Clinical Use of a 16S Ribosomal RNA Gene-Based Sanger and/or Next Generation Sequencing Assay to Test Preoperative Synovial Fluid for Periprosthetic Joint Infection Diagnosis

Laure Flurin, Joseph J. Hemenway, Cody R. Fisher, James J. Vaillant, Marisa Azad, Matthew J. Wolf, Kerryl E. Greenwood-Quaintance, Matthew P. Abdel, Robin Patel

Research output: Contribution to journalArticlepeer-review


Preoperative pathogen identification in patients with periprosthetic joint infection (PJI) is typically limited to synovial fluid culture. Whether sequencing-based approaches are of potential use in identification of pathogens in PJI, and if so which approach is ideal, is incompletely defined. The objective of the study was to analyze the accuracy of a 16S rRNA (rRNA) gene-based PCR followed by Sanger sequencing and/or targeted metagenomic sequencing approach (tMGS) performed on synovial fluid for PJI diagnosis. A retrospective study was conducted, analyzing synovial fluids tested between August 2020 and May 2021 at a single center. Subjects with hip, knee, shoulder, and elbow arthroplasties who had synovial fluid aspirated and clinically subjected to sequence-based testing and conventional culture were studied. A total of 154 subjects were included in the study; 118 had noninfectious arthroplasty failure (NIAF), while 36 had PJI. Clinical sensitivity and specificity for diagnosis of PJI were 69% and 100%, respectively, for the sequencing-based approach and 72% and 100%, respectively, for conventional culture (P = 0.74). The combination of both tests was more sensitive (83%) than culture alone (P = 0.04). Results of sequencing-based testing led to changes in treatment in four of 36 (11%) PJI subjects. Microbial identification was achieved using Sanger and next generation sequencing in 19 and 6 subjects, respectively. When combined with culture, the described 16S rRNA gene sequencing-based approach increased sensitivity compared to culture alone, suggesting its potential use in the diagnosis of PJI when synovial fluid culture is negative. IMPORTANCE Periprosthetic joint infection (PJI) is a dreadful complication of joint replacement. Noninvasive identification of infectious pathogens has been traditionnally limited to culture-based testing of synovial fluid which has poor sensitivity. Sanger and Next-generation sequencing (NGS) may be used for synovial fluid testing in PJI, but experience in routine practice is sparse. We used a targeted metagenomic sequencing approach for routine testing of synovial fluid involving NGS when Sanger sequencing had failed or was likely to fail. The objective of this study was to analyze the approach's performance for diagnosis of PJI in comparison to culture for testing synovial fluid. Overall, the sequencing-based approach was not superior to culture for diagnosis of PJI, but yielded positive results in some culture-negative samples.

Original languageEnglish (US)
Issue number6
StatePublished - Dec 2022


  • 16S rRNA PCR
  • metagenomics
  • next-generation sequencing
  • periprosthetic joint infection
  • synovial fluid

ASJC Scopus subject areas

  • Microbiology
  • Virology


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