Structural and functional insights to Bacillus spore germination and toxin engineering for sustainable agriculture

Bacillus spores exhibit extraordinary resilience, surviving extreme environmental conditions through dormancy and reactivating under favourable conditions via germination. Germination is triggered by specific environmental signals detected by germinant receptors, initiating a cascade of events that culminate in the return to vegetative growth. This thesis focuses on the molecular mechanisms of germination in Bacillus cereus; an important human pathogen associated with foodborne illnesses and opportunistic infections. Specifically, the study investigates the roles of germinant receptors (GRs), the SpoVA CaDPA channel, and the SleB cortex-lytic enzyme, which are all major components of the germination apparatus. The presented data indicate that the GerI GR can function independently of the GerQ GR in triggering a germination response to inosine in B. cereus spores, whereas GerQ appears to have an essential requirement for GerI to cooperatively enable more efficient spore germinative responses. Amino acid residues critical to the recognition of the inosine, alanine and sodium (co)germinants and to receptor stability are identified in the GerI receptor. The work also reveals that deletion of either of B. cereus’ two spoVA operons has minimal impact on the ability to form heat resistant spores, whereas deletion of both operons essentially ablates sporulation. A more detailed study on the B. cereus SpoVAF protein reveals residues critical to function and incorporation into spore germinosomes but little evidence that the putative SpoVAF channel contributes significantly to germination responses at sub optimal germinant concentrations. The thesis additionally presents a molecular model of the SleB cortex lytic enzyme complexed with the YpeB and YlaJ proteins. Site directed mutagenesis guided by EV Couplings analysis lends weight to the model potentially revealing insight to how this critical enzyme is held in an inactive state during spore dormancy. Finally, the thesis additionally explores novel hybrid Cry proteins with potential bioinsecticide applications. Ultimately, the thesis contributes to a deeper understanding of spore biology presenting findings with potentially broad implications in agricultural sector.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2025

Author dc:creator

• Ow Young Villarreal, Giannina Del Carmen

Advisor dc:contributor.advisor

• Christie, Graham

• Bacillus cereus
• spore germination
• germinant receptors
• SpoVA channel
• SleB cortex-lytic enzyme
• germinosome
• site-directed mutagenesis
• Cry hybrid proteins
• molecular mechanisms
• germinant receptors (GerI, GerQ)
• bioinsecticide potential

Licence

• https://www.repository.cam.ac.uk/bitstreams/5c884df5-e0b4-4d19-a037-c7bc33fef260/download
• http://purl.org/NET/rdflicense/allrightsreserved

Language dc:language

eng

DOI dc:identifier.doi

https://doi.org/10.17863/CAM.122425

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/391216