Edward D. Chan

Mycobacterial lung diseases are caused by Mycobacterium tuberculosis (Mtb) or non-tuberculous mycobacteria (NTM). Mtb is acquired by inhalation of droplet nuclei (<5 µm) generated from persons with...

Nontuberculous mycobacteria (NTM) are environmental microorganisms for which large, systematic studies of niche diversity are lacking. We performed a semi-longitudinal state-wide sampling campaign (2015–2019) for environmental NTM across Hawai’i. A volunteer network collected 2,334 water biofilms, soil, and dust samples from built ( n = 1,946) and natural ( n = 388) sites. Of these, 541 contained culturable NTM (23%) and per island hotspots identified. Of 74 NTM species recovered, the most prevalent rapid growing mycobacteria (RGM) were Mycobacterium porcinum , Mycobacterium chelonae, and Mycobacterium abscessus. Mycobacterium intracellulare subsp. chimaera was the most frequently isolated slow growing mycobacteria (SGM). Our longitudinal analyses indicate widespread colonization of diverse niches by species less associated with lung infections such as M. chelonae . In contrast, household water biofilms tended to be reliable niches for M. abscessus and M. avium complex species colonization across the 5-year study. Analysis of 590 deidentified lung samples from Hawai’i and other Pacific Islands revealed M. chimaera as the most frequently isolated species (40%, 238/590). Phylogenetic analysis of environmental and lung M. abscessus suggests most cluster within dominant circulating clone 1 (DCC1). Contrastingly , most Hawai’i and other Pacific Island M. chimaera were distinct from previously studied European isolates, leading to the identification of two novel clusters of phylogenetically related strains we have termed Pacific Island Circulating Cluster 1 and 2 (PCC1, PCC2). PCC1 consists exclusively of Hawai’i/Pacific Island isolates, while PCC2 was enriched by lung samples and was mostly collected from Hawai’i/Pacific Islands. These data reveal the genetic diversity, ecological niches, and potential reservoirs of NTM in varied Hawai’i ecosystems. IMPORTANCE: Nearly one in four environmental Hawai’i samples tested positive for any NTM species, with hotspots often overlapping population centers. Recovery of any NTM species occurred just as often from natural settings as homes and public buildings, highlighting exposures as a normal part of life. Soil was the most common reservoir for NTM colonization, but we distinguish NTM species pertinent to lung disease that were far more likely to be found in water biofilms, such as showerheads and kitchen sinks. No single species dominated the environment; yet, the type of NTM found in water systems closely mirrored those recovered from patients’ lung samples. Genetic analyses revealed that Hawai’i harbors distinct, locally circulating strains, including lineages not linked to known hospital outbreaks. Together, these findings improve our understanding of where precarious exposures can occur and inform public health strategies to reduce exposures by highlighting niches that are common hotspots for NTM colonization.

RATIONALE: The host-protective role of alpha-1-antitrypsin (AAT) against mycobacteria may be partly attributed to its binding to the cytoplasmic glucocorticoid receptor (GR) that results in gene regulation in macrophages that favors killing of ingested mycobacteria. The AAT–GR complex was found to be significantly responsible for limiting Mycobacterium avium complex ( MAC ) burden in macrophages; this host-protective function of AAT–GR is due, in part, to induction of COLONY STIMULATING FACTOR-2 ( CSF-2 ) gene which encodes for granulocyte-monocyte colony stimulating factor (GM-CSF). METHODS: To better understand the role of AAT–GR binding during mycobacterial infection, we performed bulk RNA sequencing (RNA-seq) on four different groups of cells: (i) control THP-1 cells (THP-1 ^control ); (ii) THP-1 ^control cells infected with Mycobacterium intracellulare ( MAC ); (iii) THP-1 ^control cells incubated with MAC + AAT; and (iv) THP-1 cells knocked down for GR (THP-1 ^GR-KD ) incubated with MAC + AAT. RESULTS: Our analyses revealed that MAC infection significantly upregulated 1,977 genes and significantly downregulated 2,303 genes in THP-1 ^control cells. Additionally, AAT significantly upregulated 1,200 genes and downregulated 890 genes in MAC -infected THP-1 ^control cells. Furthermore, the regulation of 1,624 genes that are regulated by AAT+GR in THP-1 ^control cells was augmented in THP-1 ^GR-KD cells, indicating that the regulation of these genes by AAT+ MAC is inhibited by GR. Conversely, the regulation of 1,683 genes by AAT+ MAC in THP-1 ^control cells was attenuated in THP-1 ^GR-KD cells, indicating that the regulation of these genes by AAT+ MAC is enhanced by GR. MAC also induced both CSF2 (GM-CSF) and CSF1 (encodes for monocyte colony stimulating factor, M-CSF) expression. Whereas AAT inhibited MAC -induced M-CSF mRNA was dependent on GR, this inhibition of M-CSF protein was not dependent on GR. In contrast, AAT did not inhibit MAC -induced CSF2 (GM-CSF) expression. Since either MAC or AAT induced GM-CSF expression in macrophages, further investigation revealed that AAT-inhibition of cell-associated MAC burden was abrogated upon neutralization of endogenous GM-CSF. CONCLUSIONS: The ability of AAT to induce GM-CSF and to inhibit MAC -induced M-CSF may skew macrophages to a phenotype that is better endowed to control mycobacterial infection.

