bluuebunny/crossref_metadata_2025 huggingface dataset:
Implementation of a specialist pneumonia intervention nurse service significantly lowers mortality for community acquired pneumonia
Authors: Free Robert C. , Richardson Matthew , Pillay Camilla , Skeemer Julie , Hawkes Kayleigh , Broughton Rebecca , Haldar Pranabashis , Woltmann Gerrit
Published: October 2019

Objectives

Evaluate clinical outcomes associated with implementing a specialist pneumonia intervention nursing (SPIN) service, to improve adherence with BTS guidelines for hospitalised community acquired pneumonia (CAP).

Design

Retrospective cohort study, comparing periods before (2011-13) and after (2014-16) SPIN service implementation.

Setting

Single NHS trust across two hospital sites in Leicester City, England

Participants

13,496 adult (aged ≥16) admissions to hospital with a primary diagnosis of CAP

Interventions

The SPIN service was set up in 2013/2014 to provide clinical review of new CAP admissions; assurance of guidelines adherence; delivery of CAP clinical education and clinical follow up after discharge.

Main outcome measures

The primary outcomes were proportions of CAP cases receiving antibiotic treatment within 4 hours of admission and change in crude in-hospital mortality rate. Secondary outcomes were adjusted mortality rate and length of stay (LOS).

Results

The SPIN service reviewed 38% of CAP admissions in 2014-16. 82% of these admissions received antibiotic treatment in <4 hours (68.5% in the national audit). Compared with the pre-SPIN period, there was a significant reduction in both 30-day (OR=0.77 [0.70-0.85], p<0.0001) and in-hospital mortality (OR=0.66 [0.60-0.73], p<0.0001) after service implementation, with a review by the service having the largest independent 30-day mortality benefit (HR=0.60 [0.53-0.67], p<0.0001). There was no change in LOS (median 6 days).

Conclusions

Implementation of a SPIN service improves adherence with BTS guidelines and achieves significant reductions in CAP associated mortality. This enhanced model of care is low cost, highly effective and readily adoptable in secondary care.

Key Messages

What is the key question?

Does a specialist nurse-led intervention affect BTS guideline adherence and mortality for patients admitted to hospital with community acquired pneumonia (CAP)?

What is the bottom line?

Implementing specialist nurse teams for CAP delivers improved guideline adherence and survival for patients admitted with the condition.

Why read on?

This study shows a low-cost specialist nursing service focussed on CAP is associated with a significant improvement in BTS guidelines adherence and patient survival.
PERIPHERAL BLOOD AS TOOL TO DETERMINE GENE EXPRESSION PATTERNS IN PATIENTS WITH PSYCHIATRIC, NEUROLOGICAL AND OTHER COMMON DISORDERS: A SYSTEMATIC REVIEW AND META-ANALYSIS PROTOCOL
Authors: Panzenhagen Alana Castro , Alves-Teixeira Alexsander , Wissmann Martina Schroeder , Girardi Carolina Saibro , Santos Lucas , Silveira Alexandre Kleber , Gelain Daniel Pens , Moreira José Cláudio Fonseca
Published: October 2019

Introduction

Common diseases are influenced by a variety of factors that can enhance one person’s susceptibility to developing a specific condition. Complex traits have been investigated in several biological levels. One that reflects the high interconnectivity and interaction of genes, proteins and transcription factors is the transcriptome. In this study, we disclose the protocol for a systematic review and meta-analysis aiming at summarizing the available evidence regarding transcriptomic gene expression levels of peripheral blood samples comparing subjects with psychiatric, neurological and other common disorders to healthy controls.

Methods and analysis

The investigation of the transcriptomic levels in the peripheral blood enables the unique opportunity to unravel the etiology of common diseases in patients ex-vivo. However, the experimental results should be minimally consistent across studies for them to be considered as the best approximation of the true effect. In order to test this, we will systematically identify all transcriptome studies that compared subjects with common disorders to their respective control samples. We will apply meta-analyses to assess the overall differentially expressed genes throughout the studies of each condition.

Ethics and dissemination

The data that will be used to conduct this study are available online and have already been published following their own ethical laws. Therefore this study requires no further ethical approval. The results of this study will be published in leading peer-reviewed journals of the area and also presented at relevant national and international conferences.

Strengths and limitations of this study

➣

We present a new and systematically centered method to assess the overall effect of transcriptomic levels in the blood of subjects with common conditions.

➣

Meta-analyses are a robust statistical method to assess effect sizes across studies.

➣

The analysis is limited by the availability of studies, as well as their quality and comprehensiveness.

➣

Subgroup and meta-regression analyses will be also limited by the amount and quality of sample characterization variables made available by original studies.
A machine learning approach for precision diagnosis of juvenile-onset SLE
Authors: Robinson George A , Peng Junjie , Dönnes Pierre , Coelewij Leda , Radziszewska Anna , Wincup Chris , Peckham Hannah , Isenberg David A , Ioannou Yiannis , Ciurtin Coziana , Pineda-Torra Ines , Jury Elizabeth C
Published: October 2019

