The genetically engineered, live-attenuated Venezuelan equine encephalitis (VEE) virus vaccine candidate, V3526, was evaluated as a replacement for the TC-83 virus vaccine. Protection from lethal subcutaneous or aerosol challenge was evaluated in vaccinated mice clinically and immunohistochemically. Subcutaneous administration of V3526 induced systemic and mucosal protection more efficiently than did the TC-83 vaccine. The bronchial IgA responses induced in mice by subcutaneous administration of vaccines significantly corresponded to the ability to survive aerosol challenge with virulent virus. Furthermore, V3526 delivered by aerosol induced more complete mucosal protection than either vaccine administered subcutaneously. The ability of V3526 to induce protection in mice warrants its consideration for further testing as a potential vaccine candidate for human use.