Halomonas desiderata as a bacterial model to predict the possible biological nitrate reduction in concrete cells of nuclear waste disposals

Abstract

After closure of a waste disposal cell in a repository for radioactive waste, resaturation is likely to cause the release of soluble species contained in cement and bituminous matrices, such as ionic species (nitrates, sulfates, calcium and alkaline ions, etc.), organic matter (mainly organic acids), or gases (from steel containers and reinforced concrete structures as well as from radiolysis within the waste packages). However, in the presence of nitrates in the near-field of waste, the waste cell can initiate oxidative conditions leading to enhanced mobility of redox-sensitive radionuclides (RN). In biotic conditions and in the presence of organic matter and/or hydrogen as electron donors, nitrates may be microbiologically reduced, allowing a return to reducing conditions that promote the safety of storage. Our work aims to analyze the possible microbial reactivity of nitrates at the bitumen - concrete interface in conditions as close as possible to radioactive waste storage conditions in order (i) to evaluate the nitrate reaction kinetics; (ii) to identify the by-products (NO2(-), NH4(+), N2, N2O, etc.); and (iii) to discriminate between the roles of planktonic bacteria and those adhering as a biofilm structure in the denitrifying activity. Leaching experiments on solid matrices (bitumen and cement pastes) were first implemented to define the physicochemical conditions that microorganisms are likely to meet at the bitumen-concrete interface, e.g. highly alkaline pH conditions (10 < pH < 11) imposed by the cement matrix. The screening of a range of anaerobic denitrifying bacterial strains led us to select Halomonas desiderata as a model bacterium capable of catalyzing the reaction of nitrate reduction in these particular conditions of pH. The denitrifying activity of H. desiderata was quantified in a batch bioreactor in the presence of solid matrices and/or leachate from bitumen and cement matrices. Denitrification was relatively fast in the presence of cement matrix (<100 h) and 2-3 times slower in the presence of bituminous matrix (pH 9.7). The maximal rate of denitrification was approximately 0.063 mM h(-1) and some traces of nitrite were detected for a few hours (<2%). Overall, the presence of solid cement promoted the kinetics of denitrification. The inspection of the solid surfaces at the end of the experiment revealed the presence of a biofilm of H. desiderata on the cement paste surface. These attached bacteria showed a comparable denitrifying activity to planktonic bacterial culture. However, no colonization of bitumen was observed either by SEM or by epifluorescence microscopy.

Interactions between hydrated cement paste and organic acids: Thermodynamic data and speciation modeling

“…After closure and water re-saturation of the cell, bituminous wastes are expected to release various organic compounds, including organic acids such as acetic, formic and oxalic acids, as a result of leaching and radiolysis [4 -7]. It has been shown that strong interactions can occur between these organic acids and the concrete of the waste cell [8] thus modifying the bio-chemical conditions in the vicinity of the bitumen-concrete interface [8,9]. This could have a direct influence on safety [10].…”

Understanding interactions between cementitious materials and microorganisms: a key to sustainable and safe concrete structures in various contexts

“…organic acids released by bitumen, such as acetic or oxalic acids, H 2 , the various metallic phases in the repository, etc.) in the reactions [129][130][131]. The environment under consideration is complex, multicomponent and multi-phase.…”

“…The environment under consideration is complex, multicomponent and multi-phase. A step-wise approach was thus adopted, starting with simplified decoupled model systems (model media studied in batch conditions) [129,130,132], the complexity of which was progressively increased to move to close-to-real systems using an experimental pilot specifically developed for the study [131].…”

“…These reactions may, on the one hand, alter the durability of concrete cells [84,88,129,132,144], and, on the other hand, change the availability of organic matter usable by heterotrophic microorganisms for the reduction of oxyanions such as nitrates. It should also be noted that other sources of organic acids than bitumen can be found in nuclear waste storage: when cellulosic materials are immobilized with other lowand intermediate level radioactive wastes in cement matrices, cellulose may degrade, mainly to isosaccharinic acid, but also to formic and acetic acids following the reactions of polymers which break down under alkaline conditions [145,146].…”

“…notably at high pH (in the range of pH 9-11) and anoxic conditions (conditions that will prevail in the storage in the long term). Works carried out to explore the biologically induced reduction of nitrates in such conditions have considered selected pure strains and wild consortia (using natural analogs) [120,129,131,[151][152][153][154]. The first step of such studies was therefore to identify microorganisms capable of (i) growing in alkaline conditions, (ii) using nitrate as electron acceptor and (iii) using one or more electron donors present in the repository.…”