Tuberculosis (TB) affects around 10 million people worldwide in 2019. Approximately 3.4 % of new TB cases are multidrug-resistant. The gold standard method for detecting Mycobacterium tuberculosis, which is the aetiological agent of TB, is still based on microbiological culture procedures, followed by species identification and drug sensitivity testing. Sputum is the most commonly obtained clinical specimen from patients with pulmonary TB. Although smear microscopy is a low-cost and widely used method, its sensitivity is 50–60 %. Thus, owing to the need to improve the performance of current microbiological tests to provide prompt treatment, different methods with varied sensitivity and specificity for TB diagnosis have been developed. Here we discuss the existing methods developed over the past 20 years, including their strengths and weaknesses. In-house and commercial methods have been shown to be promising to achieve rapid diagnosis. Combining methods for mycobacterial detection systems demonstrates a correlation of 100 %. Other assays are useful for the simultaneous detection of M. tuberculosis species and drug-related mutations. Novel approaches have also been employed to rapidly identify and quantify total mycobacteria RNA, including assessments of global gene expression measured in whole blood to identify the risk of TB. Spoligotyping, mass spectrometry and next-generation sequencing are also promising technologies; however, their cost needs to be reduced so that low- and middle-income countries can access them. Because of the large impact of M. tuberculosis infection on public health, the development of new methods in the context of well-designed and -controlled clinical trials might contribute to the improvement of TB infection control.