†These authors contributed equally.
Academic Editor: Josef Jampílek
Background: To evaluate the concordance between QuantiFERON-TB Gold
in-tube test (QFT-GIT) and T-SPOT.TB test (T-SPOT)
for the screening of latent tuberculosis
infection (LTBI) in patients with rheumatic diseases (RDs).
Methods: Patients diagnosed as rheumatic diseases (RDs) with
clinical indications for test of interferon gamma release test (IGRA) were
prospectively recruited from 2019 to 2020. The consistency of QFT-GIT and T-SPOT
was assessed by Kappa analysis and the factors associated with the indeterminate
results were explored by multivariable logistic analysis. Results: A
total of 108 patients with RDs were enrolled, including 64 patients with systemic
lupus erythematosus (SLE) and 44 with inflammatory arthritis (26 with rheumatoid
arthritis (RA) and 18 with ankylosing spondylitis (AS)). Poor concordance was
confirmed between QFT-GIT and T-SPOT results in patients with SLE (K = 0.175,
95% confidence interval [95% CI] [–0.06, 0.40], p
Mycobacterium tuberculosis (MTB) infection could be a lethal complication in patients with rheumatic diseases (RDs), e.g., systemic lupus erythematosus (SLE) and inflammatory arthritis, due to autoimmune disorder and the treatment with glucocorticoid and immunosuppressants [1, 2, 3]. SLE patients are 5–15 times more likely to develop tuberculosis infection than the general population [3, 4], while patients with inflammatory arthritis had a 4 to 8-fold increased risk of tuberculosis compared to the general population . Early recognition of latent tuberculosis infection (LTBI) and prophylaxis in patients at high risk is fundamental in order to improve the prognosis and life quality of patients, as well as to reduce the costs associated with the disease. Therefore, the screening of LTBI and tuberculosis prophylaxis prior to immunosuppressive treatment are of a central importance in these population.
Latent tuberculosis infection is “a state of persistent immune response to
stimulation by MTB antigens without evidence of clinically manifested active
tuberculosis” . Both of tuberculin test (TST) and interferon gamma release
test (IGRA) are used in the screening of LTBI. IGRA is an interferon-gamma
This prospective cohort study was conducted from 1 January 2019 to 31 December 2020 at Renji Hospital, Shanghai, China. The research protocol was approved by Shanghai Jiao Tong University, School of Medicine, Renji Hospital Ethics Committee (No. 2016-Clinical-Res-011). All participating patients provided written informed consent. Eligible patients were recruited when the following inclusion criteria were met: (1) they fulfilled the American College of Rheumatology (ACR) or European League Against Rheumatism (EULAR) criteria for SLE, rheumatoid arthritis (RA) and ankylosing spondylitis (AS) [12, 13, 14]; (2) they had clinical indications for the test of IGRA. Exclusion criteria were: presence of active tuberculosis, patients undergoing anti-tuberculosis treatment, incomplete medical history, or patients who refused QFT-GIT and T-SPOT testing.
QFT-GIT was used for testing LTBI in the
study. Peripheral blood samples were collected and processed following the
instructions of manufacturer, Cellestis/Qiagen, Carnegie, VIC, Australia. While
performing QFT-GIT test, patients’ whole blood was collected into three QFT-GIT
collection tubes, including blank control (Nil) tubes, mitogen (M) tubes and
tuberculosis antigen (TB) tubes with an enzyme-linked immunosorbent assay to
detect interferon values. The Nil tube was a negative control, and IFN-
T-SPOT was preformed simultaneously. Specifically, the peripheral blood
mononuclear cell (PBMC) samples were stimulated with Panel A with ESAT-6, Panel B
with CFP-10, negative control and positive control. IFN-
We recorded demographic data (age, gender), laboratory indicators, including C Reactive Protein (CRP), Erythrocyte Sedimentation Rate (ESR), and CD4+ lymphocyte count, which were operated on the same day with IGRAs. For SLE patients, neutrophil counts, lymphocyte counts, hemoglobin, platelet counts, and complement 3/4 (C3/4) and anti-double-stranded DNA (anti-dsDNA) antibody were assessed additionally. Medication was reviewed, including the treatment with glucocorticoids and immunosuppressive agents. SLE disease activity was assessed with the Systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI 2000) . The concordance between QFT-GIT and T-SPOT were calculated and factors associated with indeterminate results were also analyzed in the SLE patients.
