The Legal Questing Beast: Vocational Students’ Research Strategies, Motivations and Emotions

In the context of the Bar Vocational Course, this project enquired how students perceive their research strategies when they interact with repositories of legal information. The research captured and analysed the students' descriptions of these engagements and their motivations for as well as their emotional responses to undertaking research tasks. Some tentative comparisons are made between the students' strategies and those reported by a small number of junior barristers

The Well-educated Witness: Witness Familiarisation Training in England & Wales

The presentation at the 4th Applied Legal Storytelling Conference promoted respect for witnesses’ ownership of their narratives and explained how effective witness training can successfully avoid unethical coaching whilst remaining client-centred. Witnesses’ roles within the Common Law adversarial trial tradition were analysed, and the English and Welsh Court of Appeal’s guidance to lawyers on acceptable and unacceptable forms of trial preparation for witnesses was outlined and critiqued in its historical context and compared with other common law jurisdictions’ practices. It was argued that witnesses can be educated ethically to enhance their effectiveness in the witness box whilst respecting them as pristine evidence sources. This article develops these themes to argue that in England and Wales witness familiarisation trainers are educators rather than partisan trial strategists. Case law and research literature in the field of witness familiarisation and ethics are relied upon to support this argument. In addition, the author draws on several years’ experience of the courtroom and witness familiarisation training with witnesses of fact, expert witnesses, and criminal investigators

A Relational Database for the Discovery of Genes Encoding Amino Acid Biosynthetic Enzymes in Pathogenic Fungi

Fungal phytopathogens continue to cause major economic impact, either directly, through crop losses, or due to the costs of fungicide application. Attempts to understand these organisms are hampered by a lack of fungal genome sequence data. A need exists, however, to develop specific bioinformatics tools to collate and analyse the sequence data that currently is available. A web-accessible gene discovery database (http://cogeme.ex.ac.uk/biosynthesis.html) was developed as a demonstration tool for the analysis of metabolic and signal transduction pathways in pathogenic fungi using incomplete gene inventories. Using Bayesian probability to analyse the currently available gene information from pathogenic fungi, we provide evidence that the obligate pathogen Blumeria graminis possesses all amino acid biosynthetic pathways found in free-living fungi, such as Saccharomyces cerevisiae. Phylogenetic analysis was also used to deduce a gene history of succinate-semialdehyde dehydrogenase, an enzyme in the glutamate and lysine biosynthesis pathways. The database provides a tool and methodology to researchers to direct experimentation towards predicting pathway conservation in pathogenic microorganisms

Protein kinase C is essential for viability of the rice blast fungus Magnaporthe oryzae

This is the final version of the article. Available from Wiley via the DOI in this record.Protein kinase C constitutes a family of serine-threonine kinases found in all eukaryotes and implicated in a wide range of cellular functions, including regulation of cell growth, cellular differentiation and immunity. Here, we present three independent lines of evidence which indicate that protein kinase C is essential for viability of Magnaporthe oryzae. First, all attempts to generate a target deletion of PKC1, the single copy protein kinase C-encoding gene, proved unsuccessful. Secondly, conditional gene silencing of PKC1 by RNA interference led to severely reduced growth of the fungus, which was reversed by targeted deletion of the Dicer2-encoding gene, MDL2. Finally, selective kinase inhibition of protein kinase C by targeted allelic replacement with an analogue-sensitive PKC1(AS) allele led to specific loss of fungal viability in the presence of the PP1 inhibitor. Global transcriptional profiling following selective PKC inhibition identified significant changes in gene expression associated with cell wall re-modelling, autophagy, signal transduction and secondary metabolism. When considered together, these results suggest protein kinase C is essential for growth and development of M. oryzae with extensive downstream targets in addition to the cell integrity pathway. Targeting protein kinase C signalling may therefore prove an effective means of controlling rice blast disease.This work was funded by a BBSRC CASE PhD studentship to TJP with support from Syngenta and a European Research Council, Advanced Investigator Award to NJT under the European Union's Seventh Framework Programme (FP7/2007-2013)/ERC grant agreement no. 294702 GENBLAST

