|Institution:||University of Salford, Manchester|
|Department:||School of Environment and Life Sciences|
|Keywords:||Health and Wellbeing|
|Full text PDF:||http://usir.salford.ac.uk/32933/1/Final_copy_of_PhD_thesis_.docx|
Staphylococcus aureus is a major pathogen in both hospital and community settings and it causes infections ranging from mild skin and wound infections to life-threatening systemic illness and, together with the emergence of antibiotic resistance, has been a major cause of morbidity and mortality worldwide. The stringent response, is a stress response that bacteria display to avoid death when subjected to amino-acid starvation. This phenomenon has been observed in different species among Gram positive and Gram negative bacteria but relatively few studies have observed the stress response in Staphylococcus aureus. The stringent response can be triggered by treatment with mupirocin, an antibiotic that mimics amino-acid starvation by inhibiting isoleucyl tRNA synthetase. In this project S. aureus 8325-4 was exposed to sub-inhibitory concentrations of mupirocin (0.5 MIC = 0.25µg/ml-1) to investigate the ability of this concentration to trigger the stringent response. The treatment with mupirocin was continued up to 24 h as previous studies only examined short periods of treatment. Growth was inhibited and the stringent response nucleotide ppGpp was detected 1 h after treatment which slowly decreased in concentration for up to 4 h combined with significant growth inhibition. However, ppGpp could not be detected at 12 or 24 h whereas growth resumed. In addition, the effect of sub-inhibitory concentrations of mupirocin was observed on the TSST-1 producing S. aureus clinical strain (B49). Q-PCR showed up-regulation of tst gene, codes for TSST-1, and its regulator RNAIII transcription up to 8 h of exposure relative to controls, the toxin was not detected by Reverse Passive Latex Assay. Further, RNA-seq analysis was used to observe the global transcriptional alterations caused by the stringent response in S. aureus at 1, 12 and 24 h. From the whole transcriptome profile, differentially expressed genes relative to control as well as from comparisons between treated cell time points were observed concentrating on 60 virulence genes and stress related genes that were significantly increased through stringent response status (1 h). Although ppGpp was not detected at 12 h, cells were still under the influence of the stringent response. However, cell growth had resumed by 24 h which indicates recovery after exposure to sub-lethal concentrations of mupirocin. The effect of the sub-inhibitory concentration of mupirocin on global gene expression in S. aureus is discussed in relation to global control of gene expression and clinical use. In addition, a scenario for S. aureus recovery from stringent response has been suggested here which might open doors for drug target determination in the future.