Potential Cross-Transmission of Mycobacterium abscessus among Non-Cystic Fibrosis Patients at a Tertiary Hospital in Japan.

Mycobacterium abscessus (M. abscessus) is a highly antimicrobial-resistant pathogen that causes refractory pulmonary disease. Recently, the possibility of M. abscessus cross-transmission among cystic fibrosis (CF) patients has been reported. CF is rare in Asia, but M. abscessus pulmonary disease is common. Therefore, we investigated the possibility of M. abscessus cross-transmission in a Japanese hospital setting. Of 104 M. abscessus isolates, 25 isolates from 24 patients were classified into four clusters based on their variable number of tandem repeat profiles and were subjected to whole-genome sequencing (WGS). The epidemiological linkages among our patients were investigated by integrating the WGS data of previously reported nosocomial outbreak-related M. abscessus clinical isolates in the United Kingdom and the United States. Eight transmissible clusters (TCs) were identified. The United Kingdom and United States isolates were assigned to four clusters (TC1, TC2, TC5, and TC8) and one cluster (TC3), respectively. A total of 12 isolates from our hospital belonged to 4 clusters (TC4, TC5, TC6, and TC7). Epidemiological linkage analysis inferred direct or indirect transmission between patients in our hospital in TC4 and TC5 but not in TC6 and TC7. In TC5, the single nucleotide polymorphism distance between isolates from Japanese and United Kingdom patients was less than 21; however, there was no contact. This study revealed that genetically closely related isolates exist, even in non-CF patients. However, the transmission route remains unclear, and further research is warranted to clarify whether cross-transmission is involved. IMPORTANCE Although the possibility of Mycobacterium abscessus (M. abscessus) cross-transmission in cystic fibrosis (CF) patients has often been reported, it is not clear whether similar events have occurred in Asian non-CF patients. Whole-genome sequencing analysis of M. abscessus isolates from Fukujuji Hospital in Japan indicated that genetically closely related M. abscessus isolates exist. In addition, according to epidemiological linkage analysis, some clusters were suspected of direct or indirect transmission between patients within our hospital. However, the transmission route of M. abscessus remains unclear, because interestingly, one cluster showed a single nucleotide polymorphism distance of less than 21 from the United Kingdom isolates, but no epidemiological linkage was identified.

Genomic Analysis of a Hospital-Associated Outbreak of Mycobacterium abscessus: Implications on Transmission.

Whole-genome sequencing (WGS) has recently been used to investigate acquisition of Mycobacterium abscessus. Investigators have reached conflicting conclusions about the meaning of genetic distances for interpretation of person-to-person transmission. Existing genomic studies were limited by a lack of WGS from environmental M. abscessus isolates. In this study, we retrospectively analyzed the core and accessory genomes of 26 M. abscessus subsp. abscessus isolates collected over 7 years. Clinical isolates (n = 22) were obtained from a large hospital-associated outbreak of M. abscessus subsp. abscessus, the outbreak hospital before or after the outbreak, a neighboring hospital, and two outside laboratories. Environmental M. abscessus subsp. abscessus isolates (n = 4) were obtained from outbreak hospital water outlets. Phylogenomic analysis of study isolates revealed three clades with pairwise genetic distances ranging from 0 to 135 single-nucleotide polymorphisms (SNPs). Compared to a reference environmental outbreak isolate, all seven clinical outbreak isolates and the remaining three environmental isolates had highly similar core and accessory genomes, differing by up to 7 SNPs and a median of 1.6% accessory genes, respectively. Although genomic comparisons of 15 nonoutbreak clinical isolates revealed greater heterogeneity, five (33%) isolates had fewer than 20 SNPs compared to the reference environmental isolate, including two unrelated outside laboratory isolates with less than 4% accessory genome variation. Detailed genomic comparisons confirmed environmental acquisition of outbreak isolates of M. abscessus subsp. abscessus. SNP distances alone, however, did not clearly differentiate the mechanism of acquisition of outbreak versus nonoutbreak isolates. We conclude that successful investigation of M. abscessus subsp. abscessus clusters requires molecular and epidemiologic components, ideally complemented by environmental sampling.

Healthcare-associated links in transmission of nontuberculous mycobacteria among people with cystic fibrosis (HALT NTM) study: Rationale and study design.

