Search

ABSTRACT: During processing and storage of both conventional and lactose-hydrolyzed UHT milk (LHM), aggregation of milk proteins occurs. Protein aggregation can inter alia occur via nonreducible covalent cross-links derived from either Maillard or dehydroalanine (DHA) pathways. To study this further in relation to processing method and lactase enzyme purity, LHM was produced using 3 different lactase preparations, with lactase enzymes added in a dairy setting either before (prehydrolysis) or after (posthydrolysis) UHT treatment. The prepared LHM types were subsequently stored at either 25°C or 35°C for up to one year. Mass spectrometry was used to absolutely quantify the level of furosine, N-ε-(carboxymethyl)lysine, N-ε-(carboxyethyl)lysine, lanthionine, and lysinoalanine in these products using a newly developed method on triple quadrupole for these processing-induced markers. The results showed higher levels of Maillard-related processing markers in prehydrolyzed LHM compared with posthydrolyzed LHM and conventional UHT milk, which, on the other hand, contained higher concentrations of DHA-derived cross-links. Proteomics of collected particles from asymmetrical flow field-flow fractionation in combination with gel electrophoresis indicated the presence of intramicellar cross-links during storage, especially for larger particles involving αS1- and αS2-caseins.

Abstract α-Synuclein (α-syn) accumulates as insoluble amyloid but also forms soluble α-syn oligomers (αSOs), thought to be even more cytotoxic than fibrils. To detect and block the unwanted activities of these αSOs, we have raised 30 monoclonal antibodies (mAbs) against different forms of αSOs, ranging from unmodified αSOs to species stabilized by lipid peroxidation products and polyphenols, αSOs formed by C-terminally truncated α-syn, and multivalent display of α-syn on capsid virus-like particles (cVLPs). While the mAbs generally show a preference for αSOs, they also bind fibrils, but to variable extents. Overall, we observe great diversity in the mAbs’ relative affinities for monomers and αSOs, varied requirements for the C-terminal extension of α-syn, and only a modest effect on α-syn fibrillation. Several mAbs show several orders of magnitude preference for αSOs over monomers in in-solution studies, while the commercial antibody MJF14 only bound 10-fold more strongly to αSOs than monomeric α-syn. Gratifyingly, seven mAbs almost completely block αSO permeabilization of membrane vesicles. Five selected mAbs identified α-syn-related pathologies like Lewy bodies (LBs) and Lewy Neurites, as well as Glial Cytoplasmic Inclusions in postmortem brains from people diagnosed for PD, dementia with LBs or multiple system atrophy, although to different extents. Three mAbs were particularly useful for pathological evaluation of postmortem brain human tissue, including early stages of PD. Although there was no straightforward connection between the mAbs’ biophysical and immunohistochemical properties, it is encouraging that this comprehensive collection of mAbs able to recognize different aggregated α-syn species in vitro also holds diagnostic potential.

The role of organic compounds in atmospheric new particle formation is difficult to disentangle due to the myriad of potentially important oxygenated organic molecules (OOMs) present in the atmosphere. Using state-of-the-art quantum chemical methods, we here employ a novel approach, denoted the “cluster-of-functional-groups” approach, for studying the involvement of OOMs in atmospheric cluster formation. Instead of the usual “trial-and-error” approach of testing the ability of experimentally identified OOMs to form stable clusters with other nucleation precursors, we here study which, and how many, intermolecular interactions are required in a given OOM to form stable clusters. In this manner we can reverse engineer the elusive structure of OOM candidates that might be involved in organic enhanced atmospheric cluster formation. We calculated the binding free energies of all combinations of donor and acceptor organic functional groups to investigate which functional groups most preferentially bind with each other and with other nucleation precursors such as sulfuric acid and bases (ammonia, methyl-, dimethyl- and trimethylamine). We find that multiple carboxyl groups lead to substantially more stable clusters compared to all other combinations of functional groups. Employing cluster dynamics simulations, we investigate how a hypothetically OOM composed of multiple carboxyl groups can stabilize sulfuric acid–base clusters and provide recommendations for potential atmospheric multi-carboxylic acid tracer compounds that should be explicitly studied in the future. The presented “cluster-of-functional-groups” approach is generally applicable and can be employed in many other applications, such as ion-induced nucleation and potentially in elucidating the structural patterns in molecules that facilitate ice nucleation.

