绵羊大肠杆菌与A型产气荚膜梭菌混合感染的病原分离鉴定

Pathogens isolation and identification of co-infection between Escherichia coli and Clostridium perfringens type A in sheep

试验旨在对宁夏一绵羊场疑似大肠杆菌与A型产气荚膜梭菌混合感染的死亡病例进行确诊并提出相应的防治方案。采用产气荚膜梭菌显色培养基及伊红美蓝固体培养基进行病原分离培养,对分离菌株进行16S rRNA基因序列PCR检测,依据16S rRNA序列构建分离株分子进化树;之后对大肠杆菌分离株毒素因子进行PCR检测,对产气荚膜梭菌分离株进行毒素分型鉴定。结果显示,分离株PCR检测获得了16S rRNA特异性条带;分子进化树结果显示,大肠杆菌分离菌株NXDC001与大肠杆菌在同一分支,产气荚膜梭菌分离菌株NXSJ001与产气荚膜梭菌在同一分支。大肠杆菌分离株检测出毒素因子HPI(irp2)及LEE(eae);产气荚膜梭菌分离株携带毒素因子cpa,证明分离株为A型产气荚膜梭菌。研究表明,试验成功分离鉴定大肠杆菌及A型产气荚膜梭菌各一株,证明病死羊为大肠杆菌及A型产气荚膜梭菌混合感染。

The purpose of the study was to diagnose the death of suspected co-infection of Escherichia coli and Clostridium perfringens type A in a sheep farm in Ningxia and put forward corresponding prevention and control plans.Clostridium perfringens chromogenic medium and eosin blue solid medium were used for pathogen isolation and culture,and the isolated strains were detected by PCR with 16S rRNA gene sequence,and the molecular phylogenetic tree of the isolate was constructed based on the 16S rRNA sequence.After that,the toxin factors of Escherichia coli isolates were detected by PCR,and the toxin typing of Clostridium perfringens isolates was performed.The results showed that the PCR detection of isolates obtained 16S rRNA-specific bands.The results of molecular phylogenetic tree showed that the isolate strain of Escherichia coli was NXDC001 in the same branch as Escherichia coli,and the isolate strain of Clostridium perfringens was NXSJ001 in the same clade as Clostridium perfringens.Escherichia coli isolates detected toxin factors HPI(irp2)and LEE(eae).Clostridium perfringens isolates carried the toxin factor cpa,which proved that the isolate was Clostridium perfringens type A.The results showed that one strain of Escherichia coli and Clostridium perfringens type A were successfully isolated and identified,which proved that the dead sheep were co-infected with Escherichia coli and Clostridium perfringens type A.