Prevalence of pulmonary tuberculosis and HIV infections and risk factors associated to tuberculosis in detained persons in Antananarivo, Madagascar.

The incidence rate of tuberculosis in prisons is estimated to be 8 times greater than that in the general population in Madagascar. Our objectives were to estimate the prevalence of pulmonary tuberculosis and HIV infection among prisoners and to identify risk factors associated with tuberculosis. We conducted a cross-sectional study at the central prison of Antananarivo from March to July 2021. Individual male and female inmates aged ≥ 13 years who had lived in the prison for at least three months prior to the study period were included as participants. Acid-fast bacilli detection by microscopy and/or culture, an intradermal tuberculin test, a chest X-ray, and a rapid diagnostic orientation test for HIV were performed. Among 748 participants, 4 (0.5%) were confirmed to have pulmonary tuberculosis. Overall, 14 (1.9%) patients had "confirmed" or "probable" tuberculosis [0.90-2.84, 95% CI]. The proportion of participants with latent tuberculosis infection was 69.6% (517/743) based on a positive tuberculin test without clinical symptoms or radiography images indicating tuberculosis. Out of 745 HIV screening tests, three showed reactive results (0.4%). Age (OR = 4.4, 95% CI [1.4-14.0]) and prior tuberculosis treatment (or episodes) were found to be associated with confirmed and probable tuberculosis.

C1q and HBHA-specific IL-13 levels as surrogate plasma biomarkers for monitoring tuberculosis treatment efficacy: a cross-sectional cohort study in Paraguay.

Introduction: New diagnostic tools are needed to rapidly assess the efficacy of pulmonary tuberculosis (PTB) treatment. The aim of this study was to evaluate several immune biomarkers in an observational and cross-sectional cohort study conducted in Paraguay. Methods: Thirty-two patients with clinically and microbiologically confirmed PTB were evaluated before starting treatment (T0), after 2 months of treatment (T1) and at the end of treatment (T2). At each timepoint plasma levels of IFN-y, 17 pro- and anti-inflammatory cytokines/chemokines and complement factors C1q, C3 and C4 were assessed in unstimulated and Mtb-specific stimulated whole blood samples using QuantiFERON-TB gold plus and recombinant Mycobacterium smegmatis heparin binding hemagglutinin (rmsHBHA) as stimulation antigen. Complete blood counts and liver enzyme assays were also evaluated and correlated with biomarker levels in plasma. Results: In unstimulated plasma, C1q (P<0.001), C4 (P<0.001), hemoglobin (P<0.001), lymphocyte proportion (P<0.001) and absolute white blood cell count (P=0.01) were significantly higher in PTB patients at baseline than in cured patients. C1q and C4 levels were found to be related to Mycobacterium tuberculosis load in sputum. Finally, a combinatorial analysis identified a plasma host signature comprising the detection of C1q and IL-13 levels in response to rmsHBHA as a tool differentiating PTB patients from cured TB profiles, with an AUC of 0.92 (sensitivity 94% and specificity 79%). Conclusion: This observational study provides new insights on host immune responses throughout anti-TB treatment and emphasizes the role of host C1q and HBHA-specific IL-13 response as surrogate plasma biomarkers for monitoring TB treatment efficacy.

Concordance of targeted and whole genome sequencing for Mycobacterium tuberculosis genotypic drug susceptibility testing.

Targeted Next Generation Sequencing (tNGS) and Whole Genome Sequencing (WGS) are increasingly used for genotypic drug susceptibility testing (gDST) of Mycobacterium tuberculosis. Thirty-two multi-drugs resistant and 40 drug susceptible isolates from Madagascar were tested with Deeplex® Myc-TB and WGS using the Mykrobe analysis pipeline. Sixty-four of 72 (89 %) yielded concordant categorical gDST results for drugs tested by both assays. Mykrobe didn't detect pncA K96T, pncA Q141P, pncA H51P, pncA H82R, rrs C517T and rpsL K43R mutations, which were identified as minority variants in corresponding isolates by tNGS. One discrepancy (rrs C517T) was associated with insufficient sequencing depth on WGS. Deeplex® Myc-TB didn't detect inhA G-154A which isn't covered by the assay's amplification targets. Despite those targets being included in the Deeplex® Myc-TB assay, a pncA T47A and a deletion in gid were not identified in one isolate respectively. The evaluated WGS and tNGS gDST assays show high but imperfect concordance.

