Kinetics of the Formation and Dissociation of Actin Filament Branches Mediated by Arp2/3 Complex

AbstractThe actin filament network at the leading edge of motile cells relies on localized branching by Arp2/3 complex from “mother” filaments growing near the plasma membrane. The nucleotide bound to the mother filaments (ATP, ADP and phosphate, or ADP) may influence the branch dynamics. To determine the effect of the nucleotide bound to the subunits of the mother filament on the formation and stability of branches, we compared the time courses of actin polymerization in bulk samples measured using the fluorescence of pyrene actin with observations of single filaments by total internal reflection fluorescence microscopy. Although the branch nucleation rate in bulk samples was nearly the same regardless of the nucleotide on the mother filaments, we observed fewer branches by microscopy on ADP-bound filaments than on ADP-Pi-bound filaments. Observation of branches in the microscope depends on their binding to the slide. Since the probability that a branch binds to the slide is directly related to its lifetime, we used counts of branches to infer their rates of dissociation from mother filaments. We conclude that the nucleotide on the mother filament does not affect the initial branching event but that branches are an order of magnitude more stable on the sides of new ATP- or ADP-Pi filaments than on ADP-actin filaments

Videofluoroscopic Evaluation of the Pharynx and Upper Esophageal Sphincter in the Dog: A Systematic Review of the Literature

Background: Diseases of the pharynx and upper esophageal sphincter can result in debilitating respiratory difficulty, dysphagia or a combination of both. An exact diagnosis is essential to properly prognosticate and guide therapy. Videofluoroscopic assessment of the pharynx and upper esophageal sphincter with or without orally administered contrast material is the diagnostic of choice for many diseases as both anatomic and functional information is gleaned. The purpose of this review is to assess for continuity in imaging protocols across institutions and to record quantitative and qualitative parameters used for analysis of videofluoroscopy of the pharynx and upper esophageal sphincter in dogs.Methods: A systematic literature search was performed including articles published in peer-reviewed veterinary journals involving the topic of videofluoroscopy of the pharynx and upper esophageal sphincter through August 1, 2018. Specifics of study acquisition technique were recorded. Quantitative and qualitative videofluoroscopic parameters were recorded and compared across institutions where appropriate using one-way ANOVA with p ≤ 0.05 being considered significant.Results: Videofluoroscopy of the pharynx and upper esophageal sphincter is performed either in right lateral or standing postures depending on the institution. Bolus size and consistency used during contrast videofluoroscopy of swallowing differs between institutions. Some institutions evaluate videofluoroscopic studies using qualitative criteria while others apply quantitative measures. Reported quantitative measures include inter-swallow interval, swallow rate, jaw cycles per swallow ratio, time to upper esophageal opening, maximal pharyngeal contraction, maximum laryngeal excursion, upper esophageal closure, epiglottic re-opening, and pharyngeal constriction ratio. Measurement outcomes are significantly different between institutions and when bolus size/consistency is variable when assessing healthy dogs.Conclusions: The current peer-reviewed literature on fluoroscopic evaluation of the pharynx and UES in dogs shows a lack of standardization regarding imaging protocol. There is not a standard set of quantitative criteria applied amongst the institutions and there are significant differences in the outcomes obtained from videofluoroscopic assessment of swallowing suggesting significant inter-observer or inter-institutional variability. A consensus statement regarding imaging protocol and what parameters should be used to interpret airway and swallowing videofluoroscopic studies of the pharynx and UES in dogs is needed along with targeted analysis of observer variability

Craniomaxillofacial Trauma in Dogs-Part II: Association Between Fracture Location, Morphology and Etiology.

