We previously described the cloning and expression of the protective antigen (PA) gene of Bacillus anthracis in both vaccinia virus and a baculovirus. The antigenicity of the PA products was characterized. PA expressed by the recombinant vaccinia viruses elicited a partial protective immune response against a lethal B. anthracis spore challenge in guinea pigs and mice. The WR strain vaccinia virus recombinant (WR-PA) protected 60% of male mice and 50% of guinea pigs. WR-PA elicited high anti-PA antibody titers in mice but not in guinea pigs. Connaught strain vaccinia virus recombinants failed to protect any immunized animals. PA purified from baculovirus recombinant-infected cultures plus adjuvant partially protected male CBA/J mice and completely protected female Hartley guinea pigs from challenge. Both the recombinant and nonrecombinant PA preparations combined with adjuvant elicited high anti-PA antibody titers in Hartley guinea pigs and CBA/J mice. These data demonstrate that the recombinant baculovirus- and vaccinia virus-produced PAs were immunogenic in both guinea pigs and mice, that the baculovirus-PA recombinant was a useful source of immunogenic PA, and that vaccinia virus-PA recombinants may be feasible live anthrax vaccine candidates worthy of consideration for further development as live vaccines.