bluuebunny/crossref_metadata_2025 huggingface dataset:
The Moran effect and environmental vetoes: phenological synchrony and drought drive seed production in a Mediterranean oak
Authors: Bogdziewicz Michał , Fernández-Martínez Marcos , Bonal Raul , Belmonte Jordina , Espelta Josep Maria
Published: November 2017

Masting is the highly variable production of synchronized seed crops, and is a common reproductive strategy in plants. Weather has long been recognized as centrally involved in driving seed production in masting plants. However, the theory behind mechanisms connecting weather and seeding variation has only recently been developed, and still lacks empirical evaluation. We used 12-year long seed production data for 255 holm oaks (Quercus ilex), as well as airborne pollen and meteorological data, and tested whether masting is driven by environmental constraints: phenological synchrony and associated pollination efficiency, and drought-related acorn abscission. We found that warm springs resulted in short pollen seasons, and length of the pollen seasons was negatively related to acorn production, supporting the phenological synchrony hypothesis. Furthermore, the relationship between phenological synchrony and acorn production was modulated by spring drought, and effects of environmental vetoes on seed production were dependent on last year's environmental constraint, implying passive resource storage. Both vetoes affected among-tree synchrony in seed production. Finally, precipitation preceding acorn maturation was positively related to seed production, mitigating apparent resource depletion following high crop production in the previous year. These results provide new insights into mechanisms beyond widely reported weather and seed production correlations.
Pleistocene population expansions of shade-tolerant trees indicate fragmentation of the African rainforest during the Ice Ages
Authors: Piñeiro Rosalía , Dauby Gilles , Kaymak Esra , Hardy Olivier J.
Published: November 2017

The fossil record in tropical Africa suggests that dry conditions during the Ice Ages caused expansion of savannahs and contraction of the rainforest. Forest refugia have been proposed to be located in areas of Central Africa that currently harbour high rates of endemic species. However, to what extent the forest was fragmented remains unknown. Nuclear microsatellites and plastid sequences of 732 trees of two species occurring in the same habitat—mature lowland evergreen rainforests—but with remarkably different dispersal capacities—animal versus gravity—were analysed. Geographical information system tools revealed intraspecific lineages partially congruent across the two species, suggesting common past barriers to gene flow in Central Africa. According to approximate Bayesian computation, the intraspecific genetic clusters diverged during the Pleistocene (less than 2 Ma), so that intraspecific differentiation is the appropriate scale to test the aridification effect of the Ice Ages on tree populations. Demographic tests revealed clear genetic signals of population expansion in both taxa, possibly following bottleneck events after forest fragmentation, with stronger evidence of expansion after the Penultimate rather than after the Last Glacial Maximum. The differential dispersal capacity may have modulated the particular response of each species to climate change, as revealed by the stronger evidence of expansion found in the animal-dispersed species than in the gravity-dispersed one.
Specialized mutualisms may constrain the geographical distribution of flowering plants
Authors: Duffy Karl J. , Johnson Steven D.
Published: November 2017

It is commonly assumed that the geographical distributions of plants are governed mainly by abiotic variables. However, interactions with other organisms, such as pollinators, also have the potential to influence plant distributions. To investigate this, we developed niche models for 32 plant taxa that have specialized pollination systems and which are native to a biodiversity hotspot (South Africa). We found that the distributions of these taxa are best explained by a combination of biotic (pollinators) and abiotic factors, rather than by abiotic factors alone. For approximately 66% of these plant taxa, pollinator distributions were the factor that provided the best predictor of their niches. Furthermore, co-occurrence of these plants and their pollinators was generally not explained solely by shared abiotic niches, and thus probably reflects broad-scale positive ecological interactions. These results are consistent with the proposal that pollinator distributions may constrain plant distributions and highlight the general potential for species distributions to be shaped by positive interactions with other species. This has important consequences for predicting how distributions of species might change in the face of loss of their key mutualists.
Potential high-frequency off-target mutagenesis induced by CRISPR/Cas9 in Arabidopsis and its prevention
Authors: Zhang Qiang , Xing Hui-Li , Wang Zhi-Ping , Zhang Hai-Yan , Yang Fang , Wang Xue-Chen , Chen Qi-Jun
Published: November 2017

Specificity of CRISPR/Cas9 tools has been a major concern along with the reports of their successful applications. We report unexpected observations of high frequency off-target mutagenesis induced by CRISPR/Cas9 in T1 Arabidopsis mutants although the sgRNA was predicted to have a high specificity score. We also present evidence that the off-target effects were further exacerbated in the T2 progeny. To prevent the off-target effects, we tested and optimized two strategies in Arabidopsis, including introduction of a mCherry cassette for a simple and reliable isolation of Cas9-free mutants and the use of highly specific mutant SpCas9 variants. Optimization of the mCherry vectors and subsequent validation found that fusion of tRNA with the mutant rather than the original sgRNA scaffold significantly improves editing efficiency. We then examined the editing efficiency of eight high-specificity SpCas9 variants in combination with the improved tRNA-sgRNA fusion strategy. Our results suggest that highly specific SpCas9 variants require a higher level of expression than their wild-type counterpart to maintain high editing efficiency. Additionally, we demonstrate that T-DNA can be inserted into the cleavage sites of CRISPR/Cas9 targets with high frequency. Altogether, our results suggest that in plants, continuous attention should be paid to off-target effects induced by CRISPR/Cas9 in current and subsequent generations, and that the tools optimized in this report will be useful in improving genome editing efficiency and specificity in plants and other organisms.
Morphological changes of plasma membrane and protein assembly during clathrin-mediated endocytosis
Authors: Yoshida Aiko , Sakai Nobuaki , Uekusa Yoshitsugu , Imaoka Yuka , Itagaki Yoshitsuna , Suzuki Yuki , Yoshimura Shige H.
Published: November 2017

