Microbial metabolism in the deep subsurface:case study of Opalinus Clay

Microbes have successfully colonized the deep subsurface, thanks to their small size and their diverse metabolic activities. This part of the biosphere remains a terra incognita for microbiologists, containing innumerable unknown microbial species and processes. We depend on it for many things, i. e. water supply, oil extraction and nuclear waste disposal. In Switzerland, Opalinus Clay will be used for deep geological repositories, because of its very low permeability. This rock has been studied for 20 years, however, the potential role of microbes has long been overlooked, even though they could play a major role in controlling geochemical conditions. It was showed that they were present in pristine rock, but were not active due to the lack of space. The potential for microbial activity in disturbed rock, under repository relevant conditions, remains unexplored. This is the focus of this thesis. The first condition tested was the presence of space alone. This will occur when the rock is excavated to build the repository. The digging creates a network of large fractures in the rock zones near the galleries. This change is enough to promote microbial activity in a system, mainly composed of two species. The first one, a Peptococcaceae, oxidizes organic carbon to dioxide carbon, by reducing sulfate. The second organism is a Pseudomonas that seems to grow by fermenting organic carbon. From this model, it appears that the Peptococcaceae feeds on low molecular organic acids present in the Opalinus Clay, but also on fermentation products from Pseudomonas. What is less clear is the source of organic carbon for the latter. Is it only feeding on dead microbial cells, or is it also able to feed on reactive fossilized carbon contained in Opalinus Clay? The second case study included an additional energy source, hydrogen. This gas will be produced by anoxic steel corrosion and could damage the repository if the pressure increases significantly. However, it can be oxidized in situ by autotrophic microbial species. The dominating microbe in this system is a sulfate-reducing Desulfobulbaceae. It produces organic carbon from carbon dioxide, which can later feed heterotrophic bacteria, which are also sulfate-reducing and that release carbon dioxide. A carbon loop was thus reconstructed for the first time in the deep subsurface. It includes a fermentation step that releases the organic substrate for heterotrophic sulfate-reducing bacteria that completely oxidize simple organic carbon molecules, i.e., acetate, to carbon dioxide. In this system, hydrogen and sulfate consumption rates are 1.5 and 0.2 µmol·cm-3·day-1, respectively. This means that electrons derived from hydrogen reduce electron acceptors other than sulfate, confirming carbon fixation. These projects highlight the role that microbes can play for the safety of nuclear waste disposal. Hydrogen consumption is a beneficial process, but the overall impact of microbial activity can be negative, because it can promote corrosion and weathering of the different barriers surrounding the waste. Further work is needed in order to obtain a more complete picture of the role played by bacteria in a deep geological repository. Nonetheless, the results presented here represent a large improvement in our understanding of deep subsurface microbiology. Microbial processes were never described in this much detail in these environments, as is done here: from the metabolic pathway level, up to the ecosystem level

Rates of microbial hydrogen oxidation and sulfate reduction in Opalinus Clay rock

Hydrogen gas (H2) may be produced by the anoxic corrosion of steel components in underground structures, such as geological repositories for radioactive waste. In such environments, hydrogen was shown to serve as an electron donor for autotrophic bacteria. High gas overpressures are to be avoided in radioactive waste repositories and, thus, microbial consumption of H2 is generally viewed as beneficial. However, to fully consider this biological process in models of repository evolution over time, it is crucial to determine the in situ rates of microbial hydrogen oxidation and sulfate reduction. These rates were estimated through two distinct in situ experiments, using several measurement and calculation methods. Volumetric consumption rates were calculated to be between 1.13 and 1.93 μmol cm−3 day−1 for H2, and 0.14 and 0.20 μmol cm−3 day−1 for sulfate. Based on the stoichiometry of the reaction, there is an excess of H2 consumed, suggesting that it serves as an electron donor to reduce electron acceptors other than sulfate, and/or that some H2 is lost via diffusion. These rate estimates are critical to evaluate whether biological H2 consumption can negate H2 production in repositories, and to determine whether sulfate reduction can consume sulfate faster than it is replenished by diffusion, which could lead to methanogenic conditions

