Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Thesis (Ph.D.)--Aberdeen University, 2008. / Title from web page (viewed on Apr. 21, 2009). Includes bibliographical references.

This thesis is missing page 129. Other copies of this thesis do not have the page either. -Digitization Centre / E. coli expresses an alternative sigma factor, RpoS, in response to starvation and environmental stresses. RpoS is a global regulator and it controls numerous genes, which aids in counteracting these stresses. The RpoS regulon is large but is not completely characterized. We have previously identified over one hundred RpoS-dependent fusions in a genetic screen based on the differential expression of an operon-lacZ fusion bank in rpoS mutant and wild type backgrounds. Forty-eight independent gene fusions were identified including several in well-characterized RpoS-regulated genes such as osmY, katE and otsA. Many of the fusions mapped to genes of unknown function or to genes that were not previously known to be under RpoS control. Based on the homology to other known bacterial genes, some of the RpoS regulated genes with unknown functions may be important for nutrient scavenging. To gain a better insight into the functions of these poorly characterized genes, we tested the ability of the fusion mutants to utilize various carbon sources and to utilize individual amino acids as carbon and nitrogen sources. The results indicate that most of the strains in rpos-backgrounds exhibited better growth in succinate and fumarate and in several amino acids than did the corresponding strains in wild-type backgrounds. / Thesis / Master of Science (MS)

This thesis is missing a page between pages 60 and 70, and three pages between pages 70 and 81. Since the pages are not all numbered, the specific page numbers cannot be determined. Theses missing pages are not in the other copies of the thesis. -Digitization Centre / Replicate isogenic populations of E. coli were propagated and maintained for over 4000 generations in order to investigate the adaptation of E. coli to increased levels of the mutagen methanesulfonic acid ethyl ester (EMS). Control "C" cell lines were propagated through daily serial culture in the absense of any mutagenic treatment. EMS adapted cell lines "E" / "e" were propagated through daily serial culturing and treated daily with 25Jul of EMS following serial dilution. Mutation frequency and survival assays conducted in this investigation strongly suggest that prior long-term low dose exposure to EMS results in significantly higher levels of resistance to the lethal and mutagenic effects of larger challenge doses of EMS relative to long-term evolved control cell lines "C". In addition, both survival and inhibition disk assays suggest a cross adaptive response between EMS and MNNG, showing enhanced survival and reduced growth inhibition zones in cells adapted to EMS and challenged with MNNG. Preliminary competition experiments suggest relative fitness for the EMS adapted cell lines ( "E" / "e") compared to the "C" control cell lines in both the presence of EMS. Unexpectedly the fitness estimates also suggest a higher relative fitness for the "E" / "e" EMS adapted cell lines in the absence of EMS treatment, suggesting that the EMS specific adaptation may also result in improved fitness in novel environments. Despite the adaptive advantage for the "E" / "e" cell lines suggested by the fitness estimates, the results from the competition experiments are insignificant due to the high degree of variability among replicate fitness estimates. Attempts to induce the adaptive response repair pathway were not successful in either the control "C" or the EMS adapted "E" / "e" cell lines suggesting that enhanced resistance seen in the adapted "E" / "e" cell lines could likely be a result of enhanced activity of the constitutive transferase Ogt and the constitutive glycosylase Tag. The ada and the ogt genes encode the induced and the constitutively-active DNA methyl transeferases in E. coli. As such they appeared to be the most likely candidates for genetic changes responsible for the enhanced resistance to the lethal and mutagenic effects of large doses of alkylating agents in the long-term EMS adapted "E" / "e" cell line. However, the DNA sequences analyzed for the ogt and the ada genes for both the long-term evolved control E. coli cell line "C" and the long-term-evolved EMS adapted "E" / "e" cell line indicate no sequences differences between these two cell lines. Previous studies have primarily observed E. coli's ability to phenotypically acclimate over very short time intervals to EMS. This analysis has shown that long-term genetic adaptation to low doses of EMS results in enhanced resistance to both the lethal and mutagenic effects of larger challenge doses of EMS. / Thesis / Master of Science (MS)

Poultry litter can harbor pathogenic bacteria, including avian pathogenic Escherichia coli (APEC). Applying litter amendments is one strategy to improve bird health and potentially reduce pathogens. Biochar and PLT were applied as litter amendments in a live bird trial to study their effects on E. coli populations, APEC virulence genes (VAG), and antimicrobial resistance (AMR) genes. Samples were collected at days 0, 17, 29, and 41 to enumerate E. coli and store bacterial isolates for antimicrobial-resistance gene analysis. Data analysis showed litter amendments did not significantly affect overall E. coli populations. Grow-out time impacted E. coli populations, with reductions occurring over time. Litter treatment had no impact on the prevalence of VAG or AMR. Time showed VAGs were absent at d 0 while AMR genes were prevalent at d 0. This indicates chicks may have been the source of VAG, while AMR genes were prevalent in used litter.

