A virulent strain of deformed wing virus (DWV) of honeybees (Apis mellifera) prevails after Varroa destructor-mediated, or in vitro, transmission.

The globally distributed ectoparasite Varroa destructor is a vector for viral pathogens of the Western honeybee (Apis mellifera), in particular the Iflavirus Deformed Wing Virus (DWV). In the absence of Varroa low levels DWV occur, generally causing asymptomatic infections. Conversely, Varroa-infested colonies show markedly elevated virus levels, increased overwintering colony losses, with impairment of pupal development and symptomatic workers. To determine whether changes in the virus population were due Varroa amplifying and introducing virulent virus strains and/or suppressing the host immune responses, we exposed Varroa-naïve larvae to oral and Varroa-transmitted DWV. We monitored virus levels and diversity in developing pupae and associated Varroa, the resulting RNAi response and transcriptome changes in the host. Exposed pupae were stratified by Varroa association (presence/absence) and virus levels (low/high) into three groups. Varroa-free pupae all exhibited low levels of a highly diverse DWV population, with those exposed per os (group NV) exhibiting changes in the population composition. Varroa-associated pupae exhibited either low levels of a diverse DWV population (group VL) or high levels of a near-clonal virulent variant of DWV (group VH). These groups and unexposed controls (C) could be also discriminated by principal component analysis of the transcriptome changes observed, which included several genes involved in development and the immune response. All Varroa tested contained a diverse replicating DWV population implying the virulent variant present in group VH, and predominating in RNA-seq analysis of temporally and geographically separate Varroa-infested colonies, was selected upon transmission from Varroa, a conclusion supported by direct injection of pupae in vitro with mixed virus populations. Identification of a virulent variant of DWV, the role of Varroa in its transmission and the resulting host transcriptome changes furthers our understanding of this important viral pathogen of honeybees.

MosaicSolver: a tool for determining recombinants of viral genomes from pileup data.

Viral recombination is a key evolutionary mechanism, aiding escape from host immunity, contributing to changes in tropism and possibly assisting transmission across species barriers. The ability to determine whether recombination has occurred and to locate associated specific recombination junctions is thus of major importance in understanding emerging diseases and pathogenesis. This paper describes a method for determining recombinant mosaics (and their proportions) originating from two parent genomes, using high-throughput sequence data. The method involves setting the problem geometrically and the use of appropriately constrained quadratic programming. Recombinants of the honeybee deformed wing virus and the Varroa destructor virus-1 are inferred to illustrate the method from both siRNAs and reads sampling the viral genome population (cDNA library); our results are confirmed experimentally. Matlab software (MosaicSolver) is available.

Assessing the quality of the ecological component of English Environmental Statements.

Environmental Impact Assessment (EIA) is a key tool to help ensure sustainable built development in more than 200 countries worldwide. Ecology is frequently a component of EIA and early reviews of Ecological Impact Assessment (EcIA) chapters identified scope for improvement at almost every stage of the EcIA process, regardless of country. However, there have been no reviews of UK EcIA chapters since 2000, despite important changes in biodiversity and planning legislation, policy and guidance. In addition, no UK EcIA chapter reviews have attempted to assign a grade or score to EcIA chapters (as has been done for reviews of US, Finnish and Indian EcIA chapters). Furthermore, no EcIA chapter reviews have attempted to use a scoring system to identify which variables determine EcIA chapter information content, beyond straightforward comparisons of EcIA chapters before and after the introduction of guidelines. A variant of the Biodiversity Assessment Index (BAI) was used to assign scores between zero and one to EcIA chapters based on a series of 47 questions drawn from EU legislation and professional guidance. 112 EcIA chapters for proposed developments that were subsequently granted planning permission in England were assessed. The mean BAI score was less than 0.5, indicating the presence of considerable information gaps in the majority of EcIA chapters. Of 13 predictor variables identified as having the potential to affect EcIA chapter quality, 10 were identified as significantly related to the BAI scores. A backward stepwise Generalized Linear Model identified the use of professional guidance, the ecological consultancy type and the length of the EcIA chapter as having the greatest combined explanatory power. As a result, several recommendations are made to help improve future EcIA chapter content, including formal EcIA chapter review, publicising the professional guidance to consultants, the provision of training and the introduction of an accreditation scheme for consultants involved in EcIA This approach could be replicated in other countries that conduct EIA. Context-dependent EcIA chapter review criteria (as in this paper) would help to identify targeted recommendations for improvement. Alternatively, a global set of review criteria could highlight areas of best practice that could then be exported to other countries.

