Thesis (Ph. D.)--West Virginia University, 1999. / Title from document title page. Document formatted into pages; contains x, 148 p. : ill. Includes abstract. Includes bibliographical references.

Thesis (Ph. D.)--University of Washington, 2008. / Vita. Includes bibliographical references (leaves 108-128).

Salmonella do grupo paratifóide é responsável por toxi-infecção alimentar no homem, veiculada por alimentos contaminados. Este estudo determinou o perfil genotípico de nove amostras de S. Typhimurium, a sua patogenicidade, assim como sua capacidade de colonização e invasão em aves SPF. Verificou-se pela análise dos genes: agfA, avrA, cdtB, fimA, fliC, invA, iroN, lpfC, mgtC, pefA, sefC, sifA, sipB, sipC, sitC, slyA, sopB, sopE1, sptP ou spvC em amostras de Salmonella Typhimurium que todas foram negativas para os genes sopE1 ou spvC. O gene cdtB estava presente em apenas uma amostra (11,11%) e o gene pefA em duas amostras (22,22%). O gene sefC foi encontrado em três amostras (33,33%). Em oito amostras (88,88%) os genes agfA, fimA, lpfC ou sptP estiveram presentes. Os genes avrA, fliC, invA, iroN, mgtC, sifA, sipB, sipC, sitC ou slyA ou sopB estiveram presentes em 100% das amostras analisadas. Determinou-se quatro perfis genotípicos. No ensaio de patogenicidade observou-se que as amostras inoculadas por via oral, não causaram mortalidade de pintinhos SPF de um dia de idade, com a exceção da amostra SA 633 e SA 006 que apresentaram 30 e 10%, respectivamente. No entanto, observou-se que a infecção por via subcutânea provocou uma maior mortalidade de pintinhos, sendo que as amostras SA 003, SA 004, SA 005 e a amostra vacinal mostraram somente 10% de mortalidade, a amostra SA 002 30%, as amostras SA 632 e SA 634 70% e a amostra SA 633 100%. A amostra SA 006 não provocou nenhuma mortalidade. A amostra mais patogênica pela via subcutânea foi a SA 633. O ensaio de colonização foi realizado em pintinhos SPF, com as amostras SA 002, SA 003, SA 004, SA 005, SA 006, SA 632, SA 633, SA 634 e amostra vacinal. Verificou-se ausência de invasão no fígado e baço 24 horas pós- infecção, exceto para as amostras SA 632 (30%) e amostra vacinal (20%). Após sete dias da infecção houve invasão em dois pintinhos com a amostra SA 002 e um pintinho com as amostras SA 004 e SA 005. Apenas na amostra SA 632 constatou-se colonização em ceco após 24 horas em 10% das amostras e após 7 dias em 70% pós-infecção. Concluiu-se que entre as amostras de Salmonella Typhimurium analisadas existiam diferentes perfis genotípicos baseando-se na presença ou ausência de genes de virulência, e que a amostra vacinal assemelha-se a amostras de S. Typhimurium estudadas quanto a presença dos genes. Os resultados do teste de patogenicidade das amostras de ST indicaram que a via de inoculação modifica a patogenicidade de uma mesma amostra e que a mortalidade após a inoculação pela via subcutânea é sempre superior que pela via oral. / Parathyphoid Salmonella are major food-borne pathogens spread by contaminated food products. This study determined the genotypic profile of nine samples of S. Typhimurium, pathogenicity, colonization and invasion in SPF chicks. It was found by analysis of genes: agfA, avrA, cdtB, fimA, fliC, invA, iroN, lpfC, mgtC, pefA, sefC, sifA, sipB, sipC, sitC, slyA, sopB, sopE1, sptP or spvC samples of S. Typhimurium all were negative to sopE1 or spvC. The cdtB gene was identified in one sample and pefA gene in two samples (22,22%). Sef C gene was detected in three samples (33,33%). In eight samples (88,88%) agfA, fimA, lpf or sptP were detected. AvrA, fliC, invA, iroN, mgtC, sifA, sipB, sipC, sitC, slyA and sopB were detected in all samples evaluated. Four genotypic profiles were established. Pathogenicity tests showed that samplesinoculated by oral gavage did not present mortality in one day old SPF chicks, except samples SA 633 and SA006 with 30 and 10%, respectively. However, it was observed that subcutaneous inoculation showed high mortality in SPF chicks than oral inoculation, and samples SA 003, SA004, SA005 and vaccinal strain showed 10% of mortality, 30% for sample SA002, 70% in the samples SA632 and SA 634 and 100% when the sample SA 633 was inoculated. No mortality was observed in sample SA006. Colonization test was performed in SPF chicks using the samples: SA002, SA 003, SA 004, SA005, SA006, SA 632, SA 633, SA 634 and vaccinal strain. The more pathogenic strain subcutaneously was the SA 633. There was not invasion in liver and spleen 24 hours p.i., except for sample SA 632 (30%) and vaccinal strain (20%). Seven days p.i. invasion was detected in two chicks inoculated with samples identified as SA 004 and SA 005. Sample SA 632 showed 10% of cecal colonization 24 hours p.i. and 70% after one week p.i. It was concluded that Salmonella Typhimurium strains including the vaccinal strain, showed different genotypical profiles, based on the presence or absence of genes of virulence genes. The results of the pathogenicity test indicated that inoculation route modify the pathogenicity of the same strain, and the mortality post-inoculation was always higher in chicks inoculated by subcutaneous route when compared to the oral route.

