The kinetics of the degradation process is reported to be depende

The kinetics of the degradation process is reported to be dependent largely on the concentration [6]. That is why we conducted a EVP4593 chemical structure further experiment to quantify this phenomenon. The stability of the etoposide solution in the disposable perfusion devices was studied in NaCl 0.9 % and in D5W at 600 mg/L. Sampling and Analytical Pre-treatment After preparing the devices, a sample (S1) was tested at H0 in order to determine the initial concentration of the solution.

A second sample (S2) was tested at H24 to quantify the concentration in the device after 24 h. The samples were placed in a vial and then directly into the chromatographic system. A volume of 10 μL was injected. At H24, we drilled through the balloon drug reservoir via the shell of the device and recovered 100 mL of the solution that were then placed in two 50 mL-Falcon® tubes (F1 and F2). The contents of each tube were centrifuged for 5 min at 3,000 rpm; the supernatant was then eliminated to obtain the precipitate. To obtain the whole precipitate in the device,

the inside of the shell and of the balloon was rinsed twice with 10 mL of water using a syringe with a needle (L1 and L2). L1, L2 and the precipitate were mixed and centrifuged for 5 min at 3,000 rpm. After elimination of the supernatant, the precipitate was dissolved in 25 mL of methanol. Concentrations of etoposide methanolic solutions were determined by HPLC-UV in the conditions described above. Finally, the L1 and L2 samples were

analysed by injecting 10 μL into the chromatographic system. Etoposide concentrations were determined to evaluate Dorsomorphin the efficiency of the washing and thus the reliability of the precipitate recovery method. 3 Results 3.1 PR-171 clinical trial Forced Degradation Study Exposition of etoposide solutions to studied conditions led to precipitation after 48 h for ambient and 33 °C storage conditions except for alkaline conditions, where coloration of solution was observed instead of a precipitation. Figure 3 shows results of the forced degradation study for 600-mg/L etoposide solutions in various dissolution media. Curve A shows the results of an injection of etoposide solution diluted in NaCl 0.9 %; curve B shows the chromatogram resulting from the injection of a solution of etoposide diluted in NaOH 0.1 M injected right after dilution; curve C shows the chromatogram resulting from the injection of a solution of etoposide diluted in H2O2 10 % after 48 h of exposition; curve D shows the chromatogram resulting from the injection of a solution of etoposide diluted in HCL 0.1 M after 48 h of exposition; curve E shows the chromatogram resulting from the injection of a solution of etoposide diluted in NaOH 0.1 M after 48 h of exposition. Exposition to alkaline conditions yields a main degradation product eluted around 6.0 min, its content is increased after 48 h of exposition.

3% and 20% [10, 11] Being a life threatening complication

3% and 20% [10, 11]. Being a life threatening complication buy INK 128 of peptic ulcer disease, it needs special attention with prompt resuscitation and appropriate surgical management if morbidity and mortality are to be avoided [3, 11]. The pattern of perforated PUD has been reported to vary from one geographical area

to another depending on the prevailing socio-demographic and environmental factors [12]. In the developing world, the patient population is young with male predominance, patients present late, and there is a strong association with smoking [13]. In the west the patients tend to be elderly and there is a high incidence of ulcerogenic drug ingestion [14]. The diagnosis of perforated PUD poses a diagnostic challenge in most of cases. The spillage of duodenal or gastric contents into peritoneal cavity causing selleck chemicals abdominal pain, shock, peritonitis, marked tenderness and decreased liver dullness offers little difficulty in diagnosis of perforations [15].The presence of gas under the diaphragm on plain abdominal erect X-ray is diagnostic in 75% of the cases [16]. Since the first description of surgery for acute perforated peptic ulcer disease, many techniques have been recommended. The recent advances in antiulcer therapy have shown that simple closure of perforation with omental patch followed by eradication of H. Pylori is a simple and safe option in many centers and have

