Charcoal rot caused by Macrophominaphaseolinais a major disease causing in sunflower plant. The pathogen invasion occurs from the seedling to maturity stage. To overcome this problem in vitro, sensitivity of M. phaseolina was determined through inhibition zone technique to various isolates of antagonistic bacteria like seven isolates of Pseudomonas fluorescens (EPf2, EDPf3, APf4, CPf5, MPf6, KPf7 and Pf1)and seven isolates Bacillus subtilis (EBs1, EDBs2, ABs3, CBs4, MBs5, KBs6 and Bs10) amended into PDA medium. The results showed that the entire bacterial antagonist were effective against the fungus M. phaseolina and exhibited appreciable amount of inhibition. Among these bacterial antagonist significantly compared to the control P.fluorescens (Pf1) proved to be the most effective (71.49 %) with an inhibition zone of 5.00 mm reducing the colony growth of M. Phaseolina followed B. subtilis (65.92 %)inhibition zone of 17.80 mm respectively over control. However, from these studies it is concluded that an isolate of various antagonist can vary in its sclerotia producing ability on root.