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Gan, Teck Fong Emily, and xf_dksfwm@yahoo com. "Molecular Characterisation of Salmonella enterica Serovar Sofia in Australia." RMIT University. Applied Sciences, 2008. http://adt.lib.rmit.edu.au/adt/public/adt-VIT20080821.163140.
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Despite its high isolation frequency in Australian chickens, S. II Sofia is rarely associated with animals or human salmonellosis as this serovar is avirulent in nature. The reason for its persistence and avirulence is unknown as very few studies have been conducted on the epidemiology and pathogenicity of this strain. This study details the various experimental methods utilised to investigate the genetic relatedness and molecular mechanisms involved in S. II Sofia pathogenesis. Using PFGE and Rep-PCR, the Australian S. II Sofia isolates were found to show limited genetic diversity and probably share a clonal relationship. A majority of the S. II Sofia isolates were not geographically restricted with the predominant pattern subtype spread out among the isolates from various states. Distribution and variation of the SPI-associated virulence genes within S. II Sofia was also examined. Based on RFLP and sequence analysis, most of the differences observed in SPI1 to SPI5 of S. II Sofia could be attributed to a loss or gain of restriction cleavage sites within these regions. However, a number of genes in SPI1, SPI2, SPI3 and SPI5 were found to have accumulated changes (mutations, insertions and deletions) that could have affected gene transcription and/or protein translation - these genes have been shown to be involved in different aspects of the virulence process. The avirulence of S. II Sofia is probably not the result of a single genetic change but rather a series of alterations to a large number of its virulence-associated genes. Plasmid-mediated virulence was also assessed in S. II Sofia isolates. Southern hybridisation with probes derived from the virulence plasmid of S. Typhimurium indicated either the total absence of the virulence plasmid or possible presence of a virulence plasmid containing major deletions. Clones were constructed with the missing spv operon using high-copy pCR®2.1 and low-copy pWSK29 plasmids and the adherence, invasion and intracellular survival of the mutant strain was evaluated in vitro. The presence of spvRABCD was shown to have no effect on intracellular survival and replication. Although the cloning of spv with pCR®2.1 was observed to significantly increase invasiveness of S. II Sofia, it was not capable of restoring the invasive ability of S. II Sofia to the level of pathogenic S. Typhimurium 82/6915. On the other hand, the uneven adherence and invasion ability of the other mutant strains appeared to be linked to the presence of pWSK29 and this observation is further supported by RT-PCR analysis of the clones - indicating that perhaps pWSK29 is not a suitable vector for this study. Wild-type S. II Sofia isolates are unlikely to regain full pathogenicity because of the numerous mutations in many important virulence genes: even the chance acquisition of a virulence factor (e.g. spvRABCD) is not sufficient to completely restore S. II Sofia virulence. Therefore, S. II Sofia should not be considered similar to other Salmonella spp. when monitoring Salmonellae in food samples.
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Osman, Deenah. "Copper Homeostasis in Salmonella Enterica Serovar Typhimurium." Thesis, University of Manchester, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.509383.
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Teixidó, Devesa Laura. "Estudio del regulón Fur en Salmonella enterica serovar Typhimurium." Doctoral thesis, Universitat Autònoma de Barcelona, 2013. http://hdl.handle.net/10803/117269.
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El hierro es un oligoelemento esencial para la supervivencia celular ya que es un cofactor de muchas enzimas y forma parte de la estructura de muchas proteínas. Es por este motivo que los microorganismos necesitan mecanismos eficientes para la captación de hierro cuándo carecen del mismo. Está ampliamente descrito que la captación de hierro es uno de los pasos clave en el desarrollo de un patógeno dentro de su huésped 1. Pero, aunque el Fe es indispensable, su exceso en el citoplasma es tóxico para la célula ya que cataliza la reacción de Fenton con la consiguiente formación de radicales hidroxilo 2, 3.
Por todo ello la homeóstasis del Fe2+ se encuentra estrictamente controlada. La proteína Fur (ferric uptake regulator) es el principal regulador transcripcional implicado en la respuesta celular a la concentración de hierro, controlando tanto la inducción de sistemas de captación de Fe2+ de alta afinidad, como la expresión de proteínas para el almacenamiento y enzimas que utilizan hierro 4. Normalmente Fur, asociado al ión Fe2+, se une a una secuencia concreta denominada caja Fur presente en la región promotora de los genes que regula, bloqueando de esta manera su transcripción 5. Aunque también puede activar la expresión de diferentes genes de forma directa o indirecta 1.
En el presente trabajo se estudia, mediante microarrays de DNA, el papel de la proteína Fur en el patógeno intracelular S. enterica serovar Typhimurium y la relación de este regulador con los mecanismos de virulencia de dicho microorganismo.Iron is an essential trace element for the cell since it is a cofactor for many enzymes and is a part of the structure of many proteins. It is for this reason that the microorganisms need efficient mechanisms for iron uptake when lacking it. It is widely reported that iron uptake is one of the key steps in the development of a pathogen within its host 1. Although Fe2+ is indispensable, its excess in the cytoplasm is toxic to the cell since it catalyzes the Fenton reaction leading to the formation of hydroxyl radicals 2, 3. Therefore Fe2+ homeostasis is strictly controlled. Protein Fur (ferric uptake regulator) is the major transcriptional regulator involved in the cellular response to iron concentration, by controlling the induction of Fe2+ high affinity uptake systrems and the protein expression of iron storing and utilizing enzymes 4. Normally, Fur, associated to the Fe2+ ion, binds to a specific sequence called Fur box present in the promoter region of genes the regulated genes, thus blocking its transcription 5. Fur also can activate the expression of different genes directly or indirectly 1. The role of the Fur protein in the intracellular pathogen S. enterica serovar Typhimurium and its relationship with the virulence mechanisms of this microorganism is studied in this work by DNA microarrays.
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Hu, Honghua. "Molecular typing and evolution of Salmonella enterica serovar Typhimurium." University of Sydney. School of Molecular and Microbial Biosciences, 2005. http://hdl.handle.net/2123/704.
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Salmonella enterica serovar Typhimurium is a common cause of salmonellosis among humans and animals worldwide. In Australia, Typhimurium is responsible for over half of the salmonellosis cases. The Anderson phage-typing scheme is the primary means of long-term surveillance of Typhimurium outbreak isolates, and has played an important role in epidemiology. However, there exist quite a number of strains of Typhimurium that cannot be defined by the phage-typing scheme. Furthermore, the knowledge of evolutionary relationships among isolates of different phage types is still very limited and the genetic basis of phage type variation remains largely unknown. To address these issues, this study focused on molecular typing and evolution of Typhimurium. Fluorescent amplified-fragment length polymorphism (AFLP) was applied to 46 Typhimurium isolates comprising nine phage types in Australia using the restriction enzymes MseI and EcoRI and MseI +1 / EcoRI +1 primer pair combinations. The selected phage types, DT9, DT135, DT64, DT44, DT126, DT12a, DT1, DT141 and DT108, have been dominant or frequent phage types in animal and human infections in Australia in recent years. AFLP in the present study showed a very good discrimination power with Simpson index of diversity of 0.98, 35 different AFLP patterns were observed in the 46 isolates studied. The tree based on AFLP patterns showed good correlation with phage type, grouped most Typhimurium isolates by phage type, and differentiated all nine phage types. Furthermore, 84 phage-type specific polymorphic AFLP fragments, for which presence or absence correlated with phage type (including 25 with one exception to phage-type specificity) were observed in the 46 strains studied. Eighteen phage-type specific AFLP fragments were cloned and sequenced. Sixteen are of known genes or have a homologue in the databases. It was found a predominance of phage and plasmid genes rather than mutational changes in the AFLP fragments studied. Of the 18 cloned and sequenced AFLP fragments, only four relate to mutational changes in the S. enterica chromosome, the other 14 comprise DNA of mobile elements: nine are phage related, three are plasmid related and two are gain of DNA from unknown origin. Twelve of the 18 sequenced phage-type specific AFLP markers are polymorphic because the DNA is present or absent as indicated by Southern hybridization. Two of these markers were successfully used in preliminary PCR-based typing of 30 DT9 and 29 DT135 isolates from worldwide collections. 27 of the 30 DT9 isolates and all DT135 isolates tested were correctly categorized. The results implied a good potential to use the sequence of these fragments as the basis for a multiplex PCR or a microarray based molecular �phage� typing method for Typhimurium. This thesis also studied the molecular evolutionary relationships among the same set of 46 Typhimurium isolates using mutational changes detected by AFLP, or analysis of intergenic regions and their flanking genes in genome sequences. The complete genome sequence of Typhimurium LT2 was analysed by computer modelled AFLP. The polymorphic AFLP fragments, which matched with the modelled LT2 AFLP fragments, were amplified and sequenced by LT2 genome based primers to determine the changes. Forty-nine intergenic regions with higher pairwise differences between LT2 and Typhi CT18 were amplified and sequenced using LT2 genome based primers for one isolate of each phage type. 51 polymorphic sites were detected consisting of 18 in AFLP fragments and 33 in intergenic regions or their flanking genes. PCR-RFLP (restriction fragment length polymorphism) and SNaPshot were used to further investigate the distribution of the single nucleotide polymorphisms (SNPs) detected in intergenic regions in all isolates studied. Of the 18 mutational changes detected in AFLP fragments, eight were indels (insertions / deletions) and ten single base substitutions. Of the eight indels, four were in genes, three in intergenic regions, and one covered adjacent intergenic and coding regions. The four indels in genes all caused frameshift mutations, including three single base indels and one 19 bp deletion. Of the ten substitutions, one was in an intergenic region and nine in genes comprising three synonymous and six non-synonymous substitutions. Of the 33 polymorphic sites detected from sequences of 23 intergenic regions and their flanking genes, one was IS200 insertion and 32 single nucleotide polymorphisms (SNPs), of which 30 were single base substitutions and two were single base indels. Nine of the 33 variations were found in the flanking genes, which were all single base substitutions comprising four synonymous, four non-synonymous substitutions and one non-sense mutation. More non-synonymous than synonymous substitutions were found for those in coding regions within Typhimurium, indicating that slightly deleterious intraspecies mutations can be fixed within clones, such as various lineages of Typhimurium. The 51 polymorphic sites, which were inferred from sequences of both mutation related AFLP fragments, and intergenic regions and their flanking genes, gave a single phylogenetic tree of the 46 Typhimurium isolates studied. All sequences involved were compared with the homologous sequences in the available S. enterica genome sequences for serovars Typhi, Paratyphi A, Gallinarum, Enteritidis and Pullorum and this enabled the determination of the direction of the mutational changes in the isolates studied and the root of the phylogenetic tree. There were only two events inferred to have occurred twice, the remaining 49 polymorphisms can be explained by a single event. The data indicated that Typhimurium has a very strong clonal structure with a very low level of recombination over the time for diversification of Typhimurium as majority of clonal variations are from point mutations rather than recombination. The phylogenetic tree based on mutational changes showed that most Typhimurium isolates of a given phage type are in the same evolutionary group, but that some phage types appear to have arisen more than once. Comparison of the phylogenetic tree with AFLP data gave examples of unrelated isolates of a given phage type having common AFLP fragments comprising plasmid or phage genes, supporting the view that phage type can be determined by presence of specific phages or plasmids. The mutation-based tree showed that six of the nine phage types studied appeared to have a single origin, at least for the isolates studied. It also found that DT1 and DT44 had two independent origins even for the limited set of strains used. The distribution of DT12a isolates into two groups could be explained that the group of three DT12a isolates were derived from the other group of four DT12a isolates, where the root of the tree might be. The data also confirmed that DT64 arose from DT9. The phylogenetic tree that was generated based on essentially mutational changes provides clear relationships of the closely related Typhimurium isolates with high level of consistency and reasonable confidence. This study provided one of the few analyses of relationships of isolates within a clone. Matching actual AFLP with computer modeled AFLP and sequencing intergenic regions provide very good new strategies to identify mutational polymorphisms and to study the molecular evolutionary relationships in the closely related isolates.
