Common Features of Environmental Mycobacterium chelonae from Colorado Using Partial and Whole Genomic Sequence Analyses.

Nontuberculous mycobacteria (NTM) are environmentally acquired opportunistic pathogens that cause chronic lung disease in susceptible individuals. While presumed to be ubiquitous in built and natural environments, NTM environmental studies are limited. While environmental sampling campaigns have been performed in geographic areas of high NTM disease burden, NTM species diversity is less defined among areas of lower disease burden like Colorado. In Colorado, metals such as molybdenum have been correlated with increased risk for NTM infection, yet environmental NTM species diversity has not yet been widely studied. Based on prior regression modeling, three areas of predicted high, moderate, and low NTM risk were identified for environmental sampling in Colorado. Ice, plumbing biofilms, and sink tap water samples were collected from publicly accessible freshwater sources. All samples were microbiologically cultured and NTM were identified using partial rpoB gene sequencing. From these samples, areas of moderate risk were more likely to be NTM positive. NTM recovery from ice was more common than recovery from plumbing biofilms or tap water. Overall, nine different NTM species were identified, including clinically important Mycobacterium chelonae. MinION technology was used to whole genome sequence and compare mutational differences between six M. chelonae genomes, representing three environmental isolates from this study and three other M. chelonae isolates from other sources. Drug resistance genes and prophages were common findings among environmentally derived M. chelonae, promoting the need for expanded environmental sampling campaigns to improve our current understanding of NTM species abundance while opening new avenues for improved targeted drug therapies.

Investigating Nontuberculous Mycobacteria Transmission at the Colorado Adult Cystic Fibrosis Program.

Rationale: Healthcare-associated transmission of nontuberculous mycobacteria (NTM) among people with cystic fibrosis (pwCF) has been investigated at CF centers worldwide, with conflicting conclusions. We investigated transmission at the Colorado Adult CF Program. Objectives: To systematically investigate healthcare-associated transmission and/or acquisition of NTM to determine similarity among respiratory and environmental isolates, and to compare home residence watershed mapping among pwCF having genetically similar NTM isolates. Methods: Whole-genome sequencing of NTM isolates from 80 pwCF was conducted to identify genetically similar isolate clusters (⩽30 SNP differences). Epidemiology, comparison of respiratory and environmental isolates, and home residence watershed mapping were analyzed. Measurements and Main Results: Whole-genome sequencing analysis revealed 11 clusters of NTM [6 Mycobacterium abscessus subspecies (ssp.) abscessus, 1 M. abscessus ssp. massiliense, 2 Mycobacterium avium, and 2 Mycobacterium intracellulare] among pwCF. Epidemiologic investigation demonstrated opportunities for healthcare-associated transmission in two M. abscessus and two M. avium clusters. Respiratory and healthcare environmental isolate comparisons revealed no genetic similarity. Individuals comprising one M. abscessus cluster, with no plausible healthcare-associated transmission, resided in the same watershed. Conclusions: This study suggests healthcare-associated transmission of M. abscessus is rare and includes a report of potential healthcare-associated transmission of M. avium among pwCF. One M. abscessus cluster possibly had common acquisition arising from residing in the same watershed. The presence of genetically similar isolates is insufficient to demonstrate healthcare-associated NTM transmission. Standardizing epidemiologic investigation, combined with environmental sampling and watershed analysis, will improve understanding of the frequency and nature of healthcare-associated NTM transmission among pwCF.

Soil Properties and Moisture Synergistically Influence Nontuberculous Mycobacterial Prevalence in Natural Environments of Hawai'i.