Clathrin-mediated endocytosis (CME) proceeds through a series of morphological changes of the plasma membrane induced by a number of protein components. Although the spatiotemporal assembly of these proteins has been elucidated by fluorescence-based techniques, the protein-induced morphological changes of the plasma membrane have not been fully clarified in living cells. Here we visualize membrane morphology together with protein localizations during CME by utilizing high-speed atomic force microscopy combined with a confocal laser scanning unit. The plasma membrane starts to invaginate ~30 seconds after clathrin starts to assemble, and the aperture diameter increases as clathrin accumulates. Actin rapidly accumulates around the pit and induces a small membrane swelling, which within 30 seconds rapidly covers the pit irreversibly. Inhibition of actin turnover abolishes the swelling and induces a reversible open-close motion of the pit, indicating that actin dynamics are necessary for efficient and irreversible pit closure at the end of the CME.
Incipient signs of genetic differentiation among African elephant populations in fragmenting miombo ecosystems in south-western Tanzania
Authors: Lobora Alex L. , Nahonyo Cuthbert L. , Munishi Linus K. , Caro Tim , Foley Charles , Beale Colin M. , Eggert Lori S.
Published: November 2017

Habitat fragmentation plays a major role in the reduction of genetic diversity among wildlife populations. The African savannah elephant population of the Ruaha-Rungwa and Katavi-Rukwa ecosystems in south-western Tanzania, comprises one of the world’s largest remaining elephant populations, but is increasingly threatened by loss of connectivity and poaching for ivory. We investigate whether there are incipient signs of genetic isolation (loss of heterozygosity) within the younger cohort as a result of habitat loss between the two ecosystems. To investigate the genetic structure of populations, we compared the genotypes for 11 microsatellite loci in the western (n = 81 individuals from Katavi-Rukwa), central (n = 36 individuals from Lukwati and Piti), and eastern populations (n = 193, individuals from Ruaha-Rungwa). We found evidence of significant genetic differentiation among the three populations, but the levels were low, suggesting recent divergence. Furthermore, we identified weak isolation by distance, suggesting higher gene flow among nearer individuals with samples within 50km of each other being more genetically similar to one another than beyond. Although sample sizes were small, a further analysis of genetic differences across populations and in separate age classes revealed evidence of increasing genetic structure among younger age classes across the landscape. In a long-lived species with overlapping generations, it takes a long time to develop genetic substructure even when there are substantial obstacles to migration. Thus, in these recently fragmented populations, inbreeding (and the loss of heterozygosity) may be less of an immediate concern than demography (the loss of adults due to illegal hunting).
Trends and variation in Prescribing of Low-Priority Medicines Identified by NHS England: A Cross-Sectional Study and Interactive Data Tool in English Primary Care
Authors: Walker Alex J , Curtis Helen J , Bacon Seb , Croker Richard , Goldacre Ben
Published: November 2017

Background

Routine accessible audit of prescribing data presents significant opportunities to identify cost-saving opportunities. NHS England recently announced a consultation seeking to discourage use of medicines it considers to be low-value. We set out to produce an interactive data resource to show savings in each NHS general practice, and to assess the current use of these medicines, their change in use over time, and the extent and reasons for variation in such prescribing.

Results

The total NHS spend on all low-value medicines identified by NHS England was £153.5m in the last year, across 5.8m prescriptions (mean £26 per prescription). Among individual medications, liothyronine had the highest prescribing cost at £29.6m, followed by trimipramine (£20.2m) and gluten-free foods (£18.7m). Over time, the overall total number of low-value prescriptions decreased, but the cost increased, although this varied greatly between medications. Annual practice level spending varied widely (median, £2,262 per thousand patients, IQR £1,439 to £3,298). The proportion of patients over 65 showed the strongest association with low-value prescribing; CCG was also strongly associated. Our interactive data tool was deployed to OpenPrescribing.net where monthly updated figures and graphs can be viewed.

Conclusions

Prescribing of low-value medications is extensive but varies widely by medication, geographic area and individual practice. Despite a fall in prescription numbers, the overall cost of prescribing for low-value items has risen. Prescribing behaviour is clustered by CCG, which may represent variation in medicines optimisation efficiency, or in some cases access inequality.

