鸡源鸭疫里默杆菌的分离鉴定及ompA基因遗传进化分析

从病死鸡肝脏中分离到8株革兰阴性多形杆菌,对8株分离株进行形态学和16S rDNA基因鉴定并分析ompA基因及其推导的蛋白序列遗传进化特征。16S rDNA进化分析显示,8株鸡源分离株与22株鸭疫里默杆菌(Riemerella anatipestifer,RA)参考株形成一个大的进化分支,与RA模式菌株ATCC11845和22株RA参考株之间的同源性分别为99.3%~99.8%和98.8%~100.0%。根据形态学和16S rDNA基因鉴定结果,将8株分离株鉴定为RA。ompA基因进化分析显示,8株鸡源RA分离株与3株鸭源RA分离株KML1、KML2、KML3以及1株鹅源RA分离株KS9901-G形成一个大的进化分支,同源性为94.0%~99.6%。与ATCC11845株相比较,8株鸡源RA分离株ompA基因均发生99个核苷酸位点的突变。OmpA蛋白进化分析显示,8株鸡源RA分离株形成一个独立的大的进化分支,同源性为100%。与ATCC11845株相比较,8株鸡源RA分离株OmpA蛋白均发生19个氨基酸位点的突变。8株鸡源RA分离株对青霉素、阿莫西林克拉维酸、头孢噻吩等10种抗生物敏感。本试验在国内分离到鸡源RA,证实鸡RA感染在我国的存在,对鸡RA感染的预防和控制具有重要指导意义。

Isolation,identification and phylogenetic analysis of ompA gene of Riemerella anatipestifer in chicken

To bacteriologically diagnose a suspected chicken Riemerella anatipestifer(RA)infection,eight strains of Gram-negative pleomorphic rod were isolated from the livers of dead chicken,and the morphological identification and 16 S rDNA analyses were performed.Furthermore,phylogenetic characteristics of ompA gene and its deduced protein sequence were studied.Phylogenetic analysis of the 16 S rDNA indicated that the eight strains formed a large clade with 22(RA)reference strains,and shared sequence similarity of 99.3%-99.8%and 98.8%-100.0%with the RA type strain ATCC11845 and 22 RA reference strains,respectively.Based on the results of the morphological identification and the 16 S rDNA identification,these eight strains were identified as RA.Phylogenetic analysis of the ompA gene indicated that they formed a large clade with three duck isolates and one goose isolate,and they shared sequence similarity of 94.0%-99.6%.Compared with the strain ATCC11845,the ompA gene of them has 99 nucleotide mutation sites.Phylogenetic analysis of the OmpA protein indicated that these strains formed independent clade and shared sequence similarity of 100%.Compared with the strain ATCC11845,the OmpA protein of these isolates have 19 amino acid mutation sites.These strains were sensitive to 10 antibiological agentssuch as penicillin,amoxicillin-clavulanic acid and cefalotin.To our knowledge,this is the first isolation of RA in chicken in China and it confirms the presence of RA infection in chicken.These results have important guiding significance for the prevention and control of RA infection in chicken.