Letter | Published:

An exclusive metabolic niche enables strain engraftment in the gut microbiota

Naturevolume 557pages434438 (2018) | Download Citation


The dense microbial ecosystem in the gut is intimately connected to numerous facets of human biology, and manipulation of the gut microbiota has broad implications for human health. In the absence of profound perturbation, the bacterial strains that reside within an individual are mostly stable over time1. By contrast, the fate of exogenous commensal and probiotic strains applied to an established microbiota is variable, generally unpredictable and greatly influenced by the background microbiota2,3. Therefore, analysis of the factors that govern strain engraftment and abundance is of critical importance to the emerging field of microbiome reprogramming. Here we generate an exclusive metabolic niche in mice via administration of a marine polysaccharide, porphyran, and an exogenous Bacteroides strain harbouring a rare gene cluster for porphyran utilization. Privileged nutrient access enables reliable engraftment of the exogenous strain at predictable abundances in mice harbouring diverse communities of gut microbes. This targeted dietary support is sufficient to overcome priority exclusion by an isogenic strain4, and enables strain replacement. We demonstrate transfer of the 60-kb porphyran utilization locus into a naive strain of Bacteroides, and show finely tuned control of strain abundance in the mouse gut across multiple orders of magnitude by varying porphyran dosage. Finally, we show that this system enables the introduction of a new strain into the colonic crypt ecosystem. These data highlight the influence of nutrient availability in shaping microbiota membership, expand the ability to perform a broad spectrum of investigations in the context of a complex microbiota, and have implications for cell-based therapeutic strategies in the gut.

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  1. 1.

    Faith, J. J. et al. The long-term stability of the human gut microbiota. Science 341, 1237439 (2013).

  2. 2.

    Frese, S. A., Hutkins, R. W. & Walter, J. Comparison of the colonization ability of autochthonous and allochthonous strains of lactobacilli in the human gastrointestinal tract. Adv. Microbiol. 2, 399–409 (2012).

  3. 3.

    Maldonado-Gómez, M. X. et al. Stable engraftment of Bifidobacterium longum AH1206 in the human gut depends on individualized features of the resident microbiome. Cell Host Microbe 20, 515–526 (2016).

  4. 4.

    Lee, S. M. et al. Bacterial colonization factors control specificity and stability of the gut microbiota. Nature 501, 426–429 (2013).

  5. 5.

    Ivanov, I. I. et al. Induction of intestinal Th17 cells by segmented filamentous bacteria. Cell 139, 485–498 (2009).

  6. 6.

    Atarashi, K. et al. Induction of colonic regulatory T cells by indigenous Clostridium species. Science 331, 337–341 (2011).

  7. 7.

    Lawley, T. D. et al. Targeted restoration of the intestinal microbiota with a simple, defined bacteriotherapy resolves relapsing Clostridium difficile disease in mice. PLoS Pathog. 8, e1002995 (2012).

  8. 8.

    Stecher, B. et al. Like will to like: abundances of closely related species can predict susceptibility to intestinal colonization by pathogenic and commensal bacteria. PLoS Pathog. 6, e1000711 (2010).

  9. 9.

    Ratner, M. Seres’s pioneering microbiome drug fails mid-stage trial. Nat. Biotechnol. 34, 1004–1005 (2016).

  10. 10.

    Ericsson, A. C., Personett, A. R., Turner, G., Dorfmeyer, R. A. & Franklin, C. L. Variable colonization after reciprocal fecal microbiota transfer between mice with low and high richness microbiota. Front. Microbiol. 8, 196 (2017).

  11. 11.

    Landy, J. et al. Variable alterations of the microbiota, without metabolic or immunological change, following faecal microbiota transplantation in patients with chronic pouchitis. Sci. Rep. 5, 12955 (2015).

  12. 12.

    Whitaker, W. R., Shepherd, E. S. & Sonnenburg, J. L. Tunable expression tools enable single-cell strain distinction in the gut microbiome. Cell 169, 538–546 (2017).

  13. 13.

    Sonnenburg, E. D. et al. Diet-induced extinctions in the gut microbiota compound over generations. Nature 529, 212–215 (2016).

  14. 14.

    Ridaura, V. K. et al. Gut microbiota from twins discordant for obesity modulate metabolism in mice. Science 341, 1241214 (2013).

  15. 15.

    Hehemann, J. H. et al. Transfer of carbohydrate-active enzymes from marine bacteria to Japanese gut microbiota. Nature 464, 908–912 (2010).

  16. 16.

