Metabolomic profiling of antifungal Lactiplantibacillus plantarum strains and their inhibition of Aspergillus niger and Penicillium chrysogenum in a cheese matrix.

The dairy industry has increasingly seeking natural strategies to enhance food safety and quality. Fungal contamination remains a major challenge due to spoilage, economic losses, and mycotoxin production, making bioprotective microbial cultures a promising alternative. This study aimed to investigate the antifungal potential of five Lactiplantibacillus plantarum strains (M3.1, M3.3, M3.6, R3.2, and R3.6) against Aspergillus niger and Penicillium chrysogenum. All strains significantly inhibited fungal growth, as demonstrated by in vitro assays and reductions in fungal biomass. Scanning electron microscopy (SEM) revealed severe morphological damage to fungal spores, including surface disruption, deformation, and shrinkage. Metabolomic analyses identified the production of organic acids, including lactic, acetic, succinic, malic, propionic, butyric, and formic acids, using high-performance liquid chromatography (HPLC), as well as 35 additional metabolites, mainly amino acids, fatty acids, cyclic dipeptides, phenolic compounds, and esters, using high-performance liquid chromatography-mass spectrometry (HPLC-MS), revealing a synergistic and multifactorial antifungal mechanism. In a cheese model system, all strains completely inhibited P. chrysogenum, while strains M3.3, M3.6, and R3.2 fully inhibited A. niger. These findings demonstrate the strong antifungal activity of L. plantarum strains and support their application as bioprotective cultures in dairy products.