The growing impact of nontuberculous mycobacteria: A multidisciplinary review of ecology, pathogenesis, diagnosis, and treatment

Highlights

• •NTM infections are rising globally, with increasing pulmonary and extrapulmonary cases.
• •Molecular techniques like DNA probes and gene sequencing enable precise NTM identification.
• •NTM transmission through water systems and aerosol inhalation remains a significant risk.

Abstract

Nontuberculous mycobacteria (NTM) are emerging pathogens responsible for a growing spectrum of diseases, particularly in individuals with underlying lung disorders or immune suppression. Once considered primarily environmental saprophytes, NTM are now recognized as important causes of pulmonary, cutaneous, lymphatic, and disseminated infections. With more than 200 species identified and regional variations in prevalence, their diagnosis and management present significant clinical and microbiological challenges. The lack of standardized reporting systems and overlapping features with tuberculosis complicate epidemiological understanding and case identification.

This review provides an updated and integrated overview of NTM-associated diseases, emphasizing diagnostic advancements, environmental sources, mechanisms of transmission, host immunity, genetic susceptibility, and therapeutic options. Special attention is given to molecular diagnostic techniques, species-level identification strategies, and the role of gene sequencing in differentiating NTM species. We also highlight the limitations of conventional methods, discuss antimicrobial resistance mechanisms, and summarize current treatment guidelines.

By synthesizing current knowledge across microbiology, clinical medicine, and public health, this review aims to support a multidisciplinary approach to NTM diagnosis and management and address the pressing need for increased awareness, better surveillance, and targeted research on this under-recognized group of pathogens.

Graphical abstract

1. Introduction

The term nontuberculous mycobacteria (NTM) encompasses all mycobacterial species excluding the Mycobacterium (M.) tuberculosis complex (MTBC) and M. leprae.¹ NTM are also known as atypical mycobacteria or anonymous mycobacteria, mycobacteria other than tuberculosis, and environmental mycobacteria.² These organisms are commonly found across various environmental settings, such as domestic water supplies, natural aquatic environments, and soil.³ Thereby, NTM surround humans. Notably, the species M. avium, M. chelonae, M. fortuitum, M. gordonae, M. kansasii, and M. xenopi are the most frequently reported mycobacteria in drinking water and hospital water distribution systems.⁴ These organisms are classified as opportunistic pathogens in individuals with compromised immune systems. Furthermore, evidence highlights the detection of these pathogens in individuals with intact immune systems.^5,6 NTM encompasses around 200 distinct species, and accurately identifying differences at the subspecies (subsp.) level is essential for effective treatment.⁷ The majority of these species have been isolated from various environments and may cause pulmonary and extrapulmonary infections in susceptible persons.⁸ NTM infections are categorized into several distinct clinical entities, including chronic pulmonary disease, which remains the predominant manifestation, alongside lymphadenitis, cutaneous infections, and disseminated disease, each presenting unique diagnostic and therapeutic challenges.⁹

The 2024 guidelines emphasize that the classification of these conditions is critical for tailoring treatment strategies, with chronic pulmonary disease often linked to species such as M. avium complex (MAC) and requiring prolonged antimicrobial therapy, while lymphadenitis predominantly affects pediatric populations and may necessitate surgical intervention. Cutaneous and disseminated forms, though less common, are associated with underlying immunosuppression and demand a multidisciplinary approach to manage effectively.¹⁰

While many NTM species, such as MAC and M. abscessus, are well-recognized pathogens, others, including M. chimaera and M. stomatepiae, are newly emerging, and their pathogenic potential is yet to be understood. M. chimaera causes severe healthcare-associated infections, notably after cardiac surgery, while M. stomatepiae is linked to rare human pulmonary infections with potential zoonotic origins.^11,12 In recent years, there has been a notable increase in NTM infections, prompting attention from healthcare professionals.¹³

2. Classification

The Mycobacterium genus encompasses the M. tuberculosis complex, a primary causative agent of tuberculosis (TB), a leading infectious disease globally, and the M. leprae complex, responsible for leprosy, a chronic granulomatous condition.¹⁴ Distinct from these pathogens, NTM constitute a diverse group with distinct classification criteria. According to Bergey’s Manual of Systematic Bacteriology (2nd ed.), NTM are categorized based on growth rate and pigment formation into slowly growing mycobacteria, requiring ≥ 7 days to form visible colonies, and rapidly growing mycobacteria (RGM), forming colonies in < 7 days.¹⁵ The Runyon classification further subdivides NTM into four groups: Type I (photochromogens), Type II (scotochromogens), Type III (nonchromogens), and Type IV (rapid growers), distinguished by their pigment production on Löwenstein–Jensen media. Key pathogenic species include M. kansasii, M. marinum, M. simiae, and M. xenopi among slowly growing mycobacteria, and M. abscessus complex (MABC), M. chelonae, and M. fortuitum among RGM.^16,17 Table 1 presents the classification of mycobacterial species, highlighting pathogenic NTM associated with human diseases.

Table 1.

Classification of mycobacterial species: NTM subgroups by growth rate and Runyon groups, highlighting pathogenic species.14, 15, 16

Group/Species	Runyon classification
M. tuberculosiscomplex
M. tuberculosis
M. bovis
M. africanum
M. microti
M. lepraecomplex
M. leprae
M. lepromatosis
Nontuberculous mycobacteria (NTM)
Slowly growing NTM (≥ 7 days)
M. kansasii	• Runyon I (Photochromogen)
M. marinum
M. simiae
M. gordonae	• Runyon II (Scotochromogen)
M. scrofulaceum
M. avium complex (MAC):	• Runyon III (Nonchromogen)
M. avium
M. intracellulare
M. chimaera
M. malmoense
M. ulcerans
M. xenopi
Rapidly growing NTM (< 7 days)
M. chelonae	• Runyon IV
M. fortuitum
M. abscessus complex:
M. abscessus subsp. abscessus
M. abscessus subsp. massiliense
M. abscessus subsp. bolletii

3. Epidemiology

Recent statistics show a significant rise in pulmonary NTM infections, with increases of 82.1 % for infections and 66.7 % for related diseases. The annual rate of NTM infections and diseases has risen by approximately 4.0 % and 4.1 % per 100,000 persons per year, respectively, with notable increases in infections caused by the MAC and MABC. Table 2 summarizes current prevalence and incidence rates reported in various regions and high-risk populations.¹⁸

Table 2.

Recent epidemiological estimates of nontuberculous mycobacterial pulmonary disease (NTM-PD) in general and high-risk populations.

Region/Population	Measurement	Value
United States (general population)	Pulmonary NTM disease prevalence	13.9 per 100,000 (2013 data)
Germany (2009–2014)	Pulmonary NTM disease prevalence	3.3 per 100,000
Japan (2001–2009)	Pulmonary NTM disease incidence	10.1 per 100,000
Republic of Korea (age-adjusted, 2016)	NTM infection prevalence	33.3 per 100,000
Australia (2000)	NTM pulmonary disease prevalence	0.56 per 100,000
Queensland, Australia (2005)	NTM pulmonary disease prevalence	3.2 per 100,000
New Zealand (2004)	NTM pulmonary disease prevalence	1.17 per 100,000
Adults with non-CF bronchiectasis	Global NTM prevalence	≈ 10 %–14 %
CF patients, USA (2010–2019)	NTM pulmonary infection prevalence	58 per 1,000 persons tested; ↑3.5 % per year

Abbreviations: NTM, nontuberculous mycobacteria; CF, cystic fibrosis.

