30 research outputs found

    Whole genome sequencing-based mapping and candidate identification of mutations from fixed zebrafish tissue

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    As forward genetic screens in zebrafish become more common, the number of mutants that cannot be identified by gross morphology or through transgenic approaches, such as many nervous system defects, has also increased. Screening for these difficult-to-visualize phenotypes demands techniques such as whole-mount in situ hybridization (WISH) or antibody staining, which require tissue fixation. To date, fixed tissue has not been amenable for generating libraries for whole genome sequencing (WGS). Here, we describe a method for using genomic DNA from fixed tissue and a bioinformatics suite for WGS-based mapping of zebrafish mutants. We tested our protocol using two known zebrafish mutant alleles, gpr126st49 and egr2bfh227, both of which cause myelin defects. As further proof of concept we mapped a novel mutation, stl64, identified in a zebrafish WISH screen for myelination defects. We linked stl64 to chromosome 1 and identified a candidate nonsense mutation in the F-box and WD repeat domain containing 7 (fbxw7) gene. Importantly, stl64 mutants phenocopy previously described fbxw7vu56 mutants, and knockdown of fbxw7 in wild-type animals produced similar defects, demonstrating that stl64 disrupts fbxw7. Together, these data show that our mapping protocol can map and identify causative lesions in mutant screens that require tissue fixation for phenotypic analysis

    Further characterization of Vibrio vulnificus rugose variants and identification of a capsular and rugose exopolysaccharide gene cluster

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    Capsular polysaccharide (CPS) is a major virulence factor in Vibrio vulnificus, and encapsulated strains have an opaque, smooth (OpS) colony morphology, while nonencapsulated strains have a translucent, smooth (TrS) colony morphology. Previously, we showed that OpS and TrS parental strains can yield a third colony type, rugose (R), and that the resulting strains, with the OpR and TrR phenotypes, respectively, form copious biofilms. Here we show that while OpR and TrR strains both produce three-dimensional biofilm structures that are indicative of rugose extracellular polysaccharide (rEPS) production, OpR strains also retain expression of CPS and are virulent in an iron-supplemented mouse model, while TrR strains lack CPS and are avirulent. Chlorine resistance assays further distinguished OpR and TrR isolates as exposure to 3 Όg/ml NaOCl eradicated both OpS and OpR strains, while both TrS and TrR strains survived, but at rates which were significantly different from one another. Taken together, these results further emphasize the importance of CPS for virulence of V. vulnificus and establish a correlation between CPS expression and chlorine sensitivity in this organism. Using reverse transcriptase PCR, we also identified a nine-gene cluster associated with both CPS and rEPS expression in V. vulnificus, designated the wcr (capsular and rugose polysaccharide) locus, with expression occurring primarily in R variants. The latter results set the stage for characterization of functional determinants which individually or collectively contribute to expression of multiple EPS forms in this pathogen. Copyright © 2008, American Society for Microbiology. All Rights Reserved

    Guidelines for the Management of Severe Traumatic Brain Injury: 2020 Update of the Decompressive Craniectomy Recommendations

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    © Congress of Neurological Surgeons 2020. When the fourth edition of the Brain Trauma Foundation\u27s Guidelines for the Management of Severe Traumatic Brain Injury were finalized in late 2016, it was known that the results of the RESCUEicp (Trial of Decompressive Craniectomy for Traumatic Intracranial Hypertension) randomized controlled trial of decompressive craniectomy would be public after the guidelines were released. The guideline authors decided to proceed with publication but to update the decompressive craniectomy recommendations later in the spirit of living guidelines, whereby topics are updated more frequently, and between new editions, when important new evidence is published. The update to the decompressive craniectomy chapter presented here integrates the findings of the RESCUEicp study as well as the recently published 12-mo outcome data from the DECRA (Decompressive Craniectomy in Patients With Severe Traumatic Brain Injury) trial. Incorporation of these publications into the body of evidence led to the generation of 3 new level-IIA recommendations; a fourth previously presented level-IIA recommendation remains valid and has been restated. To increase the utility of the recommendations, we added a new section entitled Incorporating the Evidence into Practice. This summary of expert opinion provides important context and addresses key issues for practitioners, which are intended to help the clinician utilize the available evidence and these recommendations. The full guideline can be found at: https://braintrauma.org/guidelines/guidelines-for-the-management-of-severe-tbi-4th-ed#/