Nontuberculous mycobacteria (NTM) are opportunistic pathogens that cause chronic pulmonary disease (PD). NTM infections are thought to be acquired from the environment; however, the basal environmental factors that drive and sustain NTM prevalence are not well understood. The highest prevalence of NTM PD cases in the United States is reported from Hawai'i, which is unique in its climate and soil composition, providing an opportunity to investigate the environmental drivers of NTM prevalence. We used microbiological sampling and spatial logistic regression complemented with fine-scale soil mineralogy to model the probability of NTM presence across the natural landscape of Hawai'i. Over 7 years, we collected and microbiologically cultured 771 samples from 422 geographic sites in natural areas across the Hawaiian Islands for the presence of NTM. NTM were detected in 210 of these samples (27%), with Mycobacterium abscessus being the most frequently isolated species. The probability of NTM presence was highest in expansive soils (those that swell with water) with a high water balance (>1-m difference between rainfall and evapotranspiration) and rich in Fe-oxides/hydroxides. We observed a positive association between NTM presence and iron in wet soils, supporting past studies, but no such association in dry soils. High soil-water balance may facilitate underground movement of NTM into the aquifer system, potentially compounded by expansive capabilities allowing crack formation under drought conditions, representing further possible avenues for aquifer infiltration. These results suggest both precipitation and soil properties are mechanisms by which surface NTM may reach the human water supply. IMPORTANCE Nontuberculous mycobacteria (NTM) are ubiquitous in the environment, being found commonly in soils and natural bodies of freshwater. However, little is known about the environmental niches of NTM and how they relate to NTM prevalence in homes and other human-dominated areas. To characterize NTM environmental associations, we collected and cultured 771 samples from 422 geographic sites in natural areas across Hawai'i, the U.S. state with the highest prevalence of NTM pulmonary disease. We show that the environmental niches of NTM are most associated with highly expansive, moist soils containing high levels of iron oxides/hydroxides. Understanding the factors associated with NTM presence in the natural environment will be crucial for identifying potential mechanisms and risk factors associated with NTM infiltration into water supplies, which are ultimately piped into homes where most exposure risk is thought to occur.

Comparative survival of environmental and clinical Mycobacterium abscessus isolates in a variety of diverse host cells.

AIMS: Mycobacterium abscessus subsp. abscessus (MABS) is an emerging, opportunistic pathogen found globally in freshwater biofilms and soil. Typically, isolates are treated as a uniform group of organisms and very little is known about their comparative survival in healthy host cells. We posit that environmentally- and clinically derived isolates, show differential infectivity in immune cells and resistance to innate defenses. METHODS AND RESULTS: Six MABS isolates were tested including three water biofilm/soil and three sputum-derived isolates. A clinical MABS type strain and an environmental isolate of Arthrobacter were also included. MABS counts were significantly higher compared to Arthrobacter after co-culture with Acanthamoeba lenticulata, BEAS-2B epithelial cells, alveolar macrophages and the THP-1 macrophage cell line. A rough sputum-derived MABS isolate emerged as an isolate with higher virulence compared to others tested, as both a pellicle and cord former, survivor in the human cell models tested, inducer of high and prolonged production of pro-inflammatory cytokines, and the capacity to evade LL-37. CONCLUSIONS: Findings support intraspecies variation between MABS isolates. SIGNIFICANCE AND IMPACT OF THE STUDY: These data indicate subversion of host immune defenses by environmental and clinical MABS isolates is nuanced and maybe isolate dependent, providing new information regarding the pathogenesis of NTM infections.

Measurable genomic changes in Mycobacterium avium subsp. hominissuis after long-term adaptation in Acanthamoeba lenticulata and reduced persistence in macrophages.

