AUTHOR=Beller Harry R., Zhou Peng , Legler Tina C., Chakicherla Anu , Kane Staci , Letain Tracy E., O'Day Peggy A.

TITLE=Genome-enabled studies of anaerobic, nitrate-dependent iron oxidation in the chemolithoautotrophic bacterium Thiobacillus denitrificans

JOURNAL=Frontiers in Microbiology

VOLUME=4

YEAR=2013

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2013.00249

DOI=10.3389/fmicb.2013.00249

ISSN=1664-302X

ABSTRACT=<p><italic>Thiobacillus denitrificans</italic> is a chemolithoautotrophic bacterium capable of anaerobic, nitrate-dependent U(IV) and Fe(II) oxidation, both of which can strongly influence the long-term efficacy of <italic>in situ</italic> reductive immobilization of uranium in contaminated aquifers. We previously identified two <italic>c</italic>-type cytochromes involved in nitrate-dependent U(IV) oxidation in <italic>T. denitrificans</italic> and hypothesized that <italic>c</italic>-type cytochromes would also catalyze Fe(II) oxidation, as they have been found to play this role in anaerobic phototrophic Fe(II)-oxidizing bacteria. Here we report on efforts to identify genes associated with nitrate-dependent Fe(II) oxidation, namely (a) whole-genome transcriptional studies [using FeCO<sub>3</sub>, Fe<sup>2</sup><sup>+</sup>, and U(IV) oxides as electron donors under denitrifying conditions], (b) Fe(II) oxidation assays performed with knockout mutants targeting primarily highly expressed or upregulated <italic>c</italic>-type cytochromes, and (c) random transposon-mutagenesis studies with screening for Fe(II) oxidation. Assays of mutants for 26 target genes, most of which were <italic>c</italic>-type cytochromes, indicated that none of the mutants tested were significantly defective in nitrate-dependent Fe(II) oxidation. The non-defective mutants included the <italic>c</italic><sub>1</sub>-cytochrome subunit of the cytochrome <italic>bc</italic><sub>1</sub> complex (complex III), which has relevance to a previously proposed role for this complex in nitrate-dependent Fe(II) oxidation and to current concepts of reverse electron transfer. A transposon mutant with a disrupted gene associated with NADH:ubiquinone oxidoreductase (complex I) was ~35% defective relative to the wild-type strain; this strain was similarly defective in nitrate reduction with thiosulfate as the electron donor. Overall, our results indicate that nitrate-dependent Fe(II) oxidation in <italic>T. denitrificans</italic> is not catalyzed by the same <italic>c</italic>-type cytochromes involved in U(IV) oxidation, nor have other <italic>c</italic>-type cytochromes yet been implicated in the process.</p>