Mycobacterium tuberculosis growth inhibition by peripheral blood mononuclear cells from household contacts is not affected by previous SARS-CoV-2 infection

IgE-Dependent Activation of T cells by Allergen in Atopic Dermatitis: Pathophysiologic Relevance

Tuberculosis (TB) is a one of the main causes of mortality and morbidity worldwide. Bactec MGIT (Mycobacteria Growth Indicator Tube) system is a rapid, reliable automated system for early diagnosis of pulmonary and extra pulmonary TB in setups where purchase of expensive instruments is not possible. The present study was thus carried out to evaluate AFB microscopy, culture on Lowenstein Jensen media and micro MGIT system for early and accurate diagnosis of Tuberculosis. A total of 280 samples were processed for direct AFB smear examination, and culture on micro MGIT and LJ media. The identification of Mycobacterium tuberculosis complex in positive cultures was done by MPT64 Ag card test (BD MGIT TBC Identification Test). Out of the processed samples, (47.1%) 132/280 were positive for Mycobacterium spp by Micro MGIT, (35%) 98/280 on LJ medium and (25.7%) 72/280 by AFB smear. A total of (48.5%) 136 samples were positive by a combination of Micro MGIT and LJ medium. Among the total positive samples (136/280), Micro MGIT was found to be positive in 97% (132/136) of samples, LJ was positive in 72% (98/136), while 52.9% (72/136) were positive by AFB smear. Manual MGIT System is a simple and efficient, safe to use the diagnostic system. It does not require any expensive/special instrumentation other than the UV lamp for the detection of fluorescence. In areas with limited resources where the purchase of expensive instruments such as the MGIT 960 is out of scope, the use of manual MGIT for rapid susceptibility testing for MDR-TB could be an option. We would recommend testing MGIT 960 using first and secondline drugs to determine DST.

Long noncoding small nucleolar RNA (LncRNA) host gene 16 (SNHG16) is associated with certain diseases, including cancers. However, its role and mechanism in Mycobacterium tuberculosis (Mtb) infection remain unclear. Here, we demonstrated that SNHG16 expression levels were suppressed in peripheral blood mononuclear cells (PBMCs) and CD14+ monocytes of tuberculosis (TB) patients. SNHG16 was up-regulated by acute Mtb infection of PBMCs from healthy control (HC) subjects. Such TB suppression of SNHG16 was consistent with an immunosuppressive-like state driven by IL-10 signaling as seen in TB patients. Notably, SNHG16 limited Mtb growth in macrophages/monocytes through autophagy and vitamin D receptor (VDR)-dependent cathelicidin (CAMP) antimicrobial pathways. Concurrently, SNHG16 was highly expressed in lymphocytes, including CD8+ and Vγ2 Vδ2 T-cell subsets in HCs. SNHG16 overexpression in lymphocytes allowed them to control Mtb infection in macrophages, and SNHG16 epigenetically increased the expression of anti-Mtb effector cytokines in lymphocytes by developing more accessible chromatin states in gene loci encoding IFN-γ, TNF-α, and Granzyme B. Furthermore, the adoptive transfer of SNHG16-overexpressing human PBMCs into Mtb-infected SCID mice conferred protective immunity against Mtb infection. Thus, SNHG16 drove the induction of pleiotropic effector functions that inhibited intracellular Mtb growth in vitro and in vivo, serving as an immunotherapy target in TB.

