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A roadmap of priority evidence gaps for the co-implementation of malaria vaccines and perennial malaria chemoprevention
Progress in malaria control will rely on deployment and effective targeting of combinations of interventions, including malaria vaccines and perennial malaria chemoprevention (PMC). Several countries with PMC programmes have introduced malaria vaccination into their essential programmes on immunizations, but empirical evidence on the impact of combining these two interventions and how best to co-implement them are lacking. At the American Society of Tropical Medicine and Hygiene 2023 annual meeting, a stakeholder meeting was convened to identify key policy, operational and research gaps for co-implementation of malaria vaccines and PMC. Participants from 11 endemic countries, including representatives from national malaria and immunization programmes, the World Health Organization, researchers, implementing organizations and funders attended. Identified evidence gaps were prioritized to select urgent issues to inform co-implementation. The output of these activities is a strategic roadmap of priority malaria vaccine and PMC co-implementation evidence gaps, and solutions to address them. The roadmap was presented to stakeholders for feedback at the 2024 Multilateral Initiative on Malaria meeting and revised accordingly. The roadmap outlines four key areas of work to address urgent evidence gaps for co-implementation: (1) support to the global and national policy process, (2) implementation support and research, (3) clinical studies, and (4) modelling. Together, these areas will provide practical guidance on the co-implementation of the interventions, and robust evidence to inform decision-making on how best to design, optimize and scale-up co-implementation in different contexts, including if and in what contexts the co-implementation is cost-effective, and the optimal schedule for co-implementation. This will work towards supporting the policy process on co-implementation of malaria vaccines and PMC, and achieving the most impactful use of available resources for the prevention of malaria in children.
Mechanistic Pathways Underlying Genetic Predisposition to Atrial Fibrillation Are Associated With Different Cardiac Phenotypes and Cardioembolic Stroke Risk.
BACKGROUND: Genome-wide association studies have clustered candidate genes associated with atrial fibrillation (AF) into biological pathways reflecting different pathophysiological mechanisms. We investigated whether these pathways associate with distinct intermediate phenotypes and confer differing risks of cardioembolic stroke. METHODS: Three distinct subsets of AF-associated genetic variants, each representing a different mechanistic pathway, that is, the cardiac muscle function and integrity pathway (15 variants), the cardiac developmental pathway (25 variants), and the cardiac ion channels pathway (12 variants), were identified from previous AF genome-wide association studies. Using genetic epidemiological methods and large-scale datasets such as UK Biobank, deCODE, and GIGASTROKE, we investigated the associations of these pathways with AF-related cardiac intermediate phenotypes, which included electrocardiogram parameters (≈16 500 electrocardiograms), left atrial and ventricular size and function (≈36 000 cardiac magnetic resonance imaging scans), and relevant plasma biomarkers (N-terminal pro-B-type natriuretic peptide, ≈70 000 samples; high-sensitivity troponin I and T, ≈87 000 samples), as well as with subtypes of ischemic stroke (≈11 000 cases). RESULTS: Genetic variants representing distinct AF-related mechanistic pathways had significantly different effects on several AF-related phenotypes. In particular, the muscle pathway was associated with a longer PR interval (P for heterogeneity between pathways [Phet]=1×10-10), lower left atrial emptying fraction (Phet=5×10-5), and higher N-terminal pro-B-type natriuretic peptide (Phet=2×10-3) per log-odds higher risk of AF compared with the developmental and ion-channel pathways. In contrast, the ion-channel pathway was associated with a lower risk of cardioembolic stroke (Phet=0.04 in European, and 7×10-3 in multiancestry populations) compared with the other pathways. CONCLUSIONS: Genetic variants representing specific mechanistic pathways for AF are associated with distinct intermediate cardiac phenotypes and a different risk of cardioembolic stroke. These findings provide a better understanding of the etiological heterogeneity underlying the development of AF and its downstream impact on disease and may offer a route to more targeted treatment strategies.
UK Public Focus Groups on Healthcare's Environmental Impacts: A Critical Analysis of Co-Benefits Approaches.