BACKGROUND: Healthcare-associated transmission of nontuberculous mycobacteria (NTM) among people with cystic fibrosis (pwCF) has been reported and is of increasing concern. No standardized epidemiologic investigation tool has been published for healthcare-associated NTM outbreak investigations. This report describes the design of an ongoing observational study to standardize the approach to NTM outbreak investigation among pwCF. METHODS: This is a parallel multi-site study of pwCF within a single Center who have respiratory NTM isolates identified as being highly-similar. Participants have a history of positive airway cultures for NTM, receive care within a single Center, and have been identified as part of a possible outbreak based on genomic analysis of NTM isolates. Participants are enrolled in the study over a 3-year period. Primary endpoints are identification of a shared healthcare-associated encounter(s) among patients in a Center and identification of environmental isolates that are genetically highly-similar to respiratory isolates recovered from pwCF. Secondary endpoints include characterization of potential transmission modes and settings, as well as incidence and prevalence of healthcare-associated environmental NTM species/subspecies by geographical region. DISCUSSION: We hypothesize that genetically highly-similar strains of NTM among pwCF cared for at the same Center may arise from healthcare sources including patient-to-patient transmission and/or acquisition from environmental sources. This novel study design will establish a standardized, evidence-based epidemiologic investigation tool for healthcare-associated NTM outbreak investigation within CF Care Centers, will broaden the scope of independent outbreak investigations and demonstrate the frequency and nature of healthcare-associated NTM transmission in CF Care Centers nationwide. Furthermore, it will provide valuable insights into modeling risk factors associated with healthcare-associated NTM transmission and better inform future infection prevention and control guidelines. This study will systematically characterize clinically-relevant NTM isolates of CF healthcare environmental dust and water biofilms and set the stage to describe the most common environmental sources within the healthcare setting harboring clinically-relevant NTM isolates. TRIAL REGISTRATION: ClinicalTrials.gov NCT04024423. Date of registry July 18, 2019.

Dissemination of Mycobacterium abscessus via global transmission networks.

Mycobacterium abscessus, a multidrug-resistant nontuberculous mycobacterium, has emerged as a major pathogen affecting people with cystic fibrosis (CF). Although originally thought to be acquired independently from the environment, most individuals are infected with one of several dominant circulating clones (DCCs), indicating the presence of global transmission networks of M. abscessus. How and when these clones emerged and spread globally is unclear. Here, we use evolutionary analyses of isolates from individuals both with and without CF to reconstruct the population history, spatiotemporal spread and recent transmission networks of the DCCs. We demonstrate synchronous expansion of six unrelated DCCs in the 1960s, a period associated with major changes in CF care and survival. Each of these clones has spread globally as a result of rare intercontinental transmission events. We show that the DCCs, but not environmentally acquired isolates, exhibit a specific smoking-associated mutational signature and that current transmission networks include individuals both with and without CF. We therefore propose that the DCCs initially emerged in non-CF populations but were then amplified and spread through the CF community. While individuals with CF are probably the most permissive host, non-CF individuals continue to play a key role in transmission networks and may facilitate long-distance transmission.

Stepwise pathogenic evolution of Mycobacterium abscessus.

Although almost all mycobacterial species are saprophytic environmental organisms, a few, such as Mycobacterium tuberculosis, have evolved to cause transmissible human infection. By analyzing the recent emergence and spread of the environmental organism M. abscessus through the global cystic fibrosis population, we have defined key, generalizable steps involved in the pathogenic evolution of mycobacteria. We show that epigenetic modifiers, acquired through horizontal gene transfer, cause saltational increases in the pathogenic potential of specific environmental clones. Allopatric parallel evolution during chronic lung infection then promotes rapid increases in virulence through mutations in a discrete gene network; these mutations enhance growth within macrophages but impair fomite survival. As a consequence, we observe constrained pathogenic evolution while person-to-person transmission remains indirect, but postulate accelerated pathogenic adaptation once direct transmission is possible, as observed for M. tuberculosis Our findings indicate how key interventions, such as early treatment and cross-infection control, might restrict the spread of existing mycobacterial pathogens and prevent new, emergent ones.

Human pathogenic Mycobacterium kansasii (former subtype I) with zoonotic potential isolated from a diseased indoor pet cat, Japan.

