High-throughput 3D tracking of Vibrio cholerae in mucus and mucus-mimicking media to assess early-stage infection’s dynamics.

• Applicant: Professorin Dr. Marianne Grognot
• Subject Area: Medical Microbiology and Mycology, Hygiene, Molecular Infection Biology ; Microbial Ecology and Applied Microbiology
• Term: since 2025
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 570781731

A wide variety of bacteria can swim by rotating one or several flagella, including an estimated half of bacterial pathogens. Swimming motility and chemotaxis enable a motile bacterium to navigate its environment, and can modulate its performances (spreading, finding food sources, accessing specific niches…). There are very strong incentives to think bacterial motility and chemotaxis are important for pathogenicity, and it is often hypothesized to be for allowing bacteria to cross the mucus barrier protecting our mucosa. However, to date there has been no direct quantification of this likely mechanism of infection, probably due to a combination of (i) a general overlooking of mucus and/or bacterial motility phenotypes and (ii) the severe limitations of 2D bacterial fluorescence tracking - especially in complex environments such as mucus. In this project, we propose to develop a 'mucus-crossing assay' in which we track bacterial pathogens in 3D as they navigate the mucus over a layer of cells. Fluorescence-based 3D tracking already exists for bacteria swimming in buffer: we aim to extend it to (highly optically complex) mucus using more recent image analysis methods. We will synergistically use a high-throughput chemotaxis assay, to test our hypotheses in simpler mucus mimicking media and provide further navigation phenotyping. With these two assays, we intend to focus on Vibrio cholerae, the causative agent of cholera, which swims using a single polar flagellum. Motility has long been recognized as an important virulence factor for V. cholerae, but only through indirect approaches that have led to several open hypotheses and/or contradictive results. The role of chemotaxis to direct movement towards infection sites is even more debated. How does V. cholerae swim in mucus? What are the most successful swimming behaviors to breach it? How do swimming behaviors couple with mucus properties and expression of other pathogenic factors (e.g. mucinases)? Does chemotaxis to bile acids drive and/or perturbate V. cholerae performances to cross the mucus barrier? Our work will provide to these debated topics a completely new approach, improve the interpretation of contradictive past macroscale experimental results, assess multiples untested hypothesis in the literature, and propose quantitative arguments to nuance existing treatment strategy suggestions. Overall, this project has great potential to deepen our understanding of host-pathogen interactions by proposing a unique and direct approach to study a likely widespread infection mechanism.

• DFG Programme: Research Grants