Contextualize trends and key findings

• •Growing burden: Pulmonary NTM prevalence is rising globally, with reports like 13.9/100,000 in the USA (2013), 33.3/100,000 in the Republic of Korea (2016), and 10.1/100,000 in Japan (2001–2009), signaling regional variations.
• •Geographic disparities: Australia's general population prevalence (≈ 0.56/100,000–3.2/100,000) is lower compared to North America and East Asia (≈ 3/100,000–14/100,000).
• •Comorbidity risk group: In adult bronchiectasis, NTM prevalence is notably high (≈ 10 %–14 %).
• •Cystic fibrosis (CF): Prevalence among USA CF patients reached 58 per 1,000 tested, with a significant annual increase of 3.5 % over a 10-year span.19, 20, 21

4. NTM in the environment

NTMs are globally distributed and widely present in the environment. Their survival and persistence are supported by several physiological traits—most notably cell surface hydrophobicity, which facilitates surface attachment, aerosolization via air bubbles, and resistance to antimicrobials. Additional survival mechanisms include biofilm formation, association with amoebae, and chlorine resistance, all of which contribute to their persistence in water distribution systems.22, 23, 24 NTM, as oligotrophs, can grow in water with very low levels of nutrients, specifically at concentrations of assimilable organic carbon ≥ 50 µg/L, making engineered water systems such as household plumbing and drinking water distribution systems ideal habitats for their persistence.²⁵ Their presence in drinking water can lead to true or pseudo-infections. NTM resist common disinfectants and tolerate a broad range of pH and temperatures, allowing them to persist in hospital settings. Inadequately disinfected reservoirs and medical equipment can become sources of nosocomial infections, highlighting the need to monitor hospital environments for NTM contamination.^26,27

Geographic variation in NTM distribution is significant—California and Hawaii are hotspots for pulmonary NTM disease in the USA. An environmental study in Hawaii found NTMs in 27 % of samples, primarily species of clinical concern such as M. chimaera, M. chelonae, and M. abscessus. Growth is enhanced in stagnant water, particularly in shower systems, while flushing can reduce bacterial load. Moreover, animals such as fish, pigs, and badgers may serve as reservoirs for pathogenic NTM species.^28,29

5. Mechanism of transmission

The presence of NTM in natural and constructed drinking water distribution systems has been extensively documented. Consequently, aerosol inhalation from urban or personal water systems is believed to be a significant route of acquisition.^30,31 The hypothesis that the rise in pulmonary infections by NTM over recent decades is linked to increased use of showers rather than baths has been advanced.³² The hypothesis of water transmission is further substantiated by a study demonstrating the genetic similarity of MAC isolates in pulmonary secretions and household water systems. The presence of MAC in shower biofilms in the USA further corroborates the notion that people are constantly exposed to NTM. Nosocomial outbreaks of NTM in patients who received solid organ transplants and hematopoietic stem cell transplants have been associated with the hospital water supply.^27,33,34 While NTM transmission has been predominantly linked to environmental sources, probable nosocomial human-to-human transmission has been documented in patients with CF admitted to a healthcare center, where identical strains of M. abscessus subsp. massiliense were identified by whole-genome sequencing.² A possible case of household human-to-human transmission of M. kansasii has also been reported.34, 35, 36 Comparative genomics is being utilized to examine the genetic diversity of NTM strains from various sources, providing insights into their transmission and resistance mechanisms.^28,29

6. Pathogenesis and virulence factors

The capacity of a microbe to bind fibronectin may promote its colonization of the host by facilitating its attachment to the extracellular matrix in areas of epithelial damage. The process of fibronectin attachment is facilitated by a protein known as fibronectin attachment protein (FAP), which is necessary for the efficient attachment and invasion of epithelial cells by M. avium subsp. paratuberculosis. FAP is present on the surface of M. avium and, by binding to fibronectin on mucosal surfaces, serves to facilitate more specific mycobacterial binding to integrin receptors. The opsonization of NTM with complement components, such as C3b and C4b, has also been observed to facilitate their entry into macrophages. Other macrophage receptors implicated in the phagocytosis of NTM include mannose receptors and scavenger receptors.37, 38, 39 The multiple pathways by which NTM can enter macrophages suggest that the intracellular niche is a favorable adaptation for the bacilli if the phagocytes are not primed to kill the bacteria. The mechanisms employed by NTM to evade the immune system and persist within macrophages include the inhibition of phagosome-lysosome fusion, the transition to an anaerobic intracellular environment, the induction of NTM-related genes that enhance replication, the direct inhibition of host macrophage function and lymphocyte proliferation, and potentially through the induction of macrophage apoptosis.^38,40

It is well established that the virulence of NTM can vary significantly between different species. For instance, the MAC has been observed to demonstrate a higher degree of pathogenicity in comparison to M. avium. Furthermore, certain strains of NTM have been shown to exhibit a phenomenon of hypervirulence, which can result in the progression of lung disease and a subsequent poor prognosis.³¹ In the context of pulmonary nontuberculous mycobacteria, numerous NTM strains have been observed to infect humans through adherence to the respiratory mucosa. In contrast to M. tuberculosis, which appears to adhere to healthy mucosa via a FAP, NTM appears to adhere to damaged mucosa. The ability of NTM to form biofilms on tissues and their capacity to inhibit inflammatory cytokine production represent mechanisms that can subvert host immunity, thereby promoting colonization and subsequent invasion of the bronchial epithelium. NTM are classified as intracellular parasites, and the effectiveness of opsonization by complement also varies by species. The cellular envelope, specifically the lipoarabinomannan moiety, has been identified as a critical factor influencing intracellular survival. This glycolipid, along with other components of the mycobacterial envelope, exhibits variation between species. Some NTM, notably RGM, have been observed to demonstrate a high degree of resistance to infections, chemicals employed in the sterilization of medical equipment, and topical antiseptics. These observations underscore the potential for differences in virulence among species and contribute to the understanding of the pattern of disease produced. Finally, some NTM can acquire virulence genes from other bacteria in specific microenvironments, such as the lungs of patients with CF, and become more dangerous.^37,41,42

7. Risk factors

Risk factors for human infection include Mendelian susceptibility to mycobacterial disease (MSMD), chronic obstructive pulmonary disease (COPD), α−1-antitrypsin deficiency, CF, heterozygosity for cystic fibrosis transmembrane conductance regulator (CFTR) mutations, gastroesophageal reflux disease, and immunodeficiency due to HIV infection.⁴² In adults, a previous history of tuberculosis disease is a risk factor for NTM pulmonary disease, probably due to structural damage of the lung (such as in bronchiectasis) altering mucociliary clearance and thereby predisposing the lung tissue to NTM isolation and disease. NTM diseases have been reported in pediatric patients with cancer, in whom bloodstream NTM infections are associated with the use of central venous catheters, and disseminated forms of NTM disease are known to have occurred in recipients of organ transplants or hematopoietic stem cell transplants and patients receiving immune therapies directed against T cells. In some cases, increased susceptibility of human hosts to NTM has been observed due to mutations in five genes (IFNGR1, IFNGR2, IL12RB1, IL12B, and STAT1).⁴³ Furthermore, clinical treatments such as chemotherapy in cases of cancer treatment and organ transplantation render these patient groups more vulnerable to infections. Additional risk groups include neonates, intensive care unit patients, and elderly persons whose immune systems have become weakened. A preponderance of literature documents infections in immunocompromised individuals, such as those with AIDS. Additionally, there is evidence of an association between NTM disease and the SLC11A1 or natural resistance-associated macrophage protein 1 (NRAMP1) gene. The function of NRAMP1 is to regulate intramacrophage iron concentrations, thereby limiting the availability of iron for intracellular bacteria, as demonstrated in M. bovis residing within the phagolysosome.^26,31,42,44, 45, 46

