Study of CD27, CD38, HLA-DR and Ki-67 immune profiles for the characterization of active tuberculosis, latent infection and end of treatment.

Background: Current blood-based diagnostic tools for TB are insufficient to properly characterize the distinct stages of TB, from the latent infection (LTBI) to its active form (aTB); nor can they assess treatment efficacy. Several immune cell biomarkers have been proposed as potential candidates for the development of improved diagnostic tools. Objective: To compare the capacity of CD27, HLA-DR, CD38 and Ki-67 markers to characterize LTBI, active TB and patients who ended treatment and resolved TB. Methods: Blood was collected from 45 patients defined according to clinical and microbiological criteria as: LTBI, aTB with less than 1 month of treatment and aTB after completing treatment. Peripheral blood mononuclear cells were stimulated with ESAT-6/CFP-10 or PPD antigens and acquired for flow cytometry after labelling with conjugated antibodies against CD3, CD4, CD8, CD27, IFN-γ, TNF-α, CD38, HLA-DR, and Ki-67. Conventional and multiparametric analyses were done with FlowJo and OMIQ, respectively. Results: The expression of CD27, CD38, HLA-DR and Ki-67 markers was analyzed in CD4+ T-cells producing IFN-γ and/or TNF-α cytokines after ESAT-6/CFP-10 or PPD stimulation. Within antigen-responsive CD4+ T-cells, CD27- and CD38+ (ESAT-6/CFP-10-specific), and HLA-DR+ and Ki-67+ (PPD- and ESAT-6/CFP-10-specific) populations were significantly increased in aTB compared to LTBI. Ki-67 demonstrated the best discriminative performance as evaluated by ROC analyses (AUC > 0.9 after PPD stimulation). Data also points to a significant change in the expression of CD38 (ESAT-6/CFP-10-specific) and Ki-67 (PPD- and ESAT-6/CFP-10-specific) after ending the anti-TB treatment regimen. Furthermore, ratio based on the CD27 median fluorescence intensity in CD4+ T-cells over Mtb-specific CD4+ T-cells showed a positive association with aTB over LTBI (ESAT-6/CFP-10-specific). Additionally, multiparametric FlowSOM analyses revealed an increase in CD27 cell clusters and a decrease in HLA-DR cell clusters within Mtb-specific populations after the end of treatment. Conclusion: Our study independently confirms that CD27-, CD38+, HLA-DR+ and Ki-67+ populations on Mtb-specific CD4+ T-cells are increased during active TB disease. Multiparametric analyses unbiasedly identify clusters based on CD27 or HLA-DR whose abundance can be related to treatment efficacy. Further studies are necessary to pinpoint the convergence between conventional and multiparametric approaches.

Tobacco Smoking and Second-Hand Smoke Exposure Impact on Tuberculosis in Children.

Little is known about whether second-hand smoke (SHS) exposure affects tuberculosis (TB). Here, we investigate the association of cigarette smoke exposure with active TB and latent TB infection (LTBI) in children, analyzing Interferon-Gamma Release Assays' (IGRAs) performance and cytokine immune responses. A total of 616 children from contact-tracing studies were included and classified regarding their smoking habits [unexposed, SHS, or smokers]. Risk factors for positive IGRAs, LTBI, and active TB were defined. GM-CSF, IFN-γ, IL-2, IL-5, IL-10, IL-13, IL-22, IL-17, TNF-α, IL-1RA and IP-10 cytokines were detected in a subgroup of patients. Being SHS exposed was associated with a positive IGRA [aOR (95% CI): 8.7 (5.9-12.8)] and was a main factor related with LTBI [aOR (95% CI): 7.57 (4.79-11.94)] and active TB [aOR (95% CI): 3.40 (1.45-7.98)]. Moreover, IGRAs' sensitivity was reduced in active TB patients exposed to tobacco. IL-22, GM-CSF, IL-5, TNF-α, IP-10, and IL-13 were less secreted in LTBI children exposed to SHS. In conclusion, SHS is associated with LTBI and active TB in children. In addition, false-negative IGRAs obtained on active TB patients exposed to SHS, together with the decrease of specific cytokines released, suggest that tobacco may alter the immune response.

Discovery and validation of an NMR-based metabolomic profile in urine as TB biomarker.

Despite efforts to improve tuberculosis (TB) detection, limitations in access, quality and timeliness of diagnostic services in low- and middle-income countries are challenging for current TB diagnostics. This study aimed to identify and characterise a metabolic profile of TB in urine by high-field nuclear magnetic resonance (NMR) spectrometry and assess whether the TB metabolic profile is also detected by a low-field benchtop NMR spectrometer. We included 189 patients with tuberculosis, 42 patients with pneumococcal pneumonia, 61 individuals infected with latent tuberculosis and 40 uninfected individuals. We acquired the urine spectra from high and low-field NMR. We characterised a TB metabolic fingerprint from the Principal Component Analysis. We developed a classification model from the Partial Least Squares-Discriminant Analysis and evaluated its performance. We identified a metabolic fingerprint of 31 chemical shift regions assigned to eight metabolites (aminoadipic acid, citrate, creatine, creatinine, glucose, mannitol, phenylalanine, and hippurate). The model developed using low-field NMR urine spectra correctly classified 87.32%, 85.21% and 100% of the TB patients compared to pneumococcal pneumonia patients, LTBI and uninfected individuals, respectively. The model validation correctly classified 84.10% of the TB patients. We have identified and characterised a metabolic profile of TB in urine from a high-field NMR spectrometer and have also detected it using a low-field benchtop NMR spectrometer. The models developed from the metabolic profile of TB identified by both NMR technologies were able to discriminate TB patients from the rest of the study groups and the results were not influenced by anti-TB treatment or TB location. This provides a new approach in the search for possible biomarkers for the diagnosis of TB.

