British Journal of Nutrition

Research Article

Molecular biological methods for studying the gut microbiota: the EU human gut flora project

M. Blauta1 c1, M.D. Collinsa2, G. W. Wellinga3, J. Doréa4, J. van Looa5 and W. de Vosa6

a1 German Institute of Human Nutrition, Potsdam-Rehbruecke, Department of Gastrointestinal Microbiology, Arthur-Scheunert-Allee 114–116, 14558 Bergholz-Rehbruecke, Germany

a2 Food Microbial Sciences Unit, School of Food Biosciences, The University of Reading, PO Box 226, Whiteknights, Reading RG6 6AP, UK

a3 Rijksuniversiteit Groningen, Laboratorium voor Medische Microbiologie, Hanzeplein 1, 9713 GZ Groningen, The Netherlands

a4 Institut National de la Recherche Agronomique, Laboratoire d'Ecologie et de Physiologie du Système Digestif, Domaine de Vilvert, 78352 Jouy-en-Josas Cedex, France

a5 ORAFTI, Aandorenstraat 1, 3300 Tienen, Belgium

a6 Wageningen University, Laboratory of Microbiology, Hesselink van Suchtelenweg 4, Wageningen 6703 CT, The Netherlands

Abstract

Seven European laboratories co-operated in a joint project (FAIR CT97-3035) to develop, refine and apply molecular methods towards facilitating elucidation of the complex composition of the human intestinal microflora and to devise robust methodologies for monitoring the gut flora in response to diet. An extensive database of 16S rRNA sequences for tracking intestinal bacteria was generated by sequencing the 16S rRNA genes of new faecal isolates and of clones obtained by amplification with polymerase chain reaction (PCR) on faecal DNA from subjects belonging to different age groups. The analyses indicated that the number of different species (diversity) present in the human gut increased with age. The sequence information generated, provided the basis for design of 16S rRNA-directed oligonucleotide probes to specifically detect bacteria at various levels of phylogenetic hierarchy. The probes were tested for their specificity and used in whole-cell and dot-blot hybridisations. The applicability of the developed methods was demonstrated in several studies and the major outcomes are described.

Correspondence:

c1 *Corresponding author: Professor M. Blaut, tel +49 33 20 08 84 70, fax +49 33 20 08 84 07, email Blaut@www.dife.de

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