Thesis (Ph.D.)--Baylor University, 2008. / Includes bibliographical references (p. 130-146).

Thesis--Göteborg University, 1997.

In contrast to the damage caused by far-UV, the damaging effects of UVA (320-400 nm) in living cells are not well understood. The damage caused by UVA irradiation is largely oxygen-dependent, suggesting UVA-mediated DNA damage involves reactive oxygen species produced through the action of an endogenous photosensitizer. Previous studies examining cellular responses to UVA irradiation in E. coli have been hindered by the fact that, at sublethal fluences, wild-type cells undergo a transient inhibition of cell growth termed a "growth delay". This effect is absent in nuvA⁻ strains, thereby facilitating the study of DNA repair factors required for the repair of UVA-mediated damage. Formamidopyrimidine (Fapy) glycosylase (encoded by fpg) and the UvrABC excinuclease are both capable of excising oxidatively damaged DNA bases. An fpg::kan mutation was placed into isogenic uvrA⁺ and uvrA⁻ strains of E. coli to evaluate the relative importance of these repair enzymes in the recovery from UVA-induced stress. In a nuvA⁻ background, the survival of fpg⁻ mutants exposed to UVA was significantly reduced relative to isogenic fpg⁺ control strains. This effect was enhanced in the absence of the UvrABC excinuclease, suggesting a role for both of these enzymes in repairing UVA-generated lesions. Survival of isogenic nuvA⁺ repair-deficient strains was significantly lower than nuvA⁻ strains, suggesting a role for the modified base 4-thiouridine in UVA-mediated lethality. An in vitro plasmid DNA irradiation assay in the presence and absence of 4-thiouridine was used to examine this possibility. When irradiated DNA was subsequently used to transform the fpg⁻ and uvrA⁻ mutant strains, no increase in DNA damage (as measured by a decrease in transformational efficiency) in the presence of 4-thiouridine was observed, suggesting that when present in solution this base does not play a photosensitizing role in UVA-mediated lethality. / Thesis / Master of Science (MSc)

Verocytotoxin-producing Escherichia coli (VTEC) are important human pathogens that may cause diarrhoea, haemorrhagic colitis and haemolytic uremic syndrome (HUS). Worldwide, non-sorbitol-fermenting (NSF) VTEC O157:H7 is the most common serogroup associated with HUS but several non-O157:H7 serogroups have emerged as causes of this disease. This research investigated the pathogenic potential of two non-O157:H7 serogroups: O26 and sorbitol-fermenting (SF) O157:NM. While VTEC O26 have emerged as a significant cause of HUS in continental Europe, human infections associated with this pathogen are uncommon in Scotland and generally only result in simple diarrhoea. The study characterised E. coli O26 isolates recovered from human infections in Europe and Scotland and isolates collected from Scottish cattle with the objectives to identify factors which may allow strains to cause more serious clinical disease and to investigate the potential of bovine VTEC O26 in Scotland to cause human infection. MLST analysis of housekeeping genes found little genetic variation in the genomic ‘backbone’ among the vast majority of E. coli O26 isolates. The gene for verocytotoxin 2 (vtx2) alone was carried by VTEC O26 isolates recovered from patients in continental Europe but was found in no Scottish human isolate, where the majority of isolates did not harbour a vtx gene. It was demonstrated that among the European VTEC O26 human isolates, 67% carried a specific allele within the promoter region for LEE1 and 87% harboured the tccP2 gene. In contrast, no Scottish VTEC O26 human isolate carried this allele or the tccP2 gene. The impact these genotypic characteristics have on the pathogenic potential of a strain remains uncertain. There were no clear differences in verocytotoxin titres, levels of LEEencoded protein secretion or levels of adherence to Caco-2 cells between VTEC O26 isolates recovered from human infections of varying severity. However, levels of LEE-encoded protein secretion from cattle isolates were generally higher than those from many of the human isolates. The differences in pathogenic potential between isolates are likely to be due to horizontally acquired DNA, including vtx2 carriage and the O-island-phage-associated effector protein repertoire. Further work is required to determine if the differences identified may also impact on shedding levels from cattle and therefore the likelihood of transmission to humans. Since 1988, SF VTEC O157:NM strains have emerged and have been associated with a higher incidence of progression to HUS than NSF VTEC O157:H7. This study investigated bacterial factors that may account for the increased pathogenic potential of SF VTEC O157:NM. While no evidence of toxin or toxin expression differences between the two VTEC O157 groups was found, the SF VTEC O157:NM strains adhered at significantly higher levels to a human colonic cell line. Under the conditions tested, curli were shown to be the main factor responsible for the increased adherence to Caco-2 cells. The capacity of SF VTEC O157:NM strains to express curli at 37C may have relevance to the epidemiology of human infections as curliated strains could promote higher levels of colonization and inflammation in the human intestine. In turn this could lead to increased toxin exposure and an increased likelihood of progression to HUS.