Tuberculosis in Lemurs and a Fossa at National Zoo, Madagascar, 2022.

We diagnosed Mycobacterium tuberculosis in captive lemurs and a fossa in Antananarivo, Madagascar. We noted clinical signs in the animals and found characteristic lesions during necropsy. The source of infection remains unknown. Our results illustrate the potential for reverse zoonotic infections and intraspecies transmission of tuberculosis in captive wildlife.

A social network analysis model approach to understand tuberculosis transmission in remote rural Madagascar.

BACKGROUND: Quality surveillance data used to build tuberculosis (TB) transmission models are frequently unavailable and may overlook community intrinsic dynamics that impact TB transmission. Social network analysis (SNA) generates data on hyperlocal social-demographic structures that contribute to disease transmission. METHODS: We collected social contact data in five villages and built SNA-informed village-specific stochastic TB transmission models in remote Madagascar. A name-generator approach was used to elicit individual contact networks. Recruitment included confirmed TB patients, followed by snowball sampling of named contacts. Egocentric network data were aggregated into village-level networks. Network- and individual-level characteristics determining contact formation and structure were identified by fitting an exponential random graph model (ERGM), which formed the basis of the contact structure and model dynamics. Models were calibrated and used to evaluate WHO-recommended interventions and community resiliency to foreign TB introduction. RESULTS: Inter- and intra-village SNA showed variable degrees of interconnectivity, with transitivity (individual clustering) values of 0.16, 0.29, and 0.43. Active case finding and treatment yielded 67%-79% reduction in active TB disease prevalence and a 75% reduction in TB mortality in all village networks. Following hypothetical TB elimination and without specific interventions, networks A and B showed resilience to both active and latent TB reintroduction, while Network C, the village network with the highest transitivity, lacked resiliency to reintroduction and generated a TB prevalence of 2% and a TB mortality rate of 7.3% after introduction of one new contagious infection post hypothetical elimination. CONCLUSION: In remote Madagascar, SNA-informed models suggest that WHO-recommended interventions reduce TB disease (active TB) prevalence and mortality while TB infection (latent TB) burden remains high. Communities' resiliency to TB introduction decreases as their interconnectivity increases. "Top down" population level TB models would most likely miss this difference between small communities. SNA bridges large-scale population-based and hyper focused community-level TB modeling.

Improving measurement of tuberculosis care cascades to enhance people-centred care.

Care cascades represent the proportion of people reaching milestones in care for a disease and are widely used to track progress towards global targets for HIV and other diseases. Despite recent progress in estimating care cascades for tuberculosis (TB) disease, they have not been routinely applied at national and subnational levels, representing a lost opportunity for public health impact. As researchers who have estimated TB care cascades in high-incidence countries (India, Madagascar, Nigeria, Peru, South Africa, and Zambia), we describe the utility of care cascades and identify measurement challenges, including the lack of population-based disease burden data and electronic data capture, the under-reporting of people with TB navigating fragmented and privatised health systems, the heterogeneity of TB tests, and the lack of post-treatment follow-up. We outline an agenda for rectifying these gaps and argue that improving care cascade measurement is crucial to enhancing people-centred care and achieving the End TB goals.

Using a multiplex serological assay to estimate time since SARS-CoV-2 infection and past clinical presentation in malagasy patients.

