Community composition of marine bacterioplankton determined by 16S rRNA gene clone libraries and fluorescence in situ hybridization

Abstract

We determined the compositions of bacterioplankton communities in surface waters of coastal California using clone libraries of 16S rRNA genes and fluorescence in situ hybridization (FISH) in order to compare the community structures inferred from these two culture-independent approaches. The compositions of two clone libraries were quite similar to those of clone libraries of marine bacterioplankton examined by previous studies. Clones from gamma-proteobacteria comprised ca. 28% of the libraries, while approximately 55% of the clones came from alpha-proteobacteria, which dominated the clone libraries. The Cytophaga-Flavobacter group and three others each comprised 10% or fewer of the clone libraries. The community composition determined by FISH differed substantially from the composition implied by the clone libraries. The Cytophaga-Flavobacter group dominated 8 of the 11 communities assayed by FISH, including the two communities assayed using clone libraries. On average only 10% of DAPI (4', 6'-diamidino-2-phenylindole)-stained bacteria were detected by FISH with a probe for alpha-proteobacteria, but 30% of DAPI-stained bacteria appeared to be in the Cytophaga-Flavobacter group as determined by FISH. alpha-Proteobacteria were greatly overrepresented in clone libraries compared to their relative abundance determined by FISH, while the Cytophaga-Flavobacter group was underrepresented in clone libraries. Our data show that the Cytophaga-Flavobacter group can be a numerically dominant component of coastal marine bacterioplankton communities.

FIG. 1

Percentages of clones represented by the major phylogenetic groups of bacteria in libraries of 16S rRNA genes in samples collected near Point Sur (station 5) (black bars) and Point Reyes (station 9) (white bars). Clones corresponding to the SAR11 cluster (SAR11), α-proteobacteria (α), β-proteobacteria (β), SAR86 cluster (SAR86), γ-proteobacteria (γ), Cytophaga-Flavobacter group (C.-F.), Planctomyces, gram-positive group (Gram +), and cyanobacteria were identified using oligonucleotide probing and nucleic acid sequence analysis. Eighty-two and 87 clones were screened in the Big Sur and Point Reyes libraries, respectively.

FIG. 2

Percentages of DAPI-stained bacteria detected with the eubacterial probe Eub338 and ³H-leucine incorporation off the coast of California. Error bars are ± 1 SD. Clone libraries were constructed at stations 5 and 9.

FIG. 3

Percentages of DAPI-stained bacteria detected by FISH with probes for α-proteobacteria (α), β-proteobacteria (β), γ-proteobacteria (γ), and the Cytophaga-Flavobacter group (C.-F.). The percentage of DAPI-stained bacteria detected with the probe for eubacteria (Eub338) corresponds to the maximum bar height. The white portions of the bars indicate cells detected with probe Eub338 but not with any group-specific probe (other eubacteria).

FIG. 4

Relationship between compositions of 16S rRNA gene clone libraries and bacterial community compositions in the coastal Pacific Ocean at Point Sur (station 5) (A) and Point Reyes (station 9) (B). Clones and DAPI-stained bacteria were classified as α-proteobacteria (α), β-proteobacteria (β), γ-proteobacteria (γ) and members of the Cytophaga-Flavobacter group (C.-F.).