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Background: Nontuberculous mycobacterial disease has emerged worldwide over the past 20 years. However, there are currently few reports on the established technique for constructing knockout mutants of nontuberculous mycobacteria. Therefore, gene recombination techniques for nontuberculous mycobacteria require further research. Results: We constructed vector pPR23LHR that harbors the ribosomal protein S12 gene (rpsL+) as a dominant negative selection marker and the hygromycin (Hyg) and lacZ cassettes as positive selection markers. We constructed knockout mutants of proteasomal genes, which we found to be required for hypoxic pellicle formation in Mycobacterium intracellulare by functional genomic analysis. The knockout mutants showed impaired hypoxic pellicle formation, consistent with previous data using epoxomicin, a proteasomal inhibitor. Conclusions: Our findings demonstrate that rpsL+ is an efficient dominant negative selection marker for gene recombination in nontuberculous mycobacteria. Our temperature‐sensitive rpsL+ method for the construction of knockout mutants will facilitate functional assays to validate the virulence factors of nontuberculous mycobacteria and the pathogenesis of nontuberculous mycobacterial disease. [ABSTRACT FROM AUTHOR]

Clinically, the incidence of nontuberculous mycobacteria (NTM) lung disease is on the rise, and Mycobacterium intracellulare (M. intracellulare) has attracted much attention as a common opportunistic pathogen in clinical practice. So it is very important to study its immunopathogenic mechanism. In this study, the mechanism of M. intracellulare induced pyroptosis of macrophage was investigated. As shown in Fig. 1, the secretion of IL-1β and IL-18 in J774A.1 cells increased with time after M. intracellulare infection and was affected by caspase-1 activation and K + efflux, while caspase-1 was significantly expressed in infected cells. Further from Fig. 2, NLRP3,AIM2,ASC proteins were significantly expressed in J774A.1 cells after infection, indicating that the NOD-like receptor thermal protein domain associated protein 3 (NLRP3) and absent in melanoma 2 (AIM2) inflammasome were involved in the infection process. In addition, when caspase-1 activity and K + efflux were inhibited, the expression of related proteins was significantly reduced. It indicates that the activation of NLRP3 and AIM2 is regulated by caspase-1 and K+. Figure 3, the percentage of dead cells with cell membrane damage increases after infection and cleavage of GSDMD proteins occurs. In summary, infection of J774A.1 cells with M. intracellulare induces pyroptosis, and this process is mediated by caspase-1. Our study provides information for further understanding of the molecular mechanism of M. intracellulare infection. [ABSTRACT FROM AUTHOR]

Non-tuberculous mycobacteria (NTM) cause skin infections, respiratory diseases, and disseminated infections. Mycobacterium avium and Mycobacterium intracellulare, which are slow grown Mycobacterium, are main agents of those NTM diseases. A silkworm infection model with Mycobacterium abscessus, a rapidly growing Mycobacterium species, was established to quantitatively evaluate its virulence within a short period. However, a silkworm infection model to quantitatively evaluate the virulence of M. intracellulare has not yet been developed. In this study, we determined the virulence of M. intracellulare subspecies within 4 days using a silkworm infection model. The subspecies of M. intracellulare strains used in this study were estimated by phylogenetic tree analysis using core gene data. The median lethal dose (LD50) values, which are the dose of a pathogen required to kill half of the silkworms in a group, were determined 4 days after infection. The LD50 value of M. intracellulare subsp. chimaera DSM44623 was higher than that of M. intracellulare subsp. intracellulare ATCC13950. These results suggest that the virulence of M. intracellulare subspecies can be compared using a silkworm model within 4 days. [ABSTRACT FROM AUTHOR]

Whole genome sequencing (WGS) of Mycobacterium avium complex (MAC) isolates in the clinical laboratory setting allows for rapid and reliable subspecies identification of a closely related complex of human pathogens. We developed a bioinformatics pipeline for accurate subspecies identification and tested 74 clinical MAC isolates from various anatomical sites. We demonstrate that reliable subspecies level identification of these common and clinically significant MAC isolates, including M. avium subsp. hominissuis (most dominant in causing lower respiratory tract infections in our cohort), M. avium subsp. avium, M. intracellulare subsp. intracellulare, and M. intracellulare subsp. chimaera, can be achieved by analysis of only two marker genes (rpoB and groEL/hsp65). We then explored the relationship between these subspecies and anatomical site of infection. Further, we conducted an in silico analysis and showed our algorithm also performed well for M. avium subsp. paratuberculosis but failed to consistently identify M. avium subsp. silvaticum and M. intracellulare subsp. yongonense, likely due to a lack of available reference genome sequences; all the 3 subspecies were not found in our clinical isolates and rarely reported to cause human infections. Accurate MAC subspecies identification may provide the tool and opportunity for better understanding of the disease-subspecies dynamics in MAC infections.

Abstract Background The Mycobacterium avium complex (MAC) comprises the most frequent non-tuberculous mycobacteria (NTM) in Central Europe and currently includes twelve species. M. avium (MAV), M. intracellulare subsp. intracellulare (MINT), and M. intracellulare subsp. chimaera (MCH) are clinically most relevant. However, the population structure and genomic landscape of MAC linked with potential pathobiological differences remain little investigated. Methods Whole genome sequencing (WGS) was performed on a multi-national set of MAC isolates from Germany, France, and Switzerland. Phylogenetic analysis was conducted, as well as plasmids, resistance, and virulence genes predicted from WGS data. Data was set into a global context with publicly available sequences. Finally, detailed clinical characteristics were associated with genomic data in a subset of the cohort. Results Overall, 610 isolates from 465 patients were included. The majority could be assigned to MAV (n = 386), MCH (n = 111), and MINT (n = 77). We demonstrate clustering with less than 12 SNPs distance of isolates obtained from different patients in all major MAC species and the identification of trans-European or even trans-continental clusters when set into relation with 1307 public sequences. However, none of our MCH isolates clustered closely with the heater-cooler unit outbreak strain Zuerich-1. Known plasmids were detected in MAV (325/1076, 30.2%), MINT (62/327, 19.0%), and almost all MCH-isolates (457/463, 98.7%). Predicted resistance to aminoglycosides or macrolides was rare. Overall, there was no direct link between phylogenomic grouping and clinical manifestations, but MCH and MINT were rarely found in patients with extra-pulmonary disease (OR 0.12 95% CI 0.04–0.28, p