The urgency of addressing climate change has accelerated the need for healthcare to mitigate its associated environmental harms. Co-benefits approaches are being used in policymaking to frame mitigation actions because they promise to deliver better health outcomes alongside environment benefits. Despite this, little empirical data exists on public perceptions about the acceptability and usefulness of this approach. We conducted 12 focus groups with 82 members of the UK public asking the question: what were participants' values, beliefs and experiences about the environmental harms associated with healthcare and how should these issues be conceptualised and addressed? Co-benefits framings resonated with participants, who perceived this approach as useful for prioritising healthcare needs while valuing the environment. However, when participants tried to frame co-benefits as a solution, they struggled to reconcile complexities. Furthermore, their discussions revealed a certain subjectivity and context-specificity in co-benefits framing, drawn from their own experiences and expectations of care. We emphasise paying attention to such subjectivities when developing co-benefits policies. This could be achieved by the inclusion of public and patient voices in policymaking. Any underlying assumptions associated with co-benefits policies-including which subjectivities are used in the framing and how tensions are resolved-must be made transparent.
Structure and stabilization of the antigenic glycoprotein building blocks of the New World mammarenavirus spike complex.
The spillover of New World (NW) arenaviruses from rodent reservoirs into human populations poses a continued risk to human health. NW arenaviruses present a glycoprotein (GP) complex on the envelope surface of the virion, which orchestrates host cell entry and is a key target of the immune response arising from infection and immunization. Each protomer of the trimeric GP is composed of a stable signal peptide, a GP1 attachment glycoprotein, and a GP2 fusion glycoprotein. To glean insights into the architecture of this key therapeutic target, we determined the crystal structures of NW GP1-GP2 heterodimeric complexes from Junín virus and Machupo virus. Due to the metastability of the interaction between GP1 and GP2, structural elucidation required the introduction of a disulfide bond at the GP1-GP2 complex interface, but no other stabilizing modifications were required. While the overall assembly of NW GP1-GP2 is conserved with that presented by Old World (OW) arenaviruses, including Lassa virus and lymphocytic choriomeningitis virus, NW GP1-GP2 complexes are structurally distinct. Indeed, we note that when compared to the OW GP1-GP2 complex, the globular portion of NW GP1 undergoes limited structural alterations upon detachment from its cognate GP2. We further demonstrate that our engineered GP1-GP2 heterodimers are antigenically relevant and recognized by neutralizing antibodies. These data provide insights into the distinct assemblies presented by NW and OW arenaviruses, as well as provide molecular-level blueprints that may guide vaccine development.IMPORTANCEAlthough the emergence of New World (NW) hemorrhagic fever mammarenaviruses poses an unceasing threat to human health, there is a paucity of reagents capable of protecting against the transmission of these pathogens from their natural rodent reservoirs. This is, in part, attributed to our limited understanding of the structure and function of the NW glycoprotein spike complex presented on the NW arenavirus surface. Here, we provide a detailed molecular-level description of how the two major components of this key therapeutic target assemble to form a key building block of the NW arenaviral spike complex. The insights gleaned from this work provide a framework for guiding the structure-based development of NW arenaviral vaccines.
Identification of cross-stage, cross-species malaria CD8+ T cell antigens.
Malaria is one of the most prevalent parasitic diseases in the world. In 2023, 263 million malaria cases were estimated worldwide. Two species of Plasmodium, P. falciparum and P. vivax, cause most human malaria. Despite the licensing of two partially protective vaccines for P. falciparum, there is no vaccine capable of providing long-term control or elimination. A major limitation for vaccine development is the lack of validated T cell epitopes for either species that could be targeted by vaccines. P. vivax is the most widespread human malaria parasite and is the major species causing malaria in the Americas and Asia while P. falciparum is more prevalent in Africa1. P. vivax exclusively infects reticulocytes in peripheral blood, which, unlike the mature erythrocytes infected by P. falciparum, still retain RNA and therefore retain host protein translation capabilities. We previously reported that P. vivax-infected reticulocytes express the major human leukocyte antigen class I (HLA-I), which allows parasite sensing by CD8+ T cells and consequent killing of parasite-infected host cells and intracellular parasites. Here we report by immunopeptidomic analysis the first unbiased identification of Plasmodium spp. antigens presented via HLA-I on infected reticulocytes. We identified 453 unique peptides that mapped to 166 different proteins. Most of these antigens were housekeeping proteins that are constitutively expressed at multiple stages of the parasite life cycle. Common peptides were presented in different individuals by the same or distinct HLA-ABC alleles as well as by non-classical HLA-E. Many peptide sequences were highly conserved in P. falciparum and P. vivax. The immunogenicity of the newly identified epitopes was validated in both P. vivax- and P. falciparum-infected patient samples. Furthermore, several of these antigens were immunogenic in the blood and liver of non-human primates following Plasmodium infection and attenuated parasite immunization. Two antigens were also the target of protective CD8+ T cell-mediated immunity in rodents. Thus, these antigens have potential for use in a cross-stage and cross-species malaria vaccine.
