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It is estimated that one third of the world population is infected with Mycobacterium tuberculosis, resulting in 2 million deaths annually. More than 8 million new cases of tuberculosis (TB) are registered per year worldwide, and Brazil ranks 19th among the top 23 countries with the highest rates of TB. The determining factors for the control of this disease include early detection, appropriate therapy and measures for avoiding transmission. The conventional diagnoses (smear and microorganism culture) have time limitations for implementation and operation, since the result may take up to 60 days to be released. Therefore, early detection is critical for blocking the chain of TB transmission. Mycobacterial diseases caused by nontuberculous mycobacteria are also having a major impact due to increased outbreaks of surgical infections. Thereby, the rapid and specific identification of microorganisms is important for the diagnosis, which will determine the type of treatment (treatment according to species). Knowledge of etiologic agents of mycobacterial diseases, as well as sensitive and specific diagnosis allows proper treatment by blocking the chain of TB transmission and controlling nontuberculous mycobacteria outbreaks.