Project description:Strains of urinary tract associated E. coli both recent isolates and from the ECOR collection and non pathogenic E. coli strains were analyzed. Replicates were performed to establish the reproduciblity, then single experiments were performed there on.
Project description:Mucosal surfaces provide ideal living conditions for the normal flora but paradoxically, they also serve as attack sites for numerous bacterial pathogens that cause extensive morbidity and mortality. Understanding this dichotomy is critical for efforts to selectively target and remove pathogens without disturbing the commensal flora or its protective effects. The complex nature of disease predicts that virulence is multifaceted and that pathogens need multiple virulence factors to initiate tissue attack, disrupt immune homeostasis and create symptoms and pathology. The urinary tract supports ABU; a commensal-like state, which has been shown to prevent super-infection with more virulent strains. To reproduce this protective effect, we have established a protocol to create ABU, by inoculation with the ABU strain E. coli 83972. The therapeutic efficacy and safety of this procedure has been documented in placebo-controlled studies in patients with incomplete bladder voiding. Genome sequencing of E. coli 83972 has revealed a general “loss of virulence” phenotype, which includes fimbrial genes. E. coli 83972 lacks functional P or type 1 fimbriae, due to attenuating point mutations in the papG adhesin gene and a large, inactivating deletion in the fim gene cluster. Both fimbrial types have been proposed to enhance bacterial persistence in the urinary tract. In an attempt to increase the efficiency of E. coli 83972 inoculation and extend its use to include UTI-prone patients with complete bladder voiding, we restored P- and type 1-fimbrial expression and addressed how fimbriae affect the gene expression in inoculated human hosts.
Project description:Uropathogenic Escherichia coli utilize a variety of adherence factors that assist in colonization of the host urinary tract. TosA (type one secretion) is a non-fimbrial adhesin that is predominately expressed during murine urinary tract infection (UTI), binds to kidney epithelial cells, and promotes survival during invasive infections. The tosRCBDAEF operon encodes the secretory machinery necessary for TosA localization to the E. coli cell surface, as well as the transcriptional regulator TosR. TosR binds upstream of the tos operon and, in a concentration dependent manner, either induces or represses tosA expression. TosR is a member of the PapB family of fimbrial regulators that can participate in crosstalk between fimbrial operons. TosR also binds upstream of the pap operon and suppresses PapA production. However, the scope of TosR-mediated crosstalk is understudied and may be underestimated. To quantify the global effects of TosR-mediated regulation on the E. coli CFT073 genome, we induced expression of tosR, collected mRNA, and performed RNA-Seq. These findings show that production of TosR affected the expression of genes involved with adhesins, including P, F1C, and Auf; nitrate/nitrite transport; microcin secretion; and promoted biofilm formation.
Project description:Urinary tract infections (UTIs) constitute a highly relevant model of microbial adaptation, in which the contrasting effects of pathogens and commensals on host tissues are clearly displayed. While virulent Escherichia coli cause severe, potentially life-threatening disease by breaking the inertia of the mucosal barrier and infecting the kidneys, the most common outcome of bacteriuria is an asymptomatic carrier state resembling commensalism at other mucosal sites. It remains unclear if the lack of destructive inflammation merely reflects low virulence or if carrier strains actively inhibit disease associated responses in the host. To address this question, we examined the effects of asymptomatic bacterial carriage on host gene expression. Therapeutic urinary tract inoculation with the prototype ABU strain E. coli 83972 is a safe alternative approach in patients with therapy-resistant recurrent UTI. The strain establishes persistent bacteriuria, protecting patients against super-infection with more virulent strains. Using this protocol, we examined if the establishment of asymptomatic bacterial carriage alters host gene expression. After antibiotic treatment to remove prior infection, patients were inoculated with E. coli 83972 through a catheter. Blood samples were obtained before and 24 h after inoculation.
Project description:Urinary tract infections (UTIs) are a very common bacterial infectious disease in humans, and uropathogenic Escherichia coli (UPEC) are the most frequent cause of UTIs. During infection, UPEC must cope with a variety of stressful conditions in the urinary tract. Here, we demonstrated that the small RNA (sRNA) RyfA of UPEC strains was required for resistance to oxidative and osmotic stresses. Inactivation of ryfA in UPEC strain CFT073 decreased urinary tract colonization in CBA/J mice and the ryfA mutant also had reduced production of type 1 and P fimbriae, which are known to be important for UTI. Transcriptomic analysis of the ryfA mutant showed changes in expression of genes associated with general stress responses, metabolism, biofilm formation and genes coding for cell surface proteins. Furthermore, loss of ryfA also reduced UPEC survival in human macrophages. Thus, ryfA plays a key regulatory role in UPEC adaptation to stress, that contributes to UTI and survival in macrophages.
Project description:Urinary tract infections (UTIs) constitute a highly relevant model of microbial adaptation, in which the contrasting effects of pathogens and commensals on host tissues are clearly displayed. While virulent Escherichia coli cause severe, potentially life-threatening disease by breaking the inertia of the mucosal barrier and infecting the kidneys, the most common outcome of bacteriuria is an asymptomatic carrier state resembling commensalism at other mucosal sites. It remains unclear if the lack of destructive inflammation merely reflects low virulence or if carrier strains actively inhibit disease associated responses in the host. To address this question, we examined the effects of asymptomatic bacterial carriage on host gene expression. Therapeutic urinary tract inoculation with the prototype ABU strain E. coli 83972 is a safe alternative approach in patients with therapy-resistant recurrent UTI. The strain establishes persistent bacteriuria, protecting patients against super-infection with more virulent strains. Using this protocol, we examined if the establishment of asymptomatic bacterial carriage alters host gene expression.