Ornithobacterium rhinotracheale is a Gram-negative bacterium associated with respiratory diseases in many avian species, with worldwide distribution, and it causes significant economic loss to the poultry industry. In this study, the isolation and characterization of O. rhinotracheale small-colony variants (SCVs) are described for the first time. O. rhinotracheale isolates (n = 27) were recovered from tracheal samples (n = 321) collected from different avian species with clinical signs of respiratory disease. Of the 27 O. rhinotracheale isolates, 21 (77.8%) showed SCVs in their primary cultures. Five O. rhinotracheale SCV isolates showed high levels of stability and were chosen for further characterization with their wild-type (WT) isolates. Stable O. rhinotracheale SCVs were oxidase negative, while their WT isolates were positive. Growth curves for stable O. rhinotracheale SCVs indicated lower growth rates and longer lag phases than for their WT isolates. Furthermore, it was possible to increase the efficacy of the broth medium in supporting the growth of O. rhinotracheale WT isolates by supplementing it with 5% fetal bovine serum (FBS) and 2% IsoVitaleX Enrichment. Antibiotic susceptibility tests showed that O. rhinotracheale SCVs had higher MIC values than their WT isolates. This study suggests that successful antibiotic treatment of respiratory diseases associated with O. rhinotracheale must take into consideration the resistance patterns of O. rhinotracheale SCVs. Intracellular persistence in murine RAW 264.7 macrophages revealed that O. rhinotracheale SCV28 had higher survival rates than its WT isolate. Finally, small-colony variants may be important contributors to the pathogenesis of O. rhinotracheale.