Mycobacterium tuberculosis latency-associated antigen Rv1733c SLP improves the accuracy of differential diagnosis of active tuberculosis and latent tuberculosis infection

Abstract

Background:Differential diagnosis of active tuberculosis (ATB) and latent tuberculosis infection (LTBI) has been a challenge for clinicians in high TB burden countries. The purpose of this study was to improve the accuracy of differential diagnosis of ATB and LTBI by using fluorescent immunospot (FluoroSpot) assay to detect specific Th1 cell immune responses. The novel mycobacterium tuberculosis (MTB) latency-associated antigens Rv1733c and synthetic long peptides derived from Rv1733c (Rv1733c SLP) were used based on virulence factors early secreting antigen target-6 (ESAT-6) and culture filtrate protein-10 (CFP-10).Methods:Fifty-seven ATB cases, including 20 pathogen-confirmed ATB and 37 clinically diagnosed ATB, and 36 LTBI cases, were enrolled between January and December 2017. FluoroSpot assay was used to detect the interferon γ (IFN-γ) and interleukin 2 (IL-2) secreted by the specific T cells after being stimulated with MTB virulence factors ESAT-6 and CFP-10, MTB latency-associated antigens Rv1733c and Rv1733c SLP. The receiver operating characteristic (ROC) curve was used to define the best cutoff value of latency-associated antigens in the use of differentiating ATB and LTBI. The sensitivity, specificity, predictive value, and likelihood ratio of ESAT-6 and CFP-10-FluoroSpot combined with latency-associated antigen in the differential diagnosis of ATB and LTBI were also calculated.Results:Following the stimulation with Rv1733c and Rv1733c SLP, the frequency of single IL-2-secreting T cells stimulated by Rv1733c SLP had the largest area under the ROC curve, which was 0.766. With a cutoff value of 1 (spot-forming cells [SFCs]/2.5 × 105 peripheral blood mononuclear cells) for frequency, the sensitivity and specificity of distinguishing ATB from LTBI were 72.2% and 73.7%, respectively. ESAT-6 and CFP-10-FluoroSpot detected the frequency and proportion of single IFN-γ-secreting T cells; the sensitivity and specificity of distinguishing ATB from LTBI were 82.5% and 66.7%, respectively. Combined with the frequency of single IL-2-secreting T cells stimulated by Rv1733c SLP on the basis of ESAT-6 and CFP-10-FluoroSpot, the sensitivity and specificity increased to 84.2% and 83.3%, respectively.Conclusion:Rv1733c SLP, combined with ESAT-6 and CFP-10, might be used as a candidate antigen for T cell-based tuberculosis diagnostic tests to differentiate ATB from LTBI.

Latent Tuberculosis Diagnostics: A Systematic Review What is the past, present, and future in the diagnosis of latent tuberculosis?

“…All studies in table five have been assessed for sensitivity and specificity (if provided). Study four (22) uses FluoroSpot in testing both IFN-γ and IL-2 levels after stimulation from ESAT-6, CFP-10 separately and together or in combination with Rv1733c SLP.…”

“…Study four (22) examined the use of FluoroSpot with Rv1733c, ESAT-6, and CFP-10. Sensitivity and specificity were found to be 84.2% and 83.3% respectively.…”

“…However, there is still an overlap in cytokine responses, which may inevitably reduce the accuracy of differential diagnosis. Study four (22) also adopted a case–control study design and probably overestimated the accuracy of the differential diagnosis as mentioned by the researchers as well. In addition, 70 Th1 cytokines have been identified in relation to tuberculosis (42).…”

“…All studies in table five have been assessed for sensitivity and specificity (if provided). Study four (22) uses FluoroSpot in testing both IFN-γ and IL-2 levels after stimulation from ESAT-6, CFP-10 separately and together or in combination with Rv1733c SLP. Study seven (25) examined; 25 cytokines under three different conditions 11 of the 25 cytokines had promising results and could distinguish LTBI from ATB.…”

Challenges and the Way forward in Diagnosis and Treatment of Tuberculosis Infection

“…A study by Serra-Vidal et al (2014) on 60 latency-related Ags also showed that Rv1773 was the most immunogenic protein that can distinguish between non-infected, TBI, and ATB patients based on the IFN-γ response after 18 h of stimulation, with higher responses in TBI [ 206 ]. Another study on stimulation with synthetic long peptides derived from Rv1773c Ag (Rv1773c SLP), IL-2 secreting T-cells were significantly higher in TBI compared to ATB, but no significant results were obtained with IFN-γ secreting T-cells [ 208 ].…”