Background: The Mycobacterium avium complex (MAC) comprises the most frequent non-tuberculous mycobacteria (NTM) in Central Europe and currently includes twelve species. M. avium (MAV), M. intracellulare subsp. intracellulare (MINT), and M. intracellulare subsp. chimaera (MCH) are clinically most relevant. However, the population structure and genomic landscape of MAC linked with potential pathobiological differences remain little investigated. Methods: Whole genome sequencing (WGS) was performed on a multi-national set of MAC isolates from Germany, France, and Switzerland. Phylogenetic analysis was conducted, as well as plasmids, resistance, and virulence genes predicted from WGS data. Data was set into a global context with publicly available sequences. Finally, detailed clinical characteristics were associated with genomic data in a subset of the cohort. Results: Overall, 610 isolates from 465 patients were included. The majority could be assigned to MAV (n = 386), MCH (n = 111), and MINT (n = 77). We demonstrate clustering with less than 12 SNPs distance of isolates obtained from different patients in all major MAC species and the identification of trans-European or even trans-continental clusters when set into relation with 1307 public sequences. However, none of our MCH isolates clustered closely with the heater-cooler unit outbreak strain Zuerich-1. Known plasmids were detected in MAV (325/1076, 30.2%), MINT (62/327, 19.0%), and almost all MCH-isolates (457/463, 98.7%). Predicted resistance to aminoglycosides or macrolides was rare. Overall, there was no direct link between phylogenomic grouping and clinical manifestations, but MCH and MINT were rarely found in patients with extra-pulmonary disease (OR 0.12 95% CI 0.04–0.28, p < 0.001 and OR 0.11 95% CI 0.02–0.4, p = 0.004, respectively) and MCH was negatively associated with fulfillment of the ATS criteria when isolated from respiratory samples (OR 0.28 95% CI 0.09-0.7, p = 0.011). With 14 out of 43 patients with available serial isolates, co-infections or co-colonizations with different strains or even species of the MAC were frequent (32.6%). Conclusions: This study demonstrates clustering and the presence of plasmids in a large proportion of MAC isolates in Europe and in a global context. Future studies need to urgently define potential ways of transmission of MAC isolates and the potential involvement of plasmids in virulence. [ABSTRACT FROM AUTHOR]

Abstract Background Mycobacterium avium complex (MAC), including Mycobacterium intracellulare is a member of slow-growing mycobacteria and contributes to a substantial proportion of nontuberculous mycobacterial lung disease in humans affecting immunocompromised and elderly populations. Adaptation of pathogens in hostile environments is crucial in establishing infection and persistence within the host. However, the sophisticated cellular and molecular mechanisms of stress response in M. intracellulare still need to be fully explored. We aimed to elucidate the transcriptional response of M. intracellulare under acidic and oxidative stress conditions. Results At the transcriptome level, 80 genes were shown [FC] ≥ 2.0 and p

Background: Mycobacterium avium complex (MAC), including Mycobacterium intracellulare is a member of slow-growing mycobacteria and contributes to a substantial proportion of nontuberculous mycobacterial lung disease in humans affecting immunocompromised and elderly populations. Adaptation of pathogens in hostile environments is crucial in establishing infection and persistence within the host. However, the sophisticated cellular and molecular mechanisms of stress response in M. intracellulare still need to be fully explored. We aimed to elucidate the transcriptional response of M. intracellulare under acidic and oxidative stress conditions. Results: At the transcriptome level, 80 genes were shown [FC] ≥ 2.0 and p < 0.05 under oxidative stress with 10 mM hydrogen peroxide. Specifically, 77 genes were upregulated, while 3 genes were downregulated. In functional analysis, oxidative stress conditions activate DNA replication, nucleotide excision repair, mismatch repair, homologous recombination, and tuberculosis pathways. Additionally, our results demonstrate that DNA replication and repair system genes, such as dnaB, dinG, urvB, uvrD2, and recA, are indispensable for resistance to oxidative stress. On the contrary, 878 genes were shown [FC] ≥ 2.0 and p < 0.05 under acidic stress with pH 4.5. Among these genes, 339 were upregulated, while 539 were downregulated. Functional analysis highlighted nitrogen and sulfur metabolism pathways as the primary responses to acidic stress. Our findings provide evidence of the critical role played by nitrogen and sulfur metabolism genes in the response to acidic stress, including narGHIJ, nirBD, narU, narK3, cysND, cysC, cysH, ferredoxin 1 and 2, and formate dehydrogenase. Conclusion: Our results suggest the activation of several pathways potentially critical for the survival of M. intracellulare under a hostile microenvironment within the host. This study indicates the importance of stress responses in M. intracellulare infection and identifies promising therapeutic targets. [ABSTRACT FROM AUTHOR]

Background: The Mycobacterium avium complex includes the commonest non-tuberculous mycobacteria associated with human infections. These infections have been associated with the production of biofilms in many cases, but there are only a few studies about biofilms produced by the species included in this group. Methods: Three collection strains (M. avium ATCC25291, M. intracellulare ATCC13950, and M. chimaera DSM756), three clinically significant strains (647, 657, and 655), and three clinically non-significant ones (717, 505, and 575) of each species were included. The clinical significance of the clinical isolates was established according to the internationally accepted criteria. The biofilm ultrastructure was studied by Confocal-Laser Scanning Microscopy by using BacLight Live–Dead and Nile Red stains. The viability, covered surface, height, and relative autofluorescence were measured in several images/strain. The effect of clarithromycin was studied by using the technique described by Muñoz-Egea et al. with modifications regarding incubation time. The study included clarithromycin in the culture medium at a concentration achievable in the lungs (11.3 mg/L), using one row of wells as the control without antibiotics. The bacterial viability inside the biofilm is expressed as a percentage of viable cells. The differences between the different parameters of the biofilm ultrastructure were analyzed by using the Kruskal–Wallis test. The correlation between bacterial viability in the biofilm and treatment time was evaluated by using Spearman's rank correlation coefficient (ρ). Results: The strains showed differences between them with all the studied parameters, but neither a species-specific pattern nor a clinical-significance-specific pattern were detected. For the effect of clarithromycin, the viability of the bacteria contained in the biofilm was inversely proportional to the exposure time of the biofilm (ρ > −0.3; p-value < 0.05), excluding two M. chimaera strains (M. chimaera DSM756 and 575), which showed a weak positive correlation with treatment time (0.2 < ρ < 0.39; p-value < 0.05). Curiously, despite a clarithromycin treatment of 216 h, the percentage of the biofilm viability of the strains evaluated here was not less than 40% at best (M. avium 717). Conclusions: All the M. avium complex strains studied can form biofilm in vitro, but the ultrastructural characteristics between them suggest that these are strain-specific characteristics unrelated to the species or the clinical significance. The clarithromycin effect on MAC species is biofilm-age/time-of-treatment-dependent and appears to be strain-specific while being independent of the clinical significance of the strain. [ABSTRACT FROM AUTHOR]

Objectives: This study aims to assess the characteristics and treatment outcomes of patients diagnosed with non-tuberculous mycobacteria (NTM) diseases at the Infectious Diseases Unit of ARNAS Garibaldi Hospital in Catania, Italy, focusing on demographics, clinical features, and treatment effectiveness. Methods: We conducted a retrospective observational study of 10 patients diagnosed with NTM diseases between 2019 and 2021. Data was collected from electronic medical records, including demographic information, comorbidities, treatment modalities, and outcomes. The study utilized descriptive statistics to analyze continuous and categorical variables. Treatment regimens were based on individual patient needs, incorporating a combination of antibiotics. Results: The median age of the patients was 55.44 years, all female, predominantly suffering from pulmonary NTM diseases. Mycobacterium intracellulare was the most common pathogen. Common comorbidities included COPD, bronchiectasis, GERD, and hypovitaminosis D. Patients showed symptoms like fever, cough, and asthenia. The treatment regimens were diverse, with macrolides, rifampicin, and ethambutol forming the core. Adverse effects were noted in 40 % of patients, including gastrointestinal and neurological disorders. All patients achieved microbiological cure, with 60 % showing clinical improvement and 36 % radiological improvement. Conclusion: The study highlights the complexity of diagnosing and treating NTM diseases, emphasizing the need for personalized treatment plans and vigilant monitoring of adverse effects. Despite achieving microbiological cure, challenges remain in achieving complete clinical and radiological resolution. Further research is needed to enhance the understanding and management of NTM diseases, particularly in diverse populations.

