AbstractMicrobial life in the deep subsurface biosphere is taxonomically and metabolically diverse, but it is vigorously debated whether the resident organisms are thriving (metabolizing, maintaining cellular integrity, and expressing division genes) or just surviving. As part of Integrated Ocean Drilling Program (IODP) Expedition 347: Baltic Sea Paleoenvironment, we extracted and sequenced RNA from organic carbon-rich, nutrient-replete, and permanently anoxic sediment. In stark contrast to the oligotrophic subsurface biosphere, Baltic Sea Basin samples provided a unique opportunity to understand the balance between metabolism and other cellular processes. Targeted sequencing of 16S rRNA transcripts showed Atribacteria (an uncultured phylum) and Chloroflexi to be among the dominant and the active members of the community. Metatranscriptomic analysis identified methane cycling, sulfur cycling, and halogenated compound utilization as active in situ respiratory metabolisms. Genes for cellular maintenance, cellular division, motility, and antimicrobial production were also transcribed. This indicates that microbial life in deep subsurface Baltic Sea Basin sediments was not only alive, but thriving.
AbstractSulfate reducing bacteria (SRB) oxidize a significant proportion of subseafloor organic carbon, but their metabolic activities and subsistence mechanisms are poorly understood. Here, we report in depth phylogenetic and metabolic analyses of SRB transcripts in the Peru Margin subseafloor and interpret these results in the context of sulfate reduction activity in the sediment. Relative abundance of overall SRB gene transcripts declines strongly whereas relative abundance of ribosomal protein transcripts from sulfate reducing δ‐Proteobacteria peak at 90 m below seafloor (mbsf) within a deep sulfate methane transition zone. This coincides with isotopically heavy δ34S values of pore water sulfate (70‰), indicating active subseafloor microbial sulfate reduction. Within the shallow sulfate reduction zone (0–5 mbsf), a transcript encoding the beta subunit of dissimilatory sulfite reductase (dsrB) was related to Desulfitobacterium dehalogenans and environmental sequences from Aarhus Bay (Denmark). At 159 mbsf we discovered a transcript encoding the reversely operating dissimilatory sulfite reductase α‐subunit (rdsrA), with basal phylogenetic relation to the chemolithoautotrophic SUP05 Group II clade. A diversity of SRB transcripts involved in cellular maintenance point toward potential subsistence mechanisms under low‐energy over long time periods, and provide a detailed new picture of SRB activities underlying sulfur cycling in the deep subseafloor.
AbstractViruses are highly abundant in marine subsurface sediments and can even exceed the number of prokaryotes. However, their activity and quantitative impact on microbial populations are still poorly understood. Here, we use gene expression data from published continental margin subseafloor metatranscriptomes to qualitatively assess viral diversity and activity in sediments up to 159 metres below seafloor (mbsf). Mining of the metatranscriptomic data revealed 4651 representative viral homologues (RVHs), representing 2.2% of all metatranscriptome sequence reads, which have close translated homology (average 77%, range 60–97% amino acid identity) to viral proteins. Archaea‐infecting RVHs are exclusively detected in the upper 30 mbsf, whereas RVHs for filamentous inoviruses predominate in the deepest sediment layers. RVHs indicative of lysogenic phage–host interactions and lytic activity, notably cell lysis, are detected at all analysed depths and suggest a dynamic virus–host association in the marine deep biosphere studied here. Ongoing lytic viral activity is further indicated by the expression of clustered, regularly interspaced, short palindromic repeat‐associated cascade genes involved in cellular defence against viral attacks. The data indicate the activity of viruses in subsurface sediment of the Peruvian margin and suggest that viruses indeed cause cell mortality and may play an important role in the turnover of subseafloor microbial biomass.
AbstractA method for the extraction of nucleic acids from a wide range of environmental samples was developed. This method consists of several modules, which can be individually modified to maximize yields in extractions of DNA and RNA or separations of DNA pools. Modules were designed based on elaborate tests, in which permutations of all nucleic acid extraction steps were compared. The final modular protocol is suitable for extractions from igneous rock, air, water, and sediments. Sediments range from high-biomass, organic rich coastal samples to samples from the most oligotrophic region of the world's oceans and the deepest borehole ever studied by scientific ocean drilling. Extraction yields of DNA and RNA are higher than with widely used commercial kits, indicating an advantage to optimizing extraction procedures to match specific sample characteristics. The ability to separate soluble extracellular DNA pools without cell lysis from intracellular and particle-complexed DNA pools may enable new insights into the cycling and preservation of DNA in environmental samples in the future. A general protocol is outlined, along with recommendations for optimizing this general protocol for specific sample types and research goals.
