A polyphasic taxonomic study, using phenotypic and phylogenetic characterization as well as genotypic methods, was performed on five unknown Gram-positive, catalase-negative, coccus-shaped Vagococcus-like bacteria isolated from the spoilage microbiota of cooked shrimp. Comparative 16S rRNA gene sequence analysis indicated that these strains belonged to the genus Vagococcus and that it formed a new branch distinct from the six species already recognized (V. fluvialis, V. salmoninarum, V. lutrae, V. fessus, V. carniphilus and V. elongatus). The polyphasic approach on the representative strain CD276T, including DNA–DNA hybridizations, pulsed-field gel electrophoresis of whole-genome DNA profiles, G+C content determination, cell-wall peptidoglycan typing, fatty acid analysis and extensive biochemical characterization, was used to clarify the taxonomic position of this strain. These data confirmed that CD276T (LMG 24833T = CIP 109914T) represents the type strain of a novel species, for which the name Vagococcus penaei sp. nov. is proposed.