Archives of Microbiology

, Volume 168, Issue 3, pp 185–192

Analysis of bacterial community structure in bulk soil by in situ hybridization

  • Boris Zarda
  • D. Hahn
  • Antonis Chatzinotas
  • Wilhelm Schönhuber
  • Alexander Neef
  • Rudolf I. Amann
  • Josef Zeyer
Original paper

DOI: 10.1007/s002030050486

Cite this article as:
Zarda, B., Hahn, D., Chatzinotas, A. et al. Arch Microbiol (1997) 168: 185. doi:10.1007/s002030050486

Abstract

In situ hybridization with rRNA-targeted, fluorescent (Cy3-labeled) oligonucleotide probes was used to analyze bacterial community structure in ethanol- or paraformaldehyde-fixed bulk soil after homogenization of soil samples in 0.1% pyrophosphate by mild ultrasonic treatment. In ethanol-fixed samples 37 ± 7%, and in paraformaldehyde 41 ± 8% of the 4′, 6-diamidino-2-phenylindole(DAPI)-stained cells were detected with the bacterial probe Eub338. The yield could not be increased by enzymatic and/or chemical pretreatments known to enhance the permeability of bacterial cells for probes. However, during storage in ethanol for 7 months, the detectability of bacteria increased in both ethanol- and paraformaldehyde-fixed samples to up to 47 ± 8% due to an increase in the detection yield of members of the α-subdivision of Proteobacteria from 2 ± 1% to 10 ± 3%. Approximately half of the bacteria detected by probe Eub338 could be affiliated to major phylogenetic groups such as the α-, β-, γ-, and δ-subdivisions of Proteobacteria, gram-positive bacteria with a high G+C DNA content, bacteria of the Cytophaga-Flavobacterium cluster of the CFB phylum, and the planctomycetes. The analysis revealed that bacteria of the α- and δ-subdivision of Proteobacteria and the planctomycetes were predominant. Here, members of the α-subdivision of Proteobacteria accounted for approximately 10 ± 3% of DAPI-stained cells, which corresponded to 44 ± 16 × 108 cells (g soil, dry wt.)–1, while members of the δ-subdivision of Proteobacteria made up 4 ± 2% of DAPI-stained cells [17 ± 9 × 108 cells (g soil, dry wt.)–1]. A large population of bacteria in bulk soil was represented by the planctomycetes, which accounted for 7 ± 3% of DAPI-stained cells [32 ± 12 × 108 cells (g soil, dry wt.)–1]. The detection of planctomycetes in soil confirms previous reports on the occurrence of planctomycetes in soil and indicates a yet unknown ecological significance of this group, which to date has never been isolated from terrestrial environments.

Key words Fluorescent oligonucleotide probesPlanctomycetesrRNAWhole-cell hybridization

Copyright information

© Springer-Verlag Berlin Heidelberg 1997

Authors and Affiliations

  • Boris Zarda
    • 1
  • D. Hahn
    • 1
  • Antonis Chatzinotas
    • 1
  • Wilhelm Schönhuber
    • 2
  • Alexander Neef
    • 2
  • Rudolf I. Amann
    • 2
  • Josef Zeyer
    • 1
  1. 1.Swiss Federal Institute of Technology (ETH Zürich), Institute of Terrestrial Ecology, Soil Biology, Grabenstrasse 3, CH-8952 Schlieren, Switzerland Tel. +41-1-633-6039; Fax +41-1-633-1122 e-mail: hahn@ito.umnw.ethz.chCH
  2. 2.Technische Universität München, Lehrstuhl für Mikrobiologie, D-80290 München, GermanyDE