changed the old trend of truncal vagotomy and drainage procedures [17]. The definitive operation for perforated PUD is performed by few surgeons. Delay in diagnosis and initiation of surgical treatment of perforated PUD has been reported to be associated

with high morbidity and mortality after surgery for perforated PUD [4, 17]. Early recognition and prompt surgical treatment of perforated PUD is of paramount importance if morbidity and mortality associated with perforated PUD are to be avoided [4, 11]. A successful outcome is obtained by prompt recognition of the diagnosis, aggressive resuscitation and early institution of surgical management. Little work has been done on the surgical management of perforated peptic ulcer disease in our local environment despite Protein tyrosine phosphatase increase in the number of admissions of this condition. The aim of this study was to describe our experience on the surgical management of perforated peptic ulcer disease in our local environment outlining the incidence, clinical presentation, management and outcome of patients with peptic ulcer perforation in our setting and to identify predictors of outcome of these patients. Methods Study design and setting This was a combined retrospective and prospective study of patients operated for peptic ulcer perforations at Bugando Medical Centre (BMC) in Northwestern Tanzania from April 2006 to March 2011. BMC is a tertiary care hospital in Mwanza City that also receives patients from its six neighboring regions around Lake Victoria.

0 (ref )   Employed 1 03 (0 36-2 91) 0 94 1 55 (0 38-6 27) 0 53 S

0 (ref.)   Employed 1.03 (0.36-2.91) 0.94 1.55 (0.38-6.27) 0.53 Surgery         Conservative 1.0 (ref.)   1.0 (ref.)   Mastectomy 1.30 (0.55-3.05) 0.54 1.07 (0.36-3.22) 0.89 Chemotherapy         No 1.0 (ref.   1.0 (ref.)   Yes 1.88 (1.10-6.24) 0.03 1.34 (0.25-7.31) 0.73 Radiotherapy         No 1.0 (ref.) BI 6727 molecular weight   1.0 (ref.)   Yes 1.88 (0.73-4.84) 0.18 2.30 (0.57-9.31) 0.24 Endocrine therapy         No 1.0 (ref.)   1.0 (ref.)   Yes 3.36 (1.57-7.22) 0.002 3.34 (1.38-8.06) 0.007 Pre-treatment sexual dysfunction         No 1.0 (ref.)   1.0 (ref.)   Yes 11.1 (3.78-33.1) < 0.0001 12.3 (3.93-39.0) < 0.0001 Time interval between pre-and

post-treatment evaluations (months) – - 1.10 (0.33-3.63) 0.21 * Obtained from univariate logistic regression analysis ** Obtained from multiple logistic regression analysis (adjusted odds ratio) Discussion The findings from this prospective study indicated that the prevalence of sexual

dysfunction among Iranian breast cancer patients was relatively high. The findings also indicated that younger age, receiving endocrine therapy and pre-treatment sexual dysfunction were independent and significant contributing variables to post-treatment sexual disorders. It is well documented that endocrine effects of adjuvant therapy, especially chemotherapy, in younger survivors causes premature menopause that is associated with poorer quality of life, decreased check details sexual functioning, menopausal symptom distress, and psychosocial distress related to infertility [17], although it is believed that as a whole most adjuvant endocrine therapy or radiation therapy for early stage breast cancer do not causes premature menopause. As noted by Cella and Fallowfield [18], recognition and management of treatment-related side-effects for breast cancer patients receiving adjuvant endocrine therapy is an important issue since such side-effects negatively affect sexual functioning, health-related quality of life and adherence to therapy. They argue that adverse events across all

adjuvant endocrine trials regardless of the treatment, vasomotor symptoms such as hot flushes are the most common side effects. Other frequently reported side-effects such as vaginal discharge, vaginal dryness, dyspareunia, and arthralgia vary in prevalence between tamoxifen and aromatase inhibitors [18]. Although there were significant decreases in all measures at post-treatment assessment compared to pre-treatment evaluation, greater decrease was observed for sexual desire (3.8 vs. 2.8) and lubrication (5.3 vs. 4.3). Perhaps these are very important aspect of sexual life for women and should receive further attention when studying sexual issues in breast cancer patients. It has been shown that sexual desire and lubrication are two important affecting factors in breast cancer survivors after mastectomy [19].

