RESUMO
Numerous archaeal lineages are known to inhabit marine subsurface sediments, although their distributions, metabolic capacities, and interspecies interactions are still not well understood. Abundant and diverse archaea were recently reported in Costa Rica (CR) margin subseafloor sediments recovered during IODP Expedition 334. Here, we recover metagenome-assembled genomes (MAGs) of archaea from the CR margin and compare them to their relatives from shallower settings. We describe 31 MAGs of six different archaeal lineages (Lokiarchaeota, Thorarchaeota, Heimdallarchaeota, Bathyarcheota, Thermoplasmatales, and Hadesarchaea) and thoroughly analyze representative MAGs from the phyla Lokiarchaeota and Bathyarchaeota. Our analysis suggests the potential capability of Lokiarchaeota members to anaerobically degrade aliphatic and aromatic hydrocarbons. We show it is genetically possible and energetically feasible for Lokiarchaeota to degrade benzoate if they associate with organisms using nitrate, nitrite, and sulfite as electron acceptors, which suggests a possibility of syntrophic relationships between Lokiarchaeota and nitrite and sulfite reducing bacteria. The novel Bathyarchaeota lineage possesses an incomplete methanogenesis pathway lacking the methyl coenzyme M reductase complex and encodes a noncanonical acetogenic pathway potentially coupling methylotrophy to acetogenesis via the methyl branch of Wood-Ljungdahl pathway. These metabolic characteristics suggest the potential of this Bathyarchaeota lineage to be a transition between methanogenic and acetogenic Bathyarchaeota lineages. This work expands our knowledge about the metabolic functional repertoire of marine benthic archaea.
Assuntos
Archaea/isolamento & purificação , Archaea/metabolismo , Sedimentos Geológicos/microbiologia , Archaea/classificação , Archaea/genética , Bactérias/classificação , Bactérias/genética , Bactérias/isolamento & purificação , Bactérias/metabolismo , Carbono/metabolismo , Ciclo do Carbono , Costa Rica , Sedimentos Geológicos/química , Metagenoma , FilogeniaRESUMO
A degenerate polymerase chain reaction (PCR)-based method of whole-genome amplification, designed to work fluidly with 454 sequencing technology, was developed and tested for use on deep marine subsurface DNA samples. While optimized here for use with Roche 454 technology, the general framework presented may be applicable to other next generation sequencing systems as well (e.g., Illumina, Ion Torrent). The method, which we have called random amplification metagenomic PCR (RAMP), involves the use of specific primers from Roche 454 amplicon sequencing, modified by the addition of a degenerate region at the 3' end. It utilizes a PCR reaction, which resulted in no amplification from blanks, even after 50 cycles of PCR. After efforts to optimize experimental conditions, the method was tested with DNA extracted from cultured E. coli cells, and genome coverage was estimated after sequencing on three different occasions. Coverage did not vary greatly with the different experimental conditions tested, and was around 62% with a sequencing effort equivalent to a theoretical genome coverage of 14.10×. The GC content of the sequenced amplification product was within 2% of the predicted values for this strain of E. coli. The method was also applied to DNA extracted from marine subsurface samples from ODP Leg 201 site 1229 (Peru Margin), and results of a taxonomic analysis revealed microbial communities dominated by Proteobacteria, Chloroflexi, Firmicutes, Euryarchaeota, and Crenarchaeota, among others. These results were similar to those obtained previously for those samples; however, variations in the proportions of taxa identified illustrates well the generally accepted view that community analysis is sensitive to both the amplification technique used and the method of assigning sequences to taxonomic groups. Overall, we find that RAMP represents a valid methodology for amplifying metagenomes from low-biomass samples.
RESUMO
The Brazos-Trinity Basin on the slope of the Gulf of Mexico passive margin was drilled during Integrated Ocean Drilling Progam Expedition 308. The buried anaerobic sediments of this basin are largely organic-poor and have few microbial inhabitants compared with the organic-rich sediments with high cell counts from the Peru Margin that were drilled during Ocean Drilling Program Leg 201. Nucleic acids were extracted from Brazos-Trinity Basin sediments and were subjected to whole-genome amplification and pyrosequencing. A comparison of the Brazos-Trinity Basin metagenome, consisting of 105 Mbp, and the existing Peru Margin metagenome revealed trends linking gene content, phylogenetic content, geological location and geochemical regime. The major microbial groups (Proteobacteria, Firmicutes, Euryarchaeota and Chloroflexi) occur consistently throughout all samples, yet their shifting abundances allow for discrimination between samples. The cluster of orthologous groups category abundances for some classes of genes are correlated with geochemical factors, such as the level of ammonia. Here we describe the sediment metagenome from the oligotrophic Brazos-Trinity Basin (Site 1320) and show similarities and differences with the dataset from the Pacific Peru Margin (Site 1229) and other pyrosequenced datasets. The microbial community found at Integrated Ocean Drilling Program Site 1320 likely represents the subsurface microbial inhabitants of turbiditic slopes that lack substantial upwelling.
