In the present study the mould incidence, ergosterol and aflatoxin B1 (AFB1) contaminations were evaluated in different food based seed samples viz., chickpea, cowpea, green gram, groundnut, Indian bean, maize, sorghum, soya bean and sunflower collected from different agro-climatic regions of Karnataka (India). The agar plate and standard blotter methods were employed for determination of the fungal incidence, and the ergosterol and AFB1 contents were estimated qualitatively and quantitatively by TLC and spectrophotometric methods. For detection of aflatoxigenic isolates of A. flavus, the target gene specific primer aflR-F and aflR-R were used in PCR amplification. The antifungal and antiaflatoxigenic activities of some selected edible vegetable extracts were evaluated by measuring mycelial dry weight and AFB1 content. The results revealed that 15 diverse fungal species belonging to 11 genera were observed. Among the seed samples analyzed, the highest fungal incidence, ergosterol and AFB1 were observed in sorghum samples followed by maize and chickpea. The PCR amplification showed positive results only for aflatoxigenic isolates of A. flavus and no amplification was observed in nonaflatoxigenic isolates and A. flavus isolate-2 produced highest AFB1, showed 99% similarity with an authenticated aflatoxigenic A. flavus isolate emb|FN398161.1|. The aqueous extract of Amorphophallus campanulatus showed highest antifungal and antiaflatoxigenic activities. The results confirmed that the ergosterol and AFB1 contents were correlated with the percent mould incidences.