Aerobic soil bacteria adapt to hypoxia by hybridizing fermentation with carbon storage

D David L. Gillett (Department of Microbiology, Monash University) T Thilini Koralegedara (Department of Microbiology, Monash University) T Tess F. Hutchinson (Department of Microbiology, Monash University) T Thanh Nguyen-Dinh (Department of Microbiology, Monash University) T Thomas D. Watts (Department of Microbiology, Monash University) J Jessica Solari (Department of Microbiology, Monash University) M Manasi Mudaliyar (Department of Biochemistry and Pharmacology, Bio21 Institute, The University of Melbourne) W William J. Jowsey (Department of Microbiology and Immunology, University of Otago) N Nadeesha Athukorala (Department of Microbiology, Monash University) W Wei Wen Wong (Water Studies, School of Chemistry, Monash University) J Jake U. S. Locop (Department of Microbiology, Monash University) N Ning Hall (Department of Microbiology, Monash University) L Luis Jimenez (Department of Microbiology, Monash University) I Iresha Hanchapola (Monash Proteomics and Metabolomics Platform, Department of Biochemistry and Molecular Biology, Monash Biomedicine Discovery Institute, Monash University) H Han-Chung Lee (Monash Proteomics and Metabolomics Platform, Department of Biochemistry and Molecular Biology, Monash Biomedicine Discovery Institute, Monash University) E Erwin Tanuwidjaya (Monash Proteomics and Metabolomics Platform, Department of Biochemistry and Molecular Biology, Monash Biomedicine Discovery Institute, Monash University) J Joel R. Steele (Monash Proteomics and Metabolomics Platform, Department of Biochemistry and Molecular Biology, Monash Biomedicine Discovery Institute, Monash University) R Ralf B. Schittenhelm (Monash Proteomics and Metabolomics Platform, Department of Biochemistry and Molecular Biology, Monash Biomedicine Discovery Institute, Monash University) C Christopher K. Barlow R Rhys Grinter (Department of Microbiology, Biomedicine Discovery Institute, Monash University) G Gregory M. Cook (Department of Microbiology and Immunology, University of Otago) M Matthew B. McNeil (Department of Microbiology and Immunology, University of Otago) D Debnath Ghosal (Department of Biochemistry and Pharmacology, Bio21 Institute, The University of Melbourne) P Pok Man Leung (Department of Microbiology, Biomedicine Discovery Institute, Monash University) P Perran L. M. Cook (Water Studies, School of Chemistry, Monash University) C Chris Greening

Abstract

In soil ecosystems, aerobic bacteria survive oxygen deprivation (hypoxia) by entering nonreplicative persistent states. In contrast to the well-studied metabolism of obligate and facultative anaerobes, little is known about how obligately aerobic bacteria adapt their metabolism to stay viable during hypoxia. The model obligate aerobe Mycobacterium smegmatis maintains redox homeostasis during hypoxia by mediating fermentative hydrogen production. However, the fate of organic carbon during fermentation is unresolved. Here we systematically profiled the metabolism of M. smegmatis during aerobic growth, hypoxic persistence, and the transition between these states. By integrating a differentially 13 C-labeled glucose isotopologue assay with paired metabolomics and proteomics, we observed M. smegmatis rerouted central carbon metabolism through the pentose phosphate pathway and/or Entner–Doudoroff pathways during hypoxia, while excreting high levels of hydrogen and acetate. Lipid and cryoelectron tomography analyses suggest M. smegmatis also stores carbon as glycerides in lipid droplets during hypoxia, which serve as a major reductant sink. Gene knockouts and knockdowns revealed that, while M. smegmatis depends on its hydrogen-producing hydrogenase for hypoxic survival, it can compensate for the disruption of acetate production and glyceride synthesis by producing and excreting other organic acids. We confirmed through an extensive genomic survey and biogeochemical measurements that diverse aerobic soil bacteria can store organic carbon and mediate fermentation during hypoxia. Altogether, this hybrid fermentative metabolism likely provides a competitive advantage in soils and other resource-variable environments by enabling bacteria, such as M. smegmatis , to simultaneously dispose excess reductant and maintain carbon stores during hypoxia.

Article Details

Volume / Issue Vol. 123, Issue 23
Published June 09, 2026
ISSN 0027-8424
Publisher National Academy of Sciences

Authors (26)

D

David L. Gillett

Department of Microbiology, Monash University

T

Thilini Koralegedara

Department of Microbiology, Monash University

T

Tess F. Hutchinson

Department of Microbiology, Monash University

T

Thanh Nguyen-Dinh

Department of Microbiology, Monash University

T

Thomas D. Watts

Department of Microbiology, Monash University

J

Jessica Solari

Department of Microbiology, Monash University

M

Manasi Mudaliyar

Department of Biochemistry and Pharmacology, Bio21 Institute, The University of Melbourne

W

William J. Jowsey

Department of Microbiology and Immunology, University of Otago

N

Nadeesha Athukorala

Department of Microbiology, Monash University

W

Wei Wen Wong

Water Studies, School of Chemistry, Monash University

J

Jake U. S. Locop

Department of Microbiology, Monash University

N

Ning Hall

Department of Microbiology, Monash University

L

Luis Jimenez

Department of Microbiology, Monash University

I

Iresha Hanchapola

Monash Proteomics and Metabolomics Platform, Department of Biochemistry and Molecular Biology, Monash Biomedicine Discovery Institute, Monash University

H

Han-Chung Lee

Monash Proteomics and Metabolomics Platform, Department of Biochemistry and Molecular Biology, Monash Biomedicine Discovery Institute, Monash University

E

Erwin Tanuwidjaya

Monash Proteomics and Metabolomics Platform, Department of Biochemistry and Molecular Biology, Monash Biomedicine Discovery Institute, Monash University

J

Joel R. Steele

Monash Proteomics and Metabolomics Platform, Department of Biochemistry and Molecular Biology, Monash Biomedicine Discovery Institute, Monash University

R

Ralf B. Schittenhelm

Monash Proteomics and Metabolomics Platform, Department of Biochemistry and Molecular Biology, Monash Biomedicine Discovery Institute, Monash University

C

Christopher K. Barlow

R

Rhys Grinter

Department of Microbiology, Biomedicine Discovery Institute, Monash University

G

Gregory M. Cook

Department of Microbiology and Immunology, University of Otago

M

Matthew B. McNeil

Department of Microbiology and Immunology, University of Otago

D

Debnath Ghosal

Department of Biochemistry and Pharmacology, Bio21 Institute, The University of Melbourne

P

Pok Man Leung

Department of Microbiology, Biomedicine Discovery Institute, Monash University

P

Perran L. M. Cook

Water Studies, School of Chemistry, Monash University

C

Chris Greening