Adult‐onset immunodeficiency (AOID) syndrome due to the presence of anti‐interferon gamma antibody (AIGA) is characterized by multiple opportunistic infections. We report a case of a 65‐year old healthy woman who suffered from Legionella pneumophila and Mycobacterium intracellulare co‐infection with clinical presentation mimicking metastatic lung cancer. She presented with chronic cough and weight loss. Her positron emission tomography scan showed a right upper lobe mass, mediastinal lymphadenopathy and multiple bone lesions. Acid fast bacilli culture of the lung mass and mediastinal lymph node revealed Mycobacterium intracellulare and she improved with prolonged antibiotic. Relapse of disseminated Mycobacterium intracellulare infection occurred 15 months post‐treatment and AIGA was positive with functional neutralizing activity on downstream immune pathway. AOID syndrome secondary to AIGA was diagnosed. This case illustrated the importance of high index of suspicion of AOID syndrome and the difficulty of early diagnosis. Further studies on its predictive factors and AIGA‐targeted treatment modalities are urgently needed. [ABSTRACT FROM AUTHOR]

Abstract Background Mycobacterium intracellulare is a major etiological agent of Mycobacterium avium-intracellulare pulmonary disease (MAC-PD). However, the characteristics of the virulence of M. intracellulare and the in vivo chemotherapeutic efficacy remain unclear. In this study, we examined the virulence of nine M. intracellulare strains with different clinical phenotypes and genotypes in C57BL/6 mice. Results We classified three types of virulence phenotypes (high, intermediate, and low) based on the kinetics of the bacterial load, histological lung inflammation, and neutrophilic infiltration. High virulence strains showed more severe neutrophilic infiltration in the lungs than intermediate and low virulence strains, with 6.27-fold and 11.0-fold differences of the average percentage of neutrophils in bronchoalveolar lavage fluid, respectively. In particular, the high virulence strain M.i.198 showed the highest mortality in mice, which corresponded to the rapid progression of clinical disease. In mice infected with the drug-sensitive high virulence strain M019, clarithromycin-containing chemotherapy showed the highest efficacy. Monotherapy with rifampicin exacerbated lung inflammation with increased lymphocytic and neutrophilic infiltration into the lungs. Conclusions The virulence phenotypes of clinical strains of M. intracellulare were diverse, with high virulence strains being associated with neutrophilic infiltration and disease progression in infected mice. These high virulence strains were proposed as a useful subject for in vivo chemotherapeutic experiments.

ABSTRACT Patients with Mycobacterium intracellulare pulmonary disease are more likely to experience poor treatment outcomes if they have been observed with microbiological persistence after 6 months of treatment. This study aims to identify the risk factors for microbiological persistence and describe the changes in the minimum inhibitory concentration (MIC) during antimycobacterial treatment. This retrospective case-control study enrolled patients diagnosed with M. intracellulare pulmonary disease between April 2017 and September 2021 at Shanghai Pulmonary Hospital. Patients with positive cultures after 6 months of treatment (positive group) were matched by age and sex in a 1:1 ratio to patients with negative conversion (negative group). Totally, 46 pairs of patients were analyzed. Risk factors for microbiological persistence at month 6 were smoking, previous tuberculosis treatment, chronic lung diseases, a positive baseline acid-fast bacilli smear, and adverse drug reactions; the risk was reduced by a regimen containing ethambutol, ≥3 effective drugs, and a higher pre-treatment absolute lymphocyte count. Regarding the drug-resistance profile, the negative group had a higher proportion of susceptibility to clarithromycin (100.0% vs 84.8%, P = 0.012). Most isolates were susceptible or intermediate to amikacin in both groups (93.5% and 84.8%, respectively). Nine patients (16.4%, 9/55) had a change in the drug-resistance profile, including four who changed from clarithromycin susceptible to clarithromycin resistant, and the other three reversed. Two pairs of isolates had a change in resistance to amikacin. In conclusion, risk factors for microbiological persistence were identified, and the change in MIC values during antimycobacterial treatment indicated the need for monitoring to enable timely adjustment of the regimen. IMPORTANCE Nontuberculous mycobacteria pulmonary disease (NTM-PD) has been recognized as an important public health issue because of its increasing incidence globally, low cure rate, and high recurrence rate. NTM-PD has innate resistance to many first-line anti-tuberculous drugs, which limits the treatment options. Mycobacterium intracellulare is reportedly the most important pathogenic NTM and accounts for the highest proportion of NTM-PD in China. A previous study suggested that poor microbiological response after 6 months of treatment is predictive of treatment failure. The present study investigated the risk factors associated with persistent positive sputum cultures by treatment month 6 in patients with M. intracellulare pulmonary disease and the variation in minimum inhibitory concentration patterns in clinical settings. This information might help to identify patients at higher risk of treatment failure and enable the timely provision of necessary interventions.

Non‐tuberculous mycobacterial infections are gradually increasing worldwide, with slow‐growing mycobacteria such as Mycobacterium avium, Mycobacterium intracellulare and Mycobacterium kansasii accounting for the majority of cases. The use of tetracyclines has received renewed attention in recent years, and this study was designed to investigate the antibacterial activity of omadacycline, eravacycline, tigecycline, sarecycline, minocycline and doxycycline against M. avium, M. intracellulare and M. kansasii. Susceptibility testing of six tetracyclines was conducted against M. avium, M. intracellulare and M. kansasii isolates, and all the clinical isolates were collected from January 2012 to December 2018. All six agents exhibited poor antibacterial activity against slowly growing mycobacteria (SGM) isolates of three subspecies with MIC50 and MIC90 ≥8 μg/mL. M. intracellulare and M. kansasii had lower resistance rates to omadacycline than the other five drugs. The severe resistance of SGM to tetracycline suggests that developing tetracycline‐class antibiotics needs to overcome existing resistance mechanisms. [ABSTRACT FROM AUTHOR]

Case summary: A 3-year-old castrated male domestic shorthaired cat, with indoor–outdoor access, was presented for chronic, progressive multinodular to generalised subcutaneous nodules covering much of its body. Previous medical treatment with doxycycline had been unhelpful. Fine-needle aspiration of the nodules revealed intra- and extracellular multibacillary negative staining rods in pyogranulomatous inflammation. Bacterial culture and susceptibility studies isolated Mycobacterium intracellulare, with zimine as the drug of choice for treatment. Initial triple therapy with rifampicin, azithromycin and pradofloxacin was ineffective, and was changed to triple therapy with clofazimine, clarithromycin and doxycycline once drug susceptibility was known, which was given for 3 months, after which long-term therapy with clofazimine and clarithromycin was continued. Relevance and novel information: Slow growing M intracellulare, a member of the Mycobacterium avium complex (MAC), has never been reported to cause disease in cats from Singapore and, by extension, Southeast Asia. The infection in this patient resulted in subcutaneous nodules, which started on the face, then spread to the feet and much of the rest of its body. This is in contrast to that commonly reported for infection with M avium, which is also a member of MAC, and may not only present with similar signs in cats, but also progress to systemic spread. Susceptibility studies suggest clofazimine as the drug of choice when treating this infection, and this case supports its use as empirical therapy for veterinarians treating this disease in this region while awaiting culture and sensitivity results. [ABSTRACT FROM AUTHOR]