Abbreviations

GP

General Practice

NHS

National Health Service

NICE

National Institute for Health and Care Excellence

PCA

Prescription cost Analysis

QOF

Quality Outcomes Framework
Uneven Data Quality and the Earliest Occupation of Europe – The Case of Untermassfeld (Germany)
Authors: Roebroeks Wil , Gaudzinski-Windheuser Sabine , Baales Michael , Kahlke Ralf-Dietrich
Published: November 2017

The database regarding the earliest occupation of Europe has increased significantly in quantity and quality of data points over the last two decades, mainly through the addition of new sites as a result of long-term systematic excavations and large-scale prospections of Early and early Middle Pleistocene exposures. The site distribution pattern suggests an ephemeral presence of hominins in the south of Europe from around one million years ago, with occasional short northward expansions along the western coastal areas when temperate conditions permitted. From around 600,000-700,000 years ago Acheulean artefacts appear in Europe and somewhat later hominin presence seems to pick up, with more sites and now some also present in colder climatic settings. It is again only later, around 350,000 years ago, that the first sites show up in more continental, central parts of Europe, east of the Rhine. A series of recent papers on the Early Pleistocene palaeontological site of Untermassfeld (Germany) makes claims that are of great interest for studies of earliest Europe and are at odds with the described pattern: the papers suggest that Untermassfeld has yielded stone tools and humanly modified faunal remains, evidence for a one million years old hominin presence in European continental mid-latitudes, and additional evidence that hominins were well-established in Europe already around that time period. Here we evaluate these claims and demonstrate that these studies are severely flawed in terms of data on provenance of the materials studied and in the interpretation of faunal remains and lithics as testifying to a hominin presence at the site. In actual fact any reference to the Untermassfeld site as an archaeological one is unwarranted. Furthermore, it is not the only European Early Pleistocene site where inferred evidence for hominin presence is problematic. The strength of the spatiotemporal patterns of hominin presence and absence depend on the quality of the data points we work with, and data base maintenance, including critical evaluation of new sites, is crucial to advance our knowledge of the expansions and contractions of hominin ranges during the Pleistocene.
The amyloidogenicity of the influenza virus PB1-derived peptide sheds light on its antiviral activity
Authors: Zabrodskaya Yana A. , Lebedev Dmitry V. , Egorova Marja A. , Shaldzhyan Aram A. , Shvetsov Alexey V. , Kuklin Alexander I. , Vinogradova Daria S. , Klopov Nikolay V. , Matusevich Oleg V. , Cheremnykh Taisiia A. , Dattani Rajeev , Egorov Vladimir V.
Published: November 2017

The influenza virus polymerase complex is a promising target for new antiviral drug development. It is known that, within the influenza virus polymerase complex, the PB1 subunit region from the 1st to the 25th amino acid residues has to be is in an alpha-helical conformation for proper interaction with the PA subunit. We have previously shown that PB1(6–13) peptide at low concentrations is able to interact with the PB1 subunit N-terminal region in a peptide model which shows aggregate formation and antiviral activity in cell cultures.

In this paper, it was shown that PB1(6–13) peptide is prone to form the amyloid-like fibrillar aggregates. The peptide homo-oligomerization kinetics were examined, and the affinity and characteristic interaction time of PB1(6–13) peptide monomers and the influenza virus polymerase complex PB1 subunit N-terminal region were evaluated by the SPR and TR-SAXS methods. Based on the data obtained, a hypothesis about the PB1(6–13) peptide mechanism of action was proposed: the peptide in its monomeric form is capable of altering the conformation of the PB1 subunit N-terminal region, causing a change from an alpha helix to a beta structure. This conformational change disrupts PB1 and PA subunit interaction and, by that mechanism, the peptide displays antiviral activity.

Graphical abstract
GPseudoRank: a permutation sampler for single cell orderings
Authors: Strauß Magdalena E , Reid John E , Wernisch Lorenz
Published: November 2017

Motivation

A number of pseudotime methods have provided point estimates of the ordering of cells for scRNA-seq data. A still limited number of methods also model the uncertainty of the pseudotime estimate. However, there is still a need for a method to sample from complicated and multi-modal distributions of orders, and to estimate changes in the amount of the uncertainty of the order during the course of a biological development, as this can support the selection of suitable cells for the clustering of genes or for network inference.

Results

In an application to a microarray data set our proposed method, GPseudoRank, identifies two modes of the distribution, each of them corresponding to point estimates of orders obtained by a different established method. In an application to scRNA-seq data we demonstrate the potential of GPseudoRank to identify phases of lower and higher pseudotime uncertainty during a biological process. GPseudoRank also correctly identifies cells precocious in their antiviral response.

Availability and implementation

Our method is available on github: https://github.com/magStra/GPseudoRank.

Contact

magdalena.strauss@mrc-bsu.cam.ac.uk

Supplementary information

Supplementary materials are available.
Mechanical strain can increase segment number in live chick embryos
Authors: Nelemans Ben K. A. , Schmitz Manuel , Tahir Hannan , Merks Roeland M. H. , Smit Theodoor H.
Published: November 2017

