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BackgroundConvergence of Klebsiella pneumoniae (KP) pathotypes has been increasingly reported in recent years. These pathogens combine features of both multidrug-resistant and hypervirulent KP. However, clinically used indicators for hypervirulent KP identification, such as hypermucoviscosity, appear to be differentially expressed in convergent KP, potential outbreak clones are difficult to identify. We aimed to fill such knowledge gaps by investigating the temperature dependence of hypermucoviscosity and virulence in a convergent KP strain isolated during a clonal outbreak and belonging to the high-risk sequence type (ST)307.MethodsHypermucoviscosity, biofilm formation, and mortality rates in Galleria mellonella larvae were examined at different temperatures (room temperature, 28°C, 37°C, 40°C and 42°C) and with various phenotypic experiments including electron microscopy. The underlying mechanisms of the phenotypic changes were explored via qPCR analysis to evaluate plasmid copy numbers, and transcriptomics.ResultsOur results show a temperature-dependent switch above 37°C towards a hypermucoviscous phenotype, consistent with increased biofilm formation and in vivo mortality, possibly reflecting a bacterial response to fever-like conditions. Furthermore, we observed an increase in plasmid copy number for a hybrid plasmid harboring carbapenemase and rmpA genes. However, transcriptomic analysis revealed no changes in rmpA expression at higher temperatures, suggesting alternative regulatory pathways.ConclusionThis study not only elucidates the impact of elevated temperatures on hypermucoviscosity and virulence in convergent KP but also sheds light on previously unrecognized aspects of its adaptive behavior, underscoring its resilience to changing environments.

ABSTRACT: Incomplete milking (IM) is one way of mitigating the negative energy balance (NEB) that is characteristic for early lactation and may increase the risk for disease. Our objectives were to test the effects of IM in early lactation on energy balance (EB), metabolic status, udder health, and subsequent performance. To facilitate the practical application, an automated system was used to remove the milking clusters once a predefined amount of milk is withdrawn. Forty-six Holstein cows were equally allocated to either the treatment (TRT, starting on 8 d in milk) or the control group (CON; conventional cluster removal at milk flow rate

Abstract Hypervirulent Klebsiella pneumoniae strains (hvKp) can cause invasive community-acquired infections in healthy patients of all ages. In this study, the prevalence of putative hvKp in a German tertiary center was investigated and hvKp were characterized by phenotypic and molecular assays. All K. pneumoniae isolates in routine microbiological diagnostics from a single center were screened by string-testing over a period of 6 months. String-test positive (≥ 0.5 mm) isolates were re-evaluated on different media and under various conditions (aerobe, anaerobe). For string-test positive isolates, genes (magA, iutA, rmpA and rmpA2) associated with hypermucoviscosity and hypervirulence were amplified by multiplex PCR. PCR-positive isolates were subjected to whole-genome sequencing and sedimentation and biofilm formation assays. From 1310 screened K. pneumoniae isolates in clinical routine 100 isolates (7.6%) were string test positive. From these, 9% (n = 9) were defined as putative hvKp (string-test+/PCR+). Highest rate of string-test-positive isolates was observed on MacConkey agar under aerobic conditions. Amongst these nine putative hvKp isolates, the international lineage ST23 carrying hvKp-plasmid pKpVP-1 was the most common, but also a rare ST86 with pKpVP-2 was identified. All nine isolates showed hypermucoviscosity and weak biofilm formation. In conclusion, 9% of string-positive, respectively 0.69% of all K. pneumoniae isolates from routine were defined as putative hypervirulent. MacConkey agar was the best medium for hvKp screening.

Abstract Mycophenolate mofetil (MMF) is applied in proteinuric kidney diseases, but the exact mechanism of its effect on podocytes is still unknown. Our previous in vitro experiments suggested that MMF can ameliorate podocyte damage via restoration of the Ca2+-actin cytoskeleton axis. The goal of this study was to characterize podocyte biology during MMF treatment in nephrotoxic serum (NTS) nephritis (NTN). NTN was induced in three-week old wild-type mice. On day 3, half of the mice were treated with MMF (100 mg/kgBW/d p.o.) for one week. On day 10, we performed proteomic analysis of glomeruli as well as super-resolution imaging of the slit diaphragm. For multiphoton imaging of Ca2+ concentration ([Ca2+]i), the experimental design was repeated in mice expressing podocyte-specific Ca2+ sensor. MMF ameliorated the proteinuria and crescent formation induced by NTS. We identified significant changes in the abundance of proteins involved in Ca2+ signaling and actin cytoskeleton regulation, which was further confirmed by direct [Ca2+]i imaging in podocytes showing decreased Ca2+ levels after MMF treatment. This was associated with a tendency to restoration of podocyte foot process structure. Here, we provide evidence that MPA has a substantial direct effect on podocytes. MMF contributes to improvement of [Ca2+]i and amelioration of the disorganized actin cytoskeleton in podocytes. These data extend the knowledge of direct effects of immunosuppressants on podocytes that may contribute to a more effective treatment of proteinuric glomerulopathies with the least possible side effects.

Polarization transfer to a bound proton in polarized electron knock-out reactions, A(e→,e′p→), is a powerful tool to look for an in-medium modification of the bound proton. It requires comparison to calculations that consider the many-body effects accompanying the quasi-free process. We report here measured components Px′, Pz′, and their ratio Px′/Pz′, of polarization transfer to protons bound in Ca40, which is described well by the shell model and for which reliable calculations are available. While the calculations capture the essence of the data, our statistical precision allows us to observe deviations that cannot be explained by simple scaling, including by varying the proton electromagnetic form factor ratio GE/GM. We further explore the deviations of the ratio of the polarization transfer components from that of a free proton, (Px′/Pz′)A/(Px′/Pz′)H, and its dependence on the bound-proton virtuality.

BackgroundThe activity, localization, and substrate specificity of the protein phosphatase 2A (PP2A) heterotrimer are controlled by various regulatory B subunits. PR72 belongs to the B'' gene family and has been shown to be upregulated in human heart failure. However, little is known about the functions of PR72 in the myocardium.MethodsTo address this issue, we generated a transgenic mouse model with heart-specific overexpression of PP2A-PR72. Biochemical and physiological methods were used to determine contractility, Ca2+ cycling parameters, and protein phosphorylation.ResultsA 2.5-fold increase in PR72 expression resulted in moderate cardiac hypertrophy. Maximal ventricular pressure was increased in catheterized transgenic mice (TG) compared to wild-type (WT) littermates. This was accompanied by an increased shortening of sarcomere length and faster relaxation at the single-cell level in TG. In parallel with these findings, the peak amplitude of Ca2+ transients was increased, and the decay in intracellular Ca2+ levels was shortened in TG compared to WT. The changes in Ca2+ cycling in TG were also evident from an increase in the full duration and width at half maximum of Ca2+ sparks. Consistent with the contractile data, phosphorylation of phospholamban at threonine-17 was higher in TG hearts. The lower expression of the Na+/Ca2+ exchanger may also contribute to the hypercontractile state in transgenic myocardium.ConclusionOur results suggest that PP2A-PR72 plays an important role in regulating cardiac contractile function and Ca2+ cycling, indicating that the upregulation of PR72 in heart failure is an attempt to compensate functionally.

