Background: Propionibacterium (P) acnes infection of the shoulder after arthroplasty is a common and
serious complication. Current detection methods for P acnes involve anaerobic cultures that require pro-
longed incubation periods (typically 7-14 days). We have developed a polymerase chain reaction (PCR)-
restriction fragment length polymorphism (RFLP) approach that sensitively and specifically identifies P
acnes in tissue specimens within a 24-hour period.
Methods: Primers were designed to amplify a unique region of the 16S rRNA gene in P acnes that con- tained a unique HaeIII restriction enzyme site. PCR and RFLP analyses were optimized to detect P acnes DNA in in vitro cultures and in arthroscopic surgical biopsy specimens from patients with P acnes infections.
Results: A564base-pairPCRampliconwasderivedfromalloftheknownPacnesstrains.HaeIIIdigests of the amplicon yielded a restriction fragment pattern that was unique to P acnes. P acnes-specific amplicons were detected in as few as 10 bacterial cells and in clinical biopsy specimens of infected shoulder tissues.
Conclusion: This PCR-RFLP assay combines the sensitivity of PCR with the specificity of RFLP mapping to identify P acnes in surgical isolates. The assay is robust and rapid, and a P acnes-positive tissue specimen can be confirmed within 24 hours of sampling, facilitating treatment decision making, targeted antibiotic therapy, and monitoring to minimize implant failure and revision surgery.
Level of evidence: Basic Science Study; Microbiologic Laboratory Study
© 2016 Journal of Shoulder and Elbow Surgery Board of Trustees. All rights reserved.
Comment: this is a significant contribution, the sample costs 5 dollars and it simplifies the process of diagnosis a low grade infection which most of the time is in "stealth mode" and it is difficult to diagnose. The protocol I follow when I suspect a p acnes infection is that I do a one stage exchange of the components, i do not remove well fixed components, I do extensive debridement and I prescribe oral Augmentin 875 mg BID for 14 days until the cultures are finalized. This window of antibiotic treatment with Augmentin can be eliminated with the use of this test. Below is an example of a different application for the diagnosis of pseudomonas.
The authors of the p acnes paper state:
"A major strength of PCR-RFLP assay described in this report is its specificity for P acnes. This specificity is predicted to be greater than routine culture-based approaches and previously published PCR approaches, because it has 2 levels of verification. The first level is PCR amplification with P acnes-specific primers, and the second is a P acnes-specific RFLP pattern. Previous reports had the first (albeit less specific than ours), but not the second. The strength of this extremely sensitive technology is the ability to detect fewer than 10 P acnes cells in the sample. The weakness is that this level of sensitivity makes aseptic sample collection very important. PCR and RFLP mapping are techniques that are routinely performed in many clinical microbiology and pathology laboratories, and so this approach is not restricted to highly specialized research laboratories."
Methods: Primers were designed to amplify a unique region of the 16S rRNA gene in P acnes that con- tained a unique HaeIII restriction enzyme site. PCR and RFLP analyses were optimized to detect P acnes DNA in in vitro cultures and in arthroscopic surgical biopsy specimens from patients with P acnes infections.
Results: A564base-pairPCRampliconwasderivedfromalloftheknownPacnesstrains.HaeIIIdigests of the amplicon yielded a restriction fragment pattern that was unique to P acnes. P acnes-specific amplicons were detected in as few as 10 bacterial cells and in clinical biopsy specimens of infected shoulder tissues.
Conclusion: This PCR-RFLP assay combines the sensitivity of PCR with the specificity of RFLP mapping to identify P acnes in surgical isolates. The assay is robust and rapid, and a P acnes-positive tissue specimen can be confirmed within 24 hours of sampling, facilitating treatment decision making, targeted antibiotic therapy, and monitoring to minimize implant failure and revision surgery.
Level of evidence: Basic Science Study; Microbiologic Laboratory Study
© 2016 Journal of Shoulder and Elbow Surgery Board of Trustees. All rights reserved.
Comment: this is a significant contribution, the sample costs 5 dollars and it simplifies the process of diagnosis a low grade infection which most of the time is in "stealth mode" and it is difficult to diagnose. The protocol I follow when I suspect a p acnes infection is that I do a one stage exchange of the components, i do not remove well fixed components, I do extensive debridement and I prescribe oral Augmentin 875 mg BID for 14 days until the cultures are finalized. This window of antibiotic treatment with Augmentin can be eliminated with the use of this test. Below is an example of a different application for the diagnosis of pseudomonas.
The authors of the p acnes paper state:
"A major strength of PCR-RFLP assay described in this report is its specificity for P acnes. This specificity is predicted to be greater than routine culture-based approaches and previously published PCR approaches, because it has 2 levels of verification. The first level is PCR amplification with P acnes-specific primers, and the second is a P acnes-specific RFLP pattern. Previous reports had the first (albeit less specific than ours), but not the second. The strength of this extremely sensitive technology is the ability to detect fewer than 10 P acnes cells in the sample. The weakness is that this level of sensitivity makes aseptic sample collection very important. PCR and RFLP mapping are techniques that are routinely performed in many clinical microbiology and pathology laboratories, and so this approach is not restricted to highly specialized research laboratories."