Physical cues, experienced during early embryonic development, can influence species-specific vertebral numbers. Here we show that mechanical stretching of live chicken embryos can induce the formation of additional somites and thereby modify early segmental patterning. Stretching deforms the somites, and results in a cellular reorganization that forms stable daughter somites. Cells from the somite core thereby undergo mesenchymal-to-epithelial transitions (MET), thus meeting the geometrical demand for more border cells. Using a Cellular Potts Model, we suggest that this MET occurs through lateral induction by the existing epithelial cells. Our results indicate that self-organizing properties of the somitic mesoderm generate phenotypic plasticity that allows it to cope with variations in the mechanical environment. This plasticity may provide a novel mechanism for explaining how vertebral numbers in species may have increased during evolution. Additionally, by preventing the formation of transitional vertebrae, these self-organization qualities of somites may be selectively advantageous.
Lamins organize the global three-dimensional genome from the nuclear periphery
Authors: Zheng Xiaobin , Hu Jiabiao , Yue Sibiao , Kristiani Lidya , Kim Miri , Sauria Michael , Taylor James , Kim Youngjo , Zheng Yixian
Published: November 2017

Lamins are structural components of the nuclear lamina (NL) that regulate genome organization and gene expression, but the mechanism remains unclear. Using Hi-C, we show that lamins maintain proper interactions among the topologically associated chromatin domains (TADs) but not their overall architecture. Combining Hi-C with fluorescence in situ hybridization (FISH) and analyses of lamina-associated domains (LADs), we reveal that lamin loss causes expansion or detachment of specific LADs in mouse ES cells. The detached LADs disrupt 3D interactions of both LADs and interior chromatin. 4C and epigenome analyses further demonstrate that lamins maintain the active and repressive chromatin domains among different TADs. By combining these studies with transcriptome analyses, we found a significant correlation between transcription changes and the changes of active and inactive chromatin domain interactions. These findings provide a foundation to further study how the nuclear periphery impacts genome organization and transcription in development and NL-associated diseases.

Highlights

Lamin loss does not affect the overall TAD structure but alters TAD-TAD interactions

Lamin null ES cells exhibit decondensation or detachment of specific LAD regions

Expansion and detachment of LADs can alter genome-wide 3D chromatin interactions

Altered chromatin domain interactions are correlated with altered transcription
Protein Moonlighting Revealed by Non-Catalytic Phenotypes of Yeast Enzymes
Authors: Espinosa-Cantú Adriana , Ascencio Diana , Herrera-Basurto Selene , Xu Jiewei , Roguev Assen , Krogan Nevan J. , DeLuna Alexander
Published: November 2017

A single gene can partake in several biological processes, and therefore gene deletions can lead to different—sometimes unexpected—phenotypes. However, it is not always clear whether such pleiotropy reflects the loss of a unique molecular activity involved in different processes or the loss of a multifunctional protein. Here, usingSaccharomyces cerevisiaemetabolism as a model, we systematically test the null hypothesis that enzyme phenotypes depend on a single annotated molecular function, namely their catalysis. We screened a set of carefully selected genes by quantifying the contribution of catalysis to gene-deletion phenotypes under different environmental conditions. While most phenotypes were explained by loss of catalysis, 30% could be readily complemented by a catalytically-inactive enzyme. Such non-catalytic phenotypes were frequent in the Alt1 and Bat2 transaminases and in the isoleucine/valine-biosynthetic enzymes Ilv1 and Ilv2, suggesting novel "moonlighting" activities in these proteins. Furthermore, differential genetic-interaction profiles of gene-deletion and catalytic mutants indicated thatILV1is functionally associated to regulatory processes, specifically to chromatin modification. Our systematic study shows that gene-loss phenotypes and their genetic interactions are frequently not driven by the loss of an annotated catalytic function, underscoring the moonlighting nature of cellular metabolism.
Comparison of the 3D-IR - BTFE method and the conventional method in the head MRI contrast
Authors: Hirahara Daisuke
Published: November 2017

MRI using gadolinium contrast media is useful in diagnosis; however, nephrogenic systemic fibrosis is a serious side effect of gadolinium exposure. Moreover, it turns out that gadolinium deposits in the brain. This has escalated the necessity for a suitable method to use gadolinium contrast media. I developed a new imaging method that had excellent contrast. This study examined the usefulness of that new imaging method and found the method is highly effective.
The role of chromosomal inversions in speciation
Authors: Fuller Z.L. , Leonard C.J. , Young R.E. , Schaeffer S.W. , Phadnis N
Published: November 2017

The chromosomal inversions ofD. persimilisandD. pseudoobscurahave deeply influenced our understanding of the evolutionary forces that shape natural variation, speciation, and selfish chromosome dynamics. Here, we perform a comprehensive reconstruction of the evolutionary histories of the chromosomal inversions in these species. We provide a solution to the puzzling origins of the selfishSex-Ratiochromosome inD. persimilisand show that thisSex-Ratiochromosome directly descends from an ancestrally-arranged chromosome. Our results further show that all fixed inversions betweenD. persimilisandD. pseudoobscurawere segregating in the ancestral population long before speciation, and that the genes contributing to reproductive barriers between these species must have evolved within them afterwards. We propose a new model for the role of chromosomal inversions in speciation and suggest that higher levels of divergence and an association with hybrid incompatibilities are emergent properties of ancestrally segregating inversions. These findings force a reconsideration of the role of chromosomal inversions in speciation, not as protectors of existing hybrid incompatibility alleles, but as fertile grounds for their formation.
Successful aggregation of Tau protein labelled on its native cysteines
Authors: Radli Martina , Verdonschot Romy E , Ferrari Luca , Rüdiger Stefan GD
Published: November 2017