Generalizing a model beyond the inherence heuristic and applying it to beliefs about objective value.

The inherence heuristic is characterized as part of an instantiation of a more general model that describes the interaction between undeveloped intuitions, produced by System 1 heuristics, and developed beliefs, constructed by System 2 reasoning. The general model is described and illustrated by examining another instantiation of the process that constructs belief in objective moral value.

Prevalence, risk factors, and effects of performance-related medical disorders (PRMD) among tertiary-trained jazz pianists in Australia and the United States.

This study explores performance-related medical disorders (PRMD) among a sample of tertiary-trained jazz pianists. Participants included both Australian and US pianists (n=214), including current and former tertiary students, professional pianists, and teachers. This mixed methods (quantitative and qualitative) exploratory and descriptive study used survey and case studies to provide baseline data for further research. Students reported a past and present period prevalence of 63% for pain and 41% for PRMD (injury attributed to practice or performance) with the forearm being the body part most affected, usually by fatigue. Diagnosis and treatment were reported as often unsatisfactory mainly due to lack of knowledge of PRMD by teachers and by professional health providers, and also to limited access to specialist PRMD services where these exist. Although teacher knowledge of PRMD is quite low, students still seek advice primarily from their teachers. The current study highlights a need to address the issue of PRMD among jazz pianists and their teachers more strategically, both in its prevention and in diagnosis and treatment.

A Closed Cavity Ultrasonic Resonator Formed by Graphene/PMMA Membrane for Acoustic Application.

A graphene/poly(methyl methacrylate) (PMMA) closed cavity resonator with a resonant frequency at around 160 kHz has been fabricated. A six-layer graphene structure with a 450 nm PMMA laminated layer has been dry-transferred onto the closed cavity with an air gap of 105 µm. The resonator has been actuated in an atmosphere and at room temperature by mechanical, electrostatic and electro-thermal methods. The (1,1) mode has been observed to dominate the resonance, which suggests that the graphene/PMMA membrane has been perfectly clamped and seals the closed cavity. The degree of linearity of the membrane's displacement versus the actuation signal has been determined. The resonant frequency has been observed to be tuned to around 4% by applying an AC voltage through the membrane. The strain has been estimated to be around 0.08%. This research puts forward a graphene-based sensor design for acoustic sensing.

Deep aspirations: towards a sustainable offshore Blue Economy.

Abstract: The ocean economy is experiencing rapid growth that will provide benefits but will also pose environmental and social risks. With limited space and degraded resources in coastal areas, offshore waters will be a particular focus of Blue Economy expansion over the next decade. When emerging and established economic sectors expand in offshore waters (within national Exclusive Economic Zones), different potential Blue Economy opportunities and challenges will arise. Following a series of interdisciplinary workshops, we imagine two technically possible futures for the offshore Blue Economy and we identify the actions required to achieve the more sustainable outcome. Under a business as usual scenario the focus will remain on economic growth, the commodification of nature, the dominance of private over public and cultural interests, and prioritisation of the interests of current over future generations. A more sustainable scenario would meet multiple UN Sustainable Development Goals and ensure inclusive economic developments, environmental sustainability, and fair and equitable access to resources and technologies across users, nations, and generations. Challenges to this more sustainable future are a lack of infrastructure and technology to support emerging offshore sectors, limited understanding of environmental impacts, inequitable outcomes, and a lack of planning and governmental oversight. Addressing these challenges will require a shift in societal values, a more balanced allocation of funding to offshore activities, transparency in information sharing between industries and across nations, and adjustment of international legal and institutional mechanisms. The sustainable and equitable offshore Blue Economy we envisage is achievable and provides a unique opportunity to build global capacity and partnership.

