Close contact interferon-gamma response to the new PstS1(285-374):CPF10: a preliminary 1-year follow-up study

Abstract

Background: The available diagnostic tools for latent tuberculosis (TB) infection (LTBI) via interferon-gamma (IFN-g) release assays (IGRA) are based on ESAT6:CFP10 stimulation. However, the mycobacterial antigen PstS1 is also highly immunogenic and some of its fragments, such as PstS1_(285-374), have shown higher immunoreactivity in LTBI than in active TB. PstS1_(285-374), therefore, could increase the accuracy of the existing IGRA to detect LTBI. Thus, a new chimeric protein has recently been developed (PstS1_(285-374):CFP10) showing potential for LTBI screening of recent close contacts (rCt) exposed to Mycobacterium tuberculosis. The aim of this study was to analyze the PstS1_(285-374):CFP10 longitudinal IFN-g profile in comparison to ESAT6:CFP10 and full PstS1/CFP10 stimulation in a rCt cohort and correlate the responses to these in-house IGRA with any clinical changes/interventions that might occur.

Methods: A free-of-cost, one-year follow up was offered to 120 rCt recruited in Rio de Janeiro, RJ, Brazil. Whole blood short-term (WBA), long-term stimulation (LSA) assays, and the tuberculin skin test (TST) were performed during follow up.

Results: Among the enrolled rCt, 44.2% (53/120) returned for re-evaluation and the control group (TST negative, n = 17) showed low IFN-g reactivity to all antigen stimulations during the entire follow up, except for one participant who had shown radiological evidence of past TB/LTBI. Both incident cases were detected by IGRA-PstS1_(285-374):CFP10 during LTBI and after disease progression. Moreover, subsequent to the prophylactic treatment for LTBI (tLTBI), a significant regression in the LSA response was predominantly observed through stimulation of the new chimeric protein (8/10, 80%) followed by ESAT6:CFP10 (5/10, 50%) and PstS1/CFP10 (4/10, 40%). No clinical or epidemiological characteristics were exclusively shared among IGRA convertors.

Conclusion: It was demonstrated that the TST negative rCt without radiological evidence of LTBI/TB did not develop an IGRA-PstS1_(285-374):CFP10 response during the one-year follow up. Moreover, all incident cases were detected by our new IGRA; and a significant decrement of LSA-PstS1_(285-374):CFP10 reactivity post-prophylactic tLTBI was found. To our knowledge, this is the first study to monitor changes in the immune response profile of IGRA-PstS1_(285-374):CFP10 among rCt during a consecutive one-year period, thus providing additional evidence of its potential in the detection of LTBI.

Keywords: Follow up; Interferon-gamma; LTBI; M. tuberculosis; Tuberculosis.

Fig. 1

Baseline of study groups (T₀) and follow-up (T₁) characteristics. Recent close contact (rCt) screening via tuberculin skin test (TST), at T₀, and clinical outcomes after 1-year follow up (T₁) resulting in five different groups. All rCt-TST^pos (≥5 mm) were offered free-of-cost treatment for latent tuberculosis infection (tLBTI). A blood specimen was collected upon TST conversion or an active TB diagnosis

Fig. 2

Mean, standard deviation (SD) of interferon-gamma responses at enrollment (T₀) and after follow up (T₁). Longitudinal analysis via Wilcoxon signed rank test of mean interferon-gamma levels after short- (WBA) and/or long-term (LSA) stimulation assays priming blood cells from recent close contacts with mycobacterial antigens [ESAT6:CFP10, PstS1/CFP10, and PstS1(285–374):CFP10] upon enrollment (T₀) and after one-year follow up (T₁). In some special cases, a blood specimen was collected upon TST conversion or an active TB diagnosis. All individuals in the study were stratified according to their initial (T₀) tuberculin skin test (TST) response and T₁ outcome, as follows: a Group I TST^neg (n, WBA = 10, LSA = 17); b Group II TST convertors (n, WBA = 2, LSA = 4); c Group III TST^pos treated for latent tuberculosis infection (tLTBI) (n, WBA = 17, LSA = 25); d Group IV TST^pos not treated for tLTBI (n, WBA = 4, LSA = 5); and e Group V TB incident cases (n = 2). Short bars standard deviation

Fig. 3

Venn diagrams for the follow up of in-house interferon gamma release assay results. Distribution of interferon-gamma response conversion (a, b) or regression (c, d) at follow-up whole-blood (WBA) and long-term (LSA) stimulation assays. Recent close contact’s blood specimens were primed with Mycobacterium tuberculosis antigens (ESAT6:CFP10, PstS1/CFP10, and PstS1_(285–374):CFP10). Each symbol represents one rCt from the respective follow-up group (Black square Group I TST^neg; Black diamond Group II TST convertors; Double dagger Group III TST^pos treated for latent tuberculosis infection (tLTBI); White triangle Group IV TST^pos not tLTBI; Black circle Group V TB incident cases)

Fig. 4

Longitudinal interferon-gamma responses post-prophylactic treatment for latent tuberculosis infection. Longitudinal analysis upon enrollment (T₀) and after 1-year follow up (T₁) of mean interferon-gamma levels via short- (WBA) and long-term (LSA) stimulation assays with blood specimens from recent close contacts of TST^pos treated for latent tuberculosis infection (Black circle) and stratified according to their baseline interferon-gamma response (non-responders and responders) to each Mycobacterium tuberculosis antigen: a ESAT6:CFP10; b PstS1/CFP10; and c PstS1_(285–374):CFP10. *p < 0.05 via Wilcoxon signed rank test. White square recent close contact exposed to a multidrug-resistant Mycobacterium tuberculosis strain