The formation of fibrillar tangles of the Tau protein is crucial in the development of Alzheimer’s disease. Biophysical methods based on labelling of the cysteines of Tau with fluorescence dyes would allow to study fibril formation with an ‘internal eye’. However, the two native cysteines of Tau at positions 291 and 322 are located in the repeat domain, which is involved in forming the fibrils. The contribution of both cysteines to this process is unclear. Here we show that blocking natural cysteines using large fluorescent dyes does not interfere with Tau fibrillation so that FRET can be used to follow structural changes during the process. We anticipate that cysteine-labelled Tau enables following structural rearrangements during fibril formation in detail. This may also allow to monitor the effect of drugs, small molecules and proteins on the process.
Local and global Cdc42 GEFs for fission yeast cell polarity are coordinated by microtubules and the Tea1/Tea4/Pom1 axis
Authors: Tay Ye Dee , Leda Marcin , Goryachev Andrew B. , Sawin Kenneth E.
Published: November 2017

The conserved Rho-family GTPase Cdc42 plays a central role in eukaryotic cell polarity. The rod-shaped fission yeastSchizosaccharomyces pombehas two Cdc42 guanine-nucleotide exchange factors (GEFs), Scd1 and Gef1, but little is known about how they are coordinated in polarized growth. Although the microtubule cytoskeleton is normally not required for polarity maintenance in fission yeast, here we show that whenscdlfunction is compromised, disruption of microtubules or the polarity landmark proteins Tea1, Tea4, or Pom1 leads to isotropic rather than polarized growth. Surprisingly, this isotropic growth is due to spatially inappropriate activity of Gef1, which is a cytosolic protein rather than a membrane-associated protein at cell tips like Scd1. Microtubules and the Tea1/Tea4/Pom1 axis counteract inappropriate Gef1 activity by regulating the localization of the Cdc42 GTPase-activating protein Rga4. Our results thus demonstrate coordination of “local” (Scd1) and “global” (Gef1) Cdc42 GEFs via microtubules and microtubule-dependent polarity landmarks.
Two-photon photoactivated voltage imaging in tissue with an Archaerhodopsin-derived reporter
Authors: Chien Miao-Ping , Brinks Daan , Adam Yoav , Bloxham William , Kheifets Simon , Cohen Adam E.
Published: November 2017

Robust voltage imaging in tissue remains a technical challenge. Existing combinations of genetically encoded voltage indicators (GEVIs) and microscopy techniques cannot simultaneously achieve sufficiently high voltage sensitivity, background rejection, and time resolution for highresolution mapping of sub-cellular voltage dynamics in intact brain tissue. We developed a pooled high-throughput screening approach to identify Archaerhodopsin mutants with unusual photophysical properties. After screening ~105 cells, we identified a novel GEVI, NovArch, whose 1-photon near infrared fluorescence is reversibly enhanced by weak 2-photon excitation. Because the 2-photon excitation acts catalytically rather than stoichiometrically, high fluorescence signals, optical sectioning, and high time resolution are achieved simultaneously, at modest 2-photon laser power. We developed a microscopy system optimized for NovArch imaging in tissue. The combination of protein and optical engineering enhanced signal contrast sufficiently to enable optical mapping of back-propagating action potentials in dendrites in acute mouse brain slice.
C11orf70 mutations causing primary ciliary dyskinesia disrupt a conserved step in the intraflagellar transport-dependent assembly of multiple axonemal dyneins
Authors: Fassad Mahmoud R. , Shoemark Amelia , Borgne Pierrick le , Koll France , Patel Mitali , Dixon Mellisa , Hayward Jane , Richardson Charlotte , Frost Emily , Jenkins Lucy , Cullup Thomas , Chung Eddie MK , Lemullois Michel , Aubusson-Fleury Anne , Hogg Claire , Mitchell David R. , Tassin Anne-Marie , Mitchison Hannah M.
Published: November 2017

Primary ciliary dyskinesia (PCD) is a genetically and phenotypically heterogeneous disorder characterized by destructive respiratory disease and laterality abnormalities due to randomised left-right body asymmetry. PCD is mostly caused by mutations affecting components of the core axoneme structure of motile cilia that are essential for cilia movement. In addition, there is a growing group of PCD genes that encode proteins essential for the assembly of the ciliary dynein motors and the active transport process that delivers them from their cytoplasmic assembly site into the axoneme. We screened a cohort of affected individuals for disease-causing mutations using a targeted next generation sequencing panel and identified 2 unrelated families (3 affected children) with mutations in the uncharacterized C11orf70 gene. The affected children share a consistent PCD phenotype from early life with laterality defects and immotile respiratory cilia displaying combined loss of inner and outer dynein arms (IDA+ODA). Phylogenetic analysis shows C11orf70 is highly conserved, distributed across species similarly to proteins involved in the intraflagellar transport (IFT)-dependant assembly of axonemal dyneins. Paramecium C11orf70 RNAi knockdown led to combined loss of ciliary IDA+ODA with reduced cilia beating and swim velocity. Fluorescently tagged C11orf70 in Paramecium and Chlamydomonas localises mainly in the cytoplasm with a small amount in the ciliary component, its abundance in the axoneme being IFT-dependant. During ciliogenesis, C11orf70 accumulates at the ciliary tips in a similar distribution to the IFT-B protein IFT46. In summary, C11orf70 is essential for IFT-dependant assembly of dynein arms and C11orf70 mutations cause defective cilia motility and PCD.
C24 sphingolipids play a surprising and central role in governing cholesterol and lateral organization of the live cell plasma membrane
Authors: Courtney K. C. , Pezeshkian W , Raghupathy R , Zhang C , Darbyson A , Ipsen J. H. , Ford D. A. , Khandelia H. , Presley J. F. , Zha X
Published: November 2017

