Three vaccines developed for protection against IA/IB subtypes of Venezuelan equine encephalitis (VEE) virus were evaluated in mice for the ability to protect against systemic and mucosal challenges with a virulent virus of the IE subtype. The vaccines were the formaldehyde-inactivated C-84 and live attenuated TC-83 vaccines currently administered to people under investigational new drug (IND) status, and a new live attenuated vaccine candidate, V3526. V3526 was superior for inducing protection to VEE IA/IB within a week of vaccination, and protection persisted for at least a year. All three vaccines induced long-term clinical protection against peripheral or mucosal challenge with IE virus, with the mucosal immunity induced by attenuated vaccines lasting longer than that induced by the inactivated vaccine. These data show that the molecularly cloned V3526 vaccine induces equivalent or improved immunity to homologous and heterologous VEE viruses than the existing vaccines.