High quality phased assembly of grape genome offer new opportunities in chimera detection

In perennial plants and especially those propagated through cuttings, several genotypes can coexist in a single individual, thus leading to chimeras. When the variant induces a noticeable phenotype modification, it can lead to a new cultivar. Viticulture already took economic advantage of this natural phenomenon: for instance, the berry skin of ‘Pinot gris’ derived from ‘Pinot noir’ by the selection of a chimera. Chimeras could also impact other crucial traits without being visually identified. Periclinal chimera where the variant has entirely colonized a cell layer is the most stable and can be propagated through cuttings. In grapevine, two functional cell layers are present in leaves, L1 and L2. However, lateral roots are formed from the L2 cell layer only. Thus, comparing DNA sequences of roots and leaves could allow chimera detection. In this study we used new generation Hifi long reads sequencing and recent bioinformatics tools applied to ‘Merlot’ to detect periclinal chimeras. Sequencing of ‘Magdeleine Noire des Charentes’ and ‘Cabernet franc’, the parents of ‘Merlot’, allowed haplotype resolved assembly. Pseudomolecules were built with few contigs, in some occasions only one per chromosome. This high resolution allowed haplotype comparison. Annotation from PN40024 was transferred to all pseudomolecules. Through variant detection, periclinal chimeras were found on both haplotypes. These results open new perspectives on chimera detection, which is an important resource to improve cultivars through clonal selection or breed new ones. Detailed results will be presented and discussed

Insights into the function of ion channels by computational electrophysiology simulations

Ion channels are of universal importance for all cell types and play key roles in cellular physiology and pathology. Increased insight into their functional mechanisms is crucial to enable drug design on this important class of membrane proteins, and to enhance our understanding of some of the fundamental features of cells. This review presents the concepts behind the recently developed simulation protocol Computational Electrophysiology (CompEL), which facilitates the atomistic simulation of ion channels in action. In addition, the review provides guidelines for its application in conjunction with the molecular dynamics software package GROMACS. We first lay out the rationale for designing CompEL as a method that models the driving force for ion permeation through channels the way it is established in cells, i.e., by electrochemical ion gradients across the membrane. This is followed by an outline of its implementation and a description of key settings and parameters helpful to users wishing to set up and conduct such simulations. In recent years, key mechanistic and biophysical insights have been obtained by employing the CompEL protocol to address a wide range of questions on ion channels and permeation. We summarize these recent findings on membrane proteins, which span a spectrum from highly ion-selective, narrow channels to wide diffusion pores. Finally we discuss the future potential of CompEL in light of its limitations and strengths. This article is part of a Special Issue entitled: Membrane Proteins edited by J.C. Gumbart and Sergei Noskov

Llama-Derived Single Domain Antibodies to Build Multivalent, Superpotent and Broadened Neutralizing Anti-Viral Molecules

For efficient prevention of viral infections and cross protection, simultaneous targeting of multiple viral epitopes is a powerful strategy. Llama heavy chain antibody fragments (VHH) against the trimeric envelope proteins of Respiratory Syncytial Virus (Fusion protein), Rabies virus (Glycoprotein) and H5N1 Influenza (Hemagglutinin 5) were selected from llama derived immune libraries by phage display. Neutralizing VHH recognizing different epitopes in the receptor binding sites on the spikes with affinities in the low nanomolar range were identified for all the three viruses by viral neutralization assays. By fusion of VHH with variable linker lengths, multimeric constructs were made that improved neutralization potencies up to 4,000-fold for RSV, 1,500-fold for Rabies virus and 75-fold for Influenza H5N1. The potencies of the VHH constructs were similar or better than best performing monoclonal antibodies. The cross protection capacity against different viral strains was also improved for all three viruses, both by multivalent (two or three identical VHH) and biparatopic (two different VHH) constructs. By combining a VHH neutralizing RSV subtype A, but not subtype B with a poorly neutralizing VHH with high affinity for subtype B, a biparatopic construct was made with low nanomolar neutralizing potency against both subtypes. Trivalent anti-H5N1 VHH neutralized both Influenza H5N1 clade1 and 2 in a pseudotype assay and was very potent in neutralizing the NIBRG-14 Influenza H5N1 strain with IC50 of 9 picomolar. Bivalent and biparatopic constructs against Rabies virus cross neutralized both 10 different Genotype 1 strains and Genotype 5. The results show that multimerization of VHH fragments targeting multiple epitopes on a viral trimeric spike protein is a powerful tool for anti-viral therapy to achieve "best-in-class" and broader neutralization capacity