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Moore, Gillian Fiona. "Factors influencing biofilm formation by Salmonella enterica serovar Enteritidis." Thesis, University of Exeter, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.248173.
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Phan, Minh-Duy. "Analysis of IncHI1 plasmids in Salmonella enterica serovar Typhi." Thesis, University of Cambridge, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.608508.
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Holden, Nicola Jean. "The cold shock response of Salmonella enterica serovar Typhimurium." Thesis, University of Edinburgh, 2000. http://hdl.handle.net/1842/28240.
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The alternative sigma factor, ss, did not appear to play a significant role in cspB expression at low temperature. In contrast, Fis appeared to act as a positive regulator of cspB in stationary phase cultures. Cell survival assays (measured by ability to form colony forming units on nutrient agar plates) showed that 4% of cells that were in early exponential phase survived a rapid cold shock to 4°C, although survival was almost complete when cells were in lag phase or in late exponential phase. The alternative sigma factor, ss, did not appear to play a significant role in cell survival in response to a rapid temperature reduction to 4°C. The addition of an osmoprotectant, 0.3 M sucrose, protected against loss of plating viability to some degree for early exponential phase cells, when diluted to 4°C. 2-D PAGE analysis showed that the response of exponential phase S. typhimurium cells incubated at 10°C consisted of an adaptive phase followed by an acclimation phase, in agreement with previous reports for E. coli. Identification of CspA was verified by N-terminal sequencing. The response was delayed at 4°C and recovery of protein synthesis in the acclimation phase was not as extensive, as observed at 10°C. CspA was synthesised throughout the period of incubation at 4°C. Growth phase was found to severely affect de novo protein synthesis at low temperature. Incubation of stationary phase cells at 10°C or 4°C resulted in repression of the synthesis of the majority of proteins, although a small set of proteins was induced. CspA was not detected at 37°C, but was highly induced at 10°C. However, prolonged incubation at 4°C led to complete repression of protein synthesis, except for CspA. This study has shown that S. typhimurium adapts to low temperature in a dynamic fashion and expression of CspA is a major feature of the response. Furthermore, it appears that exponential phase S. typhimurium cells are metabolically active even after 4 days at refrigeration temperatures.
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Perkins, Timothy Trevor. "Characterisation of the transcriptome and proteome of Salmonella enterica subspecies enterica serovar Typhi." Thesis, University of Cambridge, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.611537.
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Bjur, Eva. "Virulence of Salmonella enterica serovar typhimurium and innate antibacterial host responses /." Stockholm, 2006. http://diss.kib.ki.se/2006/91-7140-946-7/.
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Sangal, Vartul. "Multilocus sequence typing analyses of Salmonella enterica subspecies enterica." Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2009. http://dx.doi.org/10.18452/15883.
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Serovare von Salmonella enterica subspecies enterica sind im allgemeinen pathogen für Mensch und andere Säugetiere. In dieser Arbeit habe ich anhand eines “Multilocus Sequences Typing” Typisierungsschemas die Populationsstruktur einer der am häufigsten auftretenden Serovaren dieser Subspecies, das aus Menschen und Schlachttieren isolierte Serovar Newport charakterisiert. Dieses Schema wurde auch für die Charakterisierung von Isolaten derselben Subspecies aus humanen Dauerträgern und Reptilien verwandt, um zu bestimmen, ob Isolate aus diesen Quellen sich in ihrer Populationstruktur von denjenigen unterscheiden, die aus anderen Quellen isoliert wurden. Multilocus Sequences Typing ist eine weitgehend für die Untersuchung der Evolution und Populationsstruktur von einen breiten Spektrum von Organismen verwendete Technik. 400 - 600 bp lange Fragmente von 7 Haushaltsgenen wurden sequenziert, und jede einzelne Sequenz jedes einzelnen Gens wurde eine Allelnummer zugeordnet. Jede einzelne Allelkombination wurde einem Sequenztyp zugeordnet. Die so gewonnenen Daten wurden weiter analysiert. Drei “Lineages”, Newport-I, Newport-II und Newport-III, wurden innerhalb dieses Serovars identifiziert, die jeweils aus Menschen in Europa, Tieren und Menschen in Nordamerika isoliert wurden. Der Multiresistenz-Phänotyp wurde häufiger in Newport II gefunden, während die meisten Newport III Isolate pan-sensitiv waren. Verglichen mit anderen Serovaren war die Anzahl von “Lineages” innerhalb Newport höher als bei Enteritidis, Kentucky und Typhimurium, aber niedriger als bei Paratyphi B. Das heisst, die Serovare von S. enterica subspecies enterica variieren stark in ihrer Populationsstruktur. Die Sequenztypen in Isolaten aus humanen Dauerträgern waren im allgemeinen am häufigsten in Isolaten von klinischen Patienten und Tieren vorhanden. In der Mehrheit der Serovaren waren die meisten Isolate aus Patienten und Tieren genetisch identisch mit solchen, die aus gesunden Trägern isoliert wurden. Die genetische Variabilität war zwischen Isolaten aus diesen Quellen vergleichbar. Diese Ergebnissen deuten daraufhin, dass Salmonellen aus Dauerträgern sowie Isolate aus Patienten und Tieren derselben Population angehören. Die meisten Serovare aus Reptilienisolaten waren genetisch identisch mit denen von Menschen und warmblütigen Tieren. In den Serovaren Bovismorbificans, Decatur, Miami und Oranienburg hingegen waren die meisten Isolate aus Reptilien genetisch anders als Isolate aus anderen Wirten. Allerdings wurden nur wenige Isolate der Serovaren Bovismorbificans, Decatur und Miami aus Reptilien und nur wenige Isolate der Serovaren Oranienburg aus anderen Quellen getestet; eine grössere Anzahl von Isolaten müsste daher untersucht werden, um festzustellen ob diese genetischen Unterschiede statistich signifikant sind oder nicht.Serovars of Salmonella enterica subspecies enterica are generally pathogenic to humans and other mammals. In this study, I examined the population structure of one of the most common serovars of this subspecies isolated from humans and food animals, serovar Newport, using a multilocus sequence typing scheme. This scheme was also used to analyze isolates of this subspecies from chronic human carriers and reptiles to determine whether isolates from these sources represent distinct populations than those from other hosts. Multilocus sequence typing has extensively been used to study evolution and population structure of a wide range of organisms. 400-600 bp fragments of 7 housekeeping genes were sequenced and every unique sequence of each gene fragment was given a distinct allele number. Each unique combination of alleles was assigned a distinct sequence type number. The data were used in further analyses. Three lineages, namely Newport-I, Newport-II and Newport-III were identified within serovar Newport which were associated to European humans, animals and humans in North America, respectively. Multidrug resistance phenotypes were most common in Newport-II whereas most isolates in Newport-III were pan-susceptible. When compared to other serovars, the numbers of lineages within Newport were higher than for Enteritidis, Kentucky and Typhimurium but lower than for Paratyphi B. Therefore, serovars of S. enterica subspecies enterica vary greatly in their population structures. The sequence types observed for isolates from chronic human carriers were generally the most common among human-clinical and animal isolates. Most isolates from non-carrier humans plus animals were genetically identical to the carried isolates within most serovars. Genetic diversity was also comparable between isolates from these sources. These results suggest that salmonellae from chronic human carriers belong to the same population as isolates from non-carrier humans and animals. For most serovars, most isolates from reptiles were genetically identical to those from humans or other warm blooded animals. However, in serovars Bovismorbificans, Decatur, Miami and Oranienburg, most reptile isolates were genetically distinct from isolates from other hosts. Only few reptile isolates were tested from Bovismorbificans, Decatur and Miami and only few non-reptile isolates were tested from Oranienburg, and in larger numbers of such isolates would be needed to determine whether these differences are statistically significant.
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Nogueira, Letícia Amaral. "Estudo molecular da resistência a bacteriófagos líticos em Salmonella enterica subsp. enterica serovar Enteritidis." Botucatu, 2015. http://hdl.handle.net/11449/144057.
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Orientador: Paulo Eduardo Martins RibollaCoorientador: Alexandre Secorun Borges
Banca: Vera Lúcia Mores Rall
Banca: João Pessoa Araújo Junior
Banca: Aline Maria da Silva
Banca: Mario Henrique de Barros
Resumo: Salmonella Enteritidis é um patógeno pertencente à Família Enterobacteriaceae que frequentemente está relacionada a infecções alimentares em humanos, principalmente em crianças, idosos e pacientes imunossuprimidos. A importância deste micro-organismo se dá pela sua prevalência significativa, com distribuição mundial, nos lotes de frangos de corte e de postura de ovos, levando a sérias implicações na saúde pública e ao mercado avícola. O controle eficaz da salmonelose é um desafio, pois envolve um complexo manejo dos animais e tratamento com uso de antibióticos que não garantem a eliminação da infecção. O uso de bacteriófagos líticos contra infecções bacterianas (fagoterapia) vem atender uma demanda crescente por alternativas ao tratamento antimicrobiano convencional. Todavia, a literatura tem relatado o surgimento de certa resistência bacteriana a alguns bacteriófagos líticos, como por exemplo, em Salmonella sp. Diversos são os mecanismos bacterianos de resistência aos fagos, tais como a prevenção de adsorção, o bloqueio da injeção do DNA do fago, restrição-modificação, infecção abortiva e o sistema CRISPR ("clustered regularly interspaced short palindromic repeats") /Cas ("CRISPR associated proteins") de imunidade adquirida. Assim sendo, o presente trabalho objetivou a caracterização molecular da resistência bacteriana de fagos líticos isolados contra cepas de Salmonella enterica subsp. enterica serovar Enteritidis (SE). Para tal, os mecanismos-alvos escolhidos foram o sistema CRISPR/Cas e o bloqueio da adsorção de fagos, via lipopolissacarídeo (LPS). Nas 22 cepas de estudo, um total de 72 CRISPRs foi identificado, com 14 diferentes domínios repetitivos (DR), apresentando um total de 551 sequências de espaçadores e os genes associados cas1, cas2 e cas3. Cas 9 e cas 10 não foram identificados. Foi observado que as cepas resistentes aos fagos líticos possuíam um maior número total...
