洋河酒窖泥细菌群落结构与菌株产酸能力分析

【背景】窖泥微生物的种类及其代谢产物类型是影响浓香型白酒发酵过程中丁酸和己酸等白酒中主要有机酸合成的影响因素之一。【目的】揭示浓香型白酒不同窖龄窖泥细菌群落结构,研究厌氧细菌产酸性能,阐明窖泥细菌与白酒中有机酸合成的相关性。【方法】通过Illumina HiSeq高通量测序,基于16S rRNA基因序列分析不同窖龄窖泥细菌的组成。分离获得厌氧细菌,通过比较菌株产丁酸和己酸能力来分析窖泥的微生物代谢特性。【结果】洋河酒窖泥细菌主要分布于梭菌纲(Clostridia)、拟杆菌纲(Bacteroidia)、互营养菌纲(Synergistia)和芽孢杆菌纲(Bacilli)。20年窖龄的窖泥中氢孢菌属(Hydrogenispora)和瘤胃梭菌属(Ruminiclostridium)丰度显著增加。窖泥细菌间相关性分析表明,瘤胃梭菌属(Ruminiclostridium)为窖泥中影响最大的核心微生物,很多微生物与梭菌属(Clostridium)菌株之间多为相互促进关系。通过传统可培养方法共分离得到梭菌目(Clostridiales)的20株厌氧菌。其中梭菌属(Clostridium)菌株产酸能力高于其他菌属,酪丁酸梭菌(Clostridium tyrobutyricum)和丁酸梭菌(Clostridium butyricum)产丁酸和己酸的能力最强。产丁酸能力最高的菌主要分离自5年和20年窖龄窖泥,产己酸能力最高的菌分离自20年窖龄窖泥。【结论】解析了浓香型白酒不同窖龄窖泥的细菌组成,并对菌株产丁酸和己酸的能力进行了比较,为揭示窖泥微生物功能及其对白酒风味物质合成的影响奠定了相关的研究基础。

Bacterial composition in pit mud of Yanghe liquor and identification of acid producing bacteria

Background] Microbial composition of pit mud and metabolites of the endogeneous bacteria are a few of the main factors that affect the synthesis of butyrate and caproic acid during Luzhou-flavor liquor fermentation. Objective] To reveal the bacterial community of pit mud of Luzhou-flavor liquor at different ages and discover organic acid production capacity of anaerobes, and to clarify the correlation between them. Methods] The composition of bacteria in pit mud was analyzed by sequencing the 16S rRNA gene using Illumina HiSeq high-throughput platform. Anaerobes in the pit mud were isolated by selective culturing and their metabolic properties were analyzed by comparing the ability of producing butyrate and caproic acid. Results] Bacteria in the pit mud of Yanghe liquor were mainly distributed in Clostridia, Bacteroidia, Synergistia and Bacilli. The abundance of Hydrogenispora and Ruminiclostridium in 20 years pit mud increased significantly compared with that in 3 or 5 years pit mud. Correlation analysis showed that Ruminiclostridium was the most influential core microorganism. The mutual promotion relationship between bacteria associated with Clostridium was also predicted. Twenty anaerobes belonging to Clostridiales were isolated from the pit mud by traditional culturable methods. Strains of genus Clostridium produced more acids than others, Clostridium tyrobutyricum showed the most significant butyrate production capacity, while Clostridium butyricum had the strongest caproic acid production capacity. Strains with highest capability of producing butyrate were isolated from 5 and 20 years pit mud, and strains with highest capability in producing caproic acid were isolated from 20 years pit mud. Conclusion] Understanding the composition and role of bacteria in pit mud aged for different years could help better control of the synthesis of volatiles of Chinese liquor.