Tuberculosis is one of the most prevalent infectious diseases worldwide and is one of the main causes of mortality by infection. In 2011, the World Health Organization (WHO) estimated 8.7 million cases of tuberculosis, 13% of which were co-infected with the Human Immunodeficiency Virus (HIV). Two billion people present latent tuberculous infection, which corresponds to one-third of the world population. Thus, it constitutes a serious public health problem, which is why the WHO has declared it to be a global emergency. Traditionally, childhood tuberculosis diagnosis requires the presence of various criteria including epidemiological, clinical, radiological, tuberculinic, and microbiological criteria, the latter of which consists of conventional methods, such as culture, which is the gold standard, and nucleic acid amplification techniques, which are more common as they offer faster and a more accurate diagnosis; this is because the tubercle bacillus is a slow-growing microorganism and requires a minimum of 104 bacilli in the culture sample, which makes it difficult to isolate in paucibacillary patients and impedes early diagnosis. The new methods of molecular biology have become a good option for diagnosis and treatment of infections produced by the Mycobacterium tuberculosis complex and nontuberculous mycobacteria, given the speed of results, its high sensitivity and specificity, and the ability to detect resistance to anti-tuberculosis drugs.