Skip to main content
SpringerLink
Log in

Direct detection of pathogens in osteoarticular infections by polymerase chain reaction amplification and microarray hybridization

  • Original Article
  • Published:
Journal of Orthopaedic Science

Abstract

Background

Molecular biological techniques such as the polymerase chain reaction (PCR) and DNA microarray are used for the detection/identification of microorganisms; however, few reports have discussed the clinical utility of microarray analysis for identification of causative organisms of osteoarticular infections. It is important to examine the utility of PCR amplification followed by analysis of DNA microarray carrying specific oligonucleotides.

Methods

This study included 101 biological samples obtained from 96 patients who underwent conservative and/or surgical treatment for osteoarticular infections. In this double-blind comparative study, routine conventional testing and the research groups were unaware of each other’s interpretation until identical specimens were identified by culture and microarray analysis.

Results

Results of PCR microarray analysis were positive for 25 samples and negative for the remaining 76 samples within 24 h, and the results of the cultures (available after a mean of 3.54 days) were positive in 26 samples and negative for the remaining 75 samples. The sensitivity of microarray analysis was 84.6% (22/26) and specificity was 88.0% (22/25). Discrepant results were identified in seven samples, including a negative culture and a positive microarray in three cases and a positive culture and a negative microarray in four other cases.

Conclusions

The PCR microarray analysis is complementary to routine cultures in identifying causative microorganisms and should be used in patients with highly suspected infections and negative bacterial culture and in patients who require prompt diagnosis and early initiation of antibiotic therapy.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Lew DP, Waldvogel FA. Osteomyelitis. Lancet 2004;364:369–379.

    Article  PubMed  CAS  Google Scholar 

  2. Shirtliff ME, Mader JT. Acute septic arthritis. Clin Microbiol Rev 2002;15:527–544.

    Article  PubMed  Google Scholar 

  3. Zimmerli W, Trampuz A, Ochsner PE. Prosthetic-joint infections. N Engl J Med 2004;351:1645–1654.

    Article  PubMed  CAS  Google Scholar 

  4. Gupta MN, Sturrock RD, Field M. Prospective comparative study of patients with culture proven and high suspicion of adult onset septic arthritis. Ann Rheum Dis 2003;62:327–331.

    Article  PubMed  CAS  Google Scholar 

  5. Kaandorp CJ, Krijnen P, Moens HJ, Habbema JD, van Schaardenburg D. The outcome of bacterial arthritis: a prospective community-based study. Arthritis Rheum 1997;40:884–892.

    Article  PubMed  CAS  Google Scholar 

  6. Uchida K, Kobayashi S, Sato R, Nakajima H, Yayama T, Kokubo Y, et al. Anterior spinal artery syndrome complicating massive paravertebral abscess. J Orthop Sci 2008;13:85–88.

    Article  PubMed  Google Scholar 

  7. Tunney MM, Patrick S, Curran MD, Ramage G, Hanna D, Nixon JR, et al. Detection of prosthetic hip infection at revision arthropathy by immunofluorescence microscopy and PCR amplification of the bacterial 16S rRNA gene. J Clin Microbiol 1999;37:3281–3290.

    PubMed  CAS  Google Scholar 

  8. Patel JB. 16S rRNA gene sequencing for bacterial pathogen identification in the clinical laboratory. Mol Diagn 2001;6:313–321.

    PubMed  CAS  Google Scholar 

  9. Yang S, Rothman RE. PCR-based diagnostics for infectious disease: uses, limitations, and future applications in acute-care settings. Lancet Infect Dis 2004;4:337–348.

    Article  PubMed  CAS  Google Scholar 

  10. 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.

    Article  PubMed  CAS  Google Scholar 

  11. Kobayashi N, Bauer TW, Togawa D, Lieberman IH, Sakai H, Fujishiro T, et al. A molecular gram stain using broad range PCR and pyrosequencing technology: a potentially useful tool for diagnosing orthopaedic infections. Diagn Mol Pathol 2005;14:83–89.

    Article  PubMed  CAS  Google Scholar 

  12. Anthony RM, Brown TJ, French GL. DNA array technology and diagnostic microbiology. Expert Rev Mol Diagn 2001;1:30–38.

    Article  PubMed  CAS  Google Scholar 

  13. Tillib SV, Mirzabekov AD. Advances in the analysis of DNA sequence variations using oligonucleotide microchip technology. Curr Opin Biotechnol 2001;12:53–58.

    Article  PubMed  CAS  Google Scholar 

  14. Anthony RM, Brown TJ, French GL. Rapid diagnosis of bacteremia by universal amplification of 23S ribosomal DNA followed by hybridization to an oligonucleotide array. J Clin Microbiol 2000;38:781–788.

    PubMed  CAS  Google Scholar 

  15. Mitterer G, Huber M, Leidinger E, Kirisits C, Lubitz W, Muellar MW, Schmidt WM. Microarray-based identification of bacteria in clinical samples by solid-phase PCR amplification of 23S ribosomal DNA sequences. J Clin Microbiol 2004;42:1048–1057.

    Article  PubMed  CAS  Google Scholar 

  16. Hall GS. Molecular applications in the clinical laboratory. In: Mahon CR, Manuselis G, editors. Diagnostic microbiology. 2nd edn. Philadelphia: Saunders; 2000. p. 191–210.

    Google Scholar 

  17. Al Zahrani K, Al Jahdali H, Poirier L, René P, Gennaro ML, Menzies D. Accuracy and utility of commercially available amplification and serologic tests for the diagnosis of minimal pulmonary tuberculosis. Am J of Respir Crit Care Med 2000;162:1323–1329.