7.1. Climatic conditions

Recent research has advanced our understanding of environmental risks related to NTM, showing how climate and environmental factors influence their distribution. Studies from the University of Colorado reveal specific pH and temperature conditions favor different NTM species, which can be predictive of their presence and virulence. Community science initiatives in high-incidence areas like Hawaii have facilitated broader sampling in natural and indoor environments, highlighting the need to explore potential NTM exposure sources in built settings, such as plumbing and aerosols.^47,48

The incidence of NTM is rising globally, with notable increases in Canada and the USA. linked to factors like trace metals in municipal water supplies, which may enhance mycobacterial metabolism and human susceptibility. Additionally, environmental minerals like calcium and molybdenum have been associated with higher pulmonary NTM infection risks.⁴⁹ In Queensland, Australia, geospatial analysis revealed clusters of M. avium complex cases tied to temperature and rainfall patterns. Understanding NTM's incubation periods and disease association with population density remains complex. Studies emphasizing precise geolocation are being conducted to identify high-risk areas systematically. As environmental conditions change and extreme weather becomes more prevalent, regular screening in susceptible populations will be crucial for monitoring exposure risks and understanding geographic trends in NTM incidence.⁴⁸ Elevated global temperatures foster NTM growth in water and soil, increasing human exposure. Understanding these dynamics is essential for forecasting and mitigating future NTM infection risks.⁵⁰

A 2020 Queensland study (2001–2016) found a nearly 2.3-fold increase in NTM incidence (from 11.1 to 25.9 per 100,000), with strong correlations to temperature and rainfall variations—though effects varied by species and region.⁵¹ A 2024 UK modeling study projected a 6.2 % rise in NTM infections over the next decade, driven by increasing precipitation and temperatures, highlighting the climate's influence on environmental reservoirs.⁵⁰ Mechanistically, floods and storms redistribute NTM from soil into water and aerosols; warm conditions favor environmental growth, as seen after Louisiana hurricanes and in high-temperature, humid regions.52, 53 Rising global temperatures, extreme weather, and altered hydrology likely increase NTM exposure, necessitating adaptation in surveillance and prevention strategies.

7.2. Host immunity and genetic susceptibility

In immunocompetent individuals, NTM are often regarded as commensal bacteria and considered to be unlikely pathogens. Several studies have highlighted the role of genetic susceptibility, particularly mutations in the CFTR gene, in increasing the risk of developing NTM‑PD. A significant proportion of patients with NTM‑PD have been found to harbor CFTR mutations. Additionally, other genetic factors—such as deficiencies in α₁‑antitrypsin, polymorphisms in the vitamin D receptor, and variations in genes involved in immune regulation—have been implicated in disease susceptibility.54, 55, 56, 57 In comparison with M. tuberculosis, in the absence of primary or secondary immunodeficiencies, the host immune system is usually capable of eradicating NTM via innate and acquired immune mechanisms. Notable insights into the protective immune systems' response to mycobacteria have emerged from studies of patients afflicted with disseminated forms of NTM. MSMD is a rare syndrome that increases the risk of developing clinical diseases caused by weakly virulent mycobacteria, such as Bacille Calmette-Guérin (BCG) vaccines and environmental mycobacteria. MSMD-causing mutations have been identified in six autosomal genes involved in interleukin 12 and 23 dependent, IFN-γ-mediated immunity. Mutation in the genes encoding the IFN-γ receptor has been observed to lead to disseminated NTM disease, BCG-osis (disseminated BCG), or TB. The majority of genetic defects underlying MSMD are autosomal recessive mutations in the β1 subunit of the interleukin 12 receptor (IL12Rβ1) or autosomal mutations in interferon γ receptor 1 (IFNGR1), which affect the interleukin 12/23-mediated and interferon γ-mediated interactions between macrophages and T cells.^30,58 Furthermore, a mutation in the STAT1 gene has been linked to susceptibility to mycobacterial infection. The identification of mutations in 10 distinct genes in humans has contributed to our enhanced comprehension of the response to mycobacterial infection, which is contingent upon the integrity of the Th1 cytokine pathway. These findings collectively suggest that host genetic predisposition plays a crucial, though complex and multifactorial, role in the pathogenesis of NTM‑PD.

The efficacy of the BCG vaccine in preventing tuberculosis varies significantly among different regions, ranging from 0 % to 89 %. Some have posited that populations with lower BCG vaccine efficacy may exhibit a higher prevalence of cross-reactive NTM. Notably, in certain countries, such as northern and eastern Europe, where the BCG vaccination program has been discontinued, there has been an observed increase in NTM lymphadenitis among children. However, there is a paucity of research addressing the prevalence of NTM infections in the context of the BCG vaccination program, highlighting the need for further investigation to elucidate the role of the BCG vaccination in the epidemiology of NTM infections.58, 59, 60

8. Nontuberculous mycobacterial diseases

For many years, it was thought that NTM were implicated in human disease only as saprophytic contaminants in tuberculous lesions. However, recent advancements in the field have led to the recognition of several species as facultative parasites capable of inducing chronic granulomatous diseases that can be pathologically indistinguishable from tuberculosis. These infections, however, remain challenging to identify due to the complexity of distinguishing the causative bacteria from M. tuberculosis using conventional methods. However, recent attention has been drawn to these organisms for two primary reasons. Firstly, in regions where tuberculosis has declined, these infections now account for a greater proportion of granulomatous disease cases. Secondly, these infections have emerged as prevalent secondary infections among patients with AIDS. The majority of localized infections are caused by the inoculation of organisms into the skin. Pulmonary infection has been observed in patients with predisposing diseases, while disseminated infection is predominantly seen in patients with impaired immune responses.^61,62

The global impact of NTM diseases is not well understood, primarily because most NTM infections do not require mandatory reporting, except for specific cases like M. chimaera related to cardiac surgeries and M. ulcerans. There are significant challenges in assessing the incidence, prevalence, and regional trends of NTM diseases since many cases go unrecorded. Some countries monitor NTM pulmonary cases through TB programs, but these figures are likely underestimated, as they typically only include patients from TB registries, excluding those who do not seek treatment at TB clinics.⁶³

While NTMs were initially detected in the late 19th century, their role as significant human pathogens gained broader acknowledgment only in the late 1950s.³⁶ Currently, more than 90 species have been identified in human samples, and several others remain either unclassified or unidentified. Human infections are predominantly associated with slow-growing species, such as the MAC and the MABC, which account for 80 % of clinical specimens.⁶⁴

Diseases in humans caused by NTM are categorized into four main clinical types: chronic lung disease, lymphadenitis, skin infections, and widespread disseminated disease.¹ Chronic lung disease is the most prevalent form, accounting for roughly 90 % of NTM infections affecting the respiratory system. Other cases impact lymph nodes, skin, soft tissues, and bones. Rarer instances include keratitis, middle ear infections, central nervous system involvement, and systemic infections.^65,66 Pulmonary NTM infections typically present as chronic lung diseases resembling tuberculosis, with symptoms including cough, fatigue, weight loss, and dyspnea. In contrast, extrapulmonary NTM infections can affect various body parts, leading to a wide range of symptoms depending on the affected site.⁶⁷

