Browsing by Author "Phalane, Khutso Gemina"
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- ItemEvaluation of multiple cytokine levels to improve our understanding of protective immune responses against Tuberculosis and to develop novel diagnostic methods(Stellenbosch : Stellenbosch University, 2013-03) Phalane, Khutso Gemina; Walzl, Gerhard; Chegou, Novel N.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Division of Molecular Biology and Human Genetics.ENGLISH ABSTRACT: Important steps towards the global control of Tuberculosis include the improvement of diagnosis, the development of effective vaccines and the identification of correlates of protection/protective immunity to Mycobacterium tuberculosis. This study has of three objectives: 1. To validate the findings of a previous study that showed increased levels of IL-1β and decreased levels of IL-17 in children who are exposed to tuberculosis but remain uninfected compared to those who are exposed/infected and unexposed/uninfected. 2. To define the protective immunological phenotype in children with negative IGRA’s and TST following exposure to Mycobacterium tuberculosis. 3. To evaluate a number of cytokines in both serum and saliva samples of identified tuberculosis cases and controls for their diagnostic potential and to evaluate saliva as a possible new diagnostic sample type. The study designs were as follows: Objectives1, and 2: Children with documented tuberculosis exposure and with Mycobacterium tuberculosis infection as assessed through interferon gamma release assays, children with exposure but no infection and a control group with no exposure nor infection were investigated. These participants were selected according to their exposure and infection phenotypes from a larger TB household contact study that was conducted in communities in Cape Town. Whole blood was stimulated in QuantiFeron tubes overnight and ten cytokines were measured in antigen stimulated and unstimulated supernatants by Luminex multiplex Immunoassay. Differential production of cytokines in the three groups was evaluated. Objective 3. Saliva and serum samples were collected from thirty eight adults with suspected tuberculosis who were recruited from a community health centre in Cape Town, after which the levels of thirty three host markers were evaluated in the samples using the Luminex platform. The main findings of the studies included: 1. Increased levels of IL-1β and decreased levels of IL-17 in children who are tuberculosis exposed but remain uninfected compared to those who are exposed/infected and unexposed/uninfected could not be confirmed. 2. Immune responses other than IFN-γ are different in children with different exposure and infection phenotypes. Higher IL-23 and IL-33 levels in children with tuberculosis exposure without subsequent Mycobacterium tuberculosis infection compared to children with no exposure were shown. 3. In both the tuberculosis cases and controls, the levels of most markers were above the minimum detectable limit in both serum and saliva, but marker levels were not consistently higher in one sample type. The levels of fractalkine , IL-17, IL-6, IL-9, MIP-1β, CRP, VEGF and IL-5 in saliva, and those of IL-6, IL-2, SAP and SAA in serum, were significantly higher in tuberculosis patients, in comparison to the levels obtained in those without active tuberculosis (p<0.05). The area under the ROC curve was ≥ 0.70 for most of these markers, thereby confirming their diagnostic potential for TB disease. The work presented in this thesis has identified markers that may grant an improved understanding on the mechanisms that are associated with protection against Mycobacterium tuberculosis in children. The preliminary results presented show that the identification of host markers in saliva is possible and the utility of saliva for the development of rapid immune-based tests for active tuberculosis is promising.