Abstract
BackgroundA challenge in tuberculosis (TB) research is to develop a new immunological test that can help distinguish, among subjects responsive to QuantiFERON TB Gold In tube (QFT-IT), those who are able to control Mtb replication (remote LTBI, recent infection and past TB) from those who cannot (active TB disease). IFN-γ response to the Heparin-binding-hemagglutinin (HBHA) of Mtb has been associated with LTBI, but the cumbersome procedures of purifying the methylated and immunological active form of the protein from Mtb or M. bovis Bacillus Calmette et Guerin (BCG) have prevented its implementation in a diagnostic test. Therefore, the aim of the present study was to evaluate the IFN-γ response to methylated HBHA of Mtb produced in M. smegmatis (rHBHAms) in individuals at different stages of TB who scored positive to QFT-IT.Methodology/Principal Findings87 individuals at different stages of TB who scored positive to QFT-IT were selected. IFN-γ response to in vitro whole blood stimulation with rHBHAms was evaluated by short-term and long-term tests and detected by ELISA or flow cytometry. We demonstrated that the IFN-γ response to rHBHAms is mediated by CD4+ T-cells with an effector-memory phenotype. This response, evaluated by short-term-tests, is significantly lower in active TB than in remote LTBI (p = 0.0010) and past TB (p = 0.0152). These results were confirmed by long-term tests. The qualitative data confirmed that IFN-γ responses higher than the cut-off point identified by ROC analysis are associated with the status of non-active disease.ConclusionsIn this study we show that the T-cell response to a recombinant and methylated HBHA of Mtb produced in M. smegmatis is useful to discriminate between active and non-active TB disease among those responsive to QFT-IT in a whole blood system. Further studies are needed to improve the accuracy of the assay.
Highlights
Tuberculosis (TB) remains a major global health problem and is one of the leading causes of morbidity and mortality due to infection [1].The identification of biomarkers of protection and disease may be helpful for a better understanding of TB pathogenesis and eventually for diagnostic purposes
In this study we show that the T-cell response to a recombinant and methylated HBHA of Mycobacterium tuberculosis (Mtb) produced in M. smegmatis is useful to discriminate between active and non-active TB disease among those responsive to QuantiFERON TB Gold In tube (QFT-IT) in a whole blood system
The introduction of T-cell-based interferon (IFN)-c release assays (IGRAs), using antigens belonging to Mycobacterium tuberculosis (Mtb) region of difference (RD)1 [including early secreted antigenic target (ESAT)-6 and culture filtrate protein 10 (CFP)-10], have made a significant step towards improved latent TB infection (LTBI) diagnosis [4]
Summary
Tuberculosis (TB) remains a major global health problem and is one of the leading causes of morbidity and mortality due to infection (www.who.int/tb/publications/global_report/en/) [1].The identification of biomarkers of protection and disease may be helpful for a better understanding of TB pathogenesis and eventually for diagnostic purposes. The introduction of T-cell-based interferon (IFN)-c release assays (IGRAs), using antigens belonging to Mycobacterium tuberculosis (Mtb) region of difference (RD)1 [including early secreted antigenic target (ESAT)-6 and culture filtrate protein 10 (CFP)-10], have made a significant step towards improved LTBI diagnosis [4]. These tests do not discriminate between active disease, remote LTBI, recent infection and past cured TB [5,6,7,8]. The aim of the present study was to evaluate the IFN-c response to methylated HBHA of Mtb produced in M. smegmatis (rHBHAms) in individuals at different stages of TB who scored positive to QFT-IT
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