Abstract: Salmonella Enteritidis is a pathogen that belongs to the Enterobacteriaceae family and is often related to infections transmitted by food in humans, especially in children, elderly and immunosuppressed patients. The importance of this microorganism is because it's significant prevalence with worldwide distribution in lots of poultry, leading to serious implications for public health and poultry industry. Effective control of salmonellosis is a challenging because involves a complex animal handling and treatment with antibiotics that do not guarantee the elimination of infection. The use of lytic bacteriophages against bacterial infections (phage therapy) meets a demand for alternatives to the conventional antimicrobial treatment. However, the literature has reported the emergence of bacterial resistance to some lytic bacteriophages, such as in Salmonella sp. There are several mechanisms of bacterial resistance to phages, such as prevention of adsorption, blocking the injection of phage DNA, restriction-modification, abortive infection and the CRISPR/Cas System acquired immunity. Thus, this research aims to study in a molecular level the bacterial resistance of lytic phages isolated from pathogenic strains of Salmonella enterica subsp. enterica serovar Enteritidis (SE). Two mechanisms have been chosen: the CRISPR/Cas system and blocking of phage adsorption via LPS. From 22 SE strains, a total of 72 CRISPRs was identified with 14 different repetitive domains (DR), presenting a total of 551 spacers' sequences and cas1, cas2 and cas3 CRISPR associated genes. Cas 9 and cas 10 genes weren't identified. It was observed that phage lytic resistant strains have a higher total number of spacers in the CRISPR locus in relation to sensitive strains, and this difference was statistically significant. Phylogenetic analysis of cas genes from CRISPR/Cas system, of rfaC, rfaH, cpsG, manB, manc, lpxA, lpxB and lpxC genes (related to LPS) and of...
Doutor
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Sapparapu, Gopal. "Development of immunological reagents for detecting Salmonella enterica serovar Typhimurium." [Gainesville, Fla.] : University of Florida, 2003. http://purl.fcla.edu/fcla/etd/UFE0001428.
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Octavia, Sophie Biotechnology & Biomolecular Sciences Faculty of Science UNSW. "Molecular typing and evolutionary relationships of Salmonella enterica serovar Typhi." Publisher:University of New South Wales. Biotechnology & Biomolecular Sciences, 2008. http://handle.unsw.edu.au/1959.4/43115.
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The evolutionary relationship between Salmonella enterica serovar Typhi, other typhoid-like enteric fever causing serovars and 10 non-Typhoid serovars from S. enterica subspecies I, could not be determined by comparative nucleotide sequences of six genes. Phylogenetic analyses of the dataset showed that the genes of interest underwent frequent recombination, suggesting a low level of clonality within subspecies I of S. enterica. To establish the evolutionary relationships within serovar Typhi, genome-wide Single Nucleotide Polymorphism (SNP) was explored as a marker for both typing purposes and phylogenetic analysis. Thirty eight SNPs were typed in 73 global Typhi isolates, including 18 isolates expressing the special flagellar antigen z66, using restriction enzyme digestion method. The isolates were differentiated into 23 SNP profiles and grouped into four distinct clusters. The z66 isolates were divided into four SNP profiles and were all grouped into one cluster, suggesting a single origin. An alternative SNP typing method using the hairpin real time PCR assay was investigated to type four additional SNPs, termed as biallelic polymorphisms (BiP). These BiPs were found to classify 481 global Typhi isolates into five major clusters (Roumagnac et al., 2006). Typing four BiPs resulted in the identification of four additional SNP profiles. We proposed nine SNPs were required to type Typhi isolates into 13 subclusters for global epidemiology. An enzymatic-based method using CelI nuclease was evaluated to discover more SNPs from other Typhi genomes. The efficiency of the CelI was shown to be unsatisfactory and we were unable to demonstrate the effectiveness of the proposed method. Nine Variable Number of Tandem Repeats (VNTRs) were typed in the 73 Typhi isolates using fluorescent-labelled universal primers, and analysed on an automated DNA sequencer. Five isolates were unable to give PCR products in one or more VNTR loci. Nine VNTRs could differentiate 68 Typhi isolates into 65 MLVA profiles, suggesting a higher discriminating power than SNP typing. SNPs were shown to be a more appropriate marker for phylogenetic tracing for Typhi while VNTRs were highly discriminating but could not be used to establish the evolutionary relationships of diverse Typhi isolates.
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Žindul, Adam. "Salmonella enterica Serovar Typhimurium inaktyvacijos fotosensibilizacija vertinimas ir poveikio modeliavimas." Master's thesis, Lithuanian Academic Libraries Network (LABT), 2014. http://vddb.library.lt/obj/LT-eLABa-0001:E.02~2011~D_20140701_164247-99768.
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Šiame tiriamajame darbe nagrinėjama bakterijos Salmonella enterica Serovar Typhimurium inkubacijos priklausomybė nuo inkubacinio periodo. Trumpai aptariami bakterijų inaktyvavimą aprašantys dažniausiai naudojami modeliai, skaitiškai išreiškiama lag fazė, randama jos ilgį aprašantį funkciją. Toliau vertinamos tiesinė ir liekamoji dalys bei išvedama inaktyvavimą aprašanti lygtis. Darbas baigiamas išvestinės formulės praktiniu panaudojimu ir rezultatų aptarimu.Evaluation of Salmonella enterica Serovar Typhimurium inactivation by photosensitization and impact modeling The aim goal of this research is to evaluate the influence of irradiation of UV light and incubation period on Salmonella enterica Serovar Typhimurium bacteria. Shortly discussed most commonly used mathematical models of bacterial inactivation, expressed lag phase and its function. Next step is evaluation of line part and tail of inactivation (mortality) curve. At the end of the research the inactivation formula is deduced and the results are discussed.
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Owen, S. V. "Exploring the prophage biology of Salmonella enterica serovar Typhimurium ST313." Thesis, University of Liverpool, 2017. http://livrepository.liverpool.ac.uk/3011773/.
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In the past 30 years, Salmonella bloodstream infections have become a significant health problem in sub-Saharan Africa and are responsible for the deaths of ~390,000 people each year. The disease is largely caused by a recently described sequence type of Salmonella Typhimurium: ST313. Comparative genomic analysis showed that the ST313 lineage is closely-related to the ancestral gastroenteritis-associated Typhimurium sequence type ST19, but carries a distinct prophage repertoire. I hypothesised that prophages contribute to the biology of this clinically-relevant ST. In this thesis I show that the African ST313 representative strain D23580 contains 5 full length prophages. Prophage BTP1 and BTP5 are undescribed, novel prophages specific to African ST313 strains, whilst Gifsy-2, ST64B and Gifsy-1 are well-characterised prophages found in other strains of S. Typhimurium. Of the five prophages, only BTP1 and BTP5 showed evidence for functional phage production, and mutations responsible for the inactivation of Gifsy-2, ST64B and Gifsy-1 were identified. The BTP1 prophage spontaneously induced at a prolific rate, estimated to result in the phage-mediated lysis of approximately 0.2% of the lysogenic cell population. A GFP reporter system was developed to visualise the spontaneous induction of the BTP1 prophage at the single-cell level. Though the BTP5 phage could not be studied using traditional plaque assay methodology, there was evidence that the BTP5 prophage was capable of forming viable BTP5 phage that could lysogenise naïve hosts. I analysed the genomes of recently discovered ST313 isolates from the UK and show that ST313 in the UK represents a distinct population of antibiotic susceptible strains associated with gastrointestinal infection. Additionally, analysis of the UK-ST313 genomes indicated that the BTP1 and BTP5 prophages were acquired independently by the two African ST313 lineages, showing convergent evolution to acquire and conserve the BTP1 and BTP5 prophages. Finally I present evidence that the prophages, in particular BTP5, effect the global gene expression of ST313, and the ST313-td gene of BTP1 mediates lysogenic conversion by functioning as a superinfection immunity factor against infection by Salmonella phage P22. The implications of these findings for understanding the pathogen in terms of ecological niche, host range and invasiveness in humans is discussed.
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Makendi, Njock E. C. "The phylogenetic and phenotypic analysis of Salmonella enterica serovar Weltevreden." Thesis, London School of Hygiene and Tropical Medicine (University of London), 2016. http://researchonline.lshtm.ac.uk/2550034/.
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Diarrhoeal diseases remain a global health threat and are responsible for high levels of morbidity and mortality worldwide, with an estimated 1.7 billion cases every annum. Additionally, according to the World Health Organisation, diarrhoeal diseases are the second leading cause of death in children under 5 years old. Salmonella are one of the most common diarrhoeal pathogens [1] (WHO Accessed 20 February 2015) with serovars Enteritidis, Typhimurium and Typhi playing a major role in outbreaks worldwide. However, Salmonella enterica serovar Weltevreden (S. Weltevreden) has recently attracted a great deal of interest due to increasing reports of its isolation by reference laboratories around the world, with a particular high incidence in South East Asia. However, relatively little is known about the genotypic or phenotypic properties of this understudied serovar. In this study, phylogenetics and comparative genomics based on whole genome sequences were used to define the genetic diversity within a sizeable collection of S. Weltevreden isolates collected from across the globe, with a focus in South East Asia. This phylogenetic analysis confirmed that the S. Weltevreden isolates belong to a monophyletic clade formed of several sub-clades presenting distinct geographical clustering and characteristics. Phenotypic characterisation was performed on selected isolates, with an aim to dissect aspects of host-pathogen interaction during infection, providing a foundation to compare S. Weltevreden with other serovars such as S. Typhimurium. Interestingly, an overall attenuated pathology was observed both invitro (hep 2 cell line) and in-vivo (murine and zebrafish embryos) for S. Weltevreden compared to the S. Typhimurium reference strain. This is the first report of the phylogenetic analyses of S. Weltevreden and of a systematic in-vitro and in-vivo characterisation of the sub-species.
