The duodenal mucosa in patients with renal failure: Response to 1,25(OH)2D3

The duodenal mucosa in patients with renal failure: Response to 1,25(OH)2D3. The structure of the duodenal mucosa was evaluated i n duodenal biopsy samples obtained from patients with moderate renal failure (MRF) and in dialysis patients (HD) in an effort to examine the possibility that changes in duodenal mucosa may contribute to the impaired calcium absorption in renal failure (RF). The effect of therapy with 1,25(OH)2D3 on the duodenal mucosa in the HD patients was also studied. The results show that both MRF and HD patients have reduction in calcium reabsorption and in the length of their intestinal villi and crypts of Lieberkuhn. In the HD patients, these structural changes were more severe. Treatment with 1,25(OH)2D3 produced significant improvement in calcium reabsorption (P < 0.01) as well as in length of villus and crypt (P < 0.02) and increased mitotic activity in the crypts (P < 0.02). Electron microscopy revealed the microvilli to be shorter, irregularly distributed, moth-eaten, and grainy, with these abnormalities disappearing after treatment. The data show that duodenal mucosa in RF exhibits structural abnormalities, which were normalized after 1,25(OH)2D3 therapy, and suggest that these derangements may play a role in the defective calcium reabsorption in RF.La muqueuse duodénale chez les malades en insuffisance rénale: Réponse au 1,25(OH)2D3. La structure de la muqueuse duodénale a été évaluée sur des biopsies duodénales de malades atteints d'insuffisance rénale modérée (MFR) et de malades en hémodialyse (HD) afin d'étudier l'hypothèse selon laquelle des modifications de la muqueuse duodénale pourraient contribuer à l'altération de l'absorption du calcium au cours de l'insuffisance rénale. L'effet du traitement par 1,25(OH)2D3 sur la muqueuse duodénale a été étudié chez les malades HD. Les résultats montrent que les malades MRF et HD ont une diminution de l'absorption du calcium et de la longueur de leurs villosités intestinales et de leurs cryptes de Lieberkuhn. Chez les malades HD ces modifications de structure sont encore plus sévères. Le traitement par 1,25(OH)2D3 détermine une amélioration significative de l'absorption du calcium (P < 0,01) de même qu'une augmentation de la longueur des villosités et des cryptes (P < 0,02) et une augmentation de l'activité mitotique dans les cryptes (P < 0,02). La microscopie électronique montre que les micro-villosités sont raccourcies, irrégulièrement distribuées et d'aspect mité et granuleux, anomalies qui disparaissent après le traitement. Les résultats montrent que la muqueuse duodénale des malades RF a des anomalies de structure qui sont normalisées au cours du traitement par 1,25(OH)2D3 et suggèrent que ces modifications peuvent jouer un rôle dans le déficit de au cours de RF

Uranium exposure of the Swiss population based on 24-hour urinary excretion.

Important regional differences in uranium exposure exist because of varying uranium concentrations in soil, water and food. Comprehensive data on the exposure of the general population to uranium is, however, scarce. Based on the 24-hour urinary excretion, the uranium exposure of the adult Swiss population was assessed in relation to age, sex, place of residence, body mass index (BMI), smoking habit and type of drinking water, as well as risk factors in relation to kidney impairment and indicators of a possible renal dysfunction. Uranium was quantified in 24-hour urine from a nationwide population-based sample (n = 1393). The ratio 238U/233U was measured for isotope dilution calibration with a sector field inductively coupled plasma mass spectrometer (HR-ICP-MS). Overall median and 95th percentile were 15 and 67 ng/24 h, respectively. The place of residence significantly influenced urinary uranium excretion. However, most of the highest urinary uranium excretion levels could not be associated to areas known for their elevated uranium concentrations in the drinking water. Sources other than the local drinking water (e.g., bottled water) might be important, too. Gender as well as albumin excretion also had a significant effect on uranium excretion. The latter was, however, strongly dependent on the presence of diabetes mellitus. No association was found for age, BMI, smoking habit or the other examined kidney related variables. On the basis of uranium exposure, assessed via 24-hour urinary uranium excretion, and current knowledge of the toxicity of naturally occurring uranium, a substantial corresponding health risk for the general adult population is unlikely. However, as long as no specific sensitive biomarker for the biological impact of low-dose chronic uranium exposure has been identified and validated, assessing subtle health impact of such exposure will remain difficult