8.1. Localized cutaneous lesions

The injection of organisms into superficial abrasions and puncture wounds can result in the formation of localized nodular or ulcerative lesions. The organisms most commonly implicated are M. marinum, which colonizes swimming pools and fish aquaria, and M. ulcerans, which is largely restricted to some tropical regions and causes deep necrotic lesions known as Buruli ulcers. M. haemophilum has more recently been associated with similar lesions in immunosuppressed patients. Abscesses resulting from contaminated injections have most frequently been attributed to two rapidly growing species, M. chelonae and M. fortuitum. The prevalence of such cases is expected to rise among individuals with substance use disorders who are immunosuppressed due to AIDS. However, the majority of reported cases have occurred in diabetics or following the administration of contaminated drugs and vaccines.⁶⁸

8.2. Lymphadenitis

NTM-related lymphadenitis occurs most commonly in children under the age of five and typically presents as a unilateral, chronic, and self-limiting cervical lymph node enlargement. The most frequently involved site is the submandibular lymph nodes, which account for approximately 87 % of cases.⁶⁹ The primary route of entry is through the oropharyngeal mucosa or following penetrating trauma to the cervical region. In adults, the condition is less common and usually associated with underlying immunosuppressive conditions. The most common causative organism is MAC, responsible for approximately 80 % of NTM lymphadenitis cases. Other frequently reported species include M. scrofulaceum, M. kansasii, and M. fortuitum.38, 70

8.3. Disseminated disease

The development of granulomatous disease, characterized by the presence of one or multiple foci, has been observed in virtually any system or organ. In cases where cellular immunity is compromised, the dissemination of the infection can occur with a rapidity comparable to that observed in miliary tuberculosis. The majority of these cases have been attributed to the MAC and M. chelonae. Disseminated NTM disease is most commonly observed in patients with advanced HIV disease, particularly those with CD4⁺ cell counts less than 50 cells/mm³. While these infections predominantly manifest in HIV-infected patients, there has been a noted increase in the prevalence of disseminated NTM infection cases among non-HIV-infected patients. Disseminated disease in non-HIV patients is usually found in those with severe immunosuppression from other conditions, such as organ transplantation, hematologic malignancies, and chronic steroid use.^30,71

8.4. Pulmonary disease

Among the four principal clinical manifestations of NTM infections, pulmonary disease (NTM‑PD) is the most common and clinically significant form.⁷² The majority of documented cases are attributed to MAC, MABC, and M. kansasii, with M. xenopi being less frequently implicated.^73,74 However, in certain geographic regions, M. scrofulaceum, M. chelonae, M. szulgai, and M. malmoense have also been identified as important pathogens.⁷⁵ A wide array of predisposing conditions has been recognized, including chronic bronchitis, occupational dust‑related lung diseases, residual pulmonary lesions from prior tuberculosis, CF, malignancy, AIDS, and other causes of immunosuppression.^76,77

8.5. Clinical manifestations of NTM pulmonary disease

Pulmonary NTM disease presents in several distinct clinical-radiologic patterns, with the two most recognized being the nodular–bronchiectatic and fibrocavitary forms. The nodular–bronchiectatic form appears predominantly in postmenopausal women without prior lung disease—often referred to as “Lady Windermere syndrome”—and is typified by chronic cough with sputum, occasional hemoptysis, and fatigue. Characteristic imaging on high-resolution CT includes centrilobular nodules and multifocal bronchiectasis localized to the middle lobe and lingula.⁷⁶ Conversely, the fibrocavitary variant, seen more in older men with COPD or smoking history, is marked by thin-walled cavities primarily in the upper lobes, with less surrounding fibrosis compared to tuberculosis. Both phenotypes often share nonspecific systemic symptoms such as weight loss, malaise, and low-grade fever.

Other less common presentations include solitary pulmonary nodules resembling malignancy, hypersensitivity pneumonitis (“hot tub lung”), and disseminated infection in immunocompromised hosts.⁷⁸ Physical examination may reveal crackles, wheezes, or rhonchi, largely reflecting underlying bronchiectasis or COPD.⁷⁹ A Japanese multicenter study categorizes five phenotypes—fibrocavitary, nodular–bronchiectatic, solitary nodular, hot tub lung, and systemic forms—with the fibrocavitary and nodular–bronchiectatic types being most relevant for pulmonary surgical or prolonged antimicrobial intervention.⁸⁰ In addition to the above clinical presentations, the diagnosis of NTM-PD is based on a combination of clinical, radiological, and microbiological findings according to ATS/IDSA criteria.³⁸, which include: (1) Clinical symptoms such as chronic cough, fatigue, and weight loss; (2) Radiographic abnormalities like nodular-bronchiectatic or fibrocavitary patterns on chest CT; (3) Microbiological evidence, such as at least two positive sputum cultures from separate samples, or one positive culture from a bronchoalveolar lavage (BAL) specimen, or histological features (e.g., granulomatous inflammation) with at least one positive culture.

These criteria provide a practical framework for confirming NTM lung disease and distinguishing infection from colonization.

9. Diagnosis of NTM infections

9.1. Laboratory and radiologic detection

For the detection of NTM, staining and culture methods are primary laboratory techniques. Acid-fast staining methods, such as Ziehl-Neelsen or fluorochrome (auramine) staining, are used to identify acid-fast bacilli (AFB) in clinical samples. AFB staining cannot distinguish between M. tuberculosis and NTM. For sample decontamination, the standard 1 % N-acetyl-l-cysteine (NaLC)-NaOH method is used, though increasing its concentration is not recommended. In CF patients, combining NaLC—NaOH with 5 % oxalic acid reduces contamination rates but has a limited impact on improving culture yield. Chlorhexidine 1 % increases NTM culture yield on solid media compared to NaLC—NaOH, but is limited in liquid media due to interference with lecithin. Both liquid and solid media are used for NTM culture. Liquid media are preferred due to faster growth detection (typically 1–3 weeks) and higher sensitivity, particularly for samples with low bacterial loads. Cultures are incubated at 35–37°C, with growth typically detected within 2–8 weeks depending on the species.^38,81

Radiologically, imaging typically reveals nodular-bronchiectatic or fibrocavitary patterns. For diagnosing NTM-PD, sputum samples are usually sufficient for the fibrocavitary form, but BAL culture is more sensitive for the nodular bronchiectatic form. Microbiologically, culturing mycobacteria in both liquid and solid media at 37°C enhances diagnostic sensitivity, and obtaining at least three respiratory samples is recommended to confirm the diagnosis. In nodular bronchiectatic disease, where bacterial load is generally lower, studies have shown that BAL culture sensitivity exceeds that of sputum culture.81, 82, 83 Table 3 summarizes the diagnostic advantages and limitations of commonly used clinical, microbiological, and imaging-based methods for the detection of NTM-PD.^84,85

Table 3.

Diagnostic methods and criteria for NTM-PD.

