The most prevalent diseases in fish farming are bacterial infections, which drive considerable financial losses to the aquaculture sector. Figuring out an innovative, eco-friendly antimicrobial is becoming more crucial, especially in light of the escalating resistance of pathogens to the existing pharmaceuticals. Accordingly, this work is the inaugural attempt to look into the antibacterial attributes of a flavanone [naringenin (NG) dissolved in dimethyl sulfoxide (DMSO)] in assisting Oreochromis niloticus (Nile tilapia) to combat a bacterial challenge (Proteus mirabilis). This study was carried out in two phases (in vitro and in vivo). The in vitro anti-bacterial activity, minimum inhibitory concentration, and minimum bactericidal concentration of NG against P. mirabilis were 19.00 ± 1.50 mm, 15.62 μg/mL, and 31.25 μg/mL, respectively. In vivo, fish (n = 250; 24.30 ± 1.12 g) were divided into five treatments for 15 days [control, DMSO (1 %), NG (17.50 mg/L), P. mirabilis (intraperitoneally injected with 100 μL bacterial suspension containing 1.50 × 108 CFU/mL), and NG + P. mirabilis]. Findings outlined that P. mirabilis infection diminished fish survival (60.00 %) and immune response, proven by a significant decline in the level of immunoglobulin M, complement 3, globulin, albumin, and total protein, and the elevation of the C-reactive protein value (P < 0.001). A noteworthy drop in the antioxidant parameters (superoxide dismutase and reduced glutathione content) and a rise in malondialdehyde (lipid peroxide) were obvious in the P. mirabilis group (P < 0.001). The histopathological assay proved the depletion of white pulps around ellipsoids and the widening of splenic sinusoids by P. mirabilis infection. Furthermore, P. mirabilis induced a remarkable up-regulation in the expression of β-catenin-1 and tilapia piscidin 3 genes and down-regulation in the immune-related genes involving interleukin 10 and transforming growth factor beta-1a (P < 0.001). An up-regulation of the pathways linked with endoplasmic reticulum (ER) stress [protein kinase RNA-like endoplasmic reticulum kinase (PERK) and glycogen synthase kinase 3 beta (GSK3-b)] and apoptosis [cysteine-aspartic acid protease-3a (CASP-3a)] was evident in the P. mirabilis-infected fish. Interestingly, administering NG reversed all the adverse implications of P. mirabilis infection besides boosting fish survival (86.00 %). In addition, NG decreased bacterial load in the spleen of the NG + P. mirabilis group (0.38 ± 0.02 × 108 CFU/g tissue), contrasted with the P. mirabilis group (7.47 ± 0.73 × 108 CFU/g tissue). Moreover, the molecular docking modeling highlighted a direct suppressive effect of NG for PERK, GSK3-b, and CASP-3a with a potent binding affinity of −8.40, −7.50, and −8.30 kcal/mol, respectively. Inclusive, this trial points out the fundamental role of NG as an outstanding antibacterial, emphasizing its capacity to enhance immune-antioxidant response and safeguard O. niloticus against P. mirabilis infection for sustainable aquaculture.