Mycobacterium bovis causes tuberculosis in cattle. Because current therapies inadequately prevent disease spread, new therapies for M. bovis infections are urgently needed. We treated M. bovis-infected cattle with combinations of DNA vaccines, resulting in significantly reduced pulmonary bacterial loads compared with the untreated group. Further, the pathological changes of lungs were also slight and the lesions were limited in animals treated with the combined DNA vaccines, whereas untreated animals had extensive lung fibrosis and consolidated lesions. This is the first report of DNA vaccine used to treat M. bovis. Thus, combined DNA vaccines may provide enhanced immunotherapeutic efficacy for treatment of bovine tuberculosis.