Optimizing Chlorella vulgaris production and exploring its impact on germination through microalga-N₂-fixing bacteria...

Microalgal biomass is increasingly valued in industrial and agricultural sectors due to its bioactive compounds. However, large-scale production remains costly, mainly due to nitrogen fertilizer expenses. A promising sustainable alternative is co-cultivation with N₂-fixing bacteria, capable of supplying biologically available nitrogen. In this study, Chlorella vulgaris was grown in synthetic medium with and without nitrogen, as well as in co-culture with three different N₂-fixing bacteria in nitrogen-free medium. Microalgal growth was assessed by dry weight, Fv/Fm ratio, and flow cytometry, which also allowed evaluation of population dynamics and cell viability. Biomass composition (proteins, carbohydrates, lipids, chlorophyll, and carotenoids) was analyzed under all conditions. Co-cultures in nitrogen-free medium showed comparable biomass productivity to nitrogen-supplemented controls, although Fv/Fm values indicated physiological stress in some cases. Moreover, the agricultural potential of the resulting biomass and supernatants was evaluated through germination bioassays using lettuce seeds. All cultures tested at 0.2 g·L^-1 significantly improved the germination index. Also, applying the culture supernatant (biomass removed) also yielded positive effects, with GI increases exceeding 40 %. These results suggest that co-cultivation with N₂-fixing bacteria can support efficient microalgal production while generating biomass and supernatants with biostimulant potential, contributing to sustainable agriculture and circular bioeconomy strategies.