O-Acetylserine Sulfhydrylase (OASS) is a pyridoxal phosphate enzyme that catalyzes the reaction of O-acetyl-Lserine with sulfide to give L-cysteine. OASS is present as two isoforms, designated -A and -B. The kinetic mechanism of OASS-A is well known and there is also much known concerning the acid-base chemistry of the enzyme. However, little is known concerning the location of the rate determining steps, the sequencing of chemical steps that occur at the active site, or the nature of the rate determining transition states. The studies performed to help elucidate these aspects of the OASS-A mechanism included determination of the thermodynamics of both half reactions, along with studies utilizing substrate analogs of OAS halting the reaction at specific points along the reaction pathway allowing the identification of reaction intermediates. The free energy change of the first half reaction was shown to be -5.7 Kcal/mole while the second half reaction was shown to be, for all intents and purposes, irreversible. Intermediates along the reaction pathway that have been previously identified include the internal Schiff base and the a-aminoacrylate. The external Schiff base was identified using the analogs cysteine, alanine, and glycine while the geminal diamine was identified using the analog serine. Formation of the external aldimine was shown to be pH dependent with a pK of 8.1 ± 0.3 most likely representing a general base that accepts a proton from the a-amine of cysteine to facilitate a nucleophilic attack on C4r of the PLP imine. Formation of the geminal diamine was also shown to be pH dependent with two pK values having an average value of 8.1. One of the groups most likely represents the general base which accepts a proton from the a-amine of cysteine while the second group likely interacts with the amino acid side chain to orientate the amino acid into the correct configuration.

O-Acetyl-L-serine sulfhydrylase-A (OASS-A) forms acetate and L-cysteine from O-acetyl-L-serine (OAS) and sulfide. One molecule of the cofactor pyridoxal 5'- phosphate (PLP) is bound in each holoenzyme protomer.

