Streptococcosis has been emerged as major threat for mass mortality of cultured tilapia in Bangladesh. Present study aimed to investigate the effects of probiotic bacteria Lactobacillus plantarum on biochemical composition, gut microbiota, innate immune response and disease resistance of Oreochromis niloticus against Enterococcus faecalis, the causative agent of streptococcosis in tilapia farming. A total of 60 fish were randomly distributed into six different aquaria with two distinct treatment groups, viz. control and probiotic. After 7 days acclimation, control group was fed with commercial tilapia feed while probiotic group was served with 1.02 × 109 CFU/mL per kg of L. plantarum supplemented diet for 56 days. At the end of trial, the results showed no significant changes in growth and biochemical composition of fish meat, however, the probiotic diet significantly modulated the gut microbiota, improved innate immune response and disease resistance of experimental tilapia. Significant increase in the abundance of Lactobacillus and Lactococcus and upregulation of cytokine genes were detected in post-feeding probiotic fish gut, wherein differentially abundant bacteria were found positively correlated with the immune gene expression. Furthermore, dietary supplementation of probiotic bacteria revealed significant reduction of mortality following injection with 2 × 105 CFU/mL of E. faecalis. The results of this study, suggest that dietary supplementation of L. plantarum could be used as potent probiotic to control streptococcosis in commercial tilapia farming.