Abstract
Enterotoxigenic Escherichia coli (ETEC) is one of the most common causes of diarrheal illness in third world countries and it especially affects children and travelers visiting these regions. ETEC causes disease by adhering tightly to the epithelial cells in a concerted effort by adhesins, flagella, and other virulence-factors. When attached ETEC secretes toxins targeting the small intestine host-cells, which ultimately leads to osmotic diarrhea. HldE is a bifunctional protein that catalyzes the nucleotide-activated heptose precursors used in the biosynthesis of lipopolysaccharide (LPS) and in post-translational protein glycosylation. Both mechanisms have been linked to ETEC virulence: Lipopolysaccharide (LPS) is a major component of the bacterial outer membrane and is needed for transport of heat-labile toxins to the host cells, and ETEC glycoproteins have been shown to play an important role for bacterial adhesion to host epithelia. Here, we report that HldE plays an important role for ETEC virulence. Deletion of hldE resulted in markedly reduced binding to the human intestinal cells due to reduced expression of colonization factor CFA/I on the bacterial surface. Deletion of hldE also affected ETEC motility in a flagella-dependent fashion. Expression of both colonization factors and flagella was inhibited at the level of transcription. In addition, the hldE mutant displayed altered growth, increased biofilm formation and clumping in minimal growth medium. Investigation of an orthogonal LPS-deficient mutant combined with mass spectrometric analysis of protein glycosylation indicated that HldE exerts its role on ETEC virulence both through protein glycosylation and correct LPS configuration. These results place HldE as an attractive target for the development of future antimicrobial therapeutics.
Highlights
Enterotoxigenic Escherichia coli (ETEC) strains contribute significantly to diarrheal illness and mortality in third world countries (Liu et al, 2012; Platts-Mills et al, 2015)
To investigate if HldE is needed for flagella production in ETEC we examined the motility ability of wild type cells, the hldE mutant and hldE/pGH106 as well as the non-motile fliC strain using a semi-solid surface in a swimming and swarming assay
As HldE has been reported to play a role in both LPS biosynthesis and protein glycosylation, we wanted to investigate the potential link between HldE and virulence in ETEC
Summary
Enterotoxigenic Escherichia coli (ETEC) strains contribute significantly to diarrheal illness and mortality in third world countries (Liu et al, 2012; Platts-Mills et al, 2015). ETEC causes disease by adhering to epithelial cells of the upper small intestine where the delivery of heat-labile and/or. LPS and ETEC Virulence heat stable toxins to host cell receptors initiates a signaling cascade, which results in watery diarrhea (Fleckenstein et al, 2013). In ETEC H10407, the host cell interaction is promoted by a combination of the plasmid-encoded fimbrial colonization factor, CFA/I, a collaboration between the adhesin EtpA and the flagellum as well as expression of virulence genes located on chromosomal pathogenicity islands (Evans et al, 1975; Patel et al, 2004; Fleckenstein et al, 2006; Roy et al, 2009b; Crossman et al, 2010). Protein glycosylation plays an important role in mediating adhesion, colonization and invasion of host tissue and may serve either as mediators of direct interactions with host-encoded cell surface glycans, as stabilizing factors of protein structure, or as a means to create surface heterogeneity and thereby evade recognition by the host immune system (Gault et al, 2015; Tytgat et al, 2016)
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