The tick-borne encephalitis virus (TBEV) is transmitted to humans through tick bites. In recent years, the appearance of the Siberian subtype of TBEV in Ixodes ricinus in Finland, together with deaths from the normally mild European subtype in the same country, have raised concerns about a possible spread of virulent variants of TBEV in Western Europe. Thus, there is a need to monitor the spread of strains, particularly of the European and Siberian subtypes. In this study, we develop a new real-time PCR method targeting Siberian and European subtypes of TBEV. The primers amplify a 176 bp fragment of the E gene, which is suitable for subsequent strain identification by Sanger sequencing. This study pioneers a new approach to primer design where the melting temperature (Tm) of primers annealed to representative mismatched target sequences is empirically determined and used to guide improvements in primer sequence. This allowed the range of TBEV strains detected to be extended to cover most European and Siberian strains tested, in addition to a strain of the Far-Eastern subtype. The limit of detection was 10–100 DNA copies per reaction and amplification efficiency varied between 83% and 94%, depending on the TBEV strain. Experimental determination of primer Tm proved to be a fruitful approach and will be a useful tool for future primer design and diagnostics.

Simulation models are widely used in urban drainage engineering and research, but they are known to include errors and uncertainties that are not yet fully realised. Within the herein developed framework, we investigate model adequacy across multiple sites by comparing model results with measurements for three model objectives, namely surcharges (water level rises above defined critical levels related to basement flooding), overflows (water levels rise above a crest level), and everyday events (water levels stay below the top of pipes). We use multi-event hydrological signatures, i.e. metrics that extract specific characteristics of time series events in order to compare model results with the observations for the mentioned objectives through categorical and statistical data analyses. Furthermore, we assess the events with respect to sufficient or insufficient categorical performance and good, acceptable, or poor statistical performance. We also develop a method to reduce the weighting of individual events in the analyses, in order to acknowledge uncertainty in model and/or measurements in cases where the model is not expected to fully replicate the measurements. A case study including several years of water level measurements from 23 sites in two different areas shows that only few sites score a sufficient categorical performance in relation to the objective overflow and that sites do not necessarily obtain good performance scores for all the analysed objectives. The developed framework, however, highlights that it is possible to identify objectives and sites for which the model is reliable, and we also suggest methods for assessing where the model is less reliable and needs further improvement, which may be further refined in the future.

Abstract The time delay from an interplanetary driver arriving at the magnetopause to the response in the ionosphere has never been quantified separately for different types of storm drivers. This study investigates the delay for storms driven by high‐speed streams and associated stream interaction regions (HSS/SIR), or by interplanetary coronal mass ejection sheaths and magnetic clouds (MC). The total field‐aligned current (FAC) and SME index lag the Newell coupling function (NCF) by 40 ± 10 min during storms driven by HSS/SIR and sheaths, and by 60 ± 10 min for MCs. The correlation coefficient between FAC and NCF reaches maximum value as NCF is averaged over the preceding 80 min for sheath, 90 min for HSS/SIR, and 140 min for MC storms.