Method	Advantages	Limitations
Clinical + Imaging	Symptoms (cough, weight loss, haemoptysis) + HRCT showing nodules, cavities, bronchiectasis help select patients for testing.	Findings are nonspecific; overlap with TB, cancer; requires microbiological confirmation.
AFB Smear (Microscopy)	Rapid, low-cost; suggests organism burden (higher smear positivity correlates with worse outcomes).	Low sensitivity; cannot differentiate NTM vs. M. tuberculosis.
Culture (Solid/Liquid media)	Gold standard; allows species ID & DST; enables multiple sampling (≥ 2 sputa or 1 BAL).	Slow growth—weeks for results; requires multiple specimens; risk of contamination.
Histology from Biopsy	Provides granulomatous confirmation; helpful if culture negative but pathology positive.	Invasive; requires specialized procedure; may miss organisms; culture still needed.
Molecular methods (PCR, Gene sequencing: 16S rRNA, hsp65, rpoB)	High specificity; rapid species/subspecies ID; macrolide-resistance detection in M. abscessus.	High cost; limited available probes (commercial kits may miss rare species); moderate sensitivity direct from samples.
MALDI-TOF MS	Fast and cost-effective once available; good species-level ID in culture.	Dependent on quality of spectrum database; may misidentify closely related species.
Culture-free PCR/sequencing	Rapid detection (∼days); direct from specimens; avoids slow culture.	Limited sensitivity (29 %–76 % detection vs. culture); restricted to common species.

Abbreviations: NTM-PD, nontuberculous mycobacterial pulmonary disease; HRCT, high-resolution computed tomography; TB, tuberculosis; AFB, acid-fast bacilli; BAL, bronchoalveolar lavage.

Because NTMs are ubiquitous in the environment and can contaminate clinical specimens from non-sterile sites, the diagnosis of NTM infection can be challenging. Contamination can occur before, during, and even after specimen collection. For example, collected sputum samples may be contaminated if rinsed in the mouth with tap water prior to expectoration. Contamination of the fibrotic bronchoscope suction channel with M. chelonae has also been reported as a cause of pseudo-epidemics. To differentiate between infection and contamination, the diagnosis of NTM disease should be established by a combination of bacteriologic, clinical, radiologic, and histologic principles.^86,87 Laboratory data alone cannot differentiate between true infections and transient colonization, highlighting the need for additional diagnostic methods to fully understand the scope of NTM diseases.

9.2. Molecular identification of NTM species

Correct species identification is very important because NTM species differ in their clinical relevance. The limitations of conventional methods in the identification of mycobacteria have led to the development of molecular techniques that have the advantage of being rapid, sensitive, and specific. Molecular techniques—particularly gene sequencing and line probe assays—have now replaced traditional biochemical methods based on mycolic acid analysis as the preferred strategy for NTM identification.⁴¹ Conventional biochemical identification methods can be misleading due to species phenotypic variation, non-standardized assay conditions, and their time-consuming nature. Among molecular methods, partial gene sequencing allows a higher level of discrimination, often down to the subspecies level, but is only feasible for laboratories with access to sequencing facilities.^5,88 Molecular DNA probes have been used to identify MAC, M. kansasii, and M. gordonae; however, this procedure is expensive, and probes are not available for all mycobacterial species. PCR restriction analysis of the hsp65 and rpoB genes is another molecular technique for identifying mycobacteria based on differences in restriction fragments of these genes.89, 90, 91 Sequence analysis of the rpoB, hsp65, 16S-23S rRNA internal transcribed spacer region, and 16S rRNA genes has recently been expanded as another method for NTM speciation.92, 93, 94

The widespread use of these methods has also led to the discovery of new species and the taxonomic reclassification of others, increasing the number of NTM species.^4,88,95,96 A new tool for the accurate identification of the M. tuberculosis complex and different NTM species is nucleotide MALDI-TOF-MS.⁹⁷ In Table 4, a comprehensive summary of various molecular techniques for the analysis of NTM is presented, revolutionizing NTM classification and phylogeny while highlighting recent advances in the precise identification of species and subspecies.

Table 4.

Summary of different molecular techniques for NTM analysis.⁷

Method	Description	Key Features
Repetitive sequences involving NTM	Utilizes insertion sequences (e.g., IS1245, IS1311) for molecular typing, highlighting genomic diversity among NTM strains.	Distinct IS types relevant to MAC strains; useful for epidemiological studies and differentiating specific strains like M. avium subsp. paratuberculosis.
Trinucleotide repeat sequence (TRS)	Found within mycobacterial genomes and aids in genotyping via PGRS-RFLP methods for species differentiation.	Improves resolution in genotyping; occurs within PE and PPE gene families; supports comparative strain analysis.
Enterobacterial repetitive intergenic consensus (ERIC)	ERIC sequences introduce variability for ERIC-PCR typing across mycobacterial species, enhancing insights into genetic diversity.	Characteristics include imperfect palindromic sequences; strengthens molecular epidemiology and phylogenetic understanding.
Multilocus variable number of tandem repeat analysis (MLVA)	Examines tandem repeat regions in the NTM genome, supporting strain differentiation and phylogenetic analysis.	Utilizes MPTR and ETR sequences; high discriminatory capacity; well-suited for various NTM species.
Repetitive element palindromic PCR (Rep-PCR)	Creates strain-specific DNA profiles using an arbitrary primer, effective for strain analysis despite reproducibility issues.	Matches or exceeds pulsed-field gel electrophoresis (PFGE) power for species prone to fragmentation; can be used for genotyping.
Genome Analysis	Involves comprehensive analysis of an organism's complete genome, offering insights into genetic variations.	Provides detailed strain characteristics and evolutionary links; highlights regulatory elements.
PFGE	Employs restriction enzyme digestion for DNA fragment pattern generation, useful for typing slow-growing NTM species.	High repeatability and exceptional separation capacity, suited for epidemiological studies despite high costs and long processing times.
Random amplified polymorphic DNA (RAPD)	Similar to Rep-PCR, this technique generates strain-specific profiles with limitations in reproducibility.	Effective for strain analysis; rivals PFGE's discriminatory powers, though reliability issues exist.
Amplified fragment length polymorphism (AFLP)	Utilizes dual restriction enzymes to distinguish closely related mycobacterial species.	Offers refined tools for differentiation, particularly for closely related species like M. hemophilum and MAC.
Large sequence polymorphism (LSP)	Acts as a molecular marker for mycobacterial genetic diversity, employing targeted PCR and microarray technology.	Generates genomic insights but requires significant DNA amounts and targeting optimization for various NTM subtypes.
Whole genome sequencing (WGS)	Provides precise mycobacterial species identification and insights into evolutionary paths and virulence genes.	Facilitates identification of new species and outbreak mapping with high accuracy; predicts virulence-related genes.
Next-generation sequencing (NGS)	Advances in high-throughput sequencing aiding in NTM detection and genetic analysis.	Rapid and efficient, bypasses traditional culture limitations while detecting novel species.
Targeted next-generation sequencing (tNGS)	Focuses on specific genes for identifying virulence and resistance factors crucial for treatment strategies.	More cost-effective and faster than WGS, though may overlook variants outside targeted areas.
Macrolide and rifampin resistance	Resistance primarily results from genetic mutations in 23S rRNA and the rpoB gene, crucial for treatment effectiveness.	erm(41) gene variations aid subspecies differentiation of M. abscessus, significantly impacting clinical strategies.
Clinical applications of molecular techniques	Advances in detection and genotyping of NTM species enable understanding of antimicrobial resistance patterns.	Techniques include MLVA, Rep-PCR, WGS, and NGS for accurate species identification and tailored treatment strategies.
Revolutionizing NTM classification and phylogeny	Molecular techniques like 16S rRNA amplification enhance understanding of NTM classification despite resolution limits.	Integration of genomic methods allows for precise species classification and valuable phylogenetic insights.
Integration of advanced diagnosis technologies	Technologies like MALDI-TOF MS improve NTM identification while facing challenges in complex samples.	MALDI-TOF MS is rapid for pure cultures.

