An eight-week feeding trial was conducted to evaluate the effects of a basal control diet (CON), Bacillus subtilis at 107 (BS7) and at 108 CFU/g diet (BS8), Lactococcus lactis at 107 CFU/g (LL7) and at 108 CFU/g diet (LL8), and oxytetracycline (OTC) at 4 g/kg diet on Nile tilapia. Fish with initial body weight of 2.83 ± 0.05 g (mean ± SD) were fed two times a day. Weight gain, specific growth rate, feed efficiency, protein efficiency ratio and lysozyme activity of fish fed BS8, LL8 and LL7 diets were significantly higher than those of fish fed CON diet (p < 0.05). Superoxide dismutase and myeloperoxidase activity of fish fed BS8, LL8, BS7, LL7 and OTC diets were significantly higher than those of fish fed CON diet. Intestinal villi length and muscular layer thickness of fish fed BS8, LL8 and LL7 diets were significantly higher than those of fish fed CON and OTC diets. Also, heat shock protein 70 (HSP70), interleukin (IL-1β), interferon-gamma (IFN-γ) and tumour necrosis factor (TNF-α) gene expression of fish fed BS8 and LL8 diets were significantly higher than those of fish fed CON diet. After 13 days of challenge test, cumulative survival rate of fish fed BS8 and LL8 diets were significantly higher than those of fish fed CON, BS7 and OTC diets. Based on these results, B. subtilis and L. lactis at 108 (CFU/g) could replace antibiotics, and have beneficial effects on growth, immunity, histology, gene expression, and disease resistance in Nile tilapia.