SEMIQUANTITATIVE REAL-TIME PCR VERSUS OPTIMIZED CULTURE FOR BACTERIAL DETECTION IN PROSTHETIC JOINT LOOSENING

Abstract

Low virulent chronic prosthetic infection might be indistinguishable to aseptic loosening. Polymerase chain reaction (PCR) has been introduced to improve bacterial detection of implant infection. However, there is great risk of false positive results when using broad range/universal PCR primers. The source of DNA contaminants may be of environmental origin as well as the reagents employed.

To study the presence of bacterial DNA in culture negative biopsy specimens by using defined criteria for PCR positivity.

We included six specimens from each of 21 patients preoperatively considered having aseptic loosening of a hip prosthesis. These 126 specimens were culture negative after seven days of incubation. Prior to incubation, the specimens were divided, and half of each specimen was processed for PCR.

Three sets of primer pairs targeting the 16S rRNA gene were developed.

Nine specimens from culture proven prosthetic infections and nine specimens from primary prosthetic surgery served as positive and negative controls, respectively.

A specimen was considered PCR positive if:

1. bacterial DNA ≥ 2 times than the reagent control, and

2. a positive PCR signal for ≥ 2 primer pairs.

All the 126 patient specimens were PCR negative and the nine positive controls were positive. One of nine negative specimen controls was PCR positive, DNA sequencing demonstrated a non-pathogen. Five single PCR reactions (1.3 %) were positive.

This study underlines the importance of establishing stringent criteria for interpretation of PCR positivity to apply to clinical specimens. By the criteria used, five single positive PCR reactions were identified as false positives. Positive PCR reactions in all of the specimen positive controls prove the detection ability of the method. One PCR positive result in a specimen negative control demonstrates the contamination risk in collection and handling of biopsies.

In conclusion, by using a semiquantitative PCR and stringent criteria for PCR positivity we were unable to detect any infections missed by culture.