Digital twins of urban drainage systems require simulation models that can adequately replicate the physical system. All models have their limitations, and it is important to investigate when and where simulation results are acceptable and to communicate the level of performance transparently to end users. This paper first defines a classification of four possible ‘locations of uncertainty’ in integrated urban drainage models. It then develops a structured framework for identifying and diagnosing various types of errors. This framework compares model outputs with in-sewer water level observations based on hydrologic and hydraulic signatures. The approach is applied on a real case study in Odense, Denmark, with examples from three different system sites: a typical manhole, a small flushing chamber, and an internal overflow structure. This allows diagnosing different model errors ranging from issues in the underlying asset database and missing hydrologic processes to limitations in the model software implementation. Structured use of signatures is promising for continuous, iterative improvements of integrated urban drainage models. It also provides a transparent way to communicate the level of model adequacy to end users. HIGHLIGHTS Transparency of the performance of simulation models in digital twins is needed.; Observation data can indicate that reality is different from what is perceived as true.; Signatures are strong tools to diagnose model errors.;

Introduction Insomnia in depression is common and difficult to resolve. Music is commonly used as a sleep aid, and clinical trials pointing to positive effects of music as a sleep aid are increasing adding to the evidence base. There is little knowledge on the effectiveness of music for depression related insomnia. Objectives A recent RCT study conducted in psychiatry at Aalborg University Hospital examined effects of a music intervention for insomnia in depression. The intervention group listened to music at bedtime for four weeks, controls were offered music intervention post-test. Primary outcome measure was Pittsburgh Sleep Quality Index (PSQI). Secondary outcomes included Actigraphy, The Hamilton depression Rating Scale (HAMD-17) and World Health Organisation well-being questionnaires (WHO-5, WHOQOL-BREF). Methods A two-armed randomized controlled trial (n=112) and a qualitative interview study (n=4) Results The RCT study showed signficant improvements for the music intervention group in sleep quality and quality of life at four weeks according to global PSQI scores (effect size= -2.1, 95%CI -3.3; -0.9) and WHO-5 scores (effect size 8.4, 95%CI 2.7; 14.0). Actigraphy measures showed no changes and changes in depression symptoms (HAMD-17) were not detected. The interview study unfolded examples of the influences of music on sleep and relaxation. Music distracted, affected mood and arousal positively and supported formation of sleep habits. Results from the trial are discussed and merged with findings from the interview study. The results from the trial suggested moderate effects of music listening for the population while findings from the interview study showed examples of individual and highly varying outcomes. Conclusions Music is suggested as a low-cost, side-effect free and safe intervention in supplement to existing treatments improving sleep in depression. Disclosure of Interest None Declared

Abstract Background Class 1 cytokine receptors (C1CRs) are single-pass transmembrane proteins responsible for transmitting signals between the outside and the inside of cells. Remarkably, they orchestrate key biological processes such as proliferation, differentiation, immunity and growth through long disordered intracellular domains (ICDs), but without having intrinsic kinase activity. Despite these key roles, their characteristics remain rudimentarily understood. Methods The current paper asks the question of why disorder has evolved to govern signaling of C1CRs by reviewing the literature in combination with new sequence and biophysical analyses of chain properties across the family. Results We uncover that the C1CR-ICDs are fully disordered and brimming with SLiMs. Many of these short linear motifs (SLiMs) are overlapping, jointly signifying a complex regulation of interactions, including network rewiring by isoforms. The C1CR-ICDs have unique properties that distinguish them from most IDPs and we forward the perception that the C1CR-ICDs are far from simple strings with constitutively bound kinases. Rather, they carry both organizational and operational features left uncovered within their disorder, including mechanisms and complexities of regulatory functions. Conclusions Critically, the understanding of the fascinating ability of these long, completely disordered chains to orchestrate complex cellular signaling pathways is still in its infancy, and we urge a perceptional shift away from the current simplistic view towards uncovering their full functionalities and potential. Video abstract

Exosomes are used as disease biomarkers, but their characterization in biological samples is challenging. Here the authors achieve simultaneous characterization of size and zeta potential of individual nanoparticles and particle mixtures at physiological salinity conditions, exploiting a salt gradient in a capillary channel.

