A Novel Point Mutation M460I in Histidine Kinase FgOs1 Confers High Resistance to Fludioxonil in <i>Fusarium graminearum</i>

Fusarium head blight (FHB), caused by <i>Fusarium graminearum</i>, severely impacts global wheat production, reducing both the yield and quality. In China, fludioxonil, a phenylpyrrole fungicide, is used for managing FHB. This study assessed fludioxonil activity against 120 <i>F. graminearum</i> strains collected from Hubei, Zhejiang, and Jiangsu in 2024, revealing an average EC₅₀ value of 0.0273 ± 0.0062 μg/mL. We obtained two resistant mutants through chemical taming and discovered a novel point mutation of FgOs1-M460I. Site-directed mutagenesis confirmed that the FgOs1-M460I mutation greatly reduced fludioxonil sensitivity, with an EC₅₀ value greater than 100 μg/mL. These mutants also displayed reduced sexual and asexual reproduction and lower virulence and accumulated less glycerol under fludioxonil and osmotic stress compared to sensitive strain. The resistant mutants showed no cross-resistance with carbendazim, tebuconazole, phenamacril, pyraclostrobin, or pydiflumetofen. Thus, we conclude that the FgOs1-M460I substitution regulates fludioxonil resistance and plays a role in asexual reproduction, sexual reproduction, and pathogenicity.