Multistage uranium bioassociation kinetics of two Halobacterium noricense strains under highly saline conditions

Salt rock is one potential host rock formation being considered by Germany for the final disposal of radioactive waste. In addition to the characterization of geochemical and geophysical parameters, it is of great importance to take microbial activity of indigenous microorganisms into account for the safety assessment. Few studies have been conducted to investigate the microbial influence on the migration of radionuclides at high ionic strength. In this work, the extremely halophilic archaeon Halobacterium (Hbt.) noricense and its interactions with uranium were studied. Two different strains of the in salt rock common Hbt. noricense [1-4] were investigated. One was originally isolated from an Austrian salt mine (Hbt. noricense DSM-15987) [1] and the other from the US nuclear waste disposal site Waste Isolation Pilot Plant (WIPP, Carlsbad, NM) [2].
Biosorption kinetics were monitored in 3 M NaCl containing 40 or 50 µM uranium at pH 5.5 in the presence of Hbt. noricense cells. The fraction of bioassociated uranium was calculated from the uranium remaining in solu-tion analyzed by ICP-MS. Surprisingly, both strains of Hbt. noricense do not reflect typical sorption behavior; i.e. fast sorption within the first hours until reaching a stable equilibrium state. In this study, a fraction of uranium was sorbed within the first two hours of exposure time followed by a release of the radionuclide for a certain time. Subsequently, the amount of bioassociated uranium was found to increase very slowly until a maximum sorption of 80% was reached after 48 h. For a better understanding of this unique behavior, spectroscopic as well as microscopic methods were applied. Results from in situ Attenuated Total Reflection Fourier-transform Infrared (ATR FT-IR) spectroscopy evidenced that within the first two hours the actinide binds to carboxylic as well as to phosphate groups to cells of Hbt. noricense DSM-15987 simultaneously. The viability of Hbt. noricense cells was checked after contact with uranium by using the LIVE/DEAD® Bac LightTM Bacterial Viability Kit and analyzed with fluorescence microscopy. Almost all cells were alive even after one week of exposure to uranium. However, cell agglomeration was observed with increasing incubation time in both Hbt. noricense strains. For Hbt. noricense DSM-15987, a faster agglomeration process with agglomerates up to 200 µm in size was observed compared to the WIPP strain, which formed only microscopically visible agglomerates. The unusual, but reproducible bioassociation kinetics suggests that the WIPP strain and the Austrian strain react in the same way to uranium presence.

REFERENCES
[1] Gruber, C. et al. (2004) Extremophiles, Vol. 8, 431-439.
[2] Swanson, J. et al. (2012) LANL Report, LA-UR-12-22824.
[3] Gramain, A. et al. (2011) Environ. Microbiol., Vol. 13, 2105-2121.
[4] Yildiz, E. et al. (2012) Pol. J. Microbiol., Vol. 61, 111-117.