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Systematic PCR Detection in Culture-negative Osteoarticular Infections

Published:October 15, 2013DOI:https://doi.org/10.1016/j.amjmed.2013.04.027

      Abstract

      Background

      Identification of microorganisms is crucial for the successful treatment of osteoarticular infections. Molecular methods are more sensitive than culture-dependent methods but may suffer from lack of specificity.

      Methods

      We studied a large series of 3840 bone and joint culture-negative samples collected from 2308 patients hospitalized in Marseille University Hospitals from November 2007 to October 2009. The samples were systematically cultured for 15 days, and conventional broad-range polymerase chain reaction (PCR) (16S rDNA and 18S rDNA) as well as real-time PCR assays targeting human Bglobin, Staphylococcus aureus, and Kingella kingae were realized on one culture-negative specimen.

      Results

      Specimens from 741 patients (32.1%) tested positive by culture, including 38 in which bacteria grew only after 6 days of incubation. PCR was positive in 141 (9%) culture-negative specimens. Microorganisms identified by PCR were classified into 2 groups: fastidious bacteria (n = 35), mostly anaerobes in adult patients, and K. kingae in children; and nonfastidious bacteria (n = 106), mostly S. aureus (32.7%). A discrepancy between a positive PCR result for S. aureus and a negative culture were explained by previous antibiotherapy in 31.4% of cases. Our study highlights the usefulness of systematic 16S rDNA gene PCR for the diagnosis of bone and joint infections in culture-negative patients, thus enabling the administration of specific antibiotic treatments.

      Conclusions

      We recommend the use of conventional broad-range PCR for culture-negative bone and joint specimens, as well as S. aureus-specific PCR for adults and K. kingae-specific PCR for children. 18S rDNA PCR should be reserved only for specific cases.

