Poster Session I - Poster of Distinction I - A21 DISRUPTION OF MUCIN-DERIVED SUGAR METABOLISM PATHWAYS IMPAIRS ENTERIC PATHOGEN CITROBACTER RODENTIUM COLONIZATION

TL;DR

Disrupting sugar metabolism in gut bacteria weakens their ability to infect the intestines, offering a new way to fight gut infections.

Contribution

Identifies a conserved metabolic vulnerability in Enterobacteriaceae pathogens by targeting GlcNAc-6P metabolism.

Findings

Disruption of nagA in Citrobacter rodentium impairs colonization and increases sensitivity to stress.

ΔnagA mutants accumulate GlcNAc-6P and show disrupted GlcN-6P synthesis.

Targeting amino sugar catabolic pathways could weaken gut pathogens like E. coli and Salmonella.

Abstract

Enteric bacterial pathogens within the Enterobacteriaceae family, including Escherichia coli and Salmonella species, can cause acute gastroenteritis in humans. In the competitive gastrointestinal environment, these pathogens depend on specific metabolic adaptations to establish infections. The intestinal mucus barrier, composed primarily of the mucin Muc2, protects the epithelium while also serving as a nutrient reservoir rich in host-derived monosaccharides such as N-acetylglucosamine (GlcNAc) and N-acetylneuraminic acid (NeuNAc). How these mucus-derived sugars are metabolized during infection and how dysregulation of these pathways affects bacterial pathogen fitness remain poorly understood.

We sought to determine how Enterobacteriaceae exploit mucin-derived sugars for colonization and whether disrupting these pathways impairs their fitness. Citrobacter rodentium, a murine-specific enteric pathogen, was used as a model system and was hypothesized to be able to exploit GlcNAc and NeuNAc to fuel their pathogenesis.

C. rodentium mutants were generated, including ΔnagA lacking GlcNAc-6P deacetylase, its complemented strain, and Δmana lacking GlcNAc/NeuNAc import systems. Colonization and tissue pathology were examined in C57BL/6J mice. Intracellular GlcNAc-6P was quantified by the Morgan–Elson assay. Transcriptional responses were analyzed by RT-qPCR. Bacterial susceptibility to cell wall stress was tested using lysozyme, osmotic, and antibiotic challenges.

Deletion of nagA severely attenuated colonization and pathology in mice, while Δmana remained fully infective, indicating that virulence loss was not solely due to nutrient deprivation. ΔnagA accumulated GlcNAc-6P, grew poorly in vitro, upregulated nagB transcription, suggesting a disrupted glucosamine-6-phosphate (GlcN-6P) synthesis. Functionally, ΔnagA exhibited increased sensitivity to lysozyme, osmotic stress, and cell wall-targeting antibiotics, indicating that cell wall integrity was compromised.

Disrupting nagA in C. rodentium causes GlcNAc-6P accumulation and disrupted GlcN-6P synthesis, compromising cell wall integrity and in vivo fitness. As these amino sugar catabolic pathways are conserved across Enterobacteriaceae, this work identifies a previously unrecognized metabolic vulnerability that could be exploited to weaken enteric pathogens such as pathogenic E. coli and Salmonella. Targeting sugar-phosphate stress pathways offers a promising approach to limit bacterial infections of the gastrointestinal mucosa.

CIHRNatural Sciences and Engineering Research Council of Canada

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