Bacterial strains and phage used in this study.

<p>Bacterial strains and phage used in this study.</p

Capsule inhibitors sensitize UPEC K1 strain to serum-mediated killing.

<p><i>E. coli</i> UTI89 and genetic capsule mutants were grown in the presence and absence of DU003 or DU011 at 50 µM and exposed to human serum. Bacterial metabolism and viability was measured using MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide). UTI89 grown in the presence of 50 µM DU003 or DU011 were significantly more sensitive to pooled human serum compared to control UTI89 (** p = 0.0067). This was similar to the serum sensitivity of the capsule mutant UTI89 ΔRII. ΔRI and ΔRII indicate a complete deletion of Region I of the capsule <i>kps</i> and Region II capsule <i>neu</i> loci, respectively.</p

Concentration-response inhibition of K1 and T7 phage-mediated cell lysis.

<p>K1 (<b>A, C</b>) and T7 (<b>B, D</b>) phage activity in <i>E. coli</i> strain UTI89 or EV36 (K1∶K-12 hybrid strain), respectively, following treatment with various concentrations of DU003 (<b>A, B</b>) or DU011 (<b>C, D</b>).</p

Solubility, permeability and plasma and microsome stability.

1<p>Measured in 1× PBS.</p>2<p>Percent remaining at 1 hr.</p>3<p>Measured in LB medium.</p>4<p>Acceptor pH: 7.4.</p>5<p>Percent remaining at 3 hrs.</p

Biochemical and immunologic verification of <i>E. coli</i> Group 2 capsule inhibition through small molecules.

<p><b>A</b>) Orcinol reactivity of capsular material released by mild acid treatment of cultures grown with 1% DMSO vehicle (UTI89 and genetic capsule mutants) or 100 µM C7, DU001, DU003, DU005, DU007, DU008, or DU011. Data represent independent experiments performed in duplicate. Treatment of K1 strain UTI89 with compounds reduces amount of orcinol-reactive polysaccharides on surface of bacteria by ∼80%. <b>B</b>) Whole-cell anti-K1 dot blots of cultures of UTI89 or indicated genetic capsule mutants treated with 1% DMSO or 100 µM DU001, DU003, DU005, DU007, DU008, or DU011 indicate that treatment of cultures with compounds reduces K1 reactive material to levels comparable to those of genetic capsule mutants. ΔRI and ΔRII indicate a complete deletion of Region I of the capsule <i>kps</i> and Region II capsule <i>neu</i> loci, respectively.</p

Inhibitor treatment decreases capsule production in different pathogenic <i>E. coli</i> serotypes.

<p><b>A</b>) K5 <i>E. coli</i> was grown in vehicle or with different inhibitors (50 and 100 µM) and then challenged with K5 lytic phage, which results in cell death in the presence of capsule. Growth was measured by absorbance at OD₆₀₀. <b>B</b>) Capsular material was isolated from multiple strains grown with and without inhibitor DU011 (200 µM). Capsule preparations were performed in at least 3 independent trials. A single representative image is shown.</p

DU011 protects mice against a lethal dose of K1 <i>E. coli</i>.

<p><b>A</b>) C57BL/6 mice were administered subcutaneous 1% DMSO (control) or DU011 (100 µL of 1 mg/ml in 1% DMSO) 12 hours prior to lethal intraperitoneal injection with 10⁸ CFU of UTI89 prepared in media containing 1% DMSO or DU011 (200 µM in 1% DMSO). Surviving animals continued to receive DMSO or DU011 each 12 hours through the course of the experiment, according to their groups (<b>B</b>) Weight was monitored during DMSO and DU011 administration and after infection.</p