Using droplet digital polymerase chain reaction for cell-free DNA to diagnose tuberculous pleural effusion

Background: Tuberculous pleural effusion (TPE) remains a diagnostic challenge.We aimed to evaluate droplet digital PCR (ddPCR) for detecting Mycobacterium tuberculosis (Mtb)-derived cell-free DNA (cfDNA) in pleural effusion.Methods: Pleural effusion samples were prospectively collected from two hospitals in Taiwan.ddPCR targeting the IS6110 sequence of Mtb was performed to quantify cfDNA levels.Diagnostic performance was compared with Xpert Ultra.Results: Of 91 participants, 29 were diagnosed with TPE and 62 with non-TPE causes.ddPCR showed significantly higher Mtb cfDNA levels in TPE compared with the non-TPE group (1.49 vs. 0.35-0.44copies/L, p<0.001).The optimal cut-off of 10 positive droplets yielded a sensitivity of 83% and specificity of 84%.In contrast, Xpert Ultra showed only 31.8% sensitivity despite 100% specificity.The area under the ROC curve for ddPCR (AUC = 0.861) outperformed Xpert.Conclusions: ddPCR of Mtb-derived cfDNA in pleural effusion offers a sensitive and specific non-invasive diagnostic tool for TPE, outperforming conventional molecular diagnostics.