Evaluation of Simplified Leaf Inoculation Procedures for Identification of Quantitative Resistance to Sclerotinia trifoliorum in Alfalfa Seedlings

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Abstract
Previous studies established that excised leaf tissues of alfalfa can be inoculated with Sclerotinia trifoliorum to select for heritable resistance, but the original procedures were not practical for use in large-scale screening programs. In this study, simplified leaf inoculation procedures for more rapid screening for resistance, based on direct application of leaf tissues to colonies of the pathogen on agar media, were evaluated. Cotyledons, unifoliate leaves, and leaflets of trifoliolate leaves of plants 7 to 21 days old from three relatively susceptible cultivars and one resistant germ plasm were applied, with and without wounding, directly to colony margins of S. trifoliorum on cornmeal agar, V8 juice agar, and water agar. Leaves were scored according to the rate and extent of development of necrosis. Significant differences between alfalfa populations were expressed in unifoliate leaves and trifoliolate leaflets but not in cotyledons. Disease severity in the resistant germ plasm (Mississippi Sclerotinia-Resistant [MSR]) was less than in the three cultivars on all agar media. Wounding of leaf tissues increased disease severity and greatly reduced the incidence of symptomless leaves, which are indicated to be escapes, but wounding generally did not prevent expression of resistance in MSR. Results indicate that initial screening for resistance to S. trifoliorum in alfalfa seedlings may be accomplished by applying wounded unifoliate leaves and leaflets of trifoliolate leaves directly to colonies on cornmeal or V8 juice agars.