Abstract Background Candidatus Neoehrlichia mikurensis is an emerging tick-borne pathogen. It is widely distributed in Ixodes ricinus ticks in Europe, but knowledge of its distribution in Norway, where I. ricinus reaches its northern limit, is limited. In this study we have developed a real time PCR test for Ca. N. mikurensis and used it to investigate the distribution of Ca. N. mikurensis in Norway. Results Real time PCR targeting the groEL gene was developed and shown to be highly sensitive. It was used to detect Ca. N. mikurensis in 1651 I. ricinus nymphs and adults collected from twelve locations in Norway, from the eastern Oslo Fjord in the south to near the Arctic Circle in the north. The overall prevalence was 6.5% and varied locally between 0 and 16%. Prevalence in adults and nymphs was similar, suggesting that ticks acquire Ca. N. mikurensis predominantly during their first blood meal. In addition, 123 larvae were investigated; Ca. N. mikurensis was not found in larvae, suggesting that transovarial transmission is rare or absent. Sequence analysis suggests that a single variant dominates in Norway. Conclusions Ca. N. mikurensis is widespread and common in ticks in Norway and reaches up to their northern limit near the Arctic Circle. Ticks appear to acquire Ca. N. mikurensis during their first blood meal. No evidence for transovarial transmission was found.

Flaviviruses are a threat to public health and can cause major disease outbreaks. Tick-borne encephalitis (TBE) is caused by a flavivirus, and it is one of the most important causes of viral encephalitis in Europe and is on the rise in Sweden. As there is no antiviral treatment available, vaccination remains the best protective measure against TBE. Currently available TBE vaccines are based on formalin-inactivated virus produced in cell culture. These vaccines must be delivered by intramuscular injection, have a burdensome immunization schedule, and may exhibit vaccine failure in certain populations. This project aimed to develop an edible TBE vaccine to trigger a stronger immune response through oral delivery of viral antigens to mucosal surfaces. We demonstrated successful expression and post-translational processing of flavivirus structural proteins which then self-assembled to form virus-like particles in Nicotiana benthamiana. We performed oral toxicity tests in mice using various plant species as potential bioreactors and evaluated the immunogenicity of the resulting edible vaccine candidate. Mice immunized with the edible vaccine candidate did not survive challenge with TBE virus. Interestingly, immunization of female mice with a commercial TBE vaccine can protect their offspring against TBE virus infection.

Mesoscopic environments and particles diffusing in them are often studied by tracking such particles individually while their Brownian motion explores their environment. Environments may be, e.g., a domain in a cell membrane, an interior compartment of a cell, or an engineered nanopit. Particle trajectories are typically determined from time-lapse recorded movies. These are recorded with sufficient exposure time per frame to be able to detect and localize particles in each frame. Since particles move during this exposure time, particles image with motion blur. This motion blur can compromise estimates of diffusion coefficients and the size of the confining domain if not accounted for correctly. We do that here. We give explicit and exact expressions for the variance of measured positions and the mean-squared displacement of a Brownian particle confined in, respectively, a 1D box, a 2D box, a 2D circular disc, and a 3D sphere. Our expressions are valid for all exposure times, irrespective of the size of the confining space and the value of the diffusion coefficient. They apply also in the common case where the exposure time is smaller than the time-lapse due, e.g., to “dead time” caused by the readout process in the camera. These expressions permit determination of diffusion coefficients and domain sizes for given movies for the simple geometries we consider. More important, the trends observed in our exact results when parameter values are varied are valid also for more complex geometries for which no exact analytical solutions exist. Wherever the underlying physics is the same, the exact quantitative description of its consequences provided here is portable as a qualitative and semi-quantitative understanding of its consequences in general. The results may also be useful for other types of reflected Brownian motion than those occurring in single-particle tracking, e.g., in nuclear magnetic resonance imaging techniques. For use in that particular context, we briefly discuss the effects of confinement on anisotropic Brownian motion imaged with motion blur.

