N-196. Phylogenetic Analysis and a Starch-Iodide Screening Method for the Isolation of Arsenic-Oxidizing Thermus spp. from the Hot Springs of Hot Creek, CA
Arsenic (As) is a toxic metalloid present in natural waters and is regulated in potable waters by most developed nations at 10 µg/L. Its most toxic form, arsenite [As (III)], is mobile and difficult to remove from treated waters. Hot Creek, located in the northeastern part of California, is an arsenic-rich tributary to the Owens River and a source of water for the Department of Water and Power of Los Angeles. Speciation of arsenic in dormant and active pools of the creek was determined using ion-chromatography with results showing >99% oxidized species of arsenic in the dormant pools verses 60% oxidized in the active pools. Given this evidence, an attempt was made to isolate arsenite-oxidizers from the waters, sediments and biofilms of these pools. Over 100 arsenite-oxidizing thermophiles were isolated using a rapid colorimetric method based on the reaction of soluble starch and subunits of tri-iodide (I3-) and pentaiodide (I5-) with arsenic. Restriction fragment length polymerization analysis on the 16S rRNA gene indicates that 5 different groups of As-oxidizing thermophiles were isolated. Phylogenetic analysis of the sequences obtained from the 16S rRNA gene places all the isolates within the Thermus genus and related to previously uncultured species. Results obtained indicate that these new isolates do not form part of the T. thermophilus or the T. aquaticus clade. These results strongly suggest that the community of heterotrophic arsenic-oxidizing Thermus spp. in Hot Creek is well varied and plays an important role in the redox cycling of arsenic in geothermal systems.