Mammalian cell sphingolipids, primarily with C24 and C16 acyl chains, reside in the outer leaflet of the plasma membrane. Curiously, little is known how C24 sphingolipids impact cholesterol and membrane microdomains. Here, we generated giant unilamellar vesicles and live mammalian cells with C24 or C16 sphingomyelin exclusively in the outer leaflet and compared microdomain formation. In giant unilamellar vesicles, we observed that asymmetrically placed C24 sphingomyelin suppresses microdomains. Conversely, C16 sphingomyelin facilitates microdomains. Replacing endogenous sphingolipids with C24 or C16 sphingomyelin in live HeLa cells has a similar impact on microdomains, characterized by FRET between GPI-anchored proteins: C24, but not C16, sphingomyelin suppresses submicron domains in the plasma membrane. Molecular dynamics simulations indicated that, when in the outer leaflet, the acyl chain of C24 sphingomyelin interdigitates into the opposing leaflet, thereby favouring cholesterol in the inner leaflet. We indeed found that cholesterol prefers the inner over the outer leaflet of asymmetric unilamellar vesicles (80/20) when C24 sphingomyelin is in the outer leaflet. However, when C16 sphingomyelin is in the outer leaflet, cholesterol is evenly partitioned between leaflets (50/50). Interestingly, when a mixture of C24/C16 sphingomyelin is in the outer leaflet of unilamellar vesicles, cholesterol still prefers the inner leaflet (80/20). Indeed, in human erythrocyte plasma membrane, where a mixture of C24 and C16 sphingolipids are naturally in the outer leaflet, cholesterol prefers the cytoplasmic leaflet (80/20). Therefore, C24 sphingomyelin uniquely interacts with cholesterol and governs the lateral organization in asymmetric membranes, including the plasma membrane, potentially by generating cholesterol asymmetry.

Statement of Significance

The plasma membrane bilayer of mammalian cells has distinct phospholipids between the outer and inner leaflet, with sphingolipids exclusively in the outer leaflet. A large portion of mammalian sphingolipids have very long acyl chains (C24). Little is known how C24 sphingolipids function in the outer leaflet. Mutations in the ceramide synthase 2 gene is found to decrease C24. This severely perturbs homeostasis in mice and humans. Here, we investigated unilamellar vesicles and mammalian cells with C24 sphingomyelin exclusively in the outer leaflet. We provide evidence that outer leaflet C24 sphingomyelin suppresses microdomains in model membranes and live cells by partitioning cholesterol into the inner leaflet. We propose that C24 sphingolipids are critical to the function of the plasma membrane.
Preparation of Double-Stranded (Replicative Form) Bacteriophage M13 DNA
Authors: Green Michael R. , Sambrook Joseph
Published: November 2017

The double-stranded, closed-circular, replicative form (RF) of M13 DNA is present in high copy numbers in infected cells, and its physical characteristics are essentially identical to those of closed-circular plasmid DNAs. Any of the methods commonly used to purify plasmid DNA can therefore be used to isolate M13 RF DNA. This protocol describes the isolation of M13 RF DNA by alkaline lysis from small volumes (1–2 mL) of infected bacterial cultures. The yield of DNA (1–4 mg, depending on the size of the M13 clone) is more than enough for most purposes in molecular cloning. However, should more DNA be needed, the procedure can easily be scaled up.
Isotype Determination of Rodent-Derived Monoclonal Antibodies Using Sandwich ELISA
Authors: Weis-Garcia Frances , Carnahan Robert H.
Published: November 2017

For research groups needing to isotype a significant number of antibodies on a fairly regular basis, the protocol outlined here provides a cost-effective option. In this simple sandwich ELISA, the monoclonal antibody is first captured with antibodies that discriminate between heavy-chain isotypes and then is detected with antibodies specific for each light chain. This combination ensures that free light chain secreted by hybridomas does not yield a false-positive result, because in a true positive both the capture and detecting antibodies bind the monoclonal antibody.
Hospital variation in cost of childbirth and contributing factors: a cross‐sectional study
Authors: Xu X , Lee HC , Lin H , Lundsberg LS , Pettker CM , Lipkind HS , Illuzzi JL
Published: November 2017

Objective

To examine hospital variation in cost of childbirth hospitalisations and identify factors that contribute to the variation.

Design

Cross‐sectional analysis of linked birth certificate and hospital discharge data.

Setting

Two hundred and twenty hospitals in California delivering ≥ 100 births per year.

Population

A total of 405 908 nulliparous term singleton vertex births during 2010–2012.

Methods

Cost of childbirth hospitalisations was compared across hospitals after accounting for differences in patient clinical risk factors. Relative contributions of patient sociodemographic, obstetric intervention, birth attendant and institutional characteristics to variation in cost were assessed by further adjusting for these factors in hierarchical generalised linear models.