Background: The world is facing a 2019 coronavirus (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In this context, efficient serological assays are needed to accurately describe the humoral responses against the virus. These tools could potentially provide temporal and clinical characteristics and are thus paramount in developing-countries lacking sufficient ongoing COVID-19 epidemic descriptions. Methods: We developed and validated a Luminex xMAP® multiplex serological assay targeting specific IgM and IgG antibodies against the SARS-CoV-2 Spike subunit 1 (S1), Spike subunit 2 (S2), Spike Receptor Binding Domain (RBD) and the Nucleocapsid protein (N). Blood samples collected periodically for 12 months from 43 patients diagnosed with COVID-19 in Madagascar were tested for these antibodies. A random forest algorithm was used to build a predictive model of time since infection and symptom presentation. Findings: The performance of the multiplex serological assay was evaluated for the detection of SARS-CoV-2 anti-IgG and anti-IgM antibodies. Both sensitivity and specificity were equal to 100% (89.85-100) for S1, RBD and N (S2 had a lower specificity = 95%) for IgG at day 14 after enrolment. This multiplex assay compared with two commercialized ELISA kits, showed a higher sensitivity. Principal Component Analysis was performed on serologic data to group patients according to time of sample collection and clinical presentations. The random forest algorithm built by this approach predicted symptom presentation and time since infection with an accuracy of 87.1% (95% CI = 70.17-96.37, p-value = 0.0016), and 80% (95% CI = 61.43-92.29, p-value = 0.0001) respectively. Interpretation: This study demonstrates that the statistical model predicts time since infection and previous symptom presentation using IgM and IgG response to SARS-CoV2. This tool may be useful for global surveillance, discriminating recent- and past- SARS-CoV-2 infection, and assessing disease severity. Fundings: This study was funded by the French Ministry for Europe and Foreign Affairs through the REPAIR COVID-19-Africa project coordinated by the Pasteur International Network association. WANTAI reagents were provided by WHO AFRO as part of a Sero-epidemiological "Unity" Study Grant/Award Number: 2020/1,019,828-0 P·O 202546047 and Initiative 5% grant n°AP-5PC-2018-03-RO.

Complexities and benefits of adopting next-generation sequencing-based tuberculosis diagnostics: a qualitative study among stakeholders in low and high-income countries.

OBJECTIVES: To clarify perceived benefits, barriers and facilitators of Mycobacterium tuberculosis next-generation sequencing implementation in Madagascar and Canada, towards informing implementation of this diagnostic technology in public health agencies and clinical settings in and beyond these settings. DESIGN: This qualitative study involved conducting semistructured interviews with key stakeholders engaged with next-generation sequencing implementation in Madagascar and Canada. Team-based descriptive analysis supported by Nvivo V.12.0 was used to identify key themes. SETTING: The study was conducted with participants involved at the clinical, diagnostic and surveillance levels of tuberculosis (TB) management from Madagascar and Canada. PARTICIPANTS: Eighteen participants were interviewed (nine Madagascar and nine Canada) and included individuals purposively sampled based on involvement with TB surveillance, laboratory diagnosis and clinical management. RESULTS: The following five themes emerged in the analysis of Malagasy and Canadian interviews: (1) heterogeneity in experience with established TB diagnostics, (2) variable understanding of new sequencing-based diagnostics potential; (3) further evidence as being key to expand adoption; (4) ethical arguments and concerns; (5) operational and system-level considerations. CONCLUSION: There persists important lack of familiarity with TB next-generation sequencing (TB NGS) applications among stakeholders in Canada and Madagascar. This translates into skepticism on the evidence underlying its use and its true potential value added within global public health systems. If deployed, TB NGS testing should be integrated with clinical and surveillance programmes. Although this is perceived as a priority, leadership and funding responsibilities for this integration to happen remains unclear to clinical, laboratory and public health stakeholders.

Back-to-Africa introductions of Mycobacterium tuberculosis as the main cause of tuberculosis in Dar es Salaam, Tanzania.

In settings with high tuberculosis (TB) endemicity, distinct genotypes of the Mycobacterium tuberculosis complex (MTBC) often differ in prevalence. However, the factors leading to these differences remain poorly understood. Here we studied the MTBC population in Dar es Salaam, Tanzania over a six-year period, using 1,082 unique patient-derived MTBC whole-genome sequences (WGS) and associated clinical data. We show that the TB epidemic in Dar es Salaam is dominated by multiple MTBC genotypes introduced to Tanzania from different parts of the world during the last 300 years. The most common MTBC genotypes deriving from these introductions exhibited differences in transmission rates and in the duration of the infectious period, but little differences in overall fitness, as measured by the effective reproductive number. Moreover, measures of disease severity and bacterial load indicated no differences in virulence between these genotypes during active TB. Instead, the combination of an early introduction and a high transmission rate accounted for the high prevalence of L3.1.1, the most dominant MTBC genotype in this setting. Yet, a longer co-existence with the host population did not always result in a higher transmission rate, suggesting that distinct life-history traits have evolved in the different MTBC genotypes. Taken together, our results point to bacterial factors as important determinants of the TB epidemic in Dar es Salaam.

