Pregnancy Outcomes in Women Screened for Tuberculosis Infection in Swedish Antenatal Care.

BACKGROUND: Tuberculosis (TB) disease has been associated with pregnancy complications. However, the potential impact of TB infection (TBI) on pregnancy outcome is unknown. To investigate this, we conducted a register-based study in immigrant women screened with QuantiFERON assays for TBI in antenatal care in Sweden. METHODS: Women with history of immigration from TB-endemic countries were eligible for inclusion if national identification numbers and available QuantiFERON results obtained during pregnancy from 2014 to 2018 were available. QuantiFERON results were linked to data on maternal characteristics and pregnancy outcomes from the national Pregnancy and Patient Registers. TBI was defined as nil-corrected QuantiFERON result ≥0.35 IU/mL, in the absence of TB disease. Pregnancies in women with TB disease or human immunodeficiency virus were excluded, as were multiplex pregnancies, pregnancies resulting in miscarriage, and pregnancies occurring >10 years after immigration. Odds of defined adverse pregnancy outcomes were compared by maternal TBI status using mixed effects logistic regression with adjustment for maternal age and region of origin. RESULTS: In total, 7408 women with 12 443 pregnancies were included. In multivariable analysis, stillbirth (adjusted odds ratio [AOR], 1.90; 95% confidence interval [CI], 1.13-3.21; P = .016), severe preeclampsia (AOR, 1.62; 95% CI, 1.03-2.56; P = .036), low birthweight (<2500 g; AOR, 1.38; 95% CI, 1.01-1.88; P = .041), and emergency cesarean section (AOR, 1.28; 95% CI, 1.02-1.63; P = .033) were significantly associated with TBI. CONCLUSIONS: Among immigrant women seeking antenatal care in Sweden, TBI was independently associated with adverse pregnancy outcomes. Further studies are needed to corroborate these findings and to explore mechanisms involved.

Mechanisms of a Mycobacterium tuberculosis Active Peptide.

Multidrug-resistant tuberculosis (MDR) continues to pose a threat to public health. Previously, we identified a cationic host defense peptide with activity against Mycobacterium tuberculosis in vivo and with a bactericidal effect against MDR M. tuberculosis at therapeutic concentrations. To understand the mechanisms of this peptide, we investigated its interactions with live M. tuberculosis and liposomes as a model. Peptide interactions with M. tuberculosis inner membranes induced tube-shaped membranous structures and massive vesicle formation, thus leading to bubbling cell death and ghost cell formation. Liposomal studies revealed that peptide insertion into inner membranes induced changes in the peptides' secondary structure and that the membranes were pulled such that they aggregated without permeabilization, suggesting that the peptide has a strong inner membrane affinity. Finally, the peptide targeted essential proteins in M. tuberculosis, such as 60 kDa chaperonins and elongation factor Tu, that are involved in mycolic acid synthesis and protein folding, which had an impact on bacterial proliferation. The observed multifaceted targeting provides additional support for the therapeutic potential of this peptide.

Dynamics of Mycobacterium tuberculosis-Specific and Nonspecific Immune Responses in Women with Tuberculosis Infection during Pregnancy.