ABSTRACT: The objective of the current study was to develop a predictive model for calf disease detection in the preweaning period using data from automated milk feeders (AMF). A deep convolutional neural network (CNN) architecture for the detection of respiratory disease and diarrhea in dairy calves was developed. German Holstein calves were fed milk replacer either ad libitum (up to 25 L/d; n = 32) or restrictively (6 L/d; n = 32) via AMF from 10 ± 3 d of life on. Concentrate, hay, and water were freely available. Calf health parameters were scored daily. The AMF measured milk replacer (MR) intake, number of rewarded visits, number of unrewarded visits, and drinking speed. A calf was considered sick if its fecal score was 3 or 4 and its respiratory score was 2 or 3. Only data from AMF up to 47 d of age were included in the analysis. This cut in the data was made to avoid data from the weaning period. Data were split in 80:20 ratios for training and testing data sets according to the Pareto principle. A minimum sensitivity of 80% was considered an appropriate requirement for the prediction models. Considering all calves in group housing, cross-validation of the test data set showed a sensitivity of 83% and a specificity of 79%, with a positive predictive value and a negative predictive value of 37 and 97%, respectively. The area under the curve of the receiver operating characteristic for the deep CNN model was 0.81 for all group-housed calves. The CNN model yielded sensitivity and specificity of 83 and 71%, respectively (for ad libitum-fed calves), and 82 and 87%, respectively (for restricted-fed calves), with good area under the curve-receiver operating characteristic (0.77 to 0.87), indicating that the CNN models can predict calf disease in both groups with different MR allowances. The permutation feature importance was measured by the decrease in model accuracy, and features (behaviors) were summarized in descending order of their relative importance to the CNN model. Drinking speed and MR intake were the main factors to predict calf disease in calves fed ad libitum. The number of unrewarded visits to the milk feeder and MR intake were the main factors to predict calf disease in restricted-fed calves. Despite the relatively small sample size, the results provide strong evidence that daily feeding behavior data from AMF can be used to identify calves at risk for disease. In conclusion, despite a very good testing performance of the CNN model, the relatively low daily prevalence of calf disease in the present study resulted in a high proportion of false-positive alarms.

Protein phosphatase 2A (PP2A) plays a central role in myocardial ischemia-reperfusion (I/R) injury. Several studies showed a detrimental function of PP2A by using either overexpression models of the catalytic subunit (PP2Ac) or exogenous inhibitors of PP2Ac. However, all of these approaches underestimate the contribution of regulatory B subunits in modulating the PP2A holoenzyme. To better understand the influence of B subunits on a “controlled” regulation of PP2A, we tested a mouse model overexpressing PP2A-B56α (TG) in the heart under the conditions of I/R in comparison to wild-type littermates (WT). Contractility was increased after reperfusion in isolated TG hearts that were initially subjected to a 20-min no-flow ischemia. This was associated with lower phosphorylation levels of myosin-binding protein C and the ryanodine receptor 2 in TG compared to WT. The application of okadaic acid abolished the contractile and biochemical effects in TG hearts. Moreover, reperfusion resulted in the detection of higher PP2A-B56α levels in mitochondrial preparations of TG hearts. The phosphorylation of ERK1 was increased in the early reperfusion phase in TG compared to WT hearts corresponding to a transient attenuation of PP2A activity. Ischemia led to a prolonged contracture time in TG hearts and a lower acidification in isolated TG cardiomyocytes. The formation of interstitial fibrosis by transient ligation of the left anterior descending (LAD) artery was reduced in TG compared to WT hearts. Taken together, these findings indicate that overexpression of PP2A-B56α is protective against I/R injury and that B56α merits further investigation as a potential therapeutic target.

Abstract Background In patients with severe polycystic liver disease (PLD), there is a need for new treatments. Estrogens and possibly other female sex hormones stimulate growth in PLD. In some patients, liver volume decreases after menopause. Female sex hormones could therefore be a target for therapy. The AGAINST-PLD study will examine the efficacy of the GnRH agonist leuprorelin, which blocks the production of estrogen and other sex hormones, to reduce liver growth in PLD. Methods The AGAINST-PLD study is an investigator-driven, multicenter, randomized controlled trial. Institutional review board (IRB) approval was received at the University Medical Center of Groningen and will be collected in other sites before opening these sites. Thirty-six female, pre-menopausal patients, with a very large liver volume for age (upper 10% of the PLD population) and ongoing liver growth despite current treatment options will be randomized to direct start of leuprorelin or to 18 months standard of care and delayed start of leuprorelin. Leuprorelin is given as 3.75 mg subcutaneously (s.c.) monthly for the first 3 months followed by 3-monthly depots of 11.25 mg s.c. The trial duration is 36 months. MRI scans to measure liver volume will be performed at screening, 6 months, 18 months, 24 months and 36 months. In addition, blood will be drawn, DEXA-scans will be performed and questionnaires will be collected. This design enables comparison between patients on study treatment and standard of care (first 18 months) and within patients before and during treatment (whole trial). Main outcome is annualized liver growth rate compared between standard of care and study treatment. Secondary outcomes are PLD disease severity, change in liver growth within individuals and (serious) adverse events. The study is designed as a prospective open-label study with blinded endpoint assessment (PROBE). Discussion In this trial, we combined the expertise of hepatologist, nephrologists and gynecologists to study the effect of leuprorelin on liver growth in PLD. In this way, we hope to stop liver growth, reduce symptoms and reduce the need for liver transplantation in severe PLD. Trial registration Eudra CT number 2020-005949-16, registered at 15 Dec 2020. https://www.clinicaltrialsregister.eu/ctr-search/search?query=2020-005949-16 .

Pup is the bacterial analog of ubiquitin for targeting proteins to the proteasome. Here, the authors use cryoEM to visualize structures of the Mycobacterium tuberculosis proteasome translocating a Pup-tagged substrate.

Aims: Na+/Ca2+ exchanger (NCX) upregulation in cardiac diseases like heart failure promotes as an independent proarrhythmic factor early and delayed afterdepolarizations (EADs/DADs) on the single cell level. Consequently, NCX inhibition protects against EADs and DADs in isolated cardiomyocytes. We here investigate, whether these promising cellular in vitro findings likewise apply to an in vivo setup. Methods/Results: Programmed ventricular stimulation (PVS) and isoproterenol were applied to a murine heterozygous NCX-knockout model (KO) to investigate ventricular arrhythmia initiation and perpetuation compared to wild-type (WT). KO displayed a reduced susceptibility towards isoproterenol-induced premature ventricular complexes. During PVS, initiation of single or double ectopic beats was similar between KO and WT. But strikingly, perpetuation of ventricular tachycardia (VT) was significantly increased in KO (animals with VT - KO: 82 %; WT: 47 %; p = 0.0122 / median number of VTs - KO: 4.5 (1.0, 6.25); WT: 0.0 (0.0, 4.0); p = 0.0039). The median VT duration was prolonged in KO (in s; KO: 0.38 (0.19, 0.96); WT: 0.0 (0.0, 0.60); p = 0.0239). The ventricular refractory period (VRP) was shortened in KO (in ms; KO: 15.1 ± 0.7; WT: 18.7 ± 0.7; p = 0.0013). Conclusions: Not the initiation, but the perpetuation of provoked whole-heart in vivo ventricular arrhythmia was increased in KO. As a potential mechanism, we found a significantly reduced VRP, which may promote perpetuation of reentrant ventricular arrhythmia. On a translational perspective, the antiarrhythmic concept of therapeutic NCX inhibition seems to be ambivalent by protecting from initiating afterdepolarizations but favoring arrhythmia perpetuation in vivo at least in a murine model.

