Basirudeen Syed Ahamed Kabeer

Role of QuantiFERON-TB Gold, Interferon Gamma Inducible Protein-10 and Tuberculin Skin Test in Active Tuberculosis Diagnosis

Background The measurement of Interferon gamma or Interferon gamma inducible protein (IP)-10 in antigen stimulated blood samples is suggested as an alternative method for latent tuberculosis (TB) diagnosis. Nonetheless, their role in active TB diagnosis, particularly in TB endemic settings is yet to be defined. In this study, the sensitivities and specificities of Interferon gamma release assay (IGRA), IP-10 assay and tuberculin skin test (TST) in detecting active TB cases were assessed in human immunodeficiency virus (HIV) sero-negative TB patients and healthy controls respectively. Methods/Principal Findings A total of 177 adult TB patients and 100 healthy controls were included for this study. QuantiFERON-TB Gold In-tube (QFT-IT) method was used to analyze the sensitivity and specificity of IGRA. QFT-IT, IP-10 and TST yielded the diagnostic sensitivities of 90.6% (95%CI: 86.3%–94.9%), 92.5% (95%CI: 88.6%–96.4%) and 68.9% (95%CI: 60.6%–77.2%) and specificities of 55% (95% CI: 35.2%–54.8%), 48% (95% CI: 38.2%–57.8%) and 75.5% (95% CI: 66.8%–84.2%), respectively. The extent of pulmonary involvement or presence of diabetes mellitus did not appear to influence the sensitivities of any of these tests. The combination of any of the two tests among QFT-IT, IP-10 and TST showed >98% sensitivity among smear negative cases and particularly the combination of IP-10, TST and smear microscopy showed 100% sensitivity, however, the specificity was decreased to 44.8%. Conclusions/Significance QFT-IT and IP-10 were highly sensitive in detecting active TB cases. The combination with TST improved the sensitivity of QFT-IT and IP-10 significantly. Although the higher sensitivity of combination of QFT-IT/IP-10 and TST may be useful in active TB diagnosis, they are limited by their poor specificity due to the high prevalence of latent TB in our settings.

Is IP-10 an accurate marker for detecting M. tuberculosis-specific response in HIV-infected persons?

Background The suboptimal sensitivity of Interferon (IFN)-γ-based in-vitro assays, especially in immunocompromised individuals, emphasizes the need for alternative markers for diagnosing tuberculosis (TB). The objective of this study was to evaluate whether interferon-inducible protein (IP)-10, monocyte chemotactic protein (MCP)-2 and interleukin (IL)-2 can be useful biomarkers for evaluating a specific response to RD1 antigens associated to active TB disease in HIV-infected individuals. Methodology/Principal Findings The study was carried out in India, the country with the highest TB burden in the world. Sixty-six HIV-infected individuals were prospectively enrolled, 28 with active-pulmonary-TB and 38 without. The whole blood assay based on RD1-selected peptides (experimental test) and QuantiFERON-TB Gold In tube (QFT-IT) was performed. Plasma was harvested at day-1-post-culture and soluble factors were evaluated by ELISA. The results indicate that by detecting IP-10, the sensitivity of the experimental test and QFT-antigen (75% and 85.7% respectively) for active TB was higher compared to the same assays based on IFN-γ (42.9% and 60.7% respectively) and was not influenced by the ability to respond to the mitogen. By detecting IP-10, the specificity of the experimental test and QFT-antigen (57.9% and 13.2% respectively) for active TB was lower than what was reported for the same assays using IFN-γ-detection (78.9% and 68.4% respectively). On the other side, in vitro IL-2 and MCP-2 responses were not significantly associated with active TB. Conclusions HIV infection does not impair RD1-specific response detected by IP-10, while it significantly decreases IFN-γ-mediated responses. At the moment it is unclear whether higher detection is related to higher sensitivity or lower specificity of the assay. Further studies in high and low TB endemic countries are needed to elucidate this.

Role of interferon gamma release assay in active TB diagnosis among HIV infected individuals.