    David, L. A. et al. Diet rapidly and reproducibly alters the human gut microbiome. Nature 505, 559–563 (2014).

  17. 17.

    Sonnenburg, J. L. et al. Glycan foraging in vivo by an intestine-adapted bacterial symbiont. Science 307, 1955–1959 (2005).

  18. 18.

    Xu, J. et al. Evolution of symbiotic bacteria in the distal human intestine. PLoS Biol. 5, e156 (2007).

  19. 19.

    Sonnenburg, E. D. et al. Specificity of polysaccharide use in intestinal Bacteroides species determines diet-induced microbiota alterations. Cell 141, 1241–1252 (2010).

  20. 20.

    Hehemann, J. H., Kelly, A. G., Pudlo, N. A., Martens, E. C. & Boraston, A. B. Bacteria of the human gut microbiome catabolize red seaweed glycans with carbohydrate-active enzyme updates from extrinsic microbes. Proc. Natl Acad. Sci. USA 109, 19786–19791 (2012).

  21. 21.

    Drake, J. A. Community-assembly mechanics and the structure of an experimental species ensemble. Am. Nat. 137, 1–26 (1991).

  22. 22.

    Fukami, T. historical contingency in community assembly: integrating niches, species pools, and priority effects. Annu. Rev. Ecol. Evol. Syst. 46, 1–23 (2015).

  23. 23.

    Chandran, S. & Shapland, E. in Synthetic DNA Vol. 1472 (ed. Hughes, R. A.) 187–192 (Humana, New York, 2017).

  24. 24.

    Martens, E. C., Chiang, H. C. & Gordon, J. I. Mucosal glycan foraging enhances fitness and transmission of a saccharolytic human gut bacterial symbiont. Cell Host Microbe 4, 447–457 (2008).

  25. 25.

    Simon, R., Priefer, U. & Pühler, A. A broad host range mobilization system for in vivo genetic engineering: transposon mutagenesis in gram negative bacteria. Nat. Biotechnol. 1, 784–791 (1983).

  26. 26.

    Ostrov, N. et al. Design, synthesis, and testing toward a 57-codon genome. Science 353, 819–822 (2016).

  27. 27.

    Caporaso, J. G. et al. QIIME allows analysis of high-throughput community sequencing data. Nat. Methods 7, 335–336 (2010).

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We thank N. Ratnayeke for early experimental assistance, S. Higginbottom for gnotobiotic assistance, and K. Ng, Z. Russ and W. Van Treuren for analytical assistance; the Amieva and Huang laboratories for use of their microscopy resources, and D. Shepherd for valuable discussions; N. Pudlo and E. Martens for the protocol on porphyran extraction, and E. Sonnenburg, T. Fukami and M. Fischbach for commenting on this manuscript. This material is based upon work supported by the National Science Foundation under grant number 1648230, the NIDDK (R01-DK085025 to J.L.S.) and an NSF Graduate Fellowship (DGE-114747 to E.S.S.).

Reviewer information

Nature thanks D. Bolam and the other anonymous reviewer(s) for their contribution to the peer review of this work.

Author information


  1. Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, USA

    • Elizabeth Stanley Shepherd
    • , Kali M. Pruss
    •  & Justin L. Sonnenburg
  2. Novome Biotechnologies, South San Francisco, CA, USA

    • Elizabeth Stanley Shepherd
    • , William C. DeLoache
    •  & Weston R. Whitaker


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E.S.S. and J.L.S. generated the idea for the project. E.S.S. performed the in vivo studies, 16 S sample preparation and analysis and in vitro growth studies. W.C.D. sequenced and analysed the NB001 genome and constructed the porphyran PUL plasmids. K.M.P. and E.S.S. performed in vivo crypt studies, imaging and analysis. W.R.W. isolated NB001 and performed in vitro growth characterization. All authors contributed to experimental design and data analysis, and E.S.S. and J.L.S. wrote the manuscript. All authors discussed the results and commented on the manuscript.

Competing interests

E.S.S., W.C.D., W.R.W. and J.L.S. are founders at Novome Biotechnologies, Inc. and have filed a provisional patent based on the work described here (US Provisional Patent No. 62/435,048).

Corresponding author

Correspondence to Justin L. Sonnenburg.

Extended data figures and tables

  1. Extended Data Fig. 1 Three model background communities of gut microbes are distinct from each other.

    a, Principal coordinates (PC) analysis of weighted UniFrac distance for 16 S rRNA gene amplicons from faeces from the three background community groups from Fig. 1 before diet switch for conventional (RF) or humanized (Hum-1 and Hum-2) mice, n = 10. b, Comparison of weighted UniFrac distances within each group (Intra) or across groups (A × B, A × C, B × C). One-way ANOVA, P < 0.0001.

