The protective efficacy of Mycobacterium bovis-bacille Calmette Guérin (BCG) against tuberculosis (TB) is variable in both humans and cattle. Exposure to environmental mycobacteria is thought to result in inappropriate priming of host immune responses. To investigate the impact of environmental mycobacteria on BCG efficacy, cattle were infected with M. avium, vaccinated with BCG, challenged with M. bovis and skin tested prior to necropsy. Elevated levels of IFNγ were evident in M. avium-exposed animals before and after BCG vaccination with a bias towards avian purified protein derivative (PPD-A), suggesting that M. avium primed host immune responses. Exposure to M. avium also resulted in a higher frequency of circulatory IFNγ-producing cells in response to PPD antigens at the time of M. bovis challenge. After M. bovis inoculation, the IFNγ response to bovine PPD (PPD-B) increased compared to pre-challenge levels, indicating that all animals had been exposed to M. bovis. Skin test responses indicated 2/6 M. avium-BCG-M. bovis animals as reactors and 2/6 as inconclusive compared with 6/6 BCG-M. bovis animals as reactors. M. avium-exposed animals also had fewer lesions and the number of tissues containing viable M. bovis at post-mortem was significantly lower (P<0.02 compared with BCG-M. bovis animals), with two of the animals described as skin test negative with no visible lesions or viable bacteria. Thus, exposure of cattle to environmental mycobacteria such as M. avium prior to BCG vaccination did not dampen BCG-specific immune responses and resulted in lower TB pathology. However, the PPD-A bias associated with M. avium exposure is likely to undermine current TB diagnostic tests and the IFNγ test in cattle.