One of the main factors limiting tilapia’s production is the occurrence of infections caused by Aeromonas and Streptococcus species. This work intended to evaluate a bivalent vaccine against A. hydrophila and S. agalactiae by intraperitoneal (i.p) administration in Nile tilapia (Oreochromis niloticus) in Brazil. The study was carried out in two phases: one in the laboratory, on a small scale, and from the results obtained, the study was expanded to a large scale in a production system in cages. The vaccine proved to be safe and effective in laboratory tests, with a vaccine efficacy (VE) of 93.66%. However, in large-scale tests with 12,000 tilapias, the VE was 59.14%, with a better food conversion ratio (1.54 kg) in the vaccinated group compared to the control group (1.27 kg). These results corroborate the efficiency of this tested vaccine; however, they indicate the need for field tests to attest to real protection.