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Quantitative Inoculation of Eastern Cottonwood Leaf Tissue with Melampsora medusae Under Controlled Conditions. Louis Shain, Associate professor, Department of Plant Pathology, University of Kentucky, Lexington 40506; P. L. Cornelius, associate professor, Departments of Agronomy and Statistics, University of Kentucky, Lexington 40506. Phytopathology 69:301-304. Accepted for publication 5 October 1978. Copyright 1979 The American Phytopathological Society. DOI: 10.1094/Phyto-69-301.

A procedure for quantitative inoculation of excised leaf tissue of eastern cottonwood with urediospores of Melampsora medusae is described. The procedure does not require large numbers of spores or elaborate apparatus. Ten-μliter samples of a urediospore suspension in 0.1% water agar were placed on the underside of leaf disks (17 mm diameter) supported on 1% agar in petri dish incubation chambers. Favorable incubation conditions were a 14-hr photoperiod (10,000 lux) at 18 C. Uredia usually appeared within 6 days after inoculation and most were erumpent within 8 days. Disks from leaf quadrants that had been washed briefly with 52% ethanol prior to inoculation had significantly greater infection than unwashed or water washed disks from quadrants of the same leaf. A linear relationship was obtained by plotting γ1+x transformations, in which x = infections/disk, against log₂ of the initial inoculum density. The range of most efficient inoculum density was 1,250–5,000 urediospores per disk, which indicated that 25–29 urediospores could cause a single infection. Leaves of about the same size and position on the stem from the same clone sometimes differed significantly in susceptibility to rust. This intraclonal variation can be accommodated statistically by distributing disks from the same leaf among experimental treatments.