A Butyrate-Producing Probiotic, Clostridium butyricum DMZ-SG06, Improves Growth Performance, Immune Function and Gut Microbiota in Largemouth Bass (Micropterus salmoides)

Abstract

Intensive aquaculture and antibiotic overuse have rendered largemouth bass (Micropterus salmoides) prone to frequent mass mortalities and economic losses. Probiotics are promising antibiotic alternatives for improving intestinal health and immunity. This study evaluated a novel butyrate-producing strain Clostridium butyricum DMZ-SG06 to elucidate its growth-promoting and immunomodulatory mechanisms. Three dietary treatments were used: control (Con, basal diet), T1 (basal diet supplemented with 1 g C. butyricum powder containing 5 × 108 CFU per 10 g diet), and T2 (basal diet supplemented with 1 mL C. butyricum suspension containing 5 × 108 CFU per 10 g diet). A total of 270 healthy juvenile largemouth bass (5.63 ± 0.03 g) were randomly allocated to three groups (90 fish per group), with each group divided into three replicate tanks (60 × 40 × 34 cm) (30 fish per tank). Fish were fed the corresponding diets to apparent satiation for a 60-day rearing period. Results showed that T2 increased weight gain rate (WGR) by 11.35% compared with control (p < 0.05, 213.90 ± 12.30% vs. 192.10 ± 10.20%) and significantly improved the specific growth rate (SGR, p < 0.05), with T2 performing best. Probiotic treatments markedly improved intestinal morphology, nonspecific immune indices, and antioxidant status (p < 0.05), with T2 exerting the most prominent effects. C. butyricum DMZ-SG06 significantly reduced the levels of pro-inflammatory cytokines. For instance, TNF-α levels in T2 decreased 1.39-fold compared with the control group (128.57 ± 4.97 pg/g vs. 178.67 ± 7.80 pg/g, p < 0.05). Meanwhile, the levels of anti-inflammatory cytokines were upregulated. For example, IL-10 levels in T2 increased 1.32-fold relative to the control group (321.89 ± 15.03 pg/g vs. 244.31 ± 7.57 pg/g, p < 0.05). Metagenomic analysis revealed reduced Acinetobacter abundance, enriched beneficial genera (Lactobacillus, Parabacteroides, p < 0.05), and enhanced microbial functions related to the phosphotransferase system and galactose metabolism (p < 0.05). In conclusion, C. butyricum DMZ-SG06 promotes largemouth bass growth and intestinal health via butyrate metabolism, immune modulation, and microbiota remodeling. Notably, the liquid bacterial suspension formulation exerts a more significant effect on enhancing the fish’s growth performance and intestinal health than the powder counterpart, supporting its application as a safe probiotic in antibiotic-free aquaculture.