几种主要窖泥微生物丰度与窖龄的关系

Relationship between pit age and the abundance of main microbes in pit mud

浓香型白酒窖泥微生物对窖池老熟和白酒品质有着关键作用.为了解能表征不同年份窖池的微生物和窖池老熟与窖泥微生物数量之间的关系,从而加强窖池的管理和维护,提供窖池人工老熟的方法和评价标准,选取几种窖泥中主要的微生物菌属,包括3个真细菌属Sporanaerobacter、Petrimonas、Tissierella,和3个古细菌属Methanobrevibacter、Methanoculleus、Methanosarcina,设计了这6个菌属的特异性引物和真细菌、古细菌的通用引物;建立了引物特异性的验证方法,并分析验证了引物的基本性质和特异性;然后应用荧光定量PCR法,测定了这6个菌属和真细菌、古细菌在不同年份窖泥中的丰度,并对比分析了其与窖龄的关系.结果显示:真细菌、古细菌总数及各目标菌数随窖龄增加呈波动上升;水井坊酒厂(代指为B酒厂)窖泥中总微生物丰度和几种目标菌属的数目在50年以内增长较快,150年与50年数量接近;剑南春酒厂(代指为A酒厂)窖泥中的Methanobrevibacter以及B酒厂窖泥中的Methanosarcina占总古细菌数百分比与窖龄呈较好的线性关系.研究证明:正常窖池的窖泥微生物丰度与窖池所酿白酒酒质成正相关关系;不同酒厂窖池的微生物总量和各种主要菌的数目及比例相差较大;不同酒厂窖池窖龄可由其特异的微生物指示.

Microbes in pit mud play a key role in the production of Luzhou-flavor liquor. This study was to understand how microbes indicate pit age, and the relationship between pit maturing and abundances of microbes. We chose to study 6 main microbes in pits including 3 genera of eubacteria （Sporanaerobacter, Petrimonas, Tissierella） and 3 genera of archaea （Methanobrevibacter, Methanoculleus and Methanosarcina）. Specific primer pairs for these genera, total eubacteria and total archaea were designed and validated; meanwhile a new primer specificity validation method was established. Then we detected by real-time fluorescent quantitative PCR the numbers of these microbes in some pit mud with different ages from two wineries in Sichuan China, Jiannanchun Winery （A） and Swellfun Winery （B） . Results showed that 1） the abundances of these microbes increased with fluctuations with pit aging; 2） in Winery B, the abundances increased fast during the previous 50 years of pit age with little difference through the 50-150 years; 3） both the percentages of Methanobrevibacter in the pit mud from Winery A and Methanosarcina in the pit mud from Winery B to total archaea had a good linear relationship with the pit age. The study showed a positive correlation between pit microbe abundance and quality of Luzhou-flavor liquor, though the abundance and proportion of pit microbes are different in various wineries. The pit age of different wineries can be indicated by specific microbes.