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Robertson, James Stuart. "DNA and protein based vaccines against Salmonella enterica Serovar Typhimurium." Thesis, University of Edinburgh, 2003. http://hdl.handle.net/1842/14307.
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GroEL is a heat shock protein, involved in the folding of denatured proteins under stressed conditions. Flagellin is a structured protein of flagella, and can undergo phase variation so that at any one time, synthesis of flagellin occurs from either of two genes, fliC or fliB. Porins, of which OmpC is an example, allow passage of nutrients and small molecules through the bacterial cell outer membrane. All of these proteins have previously been described as being immunogenic. In the work described here, antibodies specific to each of these antigens were detected in both innately susceptible and resistant mice after infection with an attenuated strain of S. typhimurium, and after subsequent challenge with the virulent organism, suggesting that they may each be involved in protective immunity. The immunogenicity of a vaccine consisting of these four recombinant Salmonella antigens was assessed in the mouse model. GroEL- and flagellin-specific antibodies were detected in innately susceptible mice after immunisation together with the adjuvant DDA. Moreover, analysis of IgG isotypes showed increased titres of both IgG1 and IgG2a, the latter indicating that cell-mediated immunity had been generated. However, challenge of immunised mice with virulent S. typhimurium demonstrated that the protective efficacy of the vaccine was of only low-level. Immunisation of innately susceptible and resistant mice using a tetravalent DNA vaccine expressing all four antigens resulted in increased antibody against GroEL, FliC and FlijB in both mouse strains. As with the subunit vaccine, IgG isotype analysis showed increased titres of both IgG1 and IgG2 indicting that both Type 2 and Type 1 helper T-cell responses had been elicited. However, considerable variation was observed within immunised mouse groups and so the protective efficacy of the vaccine was not determined.
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Nogueira, Letícia Amaral [UNESP]. "Estudo molecular da resistência a bacteriófagos líticos em Salmonella enterica subsp. enterica serovar Enteritidis." Universidade Estadual Paulista (UNESP), 2015. http://hdl.handle.net/11449/144057.
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000868807.pdf: 3365386 bytes, checksum: ea272827c7ffe9fa7ea3385d0c22cf85 (MD5)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
Salmonella Enteritidis é um patógeno pertencente à Família Enterobacteriaceae que frequentemente está relacionada a infecções alimentares em humanos, principalmente em crianças, idosos e pacientes imunossuprimidos. A importância deste micro-organismo se dá pela sua prevalência significativa, com distribuição mundial, nos lotes de frangos de corte e de postura de ovos, levando a sérias implicações na saúde pública e ao mercado avícola. O controle eficaz da salmonelose é um desafio, pois envolve um complexo manejo dos animais e tratamento com uso de antibióticos que não garantem a eliminação da infecção. O uso de bacteriófagos líticos contra infecções bacterianas (fagoterapia) vem atender uma demanda crescente por alternativas ao tratamento antimicrobiano convencional. Todavia, a literatura tem relatado o surgimento de certa resistência bacteriana a alguns bacteriófagos líticos, como por exemplo, em Salmonella sp. Diversos são os mecanismos bacterianos de resistência aos fagos, tais como a prevenção de adsorção, o bloqueio da injeção do DNA do fago, restrição-modificação, infecção abortiva e o sistema CRISPR (clustered regularly interspaced short palindromic repeats) /Cas (CRISPR associated proteins) de imunidade adquirida. Assim sendo, o presente trabalho objetivou a caracterização molecular da resistência bacteriana de fagos líticos isolados contra cepas de Salmonella enterica subsp. enterica serovar Enteritidis (SE). Para tal, os mecanismos-alvos escolhidos foram o sistema CRISPR/Cas e o bloqueio da adsorção de fagos, via lipopolissacarídeo (LPS). Nas 22 cepas de estudo, um total de 72 CRISPRs foi identificado, com 14 diferentes domínios repetitivos (DR), apresentando um total de 551 sequências de espaçadores e os genes associados cas1, cas2 e cas3. Cas 9 e cas 10 não foram identificados. Foi observado que as cepas resistentes aos fagos líticos possuíam um maior número total...
Salmonella Enteritidis is a pathogen that belongs to the Enterobacteriaceae family and is often related to infections transmitted by food in humans, especially in children, elderly and immunosuppressed patients. The importance of this microorganism is because it's significant prevalence with worldwide distribution in lots of poultry, leading to serious implications for public health and poultry industry. Effective control of salmonellosis is a challenging because involves a complex animal handling and treatment with antibiotics that do not guarantee the elimination of infection. The use of lytic bacteriophages against bacterial infections (phage therapy) meets a demand for alternatives to the conventional antimicrobial treatment. However, the literature has reported the emergence of bacterial resistance to some lytic bacteriophages, such as in Salmonella sp. There are several mechanisms of bacterial resistance to phages, such as prevention of adsorption, blocking the injection of phage DNA, restriction-modification, abortive infection and the CRISPR/Cas System acquired immunity. Thus, this research aims to study in a molecular level the bacterial resistance of lytic phages isolated from pathogenic strains of Salmonella enterica subsp. enterica serovar Enteritidis (SE). Two mechanisms have been chosen: the CRISPR/Cas system and blocking of phage adsorption via LPS. From 22 SE strains, a total of 72 CRISPRs was identified with 14 different repetitive domains (DR), presenting a total of 551 spacers' sequences and cas1, cas2 and cas3 CRISPR associated genes. Cas 9 and cas 10 genes weren't identified. It was observed that phage lytic resistant strains have a higher total number of spacers in the CRISPR locus in relation to sensitive strains, and this difference was statistically significant. Phylogenetic analysis of cas genes from CRISPR/Cas system, of rfaC, rfaH, cpsG, manB, manc, lpxA, lpxB and lpxC genes (related to LPS) and of...
FAPESP: 2011/11761-7
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Ogunniyi, Abiodun David. "Functional characterisation of the SefA protein of Salmonella enterica serovar Enteritidis /." Title page, contents and abstract only, 1996. http://web4.library.adelaide.edu.au/theses/09PH/09pho35.pdf.
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Schroeder, Betsy. "Finding Typhoid Mary: Identifying Latent Carriers of Salmonella enterica serovar Typhimurium." Diss., Virginia Tech, 2020. http://hdl.handle.net/10919/99979.
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Salmonella enterica serovar Typhimurium (S. Typhimurium) is an important human pathogen. The Centers for Disease Control and Prevention (CDC) estimates that 1,027,561 people become ill with nontyphoidal Salmonellosis annually, and S. Typhimurium is one of the most common disease causing serovars. Quantification of the true number of cases of salmonellosis is hampered by the presence of a carrier state. These carriers are animals and humans that carry the pathogens for a variable period of time without showing any clinical signs. One of the biggest barriers to controlling and preventing salmonellosis in a population is identification of these carriers. Identifying these latent carriers of chronic infections is vital to preventing such disease transmission and creating avenues for novel control and treatments.
In my dissertation research, we developed a cell culture model to study latent Salmonella infections. By activating human monocytes with retinoic acid and vitamin D3, we were able to isolate Salmonella from such cells 45 days after inoculation. We subsequently used this model to identify genes that were upregulated in this chronic infection model. We found that aceA, a gene that codes for isocitrate lyase, is significantly upregulated on days 10 and 30 post infection.
Isocitrate lyase is part of the glyloxylate cycle. Some bacterial species have developed a mechanism to utilize acetone as a carbon source to synthesize tricarboxylic acid (TCA) cycle intermediates. This anaplerotic reaction allows organisms to conserve carbon and use alternative carbon sources. This cycle is one way in which bacteria can adapt and survive in an intracellular environment. This intracellular survival is key to latent infections persisting within a host. It is biologically plausible that, in order to survive in a latent state, S. Typhimurium would up-regulate genes that would facilitate intracellular survival.
After establishing the cell culture model, we tested the hypothesis that aceA is upregulated in latent infections of S. Typhimurium in a mouse model. We orally challenged mice that were resistant to Salmonella infection, collected their feces, and collected tissue specimens at several time points up to 135 days post-challenge. These samples were cultured and tested using quantitative polymerase chain reaction (qPCR). The qPCR results showed that tissue samples from inoculated mice had increased aceA expression 95 days after challenge.
Finally, we examined whether aceA expression could be detected in cattle lymph node samples. Supra-mammary lymph nodes from 40 dairy cattle and mesenteric lymph nodes from 100 culled cattle were sampled and submitted for culture and qPCR. None of the supra-mammary lymph nodes were positive for Salmonella via culture or aceA qPCR; however, 11 mesenteric lymph nodes showed increased aceA expression in qPCR compared to 5 culture positive lymph nodes. Further research is necessary, but these results demonstrate some of the advantages of using genetic primers to identify latent Salmonella infections in clinically normal cattle. In addition, the assay may be able to differentiate between latent and active salmonellosis, and could be used to provide targeted drug delivery.Doctor of Philosophy
Salmonella enterica serovar Typhimurium (S. Typhimurium) is an important human pathogen. Determining the true number of cases of salmonellosis is made more difficult because of the presence of a carrier state. These carriers are animals and humans that carry the pathogens for a variable period of time without showing any clinical signs. Identifying these latent carriers of chronic infections is vital to preventing such disease transmission and creating avenues for novel control and treatments. In my dissertation research, we looked at genetic markers from an offshoot of the TCA cycle, the glyoxylate pathway. We used these markers to test the hypothesis that these glyoxylate pathway genes would be upregulated in latent S. Typhimurium infections. Our research involved developing a cell culture model, then using the results from the cell culture model to inform a mouse model, and then a cattle lymph node diagnostic study. The cell culture model indicated that the gene for isocitrate lyase, aceA, is significantly upregulated compared to housekeeping genes. We found the presence of aceA in chronically infected mice, as well as cattle lymph node samples. Further research is necessary, but these results demonstrate some of the advantages of using genetic primers to identify latent Salmonella infections in clinically normal cattle.
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Lundgren, Hans. "Formation of Thiolated Nucleosides in tRNA in Salmonella enterica serovar typhimurium." Doctoral thesis, Umeå universitet, Molekylärbiologi (Teknat- och Medfak), 2006. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-857.