The immune control of tuberculosis (TB) infection could be influenced by pregnancy. To elucidate this, we longitudinally characterized Mycobacterium tuberculosis (Mtb)-specific and nonspecific immune responses in women during pregnancy and postpartum. HIV-uninfected women without past or current active TB, and with blood samples available from the 1st/2nd trimester, 3rd trimester, and 9 months postpartum, were identified at Ethiopian antenatal care clinics. Twenty-two TB+ women and 10 TB- women, defined according to Mtb-stimulated interferon-γ levels (≥0.35 and <0.20 IU/mL, respectively, in the Quantiferon-TB Gold-Plus assay), were included in the study. Longitudinal dynamics of six cytokines (IL-1ra, IL-2, IP-10, MCP-2, MCP-3, and TGF-ß1) were analyzed in supernatants from Mtb-stimulated and unstimulated whole blood. In TB+ women, Mtb-specific expression of IL-2 and IP-10 was higher at 3rd compared to 1st/2nd trimester (median 139 pg/mL versus 62 pg/mL, P = 0.006; 4,999 pg/mL versus 2,310 pg/mL, P = 0.031, respectively), whereas level of Mtb-triggered TGF-ß1 was lower at 3rd compared to 1st/2nd trimester (-6.8 ng/mL versus 2.3 ng/mL, P = 0.020). Unstimulated IL-2, IP-10, and MCP-2 levels were increased postpartum, compared with those noted during pregnancy, in TB+ women. Additionally, postpartum levels of proinflammatory cytokines in unstimulated blood were higher in TB+ women, than in TB- women. None of the women developed active TB during follow-up. Taken together, dynamic changes of Mtb-specific cytokine expression revealed during the 3rd trimester in TB+ women indicate increased Mtb-antigen stimulation at later stages of pregnancy. This could reflect elevated bacterial activity, albeit without transition to active TB, during pregnancy. IMPORTANCE Tuberculosis (TB) is globally one of the most common causes of death, and a quarter of the world's population is estimated to have TB infection. The risk of active TB is increased in connection to pregnancy, a phenomenon that could be due to physiological immune changes. Here, we studied the effect of pregnancy on immune responses triggered in HIV-uninfected women with TB infection, by analyzing blood samples obtained longitudinally during pregnancy and after childbirth. We found that the dynamics of Mtb-specific and nonspecific immune responses changed during pregnancy, especially in later stages of pregnancy, although none of the women followed in this study developed active TB. This suggests that incipient TB, with elevated bacterial activity, occurs during pregnancy, but progression of infection appears to be counteracted by Mtb-specific immune responses. Thus, this study sheds light on immune control of TB during pregnancy, which could be of importance for future intervention strategies.

Tuberculosis infection and stillbirth in Ethiopia-A prospective cohort study.

BACKGROUND: Tuberculosis is among the leading causes of death among infectious diseases. Regions with a high incidence of tuberculosis, such as sub-Saharan Africa, are disproportionately burdened by stillbirth and other pregnancy complications. Active tuberculosis increases the risk of pregnancy complications, but the association between latent tuberculosis infection (LTBI) and pregnancy outcomes is unknown. We explored the effect of latent tuberculosis infection on the risk of stillbirth in women attending antenatal care clinics in Ethiopia, a country with >170 000 annual cases of active tuberculosis. METHOD: Pregnant women were enrolled from antenatal care at three health facilities in Adama, Ethiopia, during 2015-2018, with assessment for previous and current active tuberculosis and testing for LTBI using QuantiFERON-TB-GOLD-PLUS. Proportions of stillbirth (≥ 20 weeks of gestation) and neonatal death (< 29 days of birth) were compared with respect to categories of maternal tuberculosis infection (tuberculosis-uninfected, LTBI, previous-, and current active tuberculosis). Multivariable logistic regression was performed for stillbirth. RESULTS: Among 1463 participants enrolled, the median age was 25 years, 10.2% were HIV-positive, 34.6% were primigravidae, and the median gestational age at inclusion was 18 weeks. Four (0.3%) were diagnosed with active tuberculosis during pregnancy, 68 (4.6%) reported previous treatment for active tuberculosis, 470 (32.1%) had LTBI, and 921 (63.0%) were tuberculosis-uninfected. Stillbirth was more frequent in participants with LTBI compared to tuberculosis-uninfected participants, although not reaching statistical significance (19/470, 4.0% vs 25/921, 2.7%, adjusted [for age, gravidity and HIV serostatus] odds ratio 1.38, 95% confidence interval 0.73-2.57, p = 0.30). Rates of neonatal death (5/470, 1.1% vs 10/921, 1.1%) were similar between these categories. CONCLUSION: Latent tuberculosis infection was not significantly associated with stillbirth or neonatal death in this cohort. Studies based on larger cohorts and with details on causes of stillbirth, as well as other pregnancy outcomes, are needed to further investigate this issue.

