Although being deionized, filtered and therefore normally deeply oligotrophic, the water from a basin containing irradiating waste presented relatively high bacterial concentrations (ca 10 5 cfu ml À1) and biofilm development at its surface and on the walls. This water was characterized by a high concentration of molecular H 2 due to water radiolysis, while its electrochemical potential was around +400 mV due the presence of dissolved O 2 and active oxygen compounds. This combination of H 2 availability and of an oxidant environment is completely original and not described in nature. From surface and wall biofilms, we enumerated the auto-trophic populations (%10 5 bacteria ml À1) able to grow in presence of H 2 as energy source and CO 2 as carbon source, and we isolated the most abundant ones among cultivable bacteria. They efficiently grew on a mineral medium, in the presence of H 2 , O 2 and CO 2 , the presence of the three gases being indispensable. Two strains were selected and identified using their rrs gene sequence as Rals-tonia sp. GGLH002 and Burkholderia sp. GGLH005. In pure culture and using isotope exchange between hydrogen and deuterium, we demonstrated that these strains are able to oxidize hydrogen as energy source, using oxygen as an electron acceptor, and to use carbon dioxide as carbon source. These chemoautotroph hydrogen-oxidizing bacteria probably represent the pioneer bacterial populations in this basin and could be primary producers in the bacterial community.