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Evaluation of an internally controlled real-time polymerase chain reaction assay targeting the groEL gene for the detection of Bartonella spp. DNA in patients with suspected cat-scratch disease

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Abstract

Bartonella (B.) henselae is the causative agent of cat-scratch disease (CSD), which usually presents as a self-limiting lymphadenopathy. This study reports the development and evaluation of an internally controlled real-time polymerase chain reaction targeting the groEL gene for detection of Bartonella spp. DNA was extracted using the MagNA Pure system. The lower detection limit was 10–100 fg DNA and the in vitro sensitivity of the assay was not affected by duplexing with an internal control PCR. The real-time PCR assay detected DNA from all five B. henselae strains tested, and from B. birtlesii, B. vinsonii subsp. vinsonii, B. vinsonii subsp. arupensis and B. doshiae. The assay generated negative results with a selection of other bacteria, including several Mycobacterium spp., Streptococcus pyogenes and Staphylococcus aureus. Results of real-time PCR in clinical samples were compared with those of a conventional 16S rDNA-based PCR assay. During the period described in the Material and methods section, real-time PCR and conventional 16S PCR were performed on 73 clinical samples. Of these samples, 29 (40%) were found to give positive results and 44 (60%) gave negative results, both by real-time PCR and by conventional PCR, with a 100% agreement between the two tests. The PCR developed in this study is a rapid, sensitive, and simple method for the detection of Bartonella spp. in CSD and is suitable for implementation in the diagnostic laboratory.

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Acknowledgements

Genomic DNA was kindly provided by Dr. R.J. Birtles (Disease Ecology Unit, Centre for Comparative Infectious Diseases, Faculty of Veterinary Science, University of Liverpool, UK). We thank Ingrid Aarts and Caroline de Jong for providing technical support.

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Authors and Affiliations

  1. Regional Laboratory of Public Health Haarlem, Boerhaavelaan 26, 2035 RC, Haarlem, The Netherlands

    B. M. W. Diederen

  2. Laboratory for Medical Microbiology and Immunology, St. Elisabeth Hospital, Tilburg, The Netherlands

    H. Verbakel, A. van der Zee & M. F. Peeters

  3. Paediatric Department, VU Medical Centre, Amsterdam, The Netherlands

    M. J. Vermeulen

  4. Laboratory for Medical Microbiology, Franciscus Hospital, Roosendaal, The Netherlands

    A. Bergmans

  5. Laboratory for Microbiology and Infection Control, Amphia Hospital, Breda, The Netherlands

    A. Bergmans

Authors
  1. B. M. W. Diederen
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  2. M. J. Vermeulen
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  3. H. Verbakel
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  4. A. van der Zee
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  5. A. Bergmans
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  6. M. F. Peeters
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Correspondence to B. M. W. Diederen.

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Diederen, B.M.W., Vermeulen, M.J., Verbakel, H. et al. Evaluation of an internally controlled real-time polymerase chain reaction assay targeting the groEL gene for the detection of Bartonella spp. DNA in patients with suspected cat-scratch disease. Eur J Clin Microbiol Infect Dis 26, 629–633 (2007). https://doi.org/10.1007/s10096-007-0353-x

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  • DOI: https://doi.org/10.1007/s10096-007-0353-x

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