<p>Bovine norovirus (BNoV) and bovine nebovirus (BNeV) are emerging pathogens responsible for severe diarrhea in calves worldwide. In this study, we constructed a bivalent recombinant adenovirus vector vaccine, designated rAd5-BNoV + BNeV_VP1 encoding the genes for BNoV-VP1 and BNeV-VP1 based on replication-deficient human adenovirus type 5 (rAd5), and evaluated its immunogenicity in mice and calves. Serum antibody responses were assessed by enzyme-linked immunosorbent assay (ELISA) and serum blocking titer 50 (BT50). Results demonstrated that mice immunized with rAd5-BNoV + BNeV_VP1 produced significant VP1-specific IgG titers as early as two weeks post-vaccination. BNoV-VP1-specific IgG antibody titers peaked at 1:10⁵ in both intramuscular (I.M.) and oral groups (<i>n</i> = 6), while BNeV-VP1-specific IgG titers peaked at 1:10⁵ in the I.M. groups (<i>n</i> = 6) and 1:10⁴ in the oral groups (<i>n</i> = 6). The BT50 values peaked at 640 in the I.M. groups and 576 in the oral groups (<i>n</i> = 6). Furthermore, the percentages of IL-4-producing cells and CD3⁺CD8⁺ T cells in splenocytes were significantly elevated in mice immunized via the I.M. route. The vaccine also elicited a robust humoral immune response in calves. Following a booster I.M. injection with rAd5-BNoV + BNeV_VP1, serum BNoV-VP1-specific and BNeV-VP1-specific IgG antibody titers were significantly higher (<i>P</i> ≤ 0.001) compared to controls. The BT50 in immunized calves reached 1:80 at 21 and 35 days post-immunization. These results collectively indicate that rAd5-BNoV + BNeV_VP1 effectively induces immunity against BNoV and BNeV infection.</p>