Brown blotch disease of mushroom caused by Pseudomonas tolaasii , often results of severe damage to mushroom tissues. In this study, the biocontrol mechanisms mediated by endofungal bacteria Pseudomonas sp. De1, Bacillus sp. De3, Pseudomonas sp. Bi1 and Pantoea sp. Ma3 previously isolated from wild growing mushrooms against Pseudomonas tolaasii Pt18 were demonstrated. Our results provide evidence that all bacterial strains can colonize mycelia of Agaricus bisporus. All endogungal bacteria except Pseudomonas sp. Bi1 contained one or more genes encoding antibiotic production. The non-volatile compounds of endofungal bacteria significantly inhibited the biofilm formation by P. tolaasii Pt18. Results showed that all strains except Pantoea sp. Ma3 are capable of detoxify tolaasin, a major virulence factor produced by P. tolaasii Pt18. We also showed that all endofungal bacteria tested were able to significantly decrease the browning symptoms of pretreated-mushroom caps compared to inoculated caps with P. tolaasii alone. AbPPO3 gene expression levels in mycelia and fruiting bodies of A. bisporus significantly increased after P. tolaasii Pt18 inoculation. Our findings indicated that The relative expression level of AbPPO3 gene decreased in the mushroom mycelia and fruiting bodies pretreated with Pantoea sp. Ma3 and Pseudomonas sp. Bi1 strains, and the levels were significantly lower than mushroom tissues treated with P. tolaasii Pt18 alone. Overall, the findings presented in this study revealed the efficacy of the selected endofungal bacteria in biocontrol activity against P. tolaasii and the possible use of these strains as a safe microbial agents in mushroom brown blotch disease management. • The non-volatile compounds of endofungal bacteria significantly inhibited the biofilm formation by P. tolaasii. • Endofungal bacteria belongs to Pseudomonas, Bacillus and Pantoea genus significantly reduced mushroom brown blotch symptoms. • Bacillus and Pseudomonas strains detoxify tolaasin produced by Pseudomonas tolaasii under in vivo conditions. • The relative expression level of AbPPO3 gene decreased in the mushroom tissues pretreated with endofungal bacteria. [ABSTRACT FROM AUTHOR]