Transcriptome analysis of long non-coding RNAs in Mycobacterium avium complex-infected macrophages.

Mycobacterium avium complex (MAC) is a non-tuberculous mycobacterium widely distributed in the environment. Even though MAC infection is increasing in older women and immunocompromised patients, to our knowledge there has been no comprehensive analysis of the MAC-infected host-cell transcriptome-and particularly of long non-coding RNAs (lncRNAs). By using in vitro-cultured primary mouse bone-marrow-derived macrophages (BMDMs) and Cap analysis of gene expression, we analyzed the transcriptional and kinetic landscape of macrophage genes, with a focus on lncRNAs, during MAC infection. MAC infection of macrophages induced the expression of immune/inflammatory response genes and other genes similar to those involved in M1 macrophage activation, consistent with previous reports, although Nos2 (M1 activation) and Arg1 (M2 activation) had distinct expression profiles. We identified 31 upregulated and 30 downregulated lncRNA promoters corresponding respectively to 18 and 26 lncRNAs. Upregulated lncRNAs were clustered into two groups-early and late upregulated-predicted to be associated with immune activation and the immune response to infection, respectively. Furthermore, an Ingenuity Pathway Analysis revealed canonical pathways and upstream transcription regulators associated with differentially expressed lncRNAs. Several differentially expressed lncRNAs reported elsewhere underwent expressional changes upon M1 or M2 preactivation and subsequent MAC infection. Finally, we showed that expressional change of lncRNAs in MAC-infected BMDMs was mediated by toll-like receptor 2, although there may be other mechanisms that sense MAC infection. We identified differentially expressed lncRNAs in MAC-infected BMDMs, revealing diverse features that imply the distinct roles of these lncRNAs in MAC infection and macrophage polarization.

The Diagnosis of Nontuberculous Mycobacterial Pulmonary Disease by Single Bacterial Isolation Plus Anti-GPL-Core IgA Antibody.

Although serum anti-glycopeptidolipid (GPL)-core IgA antibody is a highly specific test for infection with Mycobacterium avium complex (MAC), Mycobacterium abscessus, and its subspecies abscessus, subsp. massiliense, and subsp. bolletii (MAB), its use for the definitive diagnosis of MAC pulmonary disease (PD) and MAB-PD are unknown. To clarify the diagnostic accuracy of the anti-GPL-core IgA antibody test among patients with radiologically suspected MAC-PD or MAB-PD who already have a single positive sputum culture test. The first isolations of MAC and MAB from patients with radiologically suspected MAC-PD or MAB-PD at the Osaka Toneyama Medical Center between January 2006 and December 2020 were collected. Patients were enrolled when their serum anti-GPL-core IgA antibody was measured during the 3 months before and after the first isolation. We retrospectively compared the results of anti-GPL-core IgA antibody testing with the final diagnoses based on the current guidelines. We included 976 patients for analysis. The serum anti-GPL-core IgA antibody was positive in 699 patients (71.6%). The positive predictive value of anti-GPL-core IgA antibody for the diagnosis of MAC-PD or MAB-PD was 97.4%. The median time required for the second positive culture after the first isolation was 51 days (interquartile range 12 to 196 days). The positive serum anti-GPL-core IgA antibody test allowed an early and definitive diagnosis of MAC-PD or MAB-PD in those who already had a single positive sputum culture test. IMPORTANCE To satisfy the microbiologic criteria of the current diagnostic guideline for nontuberculous mycobacterial pulmonary disease (PD), at least two positive sputum cultures of the same species of mycobacteria from sputum are required to avoid the casual isolation of mycobacteria. This study showed that the positivity of a serum anti-glycopeptidolipid (GPL)-core IgA antibody test has an excellent diagnostic ability among patients with radiologically suspected Mycobacterium avium complex (MAC)-PD or Mycobacterium abscessus (MAB)-PD who already had a single positive sputum culture test. The usage of single culture isolation plus anti-GPL-core IgA antibody as another diagnostic criterion has a time, cost, and effort-saving effect. Furthermore, it will facilitate the diagnosis of MAC-PD or MAB-PD in the early stage of disease because serum anti-GPL-core IgA antibody becomes high in these patients. Therefore, we proposed adding single culture isolation plus anti-GPL-core IgA antibody as "combined microbiological and serological criteria" to the diagnostic guidelines for MAC-PD and MAB-PD.