Connecting to the oceans: supporting ocean literacy and public engagement.

Improved public understanding of the ocean and the importance of sustainable ocean use, or ocean literacy, is essential for achieving global commitments to sustainable development by 2030 and beyond. However, growing human populations (particularly in mega-cities), urbanisation and socio-economic disparity threaten opportunities for people to engage and connect directly with ocean environments. Thus, a major challenge in engaging the whole of society in achieving ocean sustainability by 2030 is to develop strategies to improve societal connections to the ocean. The concept of ocean literacy reflects public understanding of the ocean, but is also an indication of connections to, and attitudes and behaviours towards, the ocean. Improving and progressing global ocean literacy has potential to catalyse the behaviour changes necessary for achieving a sustainable future. As part of the Future Seas project (https://futureseas2030.org/), this paper aims to synthesise knowledge and perspectives on ocean literacy from a range of disciplines, including but not exclusive to marine biology, socio-ecology, philosophy, technology, psychology, oceanography and human health. Using examples from the literature, we outline the potential for positive change towards a sustainable future based on knowledge that already exists. We focus on four drivers that can influence and improve ocean literacy and societal connections to the ocean: (1) education, (2) cultural connections, (3) technological developments, and (4) knowledge exchange and science-policy interconnections. We explore how each driver plays a role in improving perceptions of the ocean to engender more widespread societal support for effective ocean management and conservation. In doing so, we develop an ocean literacy toolkit, a practical resource for enhancing ocean connections across a broad range of contexts worldwide.

Exploring the potential of template-based modelling.

MOTIVATION: Template-based modelling can approximate the unknown structure of a target protein using an homologous template structure. The core of the resulting prediction then comprises the structural regions conserved between template and target. Target prediction could be improved by rigidly repositioning such single template, structurally conserved fragment regions. The purpose of this article is to quantify the extent to which such improvements are possible and to relate this extent to properties of the target, the template and their alignment. RESULTS: The improvement in accuracy achievable when rigid fragments from a single template are optimally positioned was calculated using structure pairs from the HOMSTRAD database, as well as CASP7 and CASP8 target/best template pairs. Over the union of the structurally conserved regions, improvements of 0.7 A in root mean squared deviation (RMSD) and 6% in GDT_HA were commonly observed. A generalized linear model revealed that the extent to which a template can be improved can be predicted using four variables. Templates with the greatest scope for improvement tend to have relatively more fragments, shorter fragments, higher percentage of helical secondary structure and lower sequence identity. Optimal positioning of the template fragments offers the potential for improving loop modelling. These results demonstrate that substantial improvement could be made on many templates if the conserved fragments were to be optimally positioned. They also provide a basis for identifying templates for which modification of fragment positions may yield such improvements.

Realization of a Graphene/PMMA Acoustic Capacitive Sensor Released by Silicon Dioxide Sacrificial Layer.

