Département d'Ecophysiologie Végétale et Microbienne-Laboratoire de Bioénergétique Cellulaire. CEA-Cadarache, Bât.156, 13108 St Paul-Lez-Durance, France.
In most contaminated soils and waste waters, selenium is present in highly soluble selenite or selenate forms. The biological reduction of these oxyanions into insoluble metallic selenium is therefore of interest as a potential strategy for bioremediation. Rhodobacter sphaeroides sp. denitrificans (IL-106) and Escherichia coli (MC4100) can reduce selenite into a red precipitate under both aerobic and anaerobic conditions. Interestingly, only MC4100 transform selenate. Moreover as determined by electron microscopy, selenium granules resulting from selenite metabolism are localized in the cytoplasm of IL-106 while they are found in the periplasm of Escherichia coli. This suggests that different pathways of reduction are possible depending upon the considered species. These pathways are largely uncharacterized. Three approaches have been used to describe the microbial reduction of oxidized selenium salts. Firstly, we have measured selenate and selenite reductase activities on non-denaturating gels. The only activities observed were correlated to the nitrate reductase. However, MC4100 and IL-106 mutants deleted in nitrate reductase still perform oxyanions reduction showing the presence of multiple pathways in vivo. Secondly, we have identified by biochemical means soluble and membrane proteins induced by selenite. Western-blot analysis using cpn60 antibodies and microsequencing of the induced proteins show clearly that selenite exposure leads to a stress response closely resembling to heat shock stress. Finally, we have created a transposon library and selected mutants unable to reduce selenate or selenite for MC4100 and IL-106 species respectively. The results of the sequencing of some mutated genes are discussed in this poster.