The data were analyzed using IBM SPSS version 20.0 (Chicago, IL, USA). The quantified variables
were expressed as mean
A total of 108 patients were enrolled in this study, including 64 with SLE and 44 with inflammatory arthritis (25 with RA and 19 with AS) (Fig. 1). Patients’ characteristics, laboratory parameters and medications were summarized in Table 1. The mean age of SLE patients was 37.3 years old and 85.4% were female patients. The median disease duration for patients with SLE was 60 months with a median SLEDAI score of 6.0 (4.0–9.0). For patients with inflammatory arthritis, the mean age in years was 47.3 (16.4) and the proportion of women was 26 (59.1%).
Flow chart of the study. RDs, rheumatic diseases; LTBI, latent tuberculosis infection; SLE, systemic lupus erythematosus; T-SPOT, T-SPOT.TB test; QFT-GIT, QuantiFERON-TB Gold in-tube test.
|Characteristics||SLE (n = 64)||Inflammatory arthritis (n = 44)|
|Female, n (%)||54 (85.4%)||26 (59.1%)|
|Age, years, Mean
|Disease Duration, Median (IQR), Months||60 (0–120)||3.5 (2–8.3)|
|SLEDAI, Median (IQR)||6.0 (4.0–9.0)||/|
|Previous Tuberculosis History, N (%)||1 (1.56%)||1 (2.27%)|
|Dose of Glucocorticoids, Median (IQR), mg/day||15 (0.25–33.75)||0 (0–6.9)|
|Lymphocyte Count, Mean
|Neutrophil Count, Mean
|CD4+ T Lymphocyte Count, Mean
|Complement 3, Mean
|Complement 4, Median (IQR), mg/dL||9.65 (5.43–14.75)||/|
|C-Reactive Protein, Median (IQR) mg/dL||0.07 (0–1.24)||0.95 (0.31–2.89)|
|Erythrocyte Sedimentation Rate, Median (IQR), mm/hour||30 (20–49)||25 (14–62)|
|Anti-dsDNA Antibody Median (IQR), IU/mL||42.8 (22.5–100)||/|
|Hydroxychloroquine, n (%)||40 (62.50%)||9 (20.5%)|
|Methotrexate, n (%)||5 (7.81%)||20 (45.5%)|
|Leflunomide, n (%)||4 (6.3%)||9 (20.5%)|
|Sulfasalazine, n (%)||0||9 (20.5%)|
|Azathioprine, n (%)||1 (1.56%)||0|
|Tacrolimus, n (%)||8 (12.50%)||0|
|Mycophenolate mofetil, n (%)||9 (14.06%)||0|
|Cyclophosphamide, n (%)||5 (7.81%)||0|
|SLEDAI, Systemic Lupus Erythematosus disease activity index; Anti-dsDNA, anti-double strand DNA antibody.|
Among SLE patients, QFT-GIT was positive in 3 patients (4.7%), indeterminate in
12 (18.8%) and negative in 49 (76.6%). T-SPOT was positive in 2 (3.1%),
indeterminate in 3 (4.7%) and negative in 59 (92.2%) (Tables 2,3). In patients
with inflammatory arthritis, QFT-positive, -negative and -indeterminate results
were 7 (15.9%), 36 (81.8%) and 1 (2.3%), respectively. Positive, negative and
indeterminate diagnosis of T-SPOT were, respectively, 4 (9.1%), 37 (84.1%) and
3 (6.8%). We found that QFT-GIT and T-SPOT have poor consistency in SLE (K =
|QFT-GIT||SLE||12 (18.8%)||52 (81.3%)||0.013|
|Inflammatory arthritis||1 (2.3%)||43 (97.7%)|
|T-SPOT||SLE||3 (4.7%)||61 (95.3%)||0.686|
|Inflammatory arthritis||3 (6.8%)||41 (93.2%)|
|RA, Rheumatoid Arthritis; AS, Ankylosing Spondylitis; SLE, Systemic Lupus Erythematosus.|
|Kappa||†K = 0.175 (95% CI [–0.06, 0.40])||‡K = 0.539 (95% CI [0.11, 0.88])|
|†K: the concordance between QFT-GIT and T-SPOT in SLE
‡K: the concordance between QFT-GIT and T-SPOT in RA and AS (p
SLE, Systemic Lupus Erythematosus.