Minimal information for studies of extracellular vesicles (MISEV2023): From basic to advanced approaches

Extracellular vesicles (EVs), through their complex cargo, can reflect the state of their cell of origin and change the functions and phenotypes of other cells. These features indicate strong biomarker and therapeutic potential and have generated broad interest, as evidenced by the steady year-on-year increase in the numbers of scientific publications about EVs. Important advances have been made in EV metrology and in understanding and applying EV biology. However, hurdles remain to realising the potential of EVs in domains ranging from basic biology to clinical applications due to challenges in EV nomenclature, separation from non-vesicular extracellular particles, characterisation and functional studies. To address the challenges and opportunities in this rapidly evolving field, the International Society for Extracellular Vesicles (ISEV) updates its 'Minimal Information for Studies of Extracellular Vesicles', which was first published in 2014 and then in 2018 as MISEV2014 and MISEV2018, respectively. The goal of the current document, MISEV2023, is to provide researchers with an updated snapshot of available approaches and their advantages and limitations for production, separation and characterisation of EVs from multiple sources, including cell culture, body fluids and solid tissues. In addition to presenting the latest state of the art in basic principles of EV research, this document also covers advanced techniques and approaches that are currently expanding the boundaries of the field. MISEV2023 also includes new sections on EV release and uptake and a brief discussion of in vivo approaches to study EVs. Compiling feedback from ISEV expert task forces and more than 1000 researchers, this document conveys the current state of EV research to facilitate robust scientific discoveries and move the field forward even more rapidly

Reducing the environmental impact of surgery on a global scale: systematic review and co-prioritization with healthcare workers in 132 countries

Abstract Background Healthcare cannot achieve net-zero carbon without addressing operating theatres. The aim of this study was to prioritize feasible interventions to reduce the environmental impact of operating theatres. Methods This study adopted a four-phase Delphi consensus co-prioritization methodology. In phase 1, a systematic review of published interventions and global consultation of perioperative healthcare professionals were used to longlist interventions. In phase 2, iterative thematic analysis consolidated comparable interventions into a shortlist. In phase 3, the shortlist was co-prioritized based on patient and clinician views on acceptability, feasibility, and safety. In phase 4, ranked lists of interventions were presented by their relevance to high-income countries and low–middle-income countries. Results In phase 1, 43 interventions were identified, which had low uptake in practice according to 3042 professionals globally. In phase 2, a shortlist of 15 intervention domains was generated. In phase 3, interventions were deemed acceptable for more than 90 per cent of patients except for reducing general anaesthesia (84 per cent) and re-sterilization of ‘single-use’ consumables (86 per cent). In phase 4, the top three shortlisted interventions for high-income countries were: introducing recycling; reducing use of anaesthetic gases; and appropriate clinical waste processing. In phase 4, the top three shortlisted interventions for low–middle-income countries were: introducing reusable surgical devices; reducing use of consumables; and reducing the use of general anaesthesia. Conclusion This is a step toward environmentally sustainable operating environments with actionable interventions applicable to both high– and low–middle–income countries

High quality phased assembly of grape genome offer new opportunities in chimera detection

In perennial plants and especially those propagated through cuttings, several genotypes can coexist in a single individual, thus leading to chimeras. When the variant induces a noticeable phenotype modification, it can lead to a new cultivar. Viticulture already took economic advantage of this natural phenomenon: for instance, the berry skin of ‘Pinot gris’ derived from ‘Pinot noir’ by the selection of a chimera. Chimeras could also impact other crucial traits without being visually identified. Periclinal chimera where the variant has entirely colonized a cell layer is the most stable and can be propagated through cuttings. In grapevine, two functional cell layers are present in leaves, L1 and L2. However, lateral roots are formed from the L2 cell layer only. Thus, comparing DNA sequences of roots and leaves could allow chimera detection. In this study we used new generation Hifi long reads sequencing and recent bioinformatics tools applied to ‘Merlot’ to detect periclinal chimeras. Sequencing of ‘Magdeleine Noire des Charentes’ and ‘Cabernet franc’, the parents of ‘Merlot’, allowed haplotype resolved assembly. Pseudomolecules were built with few contigs, in some occasions only one per chromosome. This high resolution allowed haplotype comparison. Annotation from PN40024 was transferred to all pseudomolecules. Through variant detection, periclinal chimeras were found on both haplotypes. These results open new perspectives on chimera detection, which is an important resource to improve cultivars through clonal selection or breed new ones. Detailed results will be presented and discussed

