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Title: Horizontal gene transfer in Bacteroides fragilis
Author: Jobling, Kelly Louise
ISNI:       0000 0004 5353 1286
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2014
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Horizontal gene transfer (HGT) is one of the man driving forces of evolution in prokaryotes, and can also promote within-strain variation of bacterial species. The genomes of three previously sequenced Bacteroides fragilis strains, NCTC9343, 638R and YCH46 displayed evidence of extensive HGT, demonstrated by the presence of 28 divergent capsular polysaccharide-associated biosynthesis loci. The genomes of a further four B. fragilis strains, LS66, GNAB92, RD48 and BE1 were sequenced and analysed. Genomic comparisons of BE1 and GNAB92 with NCTC9343, 638R and YCH46 identified ten new divergent polysaccharide biosynthesis loci. There is consequently, the potential to express 38 different polysaccharides amongst these five strains. Such a high level of variation in capsular polysaccharides, in so few strains has not been previously observed. HGT has occurred in B. fragilis despite the presence of diverse Restriction-Modification systems. The genome sequences of NCTC9343 and 638R contained a gene, ubb, the product of which, BfUbb, has 63% identity to human ubiquitin. The closest DNA sequence homology is to a migratory grasshopper entomopox virus, suggesting acquisition of this gene was via inter-kingdom HGT. The ubb gene was also identified in the newly sequenced genomes of B. fragilis strains LS66 and RD48. BfUbb had a predicted signal sequence; both full-length and processed forms were detected in whole-cell extracts by Western blot analysis. The inability to detect BfUbb in periplasmic extracts isolated from a B. fragilis strain containing an ubb signal sequence deletion construct, supported the periplasmic location of the processed form of the protein and the requirement for the signal peptide for transport from the cytoplasm. BfUbb was also detected in concentrated supernatants containing outer membrane vesicles, suggesting a mechanism by which the protein may be delivered to the host. This is the first example of ubiquitin being produced by a prokaryote. Transduction by bacteriophages is one mechanism by which horizontal gene transfer can occur and can also be a useful tool for genetic manipulation. Fifteen potentially new B. fragilis-specific bacteriophages were isolated from filtered sewage and characterised by phage titres and restriction endonuclease cleavage profiles. Of the fifteen, seven phages appeared to be different to the previously identified phage ФED01. None of the bacteriophages were capable of transduction. B. fragilis is a predominant member of the gastrointestinal microbiota. To survive within this specific niche, bacteria must successfully compete with other organisms for nutrients and space, and withstand attacks from bacteriophages. HGT may aid in the survival of B. fragilis as a commensal.
Supervisor: Blakely, Garry; Gallagher, Maurice Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Bacteroides fragilis ; ubiquitin ; HGT ; commensal bacteria