Objective To clarify the relationship between different species of Mycobacterium avium complex (MAC), drug sensitivity and clinical prognosis in Guangzhou. Methods The clinical isolates of MAC from 2018 to 2020 were collected from the Mycobacterium Strain Bank in Guangzhou, and the MAC strains of MAC lung disease patients were sequenced to identify them by 16S rRNA, ITS, rpoB, hsp65 multi-target gene sequencing. MIC50 and MIC90 of its minimum inhibitory concentration (MIC) were determined by microplate method to determine its drug sensitivity. The success of clinical treatment can be judged according to the clinical symptoms, DR/ CT radiology and pathogen culture results. Results (1) There were 67 strains of Mycobacterium intracellulare (73.63%) and 21 strains of Mycobacterium avium (23.08%) in 91 clinical isolates of MAC. (2) The resistance rates of Mycobacterium avium and Mycobacterium intracellular to amikacin and ethambutol were different (P < 0.05). (3) There was significant difference in the treatment success rate between M.aviumpulmonary disease and M.intracellularepulmonary disease (66.67% vs. 41.27%, χ² = 6.053, P < 0.05). Conclusion M.intracellulare and M.avium were the main clinical isolates of MAC in Guangzhou, and there were differences in drug resistance and clinical prognosis among different MAC species. For patients with NTM pulmonary disease, strain identification and in vitro drug susceptibility tests should be conducted, and clinical treatment protocols should be formulated and optimized based on the above results. [ABSTRACT FROM AUTHOR]

Rice body formation is a rare response to chronic inflammation of the synovial membrane. It is most commonly associated with rheumatoid arthritis and tuberculosis. Recently, there have been reports of rice bodies caused by non-tuberculous mycobacterial infection. We describe a case of rice body formation in a 69-year-old man who presented with pain and swelling in his third finger for six months after being punctured by a wire 1 year ago. He had no other notable recent medical history. Magnetic resonance imaging showed a large amount of fluid collection with diffuse thickening and enhancement of the synovium and rice bodies along the flexor tendon of the third finger. During surgery, multiple granular white rice bodies were found from the third carpal bone to the distal phalanx. Mycobacterium intracellulare was identified through mycobacterial culture and the patient was treated with rifampin, ethambutol, and clarithromycin, without recurrence. This case reveals that Mycobacterium intracellulare infection can cause tenosynovitis with rice bodies. [ABSTRACT FROM AUTHOR]

A 65-year-old Japanese woman repeatedly withdrew and resumed antibiotics against pulmonary non-tuberculous mycobacterial infection caused by Mycobacterium intracellulare for more than 10 years. Although she continued to take medications, her respiratory symptoms and chest computed tomography indicated an enlarged infiltrative shadow in the lingular segment of the left lung that gradually worsened over the course of a year or more. Bronchoscopy was performed and mycobacterial culture of the bronchial lavage fluid was negative, whereas Exophiala dermatitidis was detected. After administration of oral voriconazole was initiated, the productive cough and infiltrative shadow resolved. There are no characteristic physical or imaging findings of E. dermatitidis , and it often mimics other chronic respiratory infections. Thus, when confronting refractory non-tuberculous mycobacterial cases, it might be better to assume other pathogenic microorganisms, including E. dermatitidis, and actively perform bronchoscopy. [ABSTRACT FROM AUTHOR]

Background: The Mycobacterium avium complex includes the commonest non-tuberculous mycobacteria associated with human infections. These infections have been associated with the production of biofilms in many cases, but there are only a few studies about biofilms produced by the species included in this group. Methods: Three collection strains (M. avium ATCC25291, M. intracellulare ATCC13950, and M. chimaera DSM756), three clinically significant strains (647, 657, and 655), and three clinically non-significant ones (717, 505, and 575) of each species were included. The clinical significance of the clinical isolates was established according to the internationally accepted criteria. The biofilm ultrastructure was studied by Confocal-Laser Scanning Microscopy by using BacLight Live–Dead and Nile Red stains. The viability, covered surface, height, and relative autofluorescence were measured in several images/strain. The effect of clarithromycin was studied by using the technique described by Muñoz-Egea et al. with modifications regarding incubation time. The study included clarithromycin in the culture medium at a concentration achievable in the lungs (11.3 mg/L), using one row of wells as the control without antibiotics. The bacterial viability inside the biofilm is expressed as a percentage of viable cells. The differences between the different parameters of the biofilm ultrastructure were analyzed by using the Kruskal–Wallis test. The correlation between bacterial viability in the biofilm and treatment time was evaluated by using Spearman’s rank correlation coefficient (ρ). Results: The strains showed differences between them with all the studied parameters, but neither a species-specific pattern nor a clinical-significance-specific pattern were detected. For the effect of clarithromycin, the viability of the bacteria contained in the biofilm was inversely proportional to the exposure time of the biofilm (ρ > −0.3; p-value < 0.05), excluding two M. chimaera strains (M. chimaera DSM756 and 575), which showed a weak positive correlation with treatment time (0.2 < ρ < 0.39; p-value < 0.05). Curiously, despite a clarithromycin treatment of 216 h, the percentage of the biofilm viability of the strains evaluated here was not less than 40% at best (M. avium 717). Conclusions: All the M. avium complex strains studied can form biofilm in vitro, but the ultrastructural characteristics between them suggest that these are strain-specific characteristics unrelated to the species or the clinical significance. The clarithromycin effect on MAC species is biofilm-age/time-of-treatment-dependent and appears to be strain-specific while being independent of the clinical significance of the strain.

A 48-year-old male presented with complaints of occasional dry cough and sneezing for the past 4 months and high-resolution computed tomography chest done showed tree in bud and patchy nodular opacities in the right upper lobe and right middle lobe. He underwent bronchoscopy and bronchoalveolar lavage Gene X-pert Mycobacterium Tuberculosis (MTB) was not detected, but cytology showed granulomas and hence he was started on empirical antituberculous therapy (Isoniazid, Rifampicin, Pyrazinamide and Ethambutol (HRZE) regimen). His acid-fast bacilli C/S reports during follow-up showed growth of Mycobacterium Avium complex (MAC). Drug sensitivity testing was done and then he was started on oral rifampicin, ethambutol, and azithromycin and this regimen was continued for the next 6 months.

