Introduction Citrobacter freundii strains, like other representatives of the same genus, are often described as aetiological factors of diseases in humans and animals, including farmed and ornamental fish. The aim of the study was the isolation and identification of C. freundii from ornamental fish in Poland and investigation of the virulence factors and antibiotic resistance among the isolates. Material and Methods Twenty C. freundii isolates were isolated from swab samples obtained from diseased ornamental fish of 16 different species in Poland. The bacteria were initially identified to the species level by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry, and the results were confirmed based on sequencing of a fragment of the 16S rRNA gene and biochemical profile analysis using the API-20E system. The antibiotic susceptibility profile was evaluated by the disc-diffusion method. Results Most isolates were resistant to tetracycline (65%), ciprofloxacin (65%), nalidixic acid (60%) and doxycycline (50%). All of them were resistant to ampicillin. As many as 13/20 isolates (65%) carried the viaB gene encoding the Vi antigen and 9/20 (45%) carried the class 1 integron-encoding gene. The hlyA gene encoding the ability to produce α-haemolysins was not detected in any of the isolates tested. Conclusion This is the first study to describe in detail the identification and occurrence of virulence factors and antibiotic resistance among C. freundii isolates of ornamental fish in Poland. The results indicate the need for further monitoring of the bacterium’s presence.