Unveiling the resistance risk and mechanism of mefentrifluconazole in Colletotrichum scovillei

BACKGROUND Colletotrichum scovillei is a major fungal pathogen responsible for anthracnose disease, posing a growing threat to global chili production. Mefentrifluconazole, a novel isopropanol‐triazole fungicide, exhibits strong efficacy against a broad spectrum of plant pathogens. However, the resistance risk of mefentrifluconazole in Colletotrichum scovillei and the associated molecular mechanisms remain poorly characterized. RESULTS In this study, we evaluated the sensitivity of 102 Colletotrichum scovillei isolates to mefentrifluconazole, revealing a half‐maximal effective concentration (EC 50 ) value of 0.6924 ± 0.1482 μg mL −1 . Through fungicide adaptation, seven stable resistant mutants were generated from four sensitive parental isolates, with resistance factors ranging from 3.73 to 26.51. Compared to their parental isolates, the resistant mutants displayed similar or reduced fitness in terms of growth, sporulation and pathogenicity. Cross‐resistance assays indicated that mefentrifluconazole exhibited positive cross‐resistance with difenoconazole, propiconazole and prochloraz, but not with pyraclostrobin, florylpicoxamid or fluazinam. Further biochemical analysis demonstrated that mefentrifluconazole treatment resulted in a significantly higher inhibition rate of ergosterol biosynthesis in parental isolates relative to resistant mutants. A similar finding was observed in cell membrane damage assessment. Molecular investigations revealed no mutations in CYP51 paralogs among resistant mutants; however, quantitative analysis confirmed the overexpression of CYP51 paralogs in these isolates following mefentrifluconazole exposure. CONCLUSION Together, there is a low risk of Colletotrichum scovillei developing resistance to mefentrifluconazole, and the induced overexpression of CYP51 paralogs may contribute to potential mefentrifluconazole resistance in this pathogen. These findings offer significant implications for formulating effective management strategies against anthracnose.