The time-zero dielectric breakdown (TZDB) tests are investigated,

The time-zero dielectric breakdown (TZDB) tests are investigated, and the current–voltage (I-V) characteristics are discussed. It is found that stacking structure owns a higher breakdown field, which would lead to

lower resistance after breakdown. Then, in order to corroborate the results, samples with different IL thicknesses are manufactured and investigated. The stacking structures still own a higher breakdown field. Nevertheless, with the decreasing thickness of IL, higher density of interfacial states and lower breakdown field are observed. The mechanism for the CBL0137 clinical trial observation is proposed, and HRTEM is given in this work. Methods Two different MOS capacitors studied in the first experiment denoted by SH/O and H/O (S stands for stacking structure, H stands for HfO2, and O stands for SiO2) were manufactured on the substrate of p-type (100) Si wafer with a resistivity of 1 ~ 10 Ω cm. The wafers were undergone the process of standard Radio Corporation of America (RCA) cleaning in order to remove impurities. Then, SiO2 as ultrathin IL was grown onto the wafers using the technique of anodization (ANO) after removing native oxides XAV-939 concentration by HF. The oxidation method of ANO could be carried out in room temperature and could provide a promising option for the preparation of low-temperature IL [32, 33]. It was reported that the anodic oxide grown in room

temperature has few pinholes

and owns a good dielectric quality [34, 35]. The samples after anodization were followed by 950°C annealing in N2 for 15 s. Then, sample H/O was undergone the deposition of Hf onto a wafer by sputtering with the power of 60 W for 210 s, followed by NAO process to form HfO2 dielectric. Then, postoxidation annealing (POA) was carried out in a furnace at 380°C for 10 min in order to improve the quality of dielectric layer. The combined procedures from the deposition of Hf to the following annealing are defined as one cycle. Under the circumstance, the sample SH/O would undergo the sputtering time of 90 s as the first cycle PLEKHM2 and that of 60 s as other two cycles. Then, 250-nm aluminum metal was evaporated onto the top of all samples. The process of photolithography was carried out to pattern the devices with square area of 2.25 × 104 μm2. Finally, the back contact was formed by the evaporation of 250-nm aluminum. In order to corroborate our investigation, another two different MOS capacitors with various IL thicknesses denoted by SH/Ox and H/Ox were manufactured. Ox represents the SiO2 that was formed with various thicknesses from ANO process. There are two main differences of the experiments for SH/Ox and H/Ox in comparison with SH/O and H/O. First, the platinum was tilted while using the ANO in order to form IL with different thicknesses, as shown in Figure 1.

Interestingly, caspase-3 activity was not observed in Aspc1 cells

Interestingly, caspase-3 activity was not observed in Aspc1 cells (Additional file 3 figure S3C), a cell line with less sensitivity to PB282 (Additional file 3 figure S3D). Figure 7 Caspase-3 inhibition by lipophilic antioxidant correlates with caspase dependence. (A) Caspase-3 inhibition by the hydrophobic antioxidant α-tocopherol

(α-toco), hydrophilic antioxidant N-acetylcyteine (NAC), or caspase-3 inhibitor DEVD-FMK (1 μM) in Bxpc3 cells following 24 hour treatment with SW43 (30 μM), PB282 (90 μM), or HCQ (90 μM). Data represents normalized inhibition compared to this website caspase-3 inducing treatment, n = 3, p < 0.05. (B) Cell viability following 24 hour treatment with SW43 or PB282 in the presence of α-toco or NAC. Data represents percent viability compared to DMSO

