Bacterial Speck Disease caused by Pseudomonas syringae pv. tomato is an important tomato bacterial disease that causes yield losses. In recent years, the disease has been a problem in tomato growing areas such as greenhouses and open fields and also commercial nurseries. Seed treatment is an important disease management strategy in the control of the disease, and seed treatments using by antagonistic microorganisms are an effective and environmentally friendly disease control method. In the research, biological control possibilities of the disease were investigated with antagonistic bacterial microorganisms isolated from tomato fields. In order to isolate the antagonists, a total of 36 soil and root samples were collected from the tomato plant growing areas in Erdemli and Toroslar districts of Mersin province. A total of 323 candidate antagonist bacterial strains were isolated from these samples in their isolations according to bacteriological techniques. The antimicrobial effects of these strains were investigated by in vitro petri dishes, and 12 antagonistic strains formed an inhibition zone between 2.3-11.0 mm and suppressed the growth of the pathogen. The strains reduced the disease incidence by 33-95% ratios and disease severity by 28-95% ratios when the twelve strains, which were successful in in vitro tests, were applied to tomato seeds as biological seed treatments. Applications did not have a negative effect on seed germination. In the second biological seed treatment experiment with the four most successful antagonistic bacterial strains, the disease was suppressed by 81-100% ratios. Thus, the effect of successful antagonists has been proven for the second time. The MALDI-TOF MS technique was used in the identification of the successful antagonistic strains, it was determined that the antagonists were identified as Bacillus subtilis, Bacillus pumilus and Paenobacillus polymyxa. It has been shown in the study that Bacillus species with antagonist properties can be used successfully as biological seed treatments in the control of Bacterial Speck Disease on tomatoes. It should be investigated in the further research that the effect of the antagonistic Bacillus strains on the inhibition of the disease on foliage parts of tomato, their mechanism of action and the effects on other antagonists, plant pathogens. There is also a need for detailed studies such as mass production, obtaining licenses and patents for its practical use. It was concluded that the antagonistic strains can be used in part of the integrated disease management programs after the additional experiments are performed.