Francisella tularensis subspecies novicida mutants have been made with deletions introduced into the purA or purF genes. These mutants demonstrated the expected growth requirement for purines and complementation with the wild type genes restored the ability to grow on purine deficient media. The mutants were at least 10,000-fold attenuated by the ip challenge route in Balb/C mice and defective for survival in J774A.1 mouse macrophages. Immunisation with the purA mutant did not provide protection against a subsequent challenge with 100 median lethal doses of F. tularensis subspecies novicida. Immunisation of mice with the purF mutant provided protection against a subsequent challenge with F. tularensis subspecies novicida but not against a subspecies tularensis challenge. These findings suggest that purine auxotrophs of F. tularensis should be further evaluated as live attenuated vaccines against tularemia, but that differential effects are seen depending on which step in the biosynthetic pathway is inactivated.