Immunogenicity of Whole Mycobacterium intracellulare Proteins and Fingding on the Cross-Reactive Proteins between M. intracellulare and M. tuberculosis.

OBJECTIVES: To evaluate the immunogenicity of Mycobacterium intracellulare proteins and determine the cross-reactive proteins between M. intracellulare and M. tuberculosis. METHODS: 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. RESULTS: 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. CONCLUSION: 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.

Association of serum antibodies against the Mycobacterium avium complex and hemoptysis: a cross-sectional study.

BACKGROUND: Hemoptysis is very common and can be life threatening in clinical practice for nontuberculous mycobacteria. The serum antibody against the Mycobacterium avium complex (MAC-Ab), the majority of nontuberculous mycobacteria species, is well known to reflect the activity of MAC lung disease; however, there is no study investigating the association between the MAC-Ab and hemoptysis in MAC patients. Therefore, we assessed whether the MAC-Ab is a good biomarker for hemoptysis among subjects with MAC lung disease. METHODS: This study was conducted as a five-year retrospective survey at the National Hospital Organization Fukuoka National Hospital. A total of 155 patients aged ≥20 years with MAC lung disease were enrolled and separated into seropositive and seronegative groups using the cutoff for MAC-Ab levels of 0.7 U/ml. The prevalence of hemoptysis and odds ratios for the presence of hemoptysis were estimated and compared between the groups. To investigate the linear trends in the relationship between MAC-Ab levels and hemoptysis, the subjects were classified into three groups using the tertile distribution of the MAC-Ab. RESULTS: The prevalence of hemoptysis was twice as high in the seropositive group than in the seronegative group (42.2 and 21.7%, respectively, P = 0.02). The multivariable-adjusted risk of hemoptysis was elevated in the seropositive group as compared with the seronegative group (odds ratio = 2.79 (95% confidence interval 1.15-7.44)). Likewise, when categorizing the subjects into three groups, the risk of hemoptysis increased with increasing MAC-Ab levels (P = 0.03 for trend). CONCLUSIONS: A positive MAC-Ab level was a significant risk factor for hemoptysis among patients with MAC lung disease. There were also positive trends in the association between the MAC-Ab titer and the likelihood of hemoptysis. Measuring the MAC-Ab may contribute not only to early detection of the risk of hemoptysis but also to early intervention with anti-NTM therapy and, as a result, to the prevention of hemoptysis in MAC patients.

Genetic Involvement of Mycobacterium avium Complex in the Regulation and Manipulation of Innate Immune Functions of Host Cells.

Mycobacterium avium complex (MAC), a collection of mycobacterial species representing nontuberculous mycobacteria, are characterized as ubiquitous and opportunistic pathogens. The incidence and prevalence of infectious diseases caused by MAC have been emerging globally due to complications in the treatment of MAC-pulmonary disease (PD) in humans and the lack of understating individual differences in genetic traits and pathogenesis of MAC species or subspecies. Despite genetically close one to another, mycobacteria species belonging to the MAC cause diseases to different host range along with a distinct spectrum of disease. In addition, unlike Mycobacterium tuberculosis, the underlying mechanisms for the pathogenesis of MAC infection from environmental sources of infection to their survival strategies within host cells have not been fully elucidated. In this review, we highlight unique genetic and genotypic differences in MAC species and the virulence factors conferring the ability to MAC for the tactics evading innate immune attacks of host cells based on the recent advances in genetic analysis by exemplifying M. avium subsp. hominissuis, a major representative pathogen causing MAC-PD in humans. Further understanding of the genetic link between host and MAC may contribute to enhance host anti-MAC immunity, but also provide novel therapeutic approaches targeting the pangenesis-associated genes of MAC.

