In the present study, natural occurrence of fungi and aflatoxin B1 (AFB1) in trout pellet feed and feed ingredients randomly obtained from feed markets was investigated. The samples were cultured on the standard isolation media for 2 weeks at 28 ºC. Identification of fungal isolates was implemented based on the macro- and microscopic morphological criteria. AFB1 was detected using high performance liquid chromatography (HPLC). Based on the results obtained, a total of 109 fungal isolates were identified of which Aspergillus was the prominent genus (57.0%), followed by Penicillium (12.84%), Absidia (11.01%) and Pseudallscheria (10.10%). The most frequent Aspergillus species was A. flavus (60.66%) isolated from all the feed ingredients as well as pellet feed. Among 37 A. flavus isolates, 19 (51.35%) were able to produce AFB1 on YES broth in the range of 10.2 to 612.8 μg/g fungal dry weight. HPLC analyses of trout feed showed that pellet feed and all feed ingredients tested except gluten were contaminated with different levels of AFB1 in the range of 1.83 to 67.35 μg/kg. Unacceptable levels of AFB1 were reported for feed including soybean, fish meal and wheat. These results indicate the importance of AF contamination of trout feed in amounts higher than the acceptable level as a risk factor for fish farming production.