Diagnostic utility of plasma metagenomic next-generation sequencing for bloodstream and localized infections in advanced HIV disease

Abstract Objectives To evaluate the diagnostic performance of plasma metagenomic next‑generation sequencing (mNGS) for suspected bloodstream infections (BSI) and localized infections in patients with advanced HIV disease. Methods We conducted a retrospective observational study of 110 patients with advanced HIV disease who underwent plasma mNGS. Plasma mNGS and conventional microbiological tests (CMTs) were performed concurrently. The diagnostic performance of plasma mNGS and CMTs was evaluated using a composite reference standard based on final clinical outcomes. In 26 patients with indeterminate plasma results, additional mNGS was performed on clinical specimens (BALF, CSF, pleural fluid, or tissue). Results Plasma mNGS detected clinically relevant pathogens in 81.8% (90/110) of patients. For BSI ( n = 69), sensitivity was 93.9% (95% CI: 84.9–98.3%) with 100% specificity (95% CI: 29.2–100% ); for localized infections ( n = 92), sensitivity was 85.9% (95% CI: 77.6–91.6%).with 100% sensitivity for talaromycosis and 96.2% for Pneumocystis jirovecii pneumonia. Of 263 microbial sequences detected, 124 (47.1%) were clinically relevant; 139 (52.9%)—almost exclusively viral (98.6%)—represented asymptomatic reactivation. All non‑viral sequences were clinically relevant. Co‑detection of multiple clinically relevant organisms occurred in 29 samples (32.2%). In 26 patients with specimen mNGS, concordance with final diagnosis was 96.2% (25/26). Plasma mNGS missed 17 confirmed infections, mostly involving compartmentalized sites (CNS, n = 5) or pathogens with complex cell walls ( n = 10). Conclusions Plasma mNGS demonstrated superior diagnostic performance over CMTs for detecting BSI and localized infections in advanced HIV disease, particularly for talaromycosis and Pneumocystis jirovecii pneumonia. Negative results do not exclude infection; clinical adjudication—particularly for viral detections—is essential. Specimen mNGS provides added value in compartmentalized infections.