Introduction Medical management of prosthetic joint infections (PJIs) relies on the identification of causative organisms through traditional culture-based approaches to guide therapy. However, diagnosis of many PJIs remains challenging, with many clinically apparent infections remaining culture-negative. Molecular diagnostics have the potential to increase diagnostic yield, particularly among culture-negative PJIs. Methods Bone, tissue, or synovial fluid from patients with clinically identified PJIs were collected for inclusion in this study. Samples were assessed with traditional cultures and classified as culture-positive or -negative after 48 h. Samples subsequently underwent a Staphylococcus aureus-/Kingella kingae-specific PCR followed by a 16s rRNA gene PCR. Results A total of 77 unique patients with clinically identified PJIs contributed a total of 89 samples for inclusion in the study. There were 54 culture-negative and 35 culture-positive samples evaluated. The sensitivity and specificity of S. aureus PCR in culture-positive samples was 57.1% (95% CI, 34.1%–78.1%) and 92.9% (95% CI, 66.1%–98.9%), respectively. Among culture-positive samples, 16s rRNA gene PCR correctly identified 3 of 21 (14.3%) samples with S. aureus and 2 of 5 (40%) samples with Streptococcus spp. All molecular tests were negative in those with clinically identified, culture-negative PJI. Conclusions Our study suggests that these diagnostic tools have a limited role in PJI diagnosis.

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