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Journal of Clinical Microbiology, March 2007, p. 762-770, Vol. 45, No. 3
0095-1137/07/$08.00+0     doi:10.1128/JCM.01342-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Identification of Alpha-Hemolytic Streptococci by Pyrosequencing the 16S rRNA Gene and by Use of VITEK 2{triangledown}

Marjo Haanperä,1* Jari Jalava,1 Pentti Huovinen,1 Olli Meurman,2 and Kaisu Rantakokko-Jalava2

Department of Bacterial and Inflammatory Diseases, National Public Health Institute,1 Clinical Microbiology Laboratory, Turku University Hospital, Turku, Finland2

Received 30 June 2006/ Returned for modification 28 August 2006/ Accepted 26 December 2006

Alpha-hemolytic streptococci are very difficult to identify by phenotypic methods. In this study, a pyrosequencing method for the identification of streptococcal species based on two variable regions of the 16S rRNA gene is described. Almost all studied streptococcal species (n = 51) represented by their type strains could be differentiated except for some closely related species of the Streptococcus bovis or S. salivarius group. The pyrosequencing results of alpha-hemolytic streptococci isolated from blood (n = 99) or from the normal pharyngeal microbiota (n = 25) were compared to the results obtained by the VITEK 2 with GP card (bioMérieux, Marcy l'Etoile, France). As expected, the results of the two methods did not completely agree, but 93 (75.0%) of the isolates assigned to the same streptococcal group by both methods and 57 (46.0%) reached consistent results at the species level. However, 10 strains remained unidentified by VITEK 2, and 4 isolates could not be assigned to any streptococcal group by pyrosequencing. Identification of members of the S. mitis and S. sanguinis groups proved difficult for both methods. Furthermore, the pyrosequencing analysis revealed great sequence variation, since only 43 (32.3%) of the 133 isolates analyzed by pyrosequencing had sequences identical to a type strain. The variation was greatest in the pharyngeal isolates, slightly lower in the blood culture isolates, and nonexistent in invasive pneumococcal isolates (n = 17) that all had the S. pneumoniae type strain sequence. The resolution of the results obtained by the two methods is impeded by the lack of a proper gold standard.

* Corresponding author. Mailing address: Human Microbial Ecology Laboratory, National Public Health Institute, Kiinamyllynkatu 13, FI-20520 Turku, Finland. Phone: 358-2-3316631. Fax: 358-2-3316699. E-mail: marjo.haanpera{at}ktl.fi.

{triangledown} Published ahead of print on 10 January 2007.

Journal of Clinical Microbiology, March 2007, p. 762-770, Vol. 45, No. 3
0095-1137/07/$08.00+0     doi:10.1128/JCM.01342-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



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