2006-2008年山东鸡传染性支气管炎病毒分离株S1与N基因的分子特征

Molecular Characterization of Avian Infectious Bronchitis Coronavirus Strains Isolated in Shandong between 2006 and 2008

为了调查2006-2008年山东省鸡传染性支气管炎的分子流行病学特征,应用RT-PCR方法分别扩增了14株传染性支气管炎病毒地方毒株S1基因和N基因,并进行了基因克隆与测序工作。对此14株病毒的S1基因序列利用限制性内切酶HaeⅢ进行了限制性片段长度多态性（RFLP）分析,结果显示可以将IBV毒株分为3种不同的基因型。其中11个毒株与LX4毒株有相同的RFLP分型,2株病毒属于Mass基因型,1个毒株有特异的RFLP分型。并将这些分离毒株与11个参考毒株分别进行了基因与氨基酸系统进化树关系分析。S1基因分析结果显示将这些毒株分成3个基因型,与RFLP分析的分型结果一致。11株病毒同属于LX4类型的毒株,分离毒株核苷酸序列相互之间有95.4%~99.7%的同源性。基因Ⅱ型与疫苗株H120和M41的同源性很高。属于基因Ⅲ型的1个毒株形成了独特的基因型。N基因分析结果表明：12株病毒与LX4属于同一基因型,有着较高的同源性。另2株病毒与疫苗株H120同源性很高。以上分析结果表明：山东省IBV分离株存在较为广泛的基因突变、插入和缺失,同时,IBV毒株间的基因重组也可能是山东省IBV变异株产生的另一主要原因。

This experiment was conducted to study characteristics of molecular epidemiology of avian infectious bronchitis virus （IBV） in Shandong province of China between 2006 and 2008. The complete S1 gene and N gene of fourteen IBV strains isolated in Shandong were amplified using RT-PCR method. Then they were sequenced and S1 gene was analyzed by restriction frag- ment length polymorphism （RFLP）. Results indicated that these Shandong IBV isolates were classified into three groups according to their RFLP patterns obtained using the restriction en- zyme HaeⅢ. Eleven of fourteen isolates were similar to the LX4 RFLP pattern, which is a com- mon pattern in China. The RFLP patterns for 2 of 14 isolates corresponded to the patterns of IBV Massachusetts strains. One isolate showed variant RFLP pattern. They were characterized by comparison with eleven reference strains. The result of S1 gene analysis demonstrates that Shan- dong IBVs, which were mainly nephropathogenic, were divided into three genotypes. The results of S1 glycoprotein genes analysis and RFLP analysis were similar. Eleven field viruses clustered together with LX4-type strains, having 95.4%--99.7% nucleotide identity within the group, and two isolates had very close sequence relationship with Massachusetts strains. One isolate formed novel genotype in Shandong. Phylogenic analysis of N gene sequences revealed that twelve field viruses clustered together with LX4-type strains with high identity and two isolates had very close sequence relationship with Massachusetts strains. These findings suggest that not only point mu- tations, insertions and deletions but also a recombination events having contributed to the genetic diversity and emergency of IBV variants in Shandong.