Cellular Metabolism and Its Effect on the Type III Secretion System of Dickeya Dadantii 3937

Nutrition, in both eukaryotes and prokaryotes, is vital to the life and well-being of the species. In organisms such as Escherichia coli, metabolism and its regulation have been well established, whereas in Dickeya dadantii 3937, the metabolic pathways and their effects on other processes have not been elucidated. Little is known is how carbon metabolism is able to regulate virulence and pathogenicity in this organism. In this work, we have investigated how the metabolic network contributes to positively and negatively regulating the pathogenicity of Dickeya dadantii 3937. Chapter 1 provides an overview of the history and virulence processes in the organism. In Chapter 2, Dickeya dadantii 3937 was tested in different carbon sources for its ability to induce the type III secretion system (T3SS). Dickeya dadantii 3937 was able to express the T3SS in carbon sources that are upstream of the dihydroxyacetone phosphate/glyceraldehyde-3-phosphate conversion point in the Embden-Meyerhof-Parnas pathway of glycolysis. A mutation was made in the pgi gene, which encodes phosphoglucoisomerase, critical for the conversion of glucose-6-phosphate to fructose-6-phosphate. The pgi mutant had reduced expression of hrpL (the master regulator of T3SS expression), hrpA (the type III secretion pilus), and rpoN (the nitrogen-related sigma factor) genes, but increased expression of gcpA (diguanylate cyclase), in comparison to the wild-type strain. Fructose-6-phosphate supplementation restored T3SS gene expression to near wild-type levels. Complementation of the pgi mutant with the pgi gene restored swimming, swarming, in vivo virulence, and expression of T3SS genes. These results suggest that the metabolic gene pgi, and its intermediate fructose-6-phosphate, may indirectly play a role in the expression of virulence factors in Dickeya dadantii 3937. In Chapter 3, a mutant of Dickeya dadantii 3937was produced by a miniHimar Tn5 RB1 transposon insertion. This mutant was unable to grow on minimal media with fructose. The insertion mapped to a 1,650bp gene product, which sequenced to the fruA gene, which corresponds to the fructose permease, which is the sugar transport component of the fructose phosphotransferase system (PTS). This mutant had increased hrpS, hrpL, and hrpA gene transcription. A fruB mutant showed slightly decreased T3SS activity, but fruB overexpression in the wild-type, and complementation in the fruB mutant showed increased hrpS expression. The FruB protein also interacted with HrpX at the protein level using a yeast two-hybrid assay, showing increased â-galactosidase activity in comparison to empty vector control, HrpX, and FruB alone. This suggests that the fructose PTS plays a part in regulating the T3SS of Dickeya dadantii 3937

Cellular Metabolism and Its Effect on the Type III Secretion System of Dickeya Dadantii 3937