    Pre-Hospital Fast Positive Cases Identified by DFB Ambulance Paramedics: Final Clinical Diagnosis.

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    Ischaemic stroke clinical outcomes are improved by earlier treatment with intravenous thrombolysis. An existing pathway at the Mater University Hospital for assessment of suspected acute stroke in the Emergency Department was updated, aiming to shorten door-to-needle time. This study examines the final clinical diagnosis of Dublin Fire Brigade Ambulance Paramedic identified Face-Arm-Speech-Test (FAST) positive patients presenting to the Emergency Department over a 7 month period. A retrospective analysis was carried out of 177 consecutive FAST positive patients presenting between March and November 2014. The final clinical diagnosis was acute stroke in 57.1% (n=101) of patients. Of these, 76 were ischaemic strokes of whom 56.5% (n=43) were thrombolysed. In the pre-hospital setting Ambulance Paramedics can identify, with reasonable accuracy, acute stroke using the FAST test. Over half of the ischaemic stroke patients presenting via this pathway can be treated with intravenous thrombolysis

    First Demonstration of Electrostatic Damping of Parametric Instability at Advanced LIGO

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    Interferometric gravitational wave detectors operate with high optical power in their arms in order to achieve high shot-noise limited strain sensitivity. A significant limitation to increasing the optical power is the phenomenon of three-mode parametric instabilities, in which the laser field in the arm cavities is scattered into higher-order optical modes by acoustic modes of the cavity mirrors. The optical modes can further drive the acoustic modes via radiation pressure, potentially producing an exponential buildup. One proposed technique to stabilize parametric instability is active damping of acoustic modes. We report here the first demonstration of damping a parametrically unstable mode using active feedback forces on the cavity mirror. A 15 538 Hz mode that grew exponentially with a time constant of 182 sec was damped using electrostatic actuation, with a resulting decay time constant of 23 sec. An average control force of 0.03 nN was required to maintain the acoustic mode at its minimum amplitude

    TRY plant trait database – enhanced coverage and open access

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    Plant traits - the morphological, anatomical, physiological, biochemical and phenological characteristics of plants - determine how plants respond to environmental factors, affect other trophic levels, and influence ecosystem properties and their benefits and detriments to people. Plant trait data thus represent the basis for a vast area of research spanning from evolutionary biology, community and functional ecology, to biodiversity conservation, ecosystem and landscape management, restoration, biogeography and earth system modelling. Since its foundation in 2007, the TRY database of plant traits has grown continuously. It now provides unprecedented data coverage under an open access data policy and is the main plant trait database used by the research community worldwide. Increasingly, the TRY database also supports new frontiers of trait‐based plant research, including the identification of data gaps and the subsequent mobilization or measurement of new data. To support this development, in this article we evaluate the extent of the trait data compiled in TRY and analyse emerging patterns of data coverage and representativeness. Best species coverage is achieved for categorical traits - almost complete coverage for ‘plant growth form’. However, most traits relevant for ecology and vegetation modelling are characterized by continuous intraspecific variation and trait–environmental relationships. These traits have to be measured on individual plants in their respective environment. Despite unprecedented data coverage, we observe a humbling lack of completeness and representativeness of these continuous traits in many aspects. We, therefore, conclude that reducing data gaps and biases in the TRY database remains a key challenge and requires a coordinated approach to data mobilization and trait measurements. This can only be achieved in collaboration with other initiatives
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