Abbreviations: NTM, nontuberculous mycobacteria; MAC, M. avium complex; TRS, trinucleotide repeat sequence; ERIC, enterobacterial repetitive intergenic consensus; MLVA, multilocus variable number of tandem repeat analysis; Rep-PCR, repetitive element palindromic PCR; PFGE, pulsed-field gel electrophoresis; RAPD, random amplified polymorphic DNA; AFLP, amplified fragment length polymorphism; LSP, large sequence polymorphism; WGS, whole genome sequencing; NGS, next-generation sequencing; tNGS, targeted next-generation sequencing.

Simultaneously, advances in molecular diagnostics are revolutionizing the clinical management of NTM infections. A recent multicenter study evaluating 115 clinical samples found that metagenomic next-generation sequencing demonstrated 44.3 % sensitivity, targeted NGS 42.6 %, and multiplex PCR 36.5 %; when combined, these methods achieved a sensitivity of 54.8 %, with approximately 80 % concordance at the species level and 45 % at the subspecies level.⁹⁸

Further, a 2024 review emphasized the clinical value of rapid molecular platforms, including targeted NGS, metagenomic next-generation sequencing, and nanopore sequencing, in identifying NTM species and subspecies directly from respiratory specimens within days. These technologies not only expedite diagnosis but also allow for early initiation of targeted therapy—particularly crucial in managing inducible macrolide resistance linked to erm(41) and rrl mutations in M. abscessus complex.⁷

As such, integrating advanced molecular diagnostics into clinical workflows holds strong potential to improve treatment outcomes and reduce diagnostic delays.

10. Disease management

10.1. Infection control and preventive approaches

Diagnostic delays and the complexity of identifying the causative species add another layer of difficulty, as accurate subspecies differentiation is critical for tailoring effective regimens. Moreover, erroneous diagnoses may result in therapeutic interventions that are ineffective for NTM infections.⁸⁸ The MTBC and NTM are two distinct bacterial groups that share certain microbiological characteristics. The NTM have been observed to induce immune responses that are similar to those triggered by tuberculosis. Furthermore, both NTM and MTBC have been found to cause diseases that manifest similarly, particularly in the lymph nodes and lungs. However, a disease caused by NTM poses a diagnostic challenge for respiratory physicians, pediatricians, and other clinicians because it cannot readily be distinguished from tuberculosis based on clinical history, tuberculin skin test results, radiological patterns, and initial laboratory reports.³⁰ An additional and increasingly recognized challenge in NTM management is their capacity to form biofilms. Biofilm-associated infections are clinically significant due to the increased antimicrobial resistance exhibited by sessile bacteria compared to their planktonic forms. Biofilm formation on tissues and medical devices is a key pathogenic factor in NTM infections, with serious implications for both diagnosis and treatment. Consequently, strategies to prevent or disrupt biofilm formation are essential components of effective management.⁹⁹ Several genes and molecular pathways—including LpqY-Sug-ABC transporter (lpqY, sugA, subB, sugC), Pks1(Rv2946c), PpiB (Rv2582), GroEL1 (Rv3417c), MprB (Rv0982), (p)ppGpp, Poly(P), and cyclic-di-GMP—have been identified as central to mycobacterial biofilm development.¹⁰⁰

10.2. Management of antimicrobial-resistant NTM

NTM, particularly RGM such as the M. abscessus complex, pose significant therapeutic challenges due to their high level of drug resistance through mechanisms such as erm(41)-mediated inducible macrolide resistance, Eps, and β-lactamase (bla_Mab) production.^101,102 Effective management of drug-resistant RGM infections requires tailored therapeutic strategies informed by these resistance mechanisms and guided by antimicrobial susceptibility testing, despite its limitations due to poor correlation with clinical outcomes.¹⁰³ Genotypic testing for erm(41), rrl, and rrs genes is critical to identify resistance profiles early and prevent the development of resistance by avoiding ineffective regimens.⁸⁴ Novel antibiofilm strategies, such as matrix-degrading enzymes, bacteriophages, and nanoparticle-based therapies, can disrupt M. abscessus biofilms, enhancing antibiotic penetration and treatment efficacy in CF patients.¹⁰⁴

The 2020 ATS/ERS/ESCMID/IDSA guideline emphasizes that early initiation of combination therapy with at least three antibiotics, such as amikacin, cefoxitin, or tigecycline, and avoidance of monotherapy, particularly with macrolides, is critical to reducing the risk of inducible resistance.⁸⁴ Cautious use of emerging drugs, such as bedaquiline and clofazimine, which show promise for refractory M. abscessus infections in CF patients, must be carefully managed to prevent resistance to these novel agents.¹⁰⁵ Furthermore, strict adherence to prescribing guidelines, avoidance of incomplete treatment, and patient education to ensure treatment compliance significantly reduce the risk of resistance development.¹⁰⁶ Table 5 summarizes current therapeutic strategies, preferred drug regimens, and the scientific rationale behind them, based on the latest clinical guidelines and experimental evidence.^84,101,107

Table 5.

Resistance management and treatment regimens for RGM.

Strategy/Drug Class	Regimens & Rationale
Macrolide-Based Therapy	Azithromycin or clarithromycin + IV amikacin + imipenem/cefoxitin + oral agents (clofazimine, linezolid, tigecycline).
Amikacin (IV & Inhaled)	IV amikacin (15 mg/kg) for severe disease; use inhaled liposomal amikacin in refractory cases.
Imipenem/Cefoxitin	Core part of empiric therapy for M. abscessus/bolletii; cefoxitin favored due to lower resistance.
Tigecycline/Eravacycline/Omadacycline	IV tigecycline used despite gastrointestinal side effects; novel tetracyclines show promising in vitro efficacy.
Oxazolidinones	Linezolid added in refractory cases; some resistance (∼15 %–39 %) reported.
Resistance mechanisms	erm(41) for inducible macrolide resistance; blaMab β-lactamase; EPs decrease drug levels.
Genotypic testing–guided therapy	Testing for erm(41), rrl, and rrs is essential to avoid ineffective regimens.

High levels of antimicrobial resistance in NTM limit treatment options and complicate infection management. The complex structure of cell walls and characteristics such as biofilm formation are responsible for the intrinsic resistance in NTM. Generally, mycobacteria employ several mechanisms to achieve antimicrobial resistance, including: (1) Barrier Mechanisms: Limiting drug uptake through reduced cell wall permeability or reverse drug transport via the expression of efflux transporters. (2) Degradation or inactivation of enzymes: Utilization of enzymes that degrade or inactivate drugs. (3) Modification of pathways involved in drug activation or metabolism: Alterations in pathways responsible for drug activation or metabolism. (4) Drug target modification or amplification: Changes in the drug target or an increase in the number of drug targets.^108,109 Table 6 summarizes drug resistance in NTM, including types of resistance, mechanisms, and new drug developments.^110,111

Table 6.

Drug resistance in NTM.

