Complexation of uranium by three different Acidithiobacillus Ferrooxidans types

In uranium mining piles a number of acidophilic chemolithoautotrophic bacteria has been identified, which are able to oxidize sulphide minerals, elemental sulfur, ferrous iron, and, in presence of uranium minerals, also U(IV). Especially one representative of this group, Acidithiobacillus ferrooxidans, is of particular interest. This organism has been used commercially in metal leaching from ores and decontamination of industrial wastes [1].
Sequence analysis of the 16S rRNA genes of several reference strains and uranium mining waste pile isolates of this bacterium revealed specific signatures which distinguish three types within the species. This allowed to develop a technique for analysis of the distribution of the A. ferrooxidans eco-types in the soil samples of a uranium mining waste pile.
The technique is based on amplification of 16S rDNA fragments in total soil DNA by the use of two A. ferrooxidans species specific primers 16S458F and 16S1473R [2]. The resulting amplicons were then digested with a frequently cutting enzyme RsaI which produced three different type-specific profiles [3; 4]. Using this direct approach we have demonstrated that one of the A. ferrooxidans types (type I) was predominant in the soil samples studied and was found in more polluted sites, whereas the type II was found in less contaminated samples. The type III was found mostly to coexist with the type II.
The objectives of the present work were to determine whether these eco-types differ in their capability to tolerate and accumulate uranium, and also to study the structural complexes formed at the surfaces of A. ferrooxidans eco-types using different spectroscopic techniques as Extended X-ray Absorption Fine Structure (EXAFS), Infra Red (IR) and time-resolved laser-induced fluorescence spectroscopy (TRLFS). In addition, the most efficient desorbing agent for the accumulated uranium was selected.
The uranium accumulation by the above mentioned three types of A. ferrooxidans was studied at different metal concentrations and different pH values (1.5 and 4). The results obtained (Fig.1) demonstrated that the strains from the different types possess different capability to accumulate uranium. The amount of uranium biosorbed by the three types increased with increasing concentration of uranium.
Fig.1: Biosorption of uranium by different types of A. ferrooxidans

Interestingly, the strains W1 (type I) and D2 (type III) are resistant to 8 and 9 mM of uranium, respectively, whereas the strain ATCC 33020 (type II) does not tolerate more than 2 mM of uranium (Table 1).

Strains

Uranium (mM)
Tolerated
MICs
A. ferrooxidans W1
8 9
A. ferrooxidans ATCC 33020
2 4
A. ferrooxidans D2
9 10
Table 1: Minimum Inhibitory Concentrations (MICs) of uranium for the growth of A. ferrooxidans type

On the basis of these results, one may speculate that the strains of the types I and III are more resistant to uranium, probably because they possess a mechanism which limits the uranium binding below the lethal amounts.

The desorption of the accumulated uranium from the bacterial cells was investigated using different desorbing agents as sodium carbonate, sodium citrate and EDTA at different concentrations. The results obtained demonstrated that the sodium carbonate is able to recuperate up to 97% of the uranium sorbed from the cells of A. ferrooxidans type III, and 88.33 and 88.50% from the cells of the types I and II, respectively.

Using EXAFS analysis we have found that no significant structural differences were observed between the uranium complexes formed by the 3 types of A. ferrooxidans. However, the EXAFS spectra are indicating formation of uranium complexes which are different from those formed by bacilli [5; 6].

Acknowledgements
This work was supported by grant 7531.50-03-FZR/607 from the Sächsisches Staatsministerium für Wissenschaft und Kunst, Dresden, Germany