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Type: Thesis
Title: Coupling stress responses and growth pathways in Haemophilus influenzae.
Author: Jiang, Changde Donald
Issue Date: 2014
School/Discipline: School of Molecular and Biomedical Science
Abstract: Haemophilus influenzae is an obligate human pathogen which requires NAD and heme to grow. Commonly a commensal within the nasopharynx, H. influenzae can be found in both encapsulated and non-encapsulated forms. Encapsulated species, primarily the serotype type b, can be invasive and can cause bacteraemia while the non-encapsulated forms, otherwise known as non-typeable H. influenzae (NTHi), infect the upper respiratory tract, and occasionally causing the recurrence of chronic airway diseases. H. influenzae requires different pathways for bacterial growth as it can move to and cause diseases within diverse sites of the body where the physical and chemical properties are different. The bacteria encounter different reactive chemical stresses that may be produced endogenously (through the bacterial metabolic pathways) and exogenously (host-genearted chemicals) within the different niches. The presence and nature of these reactive chemical stresses impact on their ability to survive, their pathogenesis and the disease outcome within the niches. This therefore highlights a link between the capacity of the bacteria to successfully respond to the environmental stresses and their selection of their own metabolic pathways for energy, biosynthesis and intracellular control of redox balance. In addition, the response to a particular host environment is not only dependent on these stresses, but also on other molecules or biochemicals present in the local environment and the interplay between these factors will highly impact on the bacteria’s colonisation and survival. This interplay is poorly understood and is central to the work presented here. A key example of such interplay is the AdhC system. This thesis represents the work done to characterise AdhC and determine its role in the metabolism of H. influenzae and its links to exogenous stress response and the environmental conditions. The adhC gene from H. influenzae encodes the glutathione-formaldehyde dehydrogenase AdhC which has been found to be essential for defence against the host-generated chemical S-nitrosogluthatione (GSNO; the product of reactive nitric oxide combining with glutathione). Studies have further suggested that AdhC might be required for the growth of the bacteria with oxygen, glucose and iron. adhC and estD are regulated by the redox sensitive NmlR but given the link to these other environmental factors, there is some indication that this pathway is coupled to the global regulatos that sense oxygen, glucose and iron. It was aimed to investigate the stress response regulation connected to environmental oxygen and carbon metabolism in H. influenzae. One of the major environmental stresses in normal growth is the generation of reactive oxygen species (ROS) in the presence of oxygen and FNR is one of the global regulators that regulate genes in response to oxygen changes. Our results showed that FNR regulates at least 279 genes in high and low oxygen tensions. Our results also implied that another global regulator, CRP, is required for utilising mannose and glucose, or the pathways for their transport and metabolism. H. influenzae has been shown to colonise anatomical niches that possess non-optimal conditions by producing biofilms. Therefore, another aspect of this work was to determine the impact of stress responses to biofilm formation in H. influenzae. Our results showed that more biofilms were produced in adhC, nmlR, fnr and crp mutants than in Rd KW20, under both normal aerobic conditions and stressed conditions, suggesting that these genes are essential for normal aerobic growth as well as stress responses. RNA sequencing was further performed on a known biofilm-forming nikQ mutants in order to gain a deeper insight into H. influenzae’s ability to form biofilms. Significant changes in the surface structures in this lifestyle state were found.
Advisor: Kidd, S.
Bent, S.
Dissertation Note: Thesis (Ph.D.) -- University of Adelaide, School of Molecular and Biomedical Science, 2014
Keywords: stress; Haemophilus influenzae
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