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

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.