Juvenile-Onset systemic lupus erythematosus (JSLE) is an autoimmune rheumatic disease characterised by systemic inflammation and organ damage, with disease onset often coinciding with puberty. JSLE is associated with more severe disease manifestations and a higher motility rate compared to adult SLE. Due to the heterogeneous clinical and immunological manifestations of JSLE, delayed diagnosis and poor treatment efficacy are major barriers for improving patient outcome. In order to define a unique immunophenotyping profile distinguishing JSLE patients from age matched healthy controls, immune-based machine learning (ML) approaches were applied. Balanced random forest analysis discriminated JSLE patients from healthy controls with an overall 91% prediction accuracy. The top-ranked immunological features were selected from the optimal ML model and were validated by partial least squares discriminant analysis and logistic regression analysis. Patients could be clustered into four distinct groups based on the top hits from the ML model, providing an opportunity for tailored therapy. Moreover, complex correlations between the JSLE immune profile and clinical features of disease were identified. Further immunological association studies are essential for developing data-driven personalised medicine approaches to aid diagnosis of JSLE for targeted therapy and improved patient outcomes.
Forecasting Seizure Risk over Days
Authors: Proix Timothée , Truccolo Wilson , Leguia Marc G. , King-Stephens David , Rao Vikram R. , Baud Maxime O.
Published: October 2019

For persons with epilepsy, much suffering stems from the apparent unpredictability of seizures. Historically, efforts to predict seizures have sought to detect changes in brain activity in the seconds to minutes preceding seizures (pre-ictal period), a timeframe that limits preventative interventions. Recently, converging evidence from studies using chronic intracranial electroencephalography revealed that brain activity in epilepsy has a robust cyclical structure over hours (circadian) and days (multidien). These cycles organize pro-ictal states, hours-to days-long periods of heightened seizure risk, raising the possibility of forecasting seizures over horizons longer than the pre-ictal period. Here, using cEEG from 18 subjects, we developed point-process generalized linear models incorporating cyclical variables at multiple time-scales to show that seizure risk can be forecasted accurately over days in most subjects. Personalized risk-stratification days in advance of seizures is unprecedented and may enable novel preventative strategies.
A deep learning approach for building multiple trees classification
Authors: Tahiri Nadia
Published: October 2019

Each gene has its own evolutionary history which can substantially differ from the evolutionary histories of other genes. For example, some individual genes or operons can be affected by specific horizontal gene transfer or hybridization events. Thus, the evolutionary history of each gene should be represented by its own phylogenetic tree which may display different evolutionary patterns from the species tree, or Tree of Life, that represents the main patterns of vertical descent. Here, we present a new efficient method for inferring single or multiple consensus trees and supertrees for a given set of phylogenetic trees (i.e. additive trees or X-trees). The output of the traditional tree consensus methods is a unique consensus tree or supertree. Here, we show how Machine Learning (ML) models, based on some interesting properties of the Robinson and Foulds topological distance, can be used to partition a given set of trees into one (when the data are homogeneous) or multiple (when the data are heterogeneous) cluster(s) of trees. We adapt the popular Accuracy, Precision, Sensitivity, and F1 scores to the tree clustering. A special attention is paid to the relevant, but very challenging, problem of inferring alternative supertrees that are built from phylogenies defined on different, but mutually overlapping, sets of species. The use of an approximate objective function in clustering makes the new method faster than the existing tree clustering techniques and thus suitable for the analysis of large genomic datasets.
Cold exposure drives weight gain and adiposity following chronic suppression of brown adipose tissue
Authors: Aldiss Peter , Lewis Jo E , Lupini Irene , Bloor Ian , Chavoshinejad Ramyar , Boocock David , Miles Amanda K , Ebling Francis J P , Budge Helen , Symonds Michael E
Published: October 2019

Therapeutic activation of thermogenic brown adipose tissue (BAT) may be feasible to prevent, or treat, cardiometabolic disease. However, rodents are commonly housed below thermoneutrality (∼20°C) which can modulate their metabolism and physiology including the hyperactivation of brown (BAT) and beige white adipose tissue. We housed animals at thermoneutrality from weaning to chronically supress BAT, mimic human physiology and explore the efficacy of chronic, mild cold-exposure and β3-adrenoreceptor agonism under these conditions. Using metabolic phenotyping and exploratory proteomics we show that transfer from 28°C to 20°C drives weight gain and a 125% increase in subcutaneous fat mass, an effect not seen with YM-178 administration thus suggesting a direct effect of a cool ambient temperature in promoting weight gain and further adiposity in obese rats. Following chronic suppression of BAT, uncoupling protein 1 mRNA was undetectable in IWAT in all groups. Using exploratory adipose tissue proteomics, we reveal novel gene ontology terms associated with cold-induced weight gain in BAT and IWAT whilst Reactome pathway analysis highlights the regulation of mitotic (i.e. G2/M transition) and metabolism of amino acids and derivatives pathways. Conversely, YM-178 had minimal metabolic-related effects but modified pathways involved in proteolysis (i.e. eukaryotic translation initiation) and RNA surveillance across both tissues. Taken together these findings are indicative of a novel mechanism whereby animals increase body weight and fat mass following chronic suppression of adaptive thermogenesis from weaning. In addition, treatment with a B3-adrenoreceptor agonist did not improve metabolic health in obese animals raised at thermoneutrality.
Developing a xenograft model of human vasculature in the mouse ear pinna
Authors: Meehan Gavin R. , Scales Hannah E. , Osii Rowland , De Niz Mariana , Lawton Jennifer C. , Marti Matthias , Garside Paul , Craig Alister , Brewer James M.
Published: October 2019

Humanised xenograft models allow for the analysis of human tissue within a physiological environmentin vivo. However, current models often rely on the angiogenesis and ingrowth of recipient vasculature to perfuse tissues, preventing analysis of biological processes and diseases involving human blood vessels. This limits the effectiveness of xenografts in replicating human physiology and may lead to issues with translating findings into human research.