Scanning in football (soccer) denotes an active head movement where a player’s face is temporarily directed away from the ball to gather information in preparation for subsequently engaging with the ball. The aim of this study was to learn more about the ways that 27 elite professional football players in an English Premier League club use scanning in competitive matches, the conditions under which this behavior is exhibited, and the relationships between these behaviors and performance. Players were filmed across 21 matches, producing a total number of 9,574 individual ball possessions for analysis. Close-up video analyses of scanning show positional differences (with central midfielders and central defenders scanning most frequently, forwards least) and contextual differences (with relatively lower scanning frequency in situations with tight opponent pressure, in positions wide in the field and closer to the opponent’s goal, and under certain game state conditions). Players scan more frequently prior to giving passes than when they dribble, shoot, or only receive it, as well as prior to more long/forward passes compared to short/backward ones, although these differences are small. A Bayesian hierarchical model, which accounts for individual player differences and pass difficulty, suggests that the more a player scans, the higher the probability of completing a pass. In conclusion, match demands are likely to constrain the extent to which highly elite players scan, and scanning seems to have a small, but positive role in elite football players’ performance.

Coronary artery disease (CAD) is an immune-mediated disease in which CCR2 attracts classical, intermediate, and non-classical monocytes to the arterial intima where they differentiate to macrophages. Balance between pro-inflammatory M1 and anti-inflammatory M2 macrophages contributes to CAD prevention. Moderate to vigorous intensity physical activity (MVPA) elicits an immune response and reduces the incidence of CAD, however, the impact of prior MVPA on monocyte subset CCR2 expression and macrophage polarization following acute exercise is unknown. Purpose: To determine the impact of physical activity status on monocyte subset CCR2 surface expression and macrophage polarization in response to an acute bout of moderate intensity cycle ergometry. Methods: 24 healthy women and men (12 high physically active [HIACT]: ≥1500 METmin/wk MVPA & 12 low physically active [LOACT]:

Purpose: This preliminary study explores Ugandan men’s knowledge and attitudes about human papillomavirus (HPV), cervical cancer, and screening. Methods: A local physician led an education session about cervical cancer for 62 men in Kisenyi, Kampala in Uganda. Trained nurse midwives administered surveys to assess knowledge and attitudes before and after the education session. Results: From the pre-education survey, only 24.6% of men had heard of HPV previously, and 59% of men had heard of cervical cancer. Posteducation, 54.5% of men believed only women could be infected with HPV and 32.7% of men believed antibiotics could cure HPV. Despite their limited knowledge, 98.2% of men stated they would support their partners to receive screening for cervical cancer, and 100% of men surveyed stated they would encourage their daughter to get the HPV vaccine if available. Conclusions: Knowledge of HPV and cervical cancer among Ugandan men is low. Even after targeted education, confusion remained about disease transmission and treatment. Ongoing education programs geared toward men and interventions to encourage spousal communication about reproductive health and shared decision making may improve awareness of cervical cancer prevention strategies.

Mutations in potassium and calcium channel genes have been associated with cardiac arrhythmias. Here, Jensen et al. show that an anion transporter chloride-bicarbonate exchanger AE3 is also responsible for the genetically-induced mechanism of cardiac arrhythmia, suggesting new therapeutic targets for this disease

