The Year-Long Development of Microorganisms in Uncompacted Bavarian Bentonite Slurries at 30 and 60 °C

Abstract

In the multibarrier concept for the deep geological disposal of high-level radioactive waste (HLW), bentonite is proposed as a potential barrier and buffer material for sealing the space between the steel canister containing the HLW and the surrounding host rock. In order to broaden the spectra of appropriate bentonites, we investigated the metabolic activity and diversity of naturally occurring microorganisms as well as their time-dependent evolution within the industrial B25 Bavarian bentonite under repository-relevant conditions. We conducted anaerobic microcosm experiments containing the B25 bentonite and a synthetic Opalinus Clay pore water solution, which were incubated for one year at 30 and 60 °C. Metabolic activity was only stimulated by the addition of lactate, acetate, or H2. The majority of lactate- and H2-containing microcosms at 30 °C were dominated by strictly anaerobic, sulfate-reducing, and spore-forming microorganisms. The subsequent generation of hydrogen sulfide led to the formation of iron–sulfur precipitations. Independent from the availability of substrates, thermophilic bacteria dominated microcosms that were incubated at 60 °C. However, in the respective microcosms, no significant metabolic activity occurred, and there was no change in the analyzed biogeochemical parameters. Our findings show that indigenous microorganisms of B25 bentonite evolve in a temperature- and substrate-dependent manner.

Bentonite Alteration in Batch Reactor Experiments with and without Organic Supplements: Implications for the Disposal of Radioactive Waste

“…The relevance of microbial activity to the repository scenario, based on similar 1-year bentonite slurry experiments, has been previously discussed [37]. Indigenous microorganisms were shown to evolve under different temperature and substrate conditions but had no significant effect on the analyzed biogeochemical parameters [37,71]. In this follow-up study on two different bentonites, similar slurry experiments with an additional focus on mineralogical changes were compared with and without the addition of organic supplements.…”

“…The relevance of microbial activity to the repository scenario, based on similar 1-year bentonite slurry experiments, has been previously discussed [37]. Indigenous microorganisms were shown to evolve under different temperature and substrate conditions but had no significant effect on the analyzed biogeochemical parameters [37,71].…”

“…To accelerate the microbial processes and to shorten the incubation period, the concentration was increased without knowing whether the tolerable concentration would be exceeded. Matschiavelli et al [37] pointed out that a lactate or acetate concentration of 50 mM is possibly too high for soil organisms, which is also likely for the organic mixture concentrations used in this study. Inherent microorganisms may be affected, since they are not used to coping with these high concentrations in their natural environment.…”

“…For analyzing the microbial diversity within the respective samples, DNA extraction and amplification of the V4-region was conducted according to the protocol of Matschiavelli et al [37] without modifications.…”

Prediction of bacterial functional diversity in clay microcosms

“…An increased share of enzymes related to intracellular accumulation of the carbohydrate phosphate esters (phosphotransferase system), fructose and mannose metabolism, sporulation and peptidases in the sequenced 16S rRNA gene libraries from H 2 -fed microcosms with MX-80 Batch 1 clay, perhaps was due to enrichment of these incubations with spore-forming bacteria (e.g., Bacillaceae, Peptostreptococcaceae, Clostridiaceae and Peptococcaceae) that need these enzymatic systems for normal sporulation/germination processes (Bobek et al, 2014). Such an enrichment is of no coincidence since spore-formers have repeatedly been observed to predominate among cultivable bacteria in bentonites (Chi Fru and Athar, 2008;Gilmour et al, 2021;Grigoryan et al, 2020;Haynes et al, 2018;Jalique et al, 2016;Matschiavelli et al, 2019;Podlech et al, 2021). The anoxic clay microcosms were highly likely driven primarily by sulfate and hydrogen, and the sulfur metabolism fell in the top 30-35% of the most significant contributor to metabolic processes.…”