Isolation and characterization of iron-oxidizing bacteria from Boiling Springs Lake and the potential role of ferrous iron in carbon and sulfur cycling

Boiling Springs Lake (BSL) is a 52˚C, pH 2, iron and sulfur-rich thermal feature in Lassen National Volcanic Park (California, USA). Previous community composition studies of small subunit rRNA and RuBisCo genes revealed an abundance of phylotypes closely related to an Acidimicrobium strain isolated from Yellowstone National Park. As an ideal candidate to examine its contribution to primary production in BSL, we attempted to isolate Acidimicrobium and related iron-oxidizing Bacteria from BSL. We obtained 23 isolates that shared 99% rRNA gene identity with their closest cultured relative: 16 were identified as Sulfobacillus acidophilus, four as Alicyclobacillus sp., and three isolates had nearly identical rRNA gene sequences to the previously identified Acidimicrobium clones. We characterized most isolates for pH and temperature growth range and optima, and we assessed their abilities to oxidize iron, pyrite, sulfur and tetrathionate. Morphology of Acidimicrobium isolates was analyzed with transmission and scanning electron microscopy. The Acidimicrobium isolates were Gram positive, non-endospore-forming rods with a complex cellular envelope. Optimal growth temperature and pH for many isolates correlated with conditions at BSL. Acidimicrobium and Sulfobacillus isolates oxidized 10 mM iron when amended with 0.01% yeast extract. Acidimicrobium isolates were inhibited at iron concentrations ≥25 mM. Acidimicrobium and some Sulfobacillus isolates were able to oxidize pyrite when amended with yeast extract. We found no evidence for sulfur or tetrathionate oxidation by any of the isolates. These results will further our understanding of the potential role of Acidimicrobium and related iron-oxidizing Bacteria in the iron and carbon cycles in BSL.