Using an adult mouse model to study active immunity against rotavirus infection, it was previously shown that oral immunization with some, but not all, animal rotavirus strains induced protection against subsequent infection following oral challenge with the murine rotavirus strain EDIM. To determine if a specific rotavirus protein could be associated with protection in this model, mice were immunized with a series of 18 reassortants between the fully protective EDIM strain and a partially protective heterologous rotavirus strain (RRV-G). Reassortants that contained genes for EDIM proteins responsible for protection were anticipated to provide complete protection; however, no EDIM proteins were found to be both necessary and sufficient for full protection. Instead, protection was found to be highly correlated with viral shedding (P = 0.005) and with serum rotavirus IgA titers stimulated by the different reassortants (P < 0.001). This indicated that protection was related to the intestinal replication properties of the different reassortants rather than to specific immunogenic properties of EDIM proteins. This conclusion was supported by the finding that the titers of serum rotavirus IgA, but not IgG, stimulated in mice following oral immunization with a series of animal rotaviruses was directly related to protection against EDIM. If these findings can be extended to humans, they suggest that the efficiency of intestinal replication following oral inoculation with a live rotavirus vaccine candidate may be the primary determinant of successful immunization.