Antagonistic Potential of Agro-Industrial Byproduct–Derived Lactic Acid Bacteria Against Mycotoxigenic Aspergillus flavus and Fusarium verticillioides

Mycotoxins pose significant threats to food security and human health, necessitating innovative approaches for fungal control. This study investigated the antifungal and antimycotoxigenic potential of lactic acid bacteria (LAB) isolated from agro-industrial byproducts against the toxigenic fungi Aspergillus flavus and Fusarium verticillioides. Then, 14 LAB isolates were phylogenetically characterized, revealing diverse species including Lactiplantibacillus pentosus, Lactiplantibacillus plantarum, Lacticaseibacillus paracasei, and Leuconostoc pseudomesenteroides. Their antagonistic activity was first screened using an overlay-streak assay, which evaluated the combined effects of competition, pH reduction, and metabolite production on mycelial growth. Subsequently, the effect of their neutralized cell-free supernatants (CFS)—containing possible pH-stable antimicrobial compounds—was tested on fungal proliferation. The results revealed a distinct, mode-of-action-dependent efficacy. In the direct coculturing assay, stronger inhibition was observed against F. verticillioides, with six L. pentosus strains achieving nearly 100% growth suppression. In contrast, CFS treatments exhibited a more pronounced inhibitory effect on A. flavus germination and growth rate, with L. plantarum 71(6)-2F showing activity comparable to a positive control. This shift in efficacy is explicitly attributed to the different mechanisms assessed in each assay: the overlay method reflects broad-spectrum inhibition driven largely by competition and acidification, to which F. verticillioides appears highly sensitive. The CFS assay, however, highlights the impact of specific, pH-neutral antimicrobial metabolites. Furthermore, several CFS extracts significantly reduced mycotoxin biosynthesis, suggesting these LAB metabolites can disrupt critical fungal physiological pathways. These findings underscore the potential of LAB from agro-industrial byproducts as a source of natural antifungal and antimycotoxigenic compounds.