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Journal of Clinical Microbiology, September 2002, p. 3334-3340, Vol. 40, No. 9
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.9.3334-3340.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Detection and Quantification of Oral Treponemes in Subgingival Plaque by Real-Time PCR

Yasuyuki Asai, Takayoshi Jinno, Hajime Igarashi, Yoshinori Ohyama, and Tomohiko Ogawa*

Department of Oral Microbiology, Asahi University School of Dentistry, 1851-1 Hozumi, Hozumi-cho, Motosu-gun, Gifu 501-0296, Japan

Received 12 March 2002/ Returned for modification 28 May 2002/ Accepted 17 June 2002

Oral treponemes have been associated with periodontal diseases. We developed a highly sensitive and specific method to detect and quantify cultivable oral treponemes (Treponema denticola, Treponema vincentii, and Treponema medium) in 50 subgingival plaque samples from 13 healthy subjects as well as 37 patients with periodontal diseases using real-time PCR assays with specific primers and a TaqMan probe for each 16S rRNA sequence. The specificity for each assay was examined by using DNA specimens from various treponemal and other bacterial species. The TaqMan real-time PCR was able to detect from 103 to 108 cells of the oral treponemes, with correlation coefficients as follows: T. denticola, 0.984; T. vincentii, 0.991; and T. medium, 0.984. The frequencies of occurrence of these three oral treponemes in subgingival plaque samples were as follows: T. denticola, 68.0%; T. vincentii, 36.0%; and T. medium, 48.0%. In addition, the number of T. denticola, T. vincentii, and T. medium cells in plaque samples detected by real-time PCR ranged from 3 to 15,184, 1 to 447, and 1 to 7,301 cells/pg of plaque DNA, respectively. Increased numbers of T. denticola cells were detected in plaque samples from deep periodontal pockets, and T. medium was also detected in deep pockets. On the other hand, T. vincentii was mainly found in shallow pockets. These results suggest that various oral treponemes are associated with the formation of each stage of periodontal disease.

* Corresponding author. Mailing address: Department of Oral Microbiology, Asahi University School of Dentistry, 1851-1 Hozumi, Hozumi-cho, Motosu-gun, Gifu 501-0296, Japan. Phone: 81-58-329-1421. Fax: 81-58-329-1421. E-mail: tomo527{at}dent.asahi-u.ac.jp.

Journal of Clinical Microbiology, September 2002, p. 3334-3340, Vol. 40, No. 9
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.9.3334-3340.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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