Genomic Sequencing from Sputum for Tuberculosis Disease Diagnosis, Lineage Determination, and Drug Susceptibility Prediction.

Universal access to drug susceptibility testing for newly diagnosed tuberculosis patients is recommended. Access to culture-based diagnostics remains limited, and targeted molecular assays are vulnerable to emerging resistance mutations. Improved protocols for direct-from-sputum Mycobacterium tuberculosis sequencing would accelerate access to comprehensive drug susceptibility testing and molecular typing. We assessed a thermo-protection buffer-based direct-from-sample M. tuberculosis whole-genome sequencing protocol. We prospectively analyzed 60 acid-fast bacilli smear-positive clinical sputum samples in India and Madagascar. A diversity of semiquantitative smear positivity-level samples were included. Sequencing was performed using Illumina and MinION (monoplex and multiplex) technologies. We measured the impact of bacterial inoculum and sequencing platforms on genomic read depth, drug susceptibility prediction performance, and typing accuracy. M. tuberculosis was identified by direct sputum sequencing in 45/51 samples using Illumina, 34/38 were identified using MinION-monoplex sequencing, and 20/24 were identified using MinION-multiplex sequencing. The fraction of M. tuberculosis reads from MinION sequencing was lower than from Illumina, but monoplexing grade 3+ samples on MinION produced higher read depth than Illumina (P < 0.05) and MinION multiplexing (P < 0.01). No significant differences in sensitivity and specificity of drug susceptibility predictions were seen across sequencing modalities or within each technology when stratified by smear grade. Illumina sequencing from sputum accurately identified 1/8 (rifampin) and 6/12 (isoniazid) resistant samples, compared to 2/3 (rifampin) and 3/6 (isoniazid) accurately identified with Nanopore monoplex. Lineage agreement levels between direct and culture-based sequencing were 85% (MinION-monoplex), 88% (Illumina), and 100% (MinION-multiplex). M. tuberculosis direct-from-sample whole-genome sequencing remains challenging. Improved and affordable sample treatment protocols are needed prior to clinical deployment.

Evaluation of Nanopore sequencing for Mycobacterium tuberculosis drug susceptibility testing and outbreak investigation: a genomic analysis.

BACKGROUND: Mycobacterium tuberculosis whole-genome sequencing (WGS) has been widely used for genotypic drug susceptibility testing (DST) and outbreak investigation. For both applications, Illumina technology is used by most public health laboratories; however, Nanopore technology developed by Oxford Nanopore Technologies has not been thoroughly evaluated. The aim of this study was to determine whether Nanopore sequencing data can provide equivalent information to Illumina for transmission clustering and genotypic DST for M tuberculosis. METHODS: In this genomic analysis, we analysed 151 M tuberculosis isolates from Madagascar, South Africa, and England, which were collected between 2011 and 2018, using phenotypic DST and matched Illumina and Nanopore data. Illumina sequencing was done with the MiSeq, HiSeq 2500, or NextSeq500 platforms and Nanopore sequencing was done on the MinION or GridION platforms. Using highly reliable PacBio sequencing assemblies and pairwise distance correlation between Nanopore and Illumina data, we optimise Nanopore variant filters for detecting single-nucleotide polymorphisms (SNPs; using BCFtools software). We then used those SNPs to compare transmission clusters identified by Nanopore with the currently used UK Health Security Agency Illumina pipeline (COMPASS). We compared Illumina and Nanopore WGS-based DST predictions using the Mykrobe software and mutation catalogue. FINDINGS: The Nanopore BCFtools pipeline identified SNPs with a median precision of 99·3% (IQR 99·1-99·6) and recall of 90·2% (88·1-94·2) compared with a precision of 99·6% (99·4-99·7) and recall of 91·9% (87·6-98·6) using the Illumina COMPASS pipeline. Using a threshold of 12 SNPs for putative transmission clusters, Illumina identified 98 isolates as unrelated and 53 as belonging to 19 distinct clusters (size range 2-7). Nanopore reproduced 15 out of 19 clusters perfectly; two clusters were merged into one cluster, one cluster had a single sample missing, and one cluster had an additional sample adjoined. Illumina-based clusters were also closely replicated using a five SNP threshold and clustering accuracy was maintained using mixed Illumina and Nanopore datasets. Genotyping resistance variants with Nanopore was highly concordant with Illumina, having zero discordant SNPs across more than 3000 SNPs and four insertions or deletions (indels), across 60 000 indels. INTERPRETATION: Illumina and Nanopore technologies can be used independently or together by public health laboratories performing M tuberculosis genotypic DST and outbreak investigations. As a result, clinical and public health institutions making decisions on which sequencing technology to adopt for tuberculosis can base the choice on cost (which varies by country), batching, and turnaround time. FUNDING: Academy for Medical Sciences, Oxford Wellcome Institutional Strategic Support Fund, and the Swiss South Africa Joint Research Award (Swiss National Science Foundation and South African National Research Foundation).

