Keeping in view the losses caused by the leaf spot disease of sesame, the present study was carried out to characterize fungal pathogenic causes leaf spot disease of sesame in Egypt. During the current study, Bipolaris sorokiniana and Alternaria sesami isolates were characterized by microscopy, colony characteristic and its pathogenicity. The identities of the fungi were confirmed by analysis of rDNA ITS (internal transcribed spacer). Fourteen RAPD primers were screened with the DNA of the fungal isolates to detect the polymorphism among these isolates. The estimated genetic similarities ranged from 74% to 82% with Bipolaris isolates, and from 85% to 96%with Alternaria isolates revealing very high levels of genetic similarity among the studied accessions. Significant differences in aggressiveness among the isolates of both pathogens were observed after artificial inoculation of sesame plants using spray inoculation methods. The disease reaction of three varieties of sesame to both fungi reveled that, the variety Shandawil-3 showed the highest degree of susceptibility to both pathogens. The host range of the isolates of both pathogens were tested on 11 plant species belonged to three families. All isolates of Bipolaris sorokiniana were pathogenic on tomato, maize, broad bean, bean, sorghum and wheat. On the other hand, the isolates of Alternaria sesami markedly varied in its pathogenicity on tomato, broad bean, bean and sorghum and were nonpathogenic to other tested plant species.In conclusions from our results, the causal pathogen Bipolaris sorokiniana is considered the most prefiling pathogen among the sesame leaf spot causal pathogen in Egypt.