treated cells, n = 3, * p < 0.05. Discussion Recent synthesis of fluorescently labeled analogs of SV119 (SW120) and PB28 (PB385), allowing live cell imaging, has MK-1775 in vitro shown sigma-2 receptor ligand subcellular localization to the membrane components of the cell ultrastructure [16, 17]. In various pancreatic cancer cell lines we have observed similar results, and hypothesized that strong uptake into the endo-lysosomal compartment induces lysosomal membrane permeabilization (LMP). In addition, weakly basic amines as a class of drugs have N-acetylglucosamine-1-phosphate transferase been shown to induce LMP [24] and cell death [25], and the amine groups present on sigma-2 receptor ligands suggest they can induce LMP. We examined here whether this could influence the caspase-3 activation in pancreatic cancer we observed earlier [8–10] and found that LMP occurs shortly following treatment with a variety of structurally diverse

sigma-2 receptor ligands, verified by both AO and LysoTracker release from the lysosome. Uptake of fluorescently labeled compounds was inhibited by blocking the lysosomal pH gradient with concanamycin A (CMA), a specific inhibitor of the V-Type ATPase [26, 27], and translated into significant viability protection following treatment. SW43 was a stronger inducer of LMP, with greater protection from CMA pretreatment than for PB282. This that some sigma-2 receptor ligands have a greater propensity to influence the lysosomal death pathway Chemical structure differences may be responsible for this difference. For instance, the structure of the N-(9-(6-Aminohexyl)-9-azabicyclo[3.3.1]-nonan-3α-yl)-N-(2-methoxy-5-methylphenyl) carbamate hydrochloride (SV119) derivatives contain an alkyl extension with terminal amine group that is not present in the 1-cyclohexyl-4-[3-(5-methoxy-1,2,3,4-tetrahydro-naphthalen-1-yl)-propyl]-piperazine dihydrochloride (PB28) derivatives, a moiety that increases lysosomal membrane insertion and permeabilization [28].

mtsA contains an lipoprotein peptidase cleavage site signal seque

mtsA contains an lipoprotein peptidase cleavage site signal sequence as defined by Linton & Higgins [25]. To confirm that MtsA is a lipoprotein, the crude cell lysate of S. iniae HD-1

was mixed with Triton X-114, and the detergent phase was analyzed by western blotting using rabbit anti-MtsA antibodies (Figure 3B). The results showed that MtsA protein was extracted by Triton X-114. Together, the results indicated that MtsA protein is a lipoprotein. Figure 3 Analysis of the lipoprotein sequence patterns of MtsA by ScanProsite and the western blotting. (A) The mtsABC lipoprotein was assessed by ScanProsite. The results showed that amino acid residues D1 to D24 (MFKKISLAFAMLLSIFCITACSSQ) hit G+LPPv2 pattern, AZ 628 supplier and amino acid residues D17 to D21 (CITAC) hit PS51257 pattern. The symbol “”<"" indicates that the pattern

is restricted to the N terminus, and X is any amino acid. (B) Western blotting analysis results of the lipoproteins extracted learn more with Triton X-114. Purification of recombinant MtsA To be able to further characterize MtsA, we first expressed recombinant MtsA consisting of amino acid residues D27 to D310 that lacked the putative signal sequence. Briefly, mtsA gene was cloned and the PCR product was isolated from the plasmid after a double digestion with restriction enzymes BamHI and XhoI, and ligated into the compatible site of pET-32a-c (+) Vector to yield recombinant protein Calpain MtsA. The expressed MtsA had a molecular mass of 49.5-kDa (Figure 4) with a tag from Trx·Tag to EcoR V of pET-32a-c (+), which has a molecular weight of 17.7-kDa. The expression level of MtsA peaked after induction with 1 mM IPTG at 37°C for 4 h. The MtsA protein was purified from E. coli BL21 (DE3) under native condition n the soluble form and immunized the

