Rapid incidence estimation from SARS-CoV-2 genomes reveals decreased case detection in Europe during summer 2020

By October 2021, 230 million SARS-CoV-2 diagnoses have been reported. Yet, a considerable proportion of cases remains undetected. Here, we propose GInPipe, a method that rapidly reconstructs SARS-CoV-2 incidence profiles solely from publicly available, time-stamped viral genomes. We validate GInPipe against simulated outbreaks and elaborate phylodynamic analyses. Using available sequence data, we reconstruct incidence histories for Denmark, Scotland, Switzerland, and Victoria (Australia) and demonstrate, how to use the method to investigate the effects of changing testing policies on case ascertainment. Specifically, we find that under-reporting was highest during summer 2020 in Europe, coinciding with more liberal testing policies at times of low testing capacities. Due to the increased use of real-time sequencing, it is envisaged that GInPipe can complement established surveillance tools to monitor the SARS-CoV-2 pandemic. In post-pandemic times, when diagnostic efforts are decreasing, GInPipe may facilitate the detection of hidden infection dynamics.Results - Method validation: in silico experiment. - Method validation: phylodynamics. - Reconstructed incidence histories. - Relative case detection rate. Discussion Method

Antiviral Resistance and Correlates of Virologic Failure in the first Cohort of HIV-Infected Children Gaining Access to Structured Antiretroviral Therapy in Lima, Peru: A Cross-Sectional Analysis

Background: The impact of extended use of ART in developing countries has been enormous. A thorough understanding of all factors contributing to the success of antiretroviral therapy is required. The current study aims to investigate the value of cross-sectional drug resistance monitoring using DNA and RNA oligonucleotide ligation assays (OLA) in treatment cohorts in low-resource settings. The study was conducted in the first cohort of children gaining access to structured ART in Peru. Methods: Between 2002–5, 46 eligible children started the standard regimen of AZT, 3TC and NFV Patients had a median age of 5.6 years (range: 0.7-14y), a median viral load of 1.7·105 RNA/ml (range: 2.1·103 – 1.2·106), and a median CD4-count of 232 cells/μL (range: 1–1591). Of these, 20 patients were classified as CDC clinical category C and 31/46 as CDC immune category 3. At the time of cross-sectional analysis in 2005, adherence questionnaires were administered. DNA OLAs and RNA OLAs were performed from frozen PBMC and plasma, RNA genotyping from dried blood spots. Results: During the first year of ART, 44% of children experienced virologic failure, with an additional 9% failing by the end of the second year. Virologic failure was significantly associated with the number of resistance mutations detected by DNA-OLA (p < 0.001) during cross-sectional analysis, but also with low immunologic CDC-scores at baseline (p < 0.001). Children who had been exposed to unsupervised short-term antiretrovirals before starting structured ART showed significantly higher numbers of resistance mutations by DNA-OLA (p = 0.01). Detection of M184V (3TC resistance) by RNA-OLA and DNA-OLA demonstrated a sensitivity of 0.93 and 0.86 and specificity of 0.67 and 0.7, respectively, for the identification of virologic failure. The RT mutations N88D and L90M (NFV resistance) detected by DNA-OLA correlated with virologic failure, whereas mutations at RT position 215 (AZT resistance) were not associated with virologic failure. Conclusions: Advanced immunosuppression at baseline and previous exposures to unsupervised brief cycles of ART significantly impaired treatment outcomes at a time when structured ART was finally introduced in his cohort. Brief maternal exposures to with AZT +/− NVP for the prevention of mother-to-child transmission did not affect treatment outcomes in this group of children. DNA-OLA from frozen PBMC provided a highly specific tool to detect archived drug resistance. RNA consensus genotyping from dried blood spots and RNA-OLA fromplasma consistently detected drug resistance mutations, but merely in association with virologic failur

HIV-1 Polymerase Inhibition by Nucleoside Analogs: Cellular- and Kinetic Parameters of Efficacy, Susceptibility and Resistance Selection

Nucleoside analogs (NAs) are used to treat numerous viral infections and cancer. They compete with endogenous nucleotides (dNTP/NTP) for incorporation into nascent DNA/RNA and inhibit replication by preventing subsequent primer extension. To date, an integrated mathematical model that could allow the analysis of their mechanism of action, of the various resistance mechanisms, and their effect on viral fitness is still lacking. We present the first mechanistic mathematical model of polymerase inhibition by NAs that takes into account the reversibility of polymerase inhibition. Analytical solutions for the model point out the cellular- and kinetic aspects of inhibition. Our model correctly predicts for HIV-1 that resistance against nucleoside analog reverse transcriptase inhibitors (NRTIs) can be conferred by decreasing their incorporation rate, increasing their excision rate, or decreasing their affinity for the polymerase enzyme. For all analyzed NRTIs and their combinations, model-predicted macroscopic parameters (efficacy, fitness and toxicity) were consistent with observations. NRTI efficacy was found to greatly vary between distinct target cells. Surprisingly, target cells with low dNTP/NTP levels may not confer hyper-susceptibility to inhibition, whereas cells with high dNTP/NTP contents are likely to confer natural resistance. Our model also allows quantification of the selective advantage of mutations by integrating their effects on viral fitness and drug susceptibility. For zidovudine triphosphate (AZT-TP), we predict that this selective advantage, as well as the minimal concentration required to select thymidine-associated mutations (TAMs) are highly cell-dependent. The developed model allows studying various resistance mechanisms, inherent fitness effects, selection forces and epistasis based on microscopic kinetic data. It can readily be embedded in extended models of the complete HIV-1 reverse transcription process, or analogous processes in other viruses and help to guide drug development and improve our understanding of the mechanisms of resistance development during treatment

