Verticillium wilt is one of the most important diseases of cotton, tomato, potato and greenhouse cucumber that causes serious losses in these crops. Biological control could be an effective strategy for controlling this disease. In this study, for the investigation of antagonistic mechanisms of Talaromyces flavus, first, pathogenic agents and antagonistic fungus were isolated from cultivated regions of abovementioned crops using Komada and TF culture media respectively. In next step, antagonistic mechanisms of T. flavus including mycoparasitism, volatile metabolites production and non-volatile metabolites production were studied separately. In this study, sixty T. flavus isolates were used from which 23, 15, 14 and 8 isolates belonged to cotton, tomato, potato and greenhouse cucumber respectively. For cotton and potato wilt disease caused by V. dahliae and V. albo-atrum respectively, maximum inhibitory percents (81.51 and 64.93%) were mediated by TF-Co-G-1 and TF-Po-V-48 respectively. Non-volatile metabolites played the most important role in their antagonistic activity.However, for tomato and greenhouse cucumber wilt disease caused by V. albo-atrom, maximum inhibitory percents (73.67 and 54.78%) were mediated by TF-To-V-31 and TF-Cu-V-60 respectively.According to the results, the most effective antagonistic mechanisms of these isolates was volatile metabolites production.