Evaluation of new fermentation and formulation strategies for a high endophytic establishment of Beauveria bassiana in oilseed rape plants

Since several years it is known that strains of the entomopathogenic fungus Beauveria bassiana (Balsamo-Crivelli) Vuillemin (Ascomycota: Hypocreales) are able to colonize plants as a true endophyte. However, so far no integrated bioprocess engineering approaches have been published where fermentation and formulation strategies are combined to optimize colonization of oilseed rape plant tissues. We therefore aimed at investigating whether and how blastospore (BS) formation can be shifted to resilient submerged conidiospores (SCS) by introducing osmotic stress in different growth phases.When 50 g/L NaCl was added after 48 h to a culture of B. bassiana a yield of 1.4 ± 0.1 × 10¹⁰ SCS/g sucrose in shake flasks and 1.8 ± 0.1 × 10¹⁰ SCS/g sucrose in a stirred tank reactor were obtained. In a bioreactor, 24 h after the addition of NaCl, the formation of BS slowed down, the respiratory quotient decreased and a shift from BS to SCS set in.Following these steps, different formulation strategies, namely encapsulation, film coating and liquid formulation were evaluated. B. bassiana grew out of beads as well as on commercial fungicide-coated seeds. Due to the complete suppression of fungal growth on non-sterile soil, the most suitable option was a foliar application. A liquid formulation consisting of 0.1% Triton X-114, 1% molasses, 1% titanium dioxide and 10⁶ spores/mL was applied on leaf tips. After 14 days, the endophyte was detected by PCR and microscopic analysis in the leaves.Further research should focus on formation of SCS and protection of plants colonized by B. bassiana against herbivorous insects.