The P. infestans isolate n. 111344 (CBS; Baarn, The Netherlands) was grown on pea agar medium (PAM, 12.5% w/v frozen peas in distilled water and 1.2% w/v bacteriological agar). To obtain sporangia suspensions, 7-day-old P. infestans mycelium grown on Rye B agar plates [57 (link)] were flooded with ice-cold pea broth medium (PB, 12.5% w/v frozen peas in distilled water) and rubbed with a glass rod to liberate the sporangia. The suspension was then filtered through a 45 μm nylon mesh, counted in a Kova chamber (Kova Inc., Garden Grove, CA, USA), and adjusted to a final concentration of 2 × 104 sporangia/mL.
P. ultimum isolate (n. 724.94; CBS; Baarn, The Netherlands) was cultured on PDA (Potato Dextrose Agar) plates (Liofilchem, Italy). Mycelial suspensions were prepared by homogenizing with a glass Potter tissue grinder (Thermo Fisher Scientific, Monza, Italy) in sterile PDB (Potato Dextrose Broth) amended with 0.1 g/L of Bacteriological agar (Difco, Franklin Lakes, NJ, USA), the mycelia from 4-day cultures, which had been grown on PDA plates overlaid with a sterile cellophane sheet. All the cultures were maintained at 20 °C in the dark.
Commercial fungicides Pergado SC (containing 250 g/L mandipropamid) and Ridomil Gold SL (465 g/L metalaxyl-M) (Syngenta, Milano, Italy) were employed in this study against P. infestans and P. ultimum, respectively. To assess P. infestans dose–response curves, five mandipropamid concentrations ranging from 0.1 to 1000 µg/L (logarithmic scale) were used. Metalaxyl-M was employed at 1000 mg/L to assess its activity against P. ultimum, using the dose suggested by the product’s label for field applications. Dilutions were prepared in the appropriate culture medium starting from 1000× stock solutions in water (400× for metalaxyl-M). Untreated controls with medium and sterile distilled water were also included in the assays.
P. ultimum isolate (n. 724.94; CBS; Baarn, The Netherlands) was cultured on PDA (Potato Dextrose Agar) plates (Liofilchem, Italy). Mycelial suspensions were prepared by homogenizing with a glass Potter tissue grinder (Thermo Fisher Scientific, Monza, Italy) in sterile PDB (Potato Dextrose Broth) amended with 0.1 g/L of Bacteriological agar (Difco, Franklin Lakes, NJ, USA), the mycelia from 4-day cultures, which had been grown on PDA plates overlaid with a sterile cellophane sheet. All the cultures were maintained at 20 °C in the dark.
Commercial fungicides Pergado SC (containing 250 g/L mandipropamid) and Ridomil Gold SL (465 g/L metalaxyl-M) (Syngenta, Milano, Italy) were employed in this study against P. infestans and P. ultimum, respectively. To assess P. infestans dose–response curves, five mandipropamid concentrations ranging from 0.1 to 1000 µg/L (logarithmic scale) were used. Metalaxyl-M was employed at 1000 mg/L to assess its activity against P. ultimum, using the dose suggested by the product’s label for field applications. Dilutions were prepared in the appropriate culture medium starting from 1000× stock solutions in water (400× for metalaxyl-M). Untreated controls with medium and sterile distilled water were also included in the assays.
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