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Quantitative whole-tissue 3D imaging reveals bacteria in close association with mouse jejunum mucosa

Poceviciute, Roberta and Bogatyrev, Said R. and Romano, Anna E. and Dilmore, Amanda H. and Mondragón-Palomino, Octavio and Takko, Heli and Ismagilov, Rustem F. (2022) Quantitative whole-tissue 3D imaging reveals bacteria in close association with mouse jejunum mucosa. . (Unpublished)

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The small intestine (SI) is the primary site of nutrient absorption, so its large surface area lacks the thick protective mucus that is characteristic of the large intestine. Because the SI epithelium is relatively exposed, any microbes that colonize the thin mucosa of the SI may exert a substantial effect on the host. Thus far, potential bacterial colonization of the SI mucosa has only been documented in disease states, suggesting mucosal colonization is a rare occurrence, likely requiring multiple perturbations.ResultsHere, we tested whether we could induce bacterial association with jejunum mucosa by a combination of malnutrition and oral co-gavage with a specific bacterial cocktail (E. coli and Bacteroides spp.) that has previously induced environmental enteropathy in mouse models. To overcome the current limitations in imaging and allow definite determination of whether bacterial colonization of the SI mucosa is occurring, we optimized our previously developed whole-tissue three-dimensional (3D) imaging tools with third-generation hybridization chain reaction (HCR v3.0) probes. Only in mice that were malnourished and gavaged with the bacterial cocktail did we detect dense bacterial clusters surrounding intestinal villi suggestive of colonization. Healthy mice gavaged with bacteria and malnourished mice not gavaged with bacteria showed no evidence of mucosal colonization. Furthermore, in malnourished mice gavaged with bacteria we detected villus loss, which may represent one possible consequence that bacterial colonization of the SI mucosa has on the host.ConclusionsOur results suggest that dense bacterial colonization of jejunum mucosa is possible in the presence of multiple perturbations and that villus loss may be one possible consequence to such colonization. Furthermore, our results demonstrate the utility of whole-tissue 3D imaging tools. Although 2D imaging of thin sections may have failed to detect and capture the full spatial complexity of such rare events, whole-tissue 3D imaging tools enabled their detection over large areas of intestinal mucosa and visualization of their spatial complexity in 3D.

Item Type:Report or Paper (Discussion Paper)
Related URLs:
URLURL TypeDescription Paper Dataset Information
Poceviciute, Roberta0000-0002-6649-2170
Bogatyrev, Said R.0000-0003-0486-9451
Romano, Anna E.0000-0002-7148-0668
Dilmore, Amanda H.0000-0001-6493-7116
Mondragón-Palomino, Octavio0000-0003-1129-4932
Takko, Heli0000-0003-2544-409X
Ismagilov, Rustem F.0000-0002-3680-4399
Additional Information:The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY 4.0 International license. We thank the Caltech Office of Laboratory Animal Resources as well as the veterinary technicians at the Wanimal facilities for animal care, personnel training, and resources. We thank Biological Imaging Facility at Caltech (including Andres Collazo, Giada Spigolon, and Steven Wilbert) for resources, training, and technical support. We thank Brett Finley (University of British Columbia) and Prof. Emma Allen Vercoe (University of Guelph) for providing bacterial isolates. We thank Prof. Jared Leadbetter and Prof. Sarkis Mazmanian for providing feedback on study design. We thank Justin Bois for introduction to data analysis in Python. We thank Emily Savela, Mary Arrastia and Eugenia Khorosheva for reviewing and filing Institutional Biosafety Committee paperwork. We thank Jacob T. Barlow for processing sequencing data. We thank Joanne Lau for maintaining anaerobic chambers for bacterial culture. We also thank Natasha Shelby for contributions to writing and editing this manuscript. This work was supported in part by the Kenneth Rainin Foundation (2018-1207), Army Research Office Multidisciplinary University Research Initiative (W911NF-17-1-0402), Defense Advanced Research Projects Agency (HR0011-17-2-0037), and the Jacobs Institute for Molecular Engineering for Medicine. OMP was supported by a Burroughs Welcome Fund Career Award at the Scientific Interface (ID# 106969). The funders had no role in the design of the study, the collection, analysis, and interpretation of data, nor in writing the manuscript. Authors’ contributions. RP Conception, animal study execution, sample collection, sample processing for imaging, imaging, data analysis, figure generation, manuscript preparation. SRB Tail cup study implementation. sample collection. AER RT-qPCR, dPCR, and sequencing data acquisition. AHD dPCR data acquisition. OMP Preliminary imaging and sequencing. HT Preliminary image analysis. RFI Project supervision and administration, acquisition of funding, manuscript review and editing. All authors read and approved the final manuscript. See Supplementary Information for detailed author contributions. Ethics approval and consent to participate. All animal husbandry and experiments were approved by the Caltech Institutional Animal Care and Use Committee (IACUC, protocol #1646). Availability of data and material. All data from this publication are available at Caltech DATA at Competing Interest Statement. This paper is the subject of a patent application filed by Caltech. RFI is a co-founder, consultant, and a director and has stock ownership of Talis Biomedical Corp. In addition, RFI is an inventor on a series of patents licensed by the University of Chicago to Bio-Rad Laboratories Inc. in the context of dPCR.
Group:Jacobs Institute for Molecular Engineering for Medicine
Funding AgencyGrant Number
Kenneth Rainin Foundation2018-1207
Army Research Office (ARO)W911NF-17-1-0402
Defense Advanced Research Projects Agency (DARPA)HR0011-17-2-0037
Joseph J. Jacobs Institute for Molecular Engineering for MedicineUNSPECIFIED
Burroughs Wellcome Fund106969
Record Number:CaltechAUTHORS:20220706-965018000
Persistent URL:
Official Citation:Quantitative whole-tissue 3D imaging reveals bacteria in close association with mouse jejunum mucosa Roberta Poceviciute, Said R. Bogatyrev, Anna E. Romano, Amanda H. Dilmore, Octavio Mondragón-Palomino, Heli Takko, Rustem F. Ismagilov bioRxiv 2022.06.17.496478; doi:
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:115349
Deposited By: George Porter
Deposited On:08 Jul 2022 22:32
Last Modified:28 Feb 2023 21:54

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