Pupylation is a bacterial post-translational protein modification, where the small ubiquitin-like protein Pup is covalently attached to lysine side chains of target proteins, which is a reversible process and depupylation is catalysed by the depupylase enzyme, Dop. Here, the authors present crystal structures of Dop in different functional states, which together with biochemical experiments provide insights into the catalytic mechanism of this enzyme.

ABSTRACT The ability of extensively drug-resistant (XDR) Klebsiella pneumoniae to rapidly acquire resistance to novel antibiotics is a global concern. Moreover, Klebsiella clonal lineages that successfully combine resistance and hypervirulence have increasingly occurred during the last years. However, the underlying mechanisms of counteracting fitness costs that accompany antibiotic resistance acquisition remain largely unexplored. Here, we investigated whether and how an XDR sequence type (ST)307 K. pneumoniae strain developed resistance against the novel drug combination ceftazidime-avibactam (CAZ-AVI) using experimental evolution. In addition, we performed in vitro and in vivo assays, molecular modeling, and bioinformatics to identify resistance-conferring processes and explore the resulting decrease in fitness and virulence. The subsequent amelioration of the initial costs was also addressed. We demonstrate that distinct mutations of the major nonselective porin OmpK36 caused CAZ-AVI resistance that persists even upon following a second experimental evolution without antibiotic selection pressure and that the Klebsiella strain compensates the resulting fitness and virulence costs. Furthermore, the genomic and transcriptomic analyses suggest the envelope stress response regulator rpoE and associated RpoE-regulated genes as drivers of this compensation. This study verifies the crucial role of OmpK36 in CAZ-AVI resistance and shows the rapid adaptation of a bacterial pathogen to compensate fitness- and virulence-associated resistance costs, which possibly contributes to the emergence of successful clonal lineages. IMPORTANCE Extensively drug-resistant Klebsiella pneumoniae causing major outbreaks and severe infections has become a significant challenge for health care systems worldwide. Rapid resistance development against last-resort therapeutics like ceftazidime-avibactam is a significant driver for the accelerated emergence of such pathogens. Therefore, it is crucial to understand what exactly mediates rapid resistance acquisition and how bacterial pathogens counteract accompanying fitness and virulence costs. By combining bioinformatics with in vitro and in vivo phenotypic approaches, this study revealed the critical role of mutations in a particular porin channel in ceftazidime-avibactam resistance development and a major metabolic regulator for ameliorating fitness and virulence costs. These results highlight underlying mechanisms and contribute to the understanding of factors important for the emergence of successful bacterial pathogens.

Wind data collection in the atmospheric boundary layer benefits from short-term wind speed measurements using unmanned aerial vehicles. Fixed-wing and rotary-wing devices with diverse anemometer technology have been used in the past to provide such data, but the accuracy still has the potential to be increased. A lightweight drone for carrying an industry-standard precision sonic anemometer was developed. Accuracy tests have been performed with the isolated anemometer at high tilt angles in a calibration wind tunnel, with the drone flying in a large wind tunnel and with the full system flying at different heights next to a bistatic lidar reference. The propeller-induced flow deflects the air to some extent, but this effect is compensated effectively. The data fusion shows a substantial reduction of crosstalk (factor of 13) between ground speed and wind speed. When compared with the bistatic lidar in very turbulent conditions, with a 10 s averaging interval and with the unmanned aerial vehicle (UAV) constantly circling around the measurement volume of the lidar reference, wind speed measurements have a bias between −2.0 % and 4.2 % (root-mean-square error (RMSE) of 4.3 % to 15.5 %), vertical wind speed bias is between −0.05 and 0.07 m s−1 (RMSE of 0.15 to 0.4 m s−1), elevation bias is between −1 and 0.7∘ (RMSE of 1.2 to 6.3∘), and azimuth bias is between −2.6 and 7.2∘ (RMSE of 2.6 to 8.0∘). Key requirements for good accuracy under challenging and dynamic conditions are the use of a full-size sonic anemometer, a large distance between anemometer and propellers, and a suitable algorithm for reducing the effect of propeller-induced flow. The system was finally flown in the wake of a wind turbine, successfully measuring the spatial velocity deficit and downwash distribution during forward flight, yielding results that are in very close agreement to lidar measurements and the theoretical distribution. We believe that the results presented in this paper can provide important information for designing flying systems for precise air speed measurements either for short duration at multiple locations (battery powered) or for long duration at a single location (power supplied via cable). UAVs that are able to accurately measure three-dimensional wind might be used as a cost-effective and flexible addition to measurement masts and lidar scans.

Super-resolution aims at increasing image resolution by algorithmic means and has progressed over the recent years due to advances in the fields of computer vision and deep learning. Convolutional Neural Networks based on a variety of architectures have been applied to the problem, e.g. autoencoders and residual networks. While most research focuses on the processing of photographs consisting only of RGB color channels, little work can be found concentrating on multi-band, analytic satellite imagery. Satellite images often include a panchromatic band, which has higher spatial resolution but lower spectral resolution than the other bands. In the field of remote sensing, there is a long tradition of applying pan-sharpening to satellite images, i.e. bringing the multispectral bands to the higher spatial resolution by merging them with the panchromatic band. To our knowledge there are so far no approaches to super-resolution which take advantage of the panchromatic band. In this paper we propose a method to train state-of-the-art CNNs using pairs of lower-resolution multispectral and high-resolution pan-sharpened image tiles in order to create super-resolved analytic images. The derived quality metrics show that the method improves information content of the processed images. We compare the results created by four CNN architectures, with RedNet30 performing best.

Alternative flooring designs in broiler housing have been the subject of intensive research. Research comparing different floor types with a focus on animal-based welfare indicators might be of special interest to meet the animal's needs. This case–control study investigated the effect of a partially perforated vs. a littered flooring system on health- and behavior-based welfare indicators of fast-growing Ross 308 broilers. Furthermore, production performance was assessed. The experimental barn was partially (50%) equipped with a perforated floor directly underneath the feeders and water lines accessible by perforated ramps. Conventional wood shavings were used in the control barn, as usual in practice. There were 4 fattening periods (repetitions) of 31 to 32 D performed with 500 animals per barn (final density of 39 kg m−2). Beside the flooring system, management conditions were identical. Health- and behavior-based welfare indicators were assessed weekly. Production performance indicators were measured continuously during animal control. During the avoidance distance test, animals were less fearful on day 21 (P = 0.010) and tended to be less fearful on day 28 (P = 0.083) in the barn with the partially perforated flooring system compared to the littered control barn. More animals around the novel object were also assessed in the barn with the partially perforated flooring system during the novel object test on day 1 (P

We present measurements of the electron helicity asymmetry in quasi-elastic proton knockout from 2H and 12C nuclei by polarized electrons. This asymmetry depends on the fifth structure function, is antisymmetric with respect to the scattering plane, and vanishes in the absence of final-state interactions, and thus it provides a sensitive tool for their study. Our kinematics cover the full range in off-coplanarity angle ϕpq, with a polar angle θpq coverage up to about 8°. The missing energy resolution enabled us to determine the asymmetries for knock-out resulting in different states of the residual 11B system. We find that the helicity asymmetry for p-shell knockout from 12C depends on the final state of the residual system and is relatively large (up to ≈0.16), especially at low missing momentum. It is considerably smaller (up to ≈0.01) for s-shell knockout from both 12C and 2H. The data for 2H are in very good agreement with theoretical calculations, while the predictions for 12C exhibit differences with respect to the data.