Nontuberculous mycobacterial (NTM) infections in humans have increased in prevalence in recent decades. Mycobacterium kansasii is one of the most prevalent human pathogenic NTM species worldwide. Herein, we report the first isolation of M. kansasii from an indoor domestic cat in Japan. Comparative genome sequence analysis of the feline isolate showed this pathogen is genetically identical to human pathogenic M. kansasii. This finding suggests that M. kansasii has a potential risk of zoonoses and requires the "One Health" approach to control NTM infection.

Whole genome sequencing of Nontuberculous Mycobacterium (NTM) isolates from sputum specimens of co-habiting patients with NTM pulmonary disease and NTM isolates from their environment.

BACKGROUND: Nontuberculous mycobacterium (NTM) species are ubiquitous microorganisms. NTM pulmonary disease (NTM-PD) is thought to be caused not by human-to-human transmission but by independent environmental acquisition. However, recent studies using next-generation sequencing (NGS) have reported trans-continental spread of Mycobacterium abscessus among patients with cystic fibrosis. RESULTS: We investigated NTM genomes through NGS to examine transmission patterns in three pairs of co-habiting patients with NTM-PD who were suspected of patient-to-patient transmission. Three pairs of patients with NTM-PD co-habiting for at least 15 years were enrolled: a mother and a daughter with M. avium-PD, a couple with M. intracellulare-PD, and a second couple, one of whom was infected with M. intracellulare and the other of whom was infected with M. abscessus. Whole genome sequencing was performed using patients' NTM isolates as well as environmental specimens. Genetic distances were estimated based on single nucleotide polymorphisms (SNPs). By comparison with the genetic distances among 78 publicly available NTM genomes, NTM isolates derived from the two pairs of patients infected with the same NTM species were not closely related to each other. In phylogenetic analysis, the NTM isolates from patients with M. avium-PD clustered with isolates from different environmental sources. CONCLUSIONS: In conclusion, considering the genetic distances between NTM strains, the likelihood of patient-to-patient transmission in pairs of co-habiting NTM-PD patients without overt immune deficiency is minimal.

Dental Unit Waterlines in Quito and Caracas Contaminated with Nontuberculous Mycobacteria: A Potential Health Risk in Dental Practice.

Three cases of severe odontogenic infections due to nontuberculous mycobacteria (NTM) in Venezuela that were directly associated with dental procedures and the finding of dental unit waterlines (DUWLs) in dental offices that were colonized with mycobacteria species was the reason for assessing the water quality of DUWLs in dental offices in two capital cities in South America, namely, Quito and Caracas. The main water supplies and the water from 143 DUWLs in both cities were sampled and especially checked for contamination with NTM. To measure the overall bacteriological quality of the water also the presence of heterotrophic bacteria, coliform bacteria, and Pseudomonas was determined. Results showed that respectively 3% and 56% of the DUWLs in Quito and Caracas yielded NTM species (up to 1000 colony-forming units (CFU)/mL). Furthermore, high and unacceptable total viable counts of heterotrophic bacteria and/or coliform bacteria and Pseudomonas were detected in 73% of the samples. We conclude that, in both cities, the water in the majority of DUWLs was contaminated with NTM and other potential pathogens, presenting a risk to human health. The detection of NTM in DUWL water with acceptable heterotrophic bacteria counts shows the need to include NTM in water quality testing. Mycobacteria are more resistant to disinfection procedures than other types of vegetative bacteria, and most testing protocols for DUWLs do not assess mycobacteria and thus do not guarantee risk-free water.

Non-tuberculous mycobacteria and the rise of Mycobacterium abscessus.

Infections caused by non-tuberculous mycobacteria (NTM) are increasing globally and are notoriously difficult to treat due to intrinsic resistance of these bacteria to many common antibiotics. NTM are diverse and ubiquitous in the environment, with only a few species causing serious and often opportunistic infections in humans, including Mycobacterium abscessus. This rapidly growing mycobacterium is one of the most commonly identified NTM species responsible for severe respiratory, skin and mucosal infections in humans. It is often regarded as one of the most antibiotic-resistant mycobacteria, leaving us with few therapeutic options. In this Review, we cover the proposed infection process of M. abscessus, its virulence factors and host interactions and highlight the commonalities and differences of M. abscessus with other NTM species. Finally, we discuss drug resistance mechanisms and future therapeutic options. Taken together, this knowledge is essential to further our understanding of this overlooked and neglected global threat.

