Purpose: To evaluate the diagnostic performance of targeted next-generation sequencing (tNGS), a multiplex PCR–based and hybrid-capture sequencing platform targeting 198 respiratory pathogens, versus Xpert MTB/RIF in diagnosing pleural tuberculosis (TB) across diverse specimen types, given the limited sensitivity of existing assays in paucibacillary disease. Patients and Methods: Consecutive patients with suspected pleural TB were prospectively enrolled. Final diagnosis was established using a composite reference standard incorporating microbiological, histopathological, and clinical criteria. Paired sputum, pleural fluid, and pleural biopsy specimens underwent tNGS, Xpert MTB/RIF, and mycobacterial culture. Diagnostic performance metrics, including sensitivity and specificity, were calculated. A predefined paired comparison (n=105) between tNGS and Xpert MTB/RIF was performed, with agreement assessed using Cohen’s kappa. Results: In the overall cohort (N = 142), tNGS demonstrated higher sensitivity than Xpert MTB/RIF and mycobacterial culture across specimen types when evaluated against the composite reference standard, while maintaining 100% specificity in the non-TB group (n = 8). In sputum samples, tNGS achieved a sensitivity of 72.73%, significantly outperforming Xpert MTB/RIF (21.43%) and MTB culture (16.30%). In pleural fluid, tNGS sensitivity (43.64%) exceeded that of Xpert MTB/RIF (30.49%) and MTB culture (24.14%), comparable to the performance of ADA ≥ 40 U/L (46.22%). The paired analysis confirmed that tNGS significantly outperformed Xpert MTB/RIF in overall diagnostic yield ( p < 0.01), detecting 26 positive cases missed by Xpert. Among patients with culture-negative pleural tuberculosis, tNGS detected 24/60 (40.0%) cases compared with 6/60 (10.0%) by Xpert MTB/RIF. Conclusion: tNGS provides incremental diagnostic value over conventional molecular and culture-based assays in pleural tuberculosis, particularly in paucibacillary and culture-negative disease. It increases microbiological confirmation in culture- and Xpert-negative cases and is best positioned as a complementary second-line assay within a tiered diagnostic framework, despite no assessment of downstream clinical impact. Keywords: targeted next-generation sequencing, pleural tuberculosis, Xpert MTB/RIF, diagnosis
Peng et al. (Fri,) studied this question.