Background A rapid and specific test is urgently needed for tuberculosis (TB) diagnosis especially among human immunodeficiency virus (HIV) infected individuals. In this study, we assessed the sensitivity of Interferon gamma release assay (IGRA) in active tuberculosis patients who were positive for HIV infection and compared it with that of tuberculin skin test (TST). Methodology/Principal Findings A total of 105 HIV-TB patients who were naïve for anti tuberculosis and anti retroviral therapy were included for this study out of which 53 (50%) were culture positive. Of 105 tested, QuantiFERON-TB Gold in-tube (QFT-G) was positive in 65% (95% CI: 56% to 74%), negative in 18% (95% CI: 11% to 25%) and indeterminate in 17% (95% CI: 10% to 24%) of patients. The sensitivity of QFT-G remained similar in pulmonary TB and extra-pulmonary TB patients. The QFT-G positivity was not affected by low CD4 count, but it often gave indeterminate results especially in individuals with CD4 count <200 cells/µl. All of the QFT-G indeterminate patients whose sputum culture were positive, showed ≤0.25 IU/ml of IFN-γ response to phytohemagglutinin (PHA). TST was performed in all the 105 patients and yielded the sensitivity of 31% (95% CI: 40% to 22%). All the TST positives were QFT-G positives. The sensitivity of TST was decreased, when CD4 cell counts declined. Conclusions/Significance Our study shows neither QFT-G alone or in combination with TST can be used to exclude the suspicion of active TB disease. However, unlike TST, QFT-G yielded fewer false negative results even in individuals with low CD4 count. The low PHA cut-off point for indeterminate results suggested in this study (≤0.25 IU/ml) may improve the proportion of valid QFT-G results.

IP-10 response to RD1 antigens might be a useful biomarker for monitoring tuberculosis therapy

BackgroundThere is an urgent need of prognosis markers for tuberculosis (TB) to improve treatment strategies. The results of several studies show that the Interferon (IFN)-γ-specific response to the TB antigens of the QuantiFERON TB Gold (QFT-IT antigens) decreases after successful TB therapy. The objective of this study was to evaluate whether there are factors other than IFN-γ [such as IFN-γ inducible protein (IP)-10 which has also been associated with TB] in response to QFT-IT antigens that can be used as biomarkers for monitoring TB treatment.MethodsIn this exploratory study we assessed the changes in IP-10 secretion in response to QFT-IT antigens and RD1 peptides selected by computational analysis in 17 patients with active TB at the time of diagnosis and after 6 months of treatment. The IFN-γ response to QFT-IT antigens and RD1 selected peptides was evaluated as a control. A non-parametric Wilcoxon signed-rank test for paired comparisons was used to compare the continuous variables at the time of diagnosis and at therapy completion. A Chi-square test was used to compare proportions.ResultsWe did not observe significant IP-10 changes in whole blood from either NIL or QFT-IT antigen tubes, after 1-day stimulation, between baseline and therapy completion (p = 0.08 and p = 0.7 respectively). Conversely, the level of IP-10 release to RD1 selected peptides was significantly different (p = 0.006). Similar results were obtained when we detected the IFN-γ in response to the QFT-IT antigens (p = 0.06) and RD1 selected peptides (p = 0.0003). The proportion of the IP-10 responders to the QFT-IT antigens did not significantly change between baseline and therapy completion (p = 0.6), whereas it significantly changed in response to RD1 selected peptides (p = 0.002). The proportion of IFN-γ responders between baseline and therapy completion was not significant for QFT-IT antigens (p = 0.2), whereas it was significant for the RD1 selected peptides (p = 0.002), confirming previous observations.ConclusionsOur preliminary study provides an interesting hypothesis: IP-10 response to RD1 selected peptides (similar to IFN-γ) might be a useful biomarker for monitoring therapy efficacy in patients with active TB. However, further studies in larger cohorts are needed to confirm the consistency of these study results.

