河南省20株布鲁氏菌鉴定与PFGE分子分型研究

目的检测与分析河南省2010-2012年20株布氏菌型别及PFGE脉冲场凝胶电泳指纹图谱特征。方法采集病人静脉血,分别以试管凝集试验(SAT)、双相血培养瓶分离培养、热裂解法制备DNA模板和AMOS-PCR鉴定4种布氏菌型别。采用脉冲场凝胶电泳技术(PFGE)对检出的布鲁氏菌进行分子分型鉴定。结果分离培养的20株布鲁氏菌经鉴定,19株为羊种,1株为牛种;羊种布鲁氏菌经XbaI酶切与脉冲场凝胶电泳后,共获得了8种不同带型,带型相似度在80%~100%之间,牛种与羊种菌株在带型相似度上差异较大,具有较好的分辨能力。结论河南省病人感染的布鲁氏菌以羊种为主要型别,AMOS-PCR与PFGE作为布鲁氏菌菌型鉴定与分子分型的技术手段,为布鲁氏菌病的病原学监测与应急检测提供依据。

Identification and PFGE molecular typing of Brucella strains in Henan Province,China

The aim was to detect and analyze Brucella strains isolated from patients in Henan Province from 2010 to 2012 with AMOS-PCR amplification and pulsed field gel electrophoresis technology, provide baseline for the monitoring, early warning, outbreak investigation, and tracing of brucellosis. We collected SAT tested positive patients blood and cultured with double phase blood bottle for at least 20 days. AMOS-PCR identification of 4 species of Brucella type was conducted using DNA template with thermal decomposition method after morphological and biochemical identification. The pulsed field gel electrophoresis (PFGE) was made for getting the molecular type characters of Brucella stains. Results showed that 20 strains of Brucella were identified by morphological, biochemical and AMOS-PCR technology, in which 19 strains were sheep species, 1 strain was cattle species. Brucella strains were digested with XbaI and made pulsed field gel electrophoresis. The result indicated that the similarity coefficient among the 19 sheep species isolates was from 80% to 100% with 9 belt types, which had a great difference from cattle species. The sheep species Brucella is the main type in patients in Henan Province. AMOS-PCR and PFGE technology provide a strong tool for the brucellosis monitoring and emergency detection.