Plasma host protein signatures correlating with Mycobacterium tuberculosis activity prior to and during antituberculosis treatment.

There is a need for rapid non-sputum-based tests to identify and treat patients infected with Mycobacterium tuberculosis (Mtb). The overall objective of this study was to measure and compare the expression of a selected panel of human plasma proteins in patients with active pulmonary tuberculosis (ATB) throughout anti-TB treatment (from baseline to the end of treatment), in Mtb-infected individuals (TBI) and healthy donors (HD) to identify a putative host-protein signature useful for both TB diagnosis and treatment monitoring. A panel of seven human host proteins CLEC3B, SELL, IGFBP3, IP10, CD14, ECM1 and C1Q were measured in the plasma isolated from an HIV-negative prospective cohort of 37 ATB, 24 TBI and 23 HD. The protein signatures were assessed using a Luminex xMAP® to quantify the plasmatic levels in unstimulated blood of the different clinical group as well as the protein levels at baseline and at three timepoints during the 6-months ATB treatment, to compare the plasma protein levels between culture slow and fast converters that may contribute to monitor the TB treatment outcome. Protein signatures were defined using the CombiROC algorithm and multivariate models. The studied plasma host proteins showed different levels between the clinical groups and during the TB treatment. Six of the plasma proteins (CLEC3B, SELL, IGFBP3, IP10, CD14 and C1Q) showed significant differences in normalised median fluorescence intensities when comparing ATB vs HD or TBI groups while ECM1 revealed a significant difference between fast and slow sputum culture converters after 2 months following treatment (p = 0.006). The expression of a four-host protein markers (CLEC3B-ECM1-IP10-SELL) was significantly different between ATB from HD or TBI groups (respectively, p < 0.05). The expression of the same signature was significantly different between the slow vs the fast sputum culture converters after 2 months of treatment (p < 0.05). The results suggest a promising 4 host-plasma marker signature that would be associated with both TB diagnostic and treatment monitoring.

Intensified tuberculosis treatment to reduce the mortality of HIV-infected and uninfected patients with tuberculosis meningitis (INTENSE-TBM): study protocol for a phase III randomized controlled trial.

BACKGROUND: Tuberculous meningitis (TBM) is the most lethal and disabling form of tuberculosis (TB), particularly in sub-Saharan Africa. Current anti-TB treatment is poorly effective since TBM mortality reaches 40% in HIV-negative patients and up to 70% in HIV-co-infected patients. To reduce TBM-induced morbidity and mortality, the INTENSE-TBM trial evaluates two interventions in both HIV-infected and uninfected patients: an anti-TB treatment intensification using oral high-dose rifampicin (35 mg/kg daily) and linezolid (1200 mg daily and then 600 mg daily) during the first 8 weeks of the anti-TB treatment and the use of adjunctive aspirin (200 mg daily). METHODS: This is a randomized controlled, phase III, multicenter, 2 × 2 factorial plan superiority trial. The trial has four arms, combining the two experimental treatments (intensified TBM regimen and aspirin) with the two reference treatments (WHO standard TB treatment and placebo), and is open-label for anti-TB treatment and double-blind placebo-controlled for aspirin treatment. This trial is conducted in adults or adolescents of age ≥15 years with TBM defined as "definite," "probable," or "possible" using Tuberculosis Meningitis International Research Consortium criteria, in four African countries: Ivory Coast, Madagascar, Uganda, and South Africa. The primary outcome is all-cause death between inclusion and week 40. DISCUSSION: The INTENSE-TBM trial represents a key opportunity to enhance TBM treatment with widely available existing drugs notably in high-incidence settings of both TB and HIV. The trial design is pragmatic and the results will permit early and effective applications in TBM patient care, in both HIV and TB high-incidence countries. TRIAL REGISTRATION: ClinicalTrials.gov NCT04145258. Registered on October 30, 2019.

