Lipopeptide-Mediated Inhibition of Aflatoxin Biosynthesis in Aspergillus flavus Revealed by DESI-MSI and Transcriptomics

Aflatoxin contamination caused by Aspergillus flavus represents a critical threat to global food safety and agricultural sustainability, yet the mechanisms underlying its effective biological control remain poorly understood, particularly at the spatial and metabolic levels. Here, we systematically investigated the biocontrol potential of Bacillus velezensis B2 and elucidated the mechanisms of its bioactive metabolites using an integrated multi-omics strategy. Chemical profiling and purification identified surfactin-iturin complexes (SLCs) as the major antifungal components. Functional assays demonstrated that SLCs completely inhibited spore germination at 160 μg/mL (MIC = 0.125 mg/mL) and exhibited approximately 10-fold higher antifungal potency than the crude extract. SLC treatment also induced severe hyphal deformities, including swelling and membrane disruption. Spatial metabolomics using DESI-MSI revealed a dynamic metabolic shift from m/z 1020 to m/z 1034 during coculture, with surfactin-type metabolites dominating the interspecies interaction interface. Transcriptomic analysis identified 3,560 differentially expressed genes (1,559 upregulated and 2,001 downregulated), and demonstrated that SLCs globally suppressed the aflatoxin biosynthetic pathway by downregulating key genes, including aflK , aflQ , aflP , aflO , aflM , aflE , aflH , and aflT . Consistently, no detectable AFB1 was observed in treated samples (LOD = 0.05 μg/kg). Collectively, these findings demonstrate that SLCs inhibit aflatoxin production through combined structural damage and metabolic reprogramming, and highlight a spatially resolved multi-omics framework for deciphering microbial interactions and developing effective biocontrol strategies in food safety. • Isolation and purification of SLCs from B. velezensis B2. • SCLs exhibit strong inhibitory effects against A. flavus. • DESI-MSI analysis of antimicrobials in B.velezensis - A.flavus coculture. • Aflatoxin pathway analysis with gene correlation heatmap.