by Ng Wai-Leung. / Thesis submitted in: August 1997. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1998. / Includes bibliographical references (leaves 135-144). / Title page --- p.i / Thesis Committee --- p.ii / Abstract --- p.iii / Acknowledgments --- p.v / Abbreviations --- p.vi / Table of contents --- p.vii / List of figures --- p.x / List of tables --- p.xiii / Chapter Chapter 1. --- General introduction --- p.1 / Chapter Chapter 2. --- Literature review / Chapter 2.1 --- Biosynthesis of deoxyribonucleotide triphosphates --- p.3 / Chapter 2.2 --- Ribonucleotide reductase --- p.6 / Chapter 2.2.1 --- Class I ribonucleotide reductase --- p.6 / Chapter 2.2.2 --- Class II ribonucleotide reductase --- p.13 / Chapter 2.2.3 --- Class III ribonucleotide reductase --- p.14 / Chapter 2.3 --- Proposed mechanism for ribonucleotide reduction --- p.17 / Chapter 2.4 --- Allosteric control of ribonucleotide reductase --- p.21 / Chapter 2.4.1 --- Allosteric control of class I ribonucleotide reductase --- p.21 / Chapter 2.4.2 --- Allosteric control of class II and class III ribonucleotide reductases --- p.23 / Chapter 2.5 --- Evolution of ribonucleotide reductase --- p.25 / Chapter 2.6 --- Central metabolism pathways of enteric bacteria --- p.28 / Chapter 2.7 --- Regulation of gene expression by oxygen in enteric bacteria --- p.33 / Chapter 2.7.1 --- Regulation of gene expression by Fnr --- p.33 / Chapter 2.7.2 --- Regulation of gene expression by AcrAB --- p.39 / Chapter 2.7.3 --- Regulation of gene expression by NarXL and NarQP --- p.42 / Chapter 2.7.4 --- Other aspects in anaerobic gene expression --- p.45 / Chapter 2.7.5 --- Relationship between NrdD and anaerobic metabolism --- p.45 / Chapter 2.8 --- Objectives --- p.46 / Chapter Chapter 3. --- Molecular cloning and sequencing of nrdDG operon from Salmonella typhimurium / Chapter 3.1 --- Introduction --- p.47 / Chapter 3.2 --- Material and methods --- p.48 / Chapter 3.2.1 --- Bacterial strains and bacteriophages --- p.48 / Chapter 3.2.2 --- Culture media --- p.48 / Chapter 3.2.3 --- Preparation of lambda lysate and phage DNA --- p.48 / Chapter 3.2.3.1 --- Plating out pf lambda phage and preparation of plate lysate --- p.48 / Chapter 3.2.3.2 --- Preparation of lambda lysate stock --- p.49 / Chapter 3.2.3.3 --- Preparation of lambda phage DNA --- p.50 / Chapter 3.2.4 --- Long distance polymerase chain reaction (LD-PCR) of nrdDG gene fragment --- p.51 / Chapter 3.2.5 --- Restriction enzyme digestion of LD-PCR products and subcloning of restriction fragments --- p.52 / Chapter 3.2.6 --- Confirmation of recombinants by colony-PCR --- p.53 / Chapter 3.2.7 --- Preparation of plasmid DNA by alkaline lysis using Wizard´ёØ Plus Miniprep DNA Purification System (Promega) --- p.54 / Chapter 3.2.8 --- DNA cycle sequencing by using dye-labeled dideoxy chain terminator and data collection --- p.55 / Chapter 3.2.9 --- Computer software for analyzing and manipulating DNA sequences --- p.57 / Chapter 3.3 --- Results --- p.59 / Chapter 3.3.1 --- Preparation of lambda DNA --- p.59 / Chapter 3.3.2 --- Long distance PCR amplification of nrdDG from lambda DNA --- p.59 / Chapter 3.3.3 --- Restriction digestion of LD-PCR products --- p.61 / Chapter 3.3.4 --- Subcloning of LD-PCR restriction fragments --- p.61 / Chapter 3.3.5 --- Miniprep of plasmid DNA from recombinants and verification of nrdDG identities --- p.64 / Chapter 3.3.6 --- Nucleotide sequence of nrdDG --- p.66 / Chapter 3.4 --- Discussions --- p.72 / Chapter 3.4.1 --- Sequence analysis of S. typhimurium nrdDG --- p.72 / Chapter 3.4.2 --- Experimental design --- p.79 / Chapter Chapter 4. --- Transcriptional regulation of anaerobic ribonucleotide reductase in Salmonella typhimurium in aerobic and anaerobic