Background: Mycobacterium intracellulare is a major etiological agent of Mycobacterium avium-intracellulare pulmonary disease (MAC-PD). However, the characteristics of the virulence of M. intracellulare and the in vivo chemotherapeutic efficacy remain unclear. In this study, we examined the virulence of nine M. intracellulare strains with different clinical phenotypes and genotypes in C57BL/6 mice. Results: We classified three types of virulence phenotypes (high, intermediate, and low) based on the kinetics of the bacterial load, histological lung inflammation, and neutrophilic infiltration. High virulence strains showed more severe neutrophilic infiltration in the lungs than intermediate and low virulence strains, with 6.27-fold and 11.0-fold differences of the average percentage of neutrophils in bronchoalveolar lavage fluid, respectively. In particular, the high virulence strain M.i.198 showed the highest mortality in mice, which corresponded to the rapid progression of clinical disease. In mice infected with the drug-sensitive high virulence strain M019, clarithromycin-containing chemotherapy showed the highest efficacy. Monotherapy with rifampicin exacerbated lung inflammation with increased lymphocytic and neutrophilic infiltration into the lungs. Conclusions: The virulence phenotypes of clinical strains of M. intracellulare were diverse, with high virulence strains being associated with neutrophilic infiltration and disease progression in infected mice. These high virulence strains were proposed as a useful subject for in vivo chemotherapeutic experiments. [ABSTRACT FROM AUTHOR]

Abstract Background Recently, Mycobacterium avium complex (MAC) infections have been increasing, especially in immunocompromised and older adults. The rapid increase has triggered a global health concern due to limited therapeutic strategies and adverse effects caused by long-term medication. To provide more evidence for the treatment of MAC, we studied the in vitro inhibitory activities of 17 antimicrobial agents against clinical MAC isolates. Results A total of 111 clinical MAC isolates were enrolled in the study and they were identified as M. intracellulare, M. avium, M. marseillense, M. colombiense, M. yongonense, and two isolates could not be identified at the species level. MAC strains had relatively low (0–21.6%) resistance to clarithromycin, amikacin, bedaquiline, rifabutin, streptomycin, and clofazimine, and the resistant rates to isoniazid, rifampin, linezolid, doxycycline, and ethionamide were very high (72.1–100%). In addition, M. avium had a significantly higher resistance rate than that of M. intracellulare for ethambutol (92.3% vs 40.7%, P

Abstract Background M. intracellulare is a frequent causative pathogen of nontuberculous mycobacteria infection that causes infections in the respiratory tract, whose incidence is increasing in many countries. This study aimed at determining the VNTR-based genetic diversity of a collection of 39 M. intracellulare human strains isolated from respiratory specimens over the last 5 years. Results The VNTR analysis showed that M. intracellulare strains displayed a high genetic diversity, indicating that the M. intracellulare genotypes are quite heterogeneous in our geographical area. Moreover, a comparison with VNTR profiles of strains from other countries confirmed that genotypes of clinical strains of M. intracellulare are not related to geographical origin. Conclusions VNTR typing has proved to be a highly discriminatory method for better understanding the molecular epidemiology of M. intracellulare.

Mycobacterium intracellulare is the most common cause of nontuberculous mycobacterial lung disease, with a rapidly growing prevalence worldwide. In this study, we performed comparative genomic analysis and antimicrobial susceptibility characteristics analysis of 117 clinical M. intracellulare strains in China. Phylogenetic analysis showed that clinical M. intracellulare strains had high genetic diversity and were not related to the geographical area. Notably, most strains (76.07%, 89/117) belonged to Mycobacterium paraintracellulare (MP) and Mycobacterium indicus pranii (MIP) in the genome, and we named them MP-MIP strains. These MP-MIP strains may be regarded as a causative agent of chronic lung disease. Furthermore, our data demonstrated that clarithromycin, amikacin, and rifabutin showed strong antimicrobial activity against both M. intracellulare and MP-MIP strains in vitro. Our findings also showed that there was no clear correlation between the rrs, rrl, and DNA gyrase genes (gyrA and gyrB) and the aminoglycosides, macrolides, and moxifloxacin resistance, respectively. In conclusion, this study highlights the high diversity of M. intracellulare in the clinical setting and suggests paying great attention to the lung disease caused by MP-MIP.

This study aimed to clarify the population-based distributions of Mycobacterium avium and Mycobacterium intracellulare in Japan. We conducted a combined analysis of the national insurance claim and microbiological databases. The incidence rates of M. avium by province were similar throughout the country, with some exceptions, such as in Okinawa, probably because the bathing customs are different from those in mainland Japan. In contrast, M. intracellulare showed a gradual increase from the central part of the country to the southwestern region, with precise gradation, which may indicate infection sources in the natural environment. This study found that assessment of the infection route of M. intracellulare in the natural environment, which is similar to the distribution of M. intracellular patients, is warranted. In conclusion, improvement of the household environment could decrease the incidence of M. avium, while environmental countermeasures will be required to decrease the incidence of M. intracellulare.

Background: Recently, Mycobacterium avium complex (MAC) infections have been increasing, especially in immunocompromised and older adults. The rapid increase has triggered a global health concern due to limited therapeutic strategies and adverse effects caused by long-term medication. To provide more evidence for the treatment of MAC, we studied the in vitro inhibitory activities of 17 antimicrobial agents against clinical MAC isolates. Results: A total of 111 clinical MAC isolates were enrolled in the study and they were identified as M. intracellulare, M. avium, M. marseillense, M. colombiense, M. yongonense, and two isolates could not be identified at the species level. MAC strains had relatively low (0–21.6%) resistance to clarithromycin, amikacin, bedaquiline, rifabutin, streptomycin, and clofazimine, and the resistant rates to isoniazid, rifampin, linezolid, doxycycline, and ethionamide were very high (72.1–100%). In addition, M. avium had a significantly higher resistance rate than that of M. intracellulare for ethambutol (92.3% vs 40.7%, P < 0.001), amikacin (15.4% vs 1.2%, P = 0.049), and cycloserine (69.2% vs 25.9%, P = 0.004). Conclusions: Our results supported the current usage of macrolides, rifabutin, and aminoglycosides in the regimens for MAC infection, and also demonstrated the low resistance rate against new drugs, such as clofazimine, tedizolid, and bedaquiline, suggesting the possible implementation of these drugs in MAC treatment. [ABSTRACT FROM AUTHOR]

Abstract Background Mycobacterium intracellulare is a representative etiological agent of emerging pulmonary M. avium-intracellulare complex disease in the industrialized countries worldwide. The recent genome sequencing of clinical strains isolated from pulmonary M. avium-intracellulare complex disease has provided insight into the genomic characteristics of pathogenic mycobacteria, especially for M. avium; however, the genomic characteristics of M. intracellulare remain to be elucidated. Results In this study, we performed comparative genomic analysis of 55 M. intracellulare and related strains such as M. paraintracellulare (MP), M. indicus pranii (MIP) and M. yonogonense. Based on the average nucleotide identity, the clinical M. intracellulare strains were phylogenetically grouped in two clusters: (1) the typical M. intracellulare (TMI) group, including ATCC13950 and virulent M.i.27 and M.i.198 that we previously reported, and (2) the MP-MIP group. The alignment of the genomic regions was mostly preserved between groups. Plasmids were identified between groups and subgroups, including a plasmid common among some strains of the M.i.27 subgroup. Several genomic regions including those encoding factors involved in lipid metabolism (e.g., fadE3, fadE33), transporters (e.g., mce3), and type VII secretion system (genes of ESX-2 system) were shown to be hypermutated in the clinical strains. M. intracellulare was shown to be pan-genomic at the species and subspecies levels. The mce genes were specific to particular subspecies, suggesting that these genes may be helpful in discriminating virulence phenotypes between subspecies. Conclusions Our data suggest that genomic diversity among M. intracellulare, M. paraintracellulare, M. indicus pranii and M. yonogonense remains at the subspecies or genovar levels and does not reach the species level. Genetic components such as mce genes revealed by the comparative genomic analysis could be the novel focus for further insight into the mechanism of human pathogenesis for M. intracellulare and related strains.