Chimeras in Merlot grapevine revealed by phased assembly

Chimerism is the phenomenon when several genotypes coexist in a single individual. Used to understand plant ontogenesis they also have been valorised through new cultivar breeding. Viticulture has been taking economic advantage out of chimeras when the variant induced an important modification of wine type such as berry skin colour. Crucial agronomic characters may also be impacted by chimeras that aren’t identified yet. Periclinal chimera where the variant has entirely colonised a cell layer is the most stable and can be propagated through cuttings. In grapevine, leaves are derived from both meristem layers, L1 and L2. However, lateral roots are formed from the L2 cell layer only. Thus, comparing DNA sequences of roots and leaves allows chimera detection. In this study we used new generation Hifi long reads sequencing, recent bioinformatics tools and trio-binning with parental sequences to detect periclinal chimeras on ‘Merlot’ grapevine cultivar. Sequencing of cv. ‘Magdeleine Noire des Charentes’ and ‘Cabernet Franc’, the parents of cv. ‘Merlot’, allowed haplotype resolved assembly. Pseudomolecules were built with a total of 33 to 47 contigs and in few occasions a unique contig for one chromosome. This high resolution allowed haplotype comparison. Annotation was transferred from PN40024 VCost.v3 to all pseudomolecules. After strong selection of variants, 51 and 53 ‘Merlot’ specific periclinal chimeras were found on the Merlot-haplotype-CF and Merlot-haplotype-MG respectively, 9 and 7 been located in a coding region. A subset of positions was analysed using Molecular Inversion Probes (MIPseq) and 69% were unambiguously validated, 25% are doubtful because of technological noise or weak depth and 6% invalidated. These results open new perspectives on chimera detection as an important resource to improve cultivars through clonal selection or breeding

Socioeconomic position and outdoor nitrogen dioxide (NO2) exposure in Western Europe: A multi-city analysis

Background: Inconsistent associations between socioeconomic position (SEP) and outdoor air pollution have been reported in Europe, but methodological differences prevent any direct between-study comparison. Objectives: Assess and compare the association between SEP and outdoor nitrogen dioxide (NO2) exposure as a marker of traffic exhaust, in 16 cities from eight Western European countries. Methods: Three SEP indicators, two defined at individual-level (education and occupation) and one at neighborhood-level (unemployment rate) were assessed in three European multicenter cohorts. NO2 annual concentration exposure was estimated at participants' addresses with land use regression models developed within the European Study of Cohorts for Air Pollution Effects (ESCAPE; http://www.escapeproject.eu/). Pooled and city-specific linear regressions were used to analyze associations between each SEP indicator and NO2. Heterogeneity across cities was assessed using the Higgins' I-squared test (I2). Results: The study population included 5692 participants. Pooled analysis showed that participants with lower individual-SEP were less exposed to NO2. Conversely, participants living in neighborhoods with higher unemployment rate were more exposed. City-specific results exhibited strong heterogeneity (I2 > 76% for the three SEP indicators) resulting in variation of the individual- and neighborhood-SEP patterns of NO2 exposure across cities. The coefficients from a model that included both individual- and neighborhood-SEP indicators were similar to the unadjusted coefficients, suggesting independent associations. Conclusions: Our study showed for the first time using homogenized measures of outcome and exposure across 16 cities the important heterogeneity regarding the association between SEP and NO2 in Western Europe. Importantly, our results showed that individual- and neighborhood-SEP indicators capture different aspects of the association between SEP and exposure to air pollution, stressing the importance of considering both in air pollution health effects studies.The ESCAPE study, funded by the European Community's Seventh Framework Program (FP7/2007-2011) under grant agreement no. 211250 (http://www.escapeproject.eu/)