Longitudinal Mycobacterium tuberculosis-Specific Interferon Gamma Responses in Ethiopian HIV-Negative Women during Pregnancy and Postpartum.

Pregnancy may influence cellular immune responses to Mycobacterium tuberculosis. We investigated M. tuberculosis-specific interferon-γ responses in women followed longitudinally during pregnancy and postpartum. Interferon-γ levels (stimulated by M. tuberculosis antigens [TB1 and TB2] and mitogen included in the QuantiFERON-TB Gold Plus assay) were measured in blood from pregnant HIV-negative women identified from a prospective cohort at Ethiopian antenatal care clinics. Longitudinal comparisons included women without active tuberculosis (TB) with M. tuberculosis-triggered interferon-γ responses of ≥ 0.20 IU/ml, sampled on two and/or three occasions (1st/2nd trimester, 3rd trimester, and 9 months postpartum). Among 2,093 women in the source cohort, 363 met inclusion criteria for longitudinal comparisons of M. tuberculosis-stimulated interferon-γ responses. Median M. tuberculosis-triggered interferon-γ concentrations were higher at 3rd than those at the 1st/2nd trimester (in 38 women with samples available from these time points; TB1: 2.8 versus 1.6 IU/ml, P = 0.005; TB2: 3.3 versus 2.8 IU/ml, P = 0.03) and postpartum (in 49 women with samples available from these time points; TB1: 3.1 versus 2.2 IU/ml, P = 0.01; TB2: 3.1 versus 2.3 IU/ml, P = 0.03). In contrast, mitogen-stimulated interferon-γ levels were lower at 3rd than those at 1st/2nd trimester (in 32 women with samples available from these time points: 21.0 versus 34.9 IU/ml, P = 0.02). Results were similar in 22 women sampled on all 3 occasions. In HIV-negative women, M. tuberculosis-stimulated interferon-γ responses were higher during the 3rd trimester than those at earlier stages of pregnancy and postpartum, despite decreased mitogen-triggered responses. These findings suggest increased M. tuberculosis-specific cellular responses due to dynamic changes of latent TB infection during pregnancy.

A broad spectrum anti-bacterial peptide with an adjunct potential for tuberculosis chemotherapy.

Alternative ways to prevent and treat infectious diseases are needed. Previously, we identified a fungal peptide, NZX, that was comparable to rifampicin in lowering M. tuberculosis load in a murine tuberculosis (TB) infection model. Here we assessed the potential synergy between this cationic host defence peptide (CHDP) and the current TB drugs and analysed its pharmacokinetics. We found additive effect of this peptide with isoniazid and ethambutol and confirmed these results with ethambutol in a murine TB-model. In vivo, the peptide remained stable in circulation and preserved lung structure better than ethambutol alone. Antibiotic resistance studies did not induce mutants with reduced susceptibility to the peptide. We further observed that this peptide was effective against nontuberculous mycobacteria (NTM), such as M. avium and M. abscessus, and several Gram-positive bacteria, including methicillin-resistant Staphylococcus aureus. In conclusion, the presented data supports a role for this CHDP in the treatment of drug resistant organisms.

Tuberculosis Infection in Women of Reproductive Age: A Cross-sectional Study at Antenatal Care Clinics in an Ethiopian City.

