Disease prevention and control are crucial for the development of intensive commercial aquaculture. For some fish species, several vaccines are available to contain the effects of pathogens. However, in Nile tilapia there are just a few vaccines available. The current study compared the efficacy of three inactivated vaccines against Streptococcus agalactiae; in Nile tilapia. Formaldehyde, hydrogen peroxide and pH manipulation were used to inactivate the bacteria. Nile tilapia fingerlings were distributed into six experimental groups in duplicate: T1: naïve group; T2: sham group (PBS-injected, non-challenged); T3: PBS-injected control group (PBS-injected, challenge-infected); T4: vaccinated with hydrogen peroxide-inactivated vaccine (vaccinated, challenge-infected); T5: vaccinated with formaldehyde-inactivated vaccine (vaccinated, challenge-infected); T6: vaccinated by pH manipulation (vaccinated, challenge-infected). Twenty-eight days after vaccination, groups T3, T4, T5 and T6 were challenged intraperitoneally with 0.1 mL of the bacteria and monitored for 21 days. Additionally, replicates of non-challenged fish were used for determination of total serum protein and anti-Streptococcus agalactiae IgM antibodies. The percentage of survival of the groups T1, T2, T3, T4, T5 and T6 were 100%, 100%, 62.5%, 80%, 75% and 85%, respectively. The group vaccinated by pH manipulation presented slight higher survival when compared to the control group (P > .05). However, there were no differences in survival between vaccinated groups. The relative percentage survival (RPS) in groups T4, T5 and T6 were 46.6%, 33.3% and 60%, respectively. No differences were observed for total serum proteins and anti-Streptococcus. agalactiae IgM antibodies. The results observed in this study indicate equivalent moderate vaccine efficacy between alternative inactivated (hydrogen peroxide and pH), and conventional formaldehyde vaccines, with slight higher values for the pH manipulated vaccine, which suggests that future studies should be performed for each alternative method in order to improve their performance.