Free-living amoebae are ubiquitous in aquatic environments and act as environmental reservoirs for nontuberculous mycobacteria. Mycobacterium avium subsp. hominissuis recovered from Acanthamoeba has been demonstrated to be more virulent in both human and murine models. Here, we investigate the persistence of M. avium subsp. hominissuis after short-term (2 weeks) and long-term (42 weeks) co-culture in Acanthamoeba lenticulata We hypothesize that A. lenticulata-adapted M. avium subsp. hominissuis demonstrate phenotypic and genomic changes facilitating intracellular persistence in naïve Acanthamoeba and human macrophages. M. avium subsp. hominissuis CFU in co-culture with A. lenticulata were recorded every 2 weeks up to 60 weeks. While A. lenticulata-associated M. avium subsp. hominissuis CFU did not significantly change across 60 weeks of co-culture, longer adaptation time in amoebae reduced colony size. Isolates recovered after 2 or 42 weeks of amoebae co-culture were referred as "early-adapted" and "late-adapted" M. avium subsp. hominissuis, respectively. Whole genome sequencing was performed on amoebae-adapted isolates with pan-genome comparisons to the original M. avium subsp. hominissuis isolate. Next, amoebae-adapted isolates were assessed for their persistence in A. lenticulata, A. castellanii, and human THP-1 macrophages. Multiplex cytokine/chemokine analyses were conducted on THP-1 culture supernatants.  Compared to the original isolate, counts of late-adapted M. avium subsp. hominissuis were reduced in Acanthamoeba and contrary to expectations, lower counts were also observed in THP-1 macrophages with concomitant decrease in TNFa, IL-6, and MIP-1b suggesting that host adaptation may influence the inflammatory properties of M. avium IMPORTANCE Short-term interaction between Acanthamoeba and M. avium has been demonstrated to increase infectivity in human and murine models of infection, establishing the paradigm that amoebae "train" M. avium in the environment by selecting for phenotypes capable of enduring in human cells. We investigate this phenomenon further by determining the consequence of long-term amoebae adaptation on M. avium subsp. hominissuis persistence in host cells. We monitored genomic changes across long-term Acanthamoeba co-culture and report significant changes to the M. avium subsp. hominissuis genome in response to amoebae-adaptation and reduced colony size. Furthermore, we examined isolates co-cultured with A. lenticulata for 2 or 42 weeks and provide biological evidence that long-term co-culture in amoebae reduces M. avium persistence in human macrophages.

Nontuberculous Mycobacteria Show Differential Infectivity and Use Phospholipids to Antagonize LL-37.

Comparisons of infectivity among the clinically important nontuberculous mycobacteria (NTM) species have not been explored in great depth. Rapid-growing mycobacteria, including Mycobacterium abscessus and M. porcinum, can cause indolent but progressive lung disease. Slow-growing members of the M. avium complex are the most common group of NTM to cause lung disease, and molecular approaches can now distinguish between several distinct species of M. avium complex including M. intracellulare, M. avium, M. marseillense, and M. chimaera. Differential infectivity among these NTM species may, in part, account for differences in clinical outcomes and response to treatment; thus, knowing the relative infectivity of particular isolates could increase prognostication accuracy and enhance personalized treatment. Using human macrophages, we investigated the infectivity and virulence of nine NTM species, as well as multiple isolates of the same species. We also assessed their capacity to evade killing by the antibacterial peptide cathelicidin (LL-37). We discovered that the ability of different NTM species to infect macrophages varied among the species and among isolates of the same species. Our biochemical assays implicate modified phospholipids, which may include a phosphatidylinositol or cardiolipin backbone, as candidate antagonists of LL-37 antibacterial activity. The high variation in infectivity and virulence of NTM strains suggests that more detailed microbiological and biochemical characterizations are necessary to increase our knowledge of NTM pathogenesis.

Assessment of Soil Features on the Growth of Environmental Nontuberculous Mycobacterial Isolates from Hawai'i.