BACKGROUND: Knowledge on tuberculosis (TB) infection epidemiology in women of reproductive age living in TB-endemic areas is limited. We used a composite definition of TB infection in a cohort of pregnant women recruited in an Ethiopian city as a model for TB exposure patterns, and to identify factors associated with TB infection. METHODS: Women seeking antenatal care at public health facilities underwent structured interviews, physical examination, and QuantiFERON-TB Gold-Plus (QFT) testing. Women with symptoms compatible with TB disease, and all human immunodeficiency virus (HIV)-positive women, were investigated for active TB by sputum bacteriological testing. TB infection (TB+) was defined as either positive QFT (≥ 0.35 IU/mL), self-reported previous active TB, or current active TB. Associations between TB infection and clinical, demographic, and socioeconomic characteristics were tested in multiple logistic regression analysis. RESULTS: Among 1834 participants, 679 (37.0%) met criteria for TB+ (80 [4.4%] previous active TB, 5 [0.3%] current active TB, and 594 [32.4%] QFT-positive without previous or current active TB). Age (annual adjusted odds ratio [AOR], 1.069 [95% confidence interval {CI}, 1.045-1.093]) and HIV infection (AOR, 1.43 [95% CI, 1.033-1.988]) were independently associated with TB+. The relationship with increasing age was only observed in HIV-negative women, and translated to an estimated annual risk of TB infection of 2.1% in HIV-negative women. CONCLUSIONS: TB infection in women of reproductive age in Ethiopia was independently associated with HIV infection and increasing age, suggesting exposure to contagious TB and continuous acquisition of TB infection in this population.

Alternative biomarkers for classification of latent tuberculosis infection status in pregnant women with borderline Quantiferon plus results.

Borderline interferon-gamma (IFN-γ) results (near the cut-off level 0.35 IU/ml) occur in QuantiFERON (QFT) assays. We investigated the performance of alternative biomarkers for classification of latent tuberculosis infection (LTBI) status in pregnant women with borderline QFT IFN-γ responses. Pregnant women (n = 96) were identified from a cohort study in Ethiopia, based on QFT-Plus IFN-γ results (QFT-low: <0.20 IU/ml, n = 33; QFT-borderline: 0.20-0.70 IU/ml, n = 31; QFT-high: >0.70 IU/ml, n = 32), including 12 HIV-positive individuals in each group and with 20 HIV-negative non-pregnant women from the same cohort with QFT IFN-γ <0.20 IU/ml as controls. Concentrations of 8 markers (IL-1ra, IL-6, IL-8, IP-10, MCP-1, MCP-2, osteopontin and resistin) were measured in whole blood QFT supernatants, stimulated separately with TB1 and TB2 antigens. K-nearest neighbor analysis (KNN) was used to classify participants with regard to likelihood of LTBI. Concentrations of MCP-2, IP-10 and IL-1ra were higher in QFT-borderline compared to QFT-low participants in both antigen stimulations (p < 0.001). KNN classification indicated high likelihood of LTBI in 13/31 (42%) women with QFT-borderline IFN-γ results. MCP-2, IP-10 and IL-1ra expressed in whole blood after TB antigen stimulation may be considered as alternative biomarkers for classification of LTBI status in pregnant women with borderline QFT IFN-γ results.

Transmission dynamics study of tuberculosis isolates with whole genome sequencing in southern Sweden.

Epidemiological contact tracing complemented with genotyping of clinical Mycobacterium tuberculosis isolates is important for understanding disease transmission. In Sweden, tuberculosis (TB) is mostly reported in migrant and homeless where epidemiologic contact tracing could pose a problem. This study compared epidemiologic linking with genotyping in a low burden country. Mycobacterium tuberculosis isolates (n = 93) collected at Scania University Hospital in Southern Sweden were analysed with the standard genotyping method mycobacterial interspersed repetitive units-variable number tandem repeats (MIRU-VNTR) and the results were compared with whole genome sequencing (WGS). Using a maximum of twelve single nucleotide polymorphisms (SNPs) as the upper threshold of genomic relatedness noted among hosts, we identified 18 clusters with WGS comprising 52 patients with overall pairwise genetic maximum distances ranging from zero to nine SNPs. MIRU-VNTR and WGS clustered the same isolates, although the distribution differed depending on MIRU-VNTR limitations. Both genotyping techniques identified clusters where epidemiologic linking was insufficient, although WGS had higher correlation with epidemiologic data. To summarize, WGS provided better resolution of transmission than MIRU-VNTR in a setting with low TB incidence. WGS predicted epidemiologic links better which could consolidate and correct the epidemiologically linked cases, avoiding thus false clustering.