A versatile element for gene addition in bacterial chromosomes

The increasing interest in genetic manipulation of bacterial host metabolic pathways for protein or small molecule production has led to a need to add new genes to a chromosome quickly and easily without leaving behind a selectable marker. The present report describes a vector and four-day procedure that enable site-specific chromosomal insertion of cloned genes in a context insulated from external transcription, usable once in a construction series. The use of rhamnose-inducible transcription from rhaBp allows regulation of the inserted genes independently of the commonly used IPTG and arabinose strategies. Using lacZ as a reporter, we first show that expression from the rhamnose promoter is tightly regulatable, exhibiting very low leakage of background expression compared with background, and moderate rhamnose-induced expression compared with IPTG-induced expression from lacp. Second, the expression of a DNA methyltransferase was used to show that rhamnose regulation yielded on-off expression of this enzyme, such that a resident high-copy plasmid was either fully sensitive or fully resistant to isoschizomer restriction enzyme cleavage. In both cases, growth medium manipulation allows intermediate levels of expression. The vehicle can also be adapted as an ORF-cloning vector

Use of day and night urinary iodine excretion to estimate the prevalence of inadequate iodine intakes via the estimated average requirement cut-point method.

The objectives were to determine urinary iodine concentration (UIC) in day and night samples collected over a 24-hour period and evaluate the usual dietary iodine intake distribution from this collection. We propose a method by which the prevalence of inadequacy can be calculated from a single 24-hour collection, reducing the burden on participants and the study costs. The samples from 1128 participants were collected between 2009 and 2013 within the framework of the Swiss Kidney Project on Genes observational cohort study; 1024 samples were suitable for statistical evaluation of iodine analysis. Participants were over 18, resident in Switzerland and of European ancestry. Over 24 hours, urine was collected as night-time (bedtime until and including first morning urine) and day-time (the remainder) samples. Associations with variables, in particular to estimated glomerular filtration rate (eGFR), were investigated using mixed models. The 24-hour median UICs were 73 and 96 &amp;micro;g/l for women (n = 542) and men (n = 482), respectively; 24-hour median intakes (derived from the corresponding excretion) were 127 and 156 &amp;micro;g/d, respectively. Day and night excretions were normalised to 24-hour excretion values and the usual intake distribution calculated by the US National Cancer Institute method. The Estimated Average Requirement cut-point method was used to calculate the prevalence of inadequacy, estimated at 14% for women and 4% for men; above the target of 2-3%. We conclude that segregating 24-hour urine into day and night collections is sufficient to determine the prevalence of iodine inadequacy in the population and reduces the burden on participants by sparing a second 24-hour collection. No association between iodine intake and eGFR was found

Nacubactam enhances meropenem activity against carbapenem-resistant klebsiella pneumoniae producing KPC

Carbapenem-resistant Enterobacteriaceae (CRE) are resistant to most antibiotics, making CRE infections extremely difficult to treat with available agents. Klebsiella pneumoniae carbapenemases (KPC-2 and KPC-3) are predominant carbapenemases in CRE in the United States. Nacubactam is a bridged diazabicyclooctane (DBO) -lactamase inhibitor that inactivates class A and C -lactamases and exhibits intrinsic antibiotic and -lactam “enhancer” activity against Enterobacteriaceae. In this study, we examined a collection of meropenem-resistant K. pneumoniae isolates carrying blaKPC-2 or blaKPC-3; meropenem-nacubactam restored susceptibility. Upon testing isogenic Escherichia coli strains producing KPC-2 variants with single-residue substitutions at important Ambler class A positions (K73, S130, R164, E166, N170, D179, K234, E276, etc.), the K234R variant increased the meropenem-nacubactam MIC compared to that for the strain producing KPC-2, without increasing the meropenem MIC. Correspondingly, nacubactam inhibited KPC-2 (apparent Ki [Kiapp] 31 3 M) more efficiently than the K234R variant (Kiapp 270 27 M) and displayed a faster acylation rate (k2/K), which was 5,815 582 M1 s1 for KPC-2 versus 247 25 M1 s1 for the K234R variant. Unlike avibactam, timed mass spectrometry revealed an intact sulfate on nacubactam and a novel peak (337 Da) with the K234R variant. Molecular modeling of the K234R variant showed significant catalytic residue (i.e., S70, K73, and S130) rearrangements that likely interfere with nacubactam binding and acylation. Nacubactam’s aminoethoxy tail formed unproductive interactions with the K234R variant’s active site. Molecular modeling and docking observations were consistent with the results of biochemical analyses. Overall, the meropenem-nacubactam combination is effective against carbapenem-resistant K. pneumoniae. Moreover, our data suggest that -lactamase inhibition by nacubactam proceeds through an alternative mechanism compared to that for avibactam