Objective: The prevalence of clinical cases of pulmonary non‐tuberculous mycobacteria (NTM) is increasing worldwide. The aim of this study was to determine the proportion and the NTM species isolated from presumptive tuberculosis patients in Lambaréné, Gabon. Method: From January 2018 to December 2020, sputum samples from presumptive TB patients were analysed at the tuberculosis reference laboratory of the Centre de Recherches Médicales de Lambaréné. Two sputum samples were collected per patient, and culture was performed using Bactec MGIT 960. The GenoType Mycobacterium CM/AS was used for NTM isolates confirmation and species differentiation. Results: Among 1363 sputum samples analysed, 285 (20.9%) were Auramin acid fast bacilli (AFB) smear‐positive. NTM were isolated in 137/1363 (10%) of the samples. The most prevalent NTM species was Mycobacterium intracellulare (n = 74; 54%). Conclusion: These results show the presence of NTM among presumptive TB patients in Gabon, which could potentially complicate TB diagnosis. This presents a new public health challenge, and emphasises the need to consider NTM in planning the prevention and management of tuberculosis control. [ABSTRACT FROM AUTHOR]

Recurrence of oestrogen receptor (ER)-positive breast cancer rarely occurs postoperatively after a long period. Breast cancer cells survive and settle in distant organs in a dormant state, a phenomenon known as “tumour dormancy.” Here, we present a 66-year-old woman with recurrence of ER-positive breast cancer in the left lung 23 years after surgery accompanied with non-tuberculous mycobacterium infection (NTM). At the age of 43 years, the patient underwent a right mastectomy and adjuvant hormonotherapy to completely cure breast cancer. Twenty-three years after the operation, when the patient was 66 years old, computed tomography presented nodular shadows in the lower lobes bilaterally with bronchiectasis and ill-defined satellite tree-in-bud nodules. Mycobacterium intracellulare was detected in cultured bronchoalveolar lavage fluid obtained from the left lower lobe by bronchoscopy. Rifampicin, ethambutol, and clarithromycin were started, which resulted in shrinkage of the nodule in the right lower lobe and satellite nodules; however, the nodule in the left lower lobe increased in size gradually. Wedge resection of the left lower lobe containing the nodule by video-assisted thoracoscopic surgery was performed, which demonstrated that the nodule was adenocarcinoma in intraoperative pathological diagnosis; therefore, a left lower lobectomy and mediastinal lymph node dissection were performed. The tumour was revealed to be consistent with recurrence of previous breast cancer according to its morphology and immunohistochemical staining. Furthermore, caseous epithelioid cell granulomas existed in the periphery of the tumour. It is reported that inflammatory cytokines induce reawakening of dormant oestrogen-dependent breast cancer and, in our case, NTM infection might have stimulated the dormant tumour cells in the lower lobe.

Acanthamoeba is an opportunistic protozoon, widespread in the aquatic environment, where it can be in endosymbiosis with over 30 pathogenic bacteria, including nontuberculous mycobacteria (NTM). Protozoa play a crucial role in mycobacterial pathogenesis and serve as a reservoir of infection. Since the first step in bacteria making contact with amoebae is adhesion, we were interested in investigating whether essential oils (EOs) can affect it. To that end we investigated the effects of juniper (Juniperus communis) and immortelle (Helichrysum italicum) EOs against Mycobacterium avium, M. intracellulare, and M. gordonae in tap water and against their adhesion to Acanthamoeba castellanii by combining them in synergistic EO concentrations. M. avium and M. intracellulare adhered to A. castellanii to a greater extent than M. gordonae. The adhesion of all NTMs was prevented by the subinhibitory concentrations of EOs. When comparing the effect of synergistic combinations of EOs and the effect of a single concentration from a combination, a higher percentage of adhesion inhibition in all synergistic combinations observed, except against M. gordonae. Neither oil was cytotoxic to A. castellanii. Our findings suggest that the EOs or their components weaken the contact of environmental NTMs and free-living amoebae and indirectly diminish their pathogenic potential, which could be of value in developing strategies for maintenance of water supply systems.

All members of the genus Mycobacterium are collectively labeled as “non-tuberculous mycobacterium” (NTM), with the exception of the Mycobacterium tuberculosis complex and M. leprae. Recently, the incidence of NTM infection and number of cases have been increasing, but their identification remains difficult in some countries. Usually, NTM infections and diseases are associated with primary immunodeficiency diseases (PIDs), and their prognoses can be improved with a timely diagnosis and appropriate treatment. Here, we report a case of a 3-year-old boy with disseminated NTM disease (Mycobacterium intracellulare) and interferon-γ receptor 1 (IFNGR1) deficiency. He presented with skin and soft-tissue disease, disseminated osteomyelitis, and pulmonary disease. Initially, we suspected an infection due to the Bacillus Calmette–Guérin vaccine but later suspected Langerhans cell histiocytosis. Following oral treatment of azithromycin, rifampicin, and ethambutol, his condition improved progressively according to clinical and imaging manifestations. This case highlights the importance of early identification of the pathogen in a timely prescription of specific treatments in PIDs patients. We also discuss our experience of treatment of M. intracellulare disease in patients with IFNGR1 deficiency.

Background: Nontuberculous mycobacterial disease has emerged worldwide over the past 20 years. However, there are currently few reports on the established technique for constructing knockout mutants of nontuberculous mycobacteria. Therefore, gene recombination techniques for nontuberculous mycobacteria require further research., Results: We constructed vector pPR23LHR that harbors the ribosomal protein S12 gene (rpsL + ) as a dominant negative selection marker and the hygromycin (Hyg) and lacZ cassettes as positive selection markers. We constructed knockout mutants of proteasomal genes, which we found to be required for hypoxic pellicle formation in Mycobacterium intracellulare by functional genomic analysis. The knockout mutants showed impaired hypoxic pellicle formation, consistent with previous data using epoxomicin, a proteasomal inhibitor., Conclusions: Our findings demonstrate that rpsL + is an efficient dominant negative selection marker for gene recombination in nontuberculous mycobacteria. Our temperature-sensitive rpsL + method for the construction of knockout mutants will facilitate functional assays to validate the virulence factors of nontuberculous mycobacteria and the pathogenesis of nontuberculous mycobacterial disease., (© 2024 The Author(s). Microbiology and Immunology published by The Societies and John Wiley & Sons Australia, Ltd.)