Environmental nontuberculous mycobacteria (NTM), with the potential to cause opportunistic lung infections, can reside in soil. This might be particularly relevant in Hawai'i, a geographic hot spot for NTM infections and whose soil composition differs from many other areas of the world. Soil components are likely to contribute to NTM prevalence in certain niches as food sources or attachment scaffolds, but the particular types of soils, clays, and minerals that impact NTM growth are not well-defined. Hawai'i soil and chemically weathered rock (saprolite) samples were examined to characterize the microbiome and quantify 11 mineralogical features as well as soil pH. Machine learning methods were applied to identify important soil features influencing the presence of NTM. Next, these features were directly tested in vitro by incubating synthetic clays and minerals in the presence of Mycobacteroides abscessus and Mycobacterium chimaera isolates recovered from the Hawai'i environment, and changes in bacterial growth were determined. Of the components examined, synthetic gibbsite, a mineral form of aluminum hydroxide, inhibited the growth of both M. abscessus and M. chimaera, while other minerals tested showed differential effects on each species. For example, M. abscessus (but not M. chimaera) growth was significantly higher in the presence of hematite, an iron oxide mineral. In contrast, M. chimaera (but not M. abscessus) counts were significantly reduced in the presence of birnessite, a manganese-containing mineral. These studies shed new light on the mineralogic features that promote or inhibit the presence of Hawai'i NTM in Hawai'i soil.IMPORTANCE Globally and in the United States, the prevalence of NTM pulmonary disease-a potentially life-threatening but underdiagnosed chronic illness-is prominently rising. While NTM are ubiquitous in the environment, including in soil, the specific soil components that promote or inhibit NTM growth have not been elucidated. We hypothesized that NTM culture-positive soil contains minerals that promote NTM growth in vitro Because Hawai'i is a hot spot for NTM and a unique geographic archipelago, we examined the composition of Hawai'i soil and identified individual clay, iron, and manganese minerals associated with NTM. Next, individual components were evaluated for their ability to directly modulate NTM growth in culture. In general, gibbsite and some manganese oxides were shown to decrease NTM, whereas iron-containing minerals were associated with higher NTM counts. These data provide new information to guide future analyses of soil-associated factors impacting persistence of these soil bacteria.

Human IL-32 expression protects mice against a hypervirulent strain of Mycobacterium tuberculosis.

Silencing of interleukin-32 (IL-32) in a differentiated human promonocytic cell line impairs killing of Mycobacterium tuberculosis (MTB) but the role of IL-32 in vivo against MTB remains unknown. To study the effects of IL-32 in vivo, a transgenic mouse was generated in which the human IL-32γ gene is expressed using the surfactant protein C promoter (SPC-IL-32γTg). Wild-type and SPC-IL-32γTg mice were infected with a low-dose aerosol of a hypervirulent strain of MTB (W-Beijing HN878). At 30 and 60 d after infection, the transgenic mice had 66% and 85% fewer MTB in the lungs and 49% and 68% fewer MTB in the spleens, respectively; the transgenic mice also exhibited greater survival. Increased numbers of host-protective innate and adaptive immune cells were present in SPC-IL-32γTg mice, including tumor necrosis factor-alpha (TNFα) positive lung macrophages and dendritic cells, and IFN-gamma (IFNγ) and TNFα positive CD4(+) and CD8(+) T cells in the lungs and mediastinal lymph nodes. Alveolar macrophages from transgenic mice infected with MTB ex vivo had reduced bacterial burden and increased colocalization of green fluorescent protein-labeled MTB with lysosomes. Furthermore, mouse macrophages made to express IL-32γ but not the splice variant IL-32ß were better able to limit MTB growth than macrophages capable of producing both. The lungs of patients with tuberculosis showed increased IL-32 expression, particularly in macrophages of granulomas and airway epithelial cells but also B cells and T cells. We conclude that IL-32γ enhances host immunity to MTB.

Epidemiology of Nontuberculous Mycobacterial Lung Disease and Tuberculosis, Hawaii, USA.

Previous studies found Hawaiians and Asian-Americans/Pacific Islanders to be independently at increased risk for nontuberculous mycobacterial pulmonary disease (NTMPD) and tuberculosis (TB). To better understand NTM infection and TB risk patterns in Hawaii, USA, we evaluated data on a cohort of patients in Hawaii for 2005-2013. Period prevalence of NTMPD was highest among Japanese, Chinese, and Vietnamese patients (>300/100,000 persons) and lowest among Native Hawaiians and Other Pacific Islanders (50/100,000). Japanese patients were twice as likely as all other racial/ethnic groups to have Mycobacterium abscessus isolated (adjusted odds ratio 2.0, 95% CI 1.2-3.2) but were not at increased risk for infection with other mycobacteria species. In contrast, incidence of TB was stable and was lowest among Japanese patients (no cases) and highest among Filipino, Korean, and Vietnamese patients (>50/100,000). Substantial differences exist in the epidemiology of NTMPD by race/ethnicity, suggesting behavioral and biologic factors that affect disease susceptibility.

