Multicentric validation of the PathoDetect™ MTB RIF & INH assay for simultaneous detection of Mycobacterium tuberculosis, & drug resistance to rifampicin & isoniazid in presumptive pulmonary tuberculosis & drug-resistant TB patients.

Background & objectives Tuberculosis (TB) remains a major global health concern, with India accounting for 26 per cent of the global burden. Despite advances, access to rapid molecular diagnostics is limited, and the assays currently used in National TB Elimination Programme (NTEP) do not detect isoniazid (INH) resistance upfront. PathoDetect™ MTB RIF & INH is an indigenous closed-system assay that simultaneously detects Mycobacterium tuberculosis (MTB) and resistance to rifampicin (RIF) and INH. This study evaluated its diagnostic characteristics. Methods In this cross-sectional multicenter study conducted at six TB reference laboratories in India, 1039 participants were enrolled (718 presumptive pulmonary TB, 321 presumptive multidrug resistant TB; MDR-TB). PathoDetect™'s discriminatory ability was assessed using the measures sensitivity and specificity, and its diagnostic performance using positive predictive value (PPV) and negative predictive value (NPV). Liquid culture served as the reference standard for MTB detection, while phenotypic drug susceptibility testing (pDST) and line probe assay (LPA) as reference standards for RIF and INH resistance detection. Results For MTB detection in presumptive pulmonary TB (PTB), PathoDetect™ showed a sensitivity of 98.1 per cent [95% confidence interval (CI): 96.1-99.2], specificity of 94.2 per cent (95% CI: 91-96.5), PPV of 94.9 per cent (95% CI: 92.2-96.9), and NPV of 97.8 per cent (95% CI: 95.5-99.1) with near-perfect agreement with Truenat® (k=0.89). Among 514 confirmed TB cases, PathoDetect™ detected RIF resistance with a sensitivity of 86.5 per cent (95% CI: 80.2-91.5), specificity of 91.6 per cent (95% CI: 88.2-94.3), PPV of 82.3 per cent (95% CI: 75.6-87.8), and NPV of 93.8 per cent (95% CI: 90.7-96.1). For INH resistance, sensitivity was 88.9 per cent (95% CI: 84.1-92.6), specificity 87 per cent (95% CI: 82.4-90.8), PPV 85.6 per cent (95% CI: 80.5-89.8), and NPV 90 per cent (95% CI: 85.7-93.4) using pDST as reference. Truenat® MTB-RIF showed comparable performance for RIF resistance detection (k=0.75). Compared to line probe assay (LPA), PathoDetect™ demonstrated higher sensitivity (93.4 vs. 88.8%), specificity (98.2 vs. 93.9%), PPV (96.1 vs. 86.8%) and NPV (97 vs. 94.9%) for RIF resistance detection over Truenat®. Interpretation & conclusions PathoDetect™ is a reliable molecular diagnostic tool for detection of MTB and resistance to RIF & INH. The assay showed better RIF resistance detection compared to INH. Its high sensitivity and specificity indicate strong discriminatory ability, while PPV and NPV demonstrate reasonably good diagnostic performance in the study population. These findings support PathoDetect™ as a promising alternative for rapid TB diagnosis, particularly in high-burden settings.