We have designed a xenograft model of human vasculature to address this issue. Human subcutaneous fat was culturedin vitroto promote blood vessel outgrowth prior to implantation into immunocompromised mice. We demonstrate that implants survived, retained human vasculature and anastomosed with the circulatory system of the recipient mouse. Significantly, by performing transplants into the ear pinna, this system enabled intravital observation of xenografts by multiphoton microscopy, allowing us to visualise the steps leading to vascular cytoadherence of erythrocytes infected with the human parasitePlasmodium falciparum.

This model represents a useful tool for imaging the interactions that occur within human tissuesin vivoand permits visualization of blood flow and cellular recruitment in a system which is amenable to intervention for various studies in basic biology together with drug evaluation and mechanism of action studies.
DominantVibrio choleraephage exhibits lysis inhibition sensitive to disruption by a defensive phage satellite
Authors: Hays Stephanie G. , Seed Kimberley D.
Published: October 2019

Bacteriophages and their bacterial hosts are locked in a dynamic evolutionary arms race. Phage satellites, selfish genomic islands which exploit both host bacterium and target phage, further complicate the evolutionary fray. One such tripartite system involves the etiological agent of the diarrheal disease cholera –Vibrio cholerae, the predominant phage isolated from cholera patients – ICP1, and a phage satellite – PLE. When ICP1 infectsV. choleraeharboring the integrated PLE genome, PLE accelerates host lysis, spreading the PLE while completely blocking phage production protectingV. choleraeat the population level. Here we identify a single PLE gene,lidI, sufficient to mediate accelerated lysis during ICP1 infection and demonstrate that LidI functions through disrupting lysis inhibition – an understudied outcome of phage infection when phages vastly outnumber their hosts. This work identifies ICP1-encoded holin and antiholin genesteaAandarrArespectively, that mediate this first example of lysis inhibition outside the T-even coliphages. Throughlysisinhibitiondisruption, LidI is sufficient to limit the number of progeny phage produced from an infection. Consequently, this disruption bottlenecks ICP1 evolution as probed by recombination and CRISPR-Cas targeting assays. These studies link novel characterization of the classic phenomenon of lysis inhibition with a conserved protein in a dominant phage satellite, highlighting the importance of lysis timing during infection and parasitization, as well as providing insight into the populations, relationships, and evolution of bacteria, phages, and phage satellites in nature.

Importance

With increasing awareness of microbiota impacting human health comes intensified examination of, not only bacteria and the bacteriophages that prey upon them, but also the mobile genetic elements (MGEs) that mediate interactions between them. Research is unveiling evolutionary strategies dependent on sensing the milieu: quorum sensing impacts phage infection, phage teamwork overcomes bacterial defenses, and abortive infections sacrifice single cells protecting populations. Yet, the first discovered environmental sensing by phages, known as lysis inhibition (LIN), has only been studied in the limited context of T-even coliphages. Here we characterize LIN in the etiological agent of the diarrheal disease cholera,Vibrio cholerae, infected by a phage ubiquitous in clinical samples. Further, we show that a specific MGE, the phage satellite PLE, collapses LIN with a conserved protein during its anti-phage program. The insights gleaned from this work add to our expanding understanding of microbial fitness in natural contexts beyond the canonical bacterial genome and into the realm of antagonistic evolution driven by phages and satellites.
Mindfulness meditation alters neural activity underpinning working memory during tactile distraction
Authors: Wang Michael Yufeng , Freedman Gabrielle , Raj Kavya , Fitzgibbon Bernadette Mary , Sullivan Caley , Tan Wei-Lin , Van Dam Nicholas , Fitzgerald Paul B , Bailey Neil W
Published: October 2019

Evidence suggests that mindfulness meditation (MM) improves selective attention and reduces distractibility by enhancing top-down neural modulation. Altered P300 and alpha neural activity from MM have been identified and may reflect the neural changes that underpin these improvements. Given the proposed role of alpha activity in supressing processing of task-irrelevant information, it is theorised that altered alpha activity may underlie increased availability of neural resources in meditators. The present study investigated attentional function in meditators using a cross-modal study design, examining the P300 during working memory (WM) and alpha activity during concurrent distracting tactile stimuli. Thirty-three meditators and 27 healthy controls participated in the study. Meditators showed a more frontal distribution of P300 neural activity following WM stimuli (p = 0.005, η² = 0.060) and more modulation of alpha activity at parietal-occipital regions between single (tactile stimulation only) and dual task demands (tactile stimulation plus WM task) (p < 0.001, η² = 0.065). Additionally, meditators performed more accurately than controls (p = 0.038, η² = 0.067). The altered distribution of neural activity concurrent with improved WM performance suggests greater attentional resources dedicated to task related functions such as WM in meditators. Thus, meditation-related neural changes are likely multi-faceted involving both altered distribution and also amplitudes of brain activity, enhancing attentional processes depending on task requirements.
Functional characterization and lineage analysis of broadly neutralizing human antibodies against dengue virus identified by single B cell transcriptomics
Authors: Durham Natasha D. , Agrawal Aditi , Waltari Eric , Croote Derek , Zanini Fabio , Davidson Edgar , Fouch Mallorie , Smith Olivia , Carabajal Esteban , Pak John E. , Doranz Benjamin J. , Robinson Makeda , Sanz Ana M. , Albornoz Ludwig L. , Rosso Fernando , Einav Shirit , Quake Stephen R. , McCutcheon Krista M. , Goo Leslie
Published: October 2019