QFT-GIT, QuantiFERON® TB Gold In-Tube; T-SPOT, T-SPOT®.TB.
The ratio of indeterminate values of QFT-GIT in SLE was higher than that in inflammatory arthritis patients (18.8% vs. 4.7%, p = 0.013) (Fig. 2A). Among the 12 patients with indeterminate results in SLE by QFT-GIT, one patient showed positive T-SPOT, one patient exhibited indeterminate T-SPOT and 10 (83.3%) patients displayed a negative T-SPOT. In QFT-GIT, indeterminate results due to no response to phytohemagglutinin A (PHA) were more frequent in lupus patients (12.5%) compared to patients with inflammatory arthritis (1.56%) (p = 0.046) (Fig. 2B). Responses to PHA were lower in lupus patients compared to inflammatory arthritis (p = 0.004). During the follow-up, no patient received a treatment for LTBI or developed an active TB in the 12 patients with indeterminate results in SLE by QFT-GIT.
Ratios of the indeterminate results in QFT-GIT and T-SPOT IGRA tests in patients with SLE and inflammatory arthritis. (*p = 0.013) (A); due to no response to PHA in patients with SLE and inflammatory arthritis by QFT-GIT (*p = 0.046) (B). SLE, systemic lupus erythematosus; QFT, QuantiFERON-TB Gold in-tube test; T-SPOT, T-SPOT.TB test; PHA, phytohaemagglutinin A.
The indeterminate results in SLE patients were associate with neutrophil counts (p = 0.041), lymphocyte counts (p = 0.004), CD4+ T Lymphocyte counts (p = 0.050), C3 (p = 0.011) according to univariable analysis (Table 4). Lymphocyte counts (OR [95% CI] 0.81 [0.68, 0.97], p = 0.020) was an independent predictor of QFT-GIT indeterminate results according to multivariable logistic analysis.
|Explanatory variables||OR (95% CI)||p-value|
|Age||1.01 (0.97, 1.05)||0.743|
|Male||1.10 (0.20, 5.99)||0.912|
|Neutrophil count||0.98 (0.97, 1.00)||0.041|
|Lymphocyte count||0.80 (0.68, 0.93)||0.004|
|CD4+ T Lymphocyte count||0.84 (0.70, 1.00)||0.050|
|Neutrophil/lymphocyte counts||1.02 (0.89, 1.17)||0.762|
|Hemoglobin||0.98 (0.96, 1.01)||0.192|
|Platelet counts||0.99 (0.98, 1.00)||0.088|
|ESR||0.99 (0.96, 1.02)||0.477|
|C3||0.40 (0.20, 0.81)||0.011|
|C4||0.85 (0.68, 1.06)||0.140|
|Anti-dsDNA antibody||1.01 (0.99, 1.03)||0.375|
|SLEDAI||1.00 (0.85, 1.18)||0.987|
|Immunosuppressive agents||2.88 (0.57, 14.54)||0.201|
|Doses of glucocorticoids||0.99 (0.96, 1.02)||0.567|
|Age||1.02 (0.96, 1.09)||0.482|
|Male||1.05 (0.09, 11.96)||0.974|
|Neutrophil count||1.00 (0.98, 1.03)||0.853|
|Lymphocyte count||0.81 (0.68, 0.97)||0.020|
|C3||0.42 (0.15, 1.14)||0.094|
|QFT, QuantiFERON-TB Gold in-tube test; SLE, Systemic Lupus Erythematosus; SLEDAI, systemic lupus erythematous disease activity index.|
This was the first study designed to compare the consistency of T-SPOT and QFT-GIT in patients with rheumatic diseases, including SLE, RA and AS. It was found that the consistency was particularly lower in patients with SLE. Low lymphocyte count was found to be associated with QFT-GIT indeterminate assay results. In addition, there was a fair-to-good agreement between QFT-GIT and T-SPOT in patients with inflammatory arthritis including RA and AS. The importance of this study lies in the comparison of efficacy between QFT-GIT and T-SPOT while screening LTBI in patients with SLE. Our data provided new evidence supporting T-SPOT could be a more suitable test for LTBI screening in SLE patients, for using T-SPOT yielded a significantly lower rate of indeterminate rates in this population.
