Investigating the influence of commensal Limosilactobacillus reuteri and Ruminococcus gnavus on infection with enteropathogenic Escherichia coli in a microaerobic human intestinal cell model.

Mcgrath, Conor (2022) Investigating the influence of commensal Limosilactobacillus reuteri and Ruminococcus gnavus on infection with enteropathogenic Escherichia coli in a microaerobic human intestinal cell model. Doctoral thesis, University of East Anglia.

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Abstract

The gut microbiota has an important role in maintaining intestinal health and protecting against enteric infections (colonisation resistance). Nevertheless, most of these interactions haven’t been explored, largely due to a lack of experimental model systems that can culture oxygen-sensitive commensals alongside intestinal cells. In this project, we have established a novel in vitro model system of the human intestinal epithelium (Vertical Diffusion Chamber, VDC) which supports growth of strictly anaerobic bacteria. We have applied this system to investigate the interactions of gut commensals Ruminococcus gnavus and Limosilactobacillus reuteri with a functioning mucus-producing epithelium, established using T84 and goblet-like LS174T cell lines, and their effect on infection with enteric pathogen enteropathogenic E. coli (EPEC).
Previously published work identified a culture medium that supports commensal and EPEC growth whilst maintaining epithelial integrity and barrier function. This was achieved by establishing bacterial growth curves in different media and assessing epithelial barrier function by transepithelial electrical resistance and immunofluorescence staining (IFS) of tight-junction protein occludin. Further IFS demonstrated that introduction of LS174T cells to the epithelium caused mucin secretion and facilitates colonisation by commensals. Co-culture of EPEC with commensals reduces numbers of viable and adherent EPEC, as well secretion of pro-inflammatory cytokine interleukin-8 by the infected epithelia. For R. gnavus, reduced EPEC viability and adherence is only observed when LS174T are present.
Here, we build on existing work showing potential colonisation resistance activities which would not be possible to study using traditional cell culture models. As the resident bacterial of the human gut are predominantly oxygen-sensitive, this system can be used to study a plethora of host-pathogen-commensal interactions and aid development of probiotic therapies.

Item Type: Thesis (Doctoral)
Faculty \ School: Faculty of Medicine and Health Sciences > Norwich Medical School
Depositing User: Kitty Laine
Date Deposited: 15 Jun 2022 10:48
Last Modified: 15 Jun 2022 10:48
URI: https://ueaeprints.uea.ac.uk/id/eprint/85628
DOI:

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