Eliciting broadly neutralizing antibodies (bNAbs) against the four dengue virus serotypes (DENV1-4) that are spreading into new territories is an important goal of vaccine design. To delineate bNAb targets, we characterized 28 monoclonal antibodies belonging to expanded and hypermutated clonal families identified by transcriptomic analysis of single plasmablasts from DENV-infected individuals. Among these, we identified two somatically related bNAbs that potently neutralized DENV1-4. Mutagenesis studies revealed that the major recognition determinants of these bNAbs are in E protein domain I, distinct from the only known class of human bNAbs against flaviviruses with a well-defined epitope. B cell repertoire analysis from acute-phase peripheral blood suggested a memory origin and divergent somatic hypermutation pathways for these bNAbs, and a limited number of mutations was sufficient for neutralizing activity. Our study suggests multiple B cell evolutionary pathways leading to DENV bNAbs targeting a novel epitope that can be exploited for vaccine design.
OncogenicGata1causes stage-specific megakaryocyte differentiation delay
Authors: Juban Gaëtan , Sakakini Nathalie , Chagraoui Hedia , Cheng Qian , Soady Kelly , Stoilova Bilyana , Garnett Catherine , Waithe Dominic , Doondeea Jess , Usukhbayar Batchimeg , Karkoulia Elena , Alexiou Maria , Strouboulis John , Morrissey Edward , Roberts Irene , Porcher Catherine , Vyas Paresh
Published: October 2019

The megakaryocyte/erythroid Transient Myeloproliferative Disorder (TMD) in newborns with Down Syndrome (DS) occurs when N-terminal truncating mutations of the hemopoietic transcription factor GATA1, that produce GATA1short protein (GATA1s), are acquired early in development. Prior work has shown that murine GATA1s, by itself, causes a transient yolk sac myeloproliferative disorder. However, it is unclear where in the hemopoietic cellular hierarchy GATA1s exerts its effects to produce this myeloproliferative state. Here, through a detailed examination of hemopoiesis from murine GATA1s ES cells and GATA1s embryos we define defects in erythroid and megakaryocytic differentiation that occur relatively in hemopoiesis. GATA1s causes an arrest late in erythroid differentiationin vivo, and even more profoundly in ES-cell derived cultures, with a marked reduction of Ter-119 cells and reduced erythroid gene expression. In megakaryopoiesis, GATA1s causes a differentiation delay at a specific stage, with accumulation of immature, kit-expressing CD41himegakaryocytic cells. In this specific megakaryocytic compartment, there are increased numbers of GATA1s cells in S-phase of cell cycle and reduced number of apoptotic cells compared to GATA1 cells in the same cell compartment. There is also a delay in maturation of these immature GATA1s megakaryocytic lineage cells compared to GATA1 cells at the same stage of differentiation. Finally, even when GATA1s megakaryocytic cells mature, they mature aberrantly with altered megakaryocyte-specific gene expression and activity of the mature megakaryocyte enzyme, acetylcholinesterase. These studies pinpoint the hemopoietic compartment where GATA1s megakaryocyte myeloproliferation occurs, defining where molecular studies should now be focussed to understand the oncogenic action of GATA1s.

Scientific Category

Haematopoiesis and Stem Cells

Key Points

GATA1s-induced stage-specific differentiation delay increases immature megakaryocytesin vivoandin vitro, during development.

Differentiation delay is associated with increased numbers of cells in S-phase and reduced apoptosis.
A simple model for the evolution of temperature-dependent sex determination explains the temperature sensitivity of embryonic mortality in imperiled reptiles
Authors: Lawson Lauren , Rollinson Njal
Published: October 2019

A common reptile conservation strategy involves artificial incubation of embryos and release of hatchlings or juveniles into wild populations. Temperature-dependent sex determination (TSD) occurs in most chelonians, permitting conservation managers to bias sex ratios towards females by incubating embryos at high temperatures, ultimately allowing the introduction of more egg-bearing individuals into populations. Here, we revisit classic sex allocation theory and hypothesize that TSD evolved in some reptile groups (specifically, chelonians and crocodilians) because male fitness is more sensitive to condition (general health, vigor) than female fitness. It follows that males benefit more than females from incubation environments that confer high-quality phenotypes, and hence high-condition individuals. We predict that female-producing temperatures, which comprise relatively high incubation temperatures in chelonians and crocodilians, are relatively stressful for embryos and subsequent life stages. We synthesize data from 28 studies to investigate how constant temperature incubation affects embryonic mortality in chelonians with TSD. We find several lines of evidence suggesting that female-producing temperatures, especially relatively warm temperatures, are more stressful than male-producing temperatures, and we find some evidence that pivotal temperatures (TPiv, the temperature that produces a 1:1 sex ratio) exhibit a correlated evolution with embryonic thermal tolerance. If patterns of temperature-sensitive embryonic mortality are also indicative of chronic thermal stress that occurs post hatching, then conservation programs may benefit from incubating eggs close to species-specific TPivs, thus avoiding high-temperature incubation. Indeed, our models predict that, on average, a sex ratio of more than 75% females can generally be achieved by incubating eggs only 1°C above TPiv. Of equal importance, we provide insight into the enigmatic evolution of TSD in chelonians, by providing support to the hypothesis that TSD evolution is related to the quality of the phenotype conferred by incubation temperature, with males produced in high-quality incubation environments.