AbstractThe ability of microorganisms to withstand long periods with extremely low energy input has gained increasing scientific attention in recent years. Starvation experiments in the laboratory have shown that a phylogenetically wide range of microorganisms evolve fitness-enhancing genetic traits within weeks of incubation under low-energy stress. Studies on natural environments that are cut off from new energy supplies over geologic time scales, such as deeply buried sediments, suggest that similar adaptations might mediate survival under energy limitation in the environment. Yet, the extent to which laboratory-based evidence of starvation survival in pure or mixed cultures can be extrapolated to sustained microbial ecosystems in nature remains unclear. In this review, we discuss past investigations on microbial energy requirements and adaptations to energy limitation, identify gaps in our current knowledge, and outline possible future foci of research on life under extreme energy limitation.
AbstractQuantifying the rates of biogeochemical processes in marine sediments is essential for understanding global element cycles and climate change. Because organic matter degradation is the engine behind benthic dynamics, deciphering the impact that various forces have on this process is central to determining the evolution of the Earth system. Therefore, recent developments in the quantitative modeling of organic matter degradation in marine sediments are critically reviewed. The first part of the review synthesizes the main chemical, biological and physical factors that control organic matter degradation in sediments while the second part provides a general review of the mathematical formulations used to model these processes and the third part evaluates their application over different spatial and temporal scales. Key transport mechanisms in sedimentary environments are summarized and the mathematical formulation of the organic matter degradation rate law is described in detail. The roles of enzyme kinetics, bioenergetics, temperature and biomass growth in particular are highlighted. Alternative model approaches that quantify the degradation rate constant are also critically compared. In the third part of the review, the capability of different model approaches to extrapolate organic matter degradation rates over a broad range of temporal and spatial scales is assessed. In addition, the structure, functions and parameterization of more than 250 published models of organic matter degradation in marine sediments are analyzed. The large range of published model parameters illustrates the complex nature of organic matter dynamics, and, thus, the limited transferability of these parameters from one site to another. Compiled model parameters do not reveal a statistically significant correlation with single environmental characteristics such as water depth, deposition rate or organic matter flux. The lack of a generic framework that allows for model parameters to be constrained in data-poor areas seriously limits the quantification of organic matter degradation on a global scale. Therefore, we explore regional patterns that emerge from the compiled more than 250 organic matter rate constants and critically discuss them in their environmental context. This review provides an interdisciplinary view on organic matter degradation in marine sediments. It contributes to an improved understanding of global patterns in benthic organic matter degradation, and helps identify outstanding questions and future directions in the modeling of organic matter degradation in marine sediments.
AbstractHalf of the microbial cells in the Earth’s oceans are found in sediments. Many of these cells are members of the Archaea, single-celled prokaryotes in a domain of life separate from Bacteria and Eukaryota. However, most of these archaea lack cultured representatives, leaving their physiologies and placement on the tree of life uncertain. Here we show that the uncultured miscellaneous crenarchaeotal group (MCG) and marine benthic group-D (MBG-D) are among the most numerous archaea in the marine sub-sea floor. Single-cell genomic sequencing of one cell of MCG and three cells of MBG-D indicated that they form new branches basal to the archaeal phyla Thaumarchaeota and Aigarchaeota, for MCG, and the order Thermoplasmatales, for MBG-D. All four cells encoded extracellular protein-degrading enzymes such as gingipain and clostripain that are known to be effective in environments chemically similar to marine sediments. Furthermore, we found these two types of peptidase to be abundant and active in marine sediments, indicating that uncultured archaea may have a previously undiscovered role in protein remineralization in anoxic marine sediments.
