Deciphering the biocontrol ability of Bacillus velezensis strains against phytopathogenic fungi in grapes by in situ and in silico approaches

Bacillus species are promising biocontrol agents to confront the emergence of resistance to commonly used fungicides in agriculture. Control of Botrytis cinerea, Colletotrichum acutatum, and Aspergillus spp. is a great challenge, as these fungi negatively affect the quality and productivity of grapes. Furthermore, some Aspergillus species produce mycotoxins, which are regulated for grape products due to their toxic potential. In this study, four Bacillus velezensis strains were tested for controlling the growth of B. cinerea, C. acutatum, Aspergillus niger, Aspergillus westerdijkiae and Aspergillus flavus following in situ tests. Furthermore, an in silico approach was carried out to find genes associated with the antifungal activity of these strains. B. velezensis P1, P7, P11 and P45 demonstrated antifungal potential by inhibiting mycelial growth and spore germination, while also preventing the formation of reproductive structures in all fungi. The treatment of grapes with cell suspensions at 1.0 × 10⁹ CFU/mL reduced the fungal colony count by at least 1 log CFU/g or caused complete inhibition, depending on the strain and fungus. Even when total suppression of fungal growth was not achieved, the biosynthesis pathways of ochratoxins and aflatoxins were disrupted. Co-inoculation of strains P1:P45 and P11:P45 was successfully proposed to ensure the absence of fungi in grapes. Genome analysis revealed six gene clusters associated with the production of antifungal compounds by Bacillus strains, such as fengycin, iturin/bacillomycin, surfactin, bacillaene, bacilysin, and bacillibactin. Therefore, these bacteria are promising in the biological control of fungi that cause severe damage in grape quality, reinforcing their potential as sustainable alternatives to chemical fungicides.