Multi-country evaluation of RISK6, a 6-gene blood transcriptomic signature, for tuberculosis diagnosis and treatment monitoring.

There is a crucial need for non-sputum-based TB tests. Here, we evaluate the performance of RISK6, a human-blood transcriptomic signature, for TB screening, triage and treatment monitoring. RISK6 performance was also compared to that of two IGRAs: one based on RD1 antigens (QuantiFERON-TB Gold Plus, QFT-P, Qiagen) and one on recombinant M. tuberculosis HBHA expressed in Mycobacterium smegmatis (IGRA-rmsHBHA). In this multicenter prospective nested case-control study conducted in Bangladesh, Georgia, Lebanon and Madagascar, adult non-immunocompromised patients with bacteriologically confirmed active pulmonary TB (ATB), latent TB infection (LTBI) and healthy donors (HD) were enrolled. ATB patients were followed-up during and after treatment. Blood RISK6 scores were assessed using quantitative real-time PCR and evaluated by area under the receiver-operating characteristic curve (ROC AUC). RISK6 performance to discriminate ATB from HD reached an AUC of 0.94 (95% CI 0.89-0.99), with 90.9% sensitivity and 87.8% specificity, thus achieving the minimal WHO target product profile for a non-sputum-based TB screening test. Besides, RISK6 yielded an AUC of 0.93 (95% CI 0.85-1) with 90.9% sensitivity and 88.5% specificity for discriminating ATB from LTBI. Moreover, RISK6 showed higher performance (AUC 0.90, 95% CI 0.85-0.94) than IGRA-rmsHBHA (AUC 0.75, 95% CI 0.69-0.82) to differentiate TB infection stages. Finally, RISK6 signature scores significantly decreased after 2 months of TB treatment and continued to decrease gradually until the end of treatment reaching scores obtained in HD. We confirmed the performance of RISK6 signature as a triage TB test and its utility for treatment monitoring.

Local adaptation in populations of Mycobacterium tuberculosis endemic to the Indian Ocean Rim.

Background: Lineage 1 (L1) and 3 (L3) are two lineages of the Mycobacterium tuberculosis complex (MTBC) causing tuberculosis (TB) in humans. L1 and L3 are prevalent around the rim of the Indian Ocean, the region that accounts for most of the world's new TB cases. Despite their relevance for this region, L1 and L3 remain understudied. Methods: We analyzed 2,938 L1 and 2,030 L3 whole genome sequences originating from 69 countries. We reconstructed the evolutionary history of these two lineages and identified genes under positive selection. Results: We found a strongly asymmetric pattern of migration from South Asia toward neighboring regions, highlighting the historical role of South Asia in the dispersion of L1 and L3. Moreover, we found that several genes were under positive selection, including genes involved in virulence and resistance to antibiotics. For L1 we identified signatures of local adaptation at the esxH locus, a gene coding for a secreted effector that targets the human endosomal sorting complex, and is included in several vaccine candidates. Conclusions: Our study highlights the importance of genetic diversity in the MTBC, and sheds new light on two of the most important MTBC lineages affecting humans.

Drug-Resistant Tuberculosis in Pet Ring-Tailed Lemur, Madagascar.

We diagnosed tuberculosis in an illegally wild-captured pet ring-tailed lemur manifesting lethargy, anorexia, and cervical lymphadenopathy. Whole-genome sequencing confirmed the Mycobacterium tuberculosis isolate belonged to lineage 3 and harbored streptomycin resistance. We recommend reverse zoonosis prevention and determination of whether lemurs are able to maintain M. tuberculosis infection.

Different PPD-stimulated cytokine responses from patients infected with genetically distinct Mycobacterium tuberculosis complex lineages.

