Influence of a halophilic archaeum to uranium migration under highly saline conditions

In Germany salt rock and clay are considered as potential host rock for the final repository of radioactive waste in deep geological formations. Both possibilities have in common that high saline conditions can occur. In clay deposits of Northern Germany pore water salt concentrations of 4.3 M were measured [1] and in salt rock the salt concentration is up to saturation. Despite these extreme environments some microbes are able to survive. To date little is known about the interactions of halophilic microorganisms with actinides and hence to the migration behavior. But for the safety assessment of the final repository it is important to know the impact of indigenous microorganisms. Microbes can interact with actinides in different ways [2]. Within this study, the sorption of uranium on the cell surface (so called biosorption) of Halobacterium noricense DSM 15987 cells was studied. This halophilic archaeum was chosen due to its worldwide occurrence in salt rock [3]. The reference strain was isolated in an Austrian salt mine [4] and similar species occurred also in the Waste Isolation Pilot Plant (WIPP, Carlsbad, New Mexico, USA) [3].
Biosorption studies were undertaken at pH 6.0 and a NaCl concentration of 3.0 M in dependence of uranium concentration, time and temperature. The uranium content in the supernatant after sorption was measured with ICP-MS (Inductively Coupled Plasma Mass Spectrometry). Both, supernatant and cell pellet, were analyzed with TRLFS (Time-resolved Laser-induced Fluorescence Spectroscopy) to get information about the formed complexes. Furthermore the cells were analyzed with Infrared Spectroscopy.
The results demonstrated that independent of the uranium concentration (10 – 120 µM) around 90 % of the added uranium was sorbed by the cells at room temperature. A time-dependent sorption study showed that this maximal sorption was reached after an incubation time of 42 h. A slightly faster sorption of added uranium could be seen at higher temperatures. Particularly at 50 °C, the maximal sorption was already reached after 24 h. In general, the obtained sorption curve indicated a two-step binding process of added uranium with a fast step within the first hours and a second slower one. For a uranium concentration of 100 µM the metal sorption rate of 37.5 ± 0.7 mg U(VI) per 1 g dry biomass of Halobacterium noricense DSM 15987 was determined .
Interestingly, with increasing time, uranium concentration and temperature the cells began to form agglomerates. Live/Dead staining (LIVE/DEAD® Bac LightTM Bacterial Viability Kit L7012, Molecular Probes) of cells after the biosorption with uranium showed that nearly all single cells were dead whereas agglomerated cells were alive. One conclusion is that this process is a kind of stress response to protect the cells themselves from environmental challenges.
The characterization of the formed cell-uranium-complexes with TRLFS indicated that uranium was bound to cellular carboxylic groups. This can be seen by comparing with literature spectra [5]. The obtained lifetimes of the uranyl complexes differed from those due to the quenching effect of the high chloride concentration to uranium luminescence. The binding of uranium to the carboxylic groups of the cell could be also verified with Infrared Spectroscopy.

[1] J. Larue, VerSi Endlagerung im Tonstein, Abschlussbericht 3607R02538 (2010).
[2] K. Morris, et al., Interactions of microorganisms with radionuclides 2, 101 (2002).
[3] J. S. Swanson, et al., Status report Los Alamos National Laboratory (2012).
[4] C. Gruber, et al., Extremophiles 8, 431 (2004).
[5] M. Vogel, et al., Sci. Total Environ. 409, 384 (2010).