Lay summary

We analyze data on embryonic mortality under constant-temperature incubation for 15 species of chelonians with temperature-dependent sex determination (TSD). Mortality is lowest near species-specific pivotal temperatures (Tpiv) but increases rapidly above TPiv, consistent with a theory that explains the adaptive significance of TSD. Conservation managers should incubate embryos near TPiv.
Secretin release after Roux-en-Y Gastric Bypass reveals a population of glucose-sensitive S-cells in distal small intestine
Authors: Modvig Ida M. , Andersen Daniel B. , Grunddal Kaare V. , Kuhre Rune E. , Martinussen Christoffer , Christiansen Charlotte B. , Ørskov Cathrine , Larraufie Pierre , Kay Richard , Reimann Frank , Gribble Fiona M. , Hartmann Bolette , Bojsen-Møller Kirstine N. , Madsbad Sten , Wewer Albrechtsen Nicolai J. , Holst Jens J.
Published: October 2019

Objective

Gastrointestinal hormones contribute to the beneficial effects of Roux-en-Y gastric bypass surgery (RYGB) on glycemic control. Secretin is secreted from duodenal S-cells in response to low luminal pH, but it is unknown whether its secretion is altered after RYGB and if secretin contributes to the post-operative improvement in glycemic control. We hypothesized that secretin secretion increases after RYGB as a result of the diversion of nutrients to more distal parts of the small intestine, and thereby affects islet hormone release.

Methods

A specific secretin radioimmunoassay was developed, evaluated biochemically, and used to quantify plasma concentrations of secretin in 13 obese individuals before, 1 week after and 3 months after RYGB. Distribution of secretin and its receptor was assessed by RNA-sequencing, mass-spectrometry andin situhybridization in human and rat tissues. Isolated, perfused rat intestine and pancreas were used to explore the molecular mechanism underlying glucose-induced secretin secretion and to study direct effects of secretin on glucagon, insulin and somatostatin secretion. Secretin was administered alone or in combination with GLP-1 to non-sedated rats to evaluate effects on glucose regulation.

Results

Plasma postprandial secretin was more than doubled in humans after RYGB (P<0.001). The distal small intestine harbored secretin expressing cells in both rats and humans. Glucose increased secretion of secretin in a sodium-glucose co-transporter dependent manner when administered to the distal part but not into the proximal part of the rat small intestine. Secretin stimulated somatostatin secretion (fold change: 1.59, P<0.05) from the perfused rat pancreas but affected neither insulin (P=0.2) nor glucagon (P=0.97) secretion. When administered to ratsin vivo, insulin secretion was attenuated and glucagon secretion increased (P=0.04), while blood glucose peak time was delayed (from 15 min to 45 min) and gastric emptying time prolonged (P=0.004).

Conclusion

Glucose-sensing secretin cells located in the distal part of the small intestine may contribute to increased plasma concentrations observed after RYGB. The metabolic role of the distal S-cells warrants further studies.
Microfluidic trapping of vesicles reveals membrane-tension dependent FtsZ cytoskeletal re-organisation
Authors: Ganzinger Kristina A. , Merino-Salomón Adrián , García-Soriano Daniela A. , Nelson Butterfield A. , Litschel Thomas , Siedler Frank , Schwille Petra
Published: October 2019

The geometry of reaction compartments can affect the outcome of chemical reactions. Synthetic biology commonly uses giant unilamellar vesicles (GUVs) to generate cell-sized, membrane-bound reaction compartments. However, these liposomes are always spherical due to surface area minimization. Here, we have developed a microfluidic chip to trap and reversibly deform GUVs into rod- or cigar-like shapes, including a constriction site in the trap mimicking the membrane furrow in cell division. When we introduce into these GUVs the bacterial tubulin homologue FtsZ, the primary protein of the bacterial Z ring, we find that FtsZ organization changes from dynamic rings to elongated filaments upon GUV deformation, and that these FtsZ filaments align preferentially with the short GUV axis, in particular at the membrane neck. In contrast, pulsing Min oscillations in GUVs remained largely unaffected. We conclude that microfluidic traps are a useful tool for deforming GUVs into non-spherical membrane shapes, akin to those seen in cell division, and for investigating the effect of confinement geometry on biochemical reactions, such as protein filament self-organization.
Repurposing theStreptococcus mutansCRISPR-Cas9 System to Understand Essential Gene Function
Authors: Shields RC , Walker AR , Maricic N , Chakraborty B , Underhill SAM , Burne RA
Published: October 2019

