Pilot-scale production of Bacillus subtilis MSCL 897 spore biomass and antifungal secondary metabolites in a low-cost medium.

Abstract

Bacillus subtilis is a plant growth promoting bacterium (PGPB) that acts as a microbial fertilizer and biocontrol agent, providing benefits such as boosting crop productivity and improving nutrient content. It is able to produce secondary metabolites and endospores simultaneously, enhancing its ability to survive in unfavorable conditions and eliminate competing microorganisms. Optimizing cultivation methods to produce B. subtilis MSCL 897 spores on an industrial scale, requires a suitable medium, typically made from food industry by-products, and optimal temperature and pH levels to achieve high vegetative cell and spore densities with maximum productivity. This research demonstrates successful pilot-scale (100L bioreactor) production of a biocontrol agent B. subtilis with good spore yields (1.5×109 spores mL-1) and a high degree of sporulation (>80%) using a low-cost cultivation medium. Culture samples showed excellent antifungal activity (1.6-2.3cm) against several phytopathogenic fungi. An improved methodology for inoculum preparation was investigated to ensure an optimal seed culture state prior to inoculation, promoting process batch-to-batch repeatability. Increasing the molasses concentration in the medium and operating the process in fed-batch mode with additional molasses feed, did not improve the overall spore yield, hence, process operation in batch mode with 10g molasses L-1 is preferred. Results also showed that the product quality was not significantly impacted for up to 12months of storage at room temperature. An economically-feasible process for B. subtilis-based biocontrol agent production was successfully developed at the pilot scale.