Feasibility of nanopore metagenomic sequencing of intraoperative deep fluid samples for rapid diagnosis of acute periprosthetic joint infection: a proof-of-concept study

Abstract Background Metagenomic next-generation sequencing (mNGS) has shown promise for the diagnosis of periprosthetic joint infection (PJI), but most studies rely on tissue biopsies, synovial fluid, or sonication fluid. This proof-of-concept feasibility study evaluated the use of nanopore-based mNGS on a single intraoperative deep fluid sample obtained during revision surgery for acute PJI. Methods Patients undergoing primary revision surgery for acute postoperative hip or knee PJI were prospectively included. Intraoperative deep fluid and tissue samples were analyzed using nanopore-based mNGS and compared with conventional microbiological diagnostics. Results were assessed for pathogen detection, antimicrobial resistance (AMR) gene identification, turnaround time, and specimen-related sampling variability. Results Fifteen patients with suspected PJI were included. Deep fluid mNGS detected pathogens in 8 of 14 (57%) patients with culture-confirmed PJI, including 7 detected within 8 h of sequencing and one additional patient detected only after completion of the 72-hour sequencing run. Tissue biopsy analysis demonstrated substantial sampling variability, with several patients showing positive mNGS findings in only a subset of biopsies. AMR genes were detected in several clinically relevant pathogens, although corresponding resistance determinants were not identified in all phenotypically resistant isolates. When detected, pathogen and AMR results were typically available within approximately 15 h from sample processing. In comparison, conventional microbiological diagnostics required a median of 40 h for pathogen identification, 91 h for phenotypic susceptibility testing, and 144 h for final microbiological reporting. Conclusions In this proof-of-concept feasibility study, nanopore-based mNGS of a single intraoperative deep fluid sample provided rapid pathogen and AMR information while substantially reducing sampling complexity. Although the current sensitivity does not support replacement of conventional microbiological diagnostics, deep fluid represents a promising and clinically practical specimen type for future mNGS-based PJI diagnostics.