Response of bentonite microbial communities to stresses relevant to geodisposal of radioactive waste

Abstract

Microbes have been isolated previously from bentonite materials that may be used as barriers for the disposal of radioactive waste. Actively respiring microbes in such barrier materials, within a repository environment, have the potential to adversely affect waste container corrosion rates. Additionally, they could potentially alter the properties of the bentonite barrier itself. This is of significance, since the integrity of the waste container and properties of the bentonite barrier are required to fulfil defined safety functions. To help identify the critical factors that affect microbial activity in bentonite materials, this study examines the impact of a range of parameters that could affect microbial metabolism in a geodisposal environment. Several bentonites from different sources (bentonite mined from locations in Spain and the USA, along with commercially-sourced bentonite) were subjected to increased pressure (74 MPa, 30 s), heat (90 °C, 24 h), and irradiation (1000 Gy, 24.17 Gy min−1), before incubation in growth media selective for sulfate-reducing bacteria (SRB) or iron-reducing bacteria (IRB). The amount of SRB, and IRB were counted using the most probable number method and identified by 16S rRNA gene sequencing. The bentonites initially contained 660–6600 SRB cells g−1, and the number of SRB was correlated with the initial water content of the bentonite. A similar number of IRB was also present (400–4000 cells g−1), and the number of IRB was correlated with the ratio of bioavailable Fe(II)/Fe(III) present in the bentonite. The bentonites hosted sulfate-reducing species from two bacterial genera, with Desulfotomaculum dominating the SRB communities in the Spanish bentonite used in the Full-scale Engineered Barriers Experiment (FEBEX), while the other communities contained Desulfosporosinus species. The nature of the SRB community played a significant role in the microbial community response to different stresses, with the FEBEX material producing high SRB cell counts in response to pressure and irradiation but yielding low numbers in response to heat. Initially, the IRB communities contained a mixture of Gram-negative bacteria such as Geobacter, and Gram-positive spore-forming bacteria such as Bacillus and Desulfosporosinus, with an increase in the number of Gram-positive spore-formers in response to stress. The ability of Gram-positive spore-formers to grow, despite exposure to pressure, heat and irradiation, highlights the need to generate a swelling pressure sufficient to minimise microbial activity. In addition, we suggest that the microbial communities naturally present in the bentonite should be considered as part of the selection process for buffer materials in a geological disposal facility for radioactive waste.