Investigations into Bacillus subtilis spore resistance

In this thesis, we study various properties resistance of spores of Bacillus subtilis. First, we examine the role of heat shock proteins in spore wet heat resistance. Analysis of the effects of specific mutations in heat shock genes and the expression of fusions to the coding gene for thermostable β-galactosidase to a number of heat shock genes has shown that heat shock proteins play no significant role in the wet heat resistance of spores. Next, we study thermal adaptation by examining the resistance and other properties of spores prepared at a range of temperatures and find that the temperature of sporulation effects a variety of spore properties including resistance to many different stress factors and also results in detectable alterations in spore coat and cortex composition. This is followed by a study of spore killing by hydrogen peroxide using a fairly low concentration of this agent to kill ∼90% of spores. We find that hydrogen peroxide-killed spores can initiate germination but cannot swell properly or begin metabolism during spore germination. Finally, experiments using spores carrying the luxAB genes from Vibrio harvevii are described and the importance of this system for studying spore germination and mechanisms of spore killing by various agents are discussed. All of these studies are aimed at gaining a better understanding of spore resistance properties and spore killing mechanisms, as well as developing new methods to study dormant spores and spore germination.