New Zealand white rabbits. The results showed that the rabbit anti-MtsA antibody titers increased from essentially zero to 1:50,000 after four rounds of immunization (Additional file 1, Table S4). The western blotting analysis was performed to show the specificity of immunized sera against purified MtsA (Figure 4, and Additional file 2, Figure S3-4). Figure 4 SDS-PAGE and western blotting analysis of expressed and purified MtsA. Lanes 1~4, SDS-PAGE showing the purification results of MtsA. The gels were stained with Coomassie brilliant blue. Lane 1, molecular mass marker; lane 2, E. coli with control pet-32a-c (+) vector; lane 3, E. coli lysate containing MtsA (approximately 49.5-kDa); lane 4, purified MtsA (approximately 49.5-kDa). Lanes 5~7, western blotting results of purified MtsA. Lane 5, E. coli with control pet-32a-c (+) vector; lane 6, E. coli lysate containing MtsA (approximately 49.5-kDa); lane 7, purified MtsA (approximately 49.5-kDa).

Note that in the wavelength region from 500 to 580 nm, the absorp

Note that in the wavelength region from 500 to 580 nm, the absorption curve of P3HT/Si NWA (T = 40 and 80 nm) overlaps with that of bare Si NWA. This is due to the fact that the bare Si NWA exhibits the absorptance close to 1 in this wavelength region. Thus, although the absorptivity is increased as the P3HTs are coated on the surface of NWA, the absorption curves do not exhibit obvious enhancement. When the incident wavelength is above 650 nm, P3HT becomes transparent and only Si absorbs incident light.

At this region, despite the size of photoactive Si NW is fixed, a certain amount of absorption enhancement can still be observed as the thickness of organic coating is increased. For example, at the wavelength of 700 nm, we note that the absorption at T = 80 nm has a factor of 1.81 higher than the case of the uncoated NWs. This can be understood Selleckchem Adriamycin by electrostatic approximation. The absorption in Si NW is proportional to the factor of |E core / E inc|2, where E core and E inc are the electric field intensity in the core and incident light of Si NW, respectively [17]. In the absence of the organic coating, |E core / E inc|2 = |2ϵ ext

/ (ϵ ext + ϵ core)|2 = 0.0169, where ϵ ext = 1 is the dielectric function of the vacuum exterior to selleck chemical the NW, and ϵ core ≈ 14.34 + 0.0985i is the dielectric function (for λ = 700 nm) of the Si NW. When an organic coating is added, |E core / E inc|2 = |2ϵ ext / (ϵ ext + ϵ coat)|2|2ϵ coat / (ϵ coat + ϵ core)|2 = 0.030, where ϵ coat = 3.75 is the dielectric function (for λ = 700 nm) of P3HT. About 1.78 times enhancement can be obtained at organic coating T = 80 nm than that of uncoated NWs, which is close to the absorptance enhancement at this wavelength Tolmetin (as shown in Figure 2c). Obviously, above the cutoff of P3HT, the organic coating can serve as a non-absorbing dielectric shell, which drastically increased the absorption in vertical semiconductor NWs. Moreover, at the wavelength larger than

650 nm, the extinction coefficient of silicon is small and interference effects exist, resulting in the oscillation of reflectance and transmittance [6]. Figure 2 Optical characteristics of the hybrid solar cells with various P3HT coating thicknesses. (a) Reflection. (b) Transmission. (c) Absorption. In order to understand the propagation of light in the hybrid solar cells, we simulated the electrical field intensity and calculated the optical generation rates within the arrays from where ϵ″ is the imaginary part of the complex permittivity and E is the electric field [18]. We give the optical generation rates for conformal coating hybrid structure with 80-nm P3HT at three typical wavelengths of 400, 600, and 700 nm. The optical generation rates of the uncoated Si NWs are used as comparison.

