Human tuberculosis (TB) is one of the world’s deadliest infectious diseases [1,2,3]. In 2022, the WHO reported approximately 1.6 million deaths, 10 million new infections, and an estimated one-quarter of the human population being latently infected [3]. Infection control is hampered by the limited efficacy of a 100-year-old Bacille Calmette–Guérin (BCG) vaccine and the emergence of multidrug-resistant strains against 70-year-old antibiotics, particularly those used in first-line therapy [3]. There is also a need to develop better diagnostic tools that are more sensitive, use simple instruments, are easy to handle in remote geographical areas, and are cost-effective for the rapid detection of Mtb. Moreover, although new methods are being implemented [4], the currently available methods do not allow differentiation between the different stages of infection [4,5,6,7,8].

Mycobacterium tuberculosis (Mtb) is considered the main causative agent of human TB [9]. Mtb infections primarily affect the lungs, a condition that contributes to high transmissibility by the respiratory route [9]. Although not transmitted from person-to-person, extrapulmonary TB is a second form of disease manifestation that particularly affects less immunocompetent individuals, namely, children or people who are co-infected with HIV [10,11,12]. In addition to Mtb, other pathogens can also infect humans and cause TB with similar clinical symptoms [13,14,15,16,17,18,19,20]. These include M. africanum, which is restricted to humans in West Africa [21,22,23], where it causes nearly half of all pulmonary TB [18,19,20], and the animal-adapted species M. bovis, which is estimated to cause TB in about 2% of the world’s population [24,25]. M.caprae is responsible for a smaller burden of zoonotic TB [26]. They all belong to the Mycobacterium tuberculosis species complex (MTBC), which also includes the more distant M. canettii group and additional species that cause TB in animals such as M. microti [27], M. pinnipedii, M. orygis, and M. mungi [20,23,28,29,30]. Although each of these animal MTBC variants causes TB in its host species, they may trigger slight or no disease outside of their adapted host, especially in immunocompetent hosts [13,30,31]. However, conditions such as the geographic prevalence of infected animals, close human contact, the route of transmission (milk, infected meat, or airborne droplets containing pathogens), and conditions such as HIV infection or other immunosuppressive conditions should be taken into account to assess the true threat to public health [32].

The animal-adapted species are more recent pathogens that emerged as crowd-associated diseases during the Neolithic demographic expansion along with the development of animal domestication [33]. Mtb ascended long before the establishment of crowded populations 70 thousand years ago and accompanied the exodus of Homo sapiens from Africa during the Neolithic expansion [34]. The initial virulence of Mtb is therefore strongly adapted to the occasional availability of the population to be infected and to an evolutionary ability to survive for a long time in the host until the opportunity for transmission arises. Consequently, Mtb infections have evolved in low human population densities, exhibiting a pattern of chronic development, accompanied by decades of latency before progressing to active disease [10,11,12]. It is a well-adapted human pathogen, requiring the induction of a strong inflammation and destruction of lung tissue for transmission and evolutionary survival [9,34,35]. This feature is unusual in most pathogens, where virulence is not associated with their spread to other hosts [36].

The pathogen does not seem to require virulence factors such as the usual pili, toxins, or capsules that are essential for the invasion of epithelial barriers [9]. Nevertheless, their potential involvement should not be ignored, as a capsule and pili have recently been identified in Mtb, although their roles during infection in vivo are still unclear [37,38,39]. Mtb virulence relies on its ability to manipulate host macrophages, where it establishes intracellular niches to cross mucosal barriers and avoid pathogen destruction. First, Mtb subverts the endocytic pathway, preventing phagolysosome fusion and proteolytic digestion [40,41,42,43,44,45]. Second, it activates innate immune responses to induce its transmigration into the lung parenchyma [46,47]. There, infected macrophages attract more permissive cells, expanding intracellular niches [9,48,49,50]. Mtb induces the adaptive responses that stimulate its containment and encourage a long life inside granulomas [51,52,53,54]. Finally, the pathogen induces necrotic cell death in macrophages, granuloma destruction, and lung cavitation for transmission [47,49,55,56]. Common to all these events is the major virulence factor: the “early secreted antigenic target of 6 kDa” (ESAT-6, also called EsxA).

2. Virulence Evolution among MTBC