Nrf2 Regulates Granuloma Formation and Macrophage Activation during Mycobacterium avium Infection via Mediating Nramp1 and HO-1 Expressions.

Nrf2 is a redox-sensitive transcription factor that is thought to be important in protection against intracellular pathogens. To determine the protective role of Nrf2 in the host defense against Mycobacterium avium complex (MAC), both wild-type and Nrf2-deficient mice were intranasally infected with MAC bacteria. Nrf2-deficient mice were highly susceptible to MAC bacteria compared with wild-type mice. There were no significant changes in the levels of oxidative stress and Th1 cytokine production between genotypes. Comprehensive transcriptome analysis showed that the expressions of Nramp1 and HO-1 were much lower in the infected lungs, and the expression of Nramp1 was especially lower in alveolar macrophages of Nrf2-deficient mice than of wild-type mice. Electron microscopy showed that many infected alveolar macrophages from Nrf2-deficient mice contained a large number of intracellular MAC bacteria with little formation of phagolysosomes, compared with those from wild-type mice. Treatment with sulforaphane, an activator of Nrf2, increased resistance to MAC with increased lung expression of Nramp1 and HO-1 in wild-type mice. These results indicate that Nramp1 and HO-1, regulated by Nrf2, are essential in defending against MAC infection due to the promotion of phagolysosome fusion and granuloma formation, respectively. Thus, Nrf2 is thought to be a critical determinant of host resistance to MAC infection.IMPORTANCE Nontuberculous mycobacteria (NTM) are an important cause of morbidity and mortality in pulmonary infections. Among them, Mycobacterium avium complex (MAC) is the most common cause of pulmonary NTM disease worldwide. It is thought that both environmental exposure and host susceptibility are required for the establishment of pulmonary MAC disease, because pulmonary MAC diseases are most commonly observed in slender, postmenopausal women without a clearly recognized immunodeficiency. However, host factors that regulate MAC susceptibility have not been elucidated until now. This study shows that Nrf2 is a critical regulator of host susceptibility to pulmonary MAC disease by promoting phagolysosome fusion and granuloma formation via activating Nramp1 and HO-1 genes, respectively. The Nrf2 system is activated in alveolar macrophages, the most important cells during MAC infection, as both the main reservoir of infection and bacillus-killing cells. Thus, augmentation of Nrf2 might be a useful therapeutic approach for protection against pulmonary MAC disease.

Mycobacterial immunevasion-Spotlight on the enemy within.

Discussion on how antibiotic treatment routes Mycobacterium avium to phagolysosomes without eliciting an immune response in human macrophages.

Immunogenicity of Whole / 生物医学与环境科学（英文）

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Evaluation of plasma anti-GPL-core IgA and IgG for diagnosis of disseminated non-tuberculous mycobacteria infection.

Detection of IgA antibody against Mycobacterium avium complex (MAC) glycopeptidolipid (GPL) has recently been shown to improve the diagnosis of MAC pulmonary disease but has yet to be tested in disseminated Non-tuberculous mycobacteria (NTM) infection. In this study, we address the diagnostic efficacies of an anti-GPL-core ELISA kit in disseminated lymphadenopathy patients positive for NTM culture and anti-IFN-γ autoantibodies. The study was conducted in a tertiary referral center in northeastern Thailand and patients with NTM, tuberculosis, melioidosis, and control subjects were enrolled. Plasma immunoglobulin A (IgA) and G (IgG) antibodies against GPL-core were detected in the subjects and the specificity and sensitivity of the assay was assessed. Anti-GPL-core IgA and IgG levels were significantly higher in NTM patients than other groups (p < 0.0001). Diagnostic efficacy for NTM patients using anti-GPL-core IgA cut-off value of 0.352 U/ml showed good sensitivity (91.18%) and intermediate specificity (70.15%). Using a cut-off value of 4.140 AU/ml for anti-GPL-core IgG showed the same sensitivity (91.18%) with increased specificity (89.55%) and an 81.58% positive predictive value. Most patients with moderate levels (4.140-7.955 AU/ml) of anti-GPL-core IgG had rapidly growing mycobacteria (RGM) infection. Taken together, the detection of anti-GPL-core antibodies could provide a novel option for the diagnosis and management of disseminated NTM infected patients.

