Members of the M. avium complex are over 95% homologous and differentiation between them is notoriously slow and difficult. Particular difficulties are encountered in discriminating M. paratuberculosis from other mycobactin-dependent members of the group eg wood pigeon bacillus (syn. M. a. silvaticum). We have identified a 40kDa protein that appears to be expressed in IS901/902+ strains of Mycobacterium avium but not in Mycobacterium paratuberculosis. The amino terminal twenty amino acids of the protein share no significant homology with any proteins currently available in the OWL database. The protein stimulates strong cell-mediated and humoral immune responses in sheep and deer. Such responses are observed only in animals infected with IS901/902+ strains of M. avium. The gene encoding the 40kDa protein has been isolated from a gtll expression library of M. avium and is currently being characterized.