Evaluation of gamma interferon release assays using Mycobacterium tuberculosis antigens for diagnosis of latent and active tuberculosis in Mycobacterium bovis BCG-vaccinated populations

Abstract

T-cell-based gamma interferon (IFN-γ) release assays (IGRAs) using Mycobacterium tuberculosis-specific antigens have shown higher sensitivity and specificity than the routine tuberculin skin test (TST). However, the effects of Mycobacterium bovis BCG vaccination and anti-tuberculosis (TB) treatment on dynamic T-cell responses to M. tuberculosis-specific antigens in active TB cases have rarely been investigated in regions where TB is endemic. Eighty-nine patients with active pulmonary TB (ATB) and 57 healthy controls (HC) from China were recruited and tested by sputum smear and culture, TSTs, and IGRAs with M. tuberculosis-specific antigens ESAT-6 and CFP-10 (T-SPOT.TB) as well as purified protein derivative (PPD) stimulation. All 146 participants were screened by the T-SPOT.TB assay at recruitment. T-SPOT.TB-positive rates in ATB and HC groups were 87.6% (78/89) and 21.1% (12/57), respectively. Of 38 ATB patients who were both TST and T-SPOT.TB tested, the positive rates were 73.7% (28/38) and 94.7% (36/38), respectively (P = 0.0215), and those in the HC group were 62.3% (33/53) and 18.9% (10/53), respectively (P < 0.0001). The T-SPOT.TB-positive rates declined during TB treatment and were 94.4% (51/54), 86.4% (19/22), and 61.5% (8/13) for ATB patients receiving 0- to 1-month, 1- to 3-month, and 3- to 6-month anti-TB treatment, respectively. The IGRA is a most promising test for both active TB and latent TB infection (LTBI) diagnosis due to the improvement of its specificity and convenience, especially in the Mycobacterium bovis BCG-vaccinated population. Furthermore, the T-SPOT.TB assay using ESAT-6 and CFP-10 in ATB patients during anti-TB treatment could serve as a potential predictor of therapeutic efficacy.

FIG. 1.

Flow chart showing numbers recruited in active tuberculosis patients and healthy control individuals. mo, month(s); TST, tuberculin skin test; T-SPOT.TB, commercial IGRA from Oxford Immunotec, Abingdon, United Kingdom.

FIG. 2.

The positive rates of T-SPOT.TB assay and TST in groups of ATB and HC. There were 38 ATB patients and 53 healthy individuals simultaneously screened by T-SPOT.TB assay and TST. *, P = 0.0215, and **, P = 0.000 comparing the positive rates of these two tests in groups of ATB and HC.

FIG. 3.

The T-SPOT.TB positive rates in groups of ATB and HC, stratified by TST induration diameters (mm). T-SPOT.TB assay and TST were simultaneously performed on 38 ATB patients and 53 healthy individuals. The location of the bubble represents the T-SPOT.TB positive rate. The size of the bubble represents the percentage of subjects with positive results. The positive rate for ATB patients with indurations of <5 mm, 5 to 10 mm, and >10 mm were 26% (10/38), 18% (7/38), and 55% (21/38), respectively. The positive rate for the HC group with indurations of <5 mm, 5 to 10 mm, and >10 mm were 38% (20/53), 28% (15/53), and 34% (18/53), respectively. *, P = 0.016, comparing the T-SPOT.TB positive rate for healthy individuals with TST induration of >10 mm to TST induration of <5 mm.

FIG. 4.

The ESAT-6-, CFP-10-, and PPD-specific SFCs of ELISPOT assay with ATB patients and HC. Ex vivo ELISPOT assay was performed on 46 ATB patients receiving anti-TB treatment equal to or shorter than 1 month and 50 healthy individuals. The short transverse line represents median of SFCs in different subgroups.

FIG. 5.

The T-SPOT.TB positive rates for ATB patients with different durations of anti-TB treatment. Eighty-nine ATB patients receiving initial anti-TB treatment shorter than 6 months (mo) were screened by T-SPOT.TB assay and divided into 3 subgroups based on the duration of anti-TB treatment: 0 to 1 mo, n = 54; 1 to 3 mo, n = 22; 3 to 6 mo, n = 13. The medians of duration of anti-TB treatment were as follows: for 0 to 1 mo, 0 mo; for 1 to 3 mo, 2 mo; for 3 to 6 mo, 5 mo. *, P < 0.005, comparing the T-SPOT.TB positive rates for the 0- to 1-mo group to the 3- to 6-mo group.

FIG. 6.

The ESAT-6- and CFP-10-specific spot-forming cells of T-SPOT.TB assay in ATB patients with different durations of anti-TB treatment and HC group. T-SPOT.TB assay was performed with 89 ATB patients receiving initial anti-TB treatment shorter than 6 months and 57 healthy individuals. The short transverse line represents median of SFCs in different subgroups. Values in parentheses indicate n.