Inhibitory Effects and Mechanisms of Thymol Against Fusarium oxysporum, a Pathogen of Stem Base Rot

Background/Objectives: Stem base rot caused by Fusarium oxysporum is a devastating soil-borne disease, yet long-term chemical pesticide application has raised serious environmental and ecological concerns. Thymol, a plant-derived fungicide, demonstrates a favorable safety profile for humans and environmental systems while exhibiting broad-spectrum antifungal efficacy. This study aimed to evaluate the inhibitory effects of thymol against F. oxysporum and to elucidate its underlying mechanisms of action. Methods: The antifungal activity of thymol was assessed in vitro against F. oxysporum by measuring colony and mycelial growth inhibition, sporulation, and spore germination at varying concentrations. Microscopic examination was performed to observe morphological changes in hyphae. Biochemical assays were conducted to quantify soluble protein content, DNA synthesis, ergosterol levels, and fusaric acid production in treated mycelia. Dose–response data were used to calculate median effective concentrations (EC50) for relevant parameters. Results: Thymol significantly inhibited F. oxysporum growth, with EC50 values of 51.02 μg/mL for colony growth and 57.30 μg/mL for mycelial growth. Microscopy revealed abnormal, shorter, thicker hyphae with enlarged vacuoles. Sporulation and spore germination were also effectively suppressed, with EC50 of 39.37 μg/mL and 53.55 μg/mL, respectively. At 100 μg/mL, thymol reduced soluble protein content by 36.73%, inhibited DNA synthesis by 47.29%, decreased ergosterol content by 48.11%, and reduced fusaric acid production by 76.86%. Conclusions: Thymol exerts multi-targeted inhibitory effects against F. oxysporum through disruption of cellular structure, impairment of macromolecular synthesis, interference with ergosterol biosynthesis, and suppression of toxin production. These findings provide a strong theoretical foundation for developing thymol as a novel biopesticide for sustainable management of stem base rot.