Bacterial cystitis is overwhelmingly a syndrome of women and the kidney or urologic disease prompting the most visits to physician offices in the US. That the pathogenesis of this disease is poorly understood is ironic. It is the intention of this new project to direct the tools of molecular biology at pertinent E. coli strains in order to examine mechanisms of pathogenesis using human bladder epthelial cells (HBEC) in culture and an in vivo animal model of UTI modified to minimize kidney involvement. The majority of cystitis strains do not have P fimbriae, S fimbriae, hemolysin, and other E. coli characteristics identified as virulence factors for pyelonephritis. Therefore, we have developed an experimental plan that slows the study not only of those genes modestly epidemiologically associated with cystitis but also the identification, by functional roles, of heretofore unknown genes. Preliminary studies demonstrate the E. coli hemolysin, a pore-forming cytotoxin is active against HBEC, that HBEC internalize E. coli, and that mouse UTI virulence is not explained solely by known E. coli virulence factors. We will test the following hypotheses: 1) That strains causing cystitis in humans adhere to HBEC via specific known or as yet unrecognized adhesins. This will be demonstrated by mutagenesis of pertinent strains: isogenic for known adhesins and transposon to identify unknown adhesins. 2) That F54, a potent E. coli cystitis strain with no known virulence factors, causes cystitis in part through internalization into bladder epithelial cells. This will be approached via transposon mutagenesis to seek mutants with diminished internalization and virulence. 3) That the activities of E. coli hemolysin and cytotoxic necrotizing factor (CNF) in the living bacterium can be dissociated by genetic manipulation. Hemolysin is epidemiologically linked to cystitis and CNF is an interesting protein identified in 2/3 of hemolytic strains which enhances bacterial internalization and causes multinucleation of host epithelial cells. Our intent will be to dissociate hemolysin and CNF via isogenic mutagenesis and to study the four permutations of these two putative virulence factors in adherence, internalization, and cytotoxicity assays in order to generate hypotheses of the action and interaction of hemolysin and CNF.

Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Buckles, Eric L; Luterbach, Courtney L; Wang, Xiaolin et al. (2015) Signature-tagged mutagenesis and co-infection studies demonstrate the importance of P fimbriae in a murine model of urinary tract infection. Pathog Dis 73:
Buckles, Eric L; Wang, Xiaolin; Lane, M Chelsea et al. (2009) Role of the K2 capsule in Escherichia coli urinary tract infection and serum resistance. J Infect Dis 199:1689-97
Lane, M Chelsea; Li, Xin; Pearson, Melanie M et al. (2009) Oxygen-limiting conditions enrich for fimbriate cells of uropathogenic Proteus mirabilis and Escherichia coli. J Bacteriol 191:1382-92
Zupancic, Margaret L; Frieman, Matthew; Smith, David et al. (2008) Glycan microarray analysis of Candida glabrata adhesin ligand specificity. Mol Microbiol 68:547-59
Jacobsen, Sandra M; Lane, Mary C; Harro, Jean M et al. (2008) The high-affinity phosphate transporter Pst is a virulence factor for Proteus mirabilis during complicated urinary tract infection. FEMS Immunol Med Microbiol 52:180-93
Ma, Biao; Pan, Shih-Jung; Zupancic, Margaret L et al. (2007) Assimilation of NAD(+) precursors in Candida glabrata. Mol Microbiol 66:14-25
Buckles, Eric L; Wang, Xiaolin; Lockatell, C Virginia et al. (2006) PhoU enhances the ability of extraintestinal pathogenic Escherichia coli strain CFT073 to colonize the murine urinary tract. Microbiology 152:153-60
Castano, Irene; Pan, Shih-Jung; Zupancic, Margaret et al. (2005) Telomere length control and transcriptional regulation of subtelomeric adhesins in Candida glabrata. Mol Microbiol 55:1246-58
Domergue, Renee; Castano, Irene; De Las Penas, Alejandro et al. (2005) Nicotinic acid limitation regulates silencing of Candida adhesins during UTI. Science 308:866-70
Jansen, Angela M; Lockatell, Virginia; Johnson, David E et al. (2004) Mannose-resistant Proteus-like fimbriae are produced by most Proteus mirabilis strains infecting the urinary tract, dictate the in vivo localization of bacteria, and contribute to biofilm formation. Infect Immun 72:7294-305

Showing the most recent 10 out of 31 publications