Novel SDH substitutions confer fluxapyroxad resistance in Fusarium graminearum through two resistance‐associated regions

BACKGROUND Fusarium graminearum , the major causal agent of Fusarium head blight, is commonly controlled using succinate dehydrogenase inhibitors (SDHIs). However, the molecular basis of fluxapyroxad (Flu) resistance in this pathogen remains poorly understood. This study aimed to characterize the genetic, biochemical, and structural basis of Flu resistance. RESULTS Nine stable Flu‐resistant isolates representing five genotypes were obtained through repeated in vitro exposure to Flu, with a mutation frequency of 9 × 10 −3 . The mutants showed no significant reduction in overall biological fitness. Three previously unreported substitutions – FgSdhB P201R , FgSdhC 2 A145D , and FgSdhD D133N – were identified, and targeted allele replacement confirmed that each independently conferred moderate Flu resistance. FgSdhB P201R and FgSdhD D133N were located in the canonical quinone‐binding region, whereas FgSdhC 2 A145D was located in a putative secondary resistance‐associated region. Cross‐resistance occurred with boscalid and sedaxane, but not with pydiflumetofen or fluopyram. Flu‐mediated SDH inhibition was significantly lower in the Flu‐resistant mutants than in PH‐1. CONCLUSION These findings identify two SDH resistance‐associated regions and demonstrate that stable Flu resistance can develop in F . graminearum without significantly compromising overall biological fitness. The results provide useful insights into SDHI resistance mechanisms and support the development of effective resistance‐management strategies.