PCR-DGGE技术分析传统臭鳜鱼发酵过程中细菌群落结构

摘要

应用聚合酶链式反应-变性梯度凝胶电泳(polymerase chain reaction-denaturing gradient gel electrophoresis,PCR-DGGE)技术对黄山臭鳜鱼发酵过程中的细菌群落结构组成进行研究.以发酵0~8 d的臭鳜鱼为研究对象,每2 d取样,提取样品的总DNA,同时进行16S rDNA V6~V8区的PCR扩增,产物纯化后进行DGGE分析.结果表明:肠球菌(Enterococcus sp.,D带)、腐败西瓦氏菌(Shewanella putrefaciens,C带)、溶酪大球菌(Macrococcus caseolyticus,A带)和乙酰微小杆菌(Exiguobacterium acetylicum,F带)在整个发酵过程,特别是发酵后期占较大比例,是黄山臭鳜鱼发酵过程中的优势菌.臭鳜鱼样品的PCR-DGGE图谱相似度分析显示,臭鳜鱼发酵后期菌群结构比较相似,微生物菌落结构趋于稳定.本研究结果为筛选适合工业化生产的发酵菌株,有效控制臭鳜鱼生产中存在的腐败菌、致病菌,对提高臭鳜鱼安全性和产品品质具有一定应用价值.%Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) was used to study the bacterial community composition in stinky mandarin fish during fermentation. Stinky mandarin fish was fermented for 8 days and sampled every two days for DNA extraction. The V6–V8 region of bacterial 16S rDNA was amplified by PCR and analyzed by DGGE. The results of DGGE showed that Enterococcus sp., Shewanella putrefaciens, Macrococcus caseolyticus and Exiguobacterium acetylicum gradually became dominant strains during the fermentation process, especially in the later stage of fermentation. The comparative analysis of DGGE fingerprints showed that the bacterial community of stinky mandarin fish was similar in later stage of fermentation, and became stable. These findings may provide helpful guidance for screening an industrial fermentation starter, controlling the growth of spoilage organisms and foodborne pathogens and consequently improving the quality and safety of stinky mandarin fish.