Apramycin susceptibility of multidrug-resistant Gram-negative blood culture isolates in five countries in South-East Asia

Bloodstream infections (BSIs) are a leading cause of sepsis, a life-threatening condition that contributes significantly to the mortality of bacterial infections. Aminoglycoside antibiotics such as gentamicin or amikacin are essential medicines in the treatment of BSIs, but their clinical efficacy is increasingly compromised by antimicrobial resistance. The aminoglycoside apramycin has demonstrated preclinical efficacy against aminoglycoside- and multidrug-resistant (MDR) Gram-negative bacilli (GNB) and is currently in clinical development for the treatment of critical systemic infections. Here, we collected a panel of 470 MDR GNB isolates from health care facilities in Cambodia, Laos, Singapore, Thailand, and Vietnam for a multi-centre assessment of their antimicrobial susceptibility to apramycin in comparison to other aminoglycosides and colistin by broth microdilution assays. Apramycin and amikacin MICs ≤ 16 µg/mL were found for 462 (98.3%) and 408 (86.8%) GNB isolates, respectively. Susceptibility to gentamicin and tobramycin (MIC ≤ 4 µg/mL) was significantly lower at 122 (26.0%) and 101 (21.5%) susceptible isolates, respectively. Of note, all carbapenem- and third-generation cephalosporin (3GC) resistant Enterobacterales, all Acinetobacter baumannii, and all Pseudomonas aeruginosa isolates tested in this study appeared to be susceptible to apramycin. Of the 65 colistin-resistant isolates tested, only four (6.2%) had an apramycin MIC > 16 µg/mL. Apramycin demonstrated best-in-class activity against a panel of GNB isolates with resistances to other aminoglycosides, carbapenems, 3GC, and colistin, warranting continued consideration of apramycin as a drug candidate for the treatment of multidrug-resistant BSIs. Keywords: Bloodstream infection; Gram negative; aminoglycoside; antimicrobial resistance; apramycin; blood culture isolates

Numerical simulation of the flexural behaviour of composite glass-GFRP beams using smeared crack models

This paper presents a numerical study about the flexural behaviour of rectangular composite glass-GFRP beams, comprising annealed glass and GFRP pultruded profiles bonded with two different adhesives: (soft) polyurethane and (stiff) epoxy. The main objectives of this study were: (i) to fully characterize the non-linear behaviour of glass using the smeared crack approach; and (ii) to assess the applicability of different options to simulate adhesively bonded glass-GFRP joints. An extensive parametric study was developed to evaluate the influence of five parameters on the glass post-cracking non-linear behaviour: (i) glass fracture energy, Gf, (ii) crack band width, h, (iii) glass tensile strength, fg,t, (iv) shape of the tension-softening diagram, and (v) shear retention factor, β. The wide range of the joints’ shear stiffness was simulated by either (i) assuming a perfect bond between glass and GFRP (i.e., neglecting the presence of the adhesive), or (ii) explicitly considering the adhesive, by means of using (ii.1) plane stress elements, or (ii.2) interface elements. For the beams analysed in this paper, the following material model for glass provided a good agreement with experimental results: Gf in the range of 3 to 300 N/m, h equal to the square root of the finite element area, fg,t = 50 MPa, linear softening diagram and β according to a power law. It was also shown that the hypothesis of perfect bond at the GFRP-glass interfaces allows for an accurate simulation of joints with high levels of interaction (epoxy), while calibrated interface elements are needed for joints with low level of interaction (polyurethane).The authors wish to acknowledge FCT, ICIST/CERIS and ISISE for funding the research, and companies SIKA, Guardian and ALTO for supplying the adhesives, the glass panes and the GFRP pultruded profiles used in the experiments. The first author also wishes to thank FCT for the financial support through his PhD scholarship SFRH/BD/80234/2011