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The presence and synthesis of transfer RNA (tRNA) is highly conserved in all organisms and a lot of genetic material is dedicated to its synthesis. tRNA contains a large number of modified nucleosides and several diverse functions have been found but much about their function is still unknown. By using a novel frameshifting system to select for tRNA modification mutants, new mutations were isolated and subsequently analyzed. This thesis examines the synthesis and function of a subset of tRNA modifications that have a sulfur (thio) -group as part of the modification. The isc operon encodes for proteins synthesizing iron sulfur centers ([Fe-S]) that are a part of the active site of many key enzymes in the cell and the thiolated nucleosides are dependant on a functional iron sulfur gene (iscS) for their synthesis. By studying thiolated tRNA it is not only possible to learn more about the synthesis of the modifications themselves, but also about the synthesis of [Fe-S] clusters. Based on an analysis of mutations in three of the isc operon genes (iscS, iscU, and iscA), a two-model pathway is proposed for the synthesis of Salmonella enterica Serovar Typhimurium thiolated tRNA modifications. The interactions of IscS with other proteins in the tRNA modification thiolation pathways suggest a more complex sulfur relay than had previously been envisioned. Some of the specificities and the effect of an iscA mutant on the levels of tRNA modifications lead to an examination of the role of IscA in [Fe-S] formation and its importance for tRNA modifications.
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Lundgren, Hans K. "Formation of thiolated nucleosides in tRNA in Salmonella enterica Serovar Typhimurium /." Umeå : Department of Molecular Biology, Umeå University, 2006. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-857.
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Cook, P. J. "Induction of cell death in macrophages by Salmonella enterica serovar Typhimurium." Thesis, University of Cambridge, 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.597921.
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The aim of this thesis was to study the response of macrophages to infection with Salmonella enterica serovar Typhimurium and to determine whether TLR signalling contributes to the induction of cell death in infected macrophages. When macrophages were infected with wild-type S. Typhimurium, cell death occurred within two hours of infection. This was dependent on the bacterial protein SipB and required caspase-1 activity. At two-hours post-infection, little or no cell death was observed in macrophages infected with a sipB-deficient mutant of S. Typhimurium C5. Cell death was triggered by this mutant by 24-hours post-infection. The role of TLR4 in S. Typhimurium-induced cell death was investigated using macrophages from TLR4 knockout mice. At 24 hours, induction of cell death by S. Typhimurium C5-sipB in TLR4-deficient cells was reduced compared to wild-type controls. Activated TLR4 recruits the adapter proteins MyD88, Mal. Trif and Tram. The induction of cell death in macrophages deficient for each of these adapters was studied. Preliminary data suggest that signalling through Trif and Tram contributes to the induction of cell death in infected macrophages. MAPK inhibitors were used to investigate the role of MAPK signalling in S. Typhimurium-induced cell death pathways. Specific inhibition of p38 MAPK by SB203580 increased the level of cytotoxicity in infected macrophages. However, PD98059, which inhibits activation of p38, ERK and JNK, had no effect on cell death suggesting that MAPK proteins are involved in both pro-survival and pro-cell death pathways. Inhibition of p38 signalling alone interferes with survival pathways and tilts the balance towards cell death.
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Arkenberg, Anke. "Mechanisms of anaerobic nitric oxide detoxification by Salmonella enterica serovar Typhimurium." Thesis, University of East Anglia, 2013. https://ueaeprints.uea.ac.uk/48768/.
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Salmonella is the cause of millions of food- and water-borne infections worldwide. Systemic infection and gastroenteritis are the main diseases and often prove fatal to immunocompromised patients. Key to Salmonella’s pathogenicity is the survival of several components of the innate immune system encountered during infection. Reactive oxygen and nitrogen species (ROS and RNS) are an integral part of this antibacterial defence of the immune system. Exposure to ROS and RNS occurs within phagocytic immune cells such as macrophages, where such generation of radicals is used to combat pathogens. NO is a radical belonging to the group of RNS that damages bacterial DNA and proteins. Detoxification of NO is essential during infection to allow Salmonella to survive and replicate within macrophages. Three enzymes are currently known to help Salmonella to detoxify NO, but their deletion, however, does not eliminate Salmonella’s survival. Therefore, it is predicted that further mechanisms for NO detoxification exist. In this study, the core NO regulon has been identified: Expression of nine genes is significantly increased during endogenous and exogenous NO exposure of S. Typhimurium. Their functions range from carbon starvation, cytochrome oxidation, iron-sulphur repair and NO reduction to putative proteins with unknown function, some of which contain domains for tellurite resistance. Single and combination deletion strains have shown that these genes are important to decrease anaerobic NO sensitivity of S. Typhimurium and for intracellular survival in murine macrophages. Furthermore, we have shown for the first time that the core NO regulon also provides protection against tellurite. Tellurite is toxic and requires detoxification when encountered. Reducing tellurite to yield the elemental tellurium results in the release of ROS, which then need to be detoxified further. Deletion strains sensitive to tellurite have also shown increased sensitivity to NO. Concurrently, tellurite resistance genes also facilitate the defence against NO.
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Zeiner, Sarah Ann. "Type 1 fimbrial structure and regulation in Salmonella enterica serovar Typhimurium." Thesis, University of Iowa, 2012. https://ir.uiowa.edu/etd/3021.
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Salmonella enterica serovar Typhimurium is a common cause of bacterial food poisoning, and S. Typhimurium expresses type 1 fimbriae that enable the bacteria to bind to eukaryotic cells. Fimbrial proteins are encoded by the fim gene cluster (fimAICDHFZYW). The structural components of the fimbriae are FimA (major subunit), FimI, FimH (adhesin), and FimF (adaptor). In order to determine the genes required for fimbrial assembly in S. Typhimurium SL1344, mutations in fimA, fimI, fimH, and fimF were constructed and examined for their ability to produce fimbriae. While SL1344δfimI was able to assemble fimbriae, SL1344δfimA, δfimH, and δfimF were afimbriate, indicating that fimA, fimH, and fimF are each required for fimbrial formation in S. Typhimurium. These results suggest differences in the genetic requirements when comparing S. Typhimurium type 1 fimbrial and E. coli type 1 and Pap fimbrial systems. S. Typhimurium fim gene regulation was also examined. FimZ and FimY are positive regulators of fimbrial gene expression, and FimW is a negative regulator. FimZ is closely related to the family of response regulators of two-component systems. The response regulator activity of FimZ was examined by substituting the conserved aspartate-56 residue, the putative site of phosphorylation, with alanine, to generate an inactive phosphorylation site, or glutamate, to mimic a phosphorylated protein. Resulting strains were examined for fimbrial production and gene expression. It was observed that when the aspartate-56 is substituted with alanine fimbriae are not produced and when glutamate replaces aspartate-56 fimbriae are produced constitutively. Bacterial two-hybrid assays were also carried out to determine the effect of FimZ aspartate-56 substitutions on the previously described FimZ/FimW protein interaction. It was found that FimZD56A is unable to interact with FimW and FimZD56E is able to interact with FimW. Additionally, complementation studies were used to examine the roles of FimZ and FimY in relation to each other and results suggest that FimY acts upstream of FimZ.
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Shah, Jigna D. "Stress Response And Pathogenesis of Salmonella enterica serovar Typhimurium." DigitalCommons@USU, 2011. https://digitalcommons.usu.edu/etd/900.
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Salmonella is a food-borne pathogen that leads to substantial illness worldwide. The clinical syndromes associated with Salmonella infection are enteric (typhoid) fever and gastroenteritis, in healthy humans. Typhoid fever is caused by host-adapted S. Typhi and S. Paratyphi. Gastroenteritis is caused by serovars usually referred to as non typhoidal Salmonellae (NTS). In recent years, an increasing number of outbreaks due to NTS, despite increased efforts in food safety, were reported because of persistence of Salmonella in the food chain. Thus I hypothesized that Salmonella is able to withstand stresses in the environment and treatments used during food processing for its elimination and thereby able to develop resistance against subsequent stress encounters. The effect of cold, peroxide, and acid was tested on survival of S. Typhimurium and the survival was persistent under cold stress (5°C) for up to 240 h. Pre-adaptation to cold stress (5°C, 5 h) also increased survival of S. Typhimurium during subsequent exposure to acid stress (pH 4.0, 90 min) by repressing hydroxyl radical formation. Cold stress (5°C, 48 h) to S. Typhimurium significantly (p < 0.05) increased its adhesion and invasion in intestinal iv epithelial cells. This phenotype was attributed to a pair of protein-protein interactorsacting as receptors on microbial (STM2699) and host cell surface (SPTAN1). Cold stress significantly (q < 0.05) induced STM2699 in S. Typhimurium and SPTAN1 was significantly (q < 0.05) induced in pithelial cells upon infection with cold-stressed S. Typhimurium. Cold stress to S. Typhimurium also significantly (q < 0.05) induced genes related to virulence such as type 3 secretion system apparatus and effectors genes, prophage genes, and plasmid genes and they remain induced upon infection of epithelial cells with additional induction of spv genes on the plasmid. Infection of epithelial cells with cold-stressed S. Typhimurium significantly (p < 0.05) increased activation of caspase 9 and 3/7. Cold-stressed S. Typhimurium switched metabolism from aerobic respiration to fermentation and it persisted during infection of epithelial cells. As a result, short chain fatty acids formate and acetate, which act as diffusible signal for invasion, were detected in significantly (q < 0.05) high amounts in extracellular media of cells infected with cold-stressed S. Typhimurium supporting the phenotype of high adhesion and invasion of cold-stressed S. Typhimurium in epithelial cells.
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Gilberthorpe, Nicola J. "The Complexities of Nitric Oxide Metabolism in Salmonella enterica serovar Typhimurium." Thesis, University of Sheffield, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.489661.
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S. typhimurium is able to survive and proliferate within macrophage cells and therefore possesses an array of defence mechanisms that allow it to resist antimicrobial stresses such as harsh nitrosative, oxidative and pH conditions. The relationship between S. typ!Iimurium and nitric oxide (NO) wa~ investigated. The resistance mechanisms that S. typhimurill11l employs to avoid nitrosative stress and the ability ofS. typ!Iimuriu11l to produce NO under certain conditions were examined. The regulation of the gene encoding flavohaemoglobin (!Imp) was studied in detail and NsrR was clearly demonstrated to have a role in the regulation of hmp in S. typ!Iimllriwn. Several other genes (ygbA, ytfE, hep, her) were also identified as being repressed by NsrR in non-nitrosating conditions. Construction and use of an nsrR lllnp double mutant demonstrated that an NsrR regulated gene(s) has the capacity to protect aerobic respiration from the inhibitory effects of NO. The requirement of functional Hmp intracellularly was also assessed; !Inlp mutants were shown to be attenuated only in IFN-r-activated macrophages, where the level ofNO produced by the macrophages was enhanced. Mutants in nsrR were also attenuated in IFN-r-activated macrophages suggesting that over-expression of !Imp is detrimental to the cells, . presumably due to production of O2- by high levels of Hmp. This demonstrates the importance of regulatory systems governing the expression of !Imp. Mutants in the iron responsive regulator, Fur, were attenuated in their ability to survive and proliferate in both nonstimulated and IFN-r-activated 1774.2 macrophages and it is suggested that these mutants are hyper-sensitive to a stress other than nitrosative, most probably oxidative stress. The ability of S, typltimurium to produce NO was investigated. The N03reductase (NarGHI) was identified as having the ability to produce NO from N02- in anaerobic conditions, in the absence of its preferred substrate, N03.
