P. medicaginis is a homothallic species. Ten isolates of P. medicaginis were used (as a mixture) in all experiments, storage, and isolate culturing as described in Bithell et al. (2022) (link). Prior to inoculum production, each isolate was passaged through plants in a glasshouse using the very susceptible chickpea variety Sonali to ensure pathogenicity. With the use of low-strength V8 media (100 ml of V8 juice, 10 g of agar, 2.5 g of calcium carbonate, and 900 ml of Milli-Q water), an oospore suspension was prepared by macerating cultures with a hand-held Braun 600W blender and then added to flooded (Milli-Q water) cups of seedlings in potting mix, which were then drained after 48 h. After the observation of wilting, chlorosis, and canker development on the seedlings, stem tissue at the margin of the canker was used to re-isolate the pathogen on corn meal agar. Cultures were hyphal tipped and then grown on low-strength V8 media. Subcultures of these freshly passaged isolates were used to produce 90-mm-diameter Petri dish cultures of each isolate, which were grown in the dark at 21°C–23°C for at least 6 weeks prior to mixing with Milli-Q water (10% V/V) and macerating using a hand-held Braun 600W blender for approximately 3 min. Average oospore concentrations for each isolate were determined using counts under a 20 * 50-mm coverslip to prepare inoculum mixtures containing equal oospore concentrations.
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