Insectivorous bats are less active near freeways

Traffic disturbances (i.e. pollution, light, noise, and vibrations) often extend into the area surrounding a road creating a 'road-effect zone'. Habitat within the road-effect zone is degraded or, in severe cases, completely unsuitable for wildlife, resulting in indirect habitat loss. This can have a disproportionate impact on wildlife in highly modified landscapes, where remaining habitat is scarce or occurs predominantly along roadside reserves. In this study, we investigated the road-effect zone for insectivorous bats in highly cleared agricultural landscapes by quantifying the change in call activity with proximity to three major freeways. The activity of seven out of 10 species of bat significantly decreased with proximity to the freeway. We defined the road-effect zone to be the proximity at which call activity declined by at least 20% relative to the maximum detected activity. The overall road-effect zone for bats in this region was 307 m, varying between 123 and 890 m for individual species. Given that this road-effect zone exceeds the typical width of the roadside verges (<50 m), it is possible that much of the vegetation adjacent to freeways in this and similar landscapes provides low-quality habitat for bats. Without accounting for the road-effect zone, the amount of habitat lost or degraded due to roads is underestimated, potentially resulting in the loss of wildlife, ecosystem services and key ecosystem processes (e.g. predator-prey or plant-pollinator interactions) from the landscape. We suggest all future environmental impact assessments include quantifying the road-effect zone for sensitive wildlife, in order to best plan and mitigate the impact of roads on the environment. Mitigating the effects of new and existing roads on wildlife is essential to ensure enough high-quality habitat persists to maintain wildlife populations

Hook is an adapter that coordinates kinesin-3 and dynein cargo attachment on early endosomes.

This is the final version of the article. Available from Rockefeller University Press via the DOI in this record.Bidirectional membrane trafficking along microtubules is mediated by kinesin-1, kinesin-3, and dynein. Several organelle-bound adapters for kinesin-1 and dynein have been reported that orchestrate their opposing activity. However, the coordination of kinesin-3/dynein-mediated transport is not understood. In this paper, we report that a Hook protein, Hok1, is essential for kinesin-3- and dynein-dependent early endosome (EE) motility in the fungus Ustilago maydis. Hok1 binds to EEs via its C-terminal region, where it forms a complex with homologues of human fused toes (FTS) and its interactor FTS- and Hook-interacting protein. A highly conserved N-terminal region is required to bind dynein and kinesin-3 to EEs. To change the direction of EE transport, kinesin-3 is released from organelles, and dynein binds subsequently. A chimaera of human Hook3 and Hok1 rescues the hok1 mutant phenotype, suggesting functional conservation between humans and fungi. We conclude that Hok1 is part of an evolutionarily conserved protein complex that regulates bidirectional EE trafficking by controlling attachment of both kinesin-3 and dynein.This work was supported by Wellcome Trust (097835/Z/11/Z) and the Biotechnology and Biological Sciences Research Council (BB/J009903/1)

A codon-optimized green fluorescent protein for live cell imaging in Zymoseptoria tritici

AbstractFluorescent proteins (FPs) are powerful tools to investigate intracellular dynamics and protein localization. Cytoplasmic expression of FPs in fungal pathogens allows greater insight into invasion strategies and the host-pathogen interaction. Detection of their fluorescent signal depends on the right combination of microscopic setup and signal brightness. Slow rates of photo-bleaching are pivotal for in vivo observation of FPs over longer periods of time. Here, we test green-fluorescent proteins, including Aequorea coerulescens GFP (AcGFP), enhanced GFP (eGFP) from Aequorea victoria and a novel Zymoseptoria tritici codon-optimized eGFP (ZtGFP), for their usage in conventional and laser-enhanced epi-fluorescence, and confocal laser-scanning microscopy. We show that eGFP, expressed cytoplasmically in Z. tritici, is significantly brighter and more photo-stable than AcGFP. The codon-optimized ZtGFP performed even better than eGFP, showing significantly slower bleaching and a 20–30% further increase in signal intensity. Heterologous expression of all GFP variants did not affect pathogenicity of Z. tritici. Our data establish ZtGFP as the GFP of choice to investigate intracellular protein dynamics in Z. tritici, but also infection stages of this wheat pathogen inside host tissue