Clinical significance of anti-glycopeptidolipid-core IgA antibodies in patients newly diagnosed with Mycobacterium avium complex lung disease.

BACKGROUND: Although recent studies have identified anti-glycopeptidolipid (GPL)-core IgA antibodies as a serodiagnostic test for Mycobacterium avium complex lung disease (MAC-LD), this test shows insufficient sensitivity. This study aimed to determine the clinical utility of these antibodies in assessing disease progression and the clinical characteristics of MAC-LD patients with negative antibody results. METHODS: We retrospectively reviewed the medical records of consecutive newly diagnosed, untreated MAC-LD patients in two referral hospitals. We evaluated the association of anti-GPL-core IgA antibody results with disease progression requiring treatment and the factors associated with negative antibody results. RESULTS: In total, 229 patients (161 females; median age, 71 years; 185 with nodular/bronchiectatic disease phenotype; 69 with cavitary lesions) were enrolled; 146 patients (64%) were anti-GPL-core IgA antibody-positive. Radiological severity scores were associated with anti-GPL-core IgA antibody titers. During the median 364-day follow-up, 114 patients (49.8%) required treatment. Multivariate Cox proportional hazards analysis showed that positive anti-GPL-core IgA antibody results, a younger age, the absence of malignancy, and the presence of cavitary lesions were associated with disease progression requiring treatment. Multivariate logistic analysis revealed that significant factors related to the negative antibody results included underlying pulmonary disease, lower radiological scores, chronic sinusitis, and macrolide monotherapy. CONCLUSION: In addition to cavitary lesions, anti-GPL-core IgA antibody positivity was associated with disease progression requiring treatment. Physicians should carefully use anti-GPL-core IgA antibody results for the diagnosis of patients with underlying pulmonary disease, chronic sinusitis, macrolide monotherapy, and lower radiological severity.

[Adenopathy and mammary carcinoma: It is sometimes in the details that one encounters hypersensitivity pneumonitis!] / Adénopathie et carcinome mammaire : c'est dans les détails que se cache la pneumopathie d'hypersensibilité !

INTRODUCTION: Hypersensitivity pneumonitis (HP) is an interstitial lung disease due to an immunological reaction to exposure, by inhalation, to a large variety of antigens. The patho-physiological mechanism remains poorly understood. The diagnosis can be challenging and requires a detailed medical history taking especially when the clinical presentation is atypical or when the causal agent remains unknown. CASE REPORT: We report the case of a 75-year-old woman with a history of mammary carcinoma who presented with recently identified intramammary adenopathy. Biopsy of the adenopathy revealed non-necrotising, giant cell epithelioid granuloma. A diagnosis of hot tub lung with extra-pulmonary granulomatous lymph node involvement was made based on the clinical, functional, radiological and microbiological investigations. The evolution was favorable following antigen avoidance. CONCLUSION: Extrapulmonary lymph node involvement is rare in HP, suggesting a systemic inflammatory involvement.

The New Frontier of Host-Directed Therapies for Mycobacterium avium Complex.

Mycobacterium avium complex (MAC) is an increasingly important cause of morbidity and mortality, and is responsible for pulmonary infection in patients with underlying lung disease and disseminated disease in patients with AIDS. MAC has evolved various virulence strategies to subvert immune responses and persist in the infected host. Current treatment for MAC is challenging, requiring a combination of multiple antibiotics given over a long time period (for at least 12 months after negative sputum culture conversion). Moreover, even after eradication of infection, many patients are left with residual lung dysfunction. In order to address similar challenges facing the management of patients with tuberculosis, recent attention has focused on the development of novel adjunctive, host-directed therapies (HDTs), with the goal of accelerating the clearance of mycobacteria by immune defenses and reducing or reversing mycobacterial-induced lung damage. In this review, we will summarize the evidence supporting specific adjunctive, HDTs for MAC, with a focus on the repurposing of existing immune-modulatory agents targeting a variety of different cellular pathways. We also highlight areas meriting further investigation.

