Sesame (Sesamum indicum L., 2n = 2x = 26), also called Indian sesame, is an annual herbaceous plant of the Pedaliaceae Lindl family that includes up to 38 species, reaches a height of 1.7-1.8 m. It has a small genome size of 337–357 Mb. Sesame seeds contain 50-65% of oil, up to 27% protein, 20% carbohydrates and are source of copper, calcium, iron, magnesium, zinc, vitamins A, B1, C, E. Sesame seeds and oil are used in food industry, in medicine, in cosmetology. In terms of taste, sesame oil belongs to the best edible oils, along with worldwide known Provencal oil. The oxidative stability of sesame oil is higher than that of other vegetable oils. Recently, a decrease in the production of sesame is observed, which is caused by various diseases. Pathogenic fungi are among the factors contributing to the yield decrease. Alternaria spp. has been reported to cause yield losses of up to 30-40%. Fungi inhibit plant growth as well as the rate of seed germination. The initial route of spread of diseases is through seeds. Fungal pathogens can be transmitted as well by being on the surface or inside the seed. To determine the infection, healthy seeds without obvious external signs of infection were selected. Total DNA was isolated from seeds using 2% SDS (sodium dodecylsulfate) buffer. Samples were analyzed using nested-PCR protocol with a set of genus-specific and species-specific primers for fungal pathogens. Seeds of six sesame cultivars - Kubanets 57, Lider, Donskoi belosemianyi, Manchurskii ulucshenyi, Biolsadovski, Zaltsadovski, were analyzed for fungal infestation with Alternaria spp., Fusarium spp, Myrothecium spp. pathogens. Alternaria spp. was found in the analyzed varieties. Myrothecium roridum was identified in Kubanets 57 variety. Pathogens of Fusarium spp. were not detected in the studied six varieties of sesame seeds (Sesamum indicum L.).