The support and information needs of adolescents and young adults with cancer when active treatment ends

Background: The end of active treatment is a period of high stress for young people with cancer, but limited literature exists about their information and support needs during this phase. This study aimed to understand the needs of young people with cancer, how these needs are currently being met, and how best to provide information and support at the end of active treatment. Methods: This was a multi-stage, mixed methods study exploring the end of treatment experience from the perspectives of young people, and the healthcare professionals caring for them. Semi-structured interviews were undertaken with healthcare professionals, which informed a survey administered nationally. Subsequently, semi-structured interviews were conducted with young people. These combined results informed a co-design workshop to develop recommendations. Results: Telephone interviews were conducted with 12 healthcare professionals and 49 completed the online survey. A total of 11 young people aged 19–26 years (female = 8; 73%) were interviewed. The stakeholder workshop was attended by both healthcare professionals (n = 8) and young people (n = 3). At the end of treatment young people experience numerous ongoing physical issues including pain, fatigue and insomnia; in addition to a range of psychosocial and emotional issues including anxiety, fear of recurrence and isolation. The top three priorities for end of treatment care were: earlier provision and preparation around on-going impact of cancer and cancer treatment; standardised and continued follow-up of young people’s emotional well-being; and development of more information and resources specific to young people. Conclusion The access and availability of appropriate information and sources of support at the end of treatment is variable and inequitable. Young people’s needs would be more effectively met by timely, structured and accessible information, and support provision at the end of treatment to both prepare and enable adaptation across their transition to living with and beyond cancer. This will require both organisational and practical adjustments in care delivery, in addition to a renewed and updated understanding of what the ‘end of treatment’ transition process means

CRISPR-Cas9 ribonucleoprotein-mediated co-editing and counterselection in the rice blast fungus

The rice blast fungus Magnaporthe oryzae is the most serious pathogen of cultivated rice and a significant threat to global food security. To accelerate targeted mutation and specific genome editing in this species, we have developed a rapid plasmid-free CRISPR-Cas9-based genome editing method. We show that stable expression of Cas9 is highly toxic to M. oryzae. However efficient gene editing can be achieved by transient introduction of purified Cas9 pre-complexed to RNA guides to form ribonucleoproteins (RNPs). When used in combination with oligonucleotide or PCR-generated donor DNAs, generation of strains with specific base pair edits, in-locus gene replacements, or multiple gene edits, is very rapid and straightforward. We demonstrate a co-editing strategy for the creation of single nucleotide changes at specific loci. Additionally, we report a novel counterselection strategy which allows creation of precisely edited fungal strains that contain no foreign DNA and are completely isogenic to the wild type. Together, these developments represent a scalable improvement in the precision and speed of genetic manipulation in M. oryzae and are likely to be broadly applicable to other fungal species

Defining marine important bird areas: Testing the foraging radius approach

International audienceRecent international initiatives have promoted a number of different approaches to identify marine Important Bird and biodiversity Areas (IBAs), which are important areas for foraging, migrating or over-wintering seabirds. The ‘Foraging Radius Approach’ is one of these and uses known foraging range and habitat preferences to predict the size and location of foraging areas around breeding colonies. Here we assess the performance of the Foraging Radius Approach using GPS tracking data from six seabird species with a variety of foraging modes. For each species we compared the population home-range areas of our six study species with the home-range areas defined using the Foraging Radius Approach. We also assessed whether basic information on depth preferences from tracking data could improve these home-range area estimates. Foraging Radius Approach home-range areas based on maximum foraging radii encompassed the entire population home-range of five out of six of our study species but overestimated the size of the population home-range area in every case. The mean maximum foraging radius overestimated the population home-range areas by a factor of 4–14 for five of the six species whilst the mean foraging radius overestimated the population home-range area for half of the species and underestimated for the rest. In the absence of other data, the Foraging Radius Approach appears to provide a reasonable basis for preliminary marine IBA identification. We suggest that using the mean value of all previously reported maximum foraging radii, informed by basic depth preferences provides the most appropriate prediction, balancing the needs of seabirds with efficient use of marine space