时期分离的HCV毒株E2基因主要抗原区域的cDNA片段并对其进行了序列测定。应用DNAstar序列分析软件对所测的12个HCV毒株与国内外已知的6个毒株Alfort株、Ald株、Brescia株、Gpe株、C株、CW株及早期已测定的HCLV株、HCVSM株和北京顺义株(BJSY2/96)3个毒株的相应片段进行了同源性比较分析。E2基因主要区域长度均为224 bp,包括从HCV 2485到2708位的E2基因B、C区域。所测的疫苗株HCLV与国外测得的疫苗株C株核苷酸及氨基酸同源性分别为991%和100%，表明目前应用的疫苗株是稳定的；用目前我国流行的部分野毒株对HCLV株免疫猪的攻击试验表明，HCLV对野毒株均具有很好的免疫力，这与序列分析结果相吻合。根据系统树分析，可将HCV分为两大群，5株90年代的野毒株及1株80年代的野毒株(其中北京3株、河南2株、广东1株)均与国内外C株、标准株属同一群(即第一群)，其核苷酸及氨基酸的同源性分别为857%～100%和838%～100%；与Alfort株同属第二群的有6个野毒株(广西北海、辽宁、河北黄骅、吉林、深圳光明、四川成都)，其中80年代与90年代的野毒株各有三株，核苷酸及氨基酸的同源性分别为843%～100%和851%～100%；21株HCV的核苷酸及氨基酸的同源性分别为781%～100%和784%～100%。两群之间的特征性差异表现在713和729位氨基酸位点的不同，经分析发现猪瘟野毒株具有复杂性与多样性。

NUCLEOTIDE SEQUENCE ANALYSIS OF E2 MAJOR PROTECTIVE ANTIGEN ENCODING REGION OF 12 STRAINS OF HOG CHOLERA VIRUS(HCV)

cDNA fragments, of HCV envelope glycoprotein E2 major gene of 11 field strains isolated in China in different time and 1 French reference strain(Thiveral) were amplified respectively with RT-PCR method and sequenced. The fragments amplified located by the 5' 2485 to 2708 of E2 major domains B and C and encoded 75 amino acid residues of E2 glycoprotein. All the products amplifield by RT-PCR from 12 strains in the study were same size of 224 bp. Comparing 12 sequences with other 9 references strains sequence reported before using software DNAstar, it was found that Hog Cholera Virus could be classified in two groups by analysis of phylogenetic tree. Strains Brescia, Gpe, Ald, Thiverval, C, CW, HCLV, HCVSM, BJCY1/96, BJTX3/96, BJSY2/96, HeNXH2/98, HeNZZ1/82 and GDGZ1/95 were assigned to group A and were 85.7%-100% for nucleotide sequence and 83.8%-100% for amino acid sequence in homology; but strains HCVF98, HCVF94, HeBHH2/95, LN1/84, SZGM1/85, SCCD1/79 and Alfort were assigned to group B and were 84.3%-100% for nucleotide sequence and 85.1%-100% for amino acid sequence in homology; and 21 strains of HCV were 78.1%-100% for nucleotide sequence and for 78.4%-100% amino acid sequence in homology. Homology were 99.1% for nucleotide sequence and 100% for amino acid sequence between strains HCLV in our study and strains C reported by Rijn's in the Netherlands. It's showed our method of sequencing is reliable. There were obvious differences between the two groups in sequences of envelope glycoprotein E2 major gene, especially in the amino acid substitutions of sites 713 and 729 respectively, and it is showed the two groups of HCV field strains might vary genetically in some extents. The results of other report of challenging of the partial field strains showed that the Chinese stock vaccine virus(HCLV) has good immunity.