Aspect	Details	Examples/Notes
Forms of resistance	Intrinsic vs. acquired	• Intrinsic: due to thick cell wall, efflux pumps (EPs), enzymes.
		• Acquired: caused by spontaneous mutations.
Cell wall / Permeability barrier	Hydrophobic mycolic-rich cell wall limiting drug uptake	Prominent in all NTM species; impedes β-lactams, glycopeptides.
Eps	Actively export drugs across cell envelope	Multiple NTM EPs (e.g. MAC, M. fortuitum, M. abscessus) reduce intracellular antibiotic concentrations.
Enzymatic drug inactivation	β-lactamases, aminoglycoside-modifying enzymes, ADP-ribosylases, erm methyltransferases	e.g. erm(41) methylates rrl gene—major macrolide resistance in M. abscessus.
Target gene mutations	Altered drug-binding sites leading to high-level resistance	23S rrl (macrolides), rpoB (rifampin), embCAB (ethambutol), gyrA/gyrB (fluoroquinolones), 16S rrs (aminoglycosides).
Regulator genes	Activation of global resistance networks	whiB7 increases drug export and modification gene expression.
Biofilms & dormancy	Reduce drug penetration & allow survival in low metabolic states	Enhances tolerance to multiple drug classes (intrinsic resistance).
New drug development	Novel antibiotics and drug repurposing targeting resistant NTM	• SPR719 (topo ATPase inhibitor)—phase II for MAC.
		• Delamanid, pretomanid—mixed in vitro results for M. abscessus.
		• Benzimidazoles active against MAC, M. kansasii.
Resistance inhibitors	Agents targeting EPs and enzymatic resistance mechanisms	Verapamil shown to reverse efflux-mediated macrolide, linezolid, bedaquiline resistance.
Drug–drug synergism & repurposing	Combining existing drugs or using adjunct compounds to overcome resistance	Natural products and drug repurposing show synergistic effects with antimicrobials.

Abbreviations: EP, efflux pump; NTM, nontuberculous mycobacteria; MAC, M. avium complex.

11. Treatment

Authoritative guidelines, including the ATS/IDSA statement (2007), the ATS/ERS/ESCMID/IDSA clinical practice guideline (2020), and the British Thoracic Society guideline (2017), provide comprehensive frameworks for managing pulmonary and extrapulmonary NTM diseases. These resources emphasize the importance of prolonged multidrug regimens, adjunctive therapies, and, in specific cases, surgical interventions, offering guidance for treating common species such as MAC and M. abscessus.^38,84,112

Treatment of NTM infections is challenging due to the diversity of NTM species, their antibiotic susceptibilities, and the complexity of clinical manifestations.⁶⁷ Patient-specific factors, such as immunosuppression, comorbidities, drug interactions, and adverse drug reactions, further complicate treatment. Typically, treatment management requires a multidisciplinary approach, coordinated by an infectious disease specialist or clinical microbiologist, with contributions from other relevant specialties such as surgery, when necessary, to improve clinical outcomes and achieve microbiological cure.¹¹³ The treatment of NTM infections is contingent upon the accurate identification of the infecting species. For example, treatment of pulmonary infections caused by NTM requires long-term administration of several drugs that are species-specific.^114,115 Various guidelines have been developed to manage NTM diseases, encompassing both pulmonary and extrapulmonary forms. The American Thoracic Society (ATS) and the Infectious Diseases Society of America (IDSA) offer detailed recommendations for different NTM infections. For patients with CF, the USA Cystic Fibrosis Foundation and the European Cystic Fibrosis Society have created consensus guidelines that emphasize screening, diagnosis, and treatment of NTM-PD.³⁸ Additionally, the British Thoracic Society, along with a joint guideline by ATS, European Respiratory Society (ERS), European Society of Clinical Microbiology and Infectious Diseases (ESCMID), and IDSA, provides specific directives on NTM-PD management.^84,112

Healthcare providers should be aware of the prevalence of NTM species in their areas to guide diagnostic and therapeutic decisions. It is essential to differentiate between actual infection and cases of laboratory contamination or colonization. The choice to begin treatment should be personalized, taking into account factors like symptom intensity, imaging results, and microbiological findings. The 2020 ATS/ERS/ESCMID/IDSA guideline advises starting treatment promptly, rather than just monitoring, for patients with NTM infections confirmed by positive sputum tests for AFB or those with cavitary lung disease.⁸⁴

Treatment plans usually consist of a combination of three to four antibiotics, given either daily or thrice weekly, based on disease severity and patient tolerance. For MAC pulmonary disease, a typical regimen includes rifampicin (or rifabutin for HIV-positive patients to prevent drug interactions), ethambutol, and a macrolide such as azithromycin or clarithromycin. Treatment generally continues for at least 12 months after sputum culture conversion. Due to the complexity and potential adverse effects of long-term antibiotic therapy, ensuring patient adherence can be challenging. Therefore, treatment strategies should be customized to individual patient requirements, and healthcare providers are advised to consult with specialists experienced in managing complex NTM infections.^2,84 Table 7 shows the differences between the 2007 ATS/IDSA and 2020 ATS/ERS/ESCMID/IDSA guidelines for NTM disease management.

Table 7.

Comparison of 2007 vs. 2020 guidelines for NTM-PD management.^38,84

Aspect	2007 ATS/IDSA Guideline	2020 ATS/ERS/ESCMID/IDSA Guideline
Geographical Scope	US-based, single-society guideline.	International (North America & Europe), multi-society collaboration.
Development Process	Based on expert opinion with minimal clinical trial data.	GRADE approach, systematic reviews, PICO questions, transparent rating of evidence, and strength of recommendations.
Number of Recommendations	General guidance, not quantified.	31 formal, graded recommendations across diagnosis, treatment, and monitoring.
Diagnostic Criteria	Clinical (symptoms + radiology) + Microbiology (≥ 2 sputum or 1 BAL culture) + exclusion of TB or other causes.	Same core criteria; clarified microbiologic definitions and emphasized standardized culture conversion definitions.
MAC Treatment Regimen	Macrolide + ethambutol ± rifampin for 12 months post-culture conversion.	Same core regimen; daily vs intermittent based on disease severity; added inhaled liposomal amikacin (ALIS) for refractory disease.
M. abscessus Management	Non-specific; emphasized macrolide-based regimens without clear guidance on susceptibility or species subspecies.	Regimens tailored by macrolide susceptibility (e.g., presence of erm gene); recommended ≥ 3 active drugs + possibly macrolide even if resistant (for immunomodulation).
Drug Additions/Changes	Limited (amikacin, clofazimine sometimes mentioned).	Added recommendations for clofazimine, linezolid, rifabutin; detailed on ALIS (FDA-approved), bedaquiline, and imipenem-cilastatin in specific settings.
Therapeutic Drug Monitoring (TDM)	Not specifically addressed in detail.	Detailed guidance on IV amikacin peak/trough goals, ototoxicity risks; supportive monitoring protocols for hepatotoxic drugs.
Evidence Certainty Levels	ATS/IDSA internal grading (Classes I–III, Levels A–C), inconsistently applied.	Full GRADE methodology: strong vs conditional recommendation + high/moderate/low/very low certainty of evidence for each.
Update Mechanism	No fixed update timeline; now outdated.	Plan to update every ≈ 4 years or as needed; living guideline model considered.
Adherence in Practice	Adherence poor in US studies (only ≈ 13 % of patients received guideline-based regimens).	Aims to improve clinical adherence via clarity and standardized terminology; highlighted gaps in real-world implementation and research needs.

Abbreviations: BAL, bronchoalveolar lavage; GRADE, grading of recommendations assessment, development, and evaluation; PICO, population, intervention, comparison, and outcome; FDA, Food and Drug Administration.