Aortic Graft Infection With Mycobacterium Avium Complex.

Aortic graft infections are uncommon complications after endovascular aortic surgery. In the majority of cases, gram-positive and then gram-negative organisms are the causative agents leading to this condition. Atypical organisms are traditionally not responsible for graft infection unless the patient is immunocompromised. We are reporting a case of culture-confirmed mycobacterium avium complex infection of an aortic graft in a well-controlled patient with HIV who had an undetected viral load and a CD4 count of 324 while on highly active antiretroviral therapy.

Alpha-1-Antitrypsin Enhances Primary Human Macrophage Immunity Against Non-tuberculous Mycobacteria.

Rationale: The association between non-tuberculous mycobacterial lung disease and alpha-1-antitrypsin (AAT) deficiency is likely due, in part, to underlying emphysema or bronchiectasis. But there is increasing evidence that AAT itself enhances host immunity against microbial pathogens and thus deficiency could compromise host protection. Objectives: The goal of this project is to determine if AAT could augment macrophage activity against non-tuberculous mycobacteria. Methods: We compared the ability of monocyte-derived macrophages cultured in autologous plasma that were obtained immediately before and soon after AAT infusion-given to individuals with AAT deficiency-to control an ex vivo Mycobacterium intracellulare infection. Measurements and Main Results: We found that compared to pre-AAT infused monocyte-derived macrophages plus plasma, macrophages, and contemporaneous plasma obtained after a session of AAT infusion were significantly better able to control M. intracellulare infection; the reduced bacterial burden was linked with greater phagosome-lysosome fusion and increased autophagosome formation/maturation, the latter due to AAT inhibition of both M. intracellulare-induced nuclear factor-kappa B activation and A20 expression. While there was a modest increase in apoptosis in the M. intracellulare-infected post-AAT infused macrophages and plasma, inhibiting caspase-3 in THP-1 cells, monocyte-derived macrophages, and alveolar macrophages unexpectedly reduced the M. intracellulare burden, indicating that apoptosis impairs macrophage control of M. intracellulare and that the host protective effects of AAT occurred despite inducing apoptosis. Conclusion: AAT augments macrophage control of M. intracellulare infection through enhancing phagosome-lysosome fusion and autophagy.

Aerosol immunization by alginate coated mycobacterium (BCG/MIP) particles provide enhanced immune response and protective efficacy than aerosol of plain mycobacterium against M.tb. H37Rv infection in mice.

BACKGROUND: With the aim of preparing a more effective, safe and economical vaccine for tuberculosis, inhalable live mycobacterium formulations were evaluated. METHODS: Alginate particles in the size range of 2-4 µm were prepared by encapsulating live Bacille Calmette-Guérin (BCG) and "Mycobacterium indicus pranii" (MIP). These particles were characterized for their size, stability and release profile. Mice were immunized with liquid aerosol or dry powder aerosol (DPA) alginate encapsulated mycobacterium particles and their in-vitro recall response and infection with mycobacterium H37Rv were investigated. RESULTS: It was found that the DPA of alginate encapsulated mycobacterium particles invoked superior immune response and provided higher protection in mice than the liquid aerosol. The BCG encapsulated in alginate particles (BEAP) and MIP encapsulated in alginate particles (MEAP) were engulfed by bone marrow dendritic cells (BMDCs) and co-localized with lysosome. The MEAP/BEAP activated BMDCs exhibited higher chemotaxis movement and had enhanced ability of antigen presentation to T cells. The in-vitro recall response of BEAP/MEAP immunized mice when compared in terms of proliferation index and Interferon gamma (IFN-gamma) released by splenocytes and mediastinal lymph node cells was found to be higher than mice immunized by liquid aerosol of BCG/MIP. Finally, different groups of immunized mice were infected with M. tb H37Rv and after 16 weeks the Colony forming units (CFUs) in lung and spleen estimated. The bacilli burden in the BEAP/MEAP immunized mice was significantly less than the respective liquid aerosol immunized mice and the histopathology of BEAP/MEAP immunized mice lungs showed very little damage. CONCLUSIONS: These inhale-able vaccines formulation of alginate coated live mycobacterium are more immunogenic as compared to the aerosol of bacilli and they provide better protection in mice when infected with H37Rv.

