During the last decade, the industrialized nations have experienced a well–publicized tuberculosis (TB) epidemic. By the same token, the occurrence of new species of nontuberculous mycobacteria (NTM) have challenged the microbiology laboratories’ capability to provide accurate results in a timely fashion. Furthermore, the economic climate puts an additional spin on the charge placed on laboratories providing clinical services.Once a diagnosis of mycobacterial disease in a patient is suspected, a series of interactions between the clinician and the laboratory are set into motion. The following three topics require attention: 1) In patients with acid–fast bacilli smear–positive sputum specimens diagnosed for the first time, the differentiation between TB and NTM is imperative (respiratory isolation, adequate drug regimen, and contact investigation); 2) in areas with increased drug resistant TB susceptibility testing results, especally for rifampin, are paramount; and 3) the emergence of clinically significant NTM, especially in immuno–comprised hosts, poses an additional challenge for patient care.