Microdiversity of Leptospirillum Ferrooxidans isolates recovered from uranium wastes and their interaction with U(VI)

A large number of acidophilic chemolithoautotrophic iron oxidizing bacterial isolates recovered from two uranium mining waste piles was classified as Leptospirillum ferrooxidans. For this purpose a combination of classical and molecular microbiological methods was applied. The letter involved Amplified Ribosomal DNA Restriction Endonucleases analysis (ARDREA) and 16S rDNA sequence analysis. In the ARDREA two sets of primers were involved: one universal -16S7f and 16S1406r, and one species specific -16S176f and 16S679r (De-Wulf-Durand et al,.1997, Tzvetkova et al., 1999). In both cases using endonuclease RsaI, two groups were discriminated among the isolates studied. These groups correspond to the recently characterized RsaI RFLP types I and II of L. ferrooxidans, which include the type strain of the species L. ferrooxidans DSM 2705T (type I) and the strains L. ferrooxidans DSM 2391 and Lf30A (type II); (Fig.1 and Tzvetkova et. al., 1999);
An additional extensive comparative sequence analysis of the variable region 3 of a series of natural isolates and the deposited sequences of this region in the EMBL allowed to distinguish seven L. ferrooxidans groups. Each group possesses a specific signature in the helix 18 of the variable region 3 (Fig.2). These signatures discriminate particular types of the species and possibly reflect the genetic adaptations of different parts of the natural L. ferrooxidans populations to different concentrations of heavy metals and other components of their natural environment. The variability of the helix 18 influences target sites of several frequently cutting endonucleases and allows discrimination of some of the above mentioned groups.
The interaction of some L. ferrooxidans natural isolates with uranium was studied at different concentrations of the metal and at two pH values. Results demonstrating accumulation of U(VI) at unusually low pH values are presented and discussed in the poster