Ramulispora sorghi causes sooty stripe of sorghum. Disease severity in irrigated and dryland plots was measured for 25 susceptible sorghum genotypes during the 2007 and 2008 growing seasons using a rating scale based upon percent leaf area infected. Disease severity ratings were approximately 1.4 points higher (P < 0.0001) on the rating scale in the irrigated plots than dryland plots for 2007 and 2008. Sooty stripe lesions were collected from each sorghum genotype in irrigated plots and assessed for mean microsclerotium production within lesions, microsclerotium size, and sporogenic germination, with significant differences apparent between genotypes for microsclerotium size (P = 0.01) and sporogenic germination (P = 0.01). There was no relationship between disease severity and microsclerotium production within leaf lesions, microsclerotium size, or sporogenic germination; however, there was a positive and significant correlation between microsclerotia production within a lesion and microsclerotium size (R2 = 0.19, P < 0.0001). Although microsclerotia from sorghum lesions varied in structural characteristics and their ability to produce spore masses, these qualities were dependent upon the sorghum genotype from which the microsclerotia were derived, because the R. sorghi population was genetically uniform as determined by internal transcribed spacer sequences and random amplified polymorphic DNA polymerase chain reaction.