environments / Chapter 4.1 --- Introduction --- p.84 / Chapter 4.2 --- Materials and methods --- p.86 / Chapter 4.2.1 --- Bacteria and bacteriophages strains / Chapter 4.2.2 --- Culture media --- p.86 / Chapter 4.2.3 --- Construction and characterization of oxrA mutant --- p.87 / Chapter 4.2.3.1 --- Preparation of P22 lysate of TN2336 --- p.87 / Chapter 4.2.3.2 --- P22 transduction for construction of oxrA mutant --- p.87 / Chapter 4.2.3.3 --- Characterization of oxrA mutant --- p.87 / Chapter 4.2.4 --- Extraction of bacterial RNA by hot phenol method --- p.88 / Chapter 4.2.5 --- Formaldehyde gel electrophoresis of RNA --- p.88 / Chapter 4.2.6 --- Reverse transcriptase polymerase chain reaction (RT-PCR) of nrdD transcript --- p.89 / Chapter 4.2.7 --- Transfer of DNA/RNA to solid support --- p.90 / Chapter 4.2.7.1 --- Transfer of DNA to solid support by Southern blotting --- p.90 / Chapter 4.2.7.2 --- Transfer of RNA to solid support by Northern blotting --- p.91 / Chapter 4.2.7.3 --- RNA Dot blot --- p.91 / Chapter 4.2.8 --- Preparation of radioactive-labeled probes for hybridization --- p.92 / Chapter 4.2.8.1 --- Synthesis of radioactive-labeled probes by labeling --- p.92 / Chapter 4.2.8.2 --- Preparation of RNA probe by in vitro transcription --- p.93 / Chapter 4.2.9 --- Hybridization and membrane washing conditions --- p.95 / Chapter 4.2.10 --- Normalization of samples by 16S ribosomal RNA (rRNA) --- p.95 / Chapter 4.3 --- Results --- p.97 / Chapter 4.3.1 --- Preparation of RNA --- p.97 / Chapter 4.3.2 --- RT-PCR of nrdD transcript --- p.97 / Chapter 4.3.3 --- Northern blot analysis of nrdD transcript --- p.103 / Chapter 4.3.4 --- Dot blot hybridization analysis of nrdD expression in an oxrA mutant --- p.103 / Chapter 4.4 --- Discussions --- p.107 / Chapter 4.4.1 --- Expression of nrdD of S. typhimurium in aerobic and anaerobic environments --- p.107 / Chapter 4.4.2 --- Experimental design --- p.110 / Chapter Chapter 5. --- Characterization of nrdD::Tn10 mutant of S. typhimurium / Chapter 5.1 --- Introduction --- p.112 / Chapter 5.2 --- Materials and methods --- p.112 / Chapter 5.2.1 --- Bacteria and bacteriophages strains --- p.113 / Chapter 5.2.2 --- Transduction of zzz-3875::Tn10 to S. typhimurium --- p.113 / Chapter 5.2.3 --- Characterization of zzz-3875::Tn10 by Southern hybridization --- p.113 / Chapter 5.2.3.1 --- Preparation of genomic DNA from S. typhimurium --- p.113 / Chapter 5.2.3.2 --- Restriction enzyme digestion of genomic DNA and Southern hybridization --- p.114 / Chapter 5.2.4 --- Characterization of growth pattern of nrdD::Tn10 mutant --- p.115 / Chapter 5.3 --- Results --- p.116 / Chapter 5.3.1 --- Characterization of zzz-3 875: :Tn7 0 in S. typhimurium --- p.116 / Chapter 5.3.2 --- Characterization of growth pattern of nrdD mutant --- p.120 / Chapter 5.4 --- Discussions --- p.125 / Chapter Chapter 6. --- General Discussions / Chapter 6.1 --- General discussions --- p.131 / Chapter 6.2 --- Further studies --- p.134 / References --- p.135

Thesis (M. Sc.)(Microbiology)--University of Pretoria, 2005. / Includes summary. Includes bibliographical references (leaves 87-108). Available on the Internet via the World Wide Web.

"Erratum" [i.e. Errata] inserted / Includes bibliography / iv, 83 leaves, [33] leaves of plates : ill ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.)--University of Adelaide, Dept. of Microbiology and Immunology, 1989

Typescript (photocopy) / xiv, 203 leaves, [2] leaves of col. plates : ill. ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.)--University of Adelaide, Dept. of Microbiology, 1983

xiv, 207 leaves : ill., tables ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.)--University of Adelaide, Dept. of Microbiology, 1976