Background: M. intracellulare is a frequent causative pathogen of nontuberculous mycobacteria infection that causes infections in the respiratory tract, whose incidence is increasing in many countries. This study aimed at determining the VNTR-based genetic diversity of a collection of 39 M. intracellulare human strains isolated from respiratory specimens over the last 5 years. Results: The VNTR analysis showed that M. intracellulare strains displayed a high genetic diversity, indicating that the M. intracellulare genotypes are quite heterogeneous in our geographical area. Moreover, a comparison with VNTR profiles of strains from other countries confirmed that genotypes of clinical strains of M. intracellulare are not related to geographical origin. Conclusions: VNTR typing has proved to be a highly discriminatory method for better understanding the molecular epidemiology of M. intracellulare. [ABSTRACT FROM AUTHOR]

Purpose: Despite decreased prevalence of tuberculosis, the incidence of the diseases associated with nontuberculous mycobacteria (NTM) has been increasing in South Korea and around the world. The present retrospective study was conducted to determine longitudinal changes in the epidemiology and distribution of NTM over 13 years at a tertiary care hospital in Korea. Materials and Methods: We retrospectively analyzed data on Mycobacterium species over 13 years (January 2007 to December 2019) by utilizing the laboratory information system. Mycobacterium species were identified using biochemical tests and PCR-restriction fragment length polymorphism and Mycobacteria GenoBlot assays. Results: After excluding duplicates from the initial pool of 17996 mycobacterial isolates, 7674 strains were analyzed and 2984 (38.9%) NTM were isolated. The proportion of NTM continuously increased over the 13-year period, from 17.0% in 2007 to 57.5% in 2019. Among the NTM isolates, the most common species were Mycobacterium intracellulare (50.6%), M. avium (18.3%), M. fortuitum complex (4.9%), M. abscessus (4.5%), M. gordonae (3.3%), M. kansasii (1.1%), M. chelonae (1.0%), and M. massiliense (0.9%). In patients over the age of 70 years, the proportion of NTM among the isolates increased from 26.6% in 2007 to 62.0% in 2019, and that of M. intracellulare isolates among the NTM increased from 13.9% (11/79) in 2007 to 37.4% (175/468) in 2019. Conclusion: The number of NTM isolates continuously increased over the study period, and the increase in the proportion of M. intracellulare in patients aged over 70 years was notable. [ABSTRACT FROM AUTHOR]

To evaluate the immunogenicity of Mycobacterium intracellulare proteins and determine the cross-reactive proteins between M. intracellulare and M. tuberculosis. Protein extracts from M. intracellulare were used to immunize BALB/c mice. The antigens were evaluated using cellular and humoral immunoassays. The common genes between M. intracellular and M. tuberculosis were identified using genome-wide comparative analysis, and cross-reactive proteins were screened using immunoproteome microarrays. Immunization with M. intracellulare proteins induced significantly higher levels of the cytokines interferon-γ (IFN-γ), interleukin-2 (IL-2), interleukin-12 (IL-12), interleukin-6 (IL-6) and immunoglobulins IgG, IgG1, IgM, and IgG2a in mouse serum. Bone marrow-derived macrophages isolated from mice immunized with M. intracellulare antigens displayed significantly lower bacillary loads than those isolated from mice immunized with adjuvants. Whole-genome sequence analysis revealed 396 common genes between M. intracellulare and M. tuberculosis. Microchip hybridization with M. tuberculosis proteins revealed the presence of 478 proteins in the serum of mice immunized with M. intracellulare protein extracts. Sixty common antigens were found using both microchip and genomic comparative analyses. This is the advanced study to investigate the immunogenicity of M. intracellulare proteins and the cross-reactive proteins between M. intracellulare and M. tuberculosis. The results revealed the presence of a number of cross-reactive proteins between M. intracellulare and M. tuberculosis. Therefore, this study provides a new way of identifying immunogenic proteins for use in tuberculosis vaccines against both M. intracellulare and M. tuberculosis in future. [ABSTRACT FROM AUTHOR]

Background Nontuberculous mycobacteria (NTM) are a rare cause of infectious tenosynovitis of the upper extremity. Using molecular methods, clinical microbiology laboratories are increasingly reporting identification down to the species level. Improved methods for speciation are revealing new insights into the clinical and epidemiologic features of rare NTM infections. Methods We encountered 3 cases of epidemiologically linked upper extremity NTM tenosynovitis associated with exposure to hurricane-damaged wood. We conducted whole-genome sequencing to assess isolate relatedness followed by a literature review of NTM infections that involved the upper extremity. Results Despite shared epidemiologic risk, the cases were caused by 3 distinct organisms. Two cases were rare infections caused by closely related but distinct species within the Mycobacterium terrae complex that could not be differentiated by traditional methods. The third case was caused by Mycobacterium intracellulare. An updated literature review that focused on research that used modern molecular speciation methods found that several species within the M. terrae complex are increasingly reported as a cause of upper extremity tenosynovitis, often in association with environmental exposures. Conclusions These cases illustrate the importance of molecular methods for speciating phenotypically similar NTM, as well as the limitations of laboratory-based surveillance in detecting point-source outbreaks when the source is environmental and may involve multiple organisms. [ABSTRACT FROM AUTHOR]

Abstract Mycobacterium avium complex (MAC)‐infected lung bulla was a rare type of pulmonary non‐tuberculous mycobacterial (NTM) infection. A 29‐year‐old man with a history of tetralogy of Fallot was admitted to our hospital because of a high fever and left chest pain. Chest computed tomography showed two bullae with intrabullous fluid in both the lower lobes and centrilobular small nodular shadow in the right upper lobe and the left lower lobe. Culture of bronchoscopic washing specimen from the right upper lobe bronchus and left lower lobe one and purulent fluid drained from the bulla in the left lower lobe revealed Mycobacterium intracellulare. Percutaneous drainage from the left bulla and anti‐NTM treatment were performed. Afterwards, symptoms improved and two intrabullous fluid disappeared. Therefore, a diagnosis of multiple infected lung bullae associated with M. intracellulare was made. This is the first documented case of multiple infected lung bullae associated with MAC.

Background: Mycobacterium intracellulare is a representative etiological agent of emerging pulmonary M. avium-intracellulare complex disease in the industrialized countries worldwide. The recent genome sequencing of clinical strains isolated from pulmonary M. avium-intracellulare complex disease has provided insight into the genomic characteristics of pathogenic mycobacteria, especially for M. avium; however, the genomic characteristics of M. intracellulare remain to be elucidated. Results: In this study, we performed comparative genomic analysis of 55 M. intracellulare and related strains such as M. paraintracellulare (MP), M. indicus pranii (MIP) and M. yonogonense. Based on the average nucleotide identity, the clinical M. intracellulare strains were phylogenetically grouped in two clusters: (1) the typical M. intracellulare (TMI) group, including ATCC13950 and virulent M.i.27 and M.i.198 that we previously reported, and (2) the MP-MIP group. The alignment of the genomic regions was mostly preserved between groups. Plasmids were identified between groups and subgroups, including a plasmid common among some strains of the M.i.27 subgroup. Several genomic regions including those encoding factors involved in lipid metabolism (e.g., fadE3, fadE33), transporters (e.g., mce3), and type VII secretion system (genes of ESX-2 system) were shown to be hypermutated in the clinical strains. M. intracellulare was shown to be pan-genomic at the species and subspecies levels. The mce genes were specific to particular subspecies, suggesting that these genes may be helpful in discriminating virulence phenotypes between subspecies. Conclusions: Our data suggest that genomic diversity among M. intracellulare, M. paraintracellulare, M. indicus pranii and M. yonogonense remains at the subspecies or genovar levels and does not reach the species level. Genetic components such as mce genes revealed by the comparative genomic analysis could be the novel focus for further insight into the mechanism of human pathogenesis for M. intracellulare and related strains. [ABSTRACT FROM AUTHOR]

