Aquaculture has become increasingly important for meeting global seafood demand, but intensive tilapia farming remains constrained by bacterial diseases and the risks associated with antibiotic-dependent disease management. Probiotics, particularly lactic acid bacteria (LAB), offer a potential sustainable alternative for improving fish health and reducing antibiotic reliance. This study evaluated the probiotic potential of kefir-derived Lacticaseibacillus paracasei on antimicrobial activity, biosafety, viability, growth performance and intestinal health. The bacteria were screened for antimicrobial properties via well diffusion assay and for biosafety via brine shrimp lethality assay prior to feeding trial where Nile tilapia were assigned to three dietary groups: negative control without probiotic supplementation, 1 × 10⁷ CFU/mL L. paracasei, and 1 × 10⁸ CFU/mL L. paracasei respectively. Growth performance, feed conversion ratio, intestinal histomorphology, external pathology, liver and spleen histopathology were measured and compared either throughout or after 8 weeks. In vivo, L. paracasei supplementation did not significantly improve specific growth rate or feed conversion ratio (P > 0.05).  The 10⁷ CFU/mL group had significantly greater villus length, villus width, crypt depth, muscularis thickness, and goblet cell number compared with both the control and 10⁸ CFU/mL groups (P < 0.05), while the control and 10⁸ CFU/mL groups did not differ significantly. Additionally, Probiotic viability was monitored over eight weeks of refrigerated storage where viability declined gradually during refrigerated storage, but both 10⁷ and 10⁸ CFU/mL preparations remained within potentially functional probiotic levels after eight weeks.