The transcriptional activator PafBC is a regulator of mycobacterial DNA damage response and upregulates genes involved in DNA repair. Here, the authors present the crystal structure of Arthrobacter aurescens PafBC and suggest that PafBC might be activated by binding of a nucleic acid ligand, and bioinformatics analysis shows that its central WYL domain is a widespread feature in bacterial transcription factors.

We present measurements of the polarization-transfer components in the H2(e→,e′p→) reaction, covering a previously unexplored kinematic region with large positive (anti-parallel) missing momentum, pmiss, up to 220MeV/c, and Q2=0.65 (GeV/c)2. These measurements, performed at the Mainz Microtron (MAMI), were motivated by theoretical calculations which predict small final-state interaction (FSI) effects in these kinematics, making them favorable for searching for medium modifications of bound nucleons in nuclei. We find in this kinematic region that the measured polarization-transfer components Px and Pz and their ratio agree with the theoretical calculations, which use free-proton form factors. Using this, we establish upper limits on possible medium effects that modify the bound proton's form factor ratio GE/GM at the level of a few percent. We also compare the measured polarization-transfer components and their ratio for 2H to those of a free (moving) proton. We find that the universal behavior of 2H, 4He and 12C in the double ratio (Px/Pz)A(Px/Pz)H1 is maintained in the positive missing-momentum region.

A comparison between polarization-transfer to a bound proton in quasi-free kinematics by the A(e→,e′p→) knockout reaction and that in elastic scattering off a free proton can provide information on the characteristics of the bound proton. In the past the reported measurements have been compared to those of a free proton with zero initial momentum. We introduce, for the first time, expressions for the polarization-transfer components when the proton is initially in motion and compare them to the 2H data measured at the Mainz Microtron (MAMI). We show the ratios of the transverse (Px) and longitudinal (Pz) components of the polarization transfer in H2(e→,e′p→)n, to those of elastic scattering off a “moving proton”, assuming the proton's initial (Fermi-motion) momentum equals the negative missing momentum in the measured reaction. We found that the correction due to the proton motion is up to 20% at high missing momentum. However the effect on the double ratio (Px/Pz)A(Px/Pz)H1 is largely canceled out, as shown for both 2H and 12C data. This implies that the difference between the resting- and the moving-proton kinematics is not the primary cause for the deviations between quasi-elastic and elastic scattering reported previously.

Abstract Background A consensus on digital ulcer (DU) definition in systemic sclerosis (SSc) has been recently reached (Suliman et al., J Scleroderma Relat Disord 2:115-20, 2017), while for their evaluation, classification and categorisation, it is still missing. The aims of this study were to identify a set of essential items for digital ulcer (DU) evaluation, to assess if the existing DU classification was useful and feasible in clinical practice and to investigate if the new categorisation was preferred to the simple distinction of DU in recurrent and not recurrent, in patients with systemic sclerosis (SSc). Methods DeSScipher is the largest European multicentre study on SSc. It consists of five observational trials (OTs), and one of them, OT1, is focused on DU management. The DeSScipher OT1 items on DU that reached ≥ 60% of completion rate were administered to EUSTAR (European Scleroderma Trials and Research group) centres via online survey. Questions about feasibility and usefulness of the existing DU classification (DU due to digital pitting scars, to loss of tissue, derived from calcinosis and gangrene) and newly proposed categorisation (episodic, recurrent and chronic) were also asked. Results A total of 84/148 (56.8%) EUSTAR centres completed the questionnaire. DeSScipher items scored by ≥ 70% of the participants as essential and feasible for DU evaluation were the number of DU defined as a loss of tissue (level of agreement 92%), recurrent DU (84%) and number of new DU (74%). For 65% of the centres, the proposed classification of DU was considered useful and feasible in clinical practice. Moreover, 80% of the centres preferred the categorisation of DU in episodic, recurrent and chronic to simple distinction in recurrent/not recurrent DU. Conclusions For clinical practice, EUSTAR centres identified only three essential items for DU evaluation and considered the proposed classification and categorisation as useful and feasible. The set of items needs to be validated while further implementation of DU classification and categorisation is warranted. Trial registration Observational trial on DU (OT1) is one of the five trials of the DeSScipher project (ClinicalTrials.gov; OT1 Identifier: NCT01836263, posted on April 19, 2013).

As part of our ongoing studies on the potential pathophysiological role of serine/threonine phosphatases (PP) in the mammalian heart, we have generated mice with cardiac-specific overexpression of PP2Cβ (PP2C-TG) and compared them with littermate wild type mice (WT) serving as a control. Cardiac fibrosis was noted histologically in PP2C-TG. Collagen 1a, interleukin-6 and the natriuretic peptides ANP and BNP were augmented in PP2C-TG vs. WT (p < 0.05). Left atrial preparations from PP2C-TG were less resistant to hypoxia than atria from WT. PP2C-TG maintained cardiac function after the injection of lipopolysaccharide (LPS, a model of sepsis) and chronic isoproterenol treatment (a model of heart failure) better than WT. Crossbreeding of PP2C-TG mice with PP2A-TG mice (a genetic model of heart failure) resulted in double transgenic (DT) mice that exhibited a pronounced increase of heart weight in contrast to the mild hypertrophy noted in the mono-transgenic mice. The ejection fraction was reduced in PP2C-TG and in PP2A-TG mice compared with WT, but the reduction was the highest in DT compared with WT. PP2A enzyme activity was enhanced in PP2A-TG and DT mice compared with WT and PP2C-TG mice. In summary, cardiac overexpression of PP2Cβ and co-overexpression of both the catalytic subunit of PP2A and PP2Cβ were detrimental to cardiac function. PP2Cβ overexpression made cardiac preparations less resistant to hypoxia than WT, leading to fibrosis, but PP2Cβ overexpression led to better adaptation to some stressors, such as LPS or chronic β-adrenergic stimulation. Hence, the effect of PP2Cβ is context sensitive.

We present the first measurements of the double ratio of the polarization-transfer components (Px′/Pz′)p/(Px′/Pz′)s for knock-out protons from the s and p shells in C12 measured by the C12(e→,e′p→) reaction in quasi-elastic kinematics. The data are compared to theoretical predictions in the relativistic distorted-wave impulse approximation. Our results show that the differences between s- and p-shell protons, observed when compared at the same initial momentum (missing momentum), largely disappear when the comparison is done at the same proton virtuality. We observe no difference in medium modifications between protons from the s and p shells with the same virtuality in spite of the large differences in the respective nuclear densities.

We report measurements of the induced polarization P→ of protons knocked out from 2H and 12C via the A(e,e′p→) reaction. We have studied the dependence of P→ on two kinematic variables: the missing momentum pmiss and the “off-coplanarity” angle ϕpq between the scattering and reaction planes. For the full 360° range in ϕpq, both the normal (Py) and, for the first time, the transverse (Px) components of the induced polarization were measured with respect to the coordinate system associated with the scattering plane. Px vanishes in coplanar kinematics, however in non-coplanar kinematics, it is on the same scale as Py.We find that the dependence on ϕpq is sine-like for Px and cosine-like for Py. For carbon, the magnitude of the induced polarization is especially large when protons are knocked out from the p3/2 shell at very small pmiss. For the deuteron, the induced polarization is near zero at small |pmiss|, and its magnitude increases with |pmiss|. For both nuclei such behavior is reproduced qualitatively by theoretical results, driven largely by the spin-orbit part of the final-state interactions. However, for both nuclei, sizeable discrepancies exist between experiment and theory.

