In Vitro Assessment of Penicillium expansum Sensitivity to Difenoconazole

<i>Penicillium expansum</i> causes blue mold, a major post-harvest disease affecting apples. This disease is commonly managed using fungicides, including Difenoconazole (Dif), a demethylation inhibitor (DMI) approved for its control. This investigation aims to evaluate the baseline sensitivity of 100 <i>P. expansum</i> isolates to Difenoconazole. The isolates were collected from symptomatic apples in 34 storage warehouses across the Fes-Meknes and Draa-Tafilalet regions over three years (2020, 2021, and 2022). The study revealed an increase in the percentage of inhibition of mycelial growth and spore germination of <i>P. expansum</i> proportional to the increasing concentration of the fungicide. Moreover, the results indicate that 46 isolates were able to develop even at a concentration of 5 µg/mL of Dif (the suggested discriminatory dose), indicating reduced sensitivity to this fungicide. The analysis of the values of the effective concentration to inhibit 50% (EC50) of mycelial growth of <i>P. expansum</i> ranging from 0.027 to 1.673 µg/mL (mean: 0.263 µg/mL, variation factor: 62.507) and for spore germination from 0.0002 to 0.787 µg/mL (mean: 0.048 µg/mL, variation factor: 4113.835). The wide variation in EC50 values indicates significant variability in the isolates' responses to Dif, likely due to diverse sampling in space and time. Our results showed that some <i>P. expansum</i> isolates could grow even at high concentrations of Dif, indicating limited efficacy of this treatment. The EC50 of five isolates exceeded 0.92 µg/mL, suggesting potential resistance. This study indicates reduced sensitivity and possible emergence of resistant strains. Notably, it is the first evaluation of <i>P. expansum</i> sensitivity to Dif in Morocco.