We report the realization of an acoustic capacitive microphone formed by graphene/poly(methyl methacrylate) (PMMA). It is the first time that the ultra-large graphene/PMMA membrane suspended fully over the cavity has been fabricated by releasing the silicon dioxide sacrificial layer underneath the membrane. The novelty in the fabrication method is that the silicon dioxide layer has been etched by hydrogen fluoride vapor from the back of the partly etched silicon substrate. Using the new process, the ultra-large graphene/PMMA membrane, with a diameter to thickness ratio of 7800, has been suspended over the cavity with a 2 µm air gap. The spacing of 2 µm is the minimum gap over the graphene-based acoustic capacitive microphones which have been reported so far. The static deformation of the suspended graphene/PMMA membrane after silicon dioxide has been etched is estimated to be 270 nm. The aspect ratio of the membrane's diameter over its static deformation is around 13,000, which shows that the graphene/PMMA membrane with a diameter of a few millimeters can be transferred and suspended over the substrate with relatively small deformation by releasing the sacrificial silicon dioxide layer. The dynamic behavior of the device under electrostatic actuation has been characterized. The acoustic response of the graphene/PMMA capacitive microphone has been measured, and the sensitivity has been observed to be -47.5 dB V (4.22 mV/Pa) ± 10%. The strain in the graphene/PMMA membrane is estimated to be 0.034%.

Directionality in protein fold prediction.

BACKGROUND: Ever since the ground-breaking work of Anfinsen et al. in which a denatured protein was found to refold to its native state, it has been frequently stated by the protein fold prediction community that all the information required for protein folding lies in the amino acid sequence. Recent in vitro experiments and in silico computational studies, however, have shown that cotranslation may affect the folding pathway of some proteins, especially those of ancient folds. In this paper aspects of cotranslational folding have been incorporated into a protein structure prediction algorithm by adapting the Rosetta program to fold proteins as the nascent chain elongates. This makes it possible to conduct a pairwise comparison of folding accuracy, by comparing folds created sequentially from each end of the protein. RESULTS: A single main result emerged: in 94% of proteins analyzed, following the sense of translation, from N-terminus to C-terminus, produced better predictions than following the reverse sense of translation, from the C-terminus to N-terminus. Two secondary results emerged. First, this superiority of N-terminus to C-terminus folding was more marked for proteins showing stronger evidence of cotranslation and second, an algorithm following the sense of translation produced predictions comparable to, and occasionally better than, Rosetta. CONCLUSIONS: There is a directionality effect in protein fold prediction. At present, prediction methods appear to be too noisy to take advantage of this effect; as techniques refine, it may be possible to draw benefit from a sequential approach to protein fold prediction.

Reference intervals for eight measurands of the blood count in a large population based study.

BACKGROUND: The blood count is widely used in clinical practice. Well defined reference intervals for each measurand are essential for correct clinical interpretation of results. Most previous studies have not been population-based. We therefore calculated reference intervals for several hematological measurands from a sample of the general adult population of Northeastern Germany. METHODS AND RESULTS: We used data from 2967 healthy individuals recruited for the population-based Study of Health in Pomerania (SHIP). Reference intervals were calculated according to the guidelines of the Clinical and Laboratory Standards Institute (CLSI) using the bootstrap method for the age range from 20 to 79 years and, in addition, stratified according to age and gender with both bootstrap and quantile regression procedures. Reference ranges for erythrocytes, hemoglobin and hematocrit increased with age in women but decreased in men. CONCLUSIONS: Our reference intervals were lower than those previously published for erythrocytes, hemoglobin, hematocrit and leukocytes but higher for Mean Corpuscular Volume (MCV) and Mean Corpuscular Hemoglobin (MCH). Different laboratory methods and study populations may lead to disparity in results.

A restriction enzyme reduced representation sequencing approach for low-cost, high-throughput metagenome profiling.