Application of a commercial serodiagnostic kit that measures the serum anti-glycopeptidolipid core IgA antibody in Mycobacterium avium complex pulmonary disease.

The diagnosis of Mycobacterium avium complex (MAC) pulmonary disease is occasionally cumbersome and time-consuming because the MAC species is ubiquitous, and therefore its detection is not necessarily indicative of a definitive diagnosis. A serodiagnostic method specific for MAC pulmonary disease that measures the serum anti-glycopeptidolipid core antigen IgA has been developed and is commercially available. Meta-analysis revealed that the test showed a good diagnostic accuracy. The estimated sensitivity and specificity values were 69.6% (95% confidence interval 62.1-76.1) and 90.6% (95% confidence interval 83.6-95.1), respectively. As antibody levels may reflect the disease activity, their serial measurement can also be used in the management of MAC disease. To justify its routine use in clinical practice, further validation in various regions and studies addressing whether serodiagnosis combined with present diagnostic criteria facilitate more rapid accurate diagnosis of MAC pulmonary disease are necessary.

Mycobacterium avium Complex Pleuritis with Elevated Anti-glycopeptidolipid-core IgA Antibody Levels in Pleural Effusion.

Pleuritis caused by nontuberculous mycobacteria is uncommon and difficult to diagnose. We herein report a case of Mycobacterium avium complex (MAC) pleuritis with elevated anti-glycopeptidolipid (GPL)-core IgA antibody levels in the pleural effusion. A 73-year-old woman with MAC pulmonary disease presented with massive left pleural effusion. A pleural biopsy by video-assisted thoracoscopic surgery was performed, revealing many noncaseating epithelioid cell granulomas. MAC was not identified by culture of the pleural effusion or specimens, but the anti-GPL-core IgA antibody level was markedly elevated in the pleural effusion. Measurement of anti-GPL-core IgA levels in the pleural fluid may be useful for diagnosing MAC pleuritis.

Mycobacteria-Specific T Cells May Be Expanded From Healthy Donors and Are Near Absent in Primary Immunodeficiency Disorders.

Mycobacterial Infections can be severe in patients with T-cell deficiency or phagocyte disorders, and treatment is frequently complicated by antimicrobial resistance. Restoration of T-cell immunity via stem cell transplantation facilitates control of mycobacterial infections, but presence of active infections during transplantation is associated with a higher risk of mortality. Adoptive T cell immunotherapy has been successful in targeting viruses, but has not been attempted to treat mycobacterial infections. We sought to expand and characterize mycobacterial-specific T-cells derived from healthy donors in order to determine suitability for adoptive immunotherapy. Mycobacteria-specific T-cells (MSTs) were generated from 10 healthy donors using a rapid ex vivo expansion protocol targeting five known mycobacterial target proteins (AG85B, PPE68, ESXA, ESXB, and ADK). MSTs were compared to T-cells expanded from the same donors using lysate from M. tuberculosis or purified protein derivative from M. avium (sensitin). MST expansion from seven patients with primary immunodeficiency disorders (PID) and two patients with IFN-γ autoantibodies and invasive M. avium infections. MSTs expanded from healthy donors recognized a median of 3 of 5 antigens, with production of IFN-γ, TNF, and GM-CSF in CD4+ T cells. Comparison of donors who received BCG vaccine (n = 6) to those who did not (n = 4) showed differential responses to PPE68 (p = 0.028) and ADK (p = 0.015) by IFN-γ ELISpot. MSTs expanded from lysate or sensitin also recognized multiple mycobacterial antigens, with a statistically significant differences noted only in the response to PPE68 (p = 0.016). MSTs expanded from patients with primary immunodeficiency (PID) and invasive mycobacterial infections showed activity against mycobacterial antigens in only two of seven subjects, whereas both patients with IFN-γ autoantibodies recognized mycobacterial antigens. Thus, MSTs can be generated from donors using a rapid expansion protocol regardless of history of BCG immunization. Most tested PID patients had no detectable T-cell immunity to mycobacteria despite history of infection. MSTs may have clinical utility for adoptive immunotherapy in T-cell deficient patients with invasive mycobacterial infections.

