[Objective] Riemerella anatipestifer (RA) is a Gram-negative pathogen that can cause duck serositis. The type Ⅸ secretion system (T9SS) of this bacterium is involved in sliding and pathogenic processes. The previous study showed that the expression ofB739_0093 inR.anatipestifer CH-1 was significantly up-regulated under the iron-limited condition. The sequence analysis showed that the protein encoded byB739_0093 contained a conserved C-terminal domain of the proteins secreted by T9SS, while its function remained unknown. This study aims to identify whether the protein encoded byB739_0093 is secreted by T9SS and the role of this protein in the pathogenesis of this bacterium. [Methods] qPCR was conducted to determine whether the transcription ofB739_0093 was regulated by iron and the ferric uptake regulator (Fur). The recombinant truncated B739_0093 protein was expressed inEscherichia coli, and the polyclonal antibody was prepared. Western blotting was employed to detect whether the protein was secreted by T9SS. Furthermore, we deletedB739_0093 from RA CH-1 and identified the role ofB739_0093 in the pathogenicity ofR.anatipestifer by virulence and colonization tests in ducklings. [Results] The expression ofB739_0093 was significantly up-regulated in the iron-restricted medium, which was mediated by Fur. Western blotting results showed that the protein encoded byB739_0093 was localized in the secretion of the parent strain RA CH-1, while it was localized in the bacterial lysate and not detected in the secretion of the T9SS-deleted strain. Compared with the parent strain RA CH-1, RA CH-1ΔB739_0093 demonstrated attenuated pathogenicity and reduced colonization ability in various tissues and organs of ducklings. [Conclusion] The protein encoded byB739_0093 is secreted by T9SS and involved in the pathogenicity ofR.anatipestifer, and its expression is regulated by iron and Fur.