| Description |
Urinary tract infections (UTIs) afflict millions of individuals yearly, constituting a tremendous global health-care burden. The primary causative agents of UTIs are the gram-negative, rod-shaped bacteria, uropathogenic Escherichia coli (UPEC). These pathogens are motile and adhesive, with a proclivity to colonize diverse niches within the urinary tract; including the kidneys, bladder, and ureters. In the bladder, UPEC grow to high levels and often associate with the superficial epithelial cells lining the lumen. UPEC can invade these superficial epithelial cells to form intracellular reservoir populations, which are thought to be a source of recurrent, or relapsing, infections. The susceptibility of these intracellular UPEC populations was tested using a panel of commonly prescribed antibiotics in a murine model of UTI. Intracellular UPEC were found to persist despite treatment with host cell-permeable antibiotics such as sparfloxacin and ciprofloxacin that effectively sterilize the urine. In a follow-up study, UPEC reservoir populations were more effectively targeted by treating infected bladders with chitosan, a chitin-based bladder exfoliant, prior to sparfloxacin treatment. Although chitosan administration prior to antibiotic treatment significantly decreased UPEC titers, mice still exhibited some relapsing UTIs, suggesting that reservoirs still persist either within the bladder or in other host tissue. To further elucidate mechanisms of bacterial persistence within the urinary tract, several underappreciated bacterial factors were examined that were hypothesized to affect UPEC virulence, stress resistance, and persistence. iv Bacterial, small, non-coding RNAs (sRNAs) are posttranscriptional regulators of gene expression in most prokaryotes and were shown to contribute to a wide variety of UPEC stress response and virulence cascades. In a follow-up study, the putative UPEC sRNA repertoire was defined using RNA-Seq technologies and bioinformatic analyses. Several novel, candidate sRNA molecules were identified and characterized, one of which seemingly repressed UPEC virulence in the murine UTI model. In a second approach to define regulators of UPEC pathogenic behaviors, the tRNA modifying enzyme MiaA was identified as a global regulator of UPEC stress response and virulence. MiaA adds a prenyl group to A-37, adjacent to the anticodon, in a subset of tRNAs to modulate ribosome fidelity and frameshifting. MiaA expression in UPEC was responsive to several environmental stresses and deletion or overexpression of MiaA interferes with the stress resistance and virulence properties of UPEC. Taken together, this thesis defines the robust nature and resilience of intracellular UPEC reservoir populations and delineates sRNAs and MiaA as important regulators of stress resistance and persistence within the host. |
