This study assessed the impact of dietary supplementation of single‐ and multi‐strain Bacillus‐based probiotics on growth performance, intestinal morphometry, blood parameters, immune‐oxidative status and transcriptomic response of Nile tilapia for 60 days. Fish (N = 150, 1.632 ± 0.035 g) were allocated into five experimental groups: (1) Control group (CG) that was fed on a basal diet; (2) Eco. 0.01%, received basal diet with 0.1 g/kg Ecobiol; (3) Eco. 0.02%, received basal diet with 0.2 g/kg Ecobiol; (4) Pro‐F FMC 0.001%, received basal diet with 0.001 g/kg Sanolife PRO‐F FMC; (5) Pro‐F FMC 0.002%, received basal diet with 0.002 g/kg Pro‐F‐FMC. Fish received Pro‐F FMC 0.002% displayed higher growth performance than CG (p < 0.05), while Pro‐F FMC 0.001% exhibited the highest intestinal weight. Sanolife PRO‐F FMC supplementation improved intestinal villi length and goblet cell number while decreased inter‐villi space (p < 0.05). The best haemoglobin, haematocrit and monocyte per cent were observed in Pro‐F FMC 0.001%, whereas Pro‐F FMC 0.002% revealed the best leukocyte, lymphocyte and neutrophil contents. Pro‐F FMC‐supplemented groups showed better phagocytic activity, phagocytic index and lysozyme activity than Ecobiol‐supplemented groups and CG. Pro‐F FMC 0.001% enrichment induced the highest upregulation level of NPYα, AgRp‐2, ghrelin and GH‐1, whereas Pro‐F FMC 0.002% significantly upregulated IGF‐1 and IGF‐2 gene expression. Eco. 0.02% induced the highest upregulation level of orexin and galanin gene expression. Conclusively, dietary enrichment with single‐ or multi‐strain Bacillus‐based probiotics is likely to improve growth, immunological parameters and transcriptomic response of Nile tilapia.