The Shewanella putrefaciens group are ubiquitous microorganisms recently isolated from different freshwater fish species and causing serious health disorders. The purpose of the study was to characterise isolates of the S. putrefaciens group with special emphasis on elucidating serological diversity and determining putative virulence factors.Isolates collected from freshwater fish (n = 44) and reference strains were used. The identification of bacteria was carried out using biochemical kits and 16S rRNA sequencing. Polyclonal antibodies were prepared against the S. putrefaciens group. The bacterium’s susceptibility to antimicrobial agents, its enzymatic properties, and its adhesion ability to fish cell lines were also tested. Finally, selected isolates were used in challenge experiments in common carp and rainbow trout.Excluding six isolates undeterminable for species, the bacteria were classified to three species: S. putrefaciens, S. xiamenensis, and S. oneidensis, and showed some phenotypic diversity. Fourteen serological variants of the S. putrefaciens group were determined with the newly developed serotyping scheme.Serodiversity may play an important role in the virulence of particular isolates. Further, S. putrefaciens group members adhere to epithelial cells and produce enzymes which may contribute to their virulence. Challenge tests confirmed the pathogenicity of the S. putrefaciens group for fish.