A bronchoscopy-associated pseudo-outbreak of Mycobacterium mucogenicum traced to use of contaminated ice used for bronchoalveolar lavage.

Clonal Mycobacterium mucogenicum isolates (determined by molecular typing) were recovered from 19 bronchoscopic specimens from 15 patients. None of these patients had evidence of mycobacterial infection. Laboratory culture materials and bronchoscopes were negative for Mycobacteria. This pseudo-outbreak was caused by contaminated ice used to provide bronchoscopic lavage. Control was achieved by transitioning to sterile ice.

Addressing the threat from within: Investigation of respiratory symptoms in a health care worker with untreated latent tuberculosis infection.

Tuberculosis (TB) in the health care worker demands orchestrated efforts from health care institutions to promptly identify cases and address community risk. We describe a pediatric intensive care unit nurse with latent TB infection who developed hemoptysis and a lung infiltrate concerning for active TB. Her evaluation and contact investigation were facilitated by our institution's command center. Although TB was ultimately ruled out, this case tested our team-based care in response to a suspected high-consequence pathogen.

Can environmental determinants explain Nontuberculous Mycobacteria geographic incidence?

INTRODUCTION AND OBJECTIVES: Nontuberculous mycobacteria (NTM) are opportunistic agents that have gained importance during the last decades due to their increasing incidence in high-risk populations. Their modes of transmission differ from person-toperson contact commonly described in Mycobacterium tuberculosis (MTB). In fact, NTM are frequently found in soil, natural waters and drinking-water distributions systems, emphasizing the contribution of environmental factors when discussing this disease's susceptibility. Our aim is to evaluate the incidence of NTM in Portugal and to identify the main environmental variables related to it. MATERIAL AND METHODS: We performed a cross-sectional study centred on 2011 (date of the latest Portuguese census) from collected personal features and environmental data available in public databases. Environmental values when only known at the district level were interpolated using inverse distance weighting. A semiparametric poisson model was used to estimate NTM incidence. The non-parametric part of the model was obtained by using thin plate smoothing splines defined on the spatial component of the data. RESULTS: 359 new NTM cases were notified during a five-year period. None of the environmental determinants studied was strong enough to predict NTM geographical incidence in Portugal (p>0.05), except for population density (p<0.001). Personal characteristics such as female sex (p<0.001), age (p<0.001) and Human Immunodeficiency Virus infection and Acquired Immune Deficiency Syndrome (HIV/AIDS) incidence (p<0.001) are associated with an increase of NTM disease incidence. CONCLUSIONS: NTM appears to be more common in elderly women, especially if they have HIV/AIDS disease or if they live in urban, highly populated areas. Overall, female sex seems to assume the most relevant role when discussing predisposition to NTM disease. However, further studies are needed to evaluate the impact on NTM geographical incidence by other environmental and personal variables not included in this one.

Prevention of transmission of Mycobacterium abscessus among patients with cystic fibrosis.

PURPOSE OF REVIEW: Pulmonary nontuberculous mycobacterial (NTM) infection is recognized as one of the most challenging infections to treat among cystic fibrosis patients. The source of NTM infection, modes of transmission, and exposure risks are poorly understood. Healthcare-associated transmission of Mycobacterium abscessus among cystic fibrosis patients has been suspected and is a growing concern for cystic fibrosis centers worldwide. This review describes our current understanding of prevention of healthcare-associated transmission of M. abscessus among patients with cystic fibrosis. RECENT FINDINGS: Multiple healthcare-associated outbreaks of M. abscessus among cystic fibrosis patients within cystic fibrosis care centers have been reported. The percentage of patients involved in the reported outbreaks, as well as the perceived impact of patient-to-patient transmission varies dramatically between the reporting centers and population surveys. Several groups have now proposed M. abscessus-specific measures to limit future outbreaks. SUMMARY: Improved NTM surveillance combined with a standardized, systematic approach to epidemiologic investigation of potential episodes of healthcare-associated transmission will help to reveal risk factors for NTM acquisition and inform future evidence-based infection prevention and control measures for patients with cystic fibrosis.

