Salivary exosomal Mycobacterium tuberculosis DNA enables sensitive detection of paucibacillary tuberculosis: a molecular diagnosis study

Background: DNA can diagnose paucibacillary tuberculosis. Methods: We developed the exosomal nucleic acid (ExoNA)-based saliva tuberculosis diagnostic, a rapid, non-sputum diagnostic method for tuberculosis by integrating enrichment of salivary exosomes, nucleic acid extraction, and dual-target qPCR melting curve analysis. In a pilot clinical study, adults with suspected pulmonary tuberculosis were enrolled to evaluate diagnostic performance against multiple reference standards. Subgroup analysis was performed in patients with negative smear microscopy. Results: DNA in saliva. ExoNA-based salivary tuberculosis diagnostic reduced the turnaround time from 16 h to 3h, and reached a limit of detection of 10 CFU/mL. In total, 130 patients with suspected pulmonary tuberculosis were included. Of them, 36·9% (48/130) were asymptomatic, and 80·8% (105/130) had negative smear microscopy results. The ExoNA-based Saliva TB assay had sensitivity of 81.2% (95% CI 53.7%-95.0%) and specificity of 82.8% (95% CI 63.5%-93.5%) against microbiological reference standards (n=45), while sensitivity of 68.4% (95% CI 54.6%-79.7%) and specificity of 82.9% (95% CI 67.4%-92.3%) under extended microbiological reference standards (n=98). In the subset of 105 patients with negative smear microscopy, the sensitivities remained within the range of 67.5% to 75.0% across different reference standards, and specificities ranged from 82.1% to 82.9%. Conclusion: ExoNA-based saliva tuberculosis assay demonstrates favorable diagnostic performance in patients with suspected pulmonary tuberculosis, with the advantages of non-invasive nature, easy sampling, and rapid detection. Multicenter validation in more heterogeneous populations is necessary prior to clinical use.