Searching for predominant soil bacteria: 16S rDNA cloning versus strain cultivation

The predominant bacteria in Dutch grassland soils, as identified by direct DNA extraction, PCR amplification of 16S rDNA and subsequent cloning and se quencing, were compared to the most abundant culturable bacteria. The 16S r DNAs of the strains from a comprehensive cultivation campaign were compared to some of the predominant cloned sequences by temperature gradient gel el ectrophoresis (TGGE). Four ribotypes were selected that were found to be ab undant in the clone library: two closely related Bacillus-like sequences, a representative from the Verrucomicrobiales cluster and an uncultured membe r of the Actinobacteria. Using a variety of cultivation approaches a total of 659 pure cultures were isolated. Initially, approximately 8% of all isol ates matched any of these ribotypes by same migration speed of their 16S rD NA amplicons on TGGE. However, sequencing analysis of matching isolates ind icated that their 16S rDNA sequences were clearly different from the cloned sequences representing the fingerprint bands. Comparing the cultivation ap proach and the molecular 16S rDNA analysis from the same soil sample, there was no correlation between the collection of cultured strains and the 16S rDNA clone library. (C) 1999 Federation of European Microbiological Societi es. Published by Elsevier Science B.V. All rights reserved.