Birds are important hosts for the first life stages of the Ixodes ricinus tick and they can transport their parasites over long distances. The aim of this study was to investigate the prevalence of Borrelia burgdorferi sensu lato, Anaplasma phagocytophilum, Neoehrlichia mikurensis and Rickettsia helvetica in ticks collected from migratory birds in Norway. A total of 815 Ixodes ricinus ticks from 216 birds trapped at Lista Bird Observatory in southern Norway during spring and autumn migration in 2008 were analysed by real-time PCR. B. burgdorferi s. l. was the most prevalent pathogen, detected in 6.1% of the ticks. The prevalence of N. mikurensis, A. phagocytophilum and R. helvetica was 1.2%, 0.9% and 0.4% respectively. In addition, one sample (0.1%) was positive for B. miyamotoi. In total, 8.2% of the ticks were infected with at least one pathogen. Co-infection with B. burgdorferi s. l. and N. mikurensis or A. phagocytophilum was found in 6.0% of the infected ticks. Our results show that all the known major tick-borne bacterial pathogens in Norway are subject to transport by migratory birds, potentially allowing spread to new areas. Our study showed a surprisingly high number of samples with PCR inhibition (57%). These samples had been extracted using standard methodology (phenol-chloroform extraction). This illustrates the need for inhibition controls to determine true prevalence rates.

We have studied strain wave generation in graphite induced by an intense ultrashort laser pulse. The study was performed in the intensity regime above the ablation threshold of graphite. The aim was to maximize the strain and, thus, also the internal pressure (stress). Laser pulses with a 1 ps temporal duration melt the surface of graphite resulting in a molten material which initially exists at the solid density. As the molten material expands, a compressive strain wave starts propagating into the crystal below the molten layer. The strain pulse was studied with time-resolved X-ray diffraction. At a temporal delay of 100 ps after laser excitation, we observed >10% compressive strain, which corresponds to a pressure of 7.2 GPa. This strain could be reproduced by hydrodynamic simulations, which also provided a temperature map as a function of time and depth.

Abstract Background High-risk HPV DNA testing has been proposed as a primary tool for cervical cancer screening (HPV-CCS) as an alternative to the Papanicolaou cytology- method. This study describes factors associated with women’s intentions to attend cervical cancer screening if high-risk HPV DNA testing (HPV-CCS) was implemented as a primary screening tool, and if screening were conducted every 4 years starting after age 25. Methods This online survey was designed using the Theory of Planned Behaviour to assess factors that impact women’s intentions to attend HPV-CCS among women aged 25–69 upon exit of the HPV FOCAL trial. Univariate and regression analyses were performed to compare the demographic, sexual history, and smoking characteristics between women willing and unwilling to screen, and scales for intention to attend HPV-CCS. A qualitative analysis was performed by compiling and coding the comments section of the survey. Results Of the 981 women who completed the survey in full, only 51.4 % responded that they intended to attend HPV-CCS with a delayed start age and extended screening interval. Women who intended to screen were more likely to have higher education (AOR 0.59, 95 % CI [0.37, 0.93]), while both positive attitudes (AOR 1.26, 95 % CI [1.23, 1.30]) and perceived behavior control (AOR 1.06, 95 % CI [1.02, 1.10]) were significant predictors of intention to screen. Among women who provided comments in the survey, a large number of women expressed fears about not being checked more than every 4 years, but 12 % stated that these fears may be alleviated by having more information. Conclusions Acceptability of increased screening intervals and starting age could be improved through enhanced education of benefits. Program planners should consider measures to assess and improve women’s knowledge, attitudes and beliefs prior to the implementation of new screening programs to avoid unintended consequences.

Putative heterotrophic bacteria carrying out N2-fixation, so-called non-cyanobacterial diazotrophs (NCDs), are widely distributed in marine waters, but details of how the O2-inhibited N2-fixation process is promoted in the oxic water column remains ambiguous. Here we carried out two experiments with water from a eutrophic temperate fjord to examine whether low-oxygen microenvironments within particulate organic matter could be loci suitable for N2-fixation. First, water enriched with natural particles or sediment showed higher N2-fixation rates than bulk water, and nitrogenase genes (nifH) revealed that specific diazotrophs were affiliated with the particulate matter. Second, pristine artificial surfaces were rapidly colonized by diverse bacteria, while putative diazotrophs emerged relatively late (after 80 h) during the colonization, and phylotypes related to Pseudomonas and to anaerobic bacteria became dominant with time. Our study pinpoints natural particles as sites of N2-fixation, and indicates that resuspension of sediment material can elevate pelagic N2-fixation. Moreover, we show that diverse natural diazotrophs can colonize artificial surfaces, but colonization by “pioneer” bacterioplankton that more rapidly associate with surfaces appears to be a prerequisite. Whereas our experimental study supports the idea of pelagic particles as sites of N2-fixation by heterotrophic bacteria, future in situ studies are needed in order to establish identity, activity and ecology of particle associated NCDs as a function of individual particle characteristics.