AbstractMicrobial communities can subsist at depth in marine sediments without fresh supply of organic matter for millions of years. At threshold sedimentation rates of 1 millimeter per 1000 years, the low rates of microbial community metabolism in the North Pacific Gyre allow sediments to remain oxygenated tens of meters below the sea floor. We found that the oxygen respiration rates dropped from 10 micromoles of O2 liter−1 year−1 near the sediment-water interface to 0.001 micromoles of O2 liter−1 year−1 at 30-meter depth within 86 million-year-old sediment. The cell-specific respiration rate decreased with depth but stabilized at around 10−3 femtomoles of O2 cell−1 day−1 10 meters below the seafloor. This result indicated that the community size is controlled by the rate of carbon oxidation and thereby by the low available energy flux.
AbstractTwo decades of scientific ocean drilling have demonstrated widespread microbial life in deep sub-seafloor sediment, and surprisingly high microbial-cell numbers. Despite the ubiquity of life in the deep biosphere, the large community sizes and the low energy fluxes in this vast buried ecosystem are not yet understood. It is not known whether organisms of the deep biosphere are specifically adapted to extremely low energy fluxes or whether most of the observed cells are in a dormant, spore-like state. Here we apply a new approach—the D:L-amino-acid model—to quantify the distributions and turnover times of living microbial biomass, endospores and microbial necromass, as well as to determine their role in the sub-seafloor carbon budget. The approach combines sensitive analyses of unique bacterial markers (muramic acid and D-amino acids) and the bacterial endospore marker, dipicolinic acid, with racemization dynamics of stereo-isomeric amino acids. Endospores are as abundant as vegetative cells and microbial activity is extremely low, leading to microbial biomass turnover times of hundreds to thousands of years. We infer from model calculations that biomass production is sustained by organic carbon deposited from the surface photosynthetic world millions of years ago and that microbial necromass is recycled over timescales of hundreds of thousands of years.
My attendance at this conference promoted oceanic subsurface microbial observatory research in general and also encouraged future collaborations with the terrestrial subsurface research community, as well as promoting the educational goals of C-DEBI. I presented results of the collaborative research in development and use of oceanic subsurface microbial observatories in the AGU Session H52: “Rocks, Fractures, Fluids and Life; Insights from Underground Research Laboratories”. I highlighted the ongoing observatory work being conducted on the Juan de Fuca Ridge flank, a central focus of the C-DEBI program. In addition, I presented information about the “Adopt A Microbe” (AAM) education and outreach project in the AGU Session ED17: “Teacher Professional Development Programs Promoting Authentic Scientific Research in the Classroom”. AAM was a custom internet-based project of IODP Expedition 327 to promote interactive learning about microbial life in the deep biosphere. With encouragement from the Deep Earth Academy, I presented information about how the project was developed and provided user-feedback to help a future generation of expedition educators develop similar programs. I also volunteered at the C-DEBI vendor booth at the AGU conference, to further promote CDEBI to the research and education communities.
I have been invited to sail as a Microbiologist on IODP Expedition 336 (X336). The purpose of this expedition is to install CORK observatory systems in the ‘North Pond’ location of the Mid-Atlantic Ridge for coupled hydrogeological, geochemical and microbiological analysis. This expedition is one of the three current focus sites of C-DEBI for deep biosphere research. During this cruise, I will be integrally involved in the assembly and installation of the microbial observatory components for these CORKs – a critical component of these experiments. I was chosen for this position based on my previous history of designing the observatory components and participation in 6 research expeditions to deploy, recover, service and analyses these instruments. I have hands-on experience with microbiology experiments in CORKs based on my participation in the recent IODP Expedition 327 to the Juan de Fuca Ridge flank.
This exchange is for travel and research support for a new collaborative project aiming to expand the utility of deep UV fluorescence microscopy techniques for working with environmental samples. Specifically, we propose to test the applicability of common fluorophore-labeled oligonucleotides, often used in environmental microbiology studies to determine the abundance of particular microbial groups, to sample scanning with current deep UV microscopes. We predict that some fluorophores should generate unique spectral signatures with deep UV excitation, and that these signatures could be utilized for mapping the distribution of specific microbial groups targeted with oligonucleotide-fluorophore probes. If successful, the combination of group-specific labeling of cells with deep UV fluorescence scanning will provide a powerful new tool for the C-DEBI community.