Mycobacterium abscessus Displays Fitness for Fomite Transmission.

Mycobacterium abscessus is a rapidly growing nontuberculous mycobacterium (NTM) increasingly reported in soft tissue infections and chronic lung diseases, including cystic fibrosis. The environmental source of M. abscessus has not been definitively identified, but NTM have been detected in soil and water. To determine the potential of soil-derived M. abscessus as an infectious source, we explored the association, growth, and survival of M. abscessus with defined mineral particulates, including kaolin, halloysite, and silicone dioxide, and house dust as possible M. abscessus fomites. M. abscessus physically associated with particulates, and the growth of M. abscessus was enhanced in the presence of both kaolin and house dust. M. abscessus survived desiccation for 2 weeks but was not viable after 3 weeks. The rate of decline of M. abscessus viability during desiccation was reduced in the presence of house dust. The evidence for enhanced growth and survival of M. abscessus during alternating growth and drying periods suggests that dissemination could occur when in wet or dry environments. These studies are important to understand environmental survival and acquisition of NTM.IMPORTANCE The environmental source of pulmonary Mycobacterium abscessus infections is not known. Fomites are nonliving carriers of infectious agents and may contribute to acquisition of M. abscessus This study provides evidence that M. abscessus growth is enhanced in the presence of particulates, using kaolin, an abundant natural clay mineral, and house dust as experimental fomites. Moreover, M. abscessus survived desiccation for up to 2 weeks in the presence of house dust, kaolin, and several chemically defined mineral particulates; mycobacterial viability during extended periods of dessication was enhanced by the presence of house dust. The growth characteristics of M. abscessus with particulates suggest that a fomite mechanism of transmission may contribute to M. abscessus acquisition, which may lead to strategies to better control infections by M. abscessus and related organisms.

Curcumin enhances human macrophage control of Mycobacterium tuberculosis infection.

BACKGROUND AND OBJECTIVE: With the worldwide emergence of highly drug-resistant tuberculosis (TB), novel agents that have direct antimycobacterial effects or that enhance host immunity are urgently needed. Curcumin is a polyphenol responsible for the bright yellow-orange colour of turmeric, a spice derived from the root of the perennial herb Curcuma longa. Curcumin is a potent inducer of apoptosis-an effector mechanism used by macrophages to kill intracellular Mycobacterium tuberculosis (MTB). METHODS: An in vitro human macrophage infection model was used to determine the effects of curcumin on MTB survival. RESULTS: We found that curcumin enhanced the clearance of MTB in differentiated THP-1 human monocytes and in primary human alveolar macrophages. We also found that curcumin was an inducer of caspase-3-dependent apoptosis and autophagy. Curcumin mediated these anti-MTB cellular functions, in part, via inhibition of nuclear factor-kappa B (NFκB) activation. CONCLUSION: Curcumin protects against MTB infection in human macrophages. The host-protective role of curcumin against MTB in macrophages needs confirmation in an animal model; if validated, the immunomodulatory anti-TB effects of curcumin would be less prone to drug resistance development.

Caspase-3-independent apoptotic pathways contribute to interleukin-32γ-mediated control of Mycobacterium tuberculosis infection in THP-1 cells.

