Fish vibriosis is among the most common diseases that is caused by a bacteria belonging to the genus Vibrio (Vibrio anguillarium). It causes considerable economic loss in the commercial cultivation of Asian sea bass (Lates calcarifer). The resistance of microorganisms to antibiotics has resulted in a growing need for developing a new antibacterial therapy that is effective in aquaculture. The aim of this study is to develop neem nanoemulsion with antibacterial activity against V. anguillarium to identify a possible alternative to the commonly used antibiotics in aquaculture. Neem nanoemulsion was prepared and the effectiveness was studied both in vitro and in vivo (agar well diffusion assay and artificial infection). Injection and immersion challenge of neem nanoemulsion formulated the fish less susceptible to V. anguillarium infection. The results confirmed the potential use of neem nanoemulsion as a source of antibacterial compounds or as a health-promoting medicine for fish culture. © 2019 National Institute of Science Communication and Information Resources (NISCAIR). All rights reserved.