Microbial community profiles have been associated with a variety of traits, including methane emissions in livestock. These profiles can be difficult and expensive to obtain for thousands of samples (e.g. for accurate association of microbial profiles with traits), therefore the objective of this work was to develop a low-cost, high-throughput approach to capture the diversity of the rumen microbiome. Restriction enzyme reduced representation sequencing (RE-RRS) using ApeKI or PstI, and two bioinformatic pipelines (reference-based and reference-free) were compared to bacterial 16S rRNA gene sequencing using repeated samples collected two weeks apart from 118 sheep that were phenotypically extreme (60 high and 58 low) for methane emitted per kg dry matter intake (n = 236). DNA was extracted from freeze-dried rumen samples using a phenol chloroform and bead-beating protocol prior to RE-RRS. The resulting sequences were used to investigate the repeatability of the rumen microbial community profiles, the effect of laboratory and analytical method, and the relationship with methane production. The results suggested that the best method was PstI RE-RRS analyzed with the reference-free approach, which accounted for 53.3±5.9% of reads, and had repeatabilities of 0.49±0.07 and 0.50±0.07 for the first two principal components (PC1 and PC2), phenotypic correlations with methane yield of 0.43±0.06 and 0.46±0.06 for PC1 and PC2, and explained 41±8% of the variation in methane yield. These results were significantly better than for bacterial 16S rRNA gene sequencing of the same samples (p<0.05) except for the correlation between PC2 and methane yield. A Sensitivity study suggested approximately 2000 samples could be sequenced in a single lane on an Illumina HiSeq 2500, meaning the current work using 118 samples/lane and future proposed 384 samples/lane are well within that threshold. With minor adaptations, our approach could be used to obtain microbial profiles from other metagenomic samples.

A multi-stage approach to clustering and imputation of gene expression profiles.

MOTIVATION: Microarray experiments have revolutionized the study of gene expression with their ability to generate large amounts of data. This article describes an alternative to existing approaches to clustering of gene expression profiles; the key idea is to cluster in stages using a hierarchy of distance measures. This method is motivated by the way in which the human mind sorts and so groups many items. The distance measures arise from the orthogonal breakup of Euclidean distance, giving us a set of independent measures of different attributes of the gene expression profile. Interpretation of these distances is closely related to the statistical design of the microarray experiment. This clustering method not only accommodates missing data but also leads to an associated imputation method. RESULTS: The performance of the clustering and imputation methods was tested on a simulated dataset, a yeast cell cycle dataset and a central nervous system development dataset. Based on the Rand and adjusted Rand indices, the clustering method is more consistent with the biological classification of the data than commonly used clustering methods. The imputation method, at varying levels of missingness, outperforms most imputation methods, based on root mean squared error (RMSE). AVAILABILITY: Code in R is available on request from the authors.

Cotranslational protein folding--fact or fiction?

MOTIVATION: Experimentalists have amassed extensive evidence over the past four decades that proteins appear to fold during production by the ribosome. Protein structure prediction methods, however, do not incorporate this property of folding. A thorough study to find the fingerprint of such sequential folding is the first step towards using it in folding algorithms, so assisting structure prediction. RESULTS: We explore computationally the existence of evidence for cotranslational folding, based on large sets of experimentally determined structures in the PDB. Our perspective is that cotranslational folding is the norm, but that the effect is masked in most classes. We show that it is most evident in alpha/beta proteins, confirming recent findings. We also find mild evidence that older proteins may fold cotranslationally. A tool is provided for determining, within a protein, where cotranslation is most evident.

Low-dose interleukin-2 in patients with stable ischaemic heart disease and acute coronary syndromes (LILACS): protocol and study rationale for a randomised, double-blind, placebo-controlled, phase I/II clinical trial.

