Year: 2012
Type: dissertation
Abstract: Mycobacterium tuberculosis, the causative agent of tuberculosis (TB) has \ninfected one third of the world’s population and continues to claim more lives \nannually than any other infectious disease agent. A significant proportion of \nindividuals carry latent TB infection (LBTI) which is characterized by the absence \nof clinical symptoms and it has been postulated that the tubercle bacilli are in a \ndormant-like state during this type of infection. Resuscitation promoting factors \n(Rpfs) are cell wall hydrolases which cleave glycosidic bonds within the \npeptidoglycan (PG), a mechanism thought to result in reactivation of bacteria \nfrom the state of dormancy. M. tuberculosis encodes five rpf-like homologues \nwhich are collectively dispensable for growth but are required for reactivation \nfrom dormancy in vitro, and for virulence in the mouse model of infection. LTBI \nthus poses a huge threat to the global burden of active disease. The purpose of \nthis study was to further investigate the biological roles of Rpfs by assessing the \neffects of rpf gene deletion in M. smegmatis, a model organism used for TB \nresearch. M. smegmatis encodes four rpf-like genes designated rpfA, rpfB, rpfC \nand rpfE, and deletion mutants that lack one or more of these genes were \nconstructed by allelic exchange mutagenesis. M. smegmatis mutant strains that \nlack either rpfA or rpfB display no significant differences in growth both on solid \nand in liquid medium when compared to wild type. However, loss of rpfA resulted \nin bacterial clumping during stationary-phase growth in broth culture and \nchanges in cell morphology. Moreover; the rpfA rpfB double mutant and its \nderivative strain lacking rpfC, rpfA rpfB rpfC displayed a ca. 2-4 log increase \nin susceptibility to erythromycin and vancomycin. Furthermore, unusual colonial \nmorphologies with reduced serpentine cording and smooth peripheries were \nobserved for these multiple mutants. Cell surface defects and cell distortions \nwere evidenced as wrinkle-textured, bent cells and polar tip bulges for the \nabovementioned mutants. The multiple deletion strains also displayed a defect in \nbiofilm formation revealing an inability for the mutants to form complex cell-cell \ninteractions. Collectively, the data are suggestive of a loss of bacterial cell wall integrity due to rpf gene deletion and it is proposed that rpfA, in combination with \nother genes, is largely responsible for cell wall integrity maintenance. Our data \nindicate that these factors play an important role in cell growth and division and \ntherefore represent an untapped source of novel targets for anti-tubercular drug \ndiscovery.
Author: Lusanda Mapela
Language: English
Subfield: Infectious Diseases
Field: Medicine
Domain: Health Sciences
Open Access status: green
OpenAlex ID: W1597063734