Thesis (Ph. D.)--University of Washington, 1996. / Vita. Includes bibliographical references (leaves [139]-158).

CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / Conselho Nacional de Desenvolvimento CientÃfico e TecnolÃgico / Porphyromonas gingivalis e Aggregatibacter actinomycetemcomitans sÃo periodontopatÃgenos associados à periodontite agressiva. A fÃmbria, uma estrutura relacionada à adesÃo e à invasÃo de cÃlulas, à um dos principais fatores de virulÃncia de P. gingivalis. Baseado na sequÃncia de nucleotÃdeos, seis genÃtipos(fimA) que codificam a fÃmbria principal dessas bactÃrias foram identificados, sendo o fimA II mais comumente relacionado à destruiÃÃo periodontal. O objetivo deste trabalho foi avaliar, por meio de reaÃÃo em cadeia da polimerase em amostras de placa subgengival dos sÃtios com maior profundidade de sondagem de pacientes com periodontite agressiva, a prevalÃncia de P. gingivalis, do genÃtipo fimA II de P. gingivalis e de A. actinomycetemcomitans, assim como relacionar a presenÃa desses patÃgenos ou genÃtipo à idade e aos parÃmetros clÃnicos periodontais (Ãndice de placa, Ãndice de sangramento gengival, profundidade de sondagem e nÃvel de inserÃÃo) encontrados nesses pacientes. Foram selecionados 45 pacientes com periodontite agressiva generalizada, com idade entre 15 e 40 anos. Nessa populaÃÃo, 64,4% apresentaram P. gingivalis e 28,8% apresentaram A. actinomycetemcomitans em sua microbiota subgengival. Dos pacientes positivos para P. gingivalis, 82,6% apresentaram o genÃtipo fimA II. Ao se relacionar a presenÃa ou ausÃncia das bactÃrias ou genÃtipo aos dados clÃnicos e idade, foi observada diferenÃa estatisticamente significante entre o nÃvel clÃnico de inserÃÃo do sÃtio coletado de pacientes com presenÃa de P. gingivalis e seu genÃtipo fimA II quando comparados aos pacientes negativos para essa bactÃria e genÃtipo, sendo a perda de inserÃÃo significativamente maior em pacientes que apresentaram P. gingivalis e em paciente com seu genÃtipo fimA II. AlÃm disso, foi encontrada mÃdia de idade significativamente mais elevada em pacientes positivos para P. gingivalis que em pacientes negativos para essa bactÃria. Concluiu-se, assim, que P. gingivalis e seu genÃtipo fimA tipo II estÃo presentes em alta prevalÃncia em pacientes com periodontite agressiva, que A. actinomycetemcomitans està presente em menor proporÃÃo de indivÃduos na populaÃÃo estudada e que P. gingivalis parece ser mais comumente encontrada em bolsas mais profundas e em indivÃduos mais velhos. / Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans are periodontal pathogens associated with aggressive periodontitis. The fimbriae, a structure related to adhesion and invasion of cells, is one of the major virulence factors of P. gingivalis. Based on the nucleotide sequence, six genotypes(fimA) encoding the major fimbriae of these bacteria were identified, and the fimA II is the most commonly associated with periodontal destruction. The objective of this study was to evaluate, by polymerase chain reaction in subgingival plaque samples from sites with highest probing depth in patients with aggressive periodontitis, the prevalence of P. gingivalis, P. gingivalis genotype fimA II and A. actinomycetemcomitans, and relate the presence of these pathogens or genotype to age and clinical periodontal parameters (plaque index, gingival bleeding index, probing depth and clinical attachment level) in these patients. We selected 45 patients with generalized aggressive periodontitis, aged from 15 to 40 years. 64.4% of these patients harbored P. gingivalis and 28.8% harbored A. actinomycetemcomitans in their subgingival microbiota. In patients positive for P. gingivalis, 82.6 % presented the genotype fimA II. In relation to the presence or absence of bacteria or gene to clinical data and age, a statistically significant difference between clinical attachment level was observed in the selected sites of patients with the presence of P. gingivalis and its genotype fimA II when compared to patients negative for these bacteria and genotype, with periodontal loss significantly higher in patients harboring P. gingivalis and in patients harboring genotype fimA II. In addition, the average age in patients positives for P. gingivalis was significantly higher than in negative ones. It is therefore concluded that P. gingivalis and its genotype fimA II are present in high prevalence in patients with aggressive periodontitis, A. actinomycetemcomitans is present in a smaller proportion of individuals in the studied population and P. gingivalis seems to be more commonly found in deeper sites and older individuals.

