The course of infection with Mycobacterium tuberculosis strains H37Rv, H37Ra and their isoniazid-resistant, hydrogen peroxide-susceptible mutants in guinea-pig spleen and lung were assessed by measuring changes in number of viable bacteria during the first and second 3-day intervals after i.v. infection of normal and BCG-vaccinated animals. Vaccination had no effect on bacterial survival in the first 3 days of infection. The peroxide-susceptible mutants were killed or inhibited more than their parent strains; in normal animals this enhanced susceptibility was expressed equally during the first and second 3-day intervals while in vaccinated animals the effect was greater in the second 3-day interval. The results suggest that hydrogen peroxide is generated in significant amounts in the environment of tubercle bacilli lodged in normal tissues and in enhanced amounts when acquired immunity becomes expressed after a few days' lodgement in the tissues of vaccinated animals. Thus hydrogen peroxide resistance may contribute to virulence by protecting against both normal resident and immunologically activated macrophages.