Patients with rheumatic diseases are reported to have increased risk of tuberculosis infection due to the pathogenesis and medication . A meta-analysis drew a conclusion that the method of T-SPOT was more sensitive than QFT-GIT in people who received immunosuppressive agents . In several studies, the QFT-indeterminate ratio in SLE population has been reported to be higher than other rheumatic diseases or immunosuppressed patients [7, 19, 20]. In line with these results, QFT-GIT may be insufficient in detecting LTBI owing to higher rate of indeterminate results and T-SPOT could have a potential advantage in this aspect. At the same time, our findings are indicating that diminished response to PHA is a major reason for indeterminate results in SLE patients by QFT-GIT.
On the other hand, our study showed a fair-to-good agreement between QFT-GIT and T-SPOT in patients with RA and AS, which featured with chronic inflammation in joints. A study reports that no significant differences were found between QFT-GIT and T-SPOT according to the proportions of positive and indeterminate in patients with RA . Kappa values were 0.6 (95% CI [0.39–0.80]) and 0.39 (95% CI [0.21–0.57]) for positive or negatives result. Another study showed that the QFT-GIT indeterminate results in RA patients were approximately 9.9% . Both of the two tests were applicable while diagnosing LTBI in patients with chronic inflammatory arthritis.
Furthermore, our study discovered that low
lymphocyte count was associated with QFT-GIT indeterminate results. However, high
SLEDAI scores seemed not related with a high proportion of indeterminate results,
and this differs from some studies [20, 21, 22, 23, 24]. Other previous researches showed
that CD4+ T cell count, lymphocyte count, and lymphocyte percentage were
influencing factors for the indeterminate results of QFT-GIT in HIV and
immunosuppressive people [25, 26]. Glucocorticoid therapy is also another reason
for the indeterminate outcome of QFT [20, 27, 28]. It was suggested that patients
with indeterminate results were more likely to be aged over 70 years, females,
suffering from SLE, lymphopenia, Anemia, thrombocytopenia, and hypoalbuminemia
. QFT-GIT results were affected by various factors and might have a
limitation in LTBI screening based on the clinical settings, and further
investigations were needed. The poor agreement between QFT-GIT and T-SPOT in
lupus patients in this study might be due to the following two reasons. First,
there was a large amount of IFN-
The results of the study suggested that T-SPOT might have an advantage as a LTBI screening test in terms of having lesser indeterminate results, especially in patients with SLE. However, there were some limitations in the study. First, a general population was not included as a control in this single-center cohort study, due to practical constraints. Second, a bias might exist due to the limited number of patients with RA and AS. In addition, the influence of the inclusion of a heterogeneous population, the differences in testing indications, disease duration, and the different medications administrated on the results could not be ruled out.
Based on the results of the study, the T-SPOT assay might be more advantageous than the QFT-GIT assay in detecting LTBI, and this is mainly related to a less proportion of indeterminate results in patients with SLE compared with inflammatory arthritis. These outcomes are supporting the application of T-SPOT as a test for LTBI screening in individuals with SLE.
(I) Conception and design—JZ, JL; (II) Administrative support—SC, LL; (III) Provision of study materials or patients—LL and LZ; (IV) Collection and assembly of data—JZ, PY and SY; (V) Data analysis and interpretation—LZ and JZ; (VI) Manuscript writing—LZ and JZ; (VII) Final approval of manuscript—All authors.
The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. The study was conducted in accordance with the Declaration of Helsinki (as revised in 2013). The study was approved by the Ethics committee of the Renji Hospital (No. 2016-Clinical-Res-011) and individual consent for this retrospective analysis was waived.
We thank Francesca Severino for English language editing and we thank Qiagen for providing the QFT-kits.
This work was supported by National Key Research and Development Program “Precision Medicine Research” Key Special Project (Ministry of Science and Technology) (2017YFC0909000), National Natural Science Foundation Youth Project (71804109), and “Rising Stars of Medical Talents” Youth Development Program-Youth Medical Talents (SHWSRS(2021)_099).
The authors declare no conflict of interest.
Publisher’s Note: IMR Press stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.