BACKGROUND: Unrecognized periprosthetic joint infections are a concern in revision surgery for aseptic failure (AF) after total hip (THA) or knee (TKA) arthroplasties. A gold diagnostic standard does not exist. The aim of the current study was to determine the prevalence of unrecognized periprosthetic joint infection (PJI) in a cohort of revision for AF, using an experimental diagnostic algorithm. METHODS: The surgeons' suspicion of AF was based primarily on patient history and clinical evaluation. X-ray imaging was used to reveal mechanical problems. To rule out an infectious aetiology standard blood biochemical tests were ordered in most patients. Evaluation followed the existing practice in the institute. Cases were included if revision surgery was planned for suspected AF. Intraoperatively, five synovial tissue biopsies were obtained routinely. PJI was defined as ≥3 positive cultures with the same microorganism(s). Patients were followed for 1 year postoperatively. Protocol samples included joint fluid, additional synovial tissue biopsies, bone biopsy, swabs from the implant surface, and sonication of retrieved components. Routine and protocol samples were cultured with extended incubation (14 days) and preserved for batchwise 16S rRNA gene amplification. Patients were stratified based on culture results and a clinical status was obtained at study end. RESULTS: A total of 72 revisions were performed on 71 patients (35 THA and 37 TKA). We found five of 72 cases of unrecognized PJI. Extended culture and protocol samples accounted for two of these. One patient diagnosed with AF was treated for a PJI during follow-up. The remaining patients did not change status from AF during follow-up. CONCLUSIONS: We found a low prevalence of unrecognized periprosthetic joint infections in patients with an AF diagnosis. The algorithm strengthens the surgeons' preoperative diagnosis of a non-infective condition. Evaluation for a failing TKA or THA is complex. Distinguishing between AF and PJI pre-operatively was a clinical decision. Our data did not support additional testing in routine revision surgery for AF.