BACKGROUND Transgenic mice (TG) with heart‐directed overexpresion of the isoform of the transcription factor cyclic adenosine monophosphate response element modulator (CREM), CREM‐IbΔC‐X, display spontaneous atrial fibrillation (AF) and action potential prolongation. The remodeling of the underlying ionic currents remains unknown. Here, we investigated the regulatory role of CREM‐IbΔC‐X on the expression of K+ channel subunits and the corresponding K+ currents in relation to AF onset in TG atrial myocytes. METHODS AND RESULTS ECG recordings documented the absence or presence of AF in 6‐week‐old (before AF onset) and 12‐week‐old TG (after AF onset) and wild‐type littermate mice before atria removal to perform patch clamp, contractility, and biochemical experiments. In TG atrial myocytes, we found reduced repolarization reserve K+ currents attributed to a decrease of transiently outward current and inward rectifier K+ current with phenotype progression, and of acetylcholine‐activated K+ current, age independent. The molecular determinants of these changes were lower mRNA levels of Kcnd2/3, Kcnip2, Kcnj2/4, and Kcnj3/5 and decreased protein levels of K+ channel interacting protein 2 (KChIP2 ), Kir2.1/3, and Kir3.1/4, respectively. After AF onset, inward rectifier K+ current contributed less to action potential repolarization, in line with the absence of outward current component, whereas the acetylcholine‐induced action potential shortening before AF onset (6‐week‐old TG mice) was smaller than in wild‐type and 12‐week‐old TG mice. Atrial force of contraction measured under combined vagal‐sympathetic stimulation revealed increased sensitivity to isoprenaline irrespective of AF onset in TG. Moreover, we identified Kcnd2, Kcnd3, Kcnj3, and Kcnh2 as novel CREM‐target genes. CONCLUSIONS Our study links the activation of cyclic adenosine monophosphate response element–mediated transcription to the proarrhythmogenic electrical remodeling of atrial inward rectifier K+ currents with a role in action potential duration, resting membrane stability, and vagal control of the electrical activity.

We report on a new measurement of the beam-normal single spin asymmetry An in the elastic scattering of 570 MeV transversely polarized electrons off 28Si and 90Zr at Q2=0.04 GeV2/c2. The studied kinematics allow for a comprehensive comparison with former results on 12C. No significant mass dependence of the beam-normal single spin asymmetry is observed in the mass regime from 12C to 90Zr.

We report the first measurements of the transverse (Px and Py) and longitudinal (Pz) components of the polarization transfer to a bound proton in the deuteron via the H2(e→,e′p→) reaction, over a wide range of missing momentum. A precise determination of the electron beam polarization reduces the systematic uncertainties on the individual components to a level that enables a detailed comparison to a state-of-the-art calculation of the deuteron using free-proton electromagnetic form factors. We observe very good agreement between the measured and the calculated Px/Pz ratios, but deviations of the individual components. Our results cannot be explained by medium modified electromagnetic form factors. They point to an incomplete description of the nuclear reaction mechanism in the calculation.

We measured the ratio Px/Pz of the transverse to longitudinal components of polarization transferred from electrons to bound protons in C12 by the C12(e→,e′p→) process at the Mainz Microtron (MAMI). We observed consistent deviations from unity of this ratio normalized to the free-proton ratio, (Px/Pz)C12/(Px/Pz)H1, for both s- and p-shell knocked out protons, even though they are embedded in averaged local densities that differ by about a factor of two. The dependence of the double ratio on proton virtuality is similar to the one for knocked out protons from H2 and He4, suggesting a universal behavior. It further implies no dependence on average local nuclear density.

We report on a new experimental method based on initial-state radiation (ISR) in e–p scattering, which exploits the radiative tail of the elastic peak to study the properties of electromagnetic processes and to extract the proton charge form factor (GEp) at extremely small Q2. The ISR technique was implemented in an experiment at the three-spectrometer facility of the Mainz Microtron (MAMI). This led to a precise validation of radiative corrections far away from elastic line and provided first measurements of GEp for 0.001≤Q2≤0.004(GeV/c)2. Keywords: Initial state radiation, Proton, Form factor, Radiative corrections

We report the measurement of the ratio of polarization-transfer components, Px/Pz, in the H2(e→,e′p→)n reaction at low and high missing momenta, in search of differences between free and bound protons. The observed deviation of Px/Pz from that of a free proton, which is similar to that observed in 4He, indicates that the effect in nuclei is a function of the virtuality of the knock-out proton and the missing momentum direction, but not the average nuclear density. There is a general agreement between the data and calculations, which assume free proton form factors, however, the measurements are consistently about 10% higher.

The cross section of the p(e,e′π+)n reaction has been measured for five kinematic settings at an invariant mass of W=1094 MeV and for a four-momentum transfer of Q2=0.078 (GeV/c)2. The measurement has been performed at MAMI using a new short-orbit spectrometer (SOS) of the A1 collaboration, intended for detection of low-energy pions. The transverse and longitudinal cross section terms were separated using the Rosenbluth method and the transverse-longitudinal interference term has been determined from the left-right asymmetry. The experimental cross section terms are compared with the calculations of three models: DMT2001, MAID2007 and χMAID. The results show that we do not yet understand the dynamics of the fundamental pion.

Background/aim: The safety of amateur and professional boxing is a contentious issue. We hypothesised that advanced magnetic resonance imaging and neuropsychological testing could provide evidence of acute and early brain injury in amateur boxers. Methods: We recruited 30 participants from a university amateur boxing club in a prospective cohort study. Magnetic resonance imaging (MRI) and neuropsychological testing was performed at three time points: prior to starting training; within 48h following a first major competition to detect acute brain injury; and one year follow-up. A single MRI acquisition was made from control participants. Imaging analysis included cortical thickness measurements with Advanced Normalization Tools (ANTS) and FreeSurfer, voxel based morphometry (VBM), and Tract Based Spatial Statistics (TBSS). A computerized battery of neuropsychological tests was performed assessing attention, learning, memory and impulsivity. Results: During the study period, one boxer developed seizures controlled with medication while another developed a chronic subdural hematoma requiring neurosurgical drainage. A total of 10 boxers contributed data at to the longitudinal assessment protocol. Reasons for withdrawal were: logistics (10), stopping boxing (7), withdrawal of consent (2), and development of a chronic subdural hematoma (1). No significant changes were detected using VBM, TBSS, cortical thickness measured with FreeSurfer or ANTS, either cross-sectionally at baseline, or longitudinally. Neuropsychological assessment of boxers found attention/concentration improved over time while planning and problem solving ability latency decreased after a bout but recovered after one year. Conclusion: While this neuroimaging and neuropsychological assessment protocol could not detect any evidence of brain injury, one boxer developed seizures and another developed a chronic sub-dural haematoma. Keywords: Boxing, Neuroimaging, Brain structure, CANTAB

