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Applied and Environmental Microbiology, September 1998, p. 3336-3345, Vol. 64, No. 9
0099-2240/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Variations of Bacterial Populations in Human Feces Measured by Fluorescent In Situ Hybridization with Group-Specific 16S rRNA-Targeted Oligonucleotide Probes

Alison H. Franks, Hermie J. M. Harmsen,* Gerwin C. Raangs, Gijsbert J. Jansen, Frits Schut,dagger and Gjalt W. Welling

Department of Medical Microbiology, University of Groningen, 9700 RB Groningen, The Netherlands

Received 7 May 1998/Accepted 23 June 1998

Six 16S rRNA-targeted oligonucleotide probes were designed, validated, and used to quantify predominant groups of anaerobic bacteria in human fecal samples. A set of two probes was specific for species of the Bacteroides fragilis group and the species Bacteroides distasonis. Two others were designed to detect species of the Clostridium histolyticum and the Clostridium lituseburense groups. Another probe was designed for the genera Streptococcus and Lactococcus, and the final probe was designed for the species of the Clostridium coccoides-Eubacterium rectale group. The temperature of dissociation of each of the probes was determined. The specificities of the probes for a collection of target and reference organisms were tested by dot blot hybridization and fluorescent in situ hybridization (FISH). The new probes were used in initial FISH experiments to enumerate human fecal bacteria. The combination of the two Bacteroides-specific probes detected a mean of 5.4 × 1010 cells per g (dry weight) of feces; the Clostridium coccoides-Eubacterium rectale group-specific probe detected a mean of 7.2 × 1010 cells per g (dry weight) of feces. The Clostridium histolyticum, Clostridium lituseburense, and Streptococcus-Lactococcus group-specific probes detected only numbers of cells ranging from 1 × 107 to 7 × 108 per g (dry weight) of feces. Three of the newly designed probes and three additional probes were used in further FISH experiments to study the fecal flora composition of nine volunteers over a period of 8 months. The combination of probes was able to detect at least two-thirds of the fecal flora. The normal biological variations within the fecal populations of the volunteers were determined and indicated that these variations should be considered when evaluating the effects of agents modulating the flora.

* Corresponding author. Mailing address: Department of Medical Microbiology, University of Groningen, P.O. Box 30001, 9700 RB Groningen, The Netherlands. Phone: 31 50 3633510. Fax: 31 50 3633528. E-mail: H.J.M.Harmsen{at}med.rug.nl.

dagger Present address: Microscreen BV, Centre for Microbial Detection and Identification Technology, 9747 AN Groningen, The Netherlands.

Applied and Environmental Microbiology, September 1998, p. 3336-3345, Vol. 64, No. 9
0099-2240/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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