A recent genome-wide screen identified ∼300 essential or growth-supporting genes in the dental caries pathogenStreptococcus mutans. To be able to study these genes, we built a CRISPR interference tool around the Cas9 nuclease (Cas9Smu) encoded in theS. mutansUA159 genome. Using a xylose-inducible dead Cas9Smuwith a constitutively active single-guide RNA (sgRNA), we observed titratable repression of GFP fluorescence that compared favorably to that ofStreptococcus pyogenesdCas9 (Cas9Spy). We then investigated sgRNA specificity and proto-spacer adjacent motif (PAM) requirements. Interference by sgRNAs did not occur with double or triple base-pair mutations, or if single base-pair mutations were in the 3’ end of the sgRNA. Bioinformatic analysis of >450S. mutansgenomes allied within vivoassays revealed a similar PAM recognition sequence as the Cas9Spy. Next, we created a comprehensive library of sgRNA plasmids that were directed at essential and growth-supporting genes. We discovered growth defects for 77% of the CRISPRi strains expressing sgRNAs. Phenotypes of CRISPRi strains, across several biological pathways, were assessed using fluorescence microscopy. A variety of cell structure anomalies were observed, including segregational instability of the chromosome, enlarged cells, and ovococci-to-rod shape transitions. CRISPRi was also employed to observe how silencing of cell wall glycopolysaccharide biosynthesis (rhamnose-glucose polysaccharide, RGP) affected both cell division and pathogenesis in a wax worm model. The CRISPRi tool and sgRNA library are valuable resources for characterizing essential genes inS. mutans, some of which could prove to be promising therapeutic targets.
SCryPTA:A web-based platform for analyzing Small-Angle Scattering curves of lyotropic liquid crystals
Authors: de Castro Raphael Dias , Casadei Bruna Renata , Santana Barbara Vasconcelos , Lotierzo Mayra , de Oliveira Natália F. , Malheiros Barbara , Mariani Paolo , Kaminski Renata C. K. , Barbosa Leandro R. S.
Published: October 2019

Small angle X-ray scattering (SAXS) is a powerful technique for the characterization of systems with highly ordered structures, such as liquid crystals and self-assembly systems. In the field of nanotechnology and nanomedicine, SAXS can be used to characterize the crystallographic properties of the crystal phase of lyotropic systems and nanoparticles with internal crystal phase, such as cubosomes, hexosomes and multi-lamellar vesicles. In this work, we introduce a new web platform namedSCryPTA: Small Angle Scattering Crystallographic Peak Treatment and Analysis, capable of reading SAXS data and providing a comprehensive visualization of the scattering curve along with the calculation of important physical parameters, such as the lattice parameter of the crystal structure, the lipidic bilayer width, among others. Cubic, hexagonal and multilamellar scattering data had their crystallographic structure characterized in SCryPTA. So far, four different cubic structures, (Pn3m(Q224),Fd3m(Q227),Im3m(Q229),Ia3d(Q230)), the hexagonal phase and also multi-lamellar vesicle systems are described in the platform. We believe that SCryPTA may help researchers from several fields, since it has a user-friendly interface. The platform is available at:www.if.usp.br/scrypta.
The consequences of surviving infection across the metamorphic boundary: tradeoff insights from RNAseq and life history measures
Authors: Keehnen Naomi L.P. , Kučerová Lucie , Nylin Sören , Theopold Ulrich , Wheat Christopher W.
Published: October 2019

The broad diversity of insect life has been shaped, in part, by pathogen pressure, yet the influence of injury and infection during critical periods of development is understudied. During development, insects undergo metamorphosis, wherein the organism experiences a dramatic shift in their overall morphology, and physiology. In temperate zones, metamorphosis is often directly followed by a developmental arrest called diapause, for which the insect needs to acquire enough energy reserves before the onset of winter. We investigated the long-term effects of injury and infection using two bacteria in the butterfly Pieris napi, revealing that the negative consequences of bacterial infection carry across the metamorphic boundary. Initial direct effects of infection were weight loss and slower development, as well as an increased mortality at higher infection levels. The detrimental effects were stronger in the gram-positive Micrococcus luteus compared to gram-negative Escherichia coli. Transcriptome-wide differences between the two bacteria were already observed in the gene expression profile of the first 24 hours after infection. Larvae infected with M. luteus showed a strong suppression of all non-immunity related processes, with several types of immune responses being activated. The impact of these transcriptomic changes, a tradeoff between homeostasis and immune response, were visible in the life history data, wherein individuals infected with M. luteus had the highest mortality rate, along with the lowest pupal weight, developmental rate and adult weight of all the treatments. Overall, we find that the cost of infection and wounding in the final larval instar carries over the metamorphic boundary, and is expected to negatively affect their lifetime fitness.
Hippocampal neural stem cells facilitate access from circulation via apical cytoplasmic processes
Authors: Licht Tamar , Keshet Eli
Published: October 2019

Blood vessels (BVs) are considered an integral component of neural stem cells (NSCs) niches. NSCs in the dentate gyrus (DG) have enigmatic elaborated apical cellular processes that are associated with BVs. Whether this contact serves as a mechanism for delivering circulating molecules is not known. Here we uncovered a previously unrecognized communication route allowing exclusive direct access of blood-borne substances to hippocampal NSCs in defiance of an intact blood-brain barrier (BBB). BBB-impermeable fluorescent tracer injected transcardially is selectively uptaken by DG NSCs within a minute, via the vessel-associated apical processes. These processes, measured >30nm in diameter, establish direct membrane-to-membrane contact with endothelial cells in areas devoid of the endothelial basement membrane. Doxorubicin, a brain-impermeable chemotherapeutic agent, is also readily and selectively uptaken by NSCs and reduces their proliferation, which might explain its problematic anti-neurogenic or cognitive side-effect. The newly-discovered NSC-BV communication route explains how circulatory neurogenic mediators are ‘sensed’ by NSCs.
Metabarcoding unsorted kick-samples facilitates macroinvertebrate-based biomonitoring with increased taxonomic resolution, while outperforming environmental DNA
Authors: Pereira-da-Conceicoa Lyndall , Elbrecht Vasco , Hall Andie , Briscoe Andrew , Barber-James Helen , Price Benjamin
Published: October 2019