Proteasomal protein degradation exists in mycobacteria and other actinobacteria, and expands their repertoire of compartmentalizing protein degradation pathways beyond the usual bacterial types. A product of horizontal gene transfer, bacterial proteasomes have evolved to support the organism's survival under challenging environmental conditions like nutrient starvation and physical or chemical stresses. Like the eukaryotic 20S proteasome, the bacterial core particle is gated and must associate with a regulator complex to form a fully active protease capable of recruiting and internalizing substrate proteins. By association with diverse regulator complexes that employ different recruitment strategies, the bacterial 20S core particle is able to act in different cellular degradation pathways. In association with the mycobacterial proteasomal ATPase Mpa, the proteasome degrades substrates post-translationally modified with prokaryotic, ubiquitin-like protein Pup in a process called pupylation. Upon interaction with the ATP-independent bacterial proteasome activator Bpa, poorly structured substrates are recruited for proteasomal degradation. A potential third degradation route might employ a Cdc48-like protein of actinobacteria (Cpa), for which interaction with the 20S core was recently demonstrated but no degradation substrates have been identified yet. The alternative interaction partners and wide range of substrate proteins suggest that the bacterial proteasome is a modular, functionally flexible and conditionally regulated degradation machine in bacteria that encounter rapidly changing and challenging conditions.

A new global synthesis and biomization of long (> 40 kyr) pollen-data records is presented and used with simulations from the HadCM3 and FAMOUS climate models and the BIOME4 vegetation model to analyse the dynamics of the global terrestrial biosphere and carbon storage over the last glacial–interglacial cycle. Simulated biome distributions using BIOME4 driven by HadCM3 and FAMOUS at the global scale over time generally agree well with those inferred from pollen data. Global average areas of grassland and dry shrubland, desert, and tundra biomes show large-scale increases during the Last Glacial Maximum, between ca. 64 and 74 ka BP and cool substages of Marine Isotope Stage 5, at the expense of the tropical forest, warm-temperate forest, and temperate forest biomes. These changes are reflected in BIOME4 simulations of global net primary productivity, showing good agreement between the two models. Such changes are likely to affect terrestrial carbon storage, which in turn influences the stable carbon isotopic composition of seawater as terrestrial carbon is depleted in 13C.

Background: Principal mechanisms of arrhythmia have been derived from ventricular but not atrial cardiomyocytes of animal models despite higher prevalence of atrial arrhythmia (e.g., atrial fibrillation). Due to significant ultrastructural and functional differences, a simple transfer of ventricular proneness toward arrhythmia to atrial arrhythmia is critical. The use of murine models in arrhythmia research is widespread, despite known translational limitations. We here directly compare atrial and ventricular mechanisms of arrhythmia to identify critical differences that should be considered in murine models for development of antiarrhythmic strategies for atrial arrhythmia.Methods and Results: Isolated murine atrial and ventricular myocytes were analyzed by wide field microscopy and subjected to a proarrhythmic protocol during patch-clamp experiments. As expected, the spindle shaped atrial myocytes showed decreased cell area and membrane capacitance compared to the rectangular shaped ventricular myocytes. Though delayed afterdepolarizations (DADs) could be evoked in a similar fraction of both cell types (80% of cells each), these led significantly more often to the occurrence of spontaneous action potentials (sAPs) in ventricular myocytes. Interestingly, numerous early afterdepolarizations (EADs) were observed in the majority of ventricular myocytes, but there was no EAD in any atrial myocyte (EADs per cell; atrial myocytes: 0 ± 0; n = 25/12 animals; ventricular myocytes: 1.5 [0–43]; n = 20/12 animals; p < 0.05). At the same time, the action potential duration to 90% decay (APD90) was unaltered and the APD50 even increased in atrial versus ventricular myocytes. However, the depolarizing L-type Ca2+ current (ICa) and Na+/Ca2+-exchanger inward current (INCX) were significantly smaller in atrial versus ventricular myocytes.Conclusion: In mice, atrial myocytes exhibit a substantially distinct occurrence of proarrhythmic afterdepolarizations compared to ventricular myocytes, since they are in a similar manner susceptible to DADs but interestingly seem to be protected against EADs and show less sAPs. Key factors in the generation of EADs like ICa and INCX were significantly reduced in atrial versus ventricular myocytes, which may offer a mechanistic explanation for the observed protection against EADs. These findings may be of relevance for current studies on atrial level in murine models to develop targeted strategies for the treatment of atrial arrhythmia.

Background: Atrial fibrillation (AF) is a significant cause of morbidity and mortality with foreseeably increasing prevalence. While large animal models of the disease are well established but resource intensive, transgenic AF mouse models are not yet widely used to develop or validate novel therapeutics for AF. Hemizygous mice with a cardiomyocyte-specific overexpression of the human cAMP response element modulator (CREM) isoform IbΔC-X spontaneously develop AF on grounds of an arrhythmogenic substrate consisting of alterations in structure, conduction, and calcium handling.Objective: We investigated if homozygous expression of the CREM-IbΔC-X transgene in mice alters the time course of AF development, and if homozygous CREM-IbΔC-X transgenics could be suitable as a disease model of AF.Methods: Southern Blot, quantitative real-time PCR, and immunoblotting were used to identify and verify homozygous transgenics. Cardiac gravimetry, quantitative real-time RT-PCR, histology, survival analysis, and repeated ECG recordings allowed assessment of phenotypic development and effects of antiarrhythmic drugs.Results: Homozygous animals could be identified by Southern blot and quantitative PCR, showing a strong trend to increased transgenic protein expression. In homozygous animals, atrial hypertrophy appeared earlier and more pronounced than in hemizygous animals, going along with an earlier onset of spontaneous AF, while no increased early mortality was observed. Application of a rate-controlling drug (esmolol) led to the expected result of a decreased heart rate. Application of a rhythm-controlling drug (flecainide) showed effects on heart rate variability, but did not lead to a definitive conversion to sinus rhythm.Conclusion: We suggest homozygous CREM-IbΔC-X overexpressing mice as a reliable model of early onset, rapidly progressive AF.

Background: Adenosine can be produced in the heart and acts on cardiac adenosine receptors. One of these receptors is the A2A-adenosine receptor (A2A-AR).Methods and Results: To better understand its role in cardiac function, we generated and characterized mice (A2A-TG) which overexpress the human A2A-AR in cardiomyocytes. In isolated atrial preparations from A2A-TG but not from WT, CGS 21680, an A2A-AR agonist, exerted positive inotropic and chronotropic effects. In ventricular preparations from A2A-TG but not WT, CGS 21680 increased the cAMP content and the phosphorylation state of phospholamban and of the inhibitory subunit of troponin in A2A-TG but not WT. Protein expression of phospholamban, SERCA, triadin, and junctin was unchanged in A2A-TG compared to WT. Protein expression of the α-subunit of the stimulatory G-protein was lower in A2A-TG than in WT but expression of the α-subunit of the inhibitory G-protein was higher in A2A-TG than in WT. While basal hemodynamic parameters like left intraventricular pressure and echocardiographic parameters like the systolic diameter of the interventricular septum were higher in A2A-TG than in WT, after β-adrenergic stimulation these differences disappeared. Interestingly, A2A-TG hearts sustained global ischemia better than WT.Conclusion: We have successfully generated transgenic mice with cardiospecific overexpression of a functional A2A-AR. This receptor is able to increase cardiac function per se and after receptor stimulation. It is speculated that this receptor may be useful to sustain contractility in failing human hearts and upon ischemia and reperfusion.