A novel derivative of the fungal antimicrobial peptide plectasin is active against Mycobacterium tuberculosis.

Tuberculosis has been reaffirmed as the infectious disease causing most deaths in the world. Co-infection with HIV and the increase in multi-drug resistant Mycobacterium tuberculosis strains complicate treatment and increases mortality rates, making the development of new drugs an urgent priority. In this study we have identified a promising candidate by screening antimicrobial peptides for their capacity to inhibit mycobacterial growth. This non-toxic peptide, NZX, is capable of inhibiting both clinical strains of M. tuberculosis and an MDR strain at therapeutic concentrations. The therapeutic potential of NZX is further supported in vivo where NZX significantly lowered the bacterial load with only five days of treatment, comparable to rifampicin treatment over the same period. NZX possesses intracellular inhibitory capacity and co-localizes with intracellular bacteria in infected murine lungs. In conclusion, the data presented strongly supports the therapeutic potential of NZX in future anti-TB treatment.

Performance of QuantiFERON-TB Gold Plus for detection of latent tuberculosis infection in pregnant women living in a tuberculosis- and HIV-endemic setting.

We evaluated the performance of QuantiFERON-TB Gold Plus (QFT-Plus), which includes two Mycobacterium tuberculosis antigen formulations (TB1 and TB2), for detection of latent tuberculosis infection during pregnancy. Eight-hundred-twenty-nine Ethiopian pregnant women (5.9% HIV-positive) were tested with QFT-Plus, with bacteriological sputum analysis performed for women with clinically suspected tuberculosis and HIV-positive women irrespective of clinical presentation. QFT-Plus read-out was categorized according to the conventional cut-off (0.35 IU/ml) for both antigen formulations. In addition, we analysed the distribution of QFT-Plus results within a borderline zone (0.20-0.70 IU/ml), and interferon-γ response in relation to HIV infection and gestational age. Two-hundred-seventy-seven women (33%) were QFT-Plus-positive (HIV-positive 16/49 [33%]; HIV-negative 261/780 [33%]). There was a strong agreement between the two antigen formulations (κ = 0.92), with discordant results in 29 cases (3.5%). Whereas discordant QFT-Plus results were rare in pregnancy, several results with both TB1 and TB2 within the borderline range were observed (11/49 [22%] vs. 43/780 [5.5%] in HIV-positive and HIV-negative women, respectively; p<0.0001). HIV-positive women had lower absolute interferon-γ levels (TB1: 0.47 vs. 2.16 IU/ml; p<0.001, TB2: 0.49 vs. 2.24 IU/ml, p<0.001, considering results ≥0.20 IU/ml) compared to HIV-negative women. QFT-Plus-positive women who submitted samples at later stages of pregnancy had lower mitogen- (p<0.001) but higher TB-antigen-specific (p = 0.031 for TB1, p = 0.061 for TB2) interferon-γ response. Considering their lower capacity to produce TB-specific interferon-γ, a lower cut-off level for defining QFT-Plus-positivity may be considered in HIV-positive pregnant women.

What Is Resistance? Impact of Phenotypic versus Molecular Drug Resistance Testing on Therapy for Multi- and Extensively Drug-Resistant Tuberculosis.