Transmission of Mycobacterium chelonae and Mycobacterium marinum in laboratory zebrafish through live feeds.

The zebrafish (Danio rerio) is a popular vertebrate model organism used in a wide range of research fields. Mycobacteriosis, caused by Mycobacterium species, is particularly concerning because it is a common disease associated with chronic infections in these fish. Infections are also a source of uncontrolled experimental variance that may influence research results. Live feeds for zebrafish are common and include paramecia (Paramecium caudatum), brine shrimp (Artemia franciscana) and rotifers (Branchionus spp.). Although nutritionally beneficial, live feeds may pose a biosecurity risk. In this study, we investigate transmission of Mycobacterium chelonae and Mycobacterium marinum through these three live feeds. We show that all three live feeds ingest both M. marinum and M. chelonae and can transmit mycobacterial infections to zebrafish. This observation emphasizes the need for live feeds to be included in the consideration of potential biosecurity risks. This study is of importance to other beyond the zebrafish community, including those of additional aquatic models and those using live feeds for other types of aquaculture.

A Global Survey of Mycobacterial Diversity in Soil.

Mycobacteria are a diverse bacterial group ubiquitous in many soil and aquatic environments. Members of this group have been associated with human and other animal diseases, including the nontuberculous mycobacteria (NTM), which are of growing relevance to public health worldwide. Although soils are often considered an important source of environmentally acquired NTM infections, the biodiversity and ecological preferences of soil mycobacteria remain largely unexplored across contrasting climates and ecosystem types. Using a culture-independent approach by combining 16S rRNA marker gene sequencing with mycobacterium-specific hsp65 gene sequencing, we analyzed the diversity, distributions, and environmental preferences of soil-dwelling mycobacteria in 143 soil samples collected from a broad range of ecosystem types. The surveyed soils harbored highly diverse mycobacterial communities that span the full extent of the known mycobacterial phylogeny, with most soil mycobacteria (97% of mycobacterial clades) belonging to previously undescribed lineages. While mycobacteria tended to have higher relative abundances in cool, wet, and acidic soil environments, several individual mycobacterial clades had contrasting environmental preferences. We identified the environmental preferences of many mycobacterial clades, including the clinically relevant Mycobacterium avium complex that was more commonly detected in wet and acidic soils. However, most of the soil mycobacteria detected were not closely related to known pathogens, calling into question previous assumptions about the general importance of soil as a source of NTM infections. Together, this work provides novel insights into the diversity, distributions, and ecological preferences of soil mycobacteria and lays the foundation for future efforts to link mycobacterial phenotypes to their distributions.IMPORTANCE Mycobacteria are common inhabitants of soil, and while most members of this bacterial group are innocuous, some mycobacteria can cause environmentally acquired infections of humans and other animals. Human infections from nontuberculous mycobacteria (NTM) are increasingly prevalent worldwide, and some areas appear to be "hotspots" for NTM disease. While exposure to soil is frequently implicated as an important mode of NTM transmission, the diversity, distributions, and ecological preferences of soil mycobacteria remain poorly understood. We analyzed 143 soils from a range of ecosystems and found that mycobacteria and lineages within the group often exhibited predictable preferences for specific environmental conditions. Soils harbor large amounts of previously undescribed mycobacterial diversity, and lineages that include known pathogens were rarely detected in soil. Together, these findings suggest that soil is an unlikely source of many mycobacterial infections. The biogeographical patterns we documented lend insight into the ecology of this important group of soil-dwelling bacteria.

Phagosome Escape of Rough Mycobacterium abscessus Strains in Murine Macrophage via Phagosomal Rupture Can Lead to Type I Interferon Production and Their Cell-To-Cell Spread.

