A Multi-species assemblages or pure cultures of a single species can serve as the foundation for microbial biotechnological activities. While more functions and increased resilience to changing environmental conditions make these assemblages beneficial, they can also be unpredictable and challenging to govern in synthetically controlled development settings. The creation of robust mutualistic systems through synthetic ecology methodologies may offer creative answers for comprehending microbial interactions in these environments, helping to overcome the unpredictable nature of these microbial assemblages. Here, we establish a durable connection between two species—a strain of the yeast Saccharomyces cerevisiae and the microalga Chlorella sorokiniana—isolated from winery effluent that are evolutionary unrelated but biotechnologically complementary. Under tailored batch co-culture growth conditions, yeast and microalgae were able to develop obligatory (interdependent) and nonobligate (facultative) mutualisms. Obligate mutualism was maintained through the reciprocal exchange of carbon and nitrogen, in which the microalga uses the carbon dioxide produced by the yeast's fermentation of mannose to feed on, and the yeast receives the nitrogen from the microalga's metabolism of nitrite to ammonium. pH was revealed to be a major factor in determining the formation of these mutualisms under obligatory settings after the impact of temperature and pH on their establishment had been assessed. Additionally, compared to singlespecies cultures produced under the same conditions, the combinations of the two species under non-obligatory growth conditions resulted in an improvement in growth rate and biomass production. As the first stage in creating stable multi-species assemblages, these manufactured mutualisms also serve as a system to produce unique insights on the evolution of mutualisms.