Cribado de la tuberculosis latente y terapia biológica / Screening for Latent Tuberculosis and Biological Therapy

No disponible

Tratamiento de la infección tuberculosa latente en una unidad clínica de tuberculosis / Treatment of Latent Tuberculosis Infection in a Tuberculosis Clinic

No disponible

Assessment of the influence of direct tobacco smoke on infection and active TB management.

BACKGROUND: Smoking is a risk factor for tuberculosis (TB) infection and disease progression. Tobacco smoking increases susceptibility to TB in a variety of ways, one of which is due to a reduction of the IFN-γ response. Consequently, an impaired immune response could affect performance of IFN-γ Release Assays (IGRAs). OBJECTIVE: In the present study, we assess the impact of direct tobacco smoking on radiological manifestations, sputum conversion and immune response to Mycobacterium tuberculosis, analyzing IFN-γ secretion by IGRAs. METHODS: A total of 525 participants were studied: (i) 175 active pulmonary TB patients and (ii) 350 individuals coming from contact tracing studies, 41 of whom were secondary TB cases. Clinical, radiological and microbiological data were collected. T-SPOT.TB and QFN-G-IT were processed according manufacturer's instructions. RESULTS: In smoking patients with active TB, QFN-G-IT (34.4%) and T-SPOT.TB (19.5%) had high frequencies of negative results. In addition, by means of an unconditional logistic regression, smoking was a main factor associated with IGRAs' false-negative results (aOR: 3.35; 95%CI:1.47-7.61; p<0.05). Smoking patients with active TB presented a high probability of having cavitary lesions (aOR: 1.88; 95%CI:1.02-3.46;p<0.05). Mean culture negativization (months) ± standard deviation (SD) was higher in smokers than in non-smokers (2.47±1.3 versus 1.69±1.4). Latent TB infection (LTBI) was favored in smoking contacts, being a risk factor associated with infection (aOR: 11.57; 95%CI:5.97-22.41; p<0.00005). The IFN-γ response was significantly higher in non-smokers than in smokers. Smoking quantity and IFN-γ response analyzed by IGRAs were dose-dependent related. CONCLUSIONS: Smoking had a negative effect on radiological manifestations, delaying time of sputum conversion. Our data establish a link between tobacco smoking and TB due to a weakened IFN-γ response caused by direct tobacco smoke.

Factors associated with smoking among tuberculosis patients in Spain.

BACKGROUND: To determine the prevalence of smoking and analyze associated factors in a cohort of patients diagnosed with tuberculosis (TB) in Spain between 2006 and 2013. METHODS: Multicenter, cross-sectional, descriptive, observational study using a national database of TB patients, using logistic regression to calculate odds ratios (OR) and confidence intervals (CI). RESULTS: We analyzed 5,846 cases (62 % men, mean age 39 years, 33 % foreigners). 23.4 % were alcohol abuser, 1.3 % were injected drug users (IDU), 4.6 % were co-infected with HIV, and 7.5 % had a history of TB treatment. 6.6 % and 0.8 % showed resistance to one and multiple drugs, respectively. The predominant clinical presentation was pulmonary (71 %) with a cavitary radiological pattern in 32.8 % of cases. 82 % of cases were confirmed microbiologically, and 54 % were smear-positive microscopy. 2,300 (39.3 %) patients were smokers. The following factors were associated with smoking: male sex (OR = 2.26;CI:1.97;2.60), Spanish origin (OR = 2.79;CI:2.40-3.24), alcoholism (OR = 2.85;CI:2.46;3.31), IDU (OR = 2.78;CI:1.48;5.52), homelessness (OR = 1.99;CI:1.14-3.57), pulmonary TB (OR = 1.61;CI:1.16;2.24), cavitary radiological pattern (OR = 1.99;CI:1.43;2.79) and a smear-positive microscopy at the time of diagnosis (OR = 1.39;CI:1.14;1.17). CONCLUSIONS: The prevalence of smoking among TB patients is high. Smokers with TB have a distinct sociodemographic, clinical, radiological and microbiological profile to non-smokers.

Correlation between tuberculin skin test and IGRAs with risk factors for the spread of infection in close contacts with sputum smear positive in pulmonary tuberculosis.

BACKGROUND: The aim of the study was to assess the correlation between the tuberculin skin test (TST) and in vitro interferon-gamma released assays (IGRAs) with risk factors for the spread of infection in smear positive pulmonary tuberculosis (TB) contacts. METHODS: We recruited prospective contacts with smear positive pulmonary TB cases. We looked at human immunodeficiency virus (HIV) infection and other conditions of immunosuppression, presence of BCG vaccination and the degree of exposure to the index case. Patients underwent the TST, chest radiography, sputum analysis when necessary, and IGRA assays (QFN-G-IT and T-SPOT.TB). Presence of cough, diagnostic delay (days between first symptoms and TB diagnostic), contact conditions: room size (square meters) and index of overcrowding (square meters per person) were investigated in the index case. RESULTS: 156 contacts (119 adults, 37 children) of 66 TB patients were enrolled, 2.4 (1-14) contacts per TB case. The positivity of the TST did not correlate with the risk factors studied: presence of cough (p = 0.929); delayed diagnosis (p = 0.244); room size (p = 0.462); overcrowding (p = 0.800). Both QFN-G-IT and T-SPOT.TB, showed significant association with cough (p = 0.001, and p = 0.007) and room size (p = 0.020, and p = 0.023), respectively. CONCLUSIONS: Both IGRA associated better than TST with certain host-related risk factors involved in the transmission of disease, such as the presence of cough.