The rapidly increasing interest of foreign investors in land in the global South, also termed land grabbing, has been widely discussed as potentially supportive, but often rather harmful for local populations. Combining a critical livelihoods perspective with access theory and a bargaining model, this study scrutinizes local people's perceptions of the land investments, power relations during land negotiations and intra-community differences. By analysing two European forestry companies in Tanzania, we have chosen a sector and a country with presumably more positive outcomes for local populations. The deals resulted in not only labour opportunities and infrastructural improvements, which are mainly perceived as positive, but also cases of violated land rights, inadequate compensation and decreased food security. Hence, even under favourable preconditions, the consequences for local people are ambivalent. With this study, we contribute to a differentiated analysis of the contested role of large-scale land deals in contemporary rural development.

Background/Objective: The cardiac Na+/Ca2+ exchanger (NCX) has been identified as a promising target to counter arrhythmia in previous studies investigating the benefit of NCX inhibition. However, the consequences of NCX inhibition have not been investigated in the setting of altered NCX expression and function, which is essential, since major cardiac diseases (heart failure/atrial fibrillation) exhibit NCX upregulation. Thus, we here investigated the effects of the NCX inhibitor SEA0400 on the Ca2+ transient amplitude and on proarrhythmia in homozygous NCX overexpressor (OE) and heterozygous NCX knockout (hetKO) mice compared to corresponding wild-types (WTOE/WThetKO).Methods/Results: Ca2+ transients of field-stimulated isolated ventricular cardiomyocytes were recorded with fluo-4-AM or indo-1-AM. SEA0400 (1 μM) significantly reduced NCX forward mode function in all mouse lines. SEA0400 (1 μM) significantly increased the amplitude of field-stimulated Ca2+ transients in WTOE, WThetKO, and hetKO, but not in OE (% of basal; OE = 98.7 ± 5.0; WTOE = 137.8 ± 5.2*; WThetKO = 126.3 ± 6.0*; hetKO = 140.6 ± 12.8*; *p < 0.05 vs. basal). SEA0400 (1 μM) significantly reduced the number of proarrhythmic spontaneous Ca2+ transients (sCR) in OE, but increased it in WTOE, WThetKO and hetKO (sCR per cell; basal/+SEA0400; OE = 12.5/3.7; WTOE = 0.2/2.4; WThetKO = 1.3/8.8; hetKO = 0.2/5.5) and induced Ca2+ overload with subsequent cell death in hetKO.Conclusion: The effects of SEA0400 on Ca2+ transient amplitude and the occurrence of spontaneous Ca2+ transients as a proxy measure for inotropy and cellular proarrhythmia depend on the NCX expression level. The antiarrhythmic effect of SEA0400 in conditions of increased NCX expression promotes the therapeutic concept of NCX inhibition in heart failure/atrial fibrillation. Conversely, in conditions of reduced NCX expression, SEA0400 suppressed the NCX function below a critical level leading to adverse Ca2+ accumulation as reflected by an increase in Ca2+ transient amplitude, proarrhythmia and cell death. Thus, the remaining NCX function under inhibition may be a critical factor determining the inotropic and antiarrhythmic efficacy of SEA0400.

The fight-or-flight response (FFR), a physiological acute stress reaction, involves positive chronotropic and inotropic effects on heart muscle cells mediated through β-adrenoceptor activation. Increased systolic calcium is required to enable stronger heart contractions whereas elevated potassium currents are to limit the duration of the action potentials and prevent arrhythmia. The latter effect is accomplished by an increased functional activity of the Kv7.1 channel encoded by KCNQ1. Current knowledge, however, does not sufficiently explain the full extent of rapid Kv7.1 activation and may hence be incomplete. Using inducible genetic KCNQ1 complementation in KCNQ1-deficient human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), we here reinvestigate the functional role of Kv7.1 in adapting human CMs to adrenergic stress. Under baseline conditions, Kv7.1 was barely detectable at the plasma membrane of hiPSC-CMs, yet it fully protected these from adrenergic stress-induced beat-to-beat variability of repolarization and torsade des pointes-like arrhythmia. Furthermore, isoprenaline treatment increased field potential durations specifically in KCNQ1-deficient CMs to cause these adverse macroscopic effects. Mechanistically, we find that the protective action by Kv7.1 resides in a rapid translocation of channel proteins from intracellular stores to the plasma membrane, induced by adrenergic signaling. Gene silencing experiments targeting RAB GTPases, mediators of intracellular vesicle trafficking, showed that fast Kv7.1 recycling under acute stress conditions is RAB4A-dependent.Our data reveal a key mechanism underlying the rapid adaptation of human cardiomyocytes to adrenergic stress. These findings moreover aid to the understanding of disease pathology in long QT syndrome and bear important implications for safety pharmacological screening.

A subthreshold signal may be detected if noise is added to the data. The noisy signal must be strong enough to exceed the threshold at least occasionally; but very strong noise tends to drown out the signal. There is an optimal noise level, called stochastic resonance. We explore the detectability of different signals, using statistical detectability measures. In the simplest setting, the signal is constant, noise is added in the form of i.i.d. random variables at uniformly spaced times, and the detector records the times at which the noisy signal exceeds the threshold. We study the best estimator for the signal from the thresholded data and determine optimal configurations of several detectors with different thresholds. In a more realistic setting, the noisy signal is described by a nonparametric regression model with equally spaced covariates and i.i.d. errors, and the detector records again the times at which the noisy signal exceeds the threshold. We study Nadaraya–Watson kernel estimators from thresholded data. We determine the asymptotic mean squared error and the asymptotic mean average squared error and calculate the corresponding local and global optimal bandwidths. The minimal asymptotic mean average squared error shows a strong stochastic resonance effect.

No abstract available.

A titanium foam for spine fusion and other applications was tested by cell culture. Its high porosity and surface roughness should enable bone cells to grow through it, resulting in a better fixation of the vertebral body.The foam was tested by in vitro experiments with human osteoblasts under static culture conditions and in a perfused system. By means of cell number, viability, scanning electron microscopy and histological staining, cell proliferation could be observed. The expression of osteogenic genes like collagen-I, alkaline phosphatase and osteocalcin was proven by reverse transcription polymerase chain reaction (RT-PCR) as well as in the case of alkaline phosphatase with biochemical methods.The conducted experiments showed that human osteoblasts could grow through the interconnected porosity of the metal foam and that they expressed an osteoblast like phenotype. The results suggest that in vivo osteoblasts are likely to form a trabecular bone bridge through this titanium foam. Consequently, with this osteoconductive material, there may be a reduced need for autologous bone in spinal fusion procedures.