Rapid and accurate drug susceptibility testing (DST) is essential for the treatment of multi- and extensively drug-resistant tuberculosis (M/XDR-TB). We compared the utility of genotypic DST assays with phenotypic DST (pDST) using Bactec 960 MGIT or Löwenstein-Jensen to construct M/XDR-TB treatment regimens for a cohort of 25 consecutive M/XDR-TB patients and 15 possible anti-TB drugs. Genotypic DST results from Cepheid GeneXpert MTB/RIF (Xpert) and line probe assays (LPAs; Hain GenoType MTBDRplus 2.0 and MTBDRsl 2.0) and whole-genome sequencing (WGS) were translated into individual algorithm-derived treatment regimens for each patient. We further analyzed if discrepancies between the various methods were due to flaws in the genotypic or phenotypic test using MIC results. Compared with pDST, the average agreement in the number of drugs prescribed in genotypic regimens ranged from just 49% (95% confidence interval [CI], 39 to 59%) for Xpert and 63% (95% CI, 56 to 70%) for LPAs to 93% (95% CI, 88 to 98%) for WGS. Only the WGS regimens did not contain any drugs to which pDST showed resistance. Importantly, MIC testing revealed that pDST likely underestimated the true rate of resistance for key drugs (rifampin, levofloxacin, moxifloxacin, and kanamycin) because critical concentrations (CCs) were too high. WGS can be used to rule in resistance even in M/XDR strains with complex resistance patterns, but pDST for some drugs is still needed to confirm susceptibility and construct the final regimens. Some CCs for pDST need to be reexamined to avoid systematic false-susceptible results in low-level resistant isolates.

Long-term Outcome of Antiretroviral Treatment in Patients With and Without Concomitant Tuberculosis Receiving Health Center-Based Care-Results From a Prospective Cohort Study.

BACKGROUND: In order to increase treatment coverage, antiretroviral treatment (ART) is provided through primary health care in low-income high-burden countries, where tuberculosis (TB) co-infection is common. We investigated the long-term outcome of health center-based ART, with regard to concomitant TB. METHODS: ART-naïve adults were included in a prospective cohort at Ethiopian health centers and followed for up to 4 years after starting ART. All participants were investigated for active TB at inclusion. The primary study outcomes were the impact of concomitant TB on all-cause mortality, loss to follow-up (LTFU), and lack of virological suppression (VS). Kaplan-Meier survival estimates and Cox proportional hazards models with multivariate adjustments were used. RESULTS: In total, 141/729 (19%) subjects had concomitant TB, 85% with bacteriological confirmation (median CD4 count TB, 169 cells/mm3; IQR, 99-265; non-TB, 194 cells/mm3; IQR, 122-275). During follow-up (median, 2.5 years), 60 (8%) died and 58 (8%) were LTFU. After ≥6 months of ART, 131/630 (21%) had lack of VS. Concomitant TB did not influence the rates of death, LTFU, or VS. Male gender and malnutrition were associated with higher risk of adverse outcomes. Regardless of TB co-infection status, even after 3 years of ART, two-thirds of participants had CD4 counts below 500 cells/mm3. CONCLUSIONS: Concomitant TB did not impact treatment outcomes in adults investigated for active TB before starting ART at Ethiopian health centers. However, one-third of patients had unsatisfactory long-term treatment outcomes and immunologic recovery was slow, illustrating the need for new interventions to optimize ART programs.

A borderline range for Quantiferon Gold In-Tube results.

OBJECTIVE: Interferon gamma release assays like Quantiferon Gold In-Tube (QFT) are used to identify individuals infected with Mycobacterium tuberculosis. A dichotomous cut-off (0.35 IU/ml) defines a positive QFT without considering test variability. Our objective was to evaluate the introduction of a borderline range under routine conditions. METHODS: Results of routine QFT samples from Sweden (2009-2014) were collected. A borderline range (0.20-0.99 IU/ml) was introduced in 2010 recommending a follow-up sample. The association between borderline results and incident active TB within 3 to 24 months was investigated through linkage with the national TB-register. RESULTS: Using the recommended QFT cut-off, 75.1% tests were negative, 21.4% positive and 3.5% indeterminate. In total, 9% (3656/40773) were within the borderline range. In follow-up samples, individuals with initial results between 0.20-0.34 IU/ml and 0.35-0.99 IU/ml displayed negative results below the borderline range (<0.20 IU/ml) in 66.1% (230/348) and 42.5% (285/671) respectively, and none developed incident TB. Among 6712 individuals with a positive initial test >0.99 IU/ml, 65 (0.97%) developed incident TB within 3-24 months. CONCLUSIONS: We recommend retesting of subjects with QFT results in the range 0.20-0.99 IU/ml to enhance reliability and validity of the test. Half of the subjects in the borderline range will be negative at a level <0.20 IU/ml when retested and have a very low risk of developing incident active TB.

