Campylobacter infections are the most common bacterial cause of human diarrhoeal illness. When stressed, C. jejuni can enter a viable but not culturable (VBNC) state, which may aid survival in harsh environments. Although resuscitation has been reported in live animal models and eggs, other investigators have been unable to replicate these experiments. The importance of the VBNC state is therefore still under debate. Development of a reliable protocol for inducing resuscitation would greatly aid further investigations. The Rpfs produced by Actinobacteria spp are molecular signals capable of promoting resuscitation. The hypothesis under investigation in this thesis was that the resuscitation of VBNC C. jejuni is controlled by a Rpf orthologue. Bioinformatics searches demonstrated that the predicted C. jejuni NCTC 11168 protein, Cj0645 shares: domain content and arrangement, sequence similarity, and genomic synteny, with M. luteus Rpf. This strongly suggests a shared ancestry. A two-pronged approach was taken to investigate whether Cj0645 was capable of resuscitating VBNC C. jejuni. Firstly, attempts were made to produce a recombinant protein for functional analysis. However, none of these attempts were effective in producing soluble protein. The second approach involved the construction and phenotypic testing of an isogenic mutant and a genetically complemented strain. Phenotypic testing demonstrated that mutation of cj0645 affects growth rate, and the process of cell division, causing the cj0645 mutant to form filaments. However, experiments testing the ability of conditioned media to promote growth from the VBNe state and low cell density found no evidence of a Rpf-like activity in exponential phase C. jejuni culture supernatants. Since the cj0645 is known to be expressed during this growth phase, it appears that, unlike M. luteus Rpf, Cj0645 does not promote growth and as such is not a orthologue.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:625273 |
| Date | January 2010 |
| Creators | Morgan, T. H. |
| Publisher | University College London (University of London) |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://discovery.ucl.ac.uk/642745/ |
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