After publication of our article [1] it came to our notice that the source of the sequence for the control plasmid, pNeo (Materials and methods: Controls) was incorrectly stated as AB094461. The correct accession number is AB074461. The authors apologize for any confusion this may have caused.

The purpose of this systematic review is to assess the evidence behind the dietary requirement of protein and to assess the health effects of varying protein intake in healthy elderly persons in order to evaluate the evidence for an optimal protein intake. The literature search covered year 2000–2011. Prospective cohort, case–control, and intervention studies of a general healthy population in settings similar to the Nordic countries with protein intake from food-based sources were included. Out of a total of 301 abstracts, 152 full papers were identified as potentially relevant. After careful scrutiny, 23 papers were quality graded as A (highest, n=1), B (n=18), or C (n=4). The grade of evidence was classified as convincing, probable, suggestive, or inconclusive. The evidence is assessed as: probable for an estimated average requirement (EAR) of 0.66 g good-quality protein/kg body weight (BW)/day based on nitrogen balance (N-balance) studies and the subsequent recommended dietary allowance (RDA) of 0.83 g good-quality protein/kg BW/day representing the minimum dietary protein needs of virtually all healthy elderly persons. Regarding the optimal level of protein related to functional outcomes like maintenance of bone mass, muscle mass, and strength, as well as for morbidity and mortality, the evidence is ranging from suggestive to inconclusive. Results from particularly prospective cohort studies suggest a safe intake of up to at least 1.2–1.5 g protein/kg BW/day or approximately 15–20 E%. Overall, many of the included prospective cohort studies were difficult to fully evaluate since results mainly were obtained by food frequency questionnaires that were flawed by underreported intakes, although some studies were ‘calibrated’ to correct for under- or over-reporting. In conclusion, the evidence is assessed as probable regarding the EAR based on N-balance studies and suggestive to inconclusive regarding an optimal protein intake higher than the estimated RDA assessed from N-balance studies, but an exact level cannot be determined. Potentially adverse effects of a protein intake exceeding 20–23 E% remain to be investigated.

The purpose of this systematic review is to assess the evidence behind the dietary requirement of protein and to assess the health effects of varying protein intake in healthy adults. The literature search covered the years 2000–2011. Prospective cohort, case-control, and intervention studies were included. Out of a total of 5,718 abstracts, 412 full papers were identified as potentially relevant, and after careful scrutiny, 64 papers were quality graded as A (highest), B, or C. The grade of evidence was classified as convincing, probable, suggestive or inconclusive. The evidence is assessed as: probable for an estimated average requirement of 0.66 g good-quality protein/kg body weight (BW)/day based on nitrogen balance studies, suggestive for a relationship between increased all-cause mortality risk and long-term low-carbohydrate–high-protein (LCHP) diets; but inconclusive for a relationship between all-cause mortality risk and protein intake per se; suggestive for an inverse relationship between cardiovascular mortality and vegetable protein intake; inconclusive for relationships between cancer mortality and cancer diseases, respectively, and protein intake; inconclusive for a relationship between cardiovascular diseases and total protein intake; suggestive for an inverse relationship between blood pressure (BP) and vegetable protein; probable to convincing for an inverse relationship between soya protein intake and LDL cholesterol; inconclusive for a relationship between protein intake and bone health, energy intake, BW control, body composition, renal function, and risk of kidney stones, respectively; suggestive for a relationship between increased risk of type 2 diabetes (T2D) and long-term LCHP-high-fat diets; inconclusive for impact of physical training on protein requirement; and suggestive for effect of physical training on whole-body protein retention. In conclusion, the evidence is assessed as probable regarding the estimated requirement based on nitrogen balance studies, and suggestive to inconclusive for protein intake and mortality and morbidity. Vegetable protein intake was associated with decreased risk in many studies. Potentially adverse effects of a protein intake exceeding 20–23 E% remain to be investigated.

