In situ measurement of Fe(III) reduction activity of Geobacter pelophilus by simultaneous in situ RT-PCR and XPS analysis

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Abstract

Geobacter pelophilus is capable of dissimilatory Fe(III)-reduction on solid phase Fe(III)-oxides by means of surface attachment and direct electron transport to Fe(III), in part mediated by outer membrane c-type cytochromes. A study was undertaken to characterize surface colonization patterns, gene expression, and mineral transformation by this organism. The gene ferA (Geobacter sulfurreducens outer membrane Fe(III) reductase cytochrome c) was used as a target for PCR based molecular detection methods for visualizing G. pelophilus surface colonization. Protein extracts were prepared from solid-phase cultures, and cytochrome c content assessed. Mineral transformations were followed by X-ray photoelectron spectroscopy (XPS). Results of in situ (IS) RT-PCR experiments demonstrate that G. pelophilus attaches and grows at ferrihydrite mineral surfaces. Fluorescently-labeled cells were observed after IS-RT-PCR experiments, suggesting that G. pelophilus contains a cytochrome c sequence similar to ferA in G. sulfurreducens which is expressed in the presence of ferrihydrite. Protein extracts possessed high mass c-type cytochromes of similar size to those found in G. sulfurreducens. In addition, unique high-mass c-type cytochromes were also detected. XPS analysis demonstrated mineral transformation to occur, mediated by the surface associated population. This study demonstrates that G. pelophilus attaches to Fe(III)-oxide surfaces, reduces the Fe(III) oxides at the surface, produces c-type cytochromes under these growth conditions, and expresses cytochrome c-encoding genes as measured by in situ molecular detection techniques.

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