Targeted metagenomic sequencing for respiratory pathogen detection in hospitalized SARI patients in Indonesia: Comparison with routine RT-qPCR.

BACKGROUND: Respiratory tract infections (RTIs) remain a major cause of morbidity and mortality. Routine RT-qPCR is widely used for respiratory pathogen detection but is restricted to predefined targets and may miss relevant pathogens and co-detections. We evaluated targeted metagenomic sequencing using the Respiratory Pathogen ID/AMR Enrichment Panel (RPIP) for respiratory pathogen detection in hospitalized patients with severe acute respiratory infection (SARI) in Indonesia and compared its performance with routine RT-qPCR. METHODS: We analyzed 189 nasopharyngeal specimens from hospitalized SARI patients at a single referral hospital in Bandung, Indonesia, in two sequencing batches (Batch 1, n = 96; Batch 2, n = 93). RPIP results were compared with commercial RT-qPCR assays using overall percent agreement, positive percent agreement (PPA), and negative percent agreement (NPA). Additional analyses assessed broader pathogen detections, co-detections, AMR-associated determinants, and surveillance-oriented microbiological patterns. RESULTS: RPIP showed high agreement with routine RT-qPCR across most shared viral targets, particularly influenza viruses, SARS-CoV-2, adenovirus, respiratory syncytial virus, and human metapneumovirus, with lower agreement observed for rhinovirus. Beyond routine viral targets, RPIP identified additional bacterial and fungal detections, frequent viral-viral and viral-bacterial co-detections, and diverse AMR-associated determinants from the same respiratory specimens. CONCLUSIONS: RPIP showed good agreement with routine RT-qPCR while providing broader respiratory pathogen and AMR-related information from the same specimens. These findings support its use as a complementary approach for respiratory infectious disease diagnostics, broader microbiological characterization, and surveillance-oriented assessment in hospital-based SARI settings.