Foodborne Enteropathogenic Escherichia coli (EPEC) infections of the small intestine cause diarrhea especially in children and are a major cause of childhood death in developing countries. EPEC infects the apical membrane of the epithelium of the small intestine by attaching, effacing the microvilli under the bacteria and then forming microcolonies on the cell surface. We first asked the question where on epithelial cells EPEC attaches and grows. Using models of polarized epithelial monolayers, we evaluated the sites of initial EPEC attachment to the apical membrane and found that EPEC preferentially attached over the cell-cell junctions and formed microcolonies preferentially where three cells come together at tricellular tight junctions. The ability of EPEC to adhere increased when host cell polarity was compromised yielding EPEC access to basolateral proteins. EPEC pedestals contain basolateral cytoskeletal proteins. Thus, we asked if attached EPEC causes reorganization the protein composition of the host cell plasma membrane at sites of microcolony formation. We found that EPEC microcolony growth at the apical membrane resulted in a local accumulation of basolateral plasma membrane proteins surrounding the microcolony. Basolateral marker protein aquaporin-3 localized to forming EPEC microcolonies. Components of the basolateral vesicle targeting machinery were re-routed. The Exocyst (Exo70) was recruited to individual EPEC as was the basolateral vesicle SNARE VAMP-3. Moreover, several Rab variants were also recruited to the infection site, and their dominant-negative equivalents were not. To quantitatively study the recruitment of basolateral proteins, we created a pulse of the temperature sensitive basolateral VSVG, VSVG3-SP-GFP, from the trans-Golgi Network. We found that after release from the TGN, significantly more VSVG3-SP-GFP accumulated at the site of microcolony growth than on equivalent membrane regions of uninfected cells. This suggests that trafficking of vesicles destined for the basolateral membrane are redirected to the apical site of microcolony growth. Thus, in addition to disrupting host cell fence function, local host cell plasma membrane protein composition is changed by altered protein trafficking and recruitment of basolateral proteins to the apical microcolony. This may aid EPEC attachment and subsequent microcolony growth.

Introduction: Community engagement and education can improve acceptability and participation in clinical trials conducted in Kisenyi, Uganda. In preparation for a randomized controlled trial exploring different methods for cervical cancer screening, we explored optimal engagement strategies from the perspective of community members and health professionals.Methods: We conducted key informant interviews followed by serial community forums with purposeful sampling and compared the perspectives of women in Kisenyi (N=26) to healthcare workers (HCW) at the local and tertiary care center levels (N=61) in a participatory, iterative process.Results: Key themes identified included format, content, language, message delivery and target population. Women in Kisenyi see demonstration as a key part of an educational intervention and not solely a didactic session whereas health professionals emphasized the biomedical content and natural history of cervical cancer. Using local language and lay leaders with locally accessible terminology was more of a priority for women in Kisenyi than clinicians. Simple language with a clear message was essential for both groups. Localization of language and reciprocal communication using demonstration between community members and HCW was a key theme.Conclusions: Although perceptions of the format are similar between women and HCW, the content, language and messaging that should be incorporated in a health education strategy differ markedly. The call for lay leaders to participate in health promotion is a clear step towards transforming this cervical cancer screening project to be a fully participatory process. This is important in scaling-up cervical cancer screening programs in Kisenyi and will be central in developing health education interventions for this purpose.