Comparative 16S rRNA analysis of lake bacterioplankton reveals globally distributed phylogenetic clusters including an abundant group of actinobacteria

Abstract

In a search for cosmopolitan phylogenetic clusters of freshwater bacteria, we recovered a total of 190 full and partial 16S ribosomal DNA (rDNA) sequences from three different lakes (Lake Gossenköllesee, Austria; Lake Fuchskuhle, Germany; and Lake Baikal, Russia). The phylogenetic comparison with the currently available rDNA data set showed that our sequences fall into 16 clusters, which otherwise include bacterial rDNA sequences of primarily freshwater and soil, but not marine, origin. Six of the clusters were affiliated with the alpha, four were affiliated with the beta, and one was affiliated with the gamma subclass of the Proteobacteria; four were affiliated with the Cytophaga-Flavobacterium-Bacteroides group; and one was affiliated with the class Actinobacteria (formerly known as the high-G+C gram-positive bacteria). The latter cluster (hgcI) is monophyletic and so far includes only sequences directly retrieved from aquatic environments. Fluorescence in situ hybridization (FISH) with probes specific for the hgcI cluster showed abundances of up to 1.7 x 10(5) cells ml(-1) in Lake Gossenköllesee, with strong seasonal fluctuations, and high abundances in the two other lakes investigated. Cell size measurements revealed that Actinobacteria in Lake Gossenköllesee can account for up to 63% of the bacterioplankton biomass. A combination of phylogenetic analysis and FISH was used to reveal 16 globally distributed sequence clusters and to confirm the broad distribution, abundance, and high biomass of members of the class Actinobacteria in freshwater ecosystems.

FIG. 1

(A to D) Unrooted phylogenetic consensus tree based on maximum likelihood (FastDNAml) analysis showing the clones in the proposed clusters. (A) α-Proteobacteria; (B) β-Proteobacteria; (C) γ-Proteobacteria and Actinobacteria; (D) CFB phylum. Bifurcations indicate branchings which appeared stable and well separated from neighboring branchings in all cases. Multifurcations indicate tree topologies which could not be unambiguously resolved based on the available data set. For clarity, only the reported clusters are shown; all sequences between the clusters have been omitted. These are included in a more detailed version of the tree which is available at ftp.mpi-bremen.de/pub/molecol_p/fog-aem. The scale bar indicates 10% estimated sequence divergence.

FIG. 1

(A to D) Unrooted phylogenetic consensus tree based on maximum likelihood (FastDNAml) analysis showing the clones in the proposed clusters. (A) α-Proteobacteria; (B) β-Proteobacteria; (C) γ-Proteobacteria and Actinobacteria; (D) CFB phylum. Bifurcations indicate branchings which appeared stable and well separated from neighboring branchings in all cases. Multifurcations indicate tree topologies which could not be unambiguously resolved based on the available data set. For clarity, only the reported clusters are shown; all sequences between the clusters have been omitted. These are included in a more detailed version of the tree which is available at ftp.mpi-bremen.de/pub/molecol_p/fog-aem. The scale bar indicates 10% estimated sequence divergence.

FIG. 1

(A to D) Unrooted phylogenetic consensus tree based on maximum likelihood (FastDNAml) analysis showing the clones in the proposed clusters. (A) α-Proteobacteria; (B) β-Proteobacteria; (C) γ-Proteobacteria and Actinobacteria; (D) CFB phylum. Bifurcations indicate branchings which appeared stable and well separated from neighboring branchings in all cases. Multifurcations indicate tree topologies which could not be unambiguously resolved based on the available data set. For clarity, only the reported clusters are shown; all sequences between the clusters have been omitted. These are included in a more detailed version of the tree which is available at ftp.mpi-bremen.de/pub/molecol_p/fog-aem. The scale bar indicates 10% estimated sequence divergence.

FIG. 1

(A to D) Unrooted phylogenetic consensus tree based on maximum likelihood (FastDNAml) analysis showing the clones in the proposed clusters. (A) α-Proteobacteria; (B) β-Proteobacteria; (C) γ-Proteobacteria and Actinobacteria; (D) CFB phylum. Bifurcations indicate branchings which appeared stable and well separated from neighboring branchings in all cases. Multifurcations indicate tree topologies which could not be unambiguously resolved based on the available data set. For clarity, only the reported clusters are shown; all sequences between the clusters have been omitted. These are included in a more detailed version of the tree which is available at ftp.mpi-bremen.de/pub/molecol_p/fog-aem. The scale bar indicates 10% estimated sequence divergence.

FIG. 2

Seasonal dynamics of the abundance and biomass of the β-Proteobacteria and the Actinobacteria counted with probes HGC69a, HG1-840, and BET42a. The horizontal bar indicates the period of ice cover.

FIG. 3

FISH of samples from Lake Gossenköllesee. DAPI (left) and epifluorescence (right) micrographs are shown for identical microscopic fields. (A) In situ hybridization with probe EUB338 labeled with Cy3. The arrow indicates a representative of a population of small, dim cells. (B) In situ hybridization with probe HGC664 labeled with Cy3. (C) In situ hybridization with probe HG1-840 labeled with Cy3. Bar, 10 μm (all panels).