Aims: Characterization of nonmotile bacteria associated with freshwater fish spoilage and that phenotypically resembled Psychrobacter spp. Methods and Results: A population of 44 nonmotile Gram-negative rods could not be assigned to the genus Psychrobacter on the basis of a definitive test (transformation assay). Conventional and commercial phenotypic systems did not help in identification. A second extensive phenotypic analysis using different temperatures and media confirmed these isolates as nonmotile although electron microscopic examination showed that all but two had one to four polar flagella and other appendages. On the basis of numerical taxonomy, this population was divided into six clusters, one of them consisting of five fluorescent strains. Sequencing of fluorescent and non fluorescent representative strains from each cluster demonstrated that strains from five clusters had between 97.8 and 98.8% sequence homology with Pseudomonas fragi IFO 3458. This and an unknown strain from deep sea were the closest organisms (80.9% sequence homology) to one aflagellated representative strain of the remaining cluster. Conclusions: Oxidase-positive, nonmotile, nonfermenter Gram-negative rods isolated from freshwater fish can be wrongly ascribed to the genus Psychrobacter. Significance and Impact of the Study: Molecular methods are necessary for the identification of environmental isolates and species with an incomplete phenotypic description. This work emphasizes the need for a sound description of Ps. fragi based on molecular and phenotypic characterization.