Mycobacteria and their cell wall components have been used with varying degrees of success to treat tumors, and Mycobacterium bovis BCG remains in use as a standard treatment for superficial bladder cancer. Mycobacterial immunotherapy is very effective in eliciting local immune responses against solid tumors when administered topically; however, its effectiveness in eliciting adaptive immune responses has been variable. Using a subcutaneous mouse thymoma model, we investigated whether immunotherapy with Mycobacterium smegmatis, a fast-growing mycobacterium of low pathogenicity, induces a systemic adaptive immune response. We found that M. smegmatis delivered adjacent to the tumor site elicited a systemic anti-tumor immune response that was primarily mediated by CD8(+) T cells. Of note, we identified a CD11c(+)CD40(int)CD11b(hi)Gr-1(+) inflammatory DC population in the tumor-draining lymph nodes that was found only in mice treated with M. smegmatis. Our data suggest that, rather than rescuing the function of the DC already present in the tumor and/or tumor-draining lymph node, M. smegmatis treatment may promote anti-tumor immune responses by inducing the involvement of a new population of inflammatory cells with intact function.