Factors Associated with Early Mortality in HIV-Positive Men and Women Investigated for Tuberculosis at Ethiopian Health Centers.

INTRODUCTION: Despite increasing access to antiretroviral treatment (ART) in low-income countries, HIV-related mortality is high, especially in the first months following ART initiation. We aimed to evaluate the impact of TB coinfection on early mortality and to assess gender-specific predictors of mortality in a cohort of Ethiopian adults subjected to intensified casefinding for active TB before starting ART. MATERIAL AND METHODS: Prospectively recruited ART-eligible adults (n = 812, 58.6% female) at five Ethiopian health centers were followed for 6 months. At inclusion sputum culture, Xpert MTB/RIF, and smear microscopy were performed (158/812 [19.5%] had TB). Primary outcome was all-cause mortality. We used multivariate Cox models to identify predictors of mortality. RESULTS: In total, 37/812 (4.6%) participants died, 12 (32.4%) of whom had TB. Karnofsky performance score (KPS) and mid-upper arm circumference (MUAC) were associated with mortality in the whole population. However, the associations were different in men and women. In men, only MUAC remained associated with mortality (adjusted hazard ratio [aHR] 0.71 [95% CI 0.57-0.88]). In women, KPS <80% was associated with mortality (aHR 10.95 [95% CI 2.33-51.49]), as well as presence of cough (aHR 3.98 [95% CI 1.10-14.36]). Cough was also associated with mortality for TB cases (aHR 8.30 [95% CI 1.06-65.14]), but not for non-TB cases. CONCLUSIONS: In HIV-positive Ethiopian adults managed at health centers, mortality was associated with reduced performance score and malnutrition, with different distribution with regard to gender and TB coinfection. These robust variables could be used at clinic registration to identify persons at increased risk of early mortality.

ESX-1 exploits type I IFN-signalling to promote a regulatory macrophage phenotype refractory to IFNγ-mediated autophagy and growth restriction of intracellular mycobacteria.

The ability of macrophages to eradicate intracellular pathogens is normally greatly enhanced by IFNγ, a cytokine produced mainly after onset of adaptive immunity. However, adaptive immunity is unable to provide sterilizing immunity against mycobacteria, suggesting that mycobacteria have evolved virulence strategies to inhibit the bactericidal effect of IFNγ-signalling in macrophages. Still, the host-pathogen interactions and cellular mechanisms responsible for this feature have remained elusive. We demonstrate that the ESX-1 type VII secretion systems of Mycobacterium tuberculosis and Mycobacterium marinum exploit type I IFN-signalling to promote an IL-12(low) /IL-10(high) regulatory macrophage phenotype characterized by secretion of IL-10, IL-27 and IL-6. This mechanism had no impact on intracellular growth in the absence of IFNγ but suppressed IFNγ-mediated autophagy and growth restriction, indicating that the regulatory phenotype extends to function. The IFNγ-refractory phenotype was partly mediated by IL-27-signalling, establishing functional relevance for this downstream cytokine. These findings identify a novel macrophage-modulating function for the ESX-1 secretion system that may contribute to suppress the efficacy of adaptive immunity and provide mechanistic insight into the antagonistic cross talk between type I IFNs and IFNγ in mycobacterial infection.

Plasma Levels of Neopterin and C-Reactive Protein (CRP) in Tuberculosis (TB) with and without HIV Coinfection in Relation to CD4 Cell Count.