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Um estudo prévio descreveu uma cepa de Escherichia coli enteropatogênica (EPEC) do sorotipo O142:H34 (Ec48/66) que apresentava o padrão de adesão agregativa (AA), característico de E. coli enteroagregativa (EAEC), e os genes eae e aatA. O objetivo deste estudo foi avaliar o papel de fímbrias no padrão AA dessa cepa. A mesma aderiu no padrão AA nas linhagens HeLa, HEp-2, HT-29 e T84, além de causar a lesão A/E em células HeLa e HT-29. Foi detectada a presença dos genes que compõem o operon da fímbria AAF/I de EAEC e da fímbria E. coli common pilus (ECP). Ambos os operons foram sequenciados mostrando alta identidade com os operons presentes em EAEC (agg) e EPEC (ecp). Mutantes nos genes aggC e ecpC mantiveram o padrão AA e a capacidade de causar a lesão A/E em células HeLa. O mutante em aggC apresentou diminuição significativa da adesão. Análises filogenéticas posicionaram essa cepa em um grupo constituído por EPEC. A cepa Ec48/66 é uma EPEC atípica que expressa o padrão AA, o qual é parcialmente dependente de uma variante de AAF/I, mas não de ECP. / A previous study described a strain of enteropathogenic Escherichia coli (EPEC), serotype O142:H34 (Ec48/66), presenting the aggregative adherence pattern (AA), typical of enteroaggregative E. coli (EAEC), and the eae and aatA genes. The aim of this study was to evaluate the role of fimbriae in the AA pattern of Ec48/66. This strain presented the AA pattern in HeLa, HEp-2, HT-29 and T84 cells, and the ability to cause the A/E lesion in HeLa and HT-29 cells. The presence of the genes comprising the operons of AAF/I fimbriae and E. coli common pilus (ECP) was detected. Both operons were sequenced showing high identity with the operons present in EAEC (agg) and EPEC (ecp). Mutants in the aggC and ecp genes maintained the AA pattern and the ability to cause the A/E lesion in HeLa cells. The aggC mutation significantly reduced the adherence. Phylogenetic analyzes located the strain in the group consisting of EPEC. The Ec48/66 strain is an atypical EPEC expressing the AA pattern, which is partially dependent on a variant of AAF/I, but not on ECP.

Mémoire numérisé par la Division de la gestion de documents et des archives de l'Université de Montréal

Thesis (Ph. D.)--University of Virginia, 2008. / Title from title page. Includes bibliographical references. Also available online through Digital Dissertations.

L’interaction avec les cellules hôtes est une étape primordiale du cycle infectieux des Salmonella. Elle implique entre autres des fimbriae permettant l’attachement des Salmonella aux cellules et des invasines permettant leur internalisation. Généralement, l’expression de ces facteurs est réprimée et ils ne sont exprimés qu’in vivo en des sites bien spécifiques. Notre objectif a été d’étudier les mécanismes de répression de l’expression des fimbriae Pef et de l’invasine Rck. Nos résultats montrent que les nucléoprotéines H-NS, Hha et YdgT régulent négativement l’expression de ces deux facteurs de virulence. Le mécanisme de répression de la transcription de l’opéron pef a été caractérisé et montre un rôle prépondérant d’H-NS. De plus, la régulation de cet opéron par Hha et YdgT semble être dépendante ou indépendante de H-NS en fonction des conditions de culture de la bactérie. La répression de l’opéron pefI-srgC, portant l’ORF rck, semble, quant à elle, plus complexe. A l’heure actuelle, seule la région cible de la répression par Hha et YdgT a pu être identifiée. Ces résultats renforcent l’hypothèse de l’existence de régulations par Hha-YdgT indépendantes de H-NS. / The interaction with host cells is an essential step of Salmonella infection cycle. Among other virulence factors, fimbriae allow attachment of Salmonella to eukaryotic cells and invasins enable cell internalisation. Generally, the expression of these factors is suppressed and they are only expressed in vivo at very specific locations. Our objective was to study the mechanisms of Pef fimbriae and Rck invasin repression. Our results show that the nucleoproteins H-NS, Hha and YdgT negatively regulate the expression of these two virulence factors. The mechanism of pef operon transcription repression was characterized and shows a predominant role of H-NS. Moreover, the regulation of this operon by Hha and YdgT appears to bedependent or independent of H-NS depending on bacterial culture conditions. Repression of the pefI-srgC operon, carrying rck ORF, shows a higher degree of complexity. Currently, only the Hha and YdgT targeted region for the repression has been identified. These results reinforce the hypothesis that Hha-YdgT can act independently of H-NS.

