Interaction Between Bacillus subtilis and Trichoderma citrinoviride: Synergistic Inhibition of Aspergillus flavus Growth and Aflatoxin B1 Production

<i>Aspergillus flavus</i> (<i>A. flavus</i>) and its potent carcinogenic metabolite, aflatoxin B₁ (AFB₁), pose severe threats to food safety and public health. This study explored the biocontrol potential of <i>Bacillus</i> and <i>Trichoderma</i> strains and their interactions. Through dual-culture assays, <i>Bacillus subtilis</i> Bs92 and <i>Trichoderma citrinoviride</i> GC-T20 were identified as the most inhibitory among 15 isolates each, with their culture filtrates inhibiting <i>A. flavus</i> growth by 48.61% and 74%, respectively. Investigation of the inter-genus interaction revealed strong mutual inhibition: <i>Trichoderma</i> filtrate suppressed <i>Bacillus</i> growth by >90%, while <i>Bacillus</i> filtrate inhibited <i>Trichoderma</i> mycelial growth by 40-90%. To circumvent this antagonism and enhance efficacy, the culture filtrates of Bs92 and GC-T20 were combined. The combined treatment demonstrated superior performance, inhibiting <i>A. flavus</i> radial growth by 69.59% and AFB₁ production by 98.69%, significantly outperforming individual applications. These results indicate that the antifungal and anti-aflatoxigenic metabolites produced by Bs92 and GC-T20 are complementary. Their combined use presents a promising synergistic strategy for the biocontrol of <i>A. flavus</i> and AFB₁ contamination in food and agricultural products.