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Chen, Peng. "Function of wobble nucleoside modifications in tRNAs of Salmonella enterica Serovar Typhimurium." Doctoral thesis, Umeå universitet, Molekylärbiologi (Teknat- och Medfak), 2004. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-328.
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Transfer RNA from all organisms has modified nucleosides and position 34 (the wobble position) is one of the most extensively modified positions. Some wobble nucleoside modifications restrict codon choice (e.g. 5-methylaminomethyl-2-thiouridine, mnm5s2U) while some extend the decoding capacity (e.g. uridine-5-oxyacetic acid, cmo5U). In this thesis the influence of wobble nucleoside modification on cell physiology and translation efficiency and accuracy is described. A mutant proL tRNA (proL207) was isolated that had an unmodified adenosine in the wobble position. Surprisingly, the proL207 mutant grows normally and is efficiently selected at the non-complementary CCC codon. The explanation of how an A34 containing tRNA can read CCC codon could be that a protonated A can form a base pair with C. cmo5U (uridine-5-oxyacetic acid) is present in the wobble position of five tRNA species in S.enterica. Two genes (cmoA and cmoB) have been identified that are involved in the synthetic pathway of cmo5U. Mutants were constructed in alanine, valine, proline, and threonine codon boxes which left only a cmo5U containing tRNA present in the cell. The influence of cmo5U on growth or on A site selection rates of the ternary complex was found to be tRNA dependent. During the study of the frameshift suppressor sufY of the hisC3737 frameshift mutation, a dominant mutation was found in YbbB protein, a selenouridine synthetase. The frameshifting occurs at CCC-CAA codon contexts and is specific for CAA codons, which are read by tRNAGlncmnm5s2UUG . The sufY204 mutation is a dominant mutation resulting in a change from Gly67 to Glu67 in the YbbB protein, and mediates the synthesis of several novel modified nucleosides/nucleotides (UKs) with unknown structure. The synthesis of these UKs is connected to the synthesis of cmnm5s2U34. The presence of UK on tRNAGlnU*UG reduced aminoacylation and therefore might account for the slow entry at CAA codons which could result in +1 frameshifting by P site tRNA. The selenourdine synthetase activity is not required for the synthesis of UKs. We hypothesize that an intrinsic activity that is low in the wild type protein has been elevated by the single amino acid substitution and results in the synthesis of UKs.
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Müller, Andreas. "Regulation der Typ III Sekretion bei Salmonella enterica serovar Typhimurium im Zellkulturmodell /." Zürich : ETH, Eidgenössische Technische Hochschule Zürich, Institut für Mikrobiologie, 2004. http://e-collection.ethbib.ethz.ch/show?type=dipl&nr=181.
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Ilg, Karin Christine. "Glycoengineering and glycomimicry : Campylobacter jejuni carbohydrate structures on Salmonella enterica serovar typhimurium /." [S.l.] : [s.n.], 2009. http://e-collection.ethbib.ethz.ch/show?type=diss&nr=18524.
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El, Mouali Benomar Youssef. "CRP-cAMP mediated silencing of virulence expression in Salmonella enterica serovar Typhimurium." Doctoral thesis, Universitat de Barcelona, 2017. http://hdl.handle.net/10803/456373.
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The regulation of the expression of virulence genes in Salmonella enterica serovar Typhimurium is an intensively studied feature of Salmonella lifestyle. How Salmonella integrates environmental signals to activate virulence-related genes within the host has been explored. The genes encoded in Salmonella pathogenicity island I (SPI-1) required for the invasion of epithelial cells are well characterized, and many regulators involved in the activation of SPI-1 genes have been described. However, little is known about mechanism involved in the repression of SPI-1 under conditions were Salmonella does not require the expression of virulence-related genes. The expression of SPI-1 encoded genes have been reported to be a burden for Salmonella physiology and therefore mechanism to control shut down of SPI-1 under non permissive conditions might play a crucial role in Salmonella physiology. Here we describe that at exponential phase, CRP-cAMP, which acts as an activator at stationary phase, represses the expression of SPI-1 genes. The overall objective of this thesis was to characterize how CRP-cAMP silences SPI-1 expression under non-permissive conditions and to describe the molecular mechanism behind this phenotypic observation. In this thesis we i) define the target gene for the CRP-mediated regulation of SPI-1 ii) elucidate at which level of regulation CRP-cAMP modulates the expression of the SPI-1 genes master regulator hilD iii) characterize the involvement of CRP-cAMP dependent sRNA in hilD regulation iv) characterize the interaction of the CRP-cAMP dependent sRNA Spot 42 with the hilD 3’UTR region to regulate SPI-1 and v) explore the role of CRP-cAMP in the modulation of the SPI-1 repressor CsrA through the regulation of the long non-coding RNA csrB and csrC.
CRP-cAMP represses hilA expression at exponential phase (non-permissive conditions for SPI-1 expression) and acts as an activator at stationary phase (permissive conditions for SPI-1 expression). CRP-cAMP mediated repression of hilA causes a concomitant attenuation in the expression level of SPI-1 encoded effector proteins. The regulation of SPI-1 during logarithmic growth phase occurs upstream of HilA by repressing hilD, hilC and rtsA expression and is mediated by the regulation of hilD expression at the post transcriptional level through the hilD 3’UTR. CRP-cAMP mediated regulation of hilD requires, in addition to the hilD 3’UTR, the sRNA chaperone Hfq and the major endonuclease RNAse E.
CRP-cAMP represses the expression of the sRNA Spot 42 at exponential phase. We show that Spot 42 positively regulates hilD expression at exponential growth phase and requires of the presence of the hilD 3’UTR, the sRNA chaperone Hfq and the major endonuclease RNAse E. Interestingly, Spot 42 and the hilD 3’UTR region physically bind to Hfq. Spot 42 physically interacts with the last 150 nt of the hilD 3’UTR and unstructured region III of Spot 42 is required for the regulation of hilD.
CRP-cAMP represses csrC but not csrB expression at exponential phase to regulate the expression of hilD. Remarkably, the CRP-cAMP dependent sRNA Spot 42 positively regulates the expression of csrC.La regulación de la expresión de genes de virulencia en Salmonella enterica serovar Typhimurium es una característica intensamente estudiada en Salmonella. La forma en que Salmonella integra señales ambientales para activar los genes relacionados con la virulencia dentro del huésped ha sido explorada. Los genes codificados en la isla de patogenicidad I de Salmonella (SPI-1) son necesarios para la invasión de células epiteliales, están bien caracterizados y se han descrito muchos reguladores implicados en su activación. Sin embargo, poco se sabe sobre el mecanismo implicado en la represión de SPI-1 en condiciones en las que Salmonella no requiere la expresión de genes relacionados con la virulencia. Es sabido que la expresión de genes codificados por SPI-1 son una carga para la fisiología de Salmonella y por lo tanto el mecanismo para controlar la represión de SPI-1 en condiciones no permisivas podría desempeñar un papel crucial en la fisiología de Salmonella. Aquí se describe que en fase exponencial, CRP-cAMP, que actúa como un activador en fase estacionaria, reprime la expresión de los genes SPI-1. El objetivo general de esta tesis fue caracterizar cómo el CRP-cAMP silencia la expresión de SPI-1 en condiciones no permisivas y describir el mecanismo molecular detrás de esta observación fenotípica. El CRP-cAMP reprime la expresión de hilA en la fase exponencial (condiciones no permisivas para la expresión de SPI-1) y actúa como un activador en fase estacionaria (condiciones permisivas para la expresión de SPI-1). La represión mediada por CRP-cAMP de hilA provoca una atenuación concomitante en el nivel de expresión de proteínas efectoras codificadas por SPI-1. La regulación de SPI-1 durante la fase de crecimiento logarítmico se produce aguas arriba de HilA mediante la represión hilD, hilC y rtsA expresión y está mediada por la regulación de hilD a nivel post transcripcional a través de la hilD 3'UTR. La regulación mediada por CRP-cAMP de hilD requiere, además de la hilD 3'UTR, la chaperona Hfq y la endonucleasa RNAsa E. CRP-cAMP reprime la expresión del sRNA Spot 42 en la fase exponencial. Mostramos que Spot 42 regula positivamente la expresión hilD en la fase de crecimiento exponencial, Spot 42 interacciona físicamente con los últimos 150 nt de la hilD 3'UTR.
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Turcot, Isabelle. "Identification and characterization of the Salmonella enterica serovar Typhimurium disulfide oxidoreductase DsbA." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp04/mq22410.pdf.
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Howell, Gillian Morag. "Proteomic analysis of the oxidative stress response of Salmonella enterica serovar Typhimurium." Thesis, University of Southampton, 2004. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.402421.
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Shrimpton, Sarah Elaine. "Antibiotic persistence in Salmonella enterica serovar Typhimurium : involvement of the CspA paralogues." Thesis, University of Edinburgh, 2011. http://hdl.handle.net/1842/9629.
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Chronic infections are often attributed to bacterial biofilms. These biofilms are extremely tolerant to antimicrobial treatment due to the presence of dormant persister cells. Whilst a number of persister genes and pathways have been identified, it is likely that others remain. Investigating persistence of S. Typhimurium was therefore undertaken. A csp null mutant of Salmonella enterica sv. Typhimurium, lacking all six cold shock protein (CspA) paralogues was previously constructed (Hutchinson 2005). At 10°C, this strain is unable to divide, but remains viable for several weeks. However it remains capable of growth at 37°C and thus is conditionally dormant. Using this strain, the link between dormancy and persistence was investigated. Treatment of stationary phase planktonic cultures with fluoroquinolones revealed persister cells in SL1344. In contrast the csp null mutant was completely eliminated by treatment at 37°C; this could be prevented by cspC or cspE expression, implicating a role for cspA paralogues in persistence. Cold shock (10°C) substantially increased persister levels, although csp null cultures remained hypersensitive. Chloramphenicol pre-treatment also reduced elimination of the csp null mutant, linking translation with the persister phenotype. Mutations in 5 genes affecting chromosomal structure and function were investigated, 3 of which (hns, hfq, rpoS) were found to reduce persister levels, suggesting a possible role for DNA supercoiling in persistence. Plasmid topologies in the csp null mutant were highly supercoiled compared to SL1344, a phenotype prevented by cspC or cspE expression. Altered supercoiling is therefore proposed as a mechanism for fluoroquinolone sensitivity in the csp null mutant. Persister levels were also characterised in biofilms of SL1344 and the csp null mutant. In contrast to stationary phase planktonic cultures, the CspA paralogues did not appear to play a role in biofilm persistence under the experimental conditions tested. However, the study revealed a novel role for CspA paralogues in pellicle formation at the air-liquid interface. A plasmid library was used to identify chromosomal regions capable of rescuing the planktonic persister phenotype of the csp null mutant. One region which delayed fluoroquinolone elimination of the csp null mutant, contained components of the hpa gene cluster, replicated in 11 isolates. This locus is involved in hydroxyphenylacetate (HPA) catabolism, indicating a possible role of cellular metabolism in the persistence. Overall this study has revealed novel information about antibiotic persistence in S. Typhimurium and the involvement of the CspA paralogues. These results provide an important foundation for further investigations and contribute towards knowledge of the complex processes of dormancy, persistence and biofilm formation in bacteria.