      Keywords

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      References

        • Zimmerli W.
        • Trampuz A.
        • Ochsner P.E.
        Prosthetic-joint infections.
        N Engl J Med. 2004; 351: 1645-1654
        • Shirtliff M.E.
        • Mader J.T.
        Acute septic arthritis.
        Clin Microbiol Rev. 2002; 15: 527-544
        • Lew D.P.
        • Waldvogel F.A.
        Osteomyelitis.
        Lancet. 2004; 364: 369-379
        • Fenollar F.
        • Roux V.
        • Stein A.
        • Drancourt M.
        • Raoult D.
        Analysis of 525 samples to determine the usefulness of PCR amplification and sequencing of the 16S rRNA gene for diagnosis of bone and joint infections.
        J Clin Microbiol. 2006; 44: 1018-1028
        • Moojen D.J.
        • Spijkers S.N.
        • Schot C.S.
        • et al.
        Identification of orthopaedic infections using broad-range polymerase chain reaction and reverse line blot hybridization.
        J Bone Joint Surg Am. 2007; 89: 1298-1305
        • Tarkin I.S.
        • Henry T.J.
        • Fey P.I.
        • Iwen P.C.
        • Hinrichs S.H.
        • Garvin K.L.
        PCR rapidly detects methicillin-resistant staphylococci periprosthetic infection.
        Clin Orthop Relat Res. 2003; 414: 89-94
        • Tunney M.M.
        • Patrick S.
        • Curran M.D.
        • et al.
        Detection of prosthetic hip infection at revision arthroplasty by immunofluorescence microscopy and PCR amplification of the bacterial 16S rRNA gene.
        J Clin Microbiol. 1999; 37: 3281-3290
        • Mariani B.D.
        • Martin D.S.
        • Levine M.J.
        • Booth Jr., R.E.
        • Tuan R.S.
        The Coventry Award. Polymerase chain reaction detection of bacterial infection in total knee arthroplasty.
        Clin Orthop Relat Res. 1996; 331: 11-22
        • Mariani B.D.
        • Tuan R.S.
        Advances in the diagnosis of infection in prosthetic joint implants.
        Mol Med Today. 1998; 4: 207-213
        • van der Heijden I.M.
        • Wilbrink B.
        • Vije A.E.
        • Schouls L.M.
        • Breedveld F.C.
        • Tak P.P.
        Detection of bacterial DNA in serial synovial samples obtained during antibiotic treatment from patients with septic arthritis.
        Arthritis Rheum. 1999; 42: 2198-2203
        • Borst A.
        • Box A.T.
        • Fluit A.C.
        False-positive results and contamination in nucleic acid amplification assays: suggestions for a prevent and destroy strategy.
        Eur J Clin Microbiol Infect Dis. 2004; 23: 289-299
        • Moumile K.
        • Merckx J.
        • Glorion C.
        • Berche P.
        • Ferroni A.
        Osteoarticular infections caused by Kingella kingae in children: contribution of polymerase chain reaction to the microbiologic diagnosis.
        Pediatr Infect Dis J. 2003; 22: 837-839
        • Stahelin J.
        • Goldenberger D.
        • Gnehm H.E.
        • Altwegg M.
        Polymerase chain reaction diagnosis of Kingella kingae arthritis in a young child.
        Clin Infect Dis. 1998; 27: 1328-1329
        • Verdier I.
        • Gayet-Ageron A.
        • Ploton C.
        • et al.
        Contribution of a broad range polymerase chain reaction to the diagnosis of osteoarticular infections caused by Kingella kingae: description of twenty-four recent pediatric diagnoses.
        Pediatr Infect Dis J. 2005; 24: 692-696
        • Verettas D.
        • Kazakos C.
        • Tilkeridis C.
        • Dermon A.
        • Petrou H.
        • Galanis V.
        Polymerase chain reaction for the detection of Mycobacterium tuberculosis in synovial fluid, tissue samples, bone marrow aspirate and peripheral blood.
        Acta Orthop Belg. 2003; 69: 396-399
        • Hunfeld K.P.
        • Rittmeister M.
        • Wichelhaus T.A.
        • Brade V.
        • Enzensberger R.
        Two cases of chronic arthritis of the forearm due to Mycobacterium tuberculosis.
        Eur J Clin Microbiol Infect Dis. 1998; 17: 344-348
        • Titov A.G.
        • Vyshnevskaya E.B.
        • Mazurenko S.I.
        • Santavirta S.
        • Konttinen Y.T.
        Use of polymerase chain reaction to diagnose tuberculous arthritis from joint tissues and synovial fluid.
        Arch Pathol Lab Med. 2004; 128: 205-209
        • La Scola B.
        • Michel G.
        • Raoult D.
        Use of amplification and sequencing of the 16S rRNA gene to diagnose Mycoplasma pneumoniae osteomyelitis in a patient with hypogammaglobulinemia.
        Clin Infect Dis. 1997; 24: 1161-1163
        • Lee A.H.
        • Ramanujam T.
        • Ware P.
        • et al.
        Molecular diagnosis of Ureaplasma urealyticum septic arthritis in a patient with hypogammaglobulinemia.
        Arthritis Rheum. 1992; 35: 443-448
        • Krause K.
        • Wenish C.
        • Fladerer P.
        • Daxbock F.
        • Krejs G.J.
        • Reisinger E.C.
        Osteomyelitis of the hip joint associated with systemic cat-scratch disease in an adult.
        Eur J Clin Microbiol Infect Dis. 2000; 19: 781-783
        • Woestyn S.
        • Moreau M.
        • Munting E.
        • Bigaignon G.
        • Delmee M.
        Osteomyelitis caused by Bartonella henselae genotype I in an immunocompetent adult woman.
        J Clin Microbiol. 2003; 41: 3430-3432
        • Harris K.A.
        • Fidler K.J.
        • Hartley J.C.
        • et al.
        Unique case of Helicobacter sp. osteomyelitis in an immunocompetent child diagnosed by broad-range 16S PCR.
        J Clin Microbiol. 2002; 40: 3100-3103
        • Ince A.
        • Rupp J.
        • Frommelt L.
        • Katzer A.
        • Gille J.
        • Lohr J.F.
        Is “aseptic” loosening of the prosthetic cup after total hip replacement due to nonculturable bacterial pathogens in patients with low-grade infection?.
