Abstract

The gut microbiota affects many important host functions, including the immune response and the nervous system1. However, while substantial progress has been made in growing diverse microorganisms of the microbiota2, 23–65% of species residing in the human gut remain uncultured3,4, which is an obstacle for understanding their biological roles. A likely reason for this unculturability is the absence in artificial media of key growth factors that are provided by neighbouring bacteria in situ5,6. In the present study, we used co-culture to isolate KLE1738, which required the presence of Bacteroides fragilis to grow. Bioassay-driven purification of B. fragilis supernatant led to the isolation of the growth factor, which, surprisingly, is the major inhibitory neurotransmitter GABA (γ-aminobutyric acid). GABA was the only tested nutrient that supported the growth of KLE1738, and a genome analysis supported a GABA-dependent metabolism mechanism. Using growth of KLE1738 as an indicator, we isolated a variety of GABA-producing bacteria, and found that Bacteroides ssp. produced large quantities of GABA. Genome-based metabolic modelling of the human gut microbiota revealed multiple genera with the predicted capability to produce or consume GABA. A transcriptome analysis of human stool samples from healthy individuals showed that GABA-producing pathways are actively expressed by Bacteroides, Parabacteroides and Escherichia species. By coupling 16S ribosmal RNA sequencing with functional magentic resonance imaging in patients with major depressive disorder, a disease associated with an altered GABA-mediated response, we found that the relative abundance levels of faecal Bacteroides are negatively correlated with brain signatures associated with depression.

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Data availability

The 16S rRNA sequence and genome data for KLE1738 are available from the NCBI (MH636586 and PRJNA482656, respectively). The American Gut sequence data are available from the EBI under accession ERP012803. fMRI data are available at the discretion of M.J.D. All other data that support the findings of this study are available from the corresponding author upon request.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

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Acknowledgements

The authors would like to thank S. Rubin for help with the cultivation of KLE1738, and J. Wang of the Small Molecule Mass Spectrometry Facility, a Harvard Faculty of Arts and Science (FAS) Division of Science Core Facility, for analysing the GC/MS samples. This work was supported by grants R01HG005824 to K.L., R01GM086158 to J.C. and F32GM108415 to T.R.R.

Author information

Author notes

  1. These authors contributed equally: Philip Strandwitz, Ki Hyun Kim.

Affiliations

  1. Antimicrobial Discovery Center, Department of Biology, Northeastern University, Boston, MA, USA

    • Philip Strandwitz
    • , Asama Lekbua
    • , Nader Mroue
    • , Eric J. Stewart
    •  & Kim Lewis
  2. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA, USA

    • Ki Hyun Kim
    • , David Dietrich
    • , Timothy R. Ramadhar
    •  & Jon Clardy
  3. School of Pharmacy, Sungkyunkwan University, Suwon, Republic of Korea

    • Ki Hyun Kim
  4. Department of Psychiatry and Brain and Mind Research Institute, Weill Cornell Medical College, New York, NY, USA

    • Darya Terekhova
    • , Conor Liston
    •  & Marc J. Dubin
  5. Bioinformatics and Systems Biology Program, University of California San Diego, La Jolla, CA, USA

    • Joanne K. Liu
  6. Department of Surgery, University of Chicago, Chicago, IL, USA

    • Anukriti Sharma
    •  & Jack A. Gilbert
  7. Biosciences Division (BIO), Argonne National Laboratory, Argonne, IL, USA

    • Anukriti Sharma
    •  & Jack A. Gilbert
  8. Center for Microbiome Innovation, University of California San Diego, La Jolla, CA, USA

    • Jennifer Levering
    • , Karsten Zengler
    •  & Rob Knight
  9. Department of Pediatrics, University of California San Diego, La Jolla, CA, USA

    • Daniel McDonald
    • , Karsten Zengler
    •  & Rob Knight
  10. Department of Chemistry, Howard University, Washington, DC, USA

    • Timothy R. Ramadhar
  11. Department of Computer Science and Engineering, University of California San Diego, La Jolla, CA, USA

    • Rob Knight
  12. Marine Biological Laboratory, Woods Hole, MA, USA

    • Jack A. Gilbert

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Contributions

P.S. and K.L. planned the study, analysed the data and wrote the paper. P.S. performed the co-culture screening for helper-dependent pairs and general KLE1738 cultivation. P.S., J.C. and K.H.K. designed and performed the bioassay-driven screening for GABA. P.S. and N.M. performed media exclusion experiments. P.S., E.J.S. and D.D. analysed the KLE1738 genome. P.S., D.D. and T.R.R. performed 13C feeding experiments. P.S. cultivated B. fragilis and D.D. analysed the supernatant for GABA and glutamate. P.S. designed and with A.L. executed the screen for GABA-producing bacteria. P.S. prepared the supernatant of GABA producers identified in the KLE1738 co-culture screen, and D.D. analysed these supernatants for GABA production. P.S., D.M., R.K., J.L., J.K.L. and K.Z. performed the metagenomic analysis for GABA producers and consumers, and P.S., A.S., and J.A.G. analysed the human transcriptome dataset. P.S., K.L., D.T. and M.J.D. designed the MDD study, and P.S., K.L., A.S., J.A.G., D.T., C.L. and M.J.D. analysed the MDD data. All authors helped edit the manuscript.

Competing interests

P.S. and K.L. declare competing financial interests as they are founders of Holobiome, Inc. All other authors have no competing interests.

Corresponding authors

Correspondence to Philip Strandwitz or Kim Lewis.

Supplementary information

  1. Supplementary Information

    Supplementary Figures 1–8, Supplementary Tables 1–4 and 6–8.

  2. Reporting Summary

  3. Supplementary Table 5

    Genome analysis of GABA modulating potential of 1,159 gut bacterial genomes, consisting of 919 species.

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DOI

https://doi.org/10.1038/s41564-018-0307-3