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Project Progress

Signatures of life on early Earth and Mars

One area of our research is directed at using chemolithoautotrophic iron-oxidizing bacteria in low-temperature areas near hydrothermal vents to study present day production of biosignatures. Electron microscopy, spatially resolved x-ray spectroscopy and diffraction, and stable isotope studies are being used to address this topic. Brandy Toner has collected and analyzed mineralogical data on (1) sulfide minerals incubated at Juan de Fuca Ridge and (2) naturally weathered chimney sulfides collected from 9° N East Pacific Rise. Biominerals can be difficult to characterize due to the absence of long-range structural order, structural defects, small particle size, and close spatial proximity to other minerals and biological material. To overcome these challenges, micro-beam synchrotron-based x-ray absorption spectroscopy (µXAS) and diffraction (µXRD) is being used to characterize the weathering of sulfide minerals. These techniques provide mineralogical information at the micrometer spatial scale for intact samples, allowing one to focus on the weathering products and identify the parent minerals. As XAS is a non-destructive technique, the mineralogical information obtained from specific sample locations can be paired with spatially resolved isotopic and electron microscopic data. While seafloor sulfides are precious samples, they are an excellent model system for optimizing sampling schemes for extremely valuable samples such as those from sample return missions to Mars.

Adaptation and molecular strategies used by microbes in the environment

We are studying the fundamental biology of iron oxidation using the lithotrophic bacterium Marinobacter aquaeolei as a model. The ocean crust represents an enormous habitat for organisms that can harness lithotrophic geochemical energy from Fe(II) oxidation. Our previous work demonstrated the presence and suggested that an abundance of Fe(II)oxidizing bacteria may be active at the ocean floor; however the mechanism by which these organisms oxidized Fe(II) has not been elucidated. Marinobacter aquaeolei, a litho-heterotroph that is closely related to many deep-sea, Fe(II)-oxidizing autotrophic bacteria, to examine a potential mechanism of Fe-oxidation by tracking changes in protein expression resulting from changing Fe concentrations. This Marinobacter strain offers several advantages for this study over the deepsea isolates. (1) The metabolic plasticity of this strain allows co-metabolism of Fe(II) and O2 in the presence of an organic substrate. In contrast to the obligately autotrophic deep-sea isolates. This metabolic flexibility also allows Fe(II) concentrations to be manipulated without drastically effecting growth rates. (2) A draft genome of Marinobacter aquaeolei has been completed by DOE’s Joint Genome Institute, therefore the genetic potential of the organism can be queried and exploited. In this study, batch cultures of the bacterium were initiated in artificial seawater medium with various concentrations of Fe or “biologically unavailable” ferrozine chelated Fe. Under all conditions in this study, the rate of Fe-oxidation was faster in the presence of the bacterium than in the abiotic controls. Protein extractions from these incubations show the presence of a circa 35-kDa protein located in the cell membrane in the Fe replete conditions. Peroxidase activity assay shows this protein contains heme. The draft genome was queried and five potential genes were found that would encode for a heme containing protein in the 35-kda size range. One of these genes is of particular interest, a di-heme cytochrome c peroxidase, occurs in an gene cluster with five other genes which encode for other proteins involved in electron transport. Primers were developed for these six genes for use in RT-PCR to determine if any of these genes are up-regulated under Fe-replete conditions. This work describes a potential mechanism for Fe-oxidation and is a first step in developing an assay to probe the activity of such organisms and metabolisms in situ. Understanding which proteins are involved in Fe-oxidation, and the genes that encode for their production could lead ultimately to an RNA or antibody assay for environmental studies.