Potential Role for IL-2 ELISpot in Differentiating Recent and Remote Infection in Tuberculosis Contact Tracing

et al. (2010) Potential Role for IL-2 ELISpot in Differentiating Recent and Remote Infection in Tuberculosis Contact Tracing. PLoS ONE 5(7): e11670. doi:10.1371/journal.pone.0011670 Potential Role for IL-2 ELISpot in Differentiating Recent and Remote Infection in Tuberculosis Contact Tracing Christoph Lange 0 Ben Marais, University of Stellenbosch, South Africa 0 1 Division of Clinical Infectious Diseases, Research Center Borstel , Borstel, Germany , 2 Division of Immune Cell Analytics, Research Center Borstel , Borstel, Germany , 3 Division of Microbial Interface Biology, Research Center Borstel , Borstel, Germany, 4 TB Surveillance , Public Health Department , Lu beck, Germany, 5 Autoimmun Diagnostika GmbH, Straberg, Germany , 6 Hygiene and Preventive Medicine Institute, University of Sassari , Sassari , Italy Interferon (IFN)-c release assays (IGRA) have improved tuberculosis contact tracing, but discrimination of recent from remote Mycobacterium tuberculosis contacts is not possible by IGRA alone. We present results of a tuberculosis contact investigation with a new early-secretory-antigenic-target (ESAT)-6 and culture-filtrate-protein (CFP)-10 specific interleukin (IL)-2 ELISpot in addition to ESAT-6 and CFP-10 specific IFN-c ELISpot and tuberculin skin testing (TST). Results of the TST, IFN-c ELISpot and IL-2 ELISpot were positive in 6/172 (3.4%), 7/167 (4.2%) and 6/196 (3.1%) of contacts, respectively. Close contact ($100 hours) to the index case increased the risk of positive results in the IFN-c ELISpot, TST, and IL-2 ELISpot by 40.8, 19.3, and 2.5 times, respectively. Individuals with a positive IFN-c ELISpot/negative IL-2 ELISpot result had a median (IQR) duration of index case exposure of 568 hours (133_1000) compared to individuals with a positive IFN-c ELISpot/ positive IL-2 ELISpot result (median = 24 hours; 20_130; p-value = 0.047). Combination of a M. tuberculosis specific IFN-c ELISpot with a M. tuberculosis specific IL-2 ELISpot significantly improved the identification of individuals with the highest risk of recent M. tuberculosis infection and is a promising method that should be explored to target tuberculosis preventive chemotherapy. - Funding: The study was funded by the Research Center Borstel, Leibniz Center for Medicine and Biomedical Research, Germany. ELISPOT plates and reagents were kindly provided free of charge by Autoimmun Diagnostika (AID), Strasberg, Germany. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: ELISPOT plates and reagents were kindly provided free of charge by Autoimmun Diagnostika (AID), Strasberg, Germany. One of the coauthors, V. Schoellhorn is the CEO of AID. However, AID does not hold any patents in the antigens used in this study (ESAT-6 or CFP-10 ). The affiliation of V. Schoellhorn with AID does not alter the authors adherence to all the PLoS ONE policies on sharing data and materials. . These authors contributed equally to this work. Identification and treatment of individuals with lasting Mycobacterium tuberculosis specific immune responses (latent tuberculosis infection -, LTBI) following recent contact to a contagious index case with pulmonary tuberculosis are crucial to tuberculosis control [1]. The diagnosis and management of LTBI has evolved over the last decade with improvements in the immunodiagnosis by M. tuberculosis specific interferon-c release assays (IGRA) in addition to the tuberculin skin test [2]. In principle, IGRA detect interferon-c release by mononuclear cells in the ELISpot (T-SPOT.TB Oxford Immunotec, Abingdon, UK) or in whole blood in the ELISA (QuantiFERON-TB GOLD in tube; QFT-G-IT; Cellestis, Carnegie, Australia) following ex vivo contact with antigens that are naturally encoded in the M. tuberculosis genomic region of difference (RD)-1 and RD11 (only QFT-G-IT). The genes encoding for these proteins are not present in most non tuberculous mycobacteria and are deleted from the genome of M. bovis Bacille Calmette Guerin (BCG) strains that are used for tuberculosis vaccinations [3,4]. The population of T lymphocytes responding rapidly with the production of IFN-c following ex vivo stimulation with these antigens belongs predominantly to the pool of CD4+ CD45RA-CCR7- effector memory cells that likely had recent antigen exposure [5]. In contrast, long-lived central memory T-cells may persist even after successful treatment of tuberculosis [6,7] and are less likely to release IFN-c after short incubation with antigen while they are more likely to produce IL-2 upon antigen exposure than effector cells [7]. Thus, assaying M. tuberculosis-specific T cell function defined solely by the quantification of IFN-c secretion after short term ex vivo incubation with M. tuberculosis-specific antigen may prove to be an insufficient indicator for recent inhalation exposure to M. tuberculosis. In contrast, simultaneous measures of other T cell function, such as M. tuberculosis-specific stimulation of IL-2 secretion may improve the discrimination of active versus remote LTBI [7]. A 28-year-old schoolteacher was referred to our hospital for diagnostic evaluations for presumptive tuberculosis. Between March and October 2007 he had consulted seven different general practitioners complaining of progressive cough and weight loss until finally in November 2007 a chest x-ray was performed and a cavitating lesion in the left upper lobe of the lung was demonstrated. In our hospital, alcohol acid fast bacilli (AAFB) were readily identified on sputum smears. The bacilli were identified as M. tuberculosis by culture and susceptibility testing revealed antibiotic drug resistance to isoniacid and streptomycin. We estimated by the presence of his symptoms that he had been infectious for a period of seven months prior to hospital admission. During this period, he had exposure to a large number of close contacts including pupils and teachers from two different schools and members of a church parish. We describe the results of tuberculosis contact tracing among well-defined contacts of a highly symptomatic sputum smear positive index case, comparing a novel M. tuberculosis antigen specific IL-2 ELISpot assay with a M. tuberculosis antigen specific IFN-c ELISpot and the TST for the diagnosis of LTBI. This outbreak provided an opportunity to test the hypothesis that simultaneous measures of M. tuberculosis-specific IFN-c and IL-2 secretion may improve identification of individuals with the highest risk of recent infection with M. tuberculosis. Materials and Methods Study population Individuals with close contact to the symptomatic index case during the period of March 2007 until October 2007 were identified by the municipal health care department of the city of Luebeck, Germany. Contacts were asked to estimate their exposure time to the index case in hours per week. We identified a group of close (with a total exposu (...truncated)