Main outcome measures

Cost of childbirth hospitalisation.

Results

Median risk‐standardised cost of childbirth was $7149 among the hospitals (10th–90th percentile range: $4760–$10,644). Maternal sociodemographic characteristics and type of birth attendant did not explain hospital variation in cost. Adjustment for obstetric interventions overall reduced within‐hospital variance by 15.8% (P < 0.001), while adjusting for caesarean delivery alone reduced within‐hospital variance by 14.4% (P < 0.001). However, obstetric interventions did not explain between‐hospital variation in cost. In contrast, adjustment for institutional characteristics reduced between‐hospital variance by 30.3% (P = 0.002). Hospital type of ownership, teaching/urban–rural status, neonatal care capacity and geographic region were most impactful. Risk‐standardised cost was positively correlated with risk‐standardised rate of severe newborn morbidities (correlation coefficient 0.22, P = 0.001), but not associated with risk‐standardised rate of severe maternal morbidities.

Conclusions

Cost of childbirth hospitalisations varied widely among hospitals in California. Institutional characteristics significantly contributed to this variation. Higher‐cost hospitals did not have better outcomes, suggesting potential opportunities to enhance value in care.

Tweetable abstract

Hospitals vary in cost of childbirth. Institutional characteristics significantly contribute to the variation.
Nonthermal Technologies for Fruit and Vegetable Juices and Beverages: Overview and Advances
Authors: Bevilacqua Antonio , Petruzzi Leonardo , Perricone Marianne , Speranza Barbara , Campaniello Daniela , Sinigaglia Milena , Corbo Maria Rosaria
Published: November 2017

In recent years, there has been a growing interest in the design of novel nonthermal processing systems that minimally modify sensory, nutritional, and functional properties of fruit and vegetable juices and beverages. The benefits of nonthermal treatments are strongly dependent on the food matrix. Thus, an understanding of the effects that these technologies exert on the properties of juices and beverages is important to design and optimize technological parameters to produce value‐added products. This review covers research on nonthermal electrical treatments, high pressure processing, ultrasound, radiation processing, inert gas treatments, cold plasma, and membrane processing. Advances towards optimization of processing conditions, and combined technologies approaches have been also extensively reviewed. This information could be useful to: (1) manage processing systems and optimize resources; (2) preserve nutritional value and organoleptic properties, and (3) provide processing conditions for validation of these technologies at the industrial scale.
Uniting emergency and inpatient clinicians across the ED–inpatient interface: The last frontier?
Authors: Staib Andrew , Sullivan Clair , Prins Johannes B , Burton‐Jones Andrew , Fitzgerald Gerry , Scott Ian
Published: November 2017

Unwell patients in the ED requiring inpatient admission must negotiate the interface between the ED and inpatient wards. Despite its importance and scale, this ED–inpatient interface (EDii) is poorly characterised. The aim of this paper is to clearly define the EDii and to describe its importance to (i) the patient: delays to admission and errors in communication across the EDii can increase adverse outcomes; (ii) the hospital: poor EDii function reduces hospital efficiency and effectiveness; and (iii) the healthcare system: half of all hospital inpatient admissions occur via the EDii and so EDii affects system‐wide performance. The EDii can be defined as the dynamic, transitional phase of patient care in which responsibility for, and delivery of care, is shared between ED and inpatient hospital services. The EDii is characterised by a complex interplay of patient, hospital and system factors. A clear definition of the EDii and an understanding of its importance will assist future research and interventions to improve patient outcomes.
A method for the direct detection of airborne dispersal in lichens
Authors: Eaton Sally , Zúñiga Catalina , Czyzewski Jakub , Ellis Christopher , Genney David R. , Haydon Daniel , Mirzai Nosrat , Yahr Rebecca
Published: November 2017

This study sets out a novel method to determine dispersal distances in lichens. Direct measurement of dispersal often remains difficult for lichens and other small inconspicuous species because of the need to track microscopic reproductive propagules, which even if they can be captured, cannot be identified using traditional morphological approaches. A low‐cost device (<£200) was developed to trap the reproductive propagules of lichens, capable of sampling around 0.1 m3 of air per minute. In parallel, molecular techniques were developed to enable species‐specific detection of propagules caught by the devices, with identification using novel species‐specific primers and optimization of a standard DNA extraction and nested PCR protocol. The methods were tested for both their sensitivity and specificity against a suite of lichen epiphytes, differing in their reproductive mechanisms, dispersal structures and rarity. Sensitivity tests showed that the molecular techniques could detect a single asexual propagule (soredium or isidium), or as few as 10 sexual spores. As proof of concept, propagule traps were deployed into a wooded landscape where the target epiphytes were present. Extractions from deployed propagule traps were sequenced, showing that the method was able to detect the presence of the target species in the atmosphere. As far as we are aware, this is the first attempt to use mechanized propagule traps in combination with DNA diagnostics to detect dispersal of lichens. The tests carried out here point the way for future dispersal studies of lichen epiphytes and other passively dispersed microscopic organisms including fungi or bryophytes.
Validation of loop‐mediated isothermal amplification for fast and portable sex determination across the phylogeny of birds
Authors: Centeno‐Cuadros A. , Tella J. L. , Delibes M. , Edelaar P. , Carrete M.
Published: November 2017

