Between June 2017 and October 2023, five cases of epididymitis caused by Pasteurella multocida were confirmed in four Japanese Black calves and one Japanese Black-Holstein crossbreed calf across five farms in three prefectures in Japan. Although P. multocida is primarily known to cause respiratory diseases in cattle, its role in reproductive disorders, particularly in epididymitis, has not been extensively studied. Histopathological examination showed pyogranulomatous lesions in the epididymal duct and surrounding areas, consisting of ductal extension, immune cell infiltration, epithelial degeneration, abscess formation, and fibrosis with angiogenesis. In the immunohistochemical analysis, positive reactions to anti-P. multocida serotype A antibodies were observed in macrophages and neutrophils in the epididymal ducts and surrounding areas, suggesting an ascending infection rather than hematogenous spread. This report is the first to directly confirm P. multocida involvement in bovine epididymitis through immunohistochemical examination. P. multocida was isolated in pure culture from the scrotal contents in all cases. The isolates were grouped into sequence types ST79 and ST13, commonly found in the respiratory tract and oral cavity of cattle, using the Rural Industries Research and Development Corporation multilocus sequence typing scheme
pulsed-field gel electrophoresis revealed unique band patterns in the isolates from each case. These results suggest that multiple genetically distinct strains, commonly found in the nasal and oral cavities, incidentally infected the epididymis via an ascending route. This bacterium warrants further investigation as a potential cause of reproductive disorders, particularly in Japanese Black cattle.