Aflatoxins are toxic and carcinogenic metabolites of several Aspergillus species. The effect of nitrate on aflatoxin production and expression of the key regulatory genes involved in aflatoxin biosynthesis, aflR and aflJ, were compared among isolates of the S-B and S-BG strains of A. flavus. Aflatoxin production by two of the three strain S-B isolates did not differ significantly between the two media tested, whereas for S-BG A. flavus isolates, the level of aflatoxins in buffered nitrate medium was as much as 20-fold lower than in ammonium salts medium. Expression of aflR was not significantly affected by growth of cultures in nitrate medium for most of the isolates. However, on nitrate medium, expression of aflJ was 2.6-fold higher for the S-B isolates than it was on ammonium salts medium, whereas for the S-BG isolates aflJ expression was 2-fold lower on nitrate than on ammonium salts medium. This difference may result from the presence in the aflJ/aflR intergenic region of S-BG isolates of fewer putative binding sites (HGATAR sites) for AreA, the positive-acting, wide domain transcription factor involved in regulation of nitrogen metabolism.