H2-fuelled microbial metabolism in Opalinus Clay

In Switzerland, the Opalinus Clay formation is considered the most likely host rock for a deep geological repository for nuclear waste. In deep geological repositories, H2 is expected to be the most abundant gas formed from the degradation of waste and from metal corrosion. The microbial community present in Opalinus Clay is capable of utilizing H2 as an electron donor and sulfate as an electron acceptor to produce hydrogen sulfide. This could be problematic due to its potential for increasing the corrosion of metal waste canisters containing radioactive waste, however, the possible impacts of these processes on the clay rock have not been fully investigated. In this study, a series of microcosm experiments were set-up containing Opalinus Clay and porewater from the Mont Terri underground research laboratory (Switzerland) as an inoculum. Uninoculated microcosms were established to investigate abiotic processes. In the presence of clay, a higher aqueous sulfate concentration was detected than in those with only porewater present and this concentration decreased over time in the inoculated experiments. However, there was no evidence of hydrogen sulfide production in the aqueous phase. In all experiments with clay, there was an increase in aqueous Fe2+ concentrations with the highest concentrations found in uninoculated experiments. The sulfur speciation of the Opalinus Clay was analysed and the results of the inoculated sample suggested that hydrogen sulfide reacted with Fe2+, precipitating iron sulfide minerals. After the incubation period, the microbial community was dominated by the sulfate-reducing Desulfobulbaceae family. The study suggests that H2-fuelled, microbially-mediated sulfate reduction can affect the mineral composition within the Opalinus Clay due to the precipitation of iron sulfide minerals. These precipitation reactions may enhance the long-term integrity of the repository by removing corrosive hydrogen sulfide from solution when sufficient Fe2+ is available and so protecting the canisters containing the nuclear waste.

Microbial metabolism in the deep subsurface

Alexandre Bagnoud

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.

EPFL2015

Utilization of silicate minerals for pH control during in situ bioremediation of chlorinated solvent

David Andrew Barry, Alessandro Brovelli, Christof Holliger, Elsa Marina Lacroix

Chloroethenes such as tetrachloroethene (PCE) and trichloroethene (TCE) are among the most prevalent contaminants in groundwater due to their extensive use in industrial processes. In situ bioremediation (ISB) is an attractive technology for removal of these compounds. It relies on an anaerobic process in which specialized bacteria obtain energy for growth using chloroethenes as an electron acceptor via organohalide respiration. Engineered bioremediation is achieved by stimulating these microorganisms through the addition of electron donor in the subsurface. This technology has been widely used for bioremediation of chloroethene plumes and recent studies have indicated promising results for bioremediation of chlorinated solvent source zones. However, application of source zone ISB is still a significant technical challenge. One of the main issues is the groundwater acidification due to organohalide respiration and fermentation processes, which can inhibit the activity of dehalogenating micro-organisms. The main objective of this work was to develop an efficient pH control strategy for chloroethene ISB by using the acid neutralizing potential of silicate minerals. To do so, modeling and experimental approaches were combined. A geochemical model, implemented within PHREEQC, was developed to select appropriate buffer candidates and to help determine main parameters influencing mineral buffering capacity. The model included chloroethene microbial degradation kinetics, mineral dissolution kinetics and chemical speciation. Second, anaerobic microcosm experiments were performed to determine the influence of pH on dehalogenation. These microcosms were inoculated with enriched consortia of dehalogenating bacteria and fed with PCE and hydrogen. Another set of microcosm experiments was carried out to compare the buffering capacity of ten silicate minerals and to investigate interactions between minerals and dehalogenating bacteria, e.g., the potential inhibitory effect of minerals on the dehalogenating activity. These microcosms were amended with 5 mmol l-1 of PCE and 4 g l-1 of mineral with grain sizes between 50 and 100 μm. The cultivation medium was modified such that the silicate mineral powder was the sole pH buffer present. Chloroethenes, pH and dissolved cation measurements were conducted to determine the system efficiency. Abiotic dissolution experiments were also performed to determine mineral dissolution rates in the absence of bacteria. The model confirmed that the efficiency of the system is dependent mainly on mineral dissolution kinetic constants, equilibrium constants and reactive surface area. The geochemical model and literature parameter data were used to pre-select minerals with a buffering capacity sufficient to counterbalance acidity produced by dehalogenating bacteria at a rate of 4 mmol l-1.d-1of chloride. Of the 31 silicate minerals for which there were published kinetic data, 10 were identified as suitable candidates. The inhibitory pH for the dehalogenating consortia was found to vary between 5 and 6. The last steps of the dechlorination from DCE to ethene were more sensitive to pH than the first steps from PCE to DCE, as has been noted in other dechlorination studies. Results of microcosm experiments with silicate minerals demonstrated that, under the selected conditions, the pH control behavior and the impact on bacterial activity exhibited strong variations depending on the mineral. Of the ten minerals tested experimentally, three (olivine, fayalite and diopside) maintained the pH in the appropriate range, i.e., between 5.5 and 6.5 and led to complete transformation of PCE. For the other minerals tested, either the acid neutralization capacity was insufficient due to slow dissolution kinetics (glaucophane and staurolite) or dechlorination activity was inhibited by (unmeasured) compounds released during mineral dissolution. Both modeling and experimental results demonstrated the feasibility of using selected silicate minerals as a buffering agent during ISB. However, the experimental results also revealed a mineral-induced potential inhibitory effect that should be investigated prior to application at a contaminated site.

2011

Utilization of silicate minerals for pH control during in situ bioremediation of chlorinated solvent

David Andrew Barry, Alessandro Brovelli, Christof Holliger, Elsa Marina Lacroix

2011

Microbial metabolism in the deep subsurface

Alexandre Bagnoud

EPFL2015