Print PagePhototrophic microbial mats that develop in the effluent channels of hot springs in Yellowstone National Park serve as models to investigate how microbial community structures are related to community functions, given their relatively limited diversity and their tractability for studying organismal interactions. It was of interest to determine whether whole community metagenomic sequencing could provide evidence revealing the functional roles of community members previously known only by small subunit ribosomal RNA (16S rRNA) sequencing. We obtained 2,355 16S rRNA clone sequences and 320 Mbp of Sanger sequence from clones randomly produced from DNA extracted from six springs. Community structure differences among springs were determined with a distance metric produced from the phylogenetic analysis of 16S rRNA sequences, and these differences correlated with geochemical parameters. Overlapping metagenomic sequences were assembled into contiguous genomic scaffolds to represent the genomes of uncultivated community members, and predicted genes were annotated. Phylogenetic marker genes and genes putatively involved in phototrophy, carbon fixation, and sulfur cycling were identified. Two methods were used to taxonomically group metagenomic sequences; i) metagenome sequences were aligned to reference genomes by BLAST recruitment, and ii) the phylogenetic signatures inherent in the oligonucleotide frequencies of scaffolds and of reference genomes were analyzed.
Results/Conclusions
A diverse range of phototrophic bacteria encompassing multiple metabolic functional strategies were found to inhabit these springs, including oxygenic cyanobacteria and anoxygenic phototrophs such as those belonging to Kingdom Chloroflexi, green-sulfur bacteria (Order Chlorobiales), purple phototrophic proteobacteria, and phototrophic acidobacteria. Differences in community structure could in part be attributed to differences in spring geochemistry. For example, Bath Lake Vista Annex Spring contained high levels of sulfide (117 μM), which caused it to be completely devoid of cyanobacteria due to their physiological constraints. Many closely-related 16S rRNA-defined clades within the Chloroflexi exhibited site-specificity, suggesting that some organisms are uniquely associated with particular springs. The metagenomic context of phylogenetic marker and functional genes within assembled scaffolds allowed for the discovery of novel phototrophic organisms within Kingdoms Chloroflexi and Chlorobi. Functional guilds were identified for a community structure model of mats from alkaline siliceous springs for which there was previous sequencing coverage available. Comparisons of the functional roles of organisms inhabiting various communities gave evidence for unique guild structures in geochemically dissimilar springs, which may have implications for differences in guild-level interactions and rates of biogeochemical cycling.