During the solid-phase PCR (SP-PCR), DNA oligonucleotides complementary to a soluble template and immobilized on a surface are extended in situ. Although primarily used for pathogen detection, SP-PCR has the potential for much broader application, including disease diagnostics, genotyping, and expression studies. Current protocols for SP-PCR in microwells are suitable for enzymatic detection of immobilized products, but yields are generally insufficient for direct detection of products using conventional fluorescent probes. Here, we quantitatively measure the outcome of tethering, hybridization, and solid-phase extension, and examine the effect of composition and length of the spacer at the 5′ end of tethered oligonucleotides. Our results indicate that steric hindrance primarily affects polymerase activity rather than the efficiency of hybridization between the template and the tethered oligonucleotide. SP-PCR yields are significantly higher for a five-unit hexaethyleneglycol (HEG) spacer than for the more commonly used 10-residue deoxythymidine spacer. The optimal 5′ HEG spacer resulted in a 60-fold increase in extension efficiency relative to a previously reported value for SP-PCR on a glass surface. Thus, optimized spacers should allow direct quantification of SP-PCR products, providing a simple, quantitative, and cost effective means of sample analysis for a variety of applications.

A number of studies have concluded that cognitive control is not fully established until late adolescence. The precise differences in brain function between adults and adolescents with respect to cognitive control, however, remain unclear. To address this issue, we conducted a study in which 185 adolescents (mean age (SD) 14.6 (0.3) years) and 28 adults (mean age (SD) 25.2 (6.3) years) performed a single task that included both a stimulus-response (S-R) interference component and a task-switching component. Behavioural responses (i.e. reaction time, RT; error rate, ER) and brain activity during correct, error and post-error trials, detected by functional magnetic resonance imaging (fMRI), were measured. Behaviourally, RT and ER were significantly higher in incongruent than in congruent trials and in switch than in repeat trials. The two groups did not differ in RT during correct trials, but adolescents had a significantly higher ER than adults. In line with similar RTs, brain responses during correct trials did not differ between groups, indicating that adolescents and adults engage the same cognitive control network to successfully overcome S-R interference or task switches. Interestingly, adolescents with stronger brain activation in the bilateral insulae during error trials and in fronto-parietal regions of the cognitive control network during post-error trials did have lower ERs. This indicates that those mid-adolescents who commit fewer errors are better at monitoring their performance, and after detecting errors are more capable of flexibly allocating further cognitive control resources. Although we did not detect a convincing neural correlate of the observed behavioural differences between adolescents and adults, the revealed interindividual differences in adolescents might at least in part be due to brain development.

Patients with schizophrenia and their siblings typically show subtle changes of brain structures, such as a reduction of hippocampal volume. Hippocampal volume is heritable, may explain a variety of cognitive symptoms of schizophrenia and is thus considered an intermediate phenotype for this mental illness. The aim of our analyses was to identify single-nucleotide polymorphisms (SNP) related to hippocampal volume without making prior assumptions about possible candidate genes. In this study, we combined genetics, imaging and neuropsychological data obtained from the Mind Clinical Imaging Consortium study of schizophrenia (n = 328). A total of 743,591 SNPs were tested for association with hippocampal volume in a genome-wide association study. Gene expression profiles of human hippocampal tissue were investigated for gene regions of significantly associated SNPs. None of the genetic markers reached genome-wide significance. However, six highly correlated SNPs (rs4808611, rs35686037, rs12982178, rs1042178, rs10406920, rs8170) on chromosome 19p13.11, located within or in close proximity to the genes NR2F6, USHBP1, and BABAM1, as well as four SNPs in three other genomic regions (chromosome 1, 2 and 10) had p-values between 6.75×10(-6) and 8.3×10(-7). Using existing data of a very recently published GWAS of hippocampal volume and additional data of a multicentre study in a large cohort of adolescents of European ancestry, we found supporting evidence for our results. Furthermore, allelic differences in rs4808611 and rs8170 were highly associated with differential mRNA expression in the cis-acting region. Associations with memory functioning indicate a possible functional importance of the identified risk variants. Our findings provide new insights into the genetic architecture of a brain structure closely linked to schizophrenia. In silico replication, mRNA expression and cognitive data provide additional support for the relevance of our findings. Identification of causal variants and their functional effects may unveil yet unknown players in the neurodevelopment and the pathogenesis of neuropsychiatric disorders.

Spatial predictions of near-surface air temperature ( T a i r ) in Antarctica are required as baseline information for a variety of research disciplines. Since the network of weather stations in Antarctica is sparse, remote sensing methods have large potential due to their capabilities and accessibility. Based on the MODIS land surface temperature (LST) data, T a i r at the exact time of satellite overpass was modelled at a spatial resolution of 1 km using data from 32 weather stations. The performance of a simple linear regression model to predict T a i r from LST was compared to the performance of three machine learning algorithms: Random Forest (RF), generalized boosted regression models (GBM) and Cubist. In addition to LST, auxiliary predictor variables were tested in these models. Their relevance was evaluated by a Cubist-based forward feature selection in conjunction with leave-one-station-out cross-validation to reduce the impact of spatial overfitting. GBM performed best to predict T a i r using LST and the month of the year as predictor variables. Using the trained model, T a i r could be estimated with a leave-one-station-out cross-validated R 2 of 0.71 and a RMSE of 10.51 ∘ C. However, the machine learning approaches only slightly outperformed the simple linear estimation of T a i r from LST ( R 2 of 0.64, RMSE of 11.02 ∘ C). Using the trained model allowed creating time series of T a i r over Antarctica for 2013. Extending the training data by including more years will allow developing time series of T a i r from 2000 on.

Saturated steam decontamination is an application for elimination of microorganisms from the surface of different materials. This technique has been optimized for the treatment of dried spices or pharmaceuticals, which could have been contaminated with microorganisms during cultivation, processing, storage or transport. The described saturated steam decontamination is based on the Lemgo process. This method does not kill microorganisms, but removes them physically from the surface. Our investigation focused on measuring the effects of steam temperatures at 120 °C and 100 °C, respectively, for 20 s with a subsequent fl ash vacuum of 20 s. Applications of fl ash vacuum as well as saturated steam heated to 120 °C were also tested separately. The impact of these parameters on the essential oil content and on the surface of different medicinal plants such as marjoram, oregano, fennel and eucalyptus was analysed using gas chromatography and scanning electron microscopy. Especially in herbal drugs with glandular trichomes such as marjoram and oregano severe surface destruction was visible accompanied by high losses of essential oil from 93 % in marjoram tissue to 59 % in oregano tissue. For fennel and eucalyptus that possess protected essential oil storage cells only minor or no reduction of volatiles has been observed during exposure to saturated steam. The experiments show clearly a positive correlation between stability of essential oil cavities and essential oil content preservation.

Im heutigen visuell dominierten Zeitalter prägen und formen visuelle (Re-)Präsentationen die Wahrnehmung und Aneignung von Regionen massgeblich. Bilder vermitteln Vorstellungen davon, welche Beziehungen zu einem bestimmten Raum eingenommen werden können und sollen. Sie zeigen, was sich in den Regionen tun lässt, welche Raumnutzungen vorhanden, möglich und erwünscht sind. In diesem Artikel wird eine Methode vorgestellt, durch die die potenzielle Raumwirksamkeit visueller Bilder erfasst werden kann. Sie ermöglicht, bildreiche Präsentationen von «Räumen» in ein übersichtliches Gesamtbild zu übersetzen und so unterschiedliche, unter Umständen in Konflikt zueinander stehende Raumaneignungsweisen herauszuarbeiten. Demonstriert wird die Methode anhand unterschiedlicher Publikationen über die UNESCO Biosphäre Entlebuch. Die Analyse offenbart die Differenz zwischen alltagsnaher Innensicht und romantisierender Aussensicht und dokumentiert einen im Ansatz unkonventionellen, Klischees aufbrechenden touristischen Auftritt.