Mycobacteria are comprised of Mycobacterium tuberculosis complex, Mycobacterium leprae, and non-tuberculous mycobacteria (NTM). While NTM contain approximately 200 distinct species and subspecies, diseases in humans are predominantly caused by three main groups: (i) Mycobacterium avium complex (which is comprised of several members, with the most clinically relevant being M. avium subsp hominissuis, M. intracellulare subsp intracellulare, and M. intracellulare subsp chimaera);(ii) Mycobacterium abscessus group (which consists of three subspecies: abscessus, massiliense, and bolletii); and (iii) Mycobacterium kansasii complex. NTM are ubiquitous in natural and built environs, especially in freshwater, soil, and biofilms, making NTM infections environmentally-acquired.Upon encountering a patient with an NTM infection, four elements should be determined: the specific NTM responsible, how the infection was contracted, the location and extent of the infection, and the presence of any host risk factors. The last two elements are interrelated because the degree of underlying host risk factors often dictates the scope of the infection. Thus, it is useful to categorize the NTM infection into one of four main types: (i) skin, soft tissue, and osteoarticular infections, which are typically due to accidental trauma or medical procedures; (ii) isolated infection of the head and neck lymph nodes (also known as cervical lymphadenitis or "scrofula"); (iii) isolated NTM lung disease; (iv) Hicks and colleagues (https://doi.org/10.3389/fimmu.2023.1078976) reported three infants with complete DiGeorge anomaly (cDGA) -a genetic condition resulting in the complete absence of the thymus resulting in severe T cell deficiency -all with disseminated NTM infections. These cases are rare, likely due to the highly infrequent intersection of two uncommon conditions (DGA and NTM infection). The authors recommend that DGA should be considered in the differential diagnoses of children with disseminated NTM infections. Currently, mutations of genes that encode for elements of the interferon-gamma (IFNg)-interleukin-12 (IL-12) axis should be suspected in very young children with disseminated NTM disease. In adults with disseminated NTM infections, acquired immune disorders like AIDS, anti-IFNg antibody syndrome, or use of immunosuppressive drugs are more likely risk factors.disseminatedVarious mouse strains have been used to explore host susceptibility to NTM infections. However, most murine infection models result in systemic NTM infection rather than exhibit isolated NTM lung disease, the latter characterized by bronchiolitis and bronchiectasis in humans. Yang and colleagues (https://doi.org/10.3389/fimmu.2023.1277745) developed a mouse model using intratracheal infection with M. abscessus embedded in agar beads, which better simulates airway infection. The infected mice developed granulomas, a common feature M. abscessus infection in humans. Others have also employed a fibrin plug model where M. abscessus is suspended in thrombin and fibrinogen, trapping the bacteria in the airways as the plugs coagulate (1). Models such as these intuitively more closely replicate isolated NTM lung disease. Future studies could investigate whether such inoculation methods also enhance NTM biofilm formation in the airways, a barrier known to resist penetration by immune cells and antibiotics.The co-evolution of humans and mycobacteria has enabled reciprocal adaptation in a mutual struggle for dominance. A recurring example is that the same host factors or mycobacterial components can be used by either side to gain an advantage, analogous to a "tug o' war" event. Wang and colleagues (https://doi.org/10.3389/fimmu.2024.1413947) reviewed the role of CD36, a cell surface scavenger receptor which mycobacteria have hijacked for their own advantage. Although their focus was on M. tuberculosis, similar mechanisms may also be involved in the pathogenesis of NTM disease. They discussed that M. tuberculosis engages CD36 leading to: (i) increased fatty acid uptake by host cells, aiding bacillary persistence; (ii) monocyte differentiation into macrophages with an immunosuppressive phenotype; and (iii) enhanced uptake of surfactant lipids by macrophages, supporting mycobacterial growth. Additionally, they highlighted studies showing CD36 expression in various cell types that are involved in granuloma formation, including macrophages, granulocytes, lymphocytes, and fibroblasts. It was previously demonstrated that the M. avium complex manipulates macrophage lipid metabolism, leading to the formation of lipid-laden foamy macrophages that create a favorable environment for mycobacterial growth (2). Recent studies using a mouse model demonstrated that CD36 expression is elevated on macrophages during M. avium infection (3). In line with this, inhibiting CD36 has been shown to reduce the growth of M. avium. Choi et al (4) confirmed that blocking CD36 leads to decreased growth of M. avium, which aligns with earlier research. These findings highlight the crucial role of CD36 in the pathology of mycobacterial diseases as well as a potential therapeutic target.Bronchiectasis, a common feature of chronic NTM lung disease, is characterized by repeated cycles of infection and inflammation. Airway injury may be exacerbated by NTM-generated biofilms, a complex matrix of extracellular polymeric substances -comprised of DNA, proteins, polysaccharides, and lipids -released from live and dead microbes (5)(6)(7)(8)(9). NTM lung cavity walls have also been demonstrated to contain biofilms (10). This biofilm matrix provides protection and nutrients for the microbes (11). Keefe and colleagues (https://doi.org/10.3389/fimmu.2025.1508584) identified gene products from M. abscessus in biofilms using a transposon library of M. abscessus along with surface proteomic analysis. They uncovered specific proteins related to the mycobacteria's ability to detach and establish new...