Introduction: Sputum negative pulmonary Tuberculosis (TB) is a major public health problem. So, the emergence of new techniques for a more precise and rapid microbiological identification of Mycobacterium tuberculosis in clinical samples is of great importance to improve the management of TB. Aim: To determine and compare the sensitivity and turnaround time for Mycobacterium tuberculosis detection by the BACTEC Mycobacteria Growth Indicator Tube (MGIT) 960 system, Lowenstein Jensen (LJ) medium and Ziehl-Neelsen (ZN) staining. Materials and Methods: An Institution based, observational, cross-sectional study was conducted at Rajendra Institute of Medical Sciences (RIMS), Ranchi, Jharkhand, India, from July 2013-March 2016. Sputum, pericardial fluid, pleural fluid, peritoneal fluid, pus and endometrial tissue samples were collected from 80 patients of suspected TB cases. All were Acid-Fast stained by ZN staining method and cultured on solid culture LJ medium and on liquid medium (MGIT). Data was analysed using Statistical Package for the Social Sciences (SPSS) software, Version 20.0 (SPSS Inc, Chicago, IL, USA). Fisher’s-Exact test was used to show association of categorical variables. Non-parametric Mann-Whitney U test was used to show median difference of non-normally distributed continuous variables of two groups. Results: Out of the 80 samples, 41 cases were positive by either of the all methods. The positive specimen for ZN staining, LJ media and MGIT were 21, 29 and 41 cases respectively. The mean Time To Detection (TTD) was shorter for MGIT system than LJ media. Both LJ medium and MGIT 960 detected all cases of sputum smear positive cases and in addition significantly higher number than ZN stain in sputum smear negative cases. MGIT 960 detected significantly higher number of cases of sputum negative cases than LJ Media. The mean TTD was also significantly shorter in case of smear positive cases than the smear negative cases by both the solid and liquid culture mediums. Conclusion: The use of liquid media (MGIT) is more accurate and rapid method for the diagnosis of TB. The combination of more than one method is also highly recommended for rapid detection and early treatment of TB.

Decision letter: Mycobacterium tuberculosis induces decelerated bioenergetic metabolism in human macrophages

