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Applied and Environmental Microbiology, December 2004, p. 7220-7228, Vol. 70, No. 12
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.12.7220-7228.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Use of 16S rRNA Gene-Targeted Group-Specific Primers for Real-Time PCR Analysis of Predominant Bacteria in Human Feces

Takahiro Matsuki,* Koichi Watanabe, Junji Fujimoto, Toshihiko Takada, and Ryuichiro Tanaka

Yakult Central Institute for Microbiological Research, Kunitachi, Tokyo, Japan

Received 10 February 2004/ Accepted 21 July 2004

16S rRNA gene-targeted group-specific primers were designed and validated for specific detection and quantification of the Clostridium leptum subgroup and the Atopobium cluster. To monitor the predominant bacteria in human feces by real-time PCR, we used these specific primers together with four sets of group-specific primers for the Clostridium coccoides group, the Bacteroides fragilis group, Bifidobacterium, and Prevotella developed in a previous study (T. Matsuki, K. Watanabe, J. Fujimoto, Y. Miyamoto, T. Takada, K. Matsumoto, H. Oyaizu, and R. Tanaka, Appl. Environ. Microbiol. 68:5445-5451, 2002). Examination of DNA extracted from the feces of 46 healthy adults showed that the C. coccoides group was present in the greatest numbers (log10 10.3 ± 0.3 cells per g [wet weight] [average ± standard deviation]), followed by the C. leptum subgroup (log10 9.9 ± 0.7 cells per g [wet weight]), the B. fragilis group (log10 9.9 ± 0.3 cells per g [wet weight]), Bifidobacterium (log10 9.4 ± 0.7 cells per g [wet weight]), and the Atopobium cluster (log10 9.3 ± 0.7 cells per g [wet weight]). These five bacterial groups were detected in all 46 volunteers. Prevotella was found in only 46% of the subjects at a level of log10 9.7 ± 0.8 cells per g (wet weight). Examination of changes in the population and the composition of the intestinal flora for six healthy adults over an 8-month period revealed that the composition of the flora of each volunteer remained stable throughout the test period.

* Corresponding author. Mailing address: Yakult Central Institute for Microbiological Research, 1796 Yaho, Kunitachi, Tokyo 186-8650, Japan. Phone: 81 (42) 577 8962. Fax: 81 (42) 577 3020. E-mail: takahiro-matsuki{at}yakult.co.jp.

Applied and Environmental Microbiology, December 2004, p. 7220-7228, Vol. 70, No. 12
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.12.7220-7228.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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