Enforced Mutualism Leads to Improved Cooperative Behavior between Saccharomyces cerevisiae and Lactobacillus plantarum

Saccharomyces cerevisiae and Lactobacillus plantarum are responsible for alcoholic and malolactic fermentation, respectively. Successful completion of both fermentations is essential for many styles of wine, and an understanding of how these species interact with each other, as well as the development of compatible pairings of these species, will help to manage the process. However, targeted improvements of species interactions are difficult to perform, in part because of the chemical and biological complexity of natural grape juice. Synthetic ecological systems reduce this complexity and can overcome these difficulties. In such synthetic systems, mutualistic growth of different species can be enforced through the reciprocal exchange of essential nutrients. Here, we implemented a novel approach to evolve mutualistic traits by establishing a co-dependent relationship between S. cerevisiae BY4742Δthi4 and Lb. plantarum IWBT B038 by omitting different combinations of amino acids from a chemically defined synthetic medium simulating standard grape juice. After optimization, the two species were able to support the growth of each other when grown in the absence of appropriate combinations of amino acids. In these obligatory mutualistic conditions, BY4742Δthi4 and IWBT B038 were co-evolved for approximately 100 generations. The selected evolved isolates showed improved mutualistic growth and the growth patterns under non-selective conditions indicate the emergence of mutually beneficial adaptations independent of the synthetic selection pressure. The combined use of synthetic ecology and co-evolution is a promising strategy to better understand and biotechnologically improve microbial interactions.