Risk Factors for False-Negative T-SPOT.TB Results in Bacteriologically Confirmed Pulmonary Tuberculosis: A...

T-SPOT.TB is an interferon-γ release assay that aids in detecting Mycobacterium tuberculosis infection but cannot distinguish latent infection from active tuberculosis. False-negative results may delay diagnosis and treatment in patients with active pulmonary tuberculosis (PTB). This study investigated factors associated with false-negative T-SPOT.TB results in bacteriologically confirmed PTB.

This retrospective cohort study included 4,931 adults with bacteriologically confirmed PTB hospitalized at Fuzhou Pulmonary Hospital, Fujian Province, China, between January 2018 and September 2025. The primary outcome was a false-negative T-SPOT.TB result. Demographic characteristics, comorbidities, radiological findings, inflammatory markers, and T-lymphocyte subset counts were collected. Multivariable logistic regression was used to identify independent predictors, while restricted cubic spline (RCS) analyses explored potential nonlinear dose-response relationships.

False-negative T-SPOT.TB results occurred in 437 patients (8.86%). Compared with patients with positive results, those with false-negative results were older (median, 61 vs 57 years), had higher neutrophil-to-lymphocyte ratios (NLR; 3.85 vs 3.25), and had lower CD4⁺ T-cell counts (540 vs 594 cells/μL) (all P<0.001). Older age (OR=1.007, 95% CI: 1.001-1.014; P=0.024), chronic obstructive pulmonary disease (OR=1.848, 95% CI: 1.273-2.682; P=0.001), and elevated NLR (OR=1.018, 95% CI: 1.002-1.035; P=0.026) were independent risk factors. Diabetes mellitus, cavitary lesions, airway lesions, and higher CD4⁺ T-cell counts were independently associated with a reduced risk of false-negative results. RCS analyses demonstrated nonlinear associations for NLR (P for nonlinearity <0.001) and CD4⁺ T-cell count (P for nonlinearity=0.002), with a marked increase in false-negative risk above an NLR of 2.96 and below a CD4⁺ T-cell count of 490.7 cells/μL.

Advanced age, chronic obstructive pulmonary disease, elevated NLR, and reduced CD4⁺ T-cell counts increase the likelihood of false-negative T-SPOT.TB results. In high-risk patients with TB-compatible imaging, a negative T-SPOT.TB result should not preclude further microbiological evaluation or diagnostic investigation.