  2. Extended Data Fig. 2 NB001 can utilize both inulin and porphyran as the only carbon source for growth.

    a, NB001 demonstrates growth in minimal medium with either glucose (blue, doubling time = 157 min), or inulin (orange, doubling time = 127 min), as the only provided carbon source. b, Schematic of porphyran PUL from NB001 based on alignment to the previously published B. plebeius PUL, based on data from whole-genome sequencing. Grey bar, the region deleted via homologous recombination to abolish the ability to utilize porphyran. c, NB001 has the ability to grow on porphyran (doubling time = 98 min) as the only carbon source (WT), and growth is abrogated when genes required for porphyran utilization are knocked out (KO).

  3. Extended Data Fig. 3 Porphyran does not significantly impact the gut microbiota in the absence of a known utilizer.

    Weighted UniFrac analysis was performed on faecal 16 S rRNA data for conventional mice colonized with a porphyran utilization knockout (as in Fig. 2c) before (Pre, n = 8) or after (Post, n = 9) addition of porphyran. a, Principal coordinates analysis. b, Weighted UniFrac analysis. Unpaired two-tailed t-test, P = 0.25 (n.s., not significant).

  4. Extended Data Fig. 4 Primary colonizer displacement is robust and contingent upon access to porphyran.

    a, Conventional mice (n = 7), which were fed a MAC-rich diet and colonized with NB001 (PUL 1) containing an eight-gene deletion abrogating its ability to utilize porphyran (Extended Data Fig. 2b, c), demonstrated resistance to subsequent challenge with an isogenic knockout strain (PUL 2) in the presence of 1% porphyran in the drinking water. Notably, our conventionally raised mice were permissive to colonization by this strain and other tested species of Bacteroides (B. thetaiotaomicron, B. fragilis, B. uniformis, B. vulgatus, stable colonization range of 8 × 105–3 × 108 c.f.u. per ml faeces), which differs from reports of tests on other conventionally raised mice, potentially reflecting inter-colony microbiota differences. b, Mice from Fig. 2e were challenged with the originally colonizing porphyran utilization knockout (PUL) that was displaced by the utilizer (PUL+) and demonstrated colonization resistance to the previously displaced knockout strain. Data are mean ± s.d. The grey-shaded boxes represent the limit of detection. Source Data

  5. Extended Data Fig. 5 Minimal porphyran utilization PULs were constructed via PCR and yeast assembly.

    Schematic representing construction of designed porphyran PULs. On the basis of the alignment to the previously published B. plebeius porphyran utilization PUL, three regions were targeted for minimal PUL assembly and amplified via PCR from the NB001 genome. The PCR fragments were assembled with digests of both a custom and commercially available vector in yeast (see Methods), after which colonies carrying correctly assembled plasmids were lysed and directly added to E. coli for electroporation.

  6. Extended Data Fig. 6 B. stercoris and B. thetaiotaomicron demonstrate different abilities to grow in minimal medium.

    a, b, Wild-type B. stercoris (a) and B. thetaiotaomicron (b) grown in SMM with glucose as the only carbon source demonstrate different maximum optical densities reached (B. stercoris maximum OD = 0.363, B. thetaiotaomicron maximum OD = 0.453). This suggests a possible explanation for why both species with the 21-gene PUL grow to different maximum optical densities as well (Fig. 3b).

  7. Extended Data Fig. 7 Abundance of B. stercoris with the 34-gene PUL can be controlled in the context of a conventional mouse microbiota.

    Conventional mice (n = 5) that were fed a MAC-rich diet were colonized with a strain of B. stercoris harbouring the designed 34-gene porphyran PUL and density of the engineered strain was tracked in the faeces. Upon administration of 1% porphyran in the drinking water (green-shaded box), density of B. stercoris increased, and subsequently decreased upon removal of porphyran. Data are mean ± s.d. The grey-shaded box represents the limit of detection. Source Data

  8. Extended Data Table 1 List of plasmids used in this work

Supplementary information

  1. Supplementary Tables 1-3

    This file contains Supplementary Table 1 (lists all oligos used for PCR and construction of fragments for yeast assembly), Supplementary Table 2 (lists the strategies for generating all PCR products used in creating PUL plasmids) and Supplementary Table 3 (lists the digests of commercial and custom vectors used in yeast assembly of PUL plasmids).

  2. Reporting Summary

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