BACKGROUND: Macrophages are the primary effector cells responsible for killing Mycobacterium tuberculosis (MTB) through various mechanisms, including apoptosis. However, MTB can evade host immunity to create a favorable environment for intracellular replication. MTB-infected human macrophages produce interleukin-32 (IL-32). IL-32 is a pro-inflammatory cytokine and has several isoforms. We previously found that IL-32γ reduced the burden of MTB in human macrophages, in part, through the induction of caspase-3-dependent apoptosis. However, based on our previous studies, we hypothesized that caspase-3-independent death pathways may also mediate IL-32 control of MTB infection. Herein, we assessed the potential roles of cathepsin-mediated apoptosis, caspase-1-mediated pyroptosis, and apoptosis-inducing factor (AIF) in mediating IL-32γ control of MTB infection in THP-1 cells. RESULTS: Differentiated human THP-1 macrophages were infected with MTB H37Rv alone or in the presence of specific inhibitors to caspase-1, cathepsin B/D, or cathepsin L for up to four days, after which TUNEL-positive cells were quantified; in addition, MTB was quantified by culture as well as by the percentage of THP-1 cells that were infected with green fluorescent protein (GFP)-labeled MTB as determined by microscopy. AIF expression was inhibited using siRNA technology. Inhibition of cathepsin B/D, cathepsin L, or caspase-1 activity significantly abrogated the IL-32γ-mediated reduction in the number of intracellular MTB and of the percentage of GFP-MTB-infected macrophages. Furthermore, inhibition of caspase-1, cathepsin B/D, or cathepsin L in the absence of exogenous IL-32γ resulted in a trend toward an increased proportion of MTB-infected THP-1 cells. Inhibition of AIF activity in the absence of exogenous IL-32γ also increased intracellular burden of MTB. However, since IL-32γ did not induce AIF and because the relative increases in MTB with inhibition of AIF were similar in the presence or absence of IL-32γ, our results indicate that AIF does not mediate the host-protective effect of IL-32γ against MTB. CONCLUSIONS: The anti-MTB effects of IL-32γ are mediated through classical caspase-3-dependent apoptosis as well as caspase-3-independent apoptosis.

Epidemiologic link between tuberculosis and cigarette/biomass smoke exposure: Limitations despite the vast literature.

The geographic overlap between the prevalence of cigarette smoke (CS) exposure and tuberculosis (TB) in the world is striking. In recent years, relatively large number of studies has linked cigarette or biomass fuel smoke exposure and various aspects of TB. Our goals are to summarize the significance of the known published studies, graphically represent reports that quantified the association and discuss their potential limitations. PubMed searches were performed using the key words 'tuberculosis' with 'cigarette', 'tobacco', 'smoke' or 'biomass fuel smoke.' The references of relevant articles were examined for additional pertinent papers. A large number of mostly case-control and cross-sectional studies significantly associate both direct and second-hand smoke exposure with tuberculous infection, active TB, and/or more severe and lethal TB. Fewer link biomass fuel smoke exposure and TB. While a number of studies interpreted the association with multivariate analysis, other confounders are often not accounted for in these analyses. It is also important to emphasize that these retrospective studies can only show an association and not any causal link. We further explored the possibility that even if CS exposure is a risk factor for TB, several mechanisms may be responsible. Numerous studies associate cigarette and biomass smoke exposure with TB but the mechanism(s) remains largely unknown. While the associative link of these two health maladies is well established, more definitive, mechanistic studies are needed to cement the effect of smoke exposure on TB pathogenesis and to utilize this knowledge in empowering public health policies.

Shower dehumidification to reduce nontuberculous mycobacteria aerosolization.

OBJECTIVE: Nontuberculous mycobacteria (NTM) are environmentally acquired opportunistic pathogens that can cause recalcitrant lung disease. Prior reports have demonstrated links between shower use and infections, yet the aerosolization of NTM from showerheads, as well as the humidity levels that may modulate NTM aerosolization from showerheads is less studied. The objective of the current study was to investigate the role of humidity in NTM aerosolization among showers in homes located in a geographic area with high lung disease incidence, Hawai'i, and test whether deployment of a dehumidifier in well-ventilated bathrooms reduce NTM exposure. RESULTS: Across two sampling events and five showers, existing NTM showerhead biofilms along with shower air were sampled at three points: pre-shower, post-shower, and post-dehumidification. In each of the sampling events, respiratory relevant NTM species were identified from shower biofilms, which were also detected in aerosolized shower air after showering events, but not after the shower was dehumidified and bathrooms vented. While sample size was small, these data suggest running a shower is a possible source of NTM aerosolization and using a commercial household dehumidifier in conjunction with opening bathroom doors and windows may be simple, cost-effective interventions to reduce environmental NTM exposures.