Treatment of craniomaxillofacial (CMF) trauma in dogs requires a thorough understanding of the CMF skeletal structures involved. The human medical literature has several examples of CMF trauma and fracture classification, including the classically described Le Fort fractures. The recent classification schemes require large studies using computed tomography (CT). In the veterinary medical literature, such studies are lacking. The aims of part II of this retrospective study were to use a large number of CT studies of dogs evaluated for CMF trauma to determine whether specific fracture locations in the CMF region occur concurrently, and whether trauma etiology influences fracture morphology. This information may then be used to form a fracture classification scheme in the future. The medical records and CT studies of 165 dogs over a 10-year period were evaluated. The skeletal location of CMF fractures as well as the severity of displacement and fragmentation of each fracture was recorded. Dogs' demographic data and trauma etiology were also recorded. Fractured portions of the mandible tended to occur with fractures of adjacent bones, with the major exception of symphyseal separation, which occurred simultaneously with fractures of the cribriform plate. Fractures of the maxillary bone were accompanied by many concurrent fractures affecting the majority of the midface, skull base, and cranial vault. When the zygomatic bone was fractured, the other bones comprising the orbit also tended to fracture. Fractures of the relatively superficially located frontal and nasal bones were often accompanied by fractures of the skull base. Fracture etiology influenced fracture morphology such that vehicular trauma resulted in a relatively higher number of severely displaced and comminuted fractures than did other trauma etiologies. This study provides examples of fractures that, when found, should prompt veterinarians to look for additional injuries in specific locations. In addition, it further highlights the need for thorough CT evaluation of the entire CMF region, even when clinically apparent fractures appear relatively superficial

Craniomaxillofacial Trauma in Dogs-Part I: Fracture Location, Morphology and Etiology.

Treatment of craniomaxillofacial (CMF) trauma in dogs often requires a multidisciplinary approach and a thorough understanding of the CMF skeletal structures involved. The aim of this retrospective study was to use a large number of CT studies of dogs evaluated for CMF trauma and to describe fracture location and morphology in relation to demographic data and trauma etiology. The medical records and CT studies of 165 dogs over a 10-year period were evaluated. The skeletal location of CMF fractures as well as the severity of displacement and fragmentation of each fracture was recorded. Patient demographic data and trauma etiology were also recorded. Animal bites accounted for the majority of trauma (50%), followed by unknown trauma (15%), vehicular accidents (13%), and blunt force trauma (13%). Small dogs, <10 kg, and juveniles accounted for the majority of patients (41.8 and 25.5%, respectively). The most likely bone or region to be fractured was the maxillary bone, followed by the premolar and molar regions of the mandible. Up to 37 bones or regions were fractured in any given patient, with an average of 8.2 fractured bones or regions per dog. The most commonly fractured location varied according to trauma etiology. Specifically, vehicular accidents tended to result in more locations with a higher probability of fracture than other trauma types. A major conclusion from this study is that every bone of the CMF region was fractured in at least one case and many cases had a large number of fractured regions. Thus, the need for comprehensive assessment of the entire CMF region, preferably using CT, is underscored

FGF4 retrogene on CFA12 is responsible for chondrodystrophy and intervertebral disc disease in dogs.

Chondrodystrophy in dogs is defined by dysplastic, shortened long bones and premature degeneration and calcification of intervertebral discs. Independent genome-wide association analyses for skeletal dysplasia (short limbs) within a single breed (PBonferroni = 0.01) and intervertebral disc disease (IVDD) across breeds (PBonferroni = 4.0 × 10-10) both identified a significant association to the same region on CFA12. Whole genome sequencing identified a highly expressed FGF4 retrogene within this shared region. The FGF4 retrogene segregated with limb length and had an odds ratio of 51.23 (95% CI = 46.69, 56.20) for IVDD. Long bone length in dogs is a unique example of multiple disease-causing retrocopies of the same parental gene in a mammalian species. FGF signaling abnormalities have been associated with skeletal dysplasia in humans, and our findings present opportunities for both selective elimination of a medically and financially devastating disease in dogs and further understanding of the ever-growing complexity of retrogene biology

Motion Corrected Cadence CPS Ultrasound for Quantifying Response to Vasoactive Drugs in a Rat Kidney Model

To establish the ability of contrast enhanced motion corrected Cadence Pulse Sequencing (CPS) to detect changes in renal blood flow induced by vasoactive substances in rats

Paediatric meningitis in the conjugate vaccine era and a novel clinical decision model to predict bacterial aetiology