OBJECTIVES: The genetic diversity of Mycobacterium tuberculosis complex (MTBC) influences the immune response of the host, which may affect the immunodiagnostic tests and biomarker assessment studies used for tuberculosis (TB). This study aimed to determine whether the mycobacterial-antigen-stimulated cytokine responses vary with the genotype of the MTBC infecting the patient. METHODS: Eighty-one patients with confirmed active pulmonary TB were recruited, and MTBC clinical strains were isolated from their sputum for bacterial lineage single-nucleotide polymorphism typing. Whole blood was drawn from the patients to measure the purified protein derivative (PPD)-stimulated cytokine responses (GM-CSF, IFN-γ, IL-1ß, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-10, TNF-α, IFN-α, IL-12, eotaxin, IL-13, IL-15, IL-17, MIP1-α, MIP1-ß, MCP1, IL1RA, IP10, IL2R, MIG) with the Luminex multiplex immunoassay. RESULTS: Of the 24 cytokines studied, three were produced differentially in whole blood dependent on the infecting lineage of MTBC. Decreased production of IL-17 was observed in patients infected with modern lineages compared with patients infected with ancestral lineages (P < 0.01), and production of IFN-γ and IL-2 was significantly decreased in patients infected with lineage 4 strains compared with patients infected with lineage 3 strains (P < 0.05). CONCLUSION: MTBC strains belonging to lineage 4 induced a decreased whole-blood PPD-stimulated pro-inflammatory cytokine response.

Longitudinal Variations of M. tuberculosis-Induced IFN-γ Responses in HIV-Negative Pregnant Women Exposed to Tuberculosis.

Introduction: Pregnancy triggers an alteration of the immune functions and increases the risk of developing the active tuberculosis (TB) symptoms in exposed women. The effect of pregnancy on the Mycobacterium tuberculosis-specific immune responses used for most of the TB immunodiagnostic assays is not well documented. Here we investigated the changes in the M. tuberculosis-specific IFN-γ production in age-matched pregnant and non-pregnant women according to their TB exposition status. Methods: We conducted a prospective cohort study on HIV-seronegative pregnant and non-pregnant women with compatible pulmonary TB symptoms addressed to TB healthcare facilities in Antananarivo, Madagascar. Active pulmonary TB was bacteriologically assessed with culture from sputum samples. Clinical data and blood samples were collected at inclusion and after 6 months of follow-up for each individual included. Whole blood samples were stimulated with QuantiFERON TB-Gold Plus (QFT-P) assay antigens. Plasma IFN-γ concentrations were then assessed by ELISA. Results: A total of 284 women were investigated for the study including 209 pregnant women without confirmed TB (pNTB), 24 pregnant women with bacteriologically confirmed active TB (pATB), 16 non-pregnant women with active TB (ATB), and 35 non-pregnant healthy donors (HC). At inclusion, IFN-γ responses are lower in the pregnant women compared to their age-matched non-pregnant counterparts and independently of their TB status. Among the pregnant women, higher concentrations of M. tuberculosis-specific IFN-γ were observed in those exposed to TB, but with a lower magnitude in the active TB compared to the latently infected pregnant women (p < 0.05 with TB1 and p < 0.01 with TB2). After 6 months of follow-up, the M. tuberculosis-specific IFN-γ responses return to their baseline concentrations except for the pregnant women treated for TB for which none of the QFT-P positive reversed to negative (0%, 0/10) at the end of their TB treatment. Conclusion: These results support the concept of specific immune priorities characterized by a concomitant reduction in inflammatory immunity during pregnancy and corroborate the important role of activating the M. tuberculosis-specific immune responses to control the infection when the pregnant women are exposed to the pathogen.

Latent tuberculosis infection prevalence in rural Madagascar.

BACKGROUND: Understanding latent Mycobacterium tuberculosis infection (LTBI) prevalence is crucial for the design of TB control strategies. There are no data on LTBI in rural Madagascar. METHODS: Tuberculin skin tests were performed in 98 adults aged >15 y in five rural villages in the Ifanadiana district, Madagascar. RESULTS: Of adults, 78.6% were positive for LTBI, ranging between 28.6% and 95.0% among villages. The majority (65.3%) showed an induration reaction of >15 mm. CONCLUSIONS: LTBI prevalence is high in rural Madagascar. Long-term TB control strategies including LTBI testing and treatment must account for high and heterogeneous prevalence in remote, underdeveloped areas.