Fifteen shades of clay: distinct microbial community profiles obtained from bentonite samples by cultivation and direct nucleic acid extraction

Abstract

Characterizing the microbiology of swelling bentonite clays can help predict the long-term behaviour of deep geological repositories (DGRs), which are proposed as a solution for the management of used nuclear fuel worldwide. Such swelling clays represent an important component of several proposed engineered barrier system designs and, although cultivation-based assessments of bentonite clay are routinely conducted, direct nucleic acid detection from these materials has been difficult due to technical challenges. In this study, we generated direct comparisons of microbial abundance and diversity captured by cultivation and direct nucleic acid analyses using 15 reference bentonite clay samples. Regardless of clay starting material, the corresponding profiles from cultivation-based approaches were consistently associated with phylogenetically similar sulfate-reducing bacteria, denitrifiers, aerobic heterotrophs, and fermenters, demonstrating that any DGR-associated growth may be consistent, regardless of the specific bentonite clay starting material selected for its construction. Furthermore, dominant nucleic acid sequences in the as-received clay microbial profiles did not correspond with the bacteria that were enriched or isolated in culture. Few core taxa were shared among cultivation and direct nucleic acid analysis profiles, yet those in common were primarily affiliated with Streptomyces, Micrococcaceae, Bacillus, and Desulfosporosinus genera. These putative desiccation-resistant bacteria associated with diverse bentonite clay samples can serve as targets for experiments that evaluate microbial viability and growth within DGR-relevant conditions. Our data will be important for global nuclear waste management organizations, demonstrating that identifying appropriate design conditions with suitable clay swelling properties will prevent growth of the same subset of clay-associated bacteria, regardless of clay origin or processing conditions.

Microbial hydrogen sinks in the sand-bentonite backfill material for the deep geological disposal of radioactive waste

“…The microbiome obtained from dry clay materials may not represent the full diversity of the community due to the inability to detect low-abundance species. This is evidenced when comparing the microbial diversity of dry clay and that of the same clay placed in water with an electron donor/acceptor ( Vachon et al, 2021 ). Therefore, it is not a trivial exercise to pinpoint the origin of the microbial community identified in the 4 reactors.…”

Investigating the effect of bentonite compaction density and environmental conditions on the corrosion of copper materials

“…[ 8 ] These changes in environmental conditions will also influence the activity of the microbes that are naturally found in bentonite clay, including sulfate‐reducing bacteria (SRB). [ 9 ] Upon emplacement, the bentonite clay's water activity will be low, suppressing microbial activity. The effects of irradiation and increased temperature from the decaying fuel inside the UFC have also been shown to eliminate spore‐forming bacteria below the detection limits of culturing.…”

“…Due to bentonite's inhospitable physical and chemical properties, the number of culturable microbes in bentonite is very low, ranging from 10 2 to 10 3 colony‐forming units per gram. [ 9 ] The low metabolic activity in bentonite clay is attributed partly to the high swelling pressure, low water activity, and small pore size. Previous studies determined that the number of metabolically active microbes decreases at higher bentonite dry densities.…”

Stable microbial community in compacted bentonite after 5 years of exposure to natural granitic groundwater

“…Sulfate-reducing bacteria (SRB) are of specific interest due to their production of hydrogen sulfide gas (H 2 S), which has potential for UFC metal corrosion. Although previous studies have explored microbial communities present in as-received, uncompacted bentonite clay and lab-scale pressure vessels ( 3 – 8 ), few SRB are detected and this may be a result of bentonite-associated inhibition. In particular, highly compacted bentonite can inhibit microbial growth when exceeding 1.6 g/cm 3 dry density, which corresponds to water activity below 0.96 and swelling pressures exceeding 2 MPa ( 9 ), and it was shown that swelling bentonites decrease sulfide production by SRB ( 10 ).…”

“…A study featuring ancient analog samples reported low cultivable heterotroph and SRB associated with a bentonite deposit that was formed ~10 million years ago and sequencing of SRB cultures revealed that the taxon present was affiliated with the genus Desulfosporosinus ( 7 ). An analysis of dry, uncompacted bentonite clay samples showed that cultivation-based approaches were consistently associated with phylogenetically similar SRB, aerobic heterotrophs, and fermenters ( 8 ). Dominant nucleic acid sequences extracted from these dry clay samples did not correspond with the bacteria that were enriched or isolated in culture, and relatively few core taxa (e.g., Streptomyces , Micrococcaceae , Bacillus , and Desulfosporosinus ) were shared among cultivation and direct nucleic acid analysis profiles ( 8 ).…”