Objectives The aims of this study were to assess aetiology and clinical characteristics in childhood meningitis, and develop clinical decision rules to distinguish bacterial meningitis from other similar clinical syndromes. Methods Children aged <16 years hospitalised with suspected meningitis/encephalitis were included, and prospectively recruited at 31 UK hospitals. Meningitis was defined as identification of bacteria/viruses from cerebrospinal fluid (CSF) and/or a raised CSF white blood cell count. New clinical decision rules were developed to distinguish bacterial from viral meningitis and those of alternative aetiology. Results The cohort included 3002 children (median age 2·4 months); 1101/3002 (36·7%) had meningitis, including 180 bacterial, 423 viral and 280 with no pathogen identified. Enterovirus was the most common pathogen in those aged <6 months and 10–16 years, with Neisseria meningitidis and/or Streptococcus pneumoniae commonest at age 6 months to 9 years. The Bacterial Meningitis Score had a negative predictive value of 95·3%. We developed two clinical decision rules, that could be used either before (sensitivity 82%, specificity 71%) or after lumbar puncture (sensitivity 84%, specificity 93%), to determine risk of bacterial meningitis. Conclusions Bacterial meningitis comprised 6% of children with suspected meningitis/encephalitis. Our clinical decision rules provide potential novel approaches to assist with identifying children with bacterial meningitis. Funding This study was funded by the Meningitis Research Foundation, Pfizer and the NIHR Programme Grants for Applied Research

Electrodynamic Dust Shield for Space Applications

Dust mitigation technology has been highlighted by NASA and the International Space Exploration Coordination Group (ISECG) as a Global Exploration Roadmap (GER) critical technology need in order to reduce life cycle cost and risk, and increase the probability of mission success. The Electrostatics and Surface Physics Lab in Swamp Works at the Kennedy Space Center has developed an Electrodynamic Dust Shield (EDS) to remove dust from multiple surfaces, including glass shields and thermal radiators. Further development is underway to improve the operation and reliability of the EDS as well as to perform material and component testing outside of the International Space Station (ISS) on the Materials on International Space Station Experiment (MISSE). This experiment is designed to verify that the EDS can withstand the harsh environment of space and will look to closely replicate the solar environment experienced on the Moon

Pneumococcal lineages associated with serotype replacement and antibiotic resistance in childhood invasive pneumococcal disease in the post-PCV13 era: an international whole-genome sequencing study

