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Applied and Environmental Microbiology, October 1998, p. 3854-3859, Vol. 64, No. 10
0099-2240/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Temperature Gradient Gel Electrophoresis Analysis of 16S rRNA from Human Fecal Samples Reveals Stable and Host-Specific Communities of Active Bacteria

Erwin G. Zoetendal, Antoon D. L. Akkermans,* and Willem M. De Vos

Laboratory of Microbiology, Department of Biomolecular Sciences, Wageningen Agricultural University, 6703 CT Wageningen, The Netherlands

Received 1 April 1998/Accepted 16 July 1998

The diversity of the predominant bacteria in the human gastrointestinal tract was studied by using 16S rRNA-based approaches. PCR amplicons of the V6 to V8 regions of fecal 16S rRNA and ribosomal DNA (rDNA) were analyzed by temperature gradient gel electrophoresis (TGGE). TGGE of fecal 16S rDNA amplicons from 16 individuals showed different profiles, with some bands in common. Fecal samples from two individuals were monitored over time and showed remarkably stable profiles over a period of at least 6 months. TGGE profiles derived from 16S rRNA and rDNA amplicons showed similar banding patterns. However, the intensities of bands with similar mobilities differed in some cases, indicating a different contribution to the total active fraction of the prominent fecal bacteria. Most 16S rRNA amplicons in the TGGE pattern of one subject were identified by cloning and sequence analysis. Forty-five of the 78 clones matched 15 bands, and 33 clones did not match any visible band in the TGGE pattern. Nested PCR of amplified 16S rDNA indicated preferential amplification of a sequence corresponding to 12 of the 33 nonmatching clones with similar mobilities in TGGE. The sequences matching 15 bands in the TGGE pattern showed 91.5 to 98.7% homology to sequences derived from different Clostridium clusters. Most of these were related to strains derived from the human intestine. The results indicate that the combination of cloning and TGGE analysis of 16S rDNA amplicons is a reliable approach to monitoring different microbial communities in feces.

* Corresponding author. Mailing address: Laboratory of Microbiology, Wageningen Agricultural University, Hesselink van Suchtelenweg 4, 6703 CT, Wageningen, The Netherlands. Phone: 31 317 483486. Fax: 31 317 483829. E-mail: antoon.akkermans{at}algemeen.micr.wau.nl.

Applied and Environmental Microbiology, October 1998, p. 3854-3859, Vol. 64, No. 10
0099-2240/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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