Arsenic methylation dynamics in a rice paddy soil anaerobic enrichment culture

Methylated arsenic (As) species represent a significant fraction of the As accumulating in rice grains, and there are geographic patterns in the abundance of methylated arsenic in rice that are not understood. The microorganisms driving As biomethylation in paddy environments, and thus the soil conditions conducive to the accumulation of methylated arsenic, are unknown. We tested the hypothesis that sulfate-reducing bacteria (SRB) are key drivers of arsenic methylation in metabolically versatile mixed anaerobic enrichments from a Mekong Delta paddy soil. We used molybdate and monofluorophosphate as inhibitors of sulfate reduction to evaluate the contribution of SRB to arsenic biomethylation, and developed degenerate primers for the amplification of arsM genes to identify methylating organisms. Enrichment cultures converted 63% of arsenite into methylated products, with dimethylarsinic acid as the major product. While molybdate inhibited As biomethylation, this effect was unrelated to its inhibition of sulfate reduction and instead inhibited the methylation pathway. Based on arsM sequences and the physiological response of cultures to media conditions, we propose that amino acid fermenting organisms are potential "drivers of As methylation in the enrichments. The lack of a demethylating capacity may have contributed to the robust methylation efficiencies in this mixed culture

A minimalistic microbial food web in an excavated deep subsurface clay rock

Clay rocks are being considered for radioactive waste disposal, but relatively little is known about the impact of microbes on the long-term safety of geological repositories. Thus, a more complete understanding of microbial community structure and function in these environments would provide further detail for the evaluation of the safety of geological disposal of radioactive waste in clay rocks. It would also provide a unique glimpse into a poorly studied deep subsurface microbial ecosystem. Previous studies concluded that microorganisms were present in pristine Opalinus Clay, but inactive. In this work, we describe the microbial community and assess the metabolic activities taking place within borehole water. Metagenomic sequencing and genome-binning of a porewater sample containing suspended clay particles revealed a remarkably simple heterotrophic microbial community, fueled by sedimentary organic carbon, mainly composed of two organisms: a Pseudomonas sp. fermenting bacterium growing on organic macromolecules and releasing organic acids and H-2, and a sulfate-reducing Peptococcaceae able to oxidize organic molecules to CO2. In Opalinus Clay, this microbial system likely thrives where pore space allows it. In a repository, this may occur where the clay rock has been locally damaged by excavation or in engineered backfills

Reconstructing a hydrogen-driven microbial metabolic network in Opalinus Clay rock

The Opalinus Clay formation will host geological nuclear waste repositories in Switzerland. It is expected that gas pressure will build-up due to hydrogen production from steel corrosion, jeopardizing the integrity of the engineered barriers. In an in situ experiment located in the Mont Terri Underground Rock Laboratory, we demonstrate that hydrogen is consumed by microorganisms, fuelling a microbial community. Metagenomic binning and metaproteomic analysis of this deep subsurface community reveals a carbon cycle driven by autotrophic hydrogen oxidizers belonging to novel genera. Necromass is then processed by fermenters, followed by complete oxidation to carbon dioxide by heterotrophic sulfate-reducing bacteria, which closes the cycle. This microbial metabolic web can be integrated in the design of geological repositories to reduce pressure build-up. This study shows that Opalinus Clay harbours the potential for chemolithoautotrophic-based system, and provides a model of microbial carbon cycle in deep subsurface environments where hydrogen and sulfate are present

The Maunakea Spectroscopic Explorer Book 2018

(Abridged) This is the Maunakea Spectroscopic Explorer 2018 book. It is intended as a concise reference guide to all aspects of the scientific and technical design of MSE, for the international astronomy and engineering communities, and related agencies. The current version is a status report of MSE's science goals and their practical implementation, following the System Conceptual Design Review, held in January 2018. MSE is a planned 10-m class, wide-field, optical and near-infrared facility, designed to enable transformative science, while filling a critical missing gap in the emerging international network of large-scale astronomical facilities. MSE is completely dedicated to multi-object spectroscopy of samples of between thousands and millions of astrophysical objects. It will lead the world in this arena, due to its unique design capabilities: it will boast a large (11.25 m) aperture and wide (1.52 sq. degree) field of view; it will have the capabilities to observe at a wide range of spectral resolutions, from R2500 to R40,000, with massive multiplexing (4332 spectra per exposure, with all spectral resolutions available at all times), and an on-target observing efficiency of more than 80%. MSE will unveil the composition and dynamics of the faint Universe and is designed to excel at precision studies of faint astrophysical phenomena. It will also provide critical follow-up for multi-wavelength imaging surveys, such as those of the Large Synoptic Survey Telescope, Gaia, Euclid, the Wide Field Infrared Survey Telescope, the Square Kilometre Array, and the Next Generation Very Large Array.Comment: 5 chapters, 160 pages, 107 figure