Background: Invasive pneumococcal disease remains an important health priority owing to increasing disease incidence caused by pneumococci expressing non-vaccine serotypes. We previously defined 621 Global Pneumococcal Sequence Clusters (GPSCs) by analysing 20 027 pneumococcal isolates collected worldwide and from previously published genomic data. In this study, we aimed to investigate the pneumococcal lineages behind the predominant serotypes, the mechanism of serotype replacement in disease, as well as the major pneumococcal lineages contributing to invasive pneumococcal disease in the post-vaccine era and their antibiotic resistant traits. Methods: We whole-genome sequenced 3233 invasive pneumococcal disease isolates from laboratory-based surveillance programmes in Hong Kong (n=78), Israel (n=701), Malawi (n=226), South Africa (n=1351), The Gambia (n=203), and the USA (n=674). The genomes represented pneumococci from before and after pneumococcal conjugate vaccine (PCV) introductions and were from children younger than 3 years. We identified predominant serotypes by prevalence and their major contributing lineages in each country, and assessed any serotype replacement by comparing the incidence rate between the pre-PCV and PCV periods for Israel, South Africa, and the USA. We defined the status of a lineage as vaccine-type GPSC (≥50% 13-valent PCV [PCV13] serotypes) or non-vaccine-type GPSC (>50% non-PCV13 serotypes) on the basis of its initial serotype composition detected in the earliest vaccine period to measure their individual contribution toward serotype replacement in each country. Major pneumococcal lineages in the PCV period were identified by pooled incidence rate using a random effects model. Findings: The five most prevalent serotypes in the PCV13 period varied between countries, with only serotypes 5, 12F, 15B/C, 19A, 33F, and 35B/D common to two or more countries. The five most prevalent serotypes in the PCV13 period varied between countries, with only serotypes 5, 12F, 15B/C, 19A, 33F, and 35B/D common to two or more countries. These serotypes were associated with more than one lineage, except for serotype 5 (GPSC8). Serotype replacement was mainly mediated by expansion of non-vaccine serotypes within vaccine-type GPSCs and, to a lesser extent, by increases in non-vaccine-type GPSCs. A globally spreading lineage, GPSC3, expressing invasive serotypes 8 in South Africa and 33F in the USA and Israel, was the most common lineage causing non-vaccine serotype invasive pneumococcal disease in the PCV13 period. We observed that same prevalent non-vaccine serotypes could be associated with distinctive lineages in different countries, which exhibited dissimilar antibiotic resistance profiles. In non-vaccine serotype isolates, we detected significant increases in the prevalence of resistance to penicillin (52 [21%] of 249 vs 169 [29%] of 575, p=0·0016) and erythromycin (three [1%] of 249 vs 65 [11%] of 575, p=0·0031) in the PCV13 period compared with the pre-PCV period. Interpretation: Globally spreading lineages expressing invasive serotypes have an important role in serotype replacement, and emerging non-vaccine serotypes associated with different pneumococcal lineages in different countries might be explained by local antibiotic-selective pressures. Continued genomic surveillance of the dynamics of the pneumococcal population with increased geographical representation in the post-vaccine period will generate further knowledge for optimising future vaccine design. Funding: Bill & Melinda Gates Foundation, Wellcome Sanger Institute, and the US Centers for Disease Control.Fil: Lo, Stephanie W.. Wellcome Sanger Institute; Reino UnidoFil: Gladstone, Rebecca A.. Wellcome Sanger Institute; Reino UnidoFil: van Tonder, Andries J.. Wellcome Sanger Institute; Reino UnidoFil: Lees, John A.. University Of New York. School Of Medicine; Estados UnidosFil: du Plessis, Mignon. National Institute For Communicable Diseases; SudáfricaFil: Benisty, Rachel. Ben Gurion University of the Negev; IsraelFil: Givon Lavi, Noga. Ben Gurion University of the Negev; IsraelFil: Hawkins, Paulina A.. University of Emory. Rollins School of Public Health; Estados UnidosFil: Cornick, Jennifer E.. Malawi liverpool wellcome trust; MalauiFil: Kwambana Adams, Brenda. University College London; Estados UnidosFil: Law, Pierra Y.. University of Hong Kong; ChinaFil: Ho, Pak Leung. University of Hong Kong; ChinaFil: Antonio, Martin. Medical Research Council Unit The Gambia; GambiaFil: Everett, Dean B.. University of Edinburgh; Reino UnidoFil: Dagan, Ron. Ben Gurion University of the Negev; IsraelFil: Von Gottberg, Anne. National Institute For Communicable Diseases; SudáfricaFil: Klugman, Keith P.. University of Emory. Rollins School of Public Health; Estados UnidosFil: McGee, Lesley. Centers for Disease Control and Prevention; Estados UnidosFil: Breiman, Robert F.. University of Emory. Rollins School of Public Health; Estados UnidosFil: Bentley, Stephen D.. Wellcome Sanger Institute; Reino UnidoFil: Brooks, Abdullah W.. The Global Pneumococcal Sequencing Consortium; Reino UnidoFil: Corso, Alejandra. The Global Pneumococcal Sequencing Consortium; Reino Unido. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas. Área de Antimicrobianos; ArgentinaFil: Davydov, Alexander. The Global Pneumococcal Sequencing Consortium; Reino UnidoFil: Maguire, Alison. The Global Pneumococcal Sequencing Consortium; Reino UnidoFil: Pollard, Andrew. The Global Pneumococcal Sequencing Consortium; Reino UnidoFil: Kiran, Anmol. The Global Pneumococcal Sequencing Consortium; Reino UnidoFil: Skoczynska, Anna. The Global Pneumococcal Sequencing Consortium; Reino UnidoFil: Moiane, Benild. The Global Pneumococcal Sequencing Consortium; Reino UnidoFil: Beall, Bernard. The Global Pneumococcal Sequencing Consortium; Reino UnidoFil: Sigauque, Betuel. The Global Pneumococcal Sequencing Consortium; Reino UnidoFil: Aanensen, David. The Global Pneumococcal Sequencing Consortium; Reino UnidoFil: Lehmann, Deborah. The Global Pneumococcal Sequencing Consortium; Reino UnidoFil: Faccone, Diego Francisco. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas. Área de Antimicrobianos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin