豫陕两省14株鸡杆菌的gyrB、16S rRNA和rpoB基因序列比较分析

Sequence Analysis of Fourteen Gallibacterium Isolates from Henan and Shanxi Provinces Based on gyrB,16S rRNA and rpoB Genes

分析河南、陕西分离的14株鸡杆菌(Gallibacterium)之间的进化关系,及gyrB基因序列比较在该菌进化分析中的作用。PCR扩增鸡杆菌分离株的gyrB、16SrRNA和rpoB3个看家基因,PCR产物纯化后直接测序。将鸡杆菌分离株、国外参考株的3个看家基因序列进行比较分析,用Phylip 3.67软件构建进化树。结果表明,14株鸡杆菌与鸭源鸡杆菌(Gallibacterium anatis)模式株间的相似性为96.3%～98.0%(gyrB)、97.7%～99.6%(16SrRNA)和97.7%～99.0%(rpoB);14株鸡杆菌与鸡杆菌复合群1(Gallibacterium genomosp.1)参考株间的相似性为88.8%～89.9%(gyrB)、96.2%～97.5%(16SrRNA)和92.6%～93.6%(rpoB)。基于3个看家基因序列的进化分析,均显示14株鸡杆菌和鸭源鸡杆菌模式株形成单独的一个群。14株鸡杆菌分离株均属于鸭源鸡杆菌种;在3个看家基因位点,鸡杆菌河南株与陕西株之间、鸡杆菌输卵管炎病鸡分离株与健康鸡分离株之间均无明显遗传上的差异;gyrB基因序列分析可用于鸡杆菌分离株的种类鉴定,且对14株鸡杆菌与复合群1参考株的区别能力优于另外2个看家基因。

The objective of this study was to analyze the evolutionary relationships of the fourteen Gallibacterium strains isolated from Henan and Shanxi Provinces,and the role of gyrB gene in phylogenesis analysis.16S rRNA,rpoB and gyrB gene of all isolates were amplified by PCR and the purified PCR product were submitted directly for sequencing.Sequences of three housekeeping genes of 14 Gallibacterium isolates were compared,and used to generate the phylogenetic tree,with that of the Gallibacterium reference strains,using the software of DNAstar,Clustalx 1.81 and Phylip 3.67.The sequence homology were 96.3%-98.0%（gyrB）,97.7%-99.6%（16S rRNA）,97.7%-99.0%（rpoB） among Gallibacterium isolates and G.anatis type strains,88.8%-89.9%（gyrB）,96.2%-97.5%（16S rRNA）,92.6%-93.6%（rpoB） among Gallibacterium isolates and Gallibacterium genomosp.1 strains.Three phylogenetic trees generated from housekeeping genes showed that 14 Gallibacterium isolates fell into the same branch with G.anatis type strains.14 Gallibacterium isolates were classified in G.anatis.In the three gene locus of gyrB,rpoB and 16S rRNA genes,there were no apparent genetic difference between Gallibacterium strains isolated from Henan and Shanxi Provinces,and from healthy chickens and diseased chickens with salpingitis.gyrB gene could be used for classification of Gallibacterium isolates,furthermore,more obvious differences was noticed in the homology analysis based on gyrB gene sequence between Gallibacterium isolates and G.genomosp.1 than that based on 16S rRNA and rpoB genes.