IFN-γ, but not IP-10, MCP-2 or IL-2 response to RD1 selected peptides associates to active tuberculosis

OBJECTIVES To evaluate whether in vitro response to Mycobacterium tuberculosis RD1 peptides selected by computational analysis, measured by IFN-gamma, IP-10, MCP-2 or IL-2 production, is associated with active tuberculosis (TB) in a country with a high incidence of TB. METHODS 129 individuals were prospectively enrolled, 41 with active-pulmonary TB and 88 without (household contacts and community controls). A whole blood assay based on RD1 selected peptides was performed. Soluble factors were evaluated by ELISA in plasma harvested at day1-post-culture. Enrolled individuals were also tested by QuantiFERON TB-Gold In tube (QFT-IT) and tuberculin skin tests (TST). RESULTS IFN-gamma response to RD1 selected peptides was significantly higher in active TB patients than in household contacts and community controls. IP-10 and MCP-2 response did not differ between active TB patients and household contacts, although it was higher in these groups compared to community controls; conversely IL-2 response did not differ among the three groups. When IFN-gamma response to RD1 selected peptides was scored based on receiver-operator-characteristic analysis, active TB was predicted with 68% sensitivity and 86% specificity. QFT-IT and TST showed a sensitivity for active TB of 90% and 68% and a specificity of 58% and 59%, respectively. CONCLUSIONS IFN-gamma (but not IP-10, MCP-2 and IL-2) response to RD1 selected peptides is associated with active TB with a higher specificity than QFT-IT and TST.

Interferon gamma and interferon gamma inducible protein-10 in detecting tuberculosis infection

OBJECTIVE This study aimed to compare the levels of TB-antigen specific Interferon gamma (IFN-γ) and IFN-γ inducible protein (IP)-10 in culture of whole blood samples from healthy controls (HC) and healthy household contacts (HHC). METHODOLOGY A total of 386 study subjects, which included 186 HC and 200 HHC, were recruited. QuantiFERON-TB Gold in-tube (QFT-IT) assay was employed to measure IFN-γ levels. IP-10 levels were measured in the supernatants collected from QFT-IT tubes. Tuberculin skin test was also performed. RESULTS The levels of TB antigen specific IFN-γ and IP-10 were significantly higher in HHC compared to HC. There was no significant difference observed between positivity of QFT-IT and IP-10 in HC and HHC. The positivity of TST was significantly lower in subjects <17 year, when compared to IP-10 (p<0.005). The reduced cut-off point 0.22IU/ml significantly increased the positivity of QFT-IT among children with high risk for latent TB infection (LTBI). CONCLUSIONS Measurement of TB antigen specific IFN-γ and IP-10 can be potential markers for the detection of LTBI.

Comparison of interferon gamma–inducible protein-10 and interferon gamma–based QuantiFERON TB Gold assays with tuberculin skin test in HIV-infected subjects

We aimed to compare the positivity of the QuantiFERON TB gold in-tube (QFT-IT antigens) specific interferon gamma (IFN-γ/QFT-IT) and IFN-γ-inducible protein-10 (IP-10/QFT-IT) assays with tuberculin skin test (TST) among human immunodeficiency virus (HIV)-infected individuals in a TB endemic setting. A total of 180 HIV-infected subjects, with no evidence of active TB, were recruited. IFN-γ and IP-10 levels specific to QFT-IT antigens were measured in plasma from QFT-IT tubes. The overall positivity of TST at the 5-mm cut-off point (19%) was significantly lower when compared to IFN-γ/QFT-IT (38%) and IP-10/QFT-IT (45%) assays. The positivity of IP-10/QFT-IT was significantly higher than that of IFN-γ/QFT-IT (P = 0.038). Indeterminate results for IFN-γ/QFT-IT and IP-10/QFT-IT were more frequent in subjects with CD4 count <100 cells/μL than in those with >100 cells/μL. IFN-γ/QFT-IT (9%) yielded significantly higher number of indeterminate results than IP-10/QFT-IT (5%). The frequency of these responses is higher than the proportion of individuals with positive TST results. However, 6 IFN-γ/QFT-IT- or IP-10/QFT-IT-negative subjects were positive for TST at the 5-mm cut-off point. Prospective and prognostic studies are required to clarify the significance of these data.