        Clin Infect Dis. 2004; 39: 1599-1603
        • Casalta J.P.
        • Gouriet F.
        • Roux V.
        • Thuny F.
        • Habib G.
        • Raoult D.
        Evaluation of the LightCycler SeptiFast test in the rapid etiologic diagnosis of infectious endocarditis.
        Eur J Clin Microbiol Infect Dis. 2009; 28: 569-573
        • Thurman K.A.
        • Warner A.K.
        • Cowart K.C.
        • Benitez A.J.
        • Winchell J.M.
        Detection of Mycoplasma pneumoniae, Chlamydia pneumoniae, and Legionella spp. in clinical specimens using a single-tube multiplex real-time PCR assay.
        Diagn Microbiol Infect Dis. 2011; 70: 1-9
        • Abdeldaim G.M.
        • Stralin K.
        • Korsgaard J.
        • Blomberg J.
        • Welinder-Olsson C.
        • Herrmann B.
        Multiplex quantitative PCR for detection of lower respiratory tract infection and meningitis caused by Streptococcus pneumoniae, Haemophilus influenzae and Neisseria meningitidis.
        BMC Microbiol. 2010; 10: 310
        • Chen C.Y.
        • Ballard R.C.
        The molecular diagnosis of sexually transmitted genital ulcer disease.
        Methods Mol Biol. 2012; 903: 103-112
        • Padmavathy B.
        • Vinoth K.R.
        • Patel A.
        • Deepika S.S.
        • Vaidehi T.
        • Jaffar Ali B.M.
        Rapid and sensitive detection of major uropathogens in a single-pot multiplex PCR assay.
        Curr Microbiol. 2012; 65: 44-53
        • Achermann Y.
        • Vogt M.
        • Leunig M.
        • Wust J.
        • Trampuz A.
        Improved diagnosis of periprosthetic joint infection by multiplex PCR of sonication fluid from removed implants.
        J Clin Microbiol. 2010; 48: 1208-1214
        • Rosey A.L.
        • Abachin E.
        • Quesnes G.
        • et al.
        Development of a broad-range 16S rDNA real-time PCR for the diagnosis of septic arthritis in chirdren.
        J Microbiol Meth. 2007; 68: 88-93
        • Fenollar F.
        • Levy P.Y.
        • Raoult D.
        Usefulness of broad-range PCR for the diagnosis of osteoarticular infections.
        Curr Opin Rheumatol. 2008; 20: 463-470
        • Fink B.
        • Makowiak C.
        • Fuerst M.
        • Berger I.
        • Schafer P.
        • Frommelt L.
        The value of synovial biopsy, joint aspiration and C-reactive protein in the diagnosis of late peri-prosthetic infection of total knee replacements.
        J Bone Joint Surg Br. 2008; 90: 874-878
        • Zannier A.
        • Drancourt M.
        • Franceschi J.P.
        • Aubaniac J.M.
        • Raoult D.
        Value of the technique of cellular lysis by thermic shock in the isolation of bacteria causing osteoarticular infections.
        Pathol Biol (Paris). 1991; 39: 543-546
        • Drancourt M.
        • Bollet C.
        • Carlioz A.
        • Martelin R.
        • Gayral J.P.
        • Raoult D.
        16S ribosomal DNA sequence analysis of a large collection of environmental and clinical unidentifiable bacterial isolates.
        J Clin Microbiol. 2000; 38: 3623-3630
        • Seng P.
        • Rolain J.M.
        • Fournier P.E.
        • La Scola B.
        • Drancourt M.
        • Raoult D.
        MALDI-TOF-mass spectrometry applications in clinical microbiology.
        Future Microbiol. 2010; 5: 1733-1754
        • Drancourt M.
        • Berger P.
        • Raoult D.
        Systematic 16S rRNA gene sequencing of atypical clinical isolates identified 27 new bacterial species associated with humans.
        J Clin Microbiol. 2004; 42: 2197-2202
        • Fihman V.
        • Hannouche D.
        • Bousson V.
        • et al.
        Improved diagnosis specificity in bone and joint infections using molecular techniques.
        J Infect. 2007; 55: 510-517
        • Levy P.Y.
        • Fenollar F.
        The role of molecular diagnostics in implant-associated bone and joint infection.
        Clin Microbiol Infect. 2012; 18: 1168-1175
        • Parvizi J.
        • Zmistowski B.
        • Berbari E.F.
        • et al.
        New definition for periprosthetic joint infection: from the Workgroup of the Musculoskeletal Infection Society.
        Clin Orthop Relat Res. 2011; 469: 2992-2994
        • Brooks Y.
        Microbiology and management of joint and bone infections to anaerobic bacteria.
        J Orthop Sci. 2008; 13: 160-169
        • Levy P.Y.
        • Fenollar F.
        • Stein A.
        • Borrione F.
        • Raoult D.
        Finegoldia magna: a forgotten pathogen in prosthetic joint infection rediscovered by molecular biology.
        Clin Infect Dis. 2009; 49: 1244-1247
        • Levy P.Y.
        • Fenollar F.
        • Stein A.
        • et al.
        Propionibacterium acnes postoperative shoulder arthritis: an emerging clinical entity.
        Clin Infect Dis. 2008; 46: 1884-1886
        • Ceroni D.
        • Cherkaoui A.
        • Ferey S.
        • Kaelin A.
        • Schrenzel J.
        Kingella kingae osteoarticular infections in young children: clinical features and contribution of a new specific real-time PCR assay to the diagnosis.
        J Pediatr Orthop. 2010; 30: 301-304
        • Fournier P.E.
        • Thuny F.
        • Richet H.
        • et al.
        Comprehensive diagnostic strategy for blood culture-negative endocarditis: a prospective study of 819 new cases.
        Clin Infect Dis. 2010; 51: 131-140
        • Rovery C.
        • Greub G.
        • Lepidi H.
        • et al.
        PCR detection of bacteria on cardiac valves of patients with treated bacterial endocarditis.
        J Clin Microbiol. 2005; 43: 163-167
        • Lecouvet F.
        • Irenge L.
        • Vandercam B.
        • Nzeusseu A.
        • Hamels S.
        • Gala J.L.
        The etiologic diagnosis of infectious discitis is improved by amplification-based DNA analysis.
        Arthritis Rheum. 2004; 50: 2985-2994