From patience to peril: how do C. difficile spores decide when it's safe to germinate?

About the Project

Background:

Clostridioides difficile is the leading cause of healthcare-associated infections and is notoriously difficult to treat. This bacterial species infects the colon following antibiotic treatment, which disrupts the protective gut microbiota. Its ability to resist treatment and spread between hosts relies on its ability to form spores – dormant cells able to survive spectacularly harsh conditions, including antibiotics, disinfectants, and stomach acid.

But how do C. difficile spores, which can remain viable for years, determine when it is safe to germinate?

Objectives:

Germination is a high-stakes decision for C. difficile: too early risks destruction in a hostile environment, too late risks missing the chance to colonise a new host. We hypothesise variation in the timing of germination is a form of “bet-hedging” — an evolutionary strategy where behavioural variability increases the chance that some individuals succeed.

Multiple chemical signals are required for C. difficile germination. In nutrient-rich media, genetically identical spores germinate at different times, but in presence of bile salts—exclusively found in the gut—spores respond more uniformly. Moreover, current evidence suggests compounds released by germinating spores can promote the germination of neighbouring spores, potentially allowing local synchrony. This project will determine how spores integrate different environmental cues to decide when to commit to germination.

Novelty:

Previous work on spore germination relies on bulk, population-level assays. Here we will instead use live microscopy and automated image analysis to individually study the behaviour of thousands of cells. This single cell approach will allow us to resolve key checkpoints in the germination pathway and the molecular processes that give rise to heterogeneity.

Timeliness:

Understanding how germination is regulated is central to developing new strategies to prevent and treat C. difficile infections. In addition, recent advances in massively parallel cell tracking now make it possible to address long-standing questions about how bacteria have adapted to survive in unpredictable environments.

Experimental Approach:

Using novel tools we developed for imaging live anaerobes, we will:

• Quantify germination dynamics in response to defined combinations of germinants (e.g. bile salts, amino acids, calcium ions).
• Determine how candidate and known germination pathway genes (e.g. cspA, cspB, sleC) influence variability.
• Parameterise simple mathematical models to understand how multiple signalling pathways interact to control germination.

This project will provide training in bacterial physiology, infection biology, quantitative microscopy and computational data analysis — a diverse skill set that will equip the student to excel in the increasing interdisciplinary research environment.

Please get in touch with William Durham (w.m.durham@sheffield.ac.uk) if you would like to have an informal chat about this project.

Funding Notes

We welcome inquiries from:

• applicants that have already secured PhD funding
• self-funded applicants