Molecular phylogeny of uncultivated Crenarchaeota in Great Basin hot springs of moderately elevated temperature

Uncultivated Crenarchaeota are distributed widely in low temperature (< 30 degrees C) environments, and it has been hypothesized that they evolved from (hyper) thermophilic species thriving in marine hydrothermal vents or terrestrial hot springs. To further our understanding of the environmental distributions of Crenarchaeota, we studied mat samples collected from hot springs of moderately elevated temperature (similar to 49-67 degrees C) in California and Nevada, USA. Clone libraries of archaeal 16S rRNA genes were constructed for selected samples using a PCR-based approach. Sequences from the Nevada hot springs ( Rick's Hot Creek and Hard to Find) were closely related to uncultivated Crenarchaeota found near deep sea hydrothermal vents or from the subsurface geothermal system; sequences from the California hot spring ( Surprise Valley), on the other hand, were closely related to sequences from freshwater sediments. Statistical analysis showed that the community structure of Archaea was significantly different between any two springs with greater differences occurring between the Nevada and California hot springs ( P = 0.002). To determine whether these sequences represent indigenous microorganisms of geothermal springs, and not soil contaminants, archaeal 16S rRNAgene clone libraries were also constructed from soil samples taken from around Rick's Hot Creek and Surprise Valley hot springs. None of the hot spring sequences was closely related to those from the surrounding soil in Nevada or California or to the predominant soil Crenarchaeota in other locations, indicating that soil contamination to the hot spring environment was insignificant. Results of this study expand the distribution of Crenarchaeota into the moderately thermobiotic environment, which has been much less intensively studied than high temperature (> 70 degrees C) or low temperature natural habitats, and demonstrates that thermophiles inhabiting moderate temperature portions of Great Basin hot springs are phylogenetically distinct from both cultivated hyperthermophilic Crenarchaeota and sympatric soil Crenarchaeota.