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.