Several genes encoding proteases and protein modification enzymes

Several genes encoding proteases and protein modification enzymes such as ClpP1, ClpP2, ClpX, Lon, HslUV, HflCKX, FtsH, HtpX and Dcp also showed significantly increased expression in the tolC mutant. In addition to protecting proteins from destruction or degradation

of the denatured ones the rpoH regulon also protects other macromolecules CP673451 molecular weight like DNA and RNA [17]. In the tolC mutant we observed increased expression of the gene encoding Mfd which recruits the DNA repair machinery to lesions, as well as genes such as mutM, recJ, topA and xerD encoding products known to maintain genomic integrity [20]. Reinforcing the idea of the tolC mutant strain being under stress, the expression of many transcripts encoding enzymes involved in detoxification and protection against oxidative stress was increased. Examples include gst1, gst4, gst7 and gst11, all of which encode glutathione S-transferases. Glutathione transferase proteins catalyze nucleophilic attack by the tripeptide glutathione (GSH) on a wide range of hydrophobic toxic compounds. They are also capable of non-catalytically binding a large number of endogenous compounds, playing an AZD5582 purchase active role in protection against oxidative stress and detoxification of harmful xenobiotics [21]. Other genes with increased expression were

katA (3.7-fold) encoding a catalase, sodB (2.4-fold) encoding a superoxide dismutase, cpo (2.5-fold) encoding a chloride peroxidase, and gor (1.8-fold) encoding a glutathione reductase. Gene thtR showed the greatest expression in this functional class with a 29.3-fold increase (Table 1). thtR encodes a protein LY294002 homologous to tiosulphate sulfurtransferases of the Rhodanese family, which catalyze the transfer of the sulphate atom of thiosulphate to cyanide, to form sulphite and thyocianate. Several studies indicate that these proteins may function as antioxidants capable of scavenging oxidative species that would otherwise lead to inactivation of enzymes such as those containing Fe-S clusters [22]. To confirm microarray data and demonstrate that the tolC mutant is under oxidative stress, enzymatic activities

of catalase, superoxide dismutase and glutathione reductase were determined in cells grown in GMS medium for 20 hours (Fig. 4). Results showed that the specific activity of glutathione reductase in the total protein extract of the tolC mutant was twice that of the wild-type strain (Fig. 4a). In-gel activity staining was used to visualize catalase activity. Despite increased expression of the katA gene and decreased katB expression compared to the wild-type strain, increased catalase activity was detected in the tolC mutant (Fig. 4b). SOD activity was also higher in the tolC mutant (Fig. 4c). The active SodB protein is a dimer [23] and corresponds probably to the lower band, while the upper band must be a multimeric form.

Thus, E195 and E368 (marked

Thus, E195 and E368 (marked CX-6258 mouse with two boxes), which located in two conserved regions, were thought to be the active site residues of Gal308 based on amino acid sequence alignment and the determined structure of β-galactosidase from T. Thermophilus (Figure 1). Figure 1 Identification of the active site residues of Gal308 by alignment of the amino acid residues with other five homologous

β-galactosidases from GH family 42. The GenBank accession numbers are as follows: Geobacillus thermocatenulatus, AAW56416; Truepera radiovictrix DSM17093, ADI14846; Thermus thermophilus, ABI35985; Alicyclobacillus acidocaldarius, AAZ81841; Bacillus circulans, AAA22260; This study (Gal308), AFD21844. The alignment was carried out using the Clustal W method. The number flanking the sequences represents amino acid positions of each sequence. Asterisks mean identity. The two putative catalytic residues (E195 and E368) of Gal308 were shown in box. Heterologous expression and purification of recombinant Epigenetics inhibitor Gal308 To investigate the biochemical properties of Gal308, E. coli expression vector pET-32a(+) was used to express recombinant protein under the conditions described in materials and methods.

