The assessment of cytokines in Quantiferon supernatants for the diagnosis of latent TB infection in a tribal population of Melghat, India.

The tuberculin skin test (TST) and interferon-gamma release assays (IGRA), namely, the QuantiFERON-TB Gold test (QFT), remain the standard immunological diagnostic tools for latent tuberculosis (TB) infection (LTBI). However, the sub-optimal detection rates of both of these tests are major impediments in recognizing the population at risk. This study was aimed at evaluating additional cytokines besides interferon-gamma (IFN-γ) as biomarkers for improving LTBI diagnosis in the tribal population of Melghat, India. Seventy-four close TB contacts were stratified by QFT and TST results into: (i) QFT+/TST+ (n = 26), (ii) QFT+/TST- (n = 12), (iii) QFT-/TST- (n = 35) and (iv) QFT-/TST+ (n = 1) groups. A panel of cytokines (IL-6, IL-10, TNF-α and IL-2R) was then evaluated in antigen-stimulated QFT cell-free culture supernatants using IMMULITE-1000, an automated immunoassay analyzer. Cytokine estimation showed significantly higher levels of IL-6 in the QFT+/TST+ group, while significantly higher levels of IL-10 were found in the QFT-/TST- group. Correlation analysis identified a positive correlation between IL-6 and the QFT response (r = 0.6723, P < 0.0001), while a negative correlation was seen between QFT and IL-10 expression (r = -0.3271, P = 0.0044). Similarly, IL-6 was positively correlated with TST levels (r = 0.6631, P <0 .0001), and conversely, a negative correlation was found between TST and IL-10 expression (r = -0.5698, P < 0.0001). The positive and negative predictive values of IL-6 were found to be 92.59 and 93.33%, respectively, and the positive and negative predictive values of IL-10 were 96.55 and 91.18%, respectively. No significant impact of the demographic characteristics on cytokine positivity was observed. Our preliminary results suggest that the evaluation of additional cytokines in QFT cell-free culture supernatants may be valuable for the identification of LTBI. Combining IL-6 and IL-10 with QFT and/or TST could markedly improve the detection accuracy of LTBI. Our observations require investigation in larger well-characterized cohorts along with follow-up studies to further confirm the study outcome.

Latent TB infection diagnosis in population exposed to TB subjects in close and poor ventilated high TB endemic zone in India.

BACKGROUND: The present study was designed to investigate the utility of Quantiferon TB gold (QFT-G) and Tuberculin skin test (TST) for diagnosis of latent TB infection (LTBI) in high crowding TB endemic zone of Nagpur, India and their comparison with associated risk factors. METHODS: Out of 342 eligible participants, QFT-G and TST were performed in 162 participants. RESULTS: The prevalence of LTBI observed according to QFT-G and TST was 48% and 42% respectively, with an agreement of 52.47%. QFT-G positivity was associated with age while TST positivity was associated with body mass index (BMI). Duration of exposure emerged as a key risk factor significantly associated with both the tests. CONCLUSION: The prevalence of LTBI was quite high in the studied zone as detected by both the evaluated tests and thus, the combination of both the tests will be best predictive for LTBI in such high TB endemic regions.

Mycobacterial dormancy regulon protein Rv2623 as a novel biomarker for the diagnosis of latent and active tuberculous meningitis.

The present study was designed to investigate Rv2623 antigen, a major dormancy regulon protein of Mycobacterium tuberculosis (MTB) in CSF of suspected latent and active tuberculous meningitis (TBM) patients. A total of 100 CSF samples from TBM (n = 31), suspected latent TBM (n = 22), and suitable noninfectious control subjects (n = 47) were collected and evaluated for Rv2623 antigen level using ELISA protocol. A significantly high (P < 0.05) mean absorbance was observed in samples of suspected latent TBM and active TBM patients as compared to non-TBM control patients. However, no significant difference in Rv2623 level was observed between suspected latent TBM and TBM patients. Our preliminary findings suggest that Rv2623 may be useful as a potential biomarker for the diagnosis of the latent as well as active TBM infection. Futher evaluation of this biomarker in large number of samples is therefore needed to confirm the result.

Assessment of immune response to repeat stimulation with BCG vaccine using in vitro PBMC model.

BACKGROUND: Tuberculosis (TB) is one of the most prevalent cause of death due to a single pathogen. Bacillus Calmette Guérin (BCG) is the only vaccine available for clinical use that protects against miliary TB; however, this vaccine has shown variable levels of efficacy against pulmonary TB. In India, a single dose of BCG vaccine is given and there are few countries where repeated doses of BCG are given. The incidence of TB in India is very high inspite of primary vaccination in neonatal period and therefore requires consideration for repeated immunization. METHODS: To improve BCG immunogenicity, we have evaluated specific antimycobacterial immune responses (anti-BCG IgG and IFN-gamma), T cell activity-ADA, CD4 and CD8 T cell count, and CD4/CD8 ratio in a peripheral blood mononuclear cells (PBMC) model using boost immunization protocols with the BCG vaccine. PBMC were induced with a repeat dose of BCG at 24 and 72 hrs of cell culture. RESULTS: At the end of the experimental time, supernatant was collected to estimate anti-BCG IgG titer, interferon gamma, ADA activity, CD 4 and CD8 T cell count, and CD4/CD8 ratio. We demonstrated that PBMC induced with a repeat dose of BCG showed an increased specific anti-mycobacterial immune responses, T cell activity, and ADA activity as compared to PBMC induced with BCG alone or without BCG induction. CONCLUSION: The repeat BCG stimulation of PBMC obtained from BCG vaccinated individuals shows enhanced immune activation with respect to increased anti-BCG titre, IFN-gamma and ADA activity without concomitant increase in CD4 and CD8 cells. This study provides some basic data in future experiments in animal models with respect to repeat BCG vaccination.

Changes in cerebrospinal fluid cytokine expression in tuberculous meningitis patients with treatment.

BACKGROUND: The prevalence of tuberculous meningitis (TBM) is very high in developing areas of the world. Inflammation and cytokine patterns produced by T lymphocytes play an important role in susceptibility to infections. The inflammatory response and production of cytokines in the cerebrospinal fluid (CSF) of patients with TBM are well documented. Conversely, little is known about the role of pro- and anti-inflammatory cytokines in the CSF of TBM patients. The goal of the present study was to estimate the level of proinflammatory cytokine and anti-inflammatory cytokine levels in CSF samples from TBM patients. MATERIALS AND METHODS: To study this, in vivo levels of IL-2 and IFN-gamma (proinflammatory cytokines), and IL-10 (anti-inflammatory cytokine) in the CSF of 60 adult TBM patients and 50 age- and sex-matched non-TBM controls were measured. These cytokines were estimated in the CSF of TBM patients before and after starting treatment. RESULTS: High levels of proinflammatory cytokines as compared to anti-inflammatory cytokines were found in TBM patients before treatment. However, CSF samples from TBM patients after treatment showed elevated levels of anti-inflammatory and low levels of proinflammatory cytokines. CONCLUSIONS: We hypothesize that an increase in anti-inflammatory cytokines during treatment may indicate a favorable response.