Retention of uranium in biofilms of a granitic nuclear waste repository research tunnel

Several mechanisms of interactions of microorganisms with actinides under aerobic conditions are known, e.g. biosorption, interactions with S-layers or bioprecipitation. Some of them have the potential of a substantial retention of actinides, like uranium. However, little is known about the retention processes in a microbial community, such as biofilms. Since bacteria do not usually occur as single individual cells in nature but as multicellular communities, we have to draw our attention on the biofilms. They are characterized by building up their own microenvironment, which can differ significantly from that of the bulk solution with the consequence that biofilms are becoming more important in potential retention processes for actinides.

At the depth of 70 m (tunnel chainage 771) of the nuclear waste repository research tunnel ONKALO (Finland), which will be part of the nuclear waste repository in the future, massive biofilms are growing next to a fracture zone in a granitic rock environment. They were described by Pedersen et al. (2008) as a pink and solid slime, consisting of Pseudomonas anguilliseptica, Arthrobacter bergeri, Hydrogenophaga sp., Methylobacter tundripaludum, Rhodoferrax ferrireducens, and Haliscomenobacter hydrossis. The samples were removed from the tunnel wall together with the fracture water for uranium sorption experiments in a flow cell. A uranium concentration of 4×10^-5 M was adjusted in the fracture water by adding UO₂(ClO₄)₂. The water was pumped through the flow cell in a closed circuit for 42 hours. Microsensor measurements of the redox potential, pH and oxygen were performed in the several millimeters thick biofilms before and after the addition of uranium in order to record the effect of uranium. The obtained data showed significant changes of redox potential, pH and oxygen. The redox potential decreased after the addition of uranium during the experiment from 70 ± 2 mV to -164 mV with an increase of the pH from 5.4 ± 0.1 to 7.3 ± 0.1 at the same time, indicating reducing conditions in the microenvironment of the biofilm. The decrease in the oxygen concentration showed that the bacteria in the top region of the biofilms, i.e. the metabolically most active biofilms zone, battled the toxic effects of aqueous uranium with an increased respiratory activity, which resulted in oxygen depleted zones. Redox processes may have been triggered, leading to a removal of uranium from the aqueous phase. Analysis, which were performed before and after the sorption experiment, clearly showed, that 63 % of the added uranium was immobilized.

The retardation of uranium in the biofilm was determined by Energy-filtered Transmission Electron Microscopy (EF-TEM) and Electron Energy Loss Spectroscopy (EELS). Elongated particles of high electron density were observed in the cytoplasm of some rod shaped gram negative bacteria, which were often found associated with large rod shaped bacteria. Analysis of the elongated particles by EELS provided spectroscopic evidence for the presence of uranium immobilization, showing unequivocally uranium ionization intensity peaks of O4,5- and N6,7-edges. Distribution analysis of uranium, phosphorus and calcium clearly showed, that a solid uranium mineral has formed intracellular, which indicates the presence of a solid U-phosphate mineral similar to Autunite (Ca[UO₂]₂[PO₄]2•10-12H₂O).

Pedersen K, Arlinger J, Eriksson S, Hallbeck M, Johansson J, Jägevall S, Karlsson L. Microbiology of Olkiluoto Groundwater. Results and Interpretations 2007. POSIVA Working Report 2008-34. Olkiluoto, Finland.