Nontuberculous Mycobacteria Persistence in a Cell Model Mimicking Alveolar Macrophages

Abstract

Nontuberculous Mycobacteria (NTM) respiratory infections have been gradually increasing. Here, THP-1 cells were used as a model to evaluate intracellular persistence of three NTM species (reference and clinical strains) in human alveolar macrophages. The contribution of phagosome acidification, nitric oxide (NO) production and cell dead on NTM intracellular fate was assessed. In addition, strains were characterized regarding their repertoire of virulence factors by whole-genome sequencing. NTM experienced different intracellular fates: M. smegmatis and M. fortuitum ATCC 6841 were cleared within 24h. In contrast, M. avium strains (reference/clinical) and M. fortuitum clinical strain were able to replicate. Despite this fact, unexpectedly high percentages of acidified phagosomes were found harbouring rab7, but not CD63. All NTM were able to survive in vitro at acidic pHs, with the exception of M. smegmatis. Our data further suggested a minor role for NO in intracellular persistence and that apoptosis mediated by caspase 8 and 3/7, but not necrosis, is triggered during NTM infection. Insights regarding the bacteria genomic backbone corroborated the virulence potential of M. avium and M. fortuitum. In conclusion, the phenotypic traits detected contrast with those described for M. tuberculosis, pointing out that NTM adopt distinct strategies to manipulate the host immune defense and persist intracellularly.

Differential Responses by Human Macrophages to Infection With Mycobacterium tuberculosis and Non-tuberculous Mycobacteria

“…They also found that, overall, M. smegmatis and M. fortuitum were better biofilm formers than M. chelonae (Sousa et al, 2015). In a subsequent study from the same group, infection with M. smegmatis, M. fortuitum (clinical and reference strains), and M. avium (clinical and reference strains) were compared in THP-1 cells (Sousa et al, 2019). They found that M. smegmatis and reference strain of M. fortuitum were cleared relatively quickly in the macrophages whereas both strains of M. avium and the clinical strain of M. fortuitum were able to replicate.…”

“…Although phagosome and lysosome markers were quantified in the cells, co-localization of mycobacteria with lysosomes was not. Caspase 8 and caspase 3/7 positive cells were quantified, showing that these caspases were induced more by M. fortuitum and M. avium up to 6 h; however, after 1 day, M. smegmatis and M. fortuitum induced greater caspase 3/7 positive cells (Sousa et al, 2019). But quantitation of actual apoptotic cells (e.g., TUNEL staining) was not performed.…”

“…Others have also compared various species of NTM with or without comparison to MTB in macrophage models of infection (Helguera-Repetto et al, 2014;Sousa et al, 2015Sousa et al, , 2019. While mycobacterial burden was consistently quantified in these studies, significant differences in the times of infection and in the assays analyzed make these studies and ours complementary.…”

“…But characterization of MTB with various species of NTM in the context of macrophage infection and the analysis of their effector functions are incomplete. Previous studies have investigated bacterial burden, biofilm formation by RGM (but not M. abscessus), apoptotic and phagosome-lysosome markers among NTM, macrophage monolayer disruption, and production of reactive oxygen intermediates (Helguera-Repetto et al, 2014;Sousa et al, 2015Sousa et al, , 2019. In this study, we compared bacterial burden in macrophages infected with a laboratory strain of MTB with various SGM and RGM species and correlated this marker with some of the known effector mechanisms of killing by macrophages.…”

Non-Tuberculous Mycobacteria: Molecular and Physiological Bases of Virulence and Adaptation to Ecological Niches

“…Inside the host, mycobacteria are found within different cell types, but tropism to macrophages where mycobacteria are internalized into phagosomes and replicate has been highlighted by the vast majority of studies [ 10 ]. The arrest of the maturation of early mycobacterium-containing phagosomes and resistance to acidic and oxidative damage are well-known protective mechanisms of these pathogens [ 10 ].…”