Virion Structure
Picornaviruses were the first animal viruses whose structure was determined in atomic detail and, as of October 2009, the Protein Data Bank (PDB) registered 53 structure depositions for picornaviruses. These data have contributed significantly to the understanding of picornavirus evolution, assembly, host-cell interaction, host adaptation, and antigenic variation and are providing the basis for novel therapeutic strategies. Subsequently classified as a picornavirus, the general morphology of FMDV could not be visualized until the advent of the electron microscope, when negative-stained images to a resolution of 4 to 5 nm revealed rather smooth round particles of ˜30 nm diameter. The current classification of picornaviruses is based on genome and protein sequence properties which are derived from the interplay of the error-prone replication mechanism of the virus with the process of natural selection. Differences in physical properties, such as buoyant density in cesium chloride and pH stability, underpinned the early classification of picornaviruses. Virus capsids recognize susceptible cells by attachment to specific receptors on the host cell membrane, thereby determining the host range and tropism of infection. The majority of antibodies are weak neutralizers that appear to operate by using the two arms of the antibody to cross-link different virus particles, causing aggregation.
Continuous Indexing of Fibrosis (CIF): improving the assessment and classification of MPN patients.
The grading of fibrosis in myeloproliferative neoplasms (MPN) is an important component of disease classification, prognostication and monitoring. However, current fibrosis grading systems are only semi-quantitative and fail to fully capture sample heterogeneity. To improve the quantitation of reticulin fibrosis, we developed a machine learning approach using bone marrow trephine (BMT) samples (n = 107) from patients diagnosed with MPN or a reactive marrow. The resulting Continuous Indexing of Fibrosis (CIF) enhances the detection and monitoring of fibrosis within BMTs, and aids MPN subtyping. When combined with megakaryocyte feature analysis, CIF discriminates between the frequently challenging differential diagnosis of essential thrombocythemia (ET) and pre-fibrotic myelofibrosis with high predictive accuracy [area under the curve = 0.94]. CIF also shows promise in the identification of MPN patients at risk of disease progression; analysis of samples from 35 patients diagnosed with ET and enrolled in the Primary Thrombocythemia-1 trial identified features predictive of post-ET myelofibrosis (area under the curve = 0.77). In addition to these clinical applications, automated analysis of fibrosis has clear potential to further refine disease classification boundaries and inform future studies of the micro-environmental factors driving disease initiation and progression in MPN and other stem cell disorders.
Exome-wide evidence of compound heterozygous effects across common phenotypes in the UK Biobank.
The phenotypic impact of compound heterozygous (CH) variation has not been investigated at the population scale. We phased rare variants (MAF ∼0.001%) in the UK Biobank (UKBB) exome-sequencing data to characterize recessive effects in 175,587 individuals across 311 common diseases. A total of 6.5% of individuals carry putatively damaging CH variants, 90% of which are only identifiable upon phasing rare variants (MAF -7) after accounting for relatedness, polygenicity, nearby common variants, and rare variant burden. Of these, just one is discovered when considering homozygosity alone. Using longitudinal health records, we additionally identify and replicate a novel association between bi-allelic variation in ATP2C2 and an earlier age at onset of chronic obstructive pulmonary disease (COPD) (p -8). Genetic phase contributes to disease risk for gene-trait pairs: ATP2C2-COPD (p = 0.000238), FLG-asthma (p = 0.00205), and USH2A-visual impairment (p = 0.0084). We demonstrate the power of phasing large-scale genetic cohorts to discover phenome-wide consequences of compound heterozygosity.
Docking for Smoothened antagonist chemotypes not susceptible to a vismodegib-resistance mutation.
The G protein-coupled receptor Smoothened (SMO) plays a pivotal role in embryonic development transducing the Hedgehog morphogen signal into the cell. Aberrant activation of the pathway is associated with various cancer types. Antagonizing SMO has been recognized as a therapeutic strategy exemplified by drugs such as vismodegib and sonidegib, but despite initial remission, cancer recurrence is frequent due to resistance mutations. Utilizing a structure-based design approach, we have identified three unprecedented chemotypes to antagonize SMO with potencies in the low micromolar range. In total, 67 compounds identified through molecular docking were assayed in four rounds with hit rates of 27% and 63% during hit identification, i.e. the first two rounds. Importantly, the potency of ligands with two of the chemotypes identified in this work is not strongly affected by the vismodegib resistance mutation D473G. The mutation affects potency and maximal inhibitory effect of these ligands only in a way similar to SANT-1, a SMO ligand unencumbered by the mutation. Our study thus shows a successful application of structure-based design for the discovery of novel SMO antagonist chemotypes.