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Johnson, Sile Ann. "The local tumour immune response following systemic Salmonella enterica serovar Typhimurium infection." Thesis, University of Glasgow, 2018. http://theses.gla.ac.uk/8982/.
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Mmolawa, Princess Tlou. "Molecular analysis of temperate phages in Salmonella enterica serovar Typhimurium DT 64 isolated in Australia." Title page, contents and summary only, 2001. http://web4.library.adelaide.edu.au/theses/09PH/09phm6855.pdf.
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Files on accompanying CD-ROM: Appendix III Phages ST64T and ST64B sequences, are in rtf format. Bibliography: leaves 279-324. System requirements for accompanying CD-ROM: IBM or compatible ; Microsoft Word or compatible to read rtf files.
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Cochrane, Brett C. "A proteomic analysis of the osmotic shock response in 'Salmonella enterica' serovar Typhimurium." Thesis, University of Southampton, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.423378.
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Perez, Gerardo Legrama. "Transport of phage P22 DNA into the cytoplasm of salmonella enterica serovar Typhimurium." Diss., [La Jolla] : [San Diego] : University of California, San Diego ; San Diego State University, 2008. http://wwwlib.umi.com/cr/ucsd/fullcit?p3319845.
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Thesis (Ph. D.)--University of California, San Diego and San Diego State University, 2008.Title from first page of PDF file (viewed Sept. 9, 2008). Available via ProQuest Digital Dissertations. Vita. Includes bibliographical references (p. 169-187).
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Martel, Alexandre. "Caractérisation de la position 48 du récepteur phoQ de Salmonella enterica serovar typhimurium." Sherbrooke : Université de Sherbrooke, 2001.
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Northen, Helen. "An investigation of mutants of the atp operon in Salmonella enterica serovar Typhimurium." Thesis, University of Cambridge, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.608857.
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Ragupathy, Roobinidevi. "Characterisation of the roles of SstR and SstA in Salmonella enterica serovar Typhimurium." Thesis, University of Manchester, 2017. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.719313.
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Salmonella enterica is an important cause of food poisoning and is responsible for approximately a billion human infections each year. Disease manifestation in humans varies from severe systemic enteric (typhoid) fever to self-limiting gastroenteritis depending upon the infecting S. enterica serovar. S. Typhimurium is responsible for acute gastroenteritis in humans but causes a typhoid-like disease in mice and thus serves as an important model for studying the pathogenesis of systemic salmonellosis. Following ingestion, S. Typhimurium employs a variety of virulence mechanisms to survive within its host and establishes infection in the intestinal tract by invading the epithelial cells. Recent studies have revealed the importance of sulfur compounds in the intestine, such as tetrathionate and thiosulfate for the disease progression. S. Typhimurium is capable of utilising these sulfur compounds as terminal electron acceptors for its anaerobic respiration and thus gains a growth advantage over host microbiota during infection. However, the regulation of sulfur availability within S. Typhimurium and the mechanisms involved in mitigating cellular sulfide toxicity are not well-defined. During this study, we have identified the sstRA operon in S. Typhimurium encoding a deduced SmtB/ArsR family of transcriptional regulatory protein (SstR) and a deduced rhodanese-family sulfurtransferase (SstA) and demonstrated a role in mitigating the effects of cellular sulfide toxicity. SstR has been confirmed to act as a transcriptional repressor from the sstRA operator-promoter and the SstR-dependent repression is alleviated by low pH and sulfide stress (sodium thiosulfate), consistent with a role for SstR in sensing sulfide stress to trigger gene expression. Electrophoretic mobility shift assays confirm binding of purified SstR to the sstRA operator-promoter region. Furthermore, a conserved pair of cysteine residues within SstR was identified to be crucial for alleviating SstR-mediated repression, with the substitution of either cysteine causing constitutive repression. This is consistent with SstR inducer-responsiveness involving a thiol-based redox switch. Importantly, S. Typhimurium mutants lacking the sstRA operon have reduced tolerance to sulfide stress, consistent with the sstRA operon having a role in cellular sulfide detoxification. Work is continuing to further characterise the roles of sstR and sstA in S. Typhimurium on their contributions to infections.
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Perry, Jennifer Jean. "Ozone based treatments for inactivation of Salmonella enterica serovar Enteritidis in shell eggs." The Ohio State University, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=osu1282680734.
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Nguyen, Tina. "Mode of Entry and Survival of Salmonella Enterica Serovar Typhimurium in Trophoblast Cells." Thesis, Université d'Ottawa / University of Ottawa, 2017. http://hdl.handle.net/10393/36032.
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Salmonella enterica species are intracellular bacteria causative agents of gastroenteritis and typhoid fever in humans. Pregnancy poses an increased risk of severe Salmonellosis in many mammalian species contributing to miscarriage and/or maternal illness. Previous studies indicated that Salmonella infection in pregnant mice caused rapid fetal and maternal death due to massive bacterial proliferation in the placenta. However, the susceptibility of human primary trophoblast cells (cTBCs) to Salmonella infection was not known. We hypothesized that human placental trophoblast cells are productively infected and provide a unique intracellular niche that permits uncontrolled Salmonella replication due to an ineffective maternal innate immune response to the virulent bacteria resulting in placental death. Firstly, we observed that S.Tm strains defective in the Salmonella pathogenicity island (SPI)-1 type III secretion system (TTSS) (S.Tm-ΔinvA) were unable to enter epithelial cells, but efficiently infected placental choriocarcinoma cell lines through scavenger receptor-mediated endocytosis. Next, we observed that S.Tm failed to grow vigorously in macrophages, but replicated rapidly within epithelial and placental trophoblast cells. Further examination of intracellular localization of S.Tm indicated that bacteria were arrested in early Rab5 expressing phagosomal vesicles within trophoblast cells, whereas phagosomal maturation progressed steadily in macrophages (with expression of lysosomal-associated membrane protein-1 (LAMP-1) and cathepsin D). Moreover, human primary cTBCs harboring S.Tm underwent rapid death of the cells. Infected cTBCs expressed phosphorylated-receptor-interacting serine/threonine-protein kinase (RIPK)-1 protein and phosphorylated-mixed lineage kinase domain-like (MLKL), suggesting induction of the necroptosis pathway of cell death. Furthermore, specific inhibition of necroptosis rescued S.Tm-induced death of cTBCs. Finally, S.Tm infected trophoblast cells produced interleukin (IL)-10, and signal transducer and activator of transcription (STAT)-3 signalling. This correlated to delayed phagosomal maturation which consequently facilitated intracellular pathogen proliferation. Overall, human trophoblast cells may act as reservoirs for S.Tm survival and may aid dissemination in the pregnant host.
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Martel, Alexandre. "Caractérisation de la position 48 du récepteur phoQ de Salmonella enterica serovar typhimurium." Mémoire, Université de Sherbrooke, 2001. http://savoirs.usherbrooke.ca/handle/11143/3284.
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La première partie de cette étude décrit la mise au point d'un test in vivo . Ce test utilise la luciférase comme gène rapporteur et permet de quantifier l'activité du récepteur PhoQ. Ce test est idéal pour la mesure de l'activité globale et de la régulation par le magnésium des différents mutants du récepteur PhoQ de Salmonella enterica serovar typhymurium obtenus par mutagenèse dirigée. De plus, cette étude démontre que l'ajout d'extension dépassant 8 acides aminés en C-terminal du récepteur PhoQ peut amener une plus grande phosphorylation de PhoP. La seconde partie de cette étude traite de la caractérisation de la position 48. Des études in vivo et in vitro démontrent que la mutation de la thréonine 48 du récepteur PhoQ conduit à la modification de l'équilibre kinase/phosphatase, et affecte le degré de phosphorylation de PhoP. Finalement, la cartographie par formation de ponts disulfures de la région environnant le résidu 48 suggère que ce dernier est positionné vers l'intérieur de l'interface de dimérisation."--Résumé abrégé par UMI
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Botten, James Alfons Desmond. "Role of sefD and sefR in the biogenesis of Salmonella enterica serovar Enteritidis SEF14 fimbriae." Title page, abstract and contents only, 2001. http://web4.library.adelaide.edu.au/theses/09PH/09phb7512.pdf.
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Mardones, Acuña Paula Carolina. "Identificación global de genes de Salmonella enterica serovar Gallinarum requeridos para la colonización sistémica de un hospedero murino." Tesis, Universidad de Chile, 2013. http://www.repositorio.uchile.cl/handle/2250/115384.
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Memoria para optar al Título Profesional de BioquímicoAutorizada por el autor, pero con restricción para ser publicada a texto completo en el Portal de Tesis Electrónicas, hasta diciembre de 2014
Salmonella enterica es un patógeno intracelular Gram negativo capaz de infectar un amplio rango de hospederos. En particular, S. enterica serovar Gallinarum infecta aves de corral causando una enfermedad sistémica que puede provocar la muerte. Una vez que entra al organismo, Salmonella utiliza una serie de factores de virulencia que le permiten sobrevivir al pH ácido del estómago, resistir a sales biliares y péptidos antimicrobiales, invadir y traspasar la barrera epitelial intestinal, sobrevivir y replicarse dentro de macrófagos y diseminarse dentro de los órganos del hospedero, principalmente a nivel del bazo e hígado. Hasta el momento, se desconoce gran parte de los factores de virulencia que utiliza S. Gallinarum para infectar un hospedero. Es por eso que en este trabajo se propuso identificar los genes de S. Gallinarum involucrados en la colonización sistémica en un modelo murino a través de un análisis global de mutantes bajo selección negativa in vivo. Para ello, se utilizó una genoteca de ~48.000 mutantes por inserción del transposón EZ-Tn5
Salmonella is a Gram negative intracelular pathogen able to infect a broad range of hosts. Specifically, S. enterica serovar Gallinarum infects poultry leading to a systemic illness that may cause death. Once Salmonella enters the organism, it uses a variety of virulence factors which allows it to survive in the gastric acid, resist bile salts and antimicrobial peptides, invade and cross the intestinal epithelium, survive and grow within macrophages, and colonize internal organs of the host, mainly spleen and liver. The main virulence factors that S. Gallinarum uses to infect a host remained unknown until know. In this work we proposed to identify genes involved in the systemic colonization of S. Gallinarum in the murine model through a genome-wide screening of mutants under negative selection in vivo. To accomplish this, we used a pool of ~48.000 mutants generated by random insertion of the EZ-Tn5
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Winter, Sebastian. "Untersuchungen zur Rolle des Regulatorproteins TviA bei der Immunevasion von Salmonella enterica Serovar Typhi." Diss., lmu, 2010. http://nbn-resolving.de/urn:nbn:de:bvb:19-108487.
