Krachler, A. | bioRxiv Science

bioRxiv2017

High-resolution analysis of spatiotemporal virulence gene regulation during food-borne infection with Escherichia coli O157:H7 within a live host

Food-borne infection with enterohemorrhagic Escherichia coli (EHEC) is a major cause of diarrheal illness in humans, and can lead to severe complications such as hemolytic uremic syndrome. Cattle and other ruminants are the main reservoir of EHEC, which enters the food-chain through contaminated meat, dairy, or vegetables. However, how EHEC transitions from the transmission vector to colonizing the intestinal tract, and how virulence-specific genes are regulated during this transition, is not well understood. Here, we describe the establishment of a vertebrate model for food-borne EHEC infection, using the protozoan Paramecium caudatum as a vector and the zebrafish (Danio rerio) as a host. At 4 days post fertilization, zebrafish have a fully developed intestinal tract, yet are fully transparent. This allows us to follow intestinal colonization, microbe-host cell interactions, and microbial gene induction within the live host and in real time throughout the infection. Additionally, this model can be adapted to compare food- and water-borne infections, under gnotobiotic conditions or against the backdrop of an endogenous (and variable) host microbiota. Finally, the zebrafish allows for investigation of factors affecting shedding and transmission of bacteria to naive hosts. High-resolution analysis of EHEC gene expression within the zebrafish host emphasizes the need for tight transcriptional regulation of virulence factors for within-host fitness.\n\nIMPORTANCEEnterohemorrhagic Escherichia coli (EHEC) is a food-borne pathogen which can cause diarrhea, vomiting and in some cases, severe complications such as kidney problems in humans. Up to 30% of cattle are colonized with EHEC, which can enter the food-chain through contaminated meat, dairy and vegetables. In order to control infections and stop transmission, it is important to understand what factors allow EHEC to colonize its hosts, cause virulence and aid transmission. Since this cannot be systematically studied in humans, it is important to develop animal models of infection and transmission. We developed a model which allows us to study food-borne infection in zebrafish, a vertebrate host that is transparent and genetically tractable. Using the zebrafish host, we can follow the bacterial infection cycle in real time, and gain important information regarding bacterial physiology and microbe-host interactions. This will allow us to identify potential new targets for infection control and prevention.

microbiology↗

bioRxiv2017

A commensal adhesin enhances E. coli retention by mucin, while mucin desulfation by mucin-foraging bacteria enhances its transmigration through the mucus barrier

Bacterial adhesion to host receptors is an early and essential step in bacterial colonization, and the nature of adhesion-receptor interactions determines bacterial localization and thus the outcome of these interactions. Here, we determine the host receptors for the Multivalent Adhesion Molecule from the gut commensal E. coli HS (MAMHS), which contains an array of seven mammalian cell entry (MCE) domains. The MAMHS adhesin interacts with a range of host receptors, through recognition of a shared 3-O-sulfo-galactosyl moiety. This functional group is also found in mucin, a component of the intestinal mucus layer and thus one of the prime adherence targets for commensal E. coli. Mucin gels impede the motility of E. coli by acting as a physical barrier, and the barrier effect is enhanced by specific interactions between mucin and MAMHS in a sulfation-dependent manner. Desulfation of mucin by pure sulfatase or the sulfatase-producing commensal Bacteroides thetaiotaomicron decreases binding of E. coli to mucin and increases attachment of bacteria to the epithelial surface, through interactions with surface-localized sulfated lipid and protein receptors. Together, our results demonstrate that the E. coli adhesin MAMHS facilitates retention of a gut commensal by mucin, and suggest that the amount of sulfatase secreted by mucin-foraging bacteria inhabiting the same niche, such as B. thetaiotaomicron, may affect the capacity of the mucus barrier to retain commensal E. coli.

biochemistry↗