Abstract

BACKGROUND Fusarium head blight (FHB) caused by Fusarium graminearum severely threatens wheat production. Although fludioxonil and tebuconazole are key fungicides for FHB control, emerging resistance has reduced their efficacy, and no dual-resistant F. graminearum strains have yet been reported. RESULTS We collected 120 F. graminearum isolates in 2024 from Shandong, Shanxi, Hebei, and Anhui provinces. Among them, 119 isolates showed sensitivities to fludioxonil ranging from 0.0084 to 0.0392 μg/mL, with an average EC50 (effective concentration 50%) value of 0.0238 ± 0.0059 μg/mL. We identified one isolate exhibiting dual resistance to both fludioxonil and tebuconazole. Its ultrahigh fludioxonil resistance was mediated by a novel point mutation, FgOs1A307V. Site-directed mutagenesis confirmed that FgOs1A307V conferred ultrahigh fludioxonil resistance (EC50 >100, RF (resistance factor) >4000). Tebuconazole resistance is attributed to FgCyp51B overexpression, conferring low-level resistance (20 >RF >5). Furthermore, the FgOs1A307V mutation caused defects in asexual and sexual reproduction and reduced pathogenicity. The mutant also exhibited osmotic stress hypersensitivity and reduced glycerol accumulation. The dual-resistant isolate showed no cross-resistance to carbendazim, phenamacril, pydiflumetofen, or pyraclostrobin. CONCLUSION This study demonstrates that F. graminearum strains with dual resistance to fludioxonil and tebuconazole have emerged in the field. It clarifies that fludioxonil resistance and tebuconazole resistance are conferred by FgOs1A307V mutation and FgCyp51B overexpression, respectively. The dual-resistant strain exhibited a significant fitness cost. These findings inform sustainable fungicide rotation and resistance management for more effective FHB control. © 2026 Society of Chemical Industry.