BACKGROUND: While the risk of TB is elevated in HIV-positive subjects with low CD4 cell counts, TB may in itself be associated with CD4 lymphocytopenia. We investigated markers of immune activation (neopterin) and inflammation (CRP) in TB patients with and without HIV coinfection and their association with CD4 cell levels, and determined their predictive capacity as alternative markers of advanced immunosuppression. METHODS: Participants selected from a cohort of adults with TB at Ethiopian health centers (195 HIV+/TB+, 170 HIV-/TB+) and 31 controls were tested for plasma levels of neopterin and CRP. Baseline levels of neopterin and CRP were correlated to CD4 cell count before and after anti-TB treatment (ATT). The performance to predict CD4 cell strata for both markers were investigated using receiver operating curves. RESULTS: Levels of both biomarkers were elevated in TB patients (neopterin: HIV+/TB+ 54 nmol/l, HIV-/TB+ 23 nmol/l, controls 3.8 nmol/l; CRP: HIV+/TB+ 36 µg/ml, HIV-/TB+ 33 µg/ml, controls 0.5 µg/ml). Neopterin levels were inversely correlated (-0.53, p<0.001) to CD4 cell count, whereas this correlation was weaker for CRP (-0.25, p<0.001). Neither of the markers had adequate predictive value for identification of subjects with CD4 cell count <100 cells/mm3 (area under the curve [AUC] 0.64 for neopterin, AUC 0.59 for CRP). CONCLUSION: Neopterin levels were high in adults with TB, both with and without HIV coinfection, with inverse correlation to CD4 cell count. This suggests that immune activation may be involved in TB-related CD4 lymphocytopenia. However, neither neopterin nor CRP showed promise as alternative tests for immunosuppression in patients coinfected with HIV and TB.

Screening for cryptococcal antigenemia using the lateral flow assay in antiretroviral therapy-naïve HIV-positive adults at an Ethiopian hospital clinic.

BACKGROUND: Since treatment for latent cryptococcal infection (CI) before starting antiretroviral therapy (ART) reduces mortality in HIV-infected subjects, screening for cryptococcal antigen (CrAg) in blood is recommended for individuals with CD4 cell counts < 100 cells/µL in regions with high CI prevalence. We assessed CrAg screening using the lateral flow assay in HIV-infected adults eligible for ART in central Ethiopia. RESULTS: HIV-positive patients (age ≥ 18 years, CD4 cell count < 350 cells/µL and/or WHO stage IV, no current or previous ART) were recruited at Adama Regional Hospital, Ethiopia (February 2013 until March 2014). CrAg was determined in plasma by lateral flow assay. Among 129 included participants (median age 35 years, 64 % female) the median CD4 cell count was 210 cells/µL (interquartile range 110-309); 29 (23 %) had CD4 cell count < 100 cells/µL. Two (1.6 %) participants were CrAg-positive (CD4 cell counts 171 vs. 250 cells/µL), one of whom had clinically manifest cryptococcal meningitis at the time of testing. CONCLUSIONS: In contrast to two recent reports from Ethiopia, we found few cases of CI among ART-naïve adults. Our study, which is the first using lateral flow assay for CrAg screening in this country, illustrates the need of larger surveys of CI prevalence among ART-naïve patients before defining recommendations on CI screening.

Impaired CXCR1-dependent oxidative defence in active tuberculosis patients.

Much of the pronounced host inflammatory response that occurs in tuberculosis (TB) is related to failed immunity against the invading pathogen. The G-protein coupled receptors CXCR1 and CXCR2 are implicated in important signal transduction pathways in lung inflammatory responses. We investigated the expression and function of these receptors in a simple whole blood model from 24 patients with pulmonary TB and in subjects with latent TB infection (LTBI). Healthy controls were recruited from close contacts to the pulmonary index patients. We found that pulmonary TB patients had significantly increased CXCR1 expression on blood cells compared to LTBI subjects and controls (p < 0.001). In contrast, LTBI subjects had a significant increase in CXCR2 expression compared to pulmonary TB patients (p < 0.001) and controls (p < 0.01). Leukocyte function, measured as oxidative capacity, was decreased in pulmonary TB patients compared to LTBI and controls (p < 0.001) and correlated with the increased CXCR1 expression. Leukocyte recruitment, measured as the expression of microRNA-223 was increased in pulmonary TB patients compared to LTBI (p < 0.05). We found that variations in receptor expression are linked to disease progression and affect the immune response against Mycobacterium tuberculosis (Mtb).