Role of Mycobacterium avium lysate INF-γ, IL-17, and IL-2 ELISPOT assays in diagnosing nontuberculous mycobacteria lymphadenitis in children.

Nontuberculous mycobacteria are the most frequent cause of chronic cervical lymphadenitis in childhood. The aim of the study was to evaluate the performance of IL-2, IL-17, and INF-γ in-house enzyme-linked immunospot assays using a Mycobacterium avium lysate, in order to identify a noninvasive diagnostic method of nontuberculous mycobacteria infection. Children with subacute and chronic lymphadenopathies or with a previous diagnosis of nontuberculous mycobacteria lymphadenitis were prospectively enrolled in the study. Sixty children with lymphadenitis were included in our study: 16 with confirmed infection (group 1), 30 probable infected (group 2) and 14 uninfected (group 3). Significantly higher median cytokine values were found in group 1 vs group 2, in group 1 vs group 3, and in group 2 vs group 3 considering IL-2-based enzyme-linked immunospot assay (p = 0.015, p < 0.001, p = 0.004, respectively). INF-γ-based enzyme-linked immunospot assay results were significantly higher in group 2 vs group 3 (p = 0.010). Differences between infected and uninfected children were not significant considering IL-17 assays (p = 0.431). Mycobacterium avium lysate IL-2 and INF-γ-based enzyme-linked immunospot assays seem to be promising noninvasive diagnostic techniques for discriminating children with nontuberculous mycobacteria lymphadenitis and noninfected subjects.

Antibody response against Mycobacterium avium complex in cystic fibrosis patients measured by a novel IgG ELISA test.

BACKGROUND: Early signs of Mycobacterium avium complex pulmonary disease can be missed in patients with cystic fibrosis due to subclinical infection or delays in mycobacterial culture. The aim of this study was to determine the diagnostic accuracy of a novel enzyme linked immunosorbent assay for immunoglobulin G against Mycobacterium avium complex, which could help stratify patients according to risk. METHODS: A retrospective cross sectional analysis of serum samples from the Copenhagen Cystic Fibrosis Center was performed. Corresponding clinical data were reviewed and patients with cystic fibrosis were assigned to one of four groups based on their mycobacterial culture results. In addition, anti-Mycobacterium avium complex immunoglobulin G levels were measured longitudinally before and after first positive culture in the period 1984-2015. RESULTS: Three-hundred and five patients with cystic fibrosis were included with a median of five nontuberculous mycobacterial cultures. Four individuals had Mycobacterium avium complex pulmonary disease at the time of cross sectional testing and their median antibody level was 22-fold higher than patients with no history of infection (1820 vs. 80 IgG units; p < 0.001). Test sensitivity was 100% (95% CI 40-100) and specificity 77% (95% CI 72-81). Longitudinal kinetics showed rising antibodies prior to first positive culture suggesting diagnostic delay. CONCLUSIONS: Antibody screening for Mycobacterium avium complex may be used as a supplement to culture. Although confirmation in a larger cohort is needed, our findings suggest that stratifying a cystic fibrosis population into high- and low-risk groups based on antibody levels may help clinicians identify patients in need of more frequent culture.