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

The Materials Corrosion Test (MaCoTe) at the Underground Research Laboratory in Grimsel, Switzerland, assesses the microbiology and corrosion behavior of engineered barrier components of a deep geological repository (DGR) for long-term disposal of high-level nuclear waste. Diversity and temporal changes of bentonite-associated microbial community profiles were assessed under DGR-like conditions for compacted Wyoming MX-80 bentonite (1.25 g/cm³ and 1.50 g/cm³ targeted dry densities) exposed to natural groundwater. Using culture-dependent and molecular techniques, samples taken from the outside layer of 5-year borehole modules revealed up to 66% and 23% of 16S rRNA gene sequences affiliated with Desulfosporosinus and Desulfovibrio, respectively. Putatively involved in sulfate reduction, these taxa were almost undetectable within the bentonite core. Instead, microbial profiles of the inner bentonite core were similar to uncompacted bentonite used to pack modules years earlier, and were consistent with a previously published 1-year time point, revealing no detectable microbial growth. Abundances of culturable aerobic and anaerobic heterotrophic bacteria in the uncompacted bentonite were relatively low, with less than 1,000 and 100 colony-forming units (CFUs) per gram dry weight, respectively. Nearly 5 years after emplacement, culturable heterotrophic bacterial CFUs and sulfate-reducing bacteria did not change significantly inside the bentonite core. Phospholipid fatty acid data indicated similar lipid abundance, and corresponding cell abundance estimates, for inner 5-year MaCoTe bentonite samples compared to those previously obtained for 1-year incubations. Collectively, our results provide complementary evidence for microbial stability inside highly compacted bentonite exposed to conditions that mimic engineered barrier components of a deep geological repository. IMPORTANCE The long-term safety of a deep geological repository for used nuclear fuel is dependent on the performance of the engineered and natural barriers. Microbial activity can produce chemical species that can influence the corrosion of the disposal containers for used nuclear fuel. Although previous studies have evaluated the microbiology of compacted bentonite clay within subsurface environments, these have been limited to relatively short incubations (i.e., 1 year). The current study provides a unique 5-year perspective that reinforces previous findings of growth inhibition for bentonite clay exposed to in situ subsurface conditions.