Role of complete blood count in the diagnosis of culture-proven neonatal sepsis: a systematic review and meta-analysis.
OBJECTIVE: Neonatal sepsis is a significant cause of morbidity and mortality, particularly in preterm infants. Despite its routine use in adults, the diagnostic utility of complete blood count (CBC) in neonatal sepsis remains debated. This systematic review and meta-analysis aimed to evaluate the diagnostic accuracy of CBC parameters for neonatal sepsis. METHODS: This review was registered at PROSPERO (CRD42023476510). MEDLINE, Embase, CINAHL and the Cochrane Library were searched from database inception to 28 October 2024. Observational studies of neonates with sepsis, published in English, were included. Pooled diagnostic accuracy metrics were calculated for CBC parameters, including the white cell count (WCC), neutrophil count and immature-to-total neutrophil ratio (ITR). Bias was assessed using a modified QUADAS-2 tool. RESULTS: Functional CBC parameters like ITR and mean neutrophil volume (MNV) showed moderate diagnostic accuracy. Pooled analysis revealed that an ITR >0.20 had 66.3% sensitivity and 85.4% specificity for neonatal sepsis. MNV also showed promising diagnostic utility, but substantial heterogeneity across studies (I2>0.80) limited its generalisability. Traditional parameters like the WCC and platelet count had lower diagnostic accuracy. CONCLUSIONS: The CBC is a rapid, cost-effective test requiring minimal blood volume, making it a practical adjunct in neonatal diagnostics. Functional parameters like ITR and MNV show the potential to complement existing approaches but are insufficient as stand-alone diagnostic tools. Further research is needed to validate their clinical utility and address heterogeneity in study designs.
Holding space for spatial biology
Spatial biology is revolutionising our understanding of cellular organisation and disease by preserving the interactions between cells within tissues. Traditional methods often disrupted these delicate structures, making it challenging to study cells in their natural environments. This article explores key technologies, such as immunohistochemistry (IHC) and in situ hybridisation, which enable precise molecular analysis while maintaining spatial context. IHC remains essential for identifying protein markers in pathology, while multiplex imaging systems significantly enhance biomarker detection and high-throughput spatial profiling. Additionally, new computational tools like MuSpAn are advancing our understanding of spatial cell organisation. Despite challenges posed by data complexity, spatial biology opens exciting new possibilities for precision medicine, facilitating targeted therapies and advancing personalised treatment strategies. As the field rapidly evolves, it continues to drive groundbreaking breakthroughs in disease research and therapeutic development.
Epithelial GREMLIN1 disrupts intestinal epithelial-mesenchymal crosstalk to induce a wnt-dependent ectopic stem cell niche through stromal remodelling
Abstract In homeostasis, counterbalanced morphogen signalling gradients along the vertical axis of the intestinal mucosa regulate the fate and function of epithelial and stromal cell compartments. Here, we use a disease-positioned mouse and human tissue to explore the consequences of pathological BMP signalling dysregulation on epithelial-mesenchymal interaction. Aberrant pan-epithelial expression of the secreted BMP antagonist Grem1 results in ectopic crypt formation, with lineage tracing demonstrating the presence of Lgr5(−) stem/progenitor cells. Isolated epithelial cell Grem1 expression has no effect on individual cell fate, indicating an intercompartmental impact of mucosal-wide BMP antagonism. Treatment with an anti-Grem1 antibody abrogates the polyposis phenotype, and triangulation of specific pathway inhibitors defines a pathological sequence of events, with Wnt-ligand-dependent ectopic stem cell niches forming through stromal remodelling following BMP disruption. These data support an emerging co-evolutionary model of intestinal cell compartmentalisation based on bidirectional regulation of epithelial-mesenchymal cell fate and function.
SARS-CoV-2 infection enhancement by amphotericin B: implications for disease management.