Many studies have highlighted the potential of DNA-based methods for the biomonitoring of freshwater macroinvertebrates, however only a few studies have investigated homogenisation of bulk samples that include debris to reduce sample-processing time. In order to explore the use of DNA-based methods in water quality assessment in South Africa, this study compares morphological and molecular-based identification of freshwater macroinvertebrates at the mixed higher taxon and mOTU level while investigating abundance and comparing mOTU recovery with historical species records. From seven sites across three rivers in South Africa, we collected a biomonitoring sample, an intensive-search comprehensive sample and an eDNA sample per site. The biomonitoring sample was picked and scored according to standard protocols and the leftover debris and comprehensive samples were homogenised including all debris. DNA-based methods recovered higher diversity than morphology, but did not always recover the same taxa, even at the family level. Regardless of the differences in taxon scores, most DNA-based methods except some eDNA samples, returned the same water quality assessment category as the standard morphology-based assessment. Homogenised comprehensive samples recovered more freshwater invertebrate diversity than all other methods. The eDNA samples recovered 2 to 10 times more mOTUs than any other method, however 90% of reads were non-target and as a result eDNA recovered the lowest target diversity. However, eDNA did find some target taxa that the other methods failed to detect. This study shows that unsorted samples recover the same water quality scores as a morphology-based assessment and much higher diversity scores than both picked and eDNA samples. As a result, there is potential to integrate DNA-based approaches into existing metrics quickly while providing much more information for the development of more refined metrics at the species or mOTU level with distributional data which can be used for conservation and biodiversity management.
Self-Incompatibility Alleles in Iranian Pear Cultivars
Authors: Bagheri Maryam , Ershadi Ahmad
Published: October 2019

In the present study, the S-alleles of eighteen pear cultivars, (including fourteen cultivars planted commercially in Iran and four controls) are determined. 34 out of 36 S-alleles are detected using nine allele-specific primers, which are designed for amplification of S101/S102, S105, S106, S107, S108, S109, S111, S112 and S114, as well as consensus primers, PycomC1F and PycomC5R. S104, S101 and S105 were the most common S-alleles observed, respectively, in eight, seven and six cultivars. In 16 cultivars, (‘Bartlett’ (S101S102), ‘Beurre Giffard’ (S101S106), ‘Comice’ (S104S105), ‘Doshes’ (S104S107), ‘Koshia’ (S104S108), ‘Paskolmar’ (S101S105), ‘Felestini’ (S101S107), ‘Domkaj’ (S104S120), ‘Ghousi’ (S104S107), ‘Kaftar Bache’ (S104S120), ‘Konjoni’ (S104S108), ‘Laleh’ (S105S108), ‘Natanzi’ (S104S105), ‘Sebri’ (S101S104), ‘Se Fasleh’ (S101S105) and ‘Louise Bonne’ (S101S108)) both alleles are identified but in two cultivars, (‘Pighambari’ (S105) and ‘Shah Miveh Esfahan’ (S107)) only one allele is recognized. It is concluded that allele-specific PCR amplification can be considered as an efficient and rapid method to identify S-genotype of Iranian pear cultivars.
Development of a multiparent advanced generation intercross (MAGIC) population for genetic exploitation of complex traits inBrassica juncea: glucosinolate content as an example
Authors: Wang Tianya , Wan Wei , Yu Kunjiang , Khattak Aimal Nawaz , Ye Botao , Yang Renqin , Tian Entang
Published: October 2019

Multiparent advanced generation intercross (MAGIC) populations have recently been developed to allow the high-resolution mapping of complex quantitative traits. This article describes the development of one MAGIC population and verifies its potential application for mapping quantitative trait loci (QTLs) inB. juncea. The population was developed from eight founders with diverse traits and composed of 408 F6recombinant inbred lines (RILs). To develop one rapid and simplified way for using the MAGIC population, a subset of 133 RILs as the primary mapping population were genotyped using 346 intron-length polymorphism (ILP) polymorphic markers. The population lacks significant signatures of population structure that are suitable for the analysis of complex traits. Genome-wide association mapping (GWAS) identified three major glucosinolate (GSL) QTLs ofQGsl.ig01.1on J01 for indole GSL (IG),QGsl.atg09.1on J09 andQGsl.atg11.1on J11 for aliphatic GSL (AG) and total GSL (TG). The candidate genes forQGsl.ig01.1, QGsl.atg09.1andQGsl.atg11.1areGSH1, GSL-ALKandMYB28, which are involved in converting glutamate and cysteine toγ–EC, the accumulation of glucoraphanin, and the whole process of AG metabolism, respectively. One effective method for association mapping of quantitative traits in theB. junceaMAGIC population is also suggested by utilization of the remaining 275 RILs and incorporation of the novel kompetitive allele specific PCR (KASP) technique. In addition to its QTL mapping purpose, the MAGIC population could also be potentially utilized in variety development by breeders.
SCRIBE: a new approach to dropout imputation and batch effects correction for single-cell RNA-seq data
Authors: Zhang Yiliang , Liang Kexuan , Liu Molei , Li Yue , Ge Hao , Zhao Hongyu
Published: October 2019