11.1. Treatment of NTM pulmonary disease

The treatment of diseases caused by NTM is challenging due to their slow growth, intrinsic antibiotic resistance, and species diversity. Treatment strategies rely heavily on observational studies, clinical experience, and guideline recommendations.^38,116 Treatment of NTM infections must be customized for each patient, taking into account the specific disease presentation, coexisting medical conditions, and the patient's age. The cavitary disease is linked to elevated mortality rates, necessitating urgent therapeutic intervention for those with fibrocavitary forms. On the other hand, nodular bronchiectatic disease typically manifests without major comorbidities and advances at a slow pace. As a result, initiating treatment early for mild, slowly progressing cases of nodular bronchiectatic disease may not be advisable due to potential adverse effects from extended use of multiple drugs.¹¹¹

11.1.1. M. avium complex treatment

Standard therapy includes a macrolide (azithromycin or clarithromycin), rifampin, and ethambutol, particularly effective in nodular‑bronchiectatic disease caused by macrolide‑susceptible strains, achieving culture conversion rates up to 82 %, though relapse occurs in approximately 48 %. In broader cohorts, success drops to ≈ 60 %.^117,118 For refractory or recurrent disease, inhaled liposomal amikacin added to standard therapy significantly improves culture conversion, leading to FDA approval for refractory MAC pulmonary disease.^119,120 In macrolide-resistant cases, antibiotic-only strategies often fail and are associated with high mortality; thus, combined medical and surgical approaches (e.g., lung lobe resection) are advised and have yielded favorable outcomes.^121,122 Other agents, including rifabutin, clofazimine, linezolid, and moxifloxacin, have been studied but achieved only around 18 % conversion rates.¹²³

11.1.2. M. abscessus complex treatment

Comprising subspecies abscessus, massiliense, and bolletii, subspecies identification is essential: abscessus and bolletii carry the inducible macrolide‑resistance gene erm(41), while massiliense does not, making it macrolide‑sensitive. Treatment typically involves a prolonged induction phase with intravenous amikacin and imipenem or cefoxitin alongside a macrolide. Culture conversion rates are approximately 35 % for abscessus and up to 79 % for massiliense. For resistant or relapsing cases, tigecycline and clofazimine have shown promise, with conversion rates between 24 % and 50 %.^117,124, 125, 126

11.1.3. New drugs and therapeutic approaches

Linezolid has demonstrated notable in vitro activity against various species of NTM, with several case reports and studies supporting its clinical efficacy in the management of NTM infections. Tedizolid (a newer oxazolidinone) exhibits enhanced in vitro potency compared to linezolid and has been reported to have a more favorable tolerability profile. Nevertheless, clinical experience with tedizolid remains extremely limited, with only a single published case documenting successful therapeutic use to date.^73,127 Amikacin liposomal inhalation suspension demonstrates enhanced microbiologic outcomes in both MAC and M. abscessus¹²⁰ Bedaquiline, has in vitro efficacy against MAC and M. abscessus, but initial clinical conversion rates are low.^128,129 Beta‑lactamase inhibitors (e.g., avibactam) restore activity of beta‑lactams like imipenem and ertapenem, and combinations like ceftazidime‑avibactam show promise against MAC.¹³⁰

11.1.4. Immunotherapy and adjunctive treatments

(1) Administration of IFN‑γ, particularly in immunocompromised hosts, has shown mixed results across trials.(2) GM‑CSF therapy has benefited CF patients with M. abscessus. (3) Inhaled nitric oxide has reduced bacterial load in patients with CF. (4) A single case demonstrated thalidomide's clinical and microbiological efficacy in disseminated MAC.^73,131,132

11.2. Treatment of extrapulmonary NTM diseases

The treatment of extrapulmonary NTM diseases, such as skin, lymph node, bone, joint, or disseminated infections, varies based on the mycobacterial species, infection site, and patient condition, often differing from pulmonary disease management. For skin and soft tissue infections caused by RGM (e.g., M. chelonae, M. fortuitum), treatment typically involves a multidrug regimen including macrolides, fluoroquinolones, or doxycycline, combined with surgical debridement for extensive lesions or abscesses. Treatment duration for mild cases ranges from 2 to 4 months, while severe cases may require 6 months or longer.¹³³ NTM lymphadenitis, often caused by MAC, is primarily treated with complete surgical excision of affected lymph nodes, achieving cure rates of up to 98.7 %. Antibiotics such as clarithromycin and ethambutol are used as adjuncts or alternatives when surgery is incomplete or not feasible, typically for 3–6 months. Bone and joint infections, often caused by M. marinum, M. kansasii, or RGM, require prolonged multidrug therapy (e.g., clarithromycin, rifampin, and ethambutol) for 6–12 months, often combined with surgical debridement or drainage to remove infected tissue. Disseminated infections, prevalent in immunocompromised patients such as those with advanced HIV (CD4⁺ < 50 cells/mm³), necessitate multidrug regimens similar to those for pulmonary disease, typically including clarithromycin (1000 mg/day), ethambutol (15 mg/kg/day), and sometimes rifabutin (150–350 mg/day) or amikacin for severe cases. These infections may require lifelong therapy or until immune reconstitution is achieved, particularly in HIV-positive patients. Unlike NTM-PD, where pharmacotherapy is the cornerstone, surgical intervention plays a more prominent role in extrapulmonary infections, particularly for lymphadenitis, skin, and bone infections. Treatment duration for localized infections (e.g., skin or lymph node) is generally shorter (3–6 months), whereas disseminated or bone/joint infections often require prolonged therapy akin to pulmonary regimens (6–12 months or longer). For specific species like M. kansasii, regimens similar to antituberculosis therapy (rifampicin, ethambutol, and isoniazid) for 18–24 months have been reported, while M. scrofulaceum lymphadenitis may also include erythromycin.^38,133

12. Conclusion

NTM are widely distributed in the environment, surrounding humans, and avoiding contact with them is practically impossible. NTM are emerging pathogens that can affect both immunocompromised and immunocompetent individuals. Diseases caused by NTM, particularly pulmonary infections, are increasing worldwide. The most common causative agents of pulmonary infections include: M. avium complex, M. kansasii, and M. abscessus complex. However, these agents may be mistaken for MTBC. Accurate and timely identification of the causative pathogen is essential for prompt treatment and minimizing side effects, as treatment options for NTM species differ from those for the MTBC. Molecular techniques, including line probe assays and gene sequencing, enable the identification of NTM. Sequencing of conserved genes is the reference method for identifying NTM species. For identification to the subspecies level, sequencing of multiple key targets such as 16S rRNA, hsp65, rpoB, and the internal transcribed spacer is required. Adherence to current guidelines for treating NTM diseases is essential.

CRediT authorship contribution statement

Mehdi Roshdi Maleki: Writing – review & editing, Writing – original draft, Supervision, Methodology, Investigation, Data curation, Conceptualization. Seyyed Reza Moaddab: Writing – review & editing, Validation, Supervision.

Informed consent

Not applicable.

Organ donation

Not applicable.

Ethics statement

Not applicable.

Data availability statement

This review did not involve any human subjects. All data included in the manuscript are from published sources or publicly available datasets.

Animal treatment

Not applicable.

Generative AI

We confirm that AI-assisted technologies were only used for language polishing, and all scientific content, data, and interpretations were created and verified by the authors.

Funding

No external funding was received for this work, and any financial interests or sponsorships related to the content of this manuscript are disclosed in the appropriate section.

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.