The prevalence of non-tuberculous mycobacteria (NTM) infections has been on the rise in recent years, especially among the elderly population and other immunocompromised groups. Risk factors for NTM infections include advanced age, preexisting pulmonary diseases, and low body mass index. This study presents a case of NTM pulmonary disease attributed to Mycobacterium intracellulare , which was rapidly identified using metagenomic next-generation sequencing (mNGS). An 82-year-old male presented with persistent fever, cough, and shortness of breath. Initial assessments revealed an elevated white blood cell count and high-sensitivity C-reactive protein, with chest CT showing newly formed nodular shadows and cavity formation. Sputum tests confirmed NTM infection through positive acid-fast staining and mNGS, which rapidly identified M. intracellulare within 48 hours. Subsequent sputum samples confirmed the diagnosis using traditional methods. The patient had a complex medical history, including pulmonary tuberculosis, chronic pancreatitis, chronic hepatitis B, diabetes, and malnutrition. The patient was treated with a combination of cefotaxime, moxifloxacin, clarithromycin, and acetylcysteine, in addition to receiving nutritional support. After the treatment, there was an improvement in symptoms, normalization of body temperature, and a decrease in cough and sputum production. This case highlights the significance of mNGS in promptly diagnosing and treating NTM pulmonary disease, especially in elderly patients with various underlying health conditions. The collaborative effort among different medical specialties enabled more thorough patient care, ultimately leading to better outcomes. Incorporating cutting-edge diagnostic techniques such as mNGS alongside a holistic treatment approach is crucial for the successful management of NTM infections in at-risk populations., Competing Interests: Human subjects: Consent was obtained or waived by all participants in this study. Conflicts of interest: In compliance with the ICMJE uniform disclosure form, all authors declare the following: Payment/services info: All authors have declared that no financial support was received from any organization for the submitted work. Financial relationships: All authors have declared that they have no financial relationships at present or within the previous three years with any organizations that might have an interest in the submitted work. Other relationships: All authors have declared that there are no other relationships or activities that could appear to have influenced the submitted work., (Copyright © 2024, Yang et al.)

The pathogen Mycobacterium avium complex (MAC) is the most common cause of nontuberculous mycobacterial pulmonary disease worldwide. The decision to initiate long-term antibiotic treatment is difficult for the physician due to inconsistent disease progression and adverse effects associated with the antibiotic treatment. The prognostic factors for the progression of MAC pulmonary disease are low body mass index, poor nutritional status, presence of cavitary lesion(s), extensive disease, and a positive acid-fast bacilli smear. A regimen consisting of macrolides (clarithromycin or azithromycin) with rifampin and ethambutol has been recommended; this regimen significantly improves the treatment of MAC pulmonary disease and should be maintained for at least 12 months after negative sputum culture conversion. However, the rates of default and disease recurrence after treatment completion are still high. Moreover, treatment failure or macrolide resistance can occur, although in some refractory cases, surgical lung resection can improve treatment outcomes. However, surgical resection should be carefully performed in a well-equipped center and be based on a rigorous risk-benefit analysis in a multidisciplinary setting. New therapies, including clofazimine, inhaled amikacin, and bedaquiline, have shown promising results for the treatment of MAC pulmonary disease, especially in patients with treatment failure or macrolide-resistant MAC pulmonary disease. However, further evidence of the efficacy and safety of these new treatment regimens is needed. Also, a new consensus is needed for treatment outcome definitions as widespread use of these definitions could increase the quality of evidence for the treatment of MAC pulmonary disease.

Acanthamoeba is an opportunistic protozoon, widespread in the aquatic environment, where it can be in endosymbiosis with over 30 pathogenic bacteria, including nontuberculous mycobacteria (NTM). Protozoa play a crucial role in mycobacterial pathogenesis and serve as a reservoir of infection. Since the first step in bacteria making contact with amoebae is adhesion, we were interested in investigating whether essential oils (EOs) can affect it. To that end we investigated the effects of juniper (Juniperus communis) and immortelle (Helichrysum italicum) EOs against Mycobacterium avium, M. intracellulare, and M. gordonae in tap water and against their adhesion to Acanthamoeba castellanii by combining them in synergistic EO concentrations. M. avium and M. intracellulare adhered to A. castellanii to a greater extent than M. gordonae. The adhesion of all NTMs was prevented by the subinhibitory concentrations of EOs. When comparing the effect of synergistic combinations of EOs and the effect of a single concentration from a combination, a higher percentage of adhesion inhibition in all synergistic combinations observed, except against M. gordonae. Neither oil was cytotoxic to A. castellanii. Our findings suggest that the EOs or their components weaken the contact of environmental NTMs and free-living amoebae and indirectly diminish their pathogenic potential, which could be of value in developing strategies for maintenance of water supply systems. [ABSTRACT FROM AUTHOR]

Recurrence of oestrogen receptor (ER)-positive breast cancer rarely occurs postoperatively after a long period. Breast cancer cells survive and settle in distant organs in a dormant state, a phenomenon known as "tumour dormancy." Here, we present a 66-year-old woman with recurrence of ER-positive breast cancer in the left lung 23 years after surgery accompanied with non-tuberculous mycobacterium infection (NTM). At the age of 43 years, the patient underwent a right mastectomy and adjuvant hormonotherapy to completely cure breast cancer. Twenty-three years after the operation, when the patient was 66 years old, computed tomography presented nodular shadows in the lower lobes bilaterally with bronchiectasis and ill-defined satellite tree-in-bud nodules. Mycobacterium intracellulare was detected in cultured bronchoalveolar lavage fluid obtained from the left lower lobe by bronchoscopy. Rifampicin, ethambutol, and clarithromycin were started, which resulted in shrinkage of the nodule in the right lower lobe and satellite nodules; however, the nodule in the left lower lobe increased in size gradually. Wedge resection of the left lower lobe containing the nodule by video-assisted thoracoscopic surgery was performed, which demonstrated that the nodule was adenocarcinoma in intraoperative pathological diagnosis; therefore, a left lower lobectomy and mediastinal lymph node dissection were performed. The tumour was revealed to be consistent with recurrence of previous breast cancer according to its morphology and immunohistochemical staining. Furthermore, caseous epithelioid cell granulomas existed in the periphery of the tumour. It is reported that inflammatory cytokines induce reawakening of dormant oestrogen-dependent breast cancer and, in our case, NTM infection might have stimulated the dormant tumour cells in the lower lobe. [ABSTRACT FROM AUTHOR]

The majority of nontuberculous mycobacterial (NTM) pulmonary infections in people with cystic fibrosis (CF) are caused by Mycobacterium avium complex (MAC) species. Data on MAC species distribution and outcomes of infection in CF are lacking. This was a single center, retrospective study. MAC isolates had species identification with MLSA of rpoB and the 16S 23S ITS region. Clinical data were compared between species. Twenty-three people with CF and 57 MAC isolates were included. Infection with M. avium was the most common (65.2%). M. intracellulare was associated with higher rates of NTM disease, younger age, and steeper decline in lung function prior to infection. We observed worse clinical outcomes in people with M. intracellulare infection relative to other MAC species. Further investigation of clinical outcomes of MAC infection among CF patients is warranted to better define the utility of species-level identification of MAC isolates in CF. [ABSTRACT FROM AUTHOR]