<p dir="ltr">This thesis presents four studies aimed at elucidating immune responses to Mycobacterium tuberculosis (Mtb) infection, with a focus on the associations with clinical features, such as tuberculosis (TB) disease severity and anemia, and the role of histone deacetylase inhibitors (HDACi) in modulating these responses.</p><p dir="ltr">In Study I, we developed a protocol for differentiation and polarization of monocyte-derived macrophages into M1 and M2 cells. Setting up a macrophage infection model with the virulent Mtb-H37Rv strain expressing green fluorescent protein (GFP), allowed assessment of the phenotype and function of Mtb-infected macrophages using flow cytometry. Results from this study indicated that Mtb modulates the polarization of macrophages and is efficiently internalized inside M2 polarized macrophages, which have lower capabilities of clearing the infection effectively. M1 macrophages, known for their robust response to bacterial infections, showed an overall lower Mtb-GFP signal potentially indicating an enhanced capacity for bacterial clearance, although limitations in bacterial containment were also noted in the M1 subset.</p><p dir="ltr">In Study II, we investigated how peripheral inflammatory mediator and cytokine levels associated with distinct features in patients with pulmonary TB, including clinical disease severity and anemia. This TB patient cohort represented a sub- group obtained from a randomized clinical trial previously conducted in Ethiopia. Multiplex Magpix data from plasma was examined with machine learning and manual analyses which identified key inflammatory markers that differentiated between severe and mild TB, anemic and non-anemic cases, and patients with extensive and limited lung involvement. These analyses revealed effective ranking of immune markers that separated the immune responses at baseline and after the start of anti-TB treatment in the different patient groups and compared to the healthy controls. Notably, while certain markers correlated with immune protection, other markers decreased with treatment, highlighting their potential as biomarkers for disease progression and response to therapy.</p><p dir="ltr">Previous work on the HDACi, phenylbutyrate (PBA), in the presence or absence of vitamin D, have demonstrated enhanced immune functions in Mtb-infected macrophages in vitro and clinical improvement of pulmonary TB upon administration to patients in vivo. However, PBA is a broad-spectrum HDACi with relatively weak HDACi activity in the millimolar-range. Therefore, Study III was designed to test a panel of commercially available HDACi compounds for their efficacy to reduce intracellular Mtb growth in immune cells. For this purpose, we used the macrophage infection model of Study I and IncuCyte live-cell imaging to enable medium-throughput screening of different compounds at different doses in cells obtained from healthy blood donors. The results demonstrated that four different SIRT1 and SIRT2 inhibitors, particularly tenovin, effectively enhanced restriction of Mtb growth in both Mtb-infected macrophages and bulk peripheral blood mononuclear cells (PBMCs). Specific SIRT2 inhibitors emerged as promising compounds useful for boosting intracellular bacterial clearance, although the efficacy of these molecules seemed dependent on the concentration and kinetics used. This study contributes to the growing body of research on HDACi as potential host-directed therapies for TB.</p><p dir="ltr">To continue the exploration of immune responses in the clinical trial cohort used in Study II, the work in Study IV was focused on phenotypical and transcriptional profiling of peripheral immune cells in TB patients using bulk RNA-sequencing (seq) on FACS-sorted lymphocytes and myeloid cells in combination with flow cytometry. Bulk RNA-seq data obtained from patients with mild, moderate or severe TB disease as compared to the controls, revealed that several genes where differentially regulated in the CD4+, CD8+, and monocyte cell subsets. Moreover, markers such as CD25, CD38, and HLA-DR were elevated in TB patients, particularly in those with severe disease and anemia, and were linked to disease severity and immune activation. Activated T cells expressing Ox40 as well as CD25+FoxP3+ and CD38+HLA-DR+ T cells were associated with disease severity and of potential relevance in chronic TB. These markers and T cell subsets decreased with effective treatment, suggesting potential use as prognostic markers of treatment response.</p><p dir="ltr">Together, these studies highlight significant insights into protective and pathological immune responses involved in TB, the impact of clinical features on immune responses, and the potential of HDAC inhibitors as adjunct host-directed therapies for modulation of TB immunity and to improve Mtb control.</p><h3>List of scientific papers</h3><p dir="ltr">I. Mily A, Kalsum S, <b>Loreti MG</b>, Rekha RS, Muvva JR, Lourda M, Brighenti S. Polarization of M1 and M2 Human Monocyte-Derived Cells and Analysis with Flow Cytometry upon M. tuberculosis Infection. Journal of Visualized Experiments, 2020 Sep 18;(163). <a href="https://doi.org/10.3791/61807" rel="noreferrer" target="_blank">https://doi.org/10.3791/61807</a></p><p dir="ltr">II. Ashenafi At, <b>Loreti MG</b>+, Bekele A, Aseffa G, Amogne W, Kassa E, Aderaye G, Brighenti S (+ Equal contribution) Inflammatory immune profiles associated with disease severity in pulmonary TB patients with moderate to severe clinical TB or anemia Frontiers in Immunology, 2023 Dec 12 (Vol 14 - 2023). <a href="https://doi.org/10.3389/fimmu.2023.1296501" rel="noreferrer" target="_blank">https://doi.org/10.3389/fimmu.2023.1296501</a></p><p dir="ltr">III. Kalsum S, Akber A, <b>Loreti MG</b>, Andersson B, Danielson E, Lerm M, Brighenti Sirtuin inhibitors reduce intracellular growth of M. tuberculosis in human macrophages via modulation of host cell immunity. [Submitted]</p><p dir="ltr">IV. <b>Loreti MG</b>, Mily A, Akber M, Ashenafi S, Bekele A, Bekele A, Aseffa G, Amogne W, Kassa E, Aderaye G, Lourda M, Brighenti S Immune profiling of peripheral T cells and myeloid cells in patients with pulmonary TB exhibiting diverse disease severity traits. [Manuscript]</p>