Nutrition, in both eukaryotes and prokaryotes, is vital to the life and well-being of the species. In organisms such as Escherichia coli, metabolism and its regulation have been well established, whereas in Dickeya dadantii 3937, the metabolic pathways and their effects on other processes have not been elucidated. Little is known is how carbon metabolism is able to regulate virulence and pathogenicity in this organism. In this work, we have investigated how the metabolic network contributes to positively and negatively regulating the pathogenicity of Dickeya dadantii 3937. Chapter 1 provides an overview of the history and virulence processes in the organism. In Chapter 2, Dickeya dadantii 3937 was tested in different carbon sources for its ability to induce the type III secretion system (T3SS). Dickeya dadantii 3937 was able to express the T3SS in carbon sources that are upstream of the dihydroxyacetone phosphate/glyceraldehyde-3-phosphate conversion point in the Embden-Meyerhof-Parnas pathway of glycolysis. A mutation was made in the pgi gene, which encodes phosphoglucoisomerase, critical for the conversion of glucose-6-phosphate to fructose-6-phosphate. The pgi mutant had reduced expression of hrpL (the master regulator of T3SS expression), hrpA (the type III secretion pilus), and rpoN (the nitrogen-related sigma factor) genes, but increased expression of gcpA (diguanylate cyclase), in comparison to the wild-type strain. Fructose-6-phosphate supplementation restored T3SS gene expression to near wild-type levels. Complementation of the pgi mutant with the pgi gene restored swimming, swarming, in vivo virulence, and expression of T3SS genes. These results suggest that the metabolic gene pgi, and its intermediate fructose-6-phosphate, may indirectly play a role in the expression of virulence factors in Dickeya dadantii 3937. In Chapter 3, a mutant of Dickeya dadantii 3937was produced by a miniHimar Tn5 RB1 transposon insertion. This mutant was unable to grow on minimal media with fructose. The insertion mapped to a 1,650bp gene product, which sequenced to the fruA gene, which corresponds to the fructose permease, which is the sugar transport component of the fructose phosphotransferase system (PTS). This mutant had increased hrpS, hrpL, and hrpA gene transcription. A fruB mutant showed slightly decreased T3SS activity, but fruB overexpression in the wild-type, and complementation in the fruB mutant showed increased hrpS expression. The FruB protein also interacted with HrpX at the protein level using a yeast two-hybrid assay, showing increased â-galactosidase activity in comparison to empty vector control, HrpX, and FruB alone. This suggests that the fructose PTS plays a part in regulating the T3SS of Dickeya dadantii 3937

The promises and challenges of pre-exposure prophylaxis as part of the emerging paradigm of combination HIV prevention

Introduction: Towards the end of the twentieth century, significant success was achieved in reducing incidence in several global HIV epidemics through ongoing prevention strategies. However, further progress in risk reduction was uncertain. For one thing, it was clear that social vulnerability had to be addressed, through research on interventions addressing health systems and other structural barriers. As soon as antiretroviral treatment became available, researchers started to conceive that antiretrovirals might play a role in decreasing either susceptibility in uninfected people or infectiousness among people living with HIV. In this paper we focus on the origin, present status, and potential contribution of pre-exposure prophylaxis (PrEP) within the combination HIV prevention framework. Discussion: After a phase of controversy, PrEP efficacy trials took off. By 2015, daily oral PrEP, using tenofovir alone or in combination with emtricitabine, has been proven efficacious, though efficacy seems heavily contingent upon adherence to pill uptake. Initial demonstration projects after release of efficacy results have shown that PrEP can be implemented in real settings and adherence can be high, leading to high effectiveness. Despite its substantial potential, beliefs persist about unfeasibility in real-life settings due to stigma, cost, adherence, and potential risk compensation barriers. Conclusions: The strategic synergy of behavioural change communication, biomedical strategies (including PrEP), and structural programmes is providing the basis for the combination HIV prevention framework. If PrEP is to ever become a key component of that framework, several negative beliefs must be confronted based on emerging evidence; moreover, research gaps regarding PrEP implementation must be filled, and appropriate prioritization strategies must be set up. Those challenges are significant, proportional to the impact that PrEP implementation may have in the global response to HIV

From personhood to citizenship: broadening the conceptual base for dementia practice and research

Personhood has provided a lens for conceptualising dementia practice and research for over ten years. It has afforded the rationale and language for improving care and for raising consciousness about the status of people with dementia, as people, intrinsically worthy of respect. However, because personhood is essentially an apolitical concept concerned with psychosocial issues it may be too limiting. Citizenship provides another possible lens. Citizenship is used in cognate disciplines to promote the status of discriminated groups of people still further, to that of a person with power entitled to the same from life as everyone else. However, as citizenship tends to assume the self-cognizance to exercise rights and responsibilities, it may not be as appropriate for people with severe dementia. Both concepts are problematic then, taking too narrow a view of the human experience. For this field to develop over the next ten years it clearly needs a wider lens that is both inclusive of personhood and citizenship, but which also recognizes the complexities of human experience. This article reviews the relevance of personhood and citizenship for dementia practice and research, and argues for a broader lens that incorporates citizenship and sociological ideas about agency and structure.<br/