Single Amino Acid Change in the FvHIK1 Protein Associated with Fludioxonil Resistance in Fusarium verticillioides

Fusarium stalk rot, caused by Fusarium verticillioides, is a major disease affecting maize production. Currently, chemical control serves as the primary management strategy, and our preliminary studies have demonstrated that fludioxonil exhibits excellent antifungal activity against F. verticillioides. However, the resistance risk and potential resistance mechanisms have not been fully established. In this study, we investigated four resistant laboratory isolates of F. verticillioides that had fludioxonil EC₅₀ values ranging from 43.3 to 53.7 μg/ml. Evaluation of their biological characteristics indicated a significant fitness cost resulting from both reduced mycelial growth and impaired pathogenicity. In addition, the isolates were extremely sensitive to osmotic stress induced by NaCl, KCl, mannitol, and glucose, suggesting a decline in their environmental adaptability. Meanwhile, molecular analysis revealed that all isolates carried a G298D substitution in FvHIK1 but little evidence of altered expression. Molecular docking analysis indicated that the G298D mutation changed the minimum binding energy from -5.6 to -5.3 kcal/mol. Cross-resistance experiments revealed no correlation between fludioxonil and other fungicides with unrelated modes of action, such as tebuconazole, prochloraz, azoxystrobin, or fluazinam, confirming a fludioxonil-specific mechanism of resistance. Taken together with the extreme sensitivity to osmotic stress, it is likely that the G298D in FvHIK1 caused the high levels of fludioxonil resistance observed in the current study. The absence of cross-resistance indicates that tebuconazole, prochloraz, azoxystrobin, and fluazinam remain viable options for F. verticillioides control if this type of resistance arose in the field and that rotation could help prevent fludioxonil resistance emergence.