<p dir="ltr">This thesis presents four studies aimed at elucidating immune responses to Mycobacterium tuberculosis (Mtb) infection, with a focus on the associations with clinical features, such as tuberculosis (TB) disease severity and anemia, and the role of histone deacetylase inhibitors (HDACi) in modulating these responses.</p><p dir="ltr">In Study I, we developed a protocol for differentiation and polarization of monocyte-derived macrophages into M1 and M2 cells. Setting up a macrophage infection model with the virulent Mtb-H37Rv strain expressing green fluorescent protein (GFP), allowed assessment of the phenotype and function of Mtb-infected macrophages using flow cytometry. Results from this study indicated that Mtb modulates the polarization of macrophages and is efficiently internalized inside M2 polarized macrophages, which have lower capabilities of clearing the infection effectively. M1 macrophages, known for their robust response to bacterial infections, showed an overall lower Mtb-GFP signal potentially indicating an enhanced capacity for bacterial clearance, although limitations in bacterial containment were also noted in the M1 subset.</p><p dir="ltr">In Study II, we investigated how peripheral inflammatory mediator and cytokine levels associated with distinct features in patients with pulmonary TB, including clinical disease severity and anemia. This TB patient cohort represented a sub- group obtained from a randomized clinical trial previously conducted in Ethiopia. Multiplex Magpix data from plasma was examined with machine learning and manual analyses which identified key inflammatory markers that differentiated between severe and mild TB, anemic and non-anemic cases, and patients with extensive and limited lung involvement. These analyses revealed effective ranking of immune markers that separated the immune responses at baseline and after the start of anti-TB treatment in the different patient groups and compared to the healthy controls. Notably, while certain markers correlated with immune protection, other markers decreased with treatment, highlighting their potential as biomarkers for disease progression and response to therapy.</p><p dir="ltr">Previous work on the HDACi, phenylbutyrate (PBA), in the presence or absence of vitamin D, have demonstrated enhanced immune functions in Mtb-infected macrophages in vitro and clinical improvement of pulmonary TB upon administration to patients in vivo. However, PBA is a broad-spectrum HDACi with relatively weak HDACi activity in the millimolar-range. Therefore, Study III was designed to test a panel of commercially available HDACi compounds for their efficacy to reduce intracellular Mtb growth in immune cells. For this purpose, we used the macrophage infection model of Study I and IncuCyte live-cell imaging to enable medium-throughput screening of different compounds at different doses in cells obtained from healthy blood donors. The results demonstrated that four different SIRT1 and SIRT2 inhibitors, particularly tenovin, effectively enhanced restriction of Mtb growth in both Mtb-infected macrophages and bulk peripheral blood mononuclear cells (PBMCs). Specific SIRT2 inhibitors emerged as promising compounds useful for boosting intracellular bacterial clearance, although the efficacy of these molecules seemed dependent on the concentration and kinetics used. This study contributes to the growing body of research on HDACi as potential host-directed therapies for TB.</p><p dir="ltr">To continue the exploration of immune responses in the clinical trial cohort used in Study II, the work in Study IV was focused on phenotypical and transcriptional profiling of peripheral immune cells in TB patients using bulk RNA-sequencing (seq) on FACS-sorted lymphocytes and myeloid cells in combination with flow cytometry. Bulk RNA-seq data obtained from patients with mild, moderate or severe TB disease as compared to the controls, revealed that several genes where differentially regulated in the CD4+, CD8+, and monocyte cell subsets. Moreover, markers such as CD25, CD38, and HLA-DR were elevated in TB patients, particularly in those with severe disease and anemia, and were linked to disease severity and immune activation. Activated T cells expressing Ox40 as well as CD25+FoxP3+ and CD38+HLA-DR+ T cells were associated with disease severity and of potential relevance in chronic TB. These markers and T cell subsets decreased with effective treatment, suggesting potential use as prognostic markers of treatment response.</p><p dir="ltr">Together, these studies highlight significant insights into protective and pathological immune responses involved in TB, the impact of clinical features on immune responses, and the potential of HDAC inhibitors as adjunct host-directed therapies for modulation of TB immunity and to improve Mtb control.</p><h3>List of scientific papers</h3><p dir="ltr">I. Mily A, Kalsum S, <b>Loreti MG</b>, Rekha RS, Muvva JR, Lourda M, Brighenti S. Polarization of M1 and M2 Human Monocyte-Derived Cells and Analysis with Flow Cytometry upon M. tuberculosis Infection. Journal of Visualized Experiments, 2020 Sep 18;(163). <a href="https://doi.org/10.3791/61807" rel="noreferrer" target="_blank">https://doi.org/10.3791/61807</a></p><p dir="ltr">II. Ashenafi At, <b>Loreti MG</b>+, Bekele A, Aseffa G, Amogne W, Kassa E, Aderaye G, Brighenti S (+ Equal contribution) Inflammatory immune profiles associated with disease severity in pulmonary TB patients with moderate to severe clinical TB or anemia Frontiers in Immunology, 2023 Dec 12 (Vol 14 - 2023). <a href="https://doi.org/10.3389/fimmu.2023.1296501" rel="noreferrer" target="_blank">https://doi.org/10.3389/fimmu.2023.1296501</a></p><p dir="ltr">III. Kalsum S, Akber A, <b>Loreti MG</b>, Andersson B, Danielson E, Lerm M, Brighenti Sirtuin inhibitors reduce intracellular growth of M. tuberculosis in human macrophages via modulation of host cell immunity. [Submitted]</p><p dir="ltr">IV. <b>Loreti MG</b>, Mily A, Akber M, Ashenafi S, Bekele A, Bekele A, Aseffa G, Amogne W, Kassa E, Aderaye G, Lourda M, Brighenti S Immune profiling of peripheral T cells and myeloid cells in patients with pulmonary TB exhibiting diverse disease severity traits. [Manuscript]</p>