Mycobacterium abscessus complex (MAB) is a rapidly growing mycobacterium(RGM) whose clinical significance as an emerging human pathogen has been increasing worldwide. It has two types of colony morphology, a smooth (S) type, producing high glycopeptidolipid (GPL) content, and a rough (R) type, which produces low levels of GPLs and is associated with increased virulence. However, the mechanism responsible for their difference in virulence is poorly known. By ultrastructural examination of murine macrophages infected, we found that MAB-R strains could replicate more actively in the macrophage phagosome than the S variants and that they could escape into cytosol via phagosomal rupture. The cytosolic access of MAB-R strains via phagosomal rupture led to enhanced Type I interferon (IFN) production and cell death, which resulted in their cell-to-cell spreading. This behavior can provide an additional niche for the survival of MAB-R strains. In addition, we found that their enhancement of cell death mediated cell spreading are dependent on Type I IFN signaling via comparison of wild-type and IFNAR1 knockout mice. In conclusion, our data indicated that a transition of MAB-S strains into MAB-R variants increased their virulence via enhanced Type I IFN production, which led to enhanced survival in infected macrophage via cell death mediated cell-to-cell spreading. This result provides not only a novel insight into the difference in virulence between MAB-R and -S variants but also hints to their treatment strategy.

Nontuberculous mycobacteria: Insights on taxonomy and evolution.

Seventy years have passed since Ernest H. Runyon presented a phenotypic classification approach for nontuberculous mycobacteria (NTM), primarily as a starting point in trying to understand their clinical relevance. From numerical taxonomy (biochemical testing) to 16S rRNA gene sequencing to whole genome sequencing (WGS), our understanding of NTM has also evolved. Novel species are described at a rapid pace, while taxonomical relationships are re-defined in large part due to the accessibility of WGS. The evolutionary course of clonal complexes within species is better known for some NTM and less for others. In contrast with M. tuberculosis, much is left to learn about NTM as a whole.

Infections with Mycobacterium chimaera and open chest surgery. An unresolved problem. / Infecciones por Mycobacterium chimaera y cirugía cardíaca. Un problema no resuelto.

Mycobacterium chimaera (M. chimaera) is a non-tuberculous mycobacterium of the Mycobacterium avium complex. Between 2013 and April 2018, more than 140 cases of severe infection have been identified, all of them in patients who had undergone cardiothoracic surgery with extracorporeal circulation. The epidemiological investigations have shown that the majority of infections have a common source: contaminated normo-hypothermia modules. These devices have a water circuit inside and cooling fans that produce an aerosol, which can be contaminated with M. chimaera, and disperse it into the air in the operating room. The clinical symptoms can take months, even years to appear after the extracorporeal surgery, with an average of 1.5 years. Sarcoidosis is the most frequently described entity as a misdiagnosis in these patients, so it is necessary to maintain a high diagnostic suspicion in all patients with a history of cardiac surgery with extracorporeal circulation.

Outbreak of Tattoo-associated Nontuberculous Mycobacterial Skin Infections.

BACKGROUND: On 29 April 2015, the Florida Department of Health in Miami-Dade County (DOH Miami-Dade) was notified by a local dermatologist of 3 patients with suspected nontuberculous mycobacterial (NTM) infection after receiving tattoos at a local tattoo studio. METHODS: DOH Miami-Dade conducted interviews and offered testing, described below, to tattoo studio clients reporting rashes. Culture of clinical isolates and identification were performed at the Florida Bureau of Public Health Laboratories. Characterization of NTM was performed by the Centers for Disease Control and Prevention and the US Food and Drug Administration (FDA), respectively. Whole-genome sequencing (WGS) and single-nucleotide polymorphism (SNP) analyses were used to construct a phylogeny among 21 Mycobacterium isolates at the FDA. RESULTS: Thirty-eight of 226 interviewed clients were identified as outbreak-associated cases. Multivariate logistic regression revealed that individuals who reported gray tattoo ink in their tattoos were 8.2 times as likely to report a rash (95% confidence interval, 3.1-22.1). Multiple NTM species were identified in clinical and environmental specimens. Phylogenetic results from environmental samples and skin biopsies indicated that 2 Mycobacterium fortuitum isolates (graywash ink and a skin biopsy) and 11 Mycobacterium abscessus isolates (5 from the implicated bottle of graywash tattoo ink, 2 from tap water, and 4 from skin biopsies) were indistinguishable. In addition, Mycobacterium chelonae was isolated from 5 unopened bottles of graywash ink provided by 2 other tattoo studios in Miami-Dade County. CONCLUSIONS: WGS and SNP analyses identified the tap water and the bottle of graywash tattoo ink as the sources of the NTM infections.