Granulomas, organized aggregates of immune cells which form in response to Mycobacterium tuberculosis (Mtb), are characteristic but not exclusive of tuberculosis (TB). Despite existing investigations on TB granulomas, the determinants that differentiate host-protective granulomas from granulomas that contribute to TB pathogenesis are often disputed. Thus, the goal of this narrative review is to help clarify the existing literature on such determinants. We adopt the a priori view that TB granulomas are host-protective organelles and discuss the molecular and cellular determinants that induce protective granulomas and those that promote their failure. While reports about protective TB granulomas and their failure may initially seem contradictory, it is increasingly recognized that either deficiencies or excesses of the molecular and cellular components in TB granuloma formation may be detrimental to the host. More specifically, insufficient or excessive expression/representation of the following components have been reported to skew granulomas toward the less protective phenotype: (i) epithelioid macrophages; (ii) type 1 adaptive immune response; (iii) type 2 adaptive immune response; (iv) tumor necrosis factor; (v) interleukin-12; (vi) interleukin-17; (vii) matrix metalloproteinases; (viii) hypoxia in the TB granulomas; (ix) hypoxia inducible factor-1 alpha; (x) aerobic glycolysis; (xi) indoleamine 2,3-dioxygenase activity; (xii) heme oxygenase-1 activity; (xiii) immune checkpoint; (xiv) leukotriene A4 hydrolase activity; (xv) nuclear-factor-kappa B; and (xvi) transforming growth factor-beta. Rather, more precise and timely coordinated immune responses appear essential for eradication or containment of Mtb infection. Since there are several animal models of infection with Mtb, other species within the Mtb complex, and the surrogate Mycobacterium marinum – whether natural (cattle, elephants) or experimental (zebrafish, mouse, guinea pig, rabbit, mini pig, goat, non-human primate) infections – we also compared the TB granulomatous response and other pathologic lung lesions in various animals infected with one of these mycobacteria with that of human pulmonary TB. Identifying components that dictate the formation of host-protective granulomas and the circumstances that result in their failure can enhance our understanding of the macrocosm of human TB and facilitate the development of novel remedies – whether they be direct therapeutics or indirect interventions – to efficiently eliminate Mtb infection and prevent its pathologic sequelae.

The controversy of immune checkpoint inhibition and risk of mycobacterial infections: the need for further studies and continued discussions on this puzzling issue

Alpha‐1‐antitrypsin (AAT) plays a homeostatic role in attenuating excessive inflammation and augmenting host defense against microbes. We demonstrated previously that AAT binds to the glucocorticoid receptor (GR) resulting in significant anti‐inflammatory and antimycobacterial consequences in macrophages. Our current investigation aims to uncover AAT‐regulated genes that rely on GR in macrophages. We incubated control THP‐1 cells (THP‐1^control) and THP‐1 cells knocked down for GR (THP‐1^GR‐KD) with AAT, performed bulk RNA sequencing, and analyzed the findings. In THP‐1^control cells, AAT significantly upregulated 408 genes and downregulated 376 genes. Comparing THP‐1^control and THP‐1^GR‐KD cells, 125 (30.6%) of the AAT‐upregulated genes and 154 (41.0%) of the AAT‐downregulated genes were significantly dependent on GR. Among the AAT‐upregulated, GR‐dependent genes, CSF‐2 that encodes for granulocyte‐monocyte colony‐stimulating factor (GM‐CSF), known to be host‐protective against nontuberculous mycobacteria, was strongly upregulated by AAT and dependent on GR. We further quantified the mRNA and protein of several AAT‐upregulated, GR‐dependent genes in macrophages and the mRNA of several AAT‐downregulated, GR‐dependent genes. We also discussed the function(s) of selected AAT‐regulated, GR‐dependent gene products largely in the context of mycobacterial infections. In conclusion, AAT regulated several genes that are dependent on GR and play roles in host immunity against mycobacteria.