A whole-cell biosensor for the detection of gold

Geochemical exploration for gold (Au) is becoming increasingly important to the mining industry. Current processes for Au analyses require sampling materials to be taken from often remote localities. Samples are then transported to a laboratory equipped with suitable analytical facilities, such as Inductively Coupled Plasma-Mass Spectrometry (ICP-MS) or Instrumental Neutron Activation Analysis (INAA). Determining the concentration of Au in samples may take several weeks, leading to long delays in exploration campaigns. Hence, a method for the on-site analysis of Au, such as a biosensor, will greatly benefit the exploration industry. The golTSB genes from Salmonella enterica serovar typhimurium are selectively induced by Au(I/III)-complexes. In the present study, the golTSB operon with a reporter gene, lacZ, was introduced into Escherichia coli. The induction of golTSB::lacZ with Au(I/III)-complexes was tested using a colorimetric β-galactosidase and an electrochemical assay. Measurements of the β-galactosidase activity for concentrations of both Au(I)- and Au(III)-complexes ranging from 0.1 to 5 µM (equivalent to 20 to 1000 ng g⁻¹ or parts-per-billion (ppb)) were accurately quantified. When testing the ability of the biosensor to detect Au(I/III)-complexes(aq) in the presence of other metal ions (Ag(I), Cu(II), Fe(III), Ni(II), Co(II), Zn, As(III), Pb(II), Sb(III) or Bi(III)), cross-reactivity was observed, i.e. the amount of Au measured was either under- or over-estimated. To assess if the biosensor would work with natural samples, soils with different physiochemical properties were spiked with Au-complexes. Subsequently, a selective extraction using 1 M thiosulfate was applied to extract the Au. The results showed that Au could be measured in these extracts with the same accuracy as ICP-MS (P<0.05). This demonstrates that by combining selective extraction with the biosensor system the concentration of Au can be accurately measured, down to a quantification limit of 20 ppb (0.1 µM) and a detection limit of 2 ppb (0.01 µM).Carla M. Zammit, Davide Quaranta, Shane Gibson, Anita J. Zaitouna, Christine Ta, Joël Brugger, Rebecca Y. Lai, Gregor Grass, Frank Reit

Systematic Dissection and Trajectory-Scanning Mutagenesis of the Molecular Interface That Ensures Specificity of Two-Component Signaling Pathways

Two-component signal transduction systems enable bacteria to sense and respond to a wide range of environmental stimuli. Sensor histidine kinases transmit signals to their cognate response regulators via phosphorylation. The faithful transmission of information through two-component pathways and the avoidance of unwanted cross-talk require exquisite specificity of histidine kinase-response regulator interactions to ensure that cells mount the appropriate response to external signals. To identify putative specificity-determining residues, we have analyzed amino acid coevolution in two-component proteins and identified a set of residues that can be used to rationally rewire a model signaling pathway, EnvZ-OmpR. To explore how a relatively small set of residues can dictate partner selectivity, we combined alanine-scanning mutagenesis with an approach we call trajectory-scanning mutagenesis, in which all mutational intermediates between the specificity residues of EnvZ and another kinase, RstB, were systematically examined for phosphotransfer specificity. The same approach was used for the response regulators OmpR and RstA. Collectively, the results begin to reveal the molecular mechanism by which a small set of amino acids enables an individual kinase to discriminate amongst a large set of highly-related response regulators and vice versa. Our results also suggest that the mutational trajectories taken by two-component signaling proteins following gene or pathway duplication may be constrained and subject to differential selective pressures. Only some trajectories allow both the maintenance of phosphotransfer and the avoidance of unwanted cross-talk