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Journal of Clinical Microbiology, November 2008, p. 3766-3771, Vol. 46, No. 11
0095-1137/08/$08.00+0     doi:10.1128/JCM.00213-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Analysis of Mixed Sequencing Chromatograms and Its Application in Direct 16S rRNA Gene Sequencing of Polymicrobial Samples{triangledown}

Øyvind Kommedal,1,2* Bjarte Karlsen,3 and Øystein Sæbø3

Department of Microbiology and Immunology, Haukeland University Hospital, Bergen, Norway,1 Section for Microbiology and Immunology, the Gade Institute, University of Bergen, Bergen, Norway,2 iSentio Ltd., Thormøhlensgate 51, Bergen, Norway3

Received 2 February 2008/ Returned for modification 4 April 2008/ Accepted 23 August 2008

Investigation of clinical samples by direct 16S rRNA gene sequencing provides the possibility to detect nonviable bacteria and bacteria with special growth requirements. This approach has been particularly valuable for the diagnosis of patients who have received antibiotics prior to sample collection. In specimens containing more than one bacterium, direct sequencing gives mixed chromatograms that complicate further interpretation. We designed an algorithm able to analyze these ambiguous chromatograms and implemented it as a Web-based service. The algorithm contains both a new base-calling procedure and a new database search procedure. 16S rRNA gene sequencing was performed on polybacterial suspensions prepared in the laboratory. The computer program identified all bacteria correctly to the species level in 23 out of 23 samples containing two different bacteria. For samples containing three different bacteria, correct identification to the species level was achieved for three out of five and to the genus level for five out of five.

* Corresponding author. Mailing address: Department of Microbiology and Immunology, Haukeland University Hospital, 5021 Bergen, Norway. Phone: 47 48 19 26 19. Fax: 47 94 76 59 36. E-mail: oyvind.kommedal{at}isentio.com

{triangledown} Published ahead of print on 3 September 2008.

Journal of Clinical Microbiology, November 2008, p. 3766-3771, Vol. 46, No. 11
0095-1137/08/$08.00+0     doi:10.1128/JCM.00213-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Kommedal, O., Kvello, K., Skjastad, R., Langeland, N., Wiker, H. G. (2009). Direct 16S rRNA Gene Sequencing from Clinical Specimens, with Special Focus on Polybacterial Samples and Interpretation of Mixed DNA Chromatograms. J. Clin. Microbiol. 47: 3562-3568 [Abstract] [Full Text]  


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