The cells were harvested and disrupted by sonication in ice-water bath. The cell lysate was found fully clear, and no inclusion bodies were formed, which suggested that the recombinant Gal308 was highly soluble. Then, the recombinant Lac308 with a six-histidine tag was purified by Ni-NTA chromatography, and the result showed that Ni-NTA chromatography of cell lysate led to 6.25-fold purification and 85% activity yield (Table 1). Furthermore, the purified

enzyme and the crude enzyme (supernatant from cell lysates) were applied to SDS-PAGE (Figure 2) together to determine the molecular mass and expression level of recombinant protein. The purified recombinant protein showed a single protein band of approximate 95 kDa, higher than its calculated molecular mass (76.77 kDa), which can be ascribed to its N-terminal fusion of 156 amino acids (about 18 kDa) corresponding to thioredoxin tag (Trx·Tag), polyhistidine tag (His·Tag), S·Tag epitope oxyclozanide (S·Tag), and a unique thrombin cleavage site (thrombin). In addition, the highest expression level of gal308 in E. coli was about 125 mg/L when the cell was induced at 30°C for 8 h. Next, the purified Gal308 was used to study its biochemical properties. Table 1 Purification of Gal308 Purification step Total protein (mg) Total activity (U) Specific activity (U/mg) Fold purification Activity yield (%) Cell lysate 37.94 1122.21 29.58 1.00 100.00% Ni-NTA chromatography 5.16 953.88 184.86 6.25 85.00% Figure 2 SDS-PAGE analysis of recombinant Gal308 from supernatant of E. coli BL21 (DE3) cell lysates and purified Gal308 by affinity chromatography. Lanes: M, standard protein molecular mass markers (sizes in kilodaltons are indicated on the left); 1, recombinant Gal308 from supernatant of E.

The wells in the second plate were carefully washed three times w

The wells in the second plate were carefully washed three times with PBS and then

used to determine the total number of adherent bacteria. All assays were performed in duplicate and repeated independently four times. Murine models of infection Six- to eight-week-old female CFW1 mice (Harlan) selleck chemicals were used for intestinal colonization experiments as described previously [64]. Briefly, mice were provided with drinking water containing 5 g/l streptomycin sulphate for 24 h and fed a 100 μl suspension containing ~109 CFU of each strain in 20% sucrose. On indicated days, faecal pellets were collected, weighed and homogenised in 0.9% NaCl and dilutions plated onto MacConkey agar supplemented Geneticin with appropriate antibiotics for faecal CFU counts.

A previously described intranasal infection model was used in a co-infection format [23]. Six- to eight-week-old female NMRi mice (Harlan) were anaesthetized and hooked on a string by their front teeth. 50 μl of bacterial suspension containing ~5 × 107 CFU of each strain was dropped onto the nares to allow for aspiration. Mice were left hooked on the string for 10 min before being returned to their cages. At sacrifice lungs, spleen and liver were collected in 0.9% NaCl and homogenised. Serial dilutions were plated on selective media for CFU counts. The ascending urinary tract infection model in which C3H mice (Harlan) were inoculated transurethrally

PDK4 with 50 μl of bacterial suspension containing ~5 × 108 CFU bacteria has been described in detail previously [22, 65]. All animal experiments were conducted under the auspices of the Animal Experiments Inspectorate, the Danish Ministry of Justice. Data analysis, statistics and nucleotide accession number Nucleotide sequences were annotated and analysed using the Integrative Services for Genomic Analysis software and manually curated [66]. The competitive index (CI) was calculated by dividing the ratio of fim2-positive to fim2-negative bacteria recovered from infected organs by the ratio of the corresponding bacteria in the initial inoculum. The non-parametric Mann–Whitney U test was used to analyse infection data. Biofilm and cell-adhesion data were analysed using the non-parametric Kruskal-Wallis test and Dunn’s posthoc analysis. The nucleotide sequence of KpGI-5 has been deposited online [GenBank: JN181158]. Acknowledgements We thank Jean-Marc Ghigo, Unité de Génétique des Biofilms, Institut Pasteur, France, for providing pKOBEG-Apra and Stefan Hyman, Centre for Core Biotechnology Services, University of Leicester, for electron microscopy analysis. This study was supported by a Medisearch research grant. JJvA was supported by a University of Leicester, 50th Anniversary PhD Scholarship. SGS was partially supported by the Danish Research Agency grant 2101-06-0009.