    Google Scholar 

  18. Berndt C, Müller U, Bergmann F, Schmitt U, Kaiser R, Müller C. Comparison between a nucleic acid sequence-based amplification and branched DNA test for quantifying HIV RNA load in blood plasma. J Virol Methods 2000;89:177–181.

    Article  PubMed  CAS  Google Scholar 

  19. Uchida K, Baba H, Wada M, Maezawa Y, Kikukawa Y, Imura S. Atypical mycobacterium osteomyelitis of the humerus in a child with chronic granulomatous disease. J Neurol Orthop Med Surg 1996;17:127–129.

    Google Scholar 

  20. Jordan JA, Durso MB. Comparison of 16S rRNA gene PCR and BACTEC 9240 for detection of neonatal bacteremia. J Clin Microbiol 2000;38:2574–2578.

    PubMed  CAS  Google Scholar 

  21. Kobayashi N, Procop GW, Krebs V, Kobayashi H, Bauer TW. Molecular identification of bacteria from aseptically loose implants. Clin Orthop 2008;466:1716–1725.

    Article  PubMed  Google Scholar 

  22. Weisburg WG, Barns SM, Pelletier DA, Lane DJ. 16S Ribosomal DNA amplification for phylogenetic study. J Bacteriol 1991;173:697–703.

    PubMed  CAS  Google Scholar 

  23. Carroll NM, Jaeger EE, Choudhury S, Dunlop AA, Matheson MM, Adamson P, et al. Detection of and discrimination between gram-positive and gram-negative bacteria in intraocular samples by using nested PCR. J Clin Microbiol 2000;38:1753–1757.

    PubMed  CAS  Google Scholar 

  24. Xu J, Millar BC, Moore JE, Murphy K, Webb H, Fox AJ, et al. Employment of broad-range 16S rRNA PCR to detect aetiological agents of infection from clinical specimens in patients with acute meningitis: rapid separation of 16S rRNA PCR amplicon without the need for cloning. J Appl Microbiol 2003;94:197–206.

    Article  PubMed  CAS  Google Scholar 

  25. Tsai JC, Teng LJ, Hsueh PR. Direct detection of bacterial pathogens in brain abscesses by polymerase chain reaction amplification and sequencing of partial 16S ribosomal deoxyribonucleic acid fragments. Neurosurgery 2004;55:1154–1162.

    Article  PubMed  Google Scholar 

  26. Wilbrink B, van der Heijden IM, Schouls LM, van Embden JD, Hazes JM, Breedveld FC, et al. Detection of bacterial DNA in joint samples from patients with undifferentiated arthritis and reactive arthritis, using polymerase chain reaction with universal 16S ribosomal RNA primers. Arthritis Rheum 1998;41:535–543.

    Article  PubMed  CAS  Google Scholar 

  27. Jackson CJ, Barton RC, Evans EG. Species identification and strain differentiation of dermatophyte fungi by analysis of ribosomal-DNA intergenic spacer regions. J Clin Microbiol 1999;37:931–936.

    PubMed  CAS  Google Scholar 

  28. Morace G, Sanguinetti M, Posteraro B, Lo Cascio L, Fadda G. Identification of various medically important Candida species in clinical specimens by PCR-restriction enzyme analysis. J Clin Microbiol 1997;35:667–672.

    PubMed  CAS  Google Scholar 

  29. Henry T, Iwen PC, Hinrichs SH. Identification of Aspergillus species using internal transcribed spacer regions 1 and 2. J Clin Microbiol 2000;38:1510–1515.

    PubMed  CAS  Google Scholar 

  30. Fujita S, Senda Y, Nakaguchi S, Hashimoto T. Multiplex PCR using internal transcribed spacer 1 and 2 regions for rapid detection and identification of yeast strains. J Clin Microbiol 2001;39:3617–3622.

    Article  PubMed  CAS  Google Scholar 

  31. Turenne CY, Sanche SE, Hoban DJ, Karlowsky JA, Kabani AM. Rapid identification of fungi by using the ITS2 genetic region and an automated fluorescent capillary electrophoresis system. J Clin Microbiol 1999;37:1846–1851.

    PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Orthopaedics and Rehabilitation Medicine, Fukui University Faculty of Medical Sciences, Matsuoka Shimoaizuki 23, Eiheiji, Fukui, 910-1193, Japan

    Kenzo Uchida, Takafumi Yayama, Yasuo Kokubo, Tsuyoshi Miyazaki, Hideaki Nakajima, Kohei Negoro, Kenichi Takeno, Elisa S. Mwaka, Norbert T. Orwotho, Shigeru Kobayashi & Hisatoshi Baba

  2. Department of Orthopaedic Surgery, Makerere University Medical School, Kampala, Republic of Uganda

    Elisa S. Mwaka & Norbert T. Orwotho

  3. R&D and Business Planning Department, Mitsubishi Chemical Medience Corporation, Tokyo, Japan

    Mitsunobu Shimadzu

Authors
  1. Kenzo Uchida
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Takafumi Yayama
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Yasuo Kokubo
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Tsuyoshi Miyazaki
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Hideaki Nakajima
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Kohei Negoro
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Kenichi Takeno
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Elisa S. Mwaka
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Norbert T. Orwotho
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Mitsunobu Shimadzu
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Shigeru Kobayashi
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Hisatoshi Baba
    View author publications

    You can also search for this author in PubMed Google Scholar

About this article

Cite this article

Uchida, K., Yayama, T., Kokubo, Y. et al. Direct detection of pathogens in osteoarticular infections by polymerase chain reaction amplification and microarray hybridization. J Orthop Sci 14, 471–483 (2009). https://doi.org/10.1007/s00776-009-1373-4

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00776-009-1373-4

Keywords

Search

Navigation