The Journal of Bone and Joint Surgery (American). 2007;89:1298-1305.
doi:10.2106/JBJS.F.00822
© 2007 The Journal of Bone and Joint Surgery, Inc.
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Identification of Orthopaedic Infections Using Broad-Range Polymerase Chain Reaction and Reverse Line Blot Hybridization

Dirk Jan F. Moojen, MD1, Sanne N.M. Spijkers, BAS2, Corrie S. Schot, BAS2, Marc W. Nijhof, MD, PhD1, H. Charles Vogely, MD, PhD1, André Fleer, MD, PhD1, Abraham J. Verbout, MD, PhD1, René M. Castelein, MD, PhD1, Wouter J.A. Dhert, MD, PhD, FBSE1 and Leo M. Schouls, PhD2

1 Departments of Orthopaedics (D.J.F.M., M.W.N., H.C.V., A.J.V., R.M.C., and W.J.A.D.) and Medical Microbiology (A.F.), University Medical Center Utrecht, P.O. Box 85500, 3508GA Utrecht, The Netherlands. E-mail address for D.J.F. Moojen: D.J.F.Moojen{at}umcutrecht.nl
2 Laboratory of Vaccine Preventable Diseases, National Institute of Public Health and the Environment (RIVM), P.O. Box 1, 3720BA Bilthoven, The Netherlands

Investigation performed at the University Medical Center Utrecht, Utrecht, and the National Institute of Public Health and the Environment, Bilthoven, The Netherlands

Disclosure: The authors did not receive any outside funding or grants in support of their research for or preparation of this work. Neither they nor a member of their immediate families received payments or other benefits or a commitment or agreement to provide such benefits from a commercial entity. A commercial entity (Stryker Orthopaedics, Mahwah, New Jersey) paid or directed in any one year, or agreed to pay or direct, benefits in excess of $10,000 to a research fund, foundation, division, center, clinical practice, or other charitable or nonprofit organization with which the authors, or a member of their immediate families, are affiliated or associated.

Background: Culture remains the gold standard in the diagnosis of bacterial infection, but molecular biological techniques have yielded promising results. In this study, we validated a combined polymerase chain reaction and reverse line blot hybridization protocol for identifying musculoskeletal infections.

Methods: Samples were obtained from seventy-six patients undergoing orthopaedic surgery for various aseptic and septic indications. The diagnosis of infection was based on a review of all available clinical and culture data. In addition to routine culture for aerobic and anaerobic growth, samples were analyzed with a broad-range 16S rRNA polymerase chain reaction and subsequent reverse line blot hybridization with use of twenty-eight group, genus, and species-specific oligonucleotide probes.

Results: An infection was diagnosed on the basis of patient data in thirty-one patients. All but one of the patients with a clinical diagnosis of infection had a positive result of the polymerase chain reaction-reverse line blot hybridization. Five of the forty-five patients in whom an infection was not suspected on the basis of patient data had at least one positive result of the polymerase chain reaction-reverse line blot hybridization. Cultures demonstrated microorganisms in twenty-five patients with an infection and in two patients in whom an infection was not suspected on the basis of the patient data. Staphylococcus aureus was the most common organism grown on culture. The species identified by the polymerase chain reaction-reverse line blot hybridization was in full accordance with that grown on culture in all but one patient.

Conclusions: Polymerase chain reaction-reverse line blot hybridization performed well in detecting and identifying the various bacterial species and was more sensitive than routine culture. It identified Staphylococcus aureus as the most frequently found microorganism. Five patients in whom an infection was not suspected on the basis of the patient data had a positive result of the polymerase chain reaction, which may have been caused by contamination of the samples. However, three of these patients had aseptic loosening of a total hip prosthesis, suggesting the presence of a low-grade bacterial infection that remained undetected by the culture but was detected by the polymerase chain reaction-reverse line blot hybridization.

Level of Evidence: Diagnostic Level III. See Instructions to Authors for a complete description of levels of evidence.


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