SREL Reprint #2770

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

Andrew L. Neal1, Lani K. Clough2, Todd D. Perkins2, Brenda J. Little3, and Timothy S. Magnuson2

1Department of Microbiology and Savannah River Ecology Laboratory, University of Georgia,
Athens, GA 30602, USA
2Department of Biological Sciences, Idaho State University, Pocatello, ID 83204, USA
3Naval Research Laboratory, Stennis Space Center, MS 39529-5494 Huntsville, AL, USA

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 fer A (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 fer A 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.

Keywords: Iron respiring bacterium; Gene detection; Surface attachment; Mineral transformation; Geobacteraceae

SREL Reprint #2770

Neal, A. L., L. K. Clough, T. D. Perkins, B. J. Little, and T. S. Magnuson. 2004. In situ measurement of Fe(III) reduction activity of Geobacter pelophilus by simultaneous in situ RT-PCR and XPS analysis. FEMS Microbiology Ecology 49:163-169.

This information was provided by the University of Georgia's Savannah River Ecology Laboratory (srel.uga.edu).