16S rRNA序列分析在多杀巴斯德氏菌鉴定中的应用

Identification of Pasteurella multocida Isolated from Rabbit by Using 16S Ribosomal RNA Sequence

目的应用16s rRNA序列分析方法,对多杀巴斯德氏菌进行鉴定。方法采集猝死家兔样本,进行病理组织学检查和病原菌分离培养。针对细菌16S rRNA基因保守区序列设计一对引物,以分离菌株抽提的DNA为模板,进行PCR扩增及16S rRNA序列分析。结果从死亡家兔肺脏中分离鉴定出了一株多杀巴斯德氏菌(PMr0901)。将分离菌株序列与GenBank中收录的序列进行比较,BLAST分析结果显示PMr0901与多杀巴斯德氏菌参考菌株PM70的16S rRNA核苷酸同源性大于99%。分离菌株对阿莫西林/克拉维酸、头孢曲松、左氧沙星、四环素敏感,而对青霉素G已产生耐药性。结论16s rRNA序列分析的分子生物学方法可用于病原菌的鉴定。

Objective To identify Pasteurella multocida using 16S ribosomal RNA gene sequence analysis. Methods Specimens of a suddenly died rabbit were collected. Histopathological examination, culture and isolation of pathogen were carried out. A pairs of primers targeting 16S ribosomal RNA gene conserved region were synthesized. DNA was extracted from isolated bacterial strain, amplified by polymerase chain reaction （PCR）, and analyzed 16S rRNA sequence. Results A strain of Pasteurella multocida named PMr0901 was isolated and identified from the lung of dead rabbit. Nucleotide sequences were compared with GenBank data base. The BLAST analysis indicated that PMr0901 was the most similar with reported 16S rRNA gene of Pasteurella multocida subsp, multocida str. Pm70 and the similarity more than 99%. The results of drug sensitivity test showed that the isolated bacterial strain was sensitive to amoxicillin/clavulanic acid, ceftriaxone, levofloxacin and tetracycline while was resistant to Penicillin G. Conclusion Molecular-biological approach using the 16S rRNA gene sequence analysis is available for identification of pathogenic bacteria.