Vibrio splendidus is a common inhabitant of marine and brackish water associated with disease outbreaks in cultured fish and bivalves. In bivalve aquaculture, V. splendidus is considered the most important bacterial pathogen responsible for severe financial losses, while in fish culture it has been reported to cause significant mortalities mostly in turbot larvae. Phages have been considered as a promising alternative to antibiotics, especially in areas where the use of antibiotics presents specific drawbacks and may not be applicable, like fish and bivalve larviculture. Here we report the isolation and characterization of pVa5, a lytic phage which infects several V. splendidus strains. pVa5 was isolated using as host an environmental V. splendidus strain obtained from mussels of a biofouled fish cage in Greece during a vibriosis incidence. Following TEM observations, pVa5 was classified to podoviridae family. It had a head measuring 85 μm in length and a short (25 μm) tail. One-step growth curve analysis showed that pVa5 had a latent period of 30 min and a large burst size of 285 virions per infected cell. The phage produced small and clear plaques on the host lawn while in vitro lysis assay indicated that it could significantly reduce bacterial growth for 8 hours even at low MOI (=1). Spot test showed that the phage could infect several V. splendidus strains but no other vibrio species. The genome of pVa5 was fully sequenced with a GS Junior System and was de novo assembled with GS Assembler software. It had a genome size of 78145 bp, 43% G+C content and 106 putative open reading frames of which 30 encode proteins of known function. Genome structure is similar to the N4-like phages while no temperateness-related nor toxin-encoded genes where found suggesting that pVa5 could be safe for use in phage therapy applications.