Um estudo prévio descreveu uma cepa de Escherichia coli enteropatogênica (EPEC) do sorotipo O142:H34 (Ec48/66) que apresentava o padrão de adesão agregativa (AA), característico de E. coli enteroagregativa (EAEC), e os genes eae e aatA. O objetivo deste estudo foi avaliar o papel de fímbrias no padrão AA dessa cepa. A mesma aderiu no padrão AA nas linhagens HeLa, HEp-2, HT-29 e T84, além de causar a lesão A/E em células HeLa e HT-29. Foi detectada a presença dos genes que compõem o operon da fímbria AAF/I de EAEC e da fímbria E. coli common pilus (ECP). Ambos os operons foram sequenciados mostrando alta identidade com os operons presentes em EAEC (agg) e EPEC (ecp). Mutantes nos genes aggC e ecpC mantiveram o padrão AA e a capacidade de causar a lesão A/E em células HeLa. O mutante em aggC apresentou diminuição significativa da adesão. Análises filogenéticas posicionaram essa cepa em um grupo constituído por EPEC. A cepa Ec48/66 é uma EPEC atípica que expressa o padrão AA, o qual é parcialmente dependente de uma variante de AAF/I, mas não de ECP. / A previous study described a strain of enteropathogenic Escherichia coli (EPEC), serotype O142:H34 (Ec48/66), presenting the aggregative adherence pattern (AA), typical of enteroaggregative E. coli (EAEC), and the eae and aatA genes. The aim of this study was to evaluate the role of fimbriae in the AA pattern of Ec48/66. This strain presented the AA pattern in HeLa, HEp-2, HT-29 and T84 cells, and the ability to cause the A/E lesion in HeLa and HT-29 cells. The presence of the genes comprising the operons of AAF/I fimbriae and E. coli common pilus (ECP) was detected. Both operons were sequenced showing high identity with the operons present in EAEC (agg) and EPEC (ecp). Mutants in the aggC and ecp genes maintained the AA pattern and the ability to cause the A/E lesion in HeLa cells. The aggC mutation significantly reduced the adherence. Phylogenetic analyzes located the strain in the group consisting of EPEC. The Ec48/66 strain is an atypical EPEC expressing the AA pattern, which is partially dependent on a variant of AAF/I, but not on ECP.

Mémoire numérisé par la Division de la gestion de documents et des archives de l'Université de Montréal.

La bactérie Escherichia coli, habituelle commensale de la microflore intestinale des mammifères, peut être responsable de diarrhées chez les veaux nouveaux-nés. Parmi les E. coli pathogènes chez le veau, les souches nécrotoxinogènes de type NTEC2 produisent des adhésines de la famille des fimbriae F17. Ces fimbriae sont caractérisés par un polymorphisme de leur piline F17A et de leur adhésine F17G, qui se traduit par l'expression de variants dont plusieurs n'ont pas encore été décrits. Cette thèse rapporte la caractérisation de deux nouveaux variants F17e-A et F17f-A de la piline et d'un nouveau variant F17-G3 de l'adhésine. Le variant F17f-A, qui correspond à une combinaison des variants F17c-A et F17d-A, a été identifié sur le plasmide Vir d'une souche NTEC2 isolée d'un veau diarrhéique. Identifiés respectivement chez des souches d'E. coli isolées de veaux diarrhéiques en Iran et une souche commensale bovine d'E. coli, les variant F17e-A et F17-G3 ont été recherchés et leurs supports génétiques caractérisés au sein de souches d'E. coli isolées de veaux sains et diarrhéiques, afin de déterminer leur association à la virulence. Alors que le gène codant le variant F17-G3 est exclusivement chromosomique ‒ détecté notamment au niveau d'un probable îlot de pathogénicité, le gène codant le variant F17e-A est chromosomique ou associé aux gènes codant les toxines CNF2 et CDT-III ‒ caractéristiques des souches NTEC2 ‒ sur des plasmides IncF appartenant à une même lignée. Ces résultats confirment le lien entre fimbriae F17 et NTEC2 et mettent en évidence le rôle évolutif de supports plasmidiques dans l'émergence de ce pathovar / The bacterium Escherichia coli is commonly found in the normal intestinal microflora in mammals but may be responsible for diarrhea outbreaks in newborn calves. Among pathogenic E. coli in calves, necrotoxigenic strains of NTEC2 type produce adhesins of the F17 fimbriae family. These fimbriae are composed of the polymorphic pilin F17A and adhesin F17G, for which several variants are unknown. In this thesis, the two new pilin variants F17e-A and F17f-A and the new adhesin variant F17-G3 were reported. F17f-A is a mix of the pilin variants F17c-A and F17d-A and was identified on a Vir plasmid in a NTEC2 strain isolated from a diarrheic calf. F17e-A and F17-G3 were identified in E. coli strains isolated from diarrheic calves in Iran and from a bovine commensal E. coli strain respectively. To determine their association to virulence, a screening of these variants was performed on E. coli strains isolated from healthy and diarrheic calves. The genetic carriage of these variants was also determined. The F17-G3 encoding gene was exclusively carried by bacterial chromosome, notably on a pathogenicity island. The F17e-A encoding gene was chromosomally located or in association with the encoding genes of the NTEC2 typical toxins CNF2 and CDT-III on IncF plasmids that belong to a same plasmidic family. These results confirm the link between F17 fimbriae and NTEC2 strains and underline the role of plasmids in NTEC2 pathovar evolution and appearance