Hawaiian Volcanic Ash, an Airborne Fomite for Nontuberculous Mycobacteria.

Nontuberculous mycobacteria (NTM) are environmentally acquired opportunistic pathogens that can cause chronic lung disease. Within the U.S., Hawai'i shows the highest prevalence rates of NTM lung infections. Here, we investigated a potential role for active volcanism at the Kilauea Volcano located on Hawai'i Island in promoting NTM growth and diversity. We recovered NTM that are known to cause lung disease from plumbing biofilms and soils collected from the Kilauea environment. We also discovered viable Mycobacterium avium, Mycobacterium abscessus, and Mycobacterium intracellulare subsp. chimaera on volcanic ash collected during the 2018 Kilauea eruption. Analysis of soil samples showed that NTM prevalence is positively associated with bulk content of phosphorus, sulfur, and total organic carbon. In growth assays, we showed that phosphorus utilization is essential for proliferation of Kilauea-derived NTM, and demonstrate that NTM cultured with volcanic ash adhere to ash surfaces and remain viable. Ambient dust collected on O'ahu concurrent with the 2018 eruption contained abundant fresh volcanic glass, suggestive of inter-island ash transport. Phylogenomic analyses using whole genome sequencing revealed that Kilauea-derived NTM are genetically similar to respiratory isolates identified on other Hawaiian Islands. Consequently, we posit that volcanic eruptions could redistribute environmental microorganisms over large scales. While additional studies are needed to confirm a direct role of ash in NTM dispersal, our results suggest that volcanic particulates harbor and can redistribute NTM and should therefore be studied as a fomite for these burgeoning, environmentally acquired respiratory infections.

Environmental Sources and Transmission of Nontuberculous Mycobacteria.

The field of environmental nontuberculous mycobacteria (NTM) is benefiting from a new era of genomics that has catapulted our understanding of preferred niches, transmission, and outbreak investigations. The ability to forecast environmental features that promote or reduce environmental NTM prevalence will greatly improve with coordinated environmental sampling and by elevating the necessity for uniform disease notifications. Studies that synergize environmental biology, isolate notifications, and comparative genomics in prospective, longitudinal studies, particularly during climate changes and weather events, will be useful to solve longstanding NTM public health quandaries.

Genomic and microbiological analyses of iron acquisition pathways among respiratory and environmental nontuberculous mycobacteria from Hawai'i.

As environmental opportunistic pathogens, nontuberculous mycobacteria (NTM) can cause severe and difficult to treat pulmonary disease. In the United States, Hawai'i has the highest prevalence of infection. Rapid growing mycobacteria (RGM) such as Mycobacterium abscessus and M. porcinum and the slow growing mycobacteria (SGM) including M. intracellulare subspecies chimaera are common environmental NTM species and subspecies in Hawai'i. Although iron acquisition is an essential process of many microorganisms, iron acquisition via siderophores among the NTM is not well-characterized. In this study, we apply genomic and microbiological methodologies to better understand iron acquisition via siderophores for environmental and respiratory isolates of M. abscessus, M. porcinum, and M. intracellulare subspecies chimaera from Hawai'i. Siderophore synthesis and transport genes, including mycobactin (mbt), mmpL/S, and esx-3 were compared among 47 reference isolates, 29 respiratory isolates, and 23 environmental Hawai'i isolates. Among all reference isolates examined, respiratory isolates showed significantly more siderophore pertinent genes compared to environmental isolates. Among the Hawai'i isolates, RGM M. abscessus and M. porcinum had significantly less esx-3 and mbt genes compared to SGM M. chimaera when stratified by growth classification. However, no significant differences were observed between the species when grown on low iron culture agar or siderophore production by the chrome azurol S (CAS) assay in vitro. These results indicate the complex mechanisms involved in iron sequestration and siderophore activity among diverse NTM species.