A collection of 68 Hafnia strains previously identified to the species level by 16S rRNA gene sequencing were investigated for simple phenotypic properties that could aid in their recognition in the clinical laboratory. Four tests, including malonate utilization, fermentation of salicin and d-arabinose, and expression of β-glucosidase activity, correctly assigned each strain to either Hafnia alvei or H. paralvei. Antibiotic susceptibility profiles were generated for 35 H. alvei and H. paralvei isolates using Etest strips for 24 antibiotics. All strains were susceptible to aminoglycosides, quinolones, carbapenems, and monobactams. Most of the Hafnia isolates had a colistin MIC of ≥2 μg/ml. Sequencing of an internal ampC gene fragment allowed genotypic differentiation of the two Hafnia species. Approximately 70% of the hafniae tested additionally produced a cytolytic toxin active on Vero cells which may play a role in gastroenteritis.