Tuberculosis kills more people than any other infection. Individuals harboring latent Mycobacterium tuberculosis infection (LTBI) carry a lifetime risk of reactivation to active disease and a better understanding of LTBI is a research priority. Studies to investigate dormant cells of M. tuberculosis when subjected to nutrient limitation, using in vitro and in vivo models to simulate LTBI, are the subject of this thesis. In vitro, extended culture with fatty acids (oleic acid) as the carbon source, leading to non-replicating persistence (NRP) was used to compare bacilli in different growth phases. Transcriptomic analyses revealed that NRP bacilli down-regulated genes involved in lipid biosynthesis, aerobic respiration, TCA (tricarboxylic acid) cycle metabolism and ATP synthases, while genes associated with 3-oxidation of lipids were induced, when compared to exponential phase (EXP) bacilli. The gene expression profiles of bacilli in vitro were compared with those from bacilli isolated from intracellular and in vivo models and the data indicated that lipid metabolism and nutrient depletion may be important during LTBI in humans. Guinea pig models of active and latent M. tuberculosis infection were established and methods of bacterial RNA extraction from infected lung were investigated, with the aim of characterising in vivo derived bacilli using transcriptomics. Technical challenges in obtaining sufficient RNA for transcriptomics could not be overcome using the available technology. The infectivity in guinea pigs, of bacilli from different growth phases generated in vitro was investigated to determine the relationship between the transcriptome and infectivity. Compared with EXP or stationary phase (STAT), NRP bacilli showed a delay in establishing infection but similar bacterial loads and pathology were observed at later time points. In summary, using an in vitro nutrient depletion model, growth phase-specific gene expression signatures and infectivity characteristics of M. tuberculosis were defined which contribute to the understanding of bacilli during latent infection.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:754060 |
| Date | January 2018 |
| Creators | Clark, Simon Oliver |
| Publisher | St George's, University of London |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
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