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Title: Regulation and secretion of virulence factors in Erwinia carotovora subspecies atroseptica
Author: Corbett, M. J.
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2006
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The twin arginine translocation (Tat) system facilitates the export of a number of proteins from the cytoplasm to the periplasm in many species, acting as an alternative to the Sec export system for several proteins. The tatC gene of Eca was disrupted by insertion of a chloramphenicol resistance gene. This mutant secreted all known virulence factors, but showed phenotypes consistent with membrane instability. The mutant showed no reduction in virulence as measured by a tuber virulence assay although a large reduction was seen in stem rot assays, indicating that Tat is essential for full Eca pathogenesis. The act of secreted proteins (the secretome) of Eca was characterised using 2D-PAGE. Proteins were identified by mass spectrometry and mapped to their cognate genes using the Eca genome sequence information. Novel putative virulence factors, as well as known exoenzymes were identified. Quorum sensing (ExpI-), and Type II secretory (Out-) transposon mutants were identified, and the secreted protein profiles investigated using difference in gel electrophoresis (DIGE). A subset of the secretome was shown to be Out-dependent, and a larger proportion of the secretome was shown to be ExpI-dependent. VirR was shown to be the repressor mediating quorum sensing-dependent regulation of virulence factors. One protein identified in the secretome is a homologue of an avirulence protein of Xanthomonas campestris. This Eca gene, svx, encodes a protein with few homologues. The secretion of Svx was Out-dependent, and transcription of svx was ExpI-dependent, mediated by VirR. An svx transposon insertion mutant was selected, and the mutant exhibited reduced virulence in potato stem assays and potato tuber assays. The tuber virulence phenotype was complemented using a plasmid carrying the svx gene in trans.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available