Brown blotch disease of Pleurotus ostreatus, caused by Pseudomonas tolaasii, substantially restricts mushroom production. Current control methods have drawbacks, including chemical residues and pathogen resistance. To address this issue, we isolated a bacterium from surface-sterilized, mildly diseased fruiting bodies, designated strain 24b, and evaluated its biocontrol potential against P. tolaasii. In vitro dual culture assays showed that strain 24b produced a stable inhibition zone against P. tolaasii. In bag cultivation experiments, preventive application of strain 24b prior to pathogen inoculation significantly reduced disease severity, with control efficacy markedly higher than that of curative application. The strain was identified as Delftia acidovorans by 16S rRNA gene sequencing and whole-genome analysis. Genomic analysis revealed multiple secondary metabolite biosynthetic gene clusters, including a complete delftibactin A/B cluster associated with siderophore synthesis. The presence of these clusters may provide a genetic basis for the observed antagonistic activity. Safety assessments showed no hemolytic activity or genes encoding known virulence factors. These findings indicate that D. acidovorans strain 24b is a promising biocontrol agent against brown blotch disease, potentially acting through niche competition and antibiosis.