Environmental contributors to bacterially dominated fermenting consortia of artisanal Mezcal

The production of spontaneously fermented beverages worldwide relies on native microorganisms acquired incidentally through cross-contamination from environmental reservoirs. We examined the microbiota involved in Mezcal fermentation, exploring their origins, dynamics, and ecology. Using shotgun metagenomics, we analyzed four batches of Mezcal, spanning the entire production process from crop to distillation. Bacterial genera such as Leuconostoc and Lentilactobacillus dominated the fermentation samples, whereas Bacillus was the most abundant in the environmental samples. Fermenting yeasts, such as Saccharomyces, accounted for only ~ 10% of the microbial abundance. No significant differences in microbial community structure were observed between the sampled batches, fermentation times, or depths of the fermentation tanks. Weevil samples clustered with fermentation and plant samples, suggesting they may serve as natural reservoirs for Leuconostoc and Lentilactobacillus. Functional differences were observed in COGs related to secondary metabolism during fermentation and correlated with sensory notes identified by a panel of expert tasters, suggesting that variations in the sensory profiles of the final spirit are directly linked to the metabolic products of genes associated with secondary metabolism. Our work analyzed the spontaneous fermentation microbiota, providing fundamental insights into its natural reservoirs and its contribution to Mezcal terroir.