Assuntos
Archaea/isolamento & purificação , Bactérias/isolamento & purificação , Sedimentos Geológicos/microbiologia , Metagenômica , Archaea/classificação , Archaea/genética , Oceano Atlântico , Bactérias/classificação , Bactérias/genética , DNA Arqueal/genética , DNA Bacteriano/genética , Sedimentos Geológicos/química , Metagenoma , México , Filogenia , Reação em Cadeia da Polimerase , RNA Ribossômico 16S/genética , Estados UnidosRESUMO
The subseafloor marine biosphere may be one of the largest reservoirs of microbial biomass on Earth and has recently been the subject of debate in terms of the composition of its microbial inhabitants, particularly on sediments from the Peru Margin. A metagenomic analysis was made by using whole-genome amplification and pyrosequencing of sediments from Ocean Drilling Program Site 1229 on the Peru Margin to further explore the microbial diversity and overall community composition within this environment. A total of 61.9 Mb of genetic material was sequenced from sediments at horizons 1, 16, 32, and 50 m below the seafloor. These depths include sediments from both primarily sulfate-reducing methane-generating regions of the sediment column. Many genes of the annotated genes, including those encoding ribosomal proteins, corresponded to those from the Chloroflexi and Euryarchaeota. However, analysis of the 16S small-subunit ribosomal genes suggests that Crenarchaeota are the abundant microbial member. Quantitative PCR confirms that uncultivated Crenarchaeota are indeed a major microbial group in these subsurface samples. These findings show that the marine subsurface is a distinct microbial habitat and is different from environments studied by metagenomics, especially because of the predominance of uncultivated archaeal groups.
Assuntos
Archaea/genética , Bactérias/genética , Sedimentos Geológicos/microbiologia , Água do Mar/microbiologia , Archaea/classificação , Bactérias/classificação , DNA Arqueal/genética , DNA Bacteriano/genética , Geografia , Peru , Filogenia , Reação em Cadeia da Polimerase , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Microbiologia da ÁguaRESUMO
Studies of deeply buried, sedimentary microbial communities and associated biogeochemical processes during Ocean Drilling Program Leg 201 showed elevated prokaryotic cell numbers in sediment layers where methane is consumed anaerobically at the expense of sulfate. Here, we show that extractable archaeal rRNA, selecting only for active community members in these ecosystems, is dominated by sequences of uncultivated Archaea affiliated with the Marine Benthic Group B and the Miscellaneous Crenarchaeotal Group, whereas known methanotrophic Archaea are not detectable. Carbon flow reconstructions based on stable isotopic compositions of whole archaeal cells, intact archaeal membrane lipids, and other sedimentary carbon pools indicate that these Archaea assimilate sedimentary organic compounds other than methane even though methanotrophy accounts for a major fraction of carbon cycled in these ecosystems. Oxidation of methane by members of Marine Benthic Group B and the Miscellaneous Crenarchaeotal Group without assimilation of methane-carbon provides a plausible explanation. Maintenance energies of these subsurface communities appear to be orders of magnitude lower than minimum values known from laboratory observations, and ecosystem-level carbon budgets suggest that community turnover times are on the order of 100-2,000 years. Our study provides clues about the metabolic functionality of two cosmopolitan groups of uncultured Archaea.
Assuntos
Archaea/isolamento & purificação , Ecossistema , Sedimentos Geológicos/microbiologia , Anaerobiose , Archaea/classificação , Archaea/genética , Archaea/metabolismo , Carbono/metabolismo , Metabolismo Energético , Lipídeos/química , Lipídeos/isolamento & purificação , Biologia Marinha , Dados de Sequência Molecular , Peru , Filogenia , RNA Arqueal/genética , RNA Arqueal/isolamento & purificação , RNA Ribossômico 16S/genética , RNA Ribossômico 16S/isolamento & purificaçãoRESUMO
Diverse microbial communities and numerous energy-yielding activities occur in deeply buried sediments of the eastern Pacific Ocean. Distributions of metabolic activities often deviate from the standard model. Rates of activities, cell concentrations, and populations of cultured bacteria vary consistently from one subseafloor environment to another. Net rates of major activities principally rely on electron acceptors and electron donors from the photosynthetic surface world. At open-ocean sites, nitrate and oxygen are supplied to the deepest sedimentary communities through the underlying basaltic aquifer. In turn, these sedimentary communities may supply dissolved electron donors and nutrients to the underlying crustal biosphere.