PCR is a universal tool for the multiplication of specific DNA sequences. For example, PCR‐based sex determination is widely used, and a diversity of primer sets is available. However, this protocol requires thermal cycling and electrophoresis, so results are typically obtained in laboratories and several days after sampling. Loop‐mediated isothermal amplification (LAMP) is an alternative to PCR that can take molecular ecology outside the laboratory. Although its application has been successfully probed for sex determination in three species of a single avian Family (raptors, Accipitridae), its generality remains untested and suitable primers across taxa are lacking. We designed and tested the first LAMP‐based primer set for sex determination across the modern birds (NEO‐W) based on a fragment of the gene chromo‐helicase‐DNA‐binding protein located on the female‐specific W chromosome. As nucleotide identity is expected to increase among more related taxa, taxonomically targeted primers were also developed for the Order Falconiformes and Families Psittacidae, Ciconiidae, Estrildidae and Icteridae as examples. NEO‐W successfully determined sex in a subset of 21 species within 17 Families and 10 Orders and is therefore a candidate primer for all modern birds. Primer sets designed specifically for the selected taxa correctly assigned sex to the evaluated species. A short troubleshooting guide for new LAMP users is provided to identify false negatives and optimize LAMP reactions. This study represents the crucial next step towards the use of LAMP for molecular sex determination in birds and other applications in molecular ecology.
Corneal backward scattering and higher‐order aberrations in children with vernal keratoconjunctivitis and normal topography
Authors: Chan Tommy C. Y. , Wong Emily S. , Chan Jason C. K. , Wang Yumeng , Yu Marco , Maeda Naoyuki , Jhanji Vishal
Published: November 2017

Purpose

To investigate the corneal backward scattering and higher‐order aberrations (HOAs) in children with vernal keratoconjunctivitis (VKC) and normal topography.

Methods

Thirty‐six eyes of 22 patients with VKC and 54 eyes of 34 normal subjects were included. All participants had clear cornea, absence of dry eyes and a normal corneal tomography. Scheimpflug imaging was used to measure corneal backward scattering in zones centred on the corneal apex (central 2‐mm zone and paracentral 2‐ to 6‐mm zone), and HOAs were compared between VKC and normal control.

Results

The mean age of participants was 12.0 ± 4.1 years in VKC group and 11.2 ± 4.1 years in control group (p = 0.339). There was no significant intergroup difference in mean keratometry, astigmatism and apex pachymetry (p ≥ 0.076). Total corneal backscatter was higher in the VKC group compared to the control group (p ≤ 0.012). Anterior and posterior cornea displayed a higher level of backward scattering in the VKC group (p < 0.001 for anterior; p ≤ 0.048 for posterior). Patients with VKC exhibited higher total HOAs and coma (p ≤ 0.036). There were significant correlations between total anterior HOAs and backward scattering measured at the central (r = 0.500; p = 0.032) and paracentral zones (r = 0.470; p = 0.024) for VKC.

Conclusion

The current study showed optical quality changes in patients with clear corneas and quiescent VKC. An increase in corneal backward scattering and HOAs was noted in patients with VKC as compared to normal patients.
The RELATION study: efficacy and safety of ranibizumab combined with laser photocoagulation treatment versus laser monotherapy in NPDR and PDR patients with diabetic macular oedema
Authors: Lang Gabriele E. , Liakopoulos Sandra , Vögeler Jessica , Weiß Claudia , Spital Georg , Gamulescu Maria‐Andreea , Lohmann Chris , Wiedemann Peter
Published: November 2017

Purpose

To assess efficacy and safety of intravitreal ranibizumab 0.5 mg plus laser (COMBI) versus laser monotherapy (LASER) in patients with visual impairment due to diabetic macular oedema (DME) in either nonproliferative diabetic retinopathy (NPDR) or proliferative diabetic retinopathy (PDR) and to analyse the relevance of inner versus outer retinal thickness.

Methods

In this double‐masked, multicentre phase IIIb study, patients (N = 128) were randomized (2:1) to receive COMBI (n = 85) versus LASER (n = 43). Patients received four initial monthly injections of ranibizumab 0.5 mg (COMBI) or sham (LASER) followed by pro re nata (PRN) injections. In both groups, patients received laser at baseline and additional laser at 3 monthly intervals, as needed. The study was started in 2010 and was prematurely terminated due to approval of ranibizumab for DME.

Results

The least squares (LS) mean change in mean best‐corrected visual acuity (BCVA) from baseline to month 12 was higher in the COMBI (6.5) versus LASER (2.3) group (LS mean difference: 4.2 [95% CI 0.9; 7.4] letters, p = 0.01, primary end‐point). There was also a tendency in the same direction for the subgroup of 26 patients with PDR (LS mean difference 14.7, p = 0.11). Mean central retinal thickness decreased by 107.3 μm in the COMBI group and by 80.3 μm in the LASER group from baseline to month 12 (p = 0.28). Ranibizumab was well tolerated.

Conclusion

This study showed that ranibizumab plus laser is a valuable treatment option for the management of DME. Patients with DME in PDR might also benefit from combined therapy compared to laser alone.