Exploratory metabolomics of Vibrio diabolicus across culture media reveals medium-dependent anti-Vibrio activity patterns

Vibriosis caused by Vibrio parahaemolyticus remains a major challenge in shrimp aquaculture, driving the search for alternative strategies to control pathogenic Vibrio proliferation. The probiotic-associated strain Vibrio diabolicus has been suggested to exhibit protective potential; however, how cultivation conditions influence its bioactive metabolite profile remains poorly characterised. Here, we explored the effect of culture medium composition on the metabolomic profiles and in vitro anti-Vibrio of V. diabolicus. The bacterium was cultivated in five media differing in carbon and nitrogen sources, followed by solvent fractionation and antibacterial testing. Untargeted LC–MS/MS metabolomics enabled the annotation of 25 metabolites across major chemical classes, including glycerophospholipids, fatty acid amides, indole derivatives, alkaloids, and aromatic compounds. The starch–yeast medium yielded extracts with greater anti-V. parahaemolyticus activity (methanolic fraction MIC 5 mg/mL) and no acute toxicity in shrimp larvae under the tested conditions. Molecular networking and multivariate analyses revealed medium-associate differences in metabolite composition, with enrichment of lipid-related and nitrogen-containing features in more active fractions. Docking analyses suggested potential interactions between selected metabolites and the β-lactamase CARB-20, although these predictions require experimental validation. Overall, the results indicate that cultivation conditions are associated with metabolite profile variation and corresponding differences in extract-level anti-Vibrio activity, supporting medium optimization as a practical parameter in developing Vibrio-based biocontrol approaches.