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Sekirov, Inna. "The role of the intestinal microbiota in host susceptibility to Salmonella enterica serovar Typhimurium." Thesis, University of British Columbia, 2009. http://hdl.handle.net/2429/7894.
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Intestinal microbiota comprise microbial communities that reside in the gastrointestinal tract and are critical to normal host physiology. Understanding the microbiota’s role in host response to invading pathogens will further expand our knowledge of host-microbe interactions, as well as foster advances in the design of novel therapeutic and prophylactic methods.
In this dissertation I used clinically relevant doses of antibiotics to disturb the intestinal microbiota balance in a murine infection model. Pre-infection perturbations in the microbiota with two antibiotics resulted in increased mouse susceptibility to Salmonella enterica serovar Typhimurium intestinal colonization, greater post-infection alterations in the microbiota, and more severe intestinal pathology. This demonstrates the importance of a balanced microbiota community in host response to an enteric pathogen.
This infection model also allowed further characterization of the host-pathogen-microbiota interactions during enteric salmonellosis. It was shown that in the presence of high numbers of indigenous microbes S. Typhimurium deficient in Salmonella pathogenicity island 2 (SPI2) is unable to trigger intestinal inflammation, while a SPI1 mutant strain promotes late typhlitis. Additionally, it was demonstrated that pathogen-induced intestinal inflammation does not always translate into extensive alterations to the host microbiota, as inflammation during a SPI1 mutant infection did not promote the same changes in host microbiota composition and numbers as inflammation induced by wild-type S. Typhimurium. Differential neutrophil recruitment by the three S. Typhimurium strains was implicated as one possible agent of microbiota perturbations.
A thorough understanding of the tripartite host-microbiota-pathogen relationship in the progression of the enteric infections is needed to fully appreciate the disease process, as well as to suggest new avenues through which to interfere with the infection progression. These studies enhance our understanding of the microbiota’s role in the progression of S. Typhimurium infection and the effects of inflammation upon the microbiota, thus broadening our knowledge of S. Typhimurium pathogenesis and associated host response.
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Panosa, Borràs Anaïs. "Ribonucleotidil Reductases de Salmonella enterica serovar Typhimurium: Regulació Transcripcional i Participació en la Patogènesi." Doctoral thesis, Universitat Autònoma de Barcelona, 2009. http://hdl.handle.net/10803/3925.
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Salmonella enterica serovar Typhimurium (S. Typhimurium) és un patògen intracel·lular gram negatiu que provoca gastroenteritis en humans però que en ratolins provoca una infecció sistèmica similar a la febre tifoidea humana (provocada en aquest cas per S. Typhi). Una de les característiques principals de la infecció per S. enterica és la capacitat que presenta per envair activament cèl·lules epitelials i sobreviure i proliferar a l'interior dels macròfags.S. Typhimurium presenta codificades en el seu genoma tres tipus de ribonucleotidil reductases (RNRs): classe Ia, Ib i III. Les RNRs són enzims essencials ja que són els responsables de la síntesi de novo dels desoxirribonucleòtids (dNTPs), necessaris per a la síntesi i reparació del DNA, a partir de la reducció dels ribonucleòtids (NTPs). Fins ara s'han descrit tres classes de RNRs que es diferencien pel mecanisme de generació del radical que utilitzen, l'estructura que presenten, la seva regulació al·lostèrica i la seva dependència de l'oxigen. Tot i així, totes tenen en comú el mecanisme de reacció i la utilització d'un radical lliure orgànic per a iniciar la catàlisi.
En aquest treball s'ha estudiat la regulació transcripcional de les tres classes de RNRs presents en S. Typhimurium, centrant-nos en dos reguladors: NrdR i Fur. S'ha estudiat l'efecte de la deleció de NrdR sobre l'expressió gènica de les tres classes de RNRs. NrdR és un repressor de les tres classes de RNRs i presenta un major efecte sobre l'expressió de l'operó nrdHIEF. També s'han descrit les seves caixes d'unió i s'han obtingut proteïnes mutants en determinats residus que afecten la funcionalitat de NrdR in vivo, possiblement degut a la participació d'aquests residus en la unió de dATP/ATP.
La mutació de Fur provoca un agument de l'expressió de l'operó nrdHIEF de fins a cinc vegades respecte la soca salvatge. A la regió promotora de l'operó nrdHIEF hi hem detectat una possible caixa d'unió de Fur, la mutació de la qual provoca un augment en l'expressió similar a l'observat en la soca amb Fur delecionat. Mitjançant assajos de retardament electroforètic hem confirmat la unió de Fur a la regió promotora de l'operó nrdHIEF.
En condicions normals, S. Typhimurium utilitza la RNR de la classe Ia per a la síntesi de novo de desoxirribonucleòtids en presència d'oxígen. La classe Ia és essencial per al seu creixement i la classe Ib no és capaç de complementar-ne la seva mutació a no ser que se li introdueixi una còpia extra. La presència de dues RNRs amb activitats redundants en un mateix microorganisme, el fet que en altres espècies bacterianes la síntesi de desoxirribonucleòtids en presència d'oxigen la dugui a terme la classe Ib, i que tant en E. coli com S. Typhimurium la classe Ib s'hagi conservat al llarg de l'evolució fa pensar que aquesta classe de RNR ha d'expressar-se en algunes condicions molt concretes de creixement.
En aquest treball s'ha estudiat la participació de les RNRs en la virulència de S. Typhimurium SL1344 mitjançant la construcció de mutants de cada una de les tres classes així com de dobles mutants i mutants en els seus reguladors (Fur, NrdR). Mitjançant assajos d'infecció de línies cel·lulars de macròfags i també de cèl·lules epitelials hem intentat desvetllar quina de les RNRs és la responsable del creixement de S. Typhimurium durant el procés d'infecció. Els resultats obtinguts indiquen que la RNR responsable de la invasió i de la proliferació a l'interior de macròfags és la classe Ia, mentre que les classes Ib i III no semblen ser essencials. No obstant, observant el comportament de la soca mutant en la classe Ia que presenta la classe Ib sobreexpressada, durant les primeres hores d'infecció (2-6 h) sí que és capaç de sobreviure. Creiem que en aquesta etapa inicial de la infecció, quan es produeix l'explosió respiratoria, es generen unes condicions que activen l'expressió de l'operó nrdHIEF, possiblement la concentració de peròxid d'hidrogen és capaç d'inhibir l'activitat repressora de Fur. La gran demanda de dNTPs a causa del dany al DNA que s'està produint fa necessari un subministrament extra de dNTPs que aportarà la reductasa NrdEF.
Salmonella enterica serovar Typhimurium (S. Typhimurium) is a gram negative intracellular human pathogen causing gastroenteritis in humans as well as a systemic infection in mice similar to human typhoid fever (which is caused by S. Typhi). One of the main features of S. enterica infection is its capacity to actively invade epithelial cells and proliferate inside macrophages.
S. Typhimurium presents three classes of ribonucleotide reductases (RNRs) in its genome: class Ia, class Ib and class III. RNRs are essential enzymes because they carry out the de novo synthesis of deoxyribonucleotides (dNTPs), needed for DNA synthesis and repair, by reducing ribonucleotides (NTPs). Up to date, three different classes of RNRs have been described, differing in their mechanism of radical generation, their three-dimensional structure, allosteric regulation and their oxygen dependence. Nevertheless, they all have in common the mechanism of reaction and the use of an organic free radical to initiate catalysis.
This work has studied the transcriptional regulation of the three RNR classes present in S. Typhimurium, giving importance to two main regulators: NrdR and Fur. We have studied the effects of NrdR deletion in each class of RNR gene expression. Our results indicate that NrdR is a repressor of all three classes, but it shows a stronger repression when regulating nrdHIEF expression. We have also described NrdR recognition sites (NrdR boxes) and we have obtained mutant proteins in some residues that affect NrdR functionality in vivo, possibly due to their participation in dATP/ATP union.
Mutation of Fur causes an upregulation of nrdHIEF expression up to five fold compared to the wild type strain. We have detected a putative Fur recognition sequence within the nrdHIEF promoter region. Mutation of this recognition sequence causes an upshift in nrdHIEF expression similar to the level observed in the fur mutant. Using electrophoretic mobility shift assays (EMSA) we have confirmed that Fur interacts with the nrdHIEF promoter region.
Under normal conditions, S. Typhimurium uses class Ia RNR to de novo synthesize dNTPs in the presence of oxygen. Class Ia is essential for normal growth and class Ib is not able to complement its mutation unless an extra copy of nrdHIEF is introduced. The presence of two RNRs with redundant activities in the same organism, the fact that other bacterial species use class Ib for dNTP synthesis in then presence of oxygen, and that both E. coli and S. Typhimurium have retained class Ib enzymes during evolution, suggest that this class might be expressed under specific growth conditions.
We have studied the role of RNRs in the virulence of S. Typhimurium SL1344 by means of different mutants and double mutants in each RNR class as well as mutants in their transcriptional regulators (Fur, NrdR). Infection assays performed in macrophage cell lines and epithelial cell lines have allowed to elucidate which RNR is responsible for S. Typhimurium growth during infection. Our results indicate that class Ia is the RNR responsible for invasion and proliferation inside macrophages, while class Ib and class III do not seem to be essential. However, class Ia mutants overexpressing class Ib are capable of surviving the first hours of infection (2-6 h). We think that is in this first stage of the infection process (when the oxidative burst takes place), specific conditions generate and activate nrdHIEF expression. It is possible that hydrogen peroxide concentrations are responsible for the inhibition of the Fur repressor activity. The highest demand of dNTPs due to DNA lesions makes it necessary for an extra dNTP supply that will be provided by the NrdEF enzyme.
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Goecke, Michelle Elisa. "A study of the regulation of expression of dsbA from Salmonella enterica serovar Typhimurium." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1998. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp01/MQ28200.pdf.
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