Severe coronavirus disease 2019 (COVID-19) patients who require hospitalization are at high risk of invasive pulmonary mucormycosis. Amphotericin B (AmB), which is the first-line therapy for invasive pulmonary mucormycosis, has been shown to promote or inhibit replication of a spectrum of viruses. In this study, we first predicted that AmB and nystatin had strong interactions with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) proteins using in silico screening, indicative of drugs with potential therapeutic activity against this virus. Subsequently, we investigated the impact of AmB, nystatin, natamycin, fluconazole, and caspofungin on SARS-CoV-2 infection and replication in vitro. Results showed that AmB and nystatin actually increased SARS-CoV-2 replication in Vero E6, Calu-3, and Huh7 cells. At optimal concentrations, AmB and nystatin increase SARS-CoV-2 replication by up to 100- and 10-fold in Vero E6 and Calu-3 cells, respectively. The other antifungals tested had no impact on SARS-CoV-2 infection in vitro. Drug kinetic studies indicate that AmB enhances SARS-CoV-2 infection by promoting viral entry into cells. Additionally, knockdown of genes encoding for interferon-induced transmembrane (IFITM) proteins 1, 2, and 3 suggests AmB enhances SARS-CoV-2 cell entry by overcoming the antiviral effect of the IFITM3 protein. This study further elucidates the role of IFITM3 in viral entry and highlights the potential dangers of treating COVID-19 patients, with invasive pulmonary mucormycosis, using AmB.IMPORTANCEAmB and nystatin are common treatments for fungal infections but were predicted to strongly interact with SARS-CoV-2 proteins, indicating their potential modulation or inhibition against the virus. However, our tests revealed that these antifungals, in fact, enhance SARS-CoV-2 infection by facilitating viral entry into cells. The magnitude of enhancement could be up to 10- or 100-fold, depending on cell lines used. These findings indicate that AmB and nystatin have the potential to enhance disease when given to patients infected with SARS-CoV-2 and therefore should not be used for treatment of fungal infections in active COVID-19 cases.
Unveiling the structural spectrum of SARS-CoV-2 fusion by in situ cryo-ET.
SARS-CoV-2 entry into host cells is mediated by the spike protein, which drives membrane fusion. While cryo-EM reveals stable prefusion and postfusion conformations of the spike, the transient fusion intermediate states during the fusion process remain poorly understood. Here, we design a near-native viral fusion system that recapitulates SARS-CoV-2 entry and use cryo-electron tomography (cryo-ET) to capture fusion intermediates leading to complete fusion. The spike protein undergoes extensive structural rearrangements, progressing through extended, partially folded, and fully folded intermediates prior to fusion-pore formation, a process that depends on protease cleavage and is inhibited by the WS6 S2 antibody. Upon interaction with ACE2 receptor dimer, spikes cluster at membrane interfaces and following S2' cleavage concurrently transition to postfusion conformations encircling the hemifusion and initial fusion pores in a distinct conical arrangement. S2' cleavage is indispensable for advancing fusion intermediates to the fully folded postfusion state, culminating in membrane integration. Subtomogram averaging reveals that the WS6 S2 antibody binds to the spike's stem-helix, crosslinks and clusters prefusion spikes, as well as inhibits refolding of fusion intermediates. These findings elucidate the entire process of spike-mediated fusion and SARS-CoV-2 entry, highlighting the neutralizing mechanism of S2-targeting antibodies.
Perivascular niche cells sense thrombocytopenia and activate hematopoietic stem cells in an IL-1 dependent manner.
Hematopoietic stem cells (HSCs) residing in specialized niches in the bone marrow are responsible for the balanced output of multiple short-lived blood cell lineages in steady-state and in response to different challenges. However, feedback mechanisms by which HSCs, through their niches, sense acute losses of specific blood cell lineages remain to be established. While all HSCs replenish platelets, previous studies have shown that a large fraction of HSCs are molecularly primed for the megakaryocyte-platelet lineage and are rapidly recruited into proliferation upon platelet depletion. Platelets normally turnover in an activation-dependent manner, herein mimicked by antibodies inducing platelet activation and depletion. Antibody-mediated platelet activation upregulates expression of Interleukin-1 (IL-1) in platelets, and in bone marrow extracellular fluid in vivo. Genetic experiments demonstrate that rather than IL-1 directly activating HSCs, activation of bone marrow Lepr+ perivascular niche cells expressing IL-1 receptor is critical for the optimal activation of quiescent HSCs upon platelet activation and depletion. These findings identify a feedback mechanism by which activation-induced depletion of a mature blood cell lineage leads to a niche-dependent activation of HSCs to reinstate its homeostasis.