? Type 1 pili, the preeminent virulence determinant in Escherichia coli-associated cystitis, promote binding to the bladder surface and facilitate the colonization and invasion of the epithelium. In their intracellular residence, E. coli amass into communal formations with biofilm-like properties termed intracellular bacterial communities (IBC) and then disperse and flux from the infected cell into the lumen of the bladder. Upon fluxing, type 1 pili may again mediate adherence to naive epithelial cells, reinitiating invasion and IBC formation. Because of the central role of type 1 pili in uropathogenesis, understanding the regulation of these fibers is essential for a complete molecular description of the infection and ultimately to identify novel targets for therapeutics. Type 1 pili are regulated by two recombinases, FimB and FimE, that together dynamically modulate the expression of the fibers through phase variation whereby the promoter region is inverted into ON and OFF positions. Preliminary studies have identified FimX, a third site-specific recombinase that, despite the absence of FimB and FimE, produces phase variation and the expression of type 1 pili and supports full virulence. Ablation of FimX and the other two recombinases renders the organism avirulent. We hypothesize that FimX, as regulator of type 1 pili expression, is a central virulence determinant in E. coli urinary tract infections.
In Aim 1 of this proposal, we will use a combination of molecular epidemiology, genetics, and animal modeling to characterize the prevalence of fimX and study the in vivo role of FimX as a regulator of type 1 pili in E. coli cystitis.
In Aim 2, through the use of genetics and biochemistry, we will study the structure-function relationships of FimX and its target DNA sequences. Given the central role of the Fim recombinases in regulating type 1 pili expression, understanding their functions and constraints may yield novel points for treatment and prevention in urinary tract infections. ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Clinical Investigator Award (CIA) (K08)
Project #
5K08DK074443-03
Application #
7447904
Study Section
Diabetes, Endocrinology and Metabolic Diseases B Subcommittee (DDK)
Program Officer
Rankin, Tracy L
Project Start
2006-07-10
Project End
2010-11-09
Budget Start
2008-07-01
Budget End
2010-11-09
Support Year
3
Fiscal Year
2008
Total Cost
$115,196
Indirect Cost
Name
Duke University
Department
Pediatrics
Type
Schools of Medicine
DUNS #
044387793
City
Durham
State
NC
Country
United States
Zip Code
27705
Madden-Fuentes, Ramiro J; Arshad, Mehreen; Ross, Sherry S et al. (2015) Efficacy of Fluoroquinolone/Probiotic Combination Therapy for Recurrent Urinary Tract Infection in Children: A Retrospective Analysis. Clin Ther 37:2143-7
Bateman, Stacey L; Seed, Patrick (2012) Intracellular Macrophage Infections with E. coli under Nitrosative Stress. Bio Protoc 2:
Bateman, Stacey L; Seed, Patrick (2012) Promoter Orientation of Prokaryotic Phase-variable Genes by PCR. Bio Protoc 2:
Bateman, Stacey L; Seed, Patrick C (2012) Epigenetic regulation of the nitrosative stress response and intracellular macrophage survival by extraintestinal pathogenic Escherichia coli. Mol Microbiol 83:908-25
Gawel, Damian; Seed, Patrick C (2011) Urinary tract infection drives genome instability in uropathogenic Escherichia coli and necessitates translesion synthesis DNA polymerase IV for virulence. Virulence 2:222-32
Bateman, Stacey L; Seed, Patrick C (2010) Procession to pediatric bacteremia and sepsis: covert operations and failures in diplomacy. Pediatrics 126:137-50
Goller, Carlos C; Seed, Patrick C (2010) High-throughput identification of chemical inhibitors of E. coli Group 2 capsule biogenesis as anti-virulence agents. PLoS One 5:e11642
Anderson, Gregory G; Goller, Carlos C; Justice, Sheryl et al. (2010) Polysaccharide capsule and sialic acid-mediated regulation promote biofilm-like intracellular bacterial communities during cystitis. Infect Immun 78:963-75
Sena, Arlene C; Seed, Patrick; Nicholson, Brad et al. (2010) Kingella kingae endocarditis and a cluster investigation among daycare attendees. Pediatr Infect Dis J 29:86-8
Cegelski, Lynette; Pinkner, Jerome S; Hammer, Neal D et al. (2009) Small-molecule inhibitors target Escherichia coli amyloid biogenesis and biofilm formation. Nat Chem Biol 5:913-9