INTRODUCTION: Inflammation and dysregulated immune responses play a crucial role in atherosclerosis, underlying ischaemic heart disease (IHD) and acute coronary syndromes (ACSs). Immune responses are also major determinants of the postischaemic injury in myocardial infarction. Regulatory T cells (CD4+CD25+FOXP3+; Treg) induce immune tolerance and preserve immune homeostasis. Recent in vivo studies suggested that low-dose interleukin-2 (IL-2) can increase Treg cell numbers. Aldesleukin is a human recombinant form of IL-2 that has been used therapeutically in several autoimmune diseases. However, its safety and efficacy is unknown in the setting of coronary artery disease. METHOD AND ANALYSIS: Low-dose interleukin-2 in patients with stable ischaemic heart disease and acute coronary syndromes is a single-centre, first-in-class, dose-escalation, two-part clinical trial. Patients with stable IHD (part A) and ACS (part B) will be randomised to receive either IL-2 (aldesleukin; dose range 0.3-3×106 IU) or placebo once daily, given subcutaneously, for five consecutive days. Part A will have five dose levels with five patients in each group. Group 1 will receive a dose of 0.3×106 IU, while the dose for the remaining four groups will be determined on completion of the preceding group. Part B will have four dose levels with eight patients in each group. The dose of the first group will be based on part A. Doses for each of the subsequent three groups will similarly be determined after completion of the previous group. The primary endpoint is safety and tolerability of aldesleukin and to determine the dose that increases mean circulating Treg levels by at least 75%. ETHICS AND DISSEMINATION: The study received a favourable opinion by the Greater Manchester Central Research Ethics Committee, UK (17/NW/0012). The results of this study will be reported through peer-reviewed journals, conference presentations and an internal organisational report. TRIAL REGISTRATION NUMBER: NCT03113773; Pre-results.

Modelling sequential protein folding under kinetic control.

MOTIVATION: This study presents a novel investigation of the effect of kinetic control on cotranslational protein folding. We demonstrate the effect using simple HP lattice models and show that the cotranslational folding of proteins under kinetic control has a significant impact on the final conformation. Differences arise if nature is not capable of pushing a partially folded protein back over a large energy barrier. For this reason we argue that such constraints should be incorporated into structure prediction techniques. We introduce a finite surmountable energy barrier which allows partially formed chains to partly unfold, and permits us to enumerate exhaustively all energy pathways. RESULTS: We compare the ground states obtained sequentially with the global ground states of designing sequences (those with a unique global ground state). We find that the sequential ground states become less numerous and more compact as the surmountable energy barrier increases. We also introduce a probabilistic model to describe the distribution of final folds and allow partial settling to the Boltzmann distribution of states at each stage. As a result, conformations with the highest probability of final occurrence are not necessarily the ones of lowest energy. AVAILABILITY: Software available on request.

The risk assessment-policy gap: an example from the UK contaminated land regime.

Risk-based regulation assumes that risk assessment can evaluate risks against policy objectives. However, policy goals are often ambiguous and require risk assessors to interpret them for use in risk assessment. This risk assessment-policy gap stems partly from normative and imprecise policy language but is rooted more fundamentally in society's uncertain expectations for the environment. Until this uncertainty is resolved, the democratic and regulatory effectiveness of risk regulation will be undermined by ad hoc policy decisions abdicated to risk assessors.

Sublingual gland excision: a dissection carried out following adjacent anatomical structures.

We describe an anatomical and simple technique for the removal of the submandibular gland that has good outcomes and minimal complications.

3C-Silicon Carbide Microresonators for Timing and Frequency Reference.

In the drive to miniaturise and integrate reference oscillator components, microelectromechanical systems (MEMS) resonators are excellent candidates to replace quartz crystals. Silicon is the most utilised resonator structural material due to its associated well-established fabrication processes. However, when operation in harsh environments is required, cubic silicon carbide (3C-SiC) is an excellent candidate for use as a structural material, due to its robustness, chemical inertness and high temperature stability. In order to actuate 3C-SiC resonators, electrostatic, electrothermal and piezoelectric methods have been explored. Both electrothermal and piezoelectric actuation can be accomplished with simpler fabrication and lower driving voltages, down to 0.5 V, compared to electrostatic actuation. The vibration amplitude at resonance can be maximised by optimising the design and location of the electrodes. Electrical read out of the resonator can be performed with electrostatic or piezoelectric transduction. Finally, a great deal of research has focused on tuning the resonant frequency of a 3C-SiC resonator by adjusting the DC bias applied to the electrodes, with a higher (up to 160-times) tuning range for electrothermal tuning compared to piezoelectric tuning. Electrothermal tuning lowers the frequency, while piezoelectric tuning can be used to raise the frequency.