A Toolbox for Tuberculosis (TB) Diagnosis: An Indian Multicentric Study (2006-2008). Evaluation of QuantiFERON-TB Gold in Tube for TB Diagnosis

Background The aim of this multicentric prospective study in India was to assess the performance of the QuantiFERON TB-Gold in tube (QFT-GIT), Tuberculin Skin Test (TST) and microbiological results as additional tools for diagnosing active tuberculosis (TB) and latent infection (LTBI) according to Human Immunodeficiency Virus (HIV) status. Methods Individuals with and without active TB and HIV infection were enrolled between 2006–2008. QFT-GIT and TST results were analyzed per se and in combination with microbiological data. Results Among the 276 individuals (96 active pulmonary TB and 180 no active TB) tested by QFT-GIT, 18 indeterminate results (6.5%) were found, more significantly numerous in the HIV-infected (15/92; 16.3%) than the HIV-uninfected (3/184; 1.6%)(p<0.0001). QFT-GIT sensitivity for active TB was 82.3% and 92.9% respectively after including or excluding indeterminate results. Clinical sensitivity was significantly lower in the HIV-infected (68.4%) than the HIV-uninfected (91.4%) patients (p = 0.0059). LTBI was detected in 49.3% of subjects without active TB but varied according to TB exposure. When the TST and QFT-GIT were concomitantly performed, the respective sensitivity for active TB diagnosis was 95.0% and 85.0% in the HIV-uninfected (p = 0.60), and 66.7% and 51.5% in the HIV-infected patients (p = 0.32). QFT-GIT and TST respective specificity for active TB in the HIV-uninfected was 25.0% and 57.1% (p = 0.028), and 64.8% and 83.3% in the HIV-infected (p = 0.047). In those with active TB, QFT-GIT results were not associated with microbiological parameters (smear grade, liquid culture status, time-to-positivity of culture) or clinical suspicion of active TB score (provided by the clinicians at enrollment). Combining microbiological tests with both immunological tests significantly increased sensitivity for active TB diagnosis (p = 0.0002), especially in the HIV-infected individuals (p = 0.0016). Conclusion QFT-GIT and TST have similar diagnostic value for active TB diagnosis. In HIV-infected patients, combining microbiological tests with both immunological tests significantly increases the sensitivity for active TB diagnosis.

A toolbox for tuberculosis diagnosis: An indian multicentric study (2006-2008): Microbiological results

Background The aim of this multicentric prospective study in India was to assess the value of several microbiological tools that contribute to the diagnosis of tuberculosis (TB) according to HIV status. Methods Standard microbiological tools on individual specimens were analyzed. Results Among the 807 patients with active TB, 131 were HIV-infected, 316 HIV-uninfected and 360 had HIV-unknown status. Among the 980 non-active TB subjects, 559 were at low risk and 421 were at high risk of M. tuberculosis (Mtb) exposure. Sensitivity of smear microscopy (SM) was significantly lower in HIV-infected (42.2%) than HIV-uninfected (75.9%) (p = 0.0001) and HIV-unknown pulmonary TB patients (61.4%) (p = 0.004). Specificity was 94.5% in non-TB patients and 100% in health care workers (HCW) and healthy family contacts. Automated liquid culture has significantly higher diagnostic performances than solid culture, measured by sensitivity (74.7% vs. 55.9%) (p = 0.0001) and shorter median time to detection (TTD) (12.0 vs. 34.0 days) (p = 0.0001). Specificity was 100% in HCW and cured-TB patients, but was lower in non-TB patients (89%) due to isolation of Mycobacteria other than tuberculosis (MOTT). TTD by both methods was related to AFB score. Contamination rate was low (1.4%). AccuProbe hybridization technique detected Mtb in almost all culture-positive specimens, but MOTT were found in 4.7% with a significantly higher frequency in HIV-infected (15%) than HIV-uninfected TB patients (0.5%) (p = 0.0007). Pre-test classification significantly increased the diagnostic value of all microbiological tests in pulmonary TB patients (p<0.0001) but to a lesser degree in extrapulmonary TB patients. Conclusions Conventional microbiological tools led to results similar to those already described in India special features for HIV-infected TB patients included lower detection by SM and culture. New microbiological assays, such as the automated liquid culture system, showed increased accuracy and speed of detection.

Increased Frequency of Antigen-Specific Polyfunctional T Cells in Tuberculosis Patients

This study assessed the polyfunctional T cells in healthy household contacts (HHCs) and TB patients. This study also assessed the memory subsets responsible for the secretion of IFN-γ during the short-term culture with Mycobacterium tuberculosis antigens. Frequencies of CD4