The identity of the patients was anonymized and after that further selleck products analysis Inhibitors,Modulators,Libraries of the bacterial strains was performed. Antimicrobial suscepti bility testing was performed as recommended by the Swedish Reference Group for Antibiotics. The blaCTX M gene was detected Inhibitors,Modulators,Libraries using real time PCR and nucleotide sequencing as previously described. The CTX M types and the antibiotic susceptibility of the different ESBL producing E. coli are shown in Table 1. UPEC strain J96, a pyelonephritis isolate, and an hmp deficient mutant of J96 were also used in the study. Deletion mutation of hmp was constructed using homologous recombination and the FLP recombinase as previously Inhibitors,Modulators,Libraries described. There was no difference in the growth ability be tween the mutant strain and the wild type strain. Bacteria were maintained on tryptic soy agar.

Antimicrobial agents All antimicrobial solutions were prepared freshly imme diately before use. DETA NO belongs to the family of diazeniumdiolates that consists of a complex of NO bound to a polyamine parent compound that spontan eously decompose in a pH dependent, first order process. Stock solutions of DETA NO were prepared Inhibitors,Modulators,Libraries in PBS, cefotaxime and polymyxin B nonapeptid were prepared in sterile water. Nitrofurantoin was pre pared in DMSO and miconazole was dis solved in 25% ethanol and 75% polyetylenglykol 400 while heating for 1 h at 60 C. Determination of MIC MIC was determined by the broth dilution test. The test substances were inocu lated with a bacterial suspension in Luria Bertani broth for 8 24 h at 37 C. All MIC tests were re peated in two isolates and at least twice.

The MIC value for cefotaxime was determined in a cephalosporin sensitive uropathogenic E. coli isolate. The MIC value for DETA NO was determined after 8 h since bacterial re growth was seen Inhibitors,Modulators,Libraries in all tubes after 24 h. A working concen tration of 4 MIC was used in all experiments. Evaluation of bacterial viability An overnight culture grown in LB broth, at 37 C on shake at 200 rpm, were diluted 1 1000 to give a bacterial count of approximately 106 CFU ml in the tube. Time zero samples were taken and the number of viable colonies determined as described below. Bacterial cultures were combined with antibiotics or relevant vehicle and incubated in darkness at 37 C. Samples were taken at different times after addition of antibiotics or vehicle depending on the experimental protocol.

The samples were diluted in PBS and at least three serial dilutions were plated on TSA plates. Following overnight culture at 37 C, bacterial CFU ml was determined by using mean from two dilu tions. Growth was calculated as the numbers of CFU ml in treated cultures or controls divided by the number of CFU ml formed upon the plating then of the initial starting inoculums and expressed as log CFU ml.