Oligo-heterotrophic activity of marinobacter subterrani creates an indirect fe(ii) oxidation phenotype in gradient tubes

Abhiney Jain, Benjamin M. Bonis, Jeffrey A. Gralnick

Research output: Contribution to journalArticlepeer-review

5 Scopus citations


Autotrophic bacteria utilizing Fe(II) as their energy and electron sources for growth affect multiple biogeochemical cycles. Some chemoheterotrophic bacteria have also been considered to exhibit an Fe(II) oxidation phenotype. For example, several Marinobacter strains have been reported to oxidize Fe(II) based on formation of oxidized iron bands in semi-solid gradient tubes that produce opposing concentration gradients of Fe(II) and oxygen. While gradient tubes are a simple and visually compelling method to test for Fe(II) oxidation, this method alone cannot confirm if, and to what extent, Fe(II) oxidation is linked to metabolism in chemoheterotrophic bacteria. Here we probe the possibility of protein-mediated and metabolic by-product-mediated Fe(II) oxidation in Marinobacter subterrani JG233, a chemoheterotroph previously proposed to oxidize Fe(II). Results from conditional and mutant studies, along with measurements of Fe(II) oxidation rates, suggest M. subterrani is unlikely to facilitate Fe(II) oxidation under microaerobic conditions. We conclude that the Fe(II) oxidation phenotype observed in gradient tubes inoculated with M. subterrani JG233 is a result of oligo-heterotrophic activity, shifting the location where oxygen dependent chemical Fe(II) oxidation occurs, rather than a biologically mediated process. IMPORTANCE Gradient tubes are the most commonly used method to isolate and identify neutrophilic Fe(II)-oxidizing bacteria. The formation of oxidized iron bands in gradient tubes provides a compelling assay to ascribe the ability to oxidize Fe(II) to autotrophic bacteria whose growth is dependent on Fe(II) oxidation. However, the physiological significance of Fe(II) oxidation in chemoheterotrophic bacteria is less well understood. Our work suggests that oligo-heterotrophic activity of certain bacteria may create a false-positive phenotype in gradient tubes by altering the location of the abiotic, oxygen-mediated oxidized iron band. Based on the results and analysis presented here, we caution against utilizing gradient tubes as the sole evidence for the capability of a strain to oxidize Fe(II) and that additional experiments are necessary to ascribe this phenotype to new isolates.

Original languageEnglish (US)
Article numbere01367-21
JournalApplied and environmental microbiology
Issue number24
StatePublished - Nov 2021

Bibliographical note

Funding Information:
This research was supported by the Office of Naval Research (Award N00014-12-1-0309 to J.A.G.) and the Minnesota Environment and Natural Resources Trust Fund.

Publisher Copyright:
© 2021 American Society for Microbiology. All rights reserved.


  • Fe(II) oxidation
  • Gradient tubes
  • Iron oxidizers
  • Marinobacter
  • Oligo-heterotrophy
  • Marinobacter/metabolism
  • Phenotype
  • Oxidation-Reduction
  • Ferrous Compounds/metabolism

PubMed: MeSH publication types

  • Research Support, U.S. Gov't, Non-P.H.S.
  • Journal Article


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