Table_1_Profiling of Fungal Diversity and Fermentative Yeasts in Traditional Chinese Xiaoqu.docx

To increase the safety and quality of baijiu and rice wine in China, controlling the use of traditional Xiaoqu by studying the beneficial yeasts present has recently been considered. The fungal diversity of six Chinese Xiaoqu including five traditional and one commercial samples was investigated to screen fermentative yeasts with low yields of higher alcohols. A high throughput sequencing approach detected fifteen fungal species with relative abundance higher than 1%, and displayed dissimilarities of fungal diversity among Xiaoqu samples. The 15 fungal species were composed of 11 filamentous fungi with Rhizopus arrhizus as the most common specie and four yeast species, containing Hyphopichia burtonii, Saccharomyces cerevisiae, Saccharomycopsis fibuligera, and Saccharomycopsis malanga. Classic culture-dependent approaches, including 5.8S-ITS-RFLP analysis and D1/D2 sequencing of the 26S rRNA gene, identified nine yeast species in the five traditional Chinese Xiaoqu. In addition to the four yeast species also detected by high throughput sequencing approach, the other five yeast species isolated were Clavispora lusitaniae, Cryptococcus neoformans, Komagataella pastoris, Trichosporon asahii, and Wickerhamomyces anomalus. Further micro-fermentations of rice wine were performed using 19 single yeast isolates, and after the fermentation of rice wine, higher alcohols and ethanol were analyzed by gas chromatography. Two yeast strains, Saccharomyces cerevisiae FBKL2.8022 and Wickerhamomyces anomalus FBKL2.8023, were found to have low yields of higher alcohols and could produce 11.70%vol and 7.10%vol ethanol separately. This study for the first time, to the best of our knowledge, explored the fungal resources in traditional Xiaoqu from different regions of Guizhou, China. The screened S. cerevisiae and W. anomalus strains could be used to establish specific starters to promote the standardization of the production of baijiu and rice wine.

为了提升中国白酒和米酒的安全性与品质，近期通过研究传统小曲中存在的有益酵母，以控制其使用，已成为一项重要考量。本研究对包括五款传统和一款商业样品在内的六个中国小曲的真菌多样性进行了调查，以筛选出产高醇量低的发酵酵母。采用高通量测序方法检测出相对丰度高于1%的十五种真菌物种，并显示出小曲样品间真菌多样性的差异。这十五种真菌物种由十一类丝状真菌组成，其中根霉属的Rhizopus arrhizus为最常见物种，以及四种酵母物种，包括毛顶毛霉属的Hyphopichia burtonii、酿酒酵母属的Saccharomyces cerevisiae、纤维状酵母属的Saccharomycopsis fibuligera和Saccharomycopsis属的Saccharomycopsis malanga。经典的文化依赖性方法，包括5.8S-ITS-RFLP分析和26S rRNA基因的D1/D2测序，在五款传统中国小曲中鉴定出九种酵母物种。除了由高通量测序方法检测出的四种酵母物种外，还分离出五种其他酵母物种，分别为Clavispora lusitaniae、Cryptococcus neoformans、毕赤酵母属的Komagataella pastoris、毛壳霉属的Trichosporon asahii和魏克汉姆酵母属的Wickerhamomyces anomalus。进一步使用19个单酵母分离株进行米酒微发酵，并通过气相色谱分析发酵后的高醇和乙醇含量。研究发现，酵母菌株Saccharomyces cerevisiae FBKL2.8022和Wickerhamomyces anomalus FBKL2.8023产高醇量低，分别能产出11.70%vol和7.10%vol的乙醇。据我们所知，本研究首次探索了来自中国贵州省不同地区传统小曲中的真菌资源。筛选出的S. cerevisiae和W. anomalus菌株可用于建立特定的启动剂，以促进白酒和米酒生产的标准化。