Abstract: To investigate the mitigating effects of Bacillus inaquosorum MC2-1 (hereinafter referred to as MC2-1) on tissue damage, antioxidant capacity, ion transport, and intestinal flora under acute salt-alkaline stress in largemouth bass (Micropterus salmoides), a one-week oral administration trial was conducted. Largemouth bass juveniles mean initial weight (8.56±0.04) g were randomly divided into five treatments: a freshwater control (CK), a saline-alkaline stress group (SC; 6 psu salinity, 4.5 mmol/L alkalinity), and three probiotic-supplemented groups receiving SC water plus MC2-1 at 2×10⁶, 2×10⁷ and 2×10⁸ CFU/tail (LT, MT, and HT, respectively). The results revealed that compared to the CK group, the SC group exhibited a significantly thinner intestinal muscle layer (P<0.05), shorter the secondary gill arch, larger gill lamella interval and wider short diameter, and increased severity of hepatic lesions including vacuolization and nuclear shift. Conversely, MC2-1 administration significantly restored intestinal muscle thickness and markedly mitigated liver damage induced by saline-alkaline stress, and the diameter and short diameter of the secondary gill arch in the MT and HT groups were restored (P<0.05). Regarding antioxidant and immune responses, the SC group significantly elevated hepatic T-AOC but decreased renal SOD activity and branchial T-AOC (P<0.05). Notably, the MT group significantly enhanced branchial SOD activity, and intestinal AKP and ACP activities compared to the CK group. Furthermore, compared to the SC group, the HT group significantly increased liver CAT and branchial SOD activities, with intestinal AMS and LPS activities being significantly higher in the HT group than that in all other groups (P<0.05). In terms of serum biochemistry, salt-alkaline stress led to a significant increase in TP and Ca²⁺ levels (P<0.05). Importantly, supplementation in the HT group successfully restored serum Cl^- and Ca²⁺ concentrations to levels comparable to those in the control group (P<0.05). Analysis of the gut microbiota showed that the relative abundance of Paracoccus was significantly higher in the CK group than that in other groups (P<0.05). Compared to the SC group, the HT group showed increased relative abundances of Aurantimicrobium, Hyphomicrobiales, and Bacillus; however, the relative abundance of the pathogenic bacterium Plesiomonas was also significantly elevated in this group (P<0.05). In conclusion, oral administration of MC2-1 can effectively alleviate histological damage to the gills, liver, and intestines of largemouth bass under acute salt-alkaline stress. Specifically, supplementation at 2×10⁷—2×10⁸ CFU/tail significantly improves the liver and branchial antioxidant systems, enhances intestinal immune parameters, and modulates the gut microbiota, thereby collectively enhancing the salt-alkaline tolerance of the fish.