Biogeographical distribution and diversity of microbes in methane hydrate-bearing deep marine sediments on the Pacific Ocean Margin

Abstract

The deep subseafloor biosphere is among the least-understood habitats on Earth, even though the huge microbial biomass therein plays an important role for potential long-term controls on global biogeochemical cycles. We report here the vertical and geographical distribution of microbes and their phylogenetic diversities in deeply buried marine sediments of the Pacific Ocean Margins. During the Ocean Drilling Program Legs 201 and 204, we obtained sediment cores from the Peru and Cascadia Margins that varied with respect to the presence of dissolved methane and methane hydrate. To examine differences in prokaryotic distribution patterns in sediments with or without methane hydrates, we studied >2,800 clones possessing partial sequences (400-500 bp) of the 16S rRNA gene and 348 representative clone sequences (approximately 1 kbp) from the two geographically separated subseafloor environments. Archaea of the uncultivated Deep-Sea Archaeal Group were consistently the dominant phylotype in sediments associated with methane hydrate. Sediment cores lacking methane hydrates displayed few or no Deep-Sea Archaeal Group phylotypes. Bacterial communities in the methane hydrate-bearing sediments were dominated by members of the JS1 group, Planctomycetes, and Chloroflexi. Results from cluster and principal component analyses, which include previously reported data from the West and East Pacific Margins, suggest that, for these locations in the Pacific Ocean, prokaryotic communities from methane hydrate-bearing sediment cores are distinct from those in hydrate-free cores. The recognition of which microbial groups prevail under distinctive subseafloor environments is a significant step toward determining the role these communities play in Earth's essential biogeochemical processes.

Fig. 1.

Relative abundances of archaeal 16S rRNA genes (•) and total prokaryotic cells (○) in sediment cores from the Peru and Cascadia Margins. Abundances of archaeal genes were evaluated by Q-PCR of 16S rRNA genes by using domain specific primers and probe sets. Total prokaryotic cell numbers were calculated by fluorescent microscopic counts of cells stained by acridine orange. The depth range of the hydrate stability zone (HSZ) is indicated by the light gray box. The locations of hydrate as confirmed by low temperature anomalies in the samples and by shipboard observations are indicated by gray lines.

Fig. 2.

Phylogenetic community structures based on 16S rRNA gene clone libraries of domains Archaea (A) and Bacteria (B) from ODP sediment core samples. Numbers of clones examined at each depth are indicated in parentheses. The phylogenetic affiliation of each clone sequence was determined by similarity analysis of 400–500 bp of 16S rRNA gene sequences. In each column diagram, the relative abundances of clones classified with the (sub)phylum level are shown. Bacterial community structures at Site 1225 were evaluated by sequencing 200 bp of PCR fragments obtained by denaturing gradient gel electrophoresis (DGGE) analysis, and the number of major bands is indicated in parentheses with asterisk. Phylogenetic trees involving ≈1 kbp sequences of 348 representative phylotypes are shown in Fig. 5.

Fig. 3.

Statistical evaluation of microbial community structures based on clonal frequencies of 16S rRNA gene phylotypes from various marine sediments on the Pacific Ocean Margin. Cluster analysis of archaeal (A) and bacterial (B) communities. Scale bars indicate the square distance determined by the Ward method. The sampling sites, where hydrates are believed to exist nearby (based on seismic surveys), are marked by asterisks, and the sites where methane hydrates were directly observed in the cores are marked by bold colored letters. Shown is principal component (PC) analysis of archaeal (C) and bacterial (D) communities. The percent variance explained by each PC is shown in parentheses. Individual samples are designated in PC space according to the ODP sites from which they were acquired. Additional data points are shown for published clonal frequencies as noted in the text. The sites where methane hydrates were observed in the cores are marked by colored dots.