Banana fruit rot by Fusarium verticillioides is one of the most important diseases of banana in Iran. In 2021-2022 this species was identified by both morphological and molecular methods in Iran. Also, its phylogenetic relationship with the reference species in the GenBank was investigated. Pathogenicity tests revealed that all isolates induced symptoms of banana fruit rot disease that were consistent with those observed in the field. In the second step of this study, control of F. verticillioides was evaluated in vitro and in vivo. Currently, the current trend is shifting from chemical pesticides toward safer methods. Antagonist microorganisms as biocontrol agents are useful substitutes. Some bacteria and fungi have been found to have inhibitory effects on F. verticillioides. This research examined the effects of bacterial and yeast antagonist isolates, Neem (Azadirachta indica) extract and a chemical fungicide (Thiabendazole) on the mycelial growth of F. verticillioides. The results showed that two bacterial isolates and two isolates of yeasts inhibited the pathogen growth. The methanol leaf extract of Neem, at a 4000 ppm concentration, showed the most antifungal activity. Integration of four bacterial and yeast antagonists with Thiabendazole showed the best activity at a 600 ppm concentration of Thiabendazole.