Structural transitions and cellular remodelling in spore germination

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Bacteria such as those causing botulism and anthrax survive harsh conditions and spread disease as spores. However for these spores to cause disease symptoms they must germinate into so-called vegetative cells. Toxins produced by the vegetative cells are most often the main factor in making infected animal or human hosts sick. We wish to understand how this germination takes place and how the active vegetative cell emerges from the dormant spore. This is a remarkable metamorphosis where one intricate type of cell structure is completely transformed into a radically different structure- it is analogous to the metamorphosis of a seed into a seedling and arguably as complex! The process is also interesting because vegetative cells are much more vulnerable to attack, for example by antibiotics or disinfectants, than spores are. Thus if we could work out how to 'germinate to exterminate', we could develop new weapons against a number of diseases and food-spoilage organisms. We will study germination in two selected organisms- Clostridium sporogenes (C. sporogenes) and Clostridioides difficile (C. difficile). C. sporogenes is non-pathogenic but very closely related to C. botulinum which causes botulism, a potentially lethal paralysis. Spores are extremely heat resistant so eradication from food requires costly heat treatment that lowers nutritional value & makes the product less appealing, yet there is increasing demand for minimally processed, safe foods. C. botulinum is also listed as a potential bio-weapon (UK Anti-terrorism, Crime & Security Act 2001). The advantage of using the harmless C. sporogenes is that we can learn a lot about C. botulinum without the need for highly specialised containment facilities. C. difficile is the leading cause of antibiotic-associated diarrhoea, especially in hospital patients and the elderly. Disease occurs when ingested spores germinate in response to bile acids in the gut. The toxins are the major cause of disease & in many cases, death. Ungerminated spores remaining in the gut can be the source of recurrent infections even after treatment.Our multidisciplinary project will apply state of the art techniques of structural and molecular biology to investigate the way these spores germinate. A crucial aspect of this work will involve mapping the proteins that make up spores in 3D molecular detail. To achieve this, we will exploit exciting new developments in microscopy - such as cryo-electron tomography - that allow us to visualise spores and cells in unprecedented detail. We will create 3D images of the spore where we can even identify and locate individual molecules. We will then go on to visualise the structures of spores as they transform into active cells, as might happen when contaminated food is ingested. The project will exploit new developments in imaging methods that allow us to see the structures more clearly. It will involve combinations of molecular genetics, use and development of electron light and scanning probe imaging approaches and computational image processing. Ultimately we aim to make a 3D movie that explains the mechanical and biochemical changes that take place when the active vegetative cell emerges out of the spore.