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<i>Botrytis cinerea</i>, a causal agent of gray mold disease, is one of the most destructive fungal pathogens that leads to substantial global economic crop losses, especially for tomato plants. The present study aims to investigate the inhibitory effect of two microbial culture filtrates (BCA filtrate alone and combined with salicylic acid) of <i>Trichoderma longibrachiatum</i> and <i>Pseudomonas</i> sp. against the phytopathogenic fungus <i>B. cinerea</i> on tomato plants. The biochemical modifications, gray mold disease incidence, and fruit quality parameters of the tomatoes were determined according to tested treatments. The results showed that both fungi and bacteria were able to solubilize phosphate and produce IAA and HCN. <i>T. longibrachiatum</i> could produce hydrolytic enzymes (chitinase, protease, and glucanase). Otherwise, <i>Pseudomonas</i> sp. showed the capacity to produce catalase and amylase enzymes. Both microbial culture filtrates inhibited the hyphae growth of <i>B. cinerea</i>. The biocontrol efficacy, in vitro, was significant: up to 50% in terms of the growth inhibition rate at a concentration of 40%. The tomato seedlings’ growth was promoted by the separate preventive treatments of each micro-organism culture filtrate. In addition, disease severity in the tomato seedlings and fruit was significantly reduced. Furthermore, the combined treatment of tomato fruit with culture filtrates and salicylic acid induced significant biochemical and physiological changes in fruit firmness, juice yield, total protein, and ROS enzyme activities. The culture filtrates of <i>T. longibrachiatum</i> and <i>Pseudomonas</i> sp. can be recommended as an effective microbial biofungicide to control gray mold disease under storage conditions.