Objective To evaluate the feasibility of Mycobacteria growth indicator tube (MGIT) liquid culture combined with rifampin resistance test real-time PCR (Xpert MTB/RIF test) in the diagnosis of tuberculosis. Methods 652 cases of suspected pulmonary tuberculosis from October 2014 to March 2015 in Quzhou People's Hospital were enrolled. The morning sputum samples were collected for acid-fast staining, Lowenstein-Jensen (L-J) culture, BACTEC MGIT culture and Xpert MTB/RIF test. Samples with positive results of MGIT liquid culture were subjected to strain identification and drug sensitivity test with fluid method.Methodological comparison between four methods was made and the performance of MGIT liquid culture combined with Xpert MTB/RIF test in the rapid diagnosis and drug resistance detection in Mycobacterium tuberculosis was evaluated by chi-square test. Results Among the samples from 399 confirmed tuberculosis patients, the diagnostic sensitivity of the 4 methods (acid-fast staining, L-J culture, MGIT culture and Xpert MTB/RIF test) were 17.0% (68/399), 23.8% (95/399), 37.8% (151/399) and 37.3% (149/399) respectively. The sensitivity and specificity of MGIT culture combined with Xpert MTB/RIF test was 39.8% (159/399) and 94.8% (240/253). The sensitivity of MGIT culture combined with Xpert MTB/RIF test was significantly higher than acid-fast staining (χ2=50.9, P<0.01) and L-J culture (χ2=23.7, P<0.01). The average detection time of MGIT culture was 7.5 days (smear positive and Xpert MTB/RIF positive), 13.4 days (smear negative and Xpert MTB/RIF positive) and 16.9 days (smear negative and Xpert MTB/RIF negative). The sensitivity and specificity of Xpert MTB/RIF test in rifampin resistance detection were 9/9 and 97.3% (129/132) respectively. The average detection time of fluid method for drug sensitivity test was 8.3 days. Conclusions MGIT liquid culture combined with Xpert MTB/RIF test can detect Mycobacterium tuberculosis and the drug sensitivity rapidly. The method is highly sensitive and specific. It is important for clinical diagnosis and treatment of tuberculosis.(Chin J Lab Med, 2016, 39: 272-276) Key words: Mycobacterium tuberculosis; Rifampin; Tuberculosis, multidrug-resistant; Polymerase chain reaction; Microbial sensitivity tests