Single-cell RNA sequencing technologies are widely used in recent years as a powerful tool allowing the observation of gene expression at the resolution of single cells. Two of the major challenges in scRNA-seq data analysis are dropout events and batch effects. The inflation of zero(dropout rate) varies substantially across single cells. Evidence has shown that technical noise, including batch effects, explains a notable proportion of this cell-to-cell variation. To capture biological variation, it is necessary to quantify and remove technical variation. Here, we introduce SCRIBE (Single-Cell Recovery Imputation with Batch Effects), a principled framework that imputes dropout events and corrects batch effects simultaneously. We demonstrate, through real examples, that SCRIBE outperforms existing scRNA-seq data analysis tools in recovering cell-specific gene expression patterns, removing batch effects and retaining biological variation across cells. Our software is freely available online at https://github.com/YiliangTracyZhang/SCRIBE.
Not all MDAs should be created equal – determinants of MDA impact and designing MDAs towards malaria elimination
Authors: Gao B. , Saralamba S. , Lubell Y. , White L. J. , Dondorp A. , Aguas R.
Published: October 2019

Malaria remains at the forefront of scientific research and global political and funding agendas. Previous malaria models of mass-interventions have consistently oversimplified how mass interventions are implemented. We present an individual based, spatially explicit model of malaria transmission that includes all the programmatic implementation details of mass drug administration (MDA) campaigns. We uncover how the impact of MDA campaigns is determined by the interaction between implementation logistics, patterns of human mobility and how transmission risk is distributed over space. This translates into a higher likelihood of malaria elimination for areas with true prevalence under 3% with a faster implementation, in highly mobile populations. If populations are more static, deploying less interventions teams would be cost optimal and predicted to be as impactful. We conclude that mass drug interventions can be an invaluable tool towards malaria elimination in the right context, specifically when paired with effective vector control.
Reproducibility warning: The curious case of Polyethylene glycol 6000 and spheroid cell culture
Authors: Serrati Simona , Martinelli Chiara , Palazzo Antonio , Iacobazzi Rosa Maria , Perrone Mara , Santoliquido Roberto , Ong Quy K. , Luo Zhi , Bekdemir Ahmet , Pinto Giulia , Cavalleri Ornella , Cutrignelli Annalisa , Laquintana Valentino , Denora Nunzio , Stellacci Francesco , Krol Silke
Published: October 2019

In this study we report about the reproducibility of three-dimensional cell culture of floating cell spheroids on PEG6000 treated cell culture dishes. Three-dimensional tumour spheroids or organoids present an interesting test platform for nanoparticulated drug delivery or nanoparticle toxicity. We tested the reproducibility of spheroid formation induced by the PEG coated surface. Interestingly we found that the results were different in a reproducible manner depending on the distributors of PEG6000.

Despite the nearly identical physicochemical properties of PEG6000 (MALDI-MS, NMR, FTIR, Triple SEC) with only minor differences, we observed only for one PEG6000 a highly reproducible formation of spheroids with different cell lines such as HT-29, HeLa, Caco2, and PANC-1. The surface coating with the different PEG6000 was studied by AFM. The surface coating as well as the physicochemical characterization showed only small differences in mass and hydrodynamic radius between the different PEGs. A direct coating of the cells with PEG from two distributors indicate that the spheroid formation in due to direct interaction of the polymer with the cell rather than by interaction of cells with the coated surface.

The experiments point out that for biological entities, such as cells or tissues, even very small differences such as impurities or batch-to-batch variations in the purchased product can have a very strong impact.
A LAMP at the end of the tunnel: a rapid, field deployable assay for the kauri dieback pathogen,Phytophthora agathidicida
Authors: Winkworth Richard C. , Nelson Briana C.W. , Bellgard Stanley E. , Probst Chantal M. , McLenachan Patricia A. , Lockhart Peter J.
Published: October 2019

The collar rot causing oomycete,Phytophthora agathidicida, threatens the long-term survival of the iconic New Zealand kauri. Currently, testing for this pathogen involves an extended soil bioassay that takes 14-20 days and requires specialised staff, consumables, and infrastructure. Here we describe a loop-mediated isothermal amplification (LAMP) assay for the detection ofP. agathidicidathat targets a portion of the mitochondrial apocytochrome b coding sequence. This assay has high specificity and sensitivity; it did not cross react with a range of otherPhytophthoraisolates and detected as little as 1 fg of totalP. agathidicidaDNA or 116 copies of the target locus. Assay performance was further investigated by testing plant tissue baits from flooded soil samples using both the extended bioassay and LAMP testing of DNA extracted from baits. In these comparisons,P. agathidicidawas detected more frequently using the LAMP assay. In addition to greater sensitivity, by removing the need for culturing, the hybrid baiting plus LAMP approach is more cost effective than the bioassay and, importantly, does not require a centralised laboratory facility with specialised staff, consumables, and equipment. Such testing will allow us to address outstanding questions aboutP. agathidicida. For example, the hybrid approach could enable monitoring of the pathogen beyond areas with visible disease symptoms, allow direct evaluation of rates and patterns of spread, and allow the effectiveness of disease control to be evaluated. The hybrid assay also has the potential to empower local communities. These communities could use this diagnostic tool to evaluate the pathogen status of local kauri stands, providing information around which to base their management and allowing informed engagement with wider initiatives.