Biocontrol of Aspergillus flavus growth and aflatoxin production by kefir-derived microorganisms under simulated storage conditions

Abstract

Kefir granules are a complex microbial consortia with numerous properties, among which their antifungal action has been reported. This study characterized the antifungal activity of water kefir granules and two microorganisms isolated from them, identified as Saccharomyces cerevisiae and Geotrichum candidum, against aflatoxigenic Aspergillus flavus. Experiments were carried out in vitro and under minisilo conditions designed to simulate the aerobic phase of commercial corn storage. In dual culture plate assays, A. flavus was inhibited in close contact with both antagonists, and mycelial weakening with lower conidia production was observed. Cell-free supernatants of both antagonists inhibited conidial germination by 92% (G. candidum) and 65% (S. cerevisiae). Non-contact assays between A. flavus and antagonists indicated that volatile organic compounds (VOCs) also contributed to inhibition, reducing mycelial growth by 29.1% (G. candidum) and 37.9% (S. cerevisiae). In corn minisilo trials water kefir granules and both antagonists showed strong antifungal activity by day 4, with qPCR of A. flavus DNA and aflatoxin measurements indicating near complete inhibition. As mixed consortia, water-kefir granules inherently include lactic acid bacteria whose rapid acidification and diffusible metabolites complement antifungal compounds produced by other components of the consortia, supporting a multi-mechanistic mode of action. Given the compositional heterogeneity of kefir granules, strain-level identification and verification of safety and regulatory status for the intended use are essential prior to food or feed applications. The demonstrated antifungal efficacy of kefir-derived microorganisms in mini-silos underscores their scale-up potential for postharvest protection of corn.