Household contacts (HHCs) of patients with active tuberculosis (ATB) are at higher risk of Mycobacterium tuberculosis (M. tuberculosis) infection. However, the immune factors responsible for different defense responses in HHCs are unknown. Hence, we aimed to evaluate transcriptome signatures in human peripheral blood mononuclear cells (PBMCs) of HHCs to aid risk stratification. We recruited 112 HHCs of ATB patients and followed them for 6 years. Among the HHCs, only 2 developed ATB, while the remaining HHCs were classified into three groups: (1) HHC-1 group (n = 23): HHCs with consistently positive T-SPOT.TB test, negative chest radiograph, and no clinical symptoms or evidence of ATB during the 6-year follow-up period; (2) HHC-2 group (n = 15): HHCs with an initial positive T-SPOT result that later became negative without evidence of ATB; (3) HHC-3 group (n = 14): HHCs with a consistently negative T-SPOT.TB test and no clinical or radiological evidence of ATB. HHC-2 and HHC-3 were combined as HHC-23 group for analysis. RNA sequencing (RNA-seq) in PBMCs, with and without purified protein derivative (PPD) stimulation, identified significant differences in gene signatures between HHC-1 and HHC-23. Gene ontology analysis revealed functions related to bacterial pathogens, leukocyte chemotaxis, and inflammatory and cytokine responses. Modules associated with clinical features in the HHC-23 group were linked to the IL-17 signaling pathway, ferroptosis, complement and coagulation cascades, and the TNF signaling pathway. Validation using real-time PCR confirmed key genes like ATG-7, CXCL-3, and TNFRSF1B associated with infection outcomes in HHCs. Our research enhances understanding of disease mechanisms in HHCs. HHCs with persistent latent tuberculosis infection (HHC-1) showed significantly different gene expression compared to HHCs with no M. tuberculosis infection (HHC-23). These findings can help identify HHCs at risk of developing ATB and guide targeted public health interventions.

Early identification and characterization of rifampicin-resistant (R(r)) Mycobacterium tuberculosis isolates recovered from the samples of tuberculosis (TB) patients in the Aegean (West Anatolian) Region was intended. Sixty isolates [47 (78.3%) multidrug-resistant (MDR)], which were identified as M. tuberculosis complex and phenotypically resistant to rifampicin by both BACTEC mycobacteria growth indicator tube (MGIT) 960 and 460 systems were analysed by a commercial line probe assay (INNO-LiPA Rif TB). The concordance of LiPA with the in vitro susceptibility test was found as 98.3%. Among the isolates, S531L (R5 pattern; 46.7%) and L511P/R, S512T, Q513L/K (DeltaS1 pattern; 11.7%) were the most frequent mutation patterns. As compared with the BACTEC systems and conventional techniques for cultivation, identification and in vitro susceptibility testing, INNO-LiPA Rif TB after cultivation in BACTEC MGIT 960 system provided an average of 20 days early diagnosis of R(r)M. tuberculosis isolates. Rapid molecular identification and characterization of R(r)M. tuberculosis isolates after BACTEC MGIT 960 cultivation would be useful for faster diagnosis, infection control and planning of accurate treatment in MDR-TB patients. Patients with MDR-TB need a specified treatment and efficient follow-up strategies. Rapid and practical methodologies to diagnose and follow these patients should be applied in routine use.

Study Design:Retrospective observational analysis.Objectives:Spinal tuberculosis accounts for about 50% of cases among extra pulmonary osteoarticular tuberculosis. Resistance to drugs in spinal tuberculosis patients is on a rise and there is inadequate literature concentrating on the precise pattern of resistance in Indian subcontinent which harbors 24% of global prevalence. The aim was to study the pattern of drug resistance in spinal tuberculosis among first- and second-line drugs. Drug resistance is common in spinal tuberculosis and we intended to find the prevalence of various drug resistance patterns.Methods:Patients with spinal tuberculosis visiting a tertiary center were assessed. Samples were taken from the affected vertebrae and sent for BACTEC mycobacterium growth indicator tube (MGIT) 960 culture. Patients with a positive growth in MGIT were included in the study. All previously treated patients (relapse, treatment after failure, treatment after loss to follow-up and other previously treated patients) were excluded.Results:A total of 150 patients with a positive growth in MGIT report were included in the study, of whom 43 patients had some kind of drug resistance. Seven were multidrug resistant (MDR), 9 had preextensive drug resistance (pre-XDR), and 4 had extensive drug resistance (XDR). Seventeen patients had mono-drug resistance, which was most frequently for isoniazid. Resistance among second-line drugs was common in the fluoroquinolone group.Conclusion:Drug resistance in spinal tuberculosis was found to be 28.6%. Of these, MDR was in 16.2%, pre-XDR in 20.9%, and XDR in 9.3% patients.

