In vivo imaging and tracking of host–microbiota interactions via metabolic labeling of gut anaerobic bacteria

Abstract

The intestine is densely populated by anaerobic commensal bacteria. These microorganisms shape immune system development, but understanding of host–commensal interactions is hampered by a lack of tools for studying the anaerobic intestinal environment. We applied metabolic oligosaccharide engineering and bioorthogonal click chemistry to label various commensal anaerobes, including Bacteroides fragilis, a common and immunologically important commensal. We studied the dissemination of B. fragilis after acute peritonitis and characterized the interactions of the intact microbe and its polysaccharide components in myeloid and B cell lineages. We were able to assess the distribution and colonization of labeled B. fragilis along the intestine, as well as niche competition after coadministration of multiple species of the microbiota. We also fluorescently labeled nine additional commensals (eight anaerobic and one microaerophilic) from three phyla common in the gut—Bacteroidetes, Firmicutes and Proteobacteria—as well as one aerobic pathogen (Staphylococcus aureus). This strategy permits visualization of the anaerobic microbial niche by various methods, including intravital two-photon microscopy and non-invasive whole-body imaging, and can be used to study microbial colonization and host–microbe interactions in real time.

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Figure 1: Generation of fluorescent anaerobic commensal gut bacteria by MOE-BCC.
Figure 2: MOE-BCC labeling preserves the immunoregulatory activity of PSA in B. fragilis.
Figure 3: In vivo uptake of AF647-labeled B. fragilis after acute infection.
Figure 4: Dissemination of AF647-labeled B. fragilis and PSA to secondary lymphoid organs.
Figure 5: In vivo tracking and visualization of B. fragilis in the intestine.
Figure 6: Labeling of various anaerobic gut commensals and assessment of species competition.

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Acknowledgements

We thank the Neurobiology Imaging Facility for consultation and instrument availability that supported this work. This facility is supported in part by the Neural Imaging Center as part of NINDS P30 Core Center grant NS072030. We also thank N. Barteneva of the Flow and Imaging Cytometry Resource for help in whole-body imaging on the IVIS Spectrum (funded by grants NIH 1 S10 OD016401 and BCH/PCMM); J. Czupryna and M. Guerra from PerkinElmer for consultation and expert advice; C. Araneo (Division of Immunology Flow Cytometry Core Facility, HMS) and L. Comstock for helpful discussions, comments and reagents; and members of the Kasper and von Andrian labs for helpful discussions. This work was also partly supported by the US National Institutes of Health (grants PO1 AI1112521, RO1 AI111595 (to U.H.v.A.) and 5T32 HL066987 (to D.A.)). Additional support to U.H.v.A. was provided by U19AI095261 and the Ragon Institute at MGH, MIT and Harvard. N.G.-Z. was supported by an EMBO Long-Term Fellowship (ALTF 251-2011), the Human Frontiers Science Program (LT000079/2012), a UNESCO L'Oreal National and International Women in Science Award, a Fulbright Award and the Weizmann Institute of Science–National Postdoctoral Award Program for Advancing Women in Science. J.E.H. was supported by the Cancer Research Institute Irvington Fellowship Program. N.C.R. was supported by a departmental Kirschstein-NRSA training grant.

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N.G.-Z. and D.A. designed the experiments, analyzed the data and wrote the manuscript with help from J.E.H. and N.C.R. D.E.-H. and S.D. provided experimental help and expertise. D.L.K. supervised the study, edited the manuscript and provided helpful comments, with assistance from U.H.v.A.

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Correspondence to Dennis L Kasper.

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Geva-Zatorsky, N., Alvarez, D., Hudak, J. et al. In vivo imaging and tracking of host–microbiota interactions via metabolic labeling of gut anaerobic bacteria. Nat Med 21, 1091–1100 (2015). https://doi.org/10.1038/nm.3929

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