The aim of the study was to clarify how HIV infection affects tuberculosis liquid and solid culture results in a resource-limited setting. We used baseline data from the Study on Outcomes Related to Tuberculosis and HIV Drug Concentrations in Uganda (SOUTH), which included 268 HIV/tuberculosis (TB)-coinfected individuals. Culture results from Löwenstein-Jensen (LJ) solid culture and mycobacteria growth indicator tube (MGIT) liquid culture systems and culture-based correlates for bacillary density from the sputum of HIV/TB-coinfected individuals at baseline were analysed. Of 268 participants, 243 had a CD4 cell count available and were included in this analysis; 72.2% of cultures showed growth on solid culture and 82.2% in liquid culture systems (P<0.015). A higher CD4 cell count was predictive of LJ positivity [adjusted odds ratio (OR) 1.14; 95% confidence interval (CI) 1.03-1.25 per 50cells/μL increase; P=0.008]. The same, but insignificant trend was observed for MGIT positivity (adjusted OR 1.09; 95% CI 0.99-1.211 per 50cells/μL increase; P=0.094). A higher CD4 cell count was associated with a higher LJ colony-forming unit grade (adjusted OR 1.14; 95% CI 1.05-1.25 per 50 cells/μL increase; P=0.011) and a shorter time to MGIT positivity [adjusted hazard ratio (HR) 1.08; 95% CI 1.04-1.12 per 50 cells/μL increase; P<0.001]. In a resource-limited setting, the MGIT liquid culture system outperformed LJ solid culture in terms of culture yield and dependence on CD4 cell counts in HIV/TB-coinfected individuals. We therefore suggest considering an adaptation of diagnostic algorithms: when resources allow only one culture method to be performed, we recommend that MGIT liquid culture should be used exclusively in HIV-positive individuals as a first-line culture method, to reduce costs and make TB culture results accessible to more patients in resource-limited settings.

CD4 Tcells are rapidly depleted from tuberculosis granulomas following acute SIV co-infection.

Tuberculosis (TB) is a chronic inflammatory disease that is often associated with alterations in systemic and cellular metabolism that resolves following successful antimicrobial drug treatment. We hypothesized that altered systemic glucose metabolism as a consequence of Mycobacterium tuberculosis (Mtb) infection, contributes to TB pathogenesis, and when normalized with anti-glycemic drugs would improve clinical outcomes. To test this hypothesis, guinea pigs were treated daily with the anti-diabetic drug metformin starting 4 weeks prior or concurrent with aerosol exposure to the H37Rv strain of Mtb. In the chronic stages of infection, Mtb infected metformin-treated animals had restored systemic insulin sensitivity but remained glucose intolerant as determined by oral glucose tolerance testing. Despite persistent glucose intolerance, metformin-treated guinea pigs had a 2.8-fold reduction in lung lesion burden and a 0.7 log decrease in CFUs. An alternative hypothesis that metformin treatment improved clinical disease by having a direct effect on immune cell energy metabolism was tested using extracellular flux analysis and flow cytometry. The proinflammatory immune response to Mtb infection in untreated guinea pigs was associated with a marked increase in energy metabolism (glycolysis and mitochondrial respiration) of peripheral blood mononuclear cells (PBMCs), which was normalized in metformin-treated guinea pigs. Moreover, both CD4+ and CD8+ T lymphocytes from Mtb infected, metformin treated animals maintained a more normal mitochondrial membrane potential while those isolated from untreated animals had persistent mitochondrial hyperpolarization. These data suggest that metformin promotes natural host resistance to Mtb infection by maintaining immune cell metabolic homeostasis and function during the chronic stages of active TB disease.

Comorbidity between counnunicable and non-communicable diseases : the example of the dual burden of tuberculosis and diabetes in Dar es Salaam, Tanzania