Mitochondrial changes within axons in multiple sclerosis

Multiple sclerosis is the most common cause of non-traumatic neurological impairment in young adults. An energy deficient state has been implicated in the degeneration of axons, the pathological correlate of disease progression, in multiple sclerosis. Mitochondria are the most efficient producers of energy and play an important role in calcium homeostasis. We analysed the density and function of mitochondria using immunohistochemistry and histochemistry, respectively, in chronic active and inactive lesions in progressive multiple sclerosis. As shown before in acute pattern III and Balo’s lesions, the mitochondrial respiratory chain complex IV activity is reduced despite the presence of mitochondria in demyelinated axons with amyloid precursor protein accumulation, which are predominantly located at the active edge of chronic active lesions. Furthermore, the strong non-phosphorylated neurofilament (SMI32) reactivity was associated with a significant reduction in complex IV activity and mitochondria within demyelinated axons. The complex IV defect associated with axonal injury may be mediated by soluble products of innate immunity, as suggested by an inverse correlation between complex IV activity and macrophage/microglial density in chronic lesions. However, in inactive areas of chronic multiple sclerosis lesions the mitochondrial respiratory chain complex IV activity and mitochondrial mass, judged by porin immunoreactivity, are increased within approximately half of large (>2.5 μm diameter) chronically demyelinated axons compared with large myelinated axons in the brain and spinal cord. The axon-specific mitochondrial docking protein (syntaphilin) and phosphorylated neurofilament-H were increased in chronic lesions. The lack of complex IV activity in a proportion of Na+/K+ ATPase α-1 positive demyelinated axons supports axonal dysfunction as a contributor to neurological impairment and disease progression. Furthermore, in vitro studies show that inhibition of complex IV augments glutamate-mediated axonal injury (amyloid precursor protein and SMI32 reactivity). Our findings have important implications for both axonal degeneration and dysfunction during the progressive stage of multiple sclerosis

Drug-Class Specific Impact of Antivirals on the Reproductive Capacity of HIV

Predictive markers linking drug efficacy to clinical outcome are a key component in the drug discovery and development process. In HIV infection, two different measures, viral load decay and phenotypic assays, are used to assess drug efficacy in vivo and in vitro. For the newly introduced class of integrase inhibitors, a huge discrepancy between these two measures of efficacy was observed. Hence, a thorough understanding of the relation between these two measures of drug efficacy is imperative for guiding future drug discovery and development activities in HIV. In this article, we developed a novel viral dynamics model, which allows for a mechanistic integration of the mode of action of all approved drugs and drugs in late clinical trials. Subsequently, we established a link between in vivo and in vitro measures of drug efficacy, and extract important determinants of drug efficacy in vivo. The analysis is based on a new quantity—the reproductive capacity—that represents in mathematical terms the in vivo analog of the read-out of a phenotypic assay. Our results suggest a drug-class specific impact of antivirals on the total amount of viral replication. Moreover, we showed that the (drug-)target half life, dominated by immune-system related clearance processes, is a key characteristic that affects both the emergence of resistance as well as the in vitro–in vivo correlation of efficacy measures in HIV treatment. We found that protease- and maturation inhibitors, due to their target half-life, decrease the total amount of viral replication and the emergence of resistance most efficiently

Canine respiratory coronavirus employs caveolin-1-mediated pathway for internalization to HRT-18G cells

Canine respiratory coronavirus (CRCoV), identified in 2003, is a member of the Coronaviridae family. The virus is a betacoronavirus and a close relative of human coronavirus OC43 and bovine coronavirus. Here, we examined entry of CRCoV into human rectal tumor cells (HRT-18G cell line) by analyzing co-localization of single virus particles with cellular markers in the presence or absence of chemical inhibitors of pathways potentially involved in virus entry. We also targeted these pathways using siRNA. The results show that the virus hijacks caveolin-dependent endocytosis to enter cells via endocytic internalization

Computational strategies to combat COVID-19: useful tools to accelerate SARS-CoV-2 and coronavirus research

SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) is a novel virus of the family Coronaviridae. The virus causesthe infectious disease COVID-19. The biology of coronaviruses has been studied for many years. However, bioinformaticstools designed explicitly for SARS-CoV-2 have only recently been developed as a rapid reaction to the need for fast detection,understanding and treatment of COVID-19. To control the ongoing COVID-19 pandemic, it is of utmost importance to getinsight into the evolution and pathogenesis of the virus. In this review, we cover bioinformatics workflows and tools for theroutine detection of SARS-CoV-2 infection, the reliable analysis of sequencing data, the tracking of the COVID-19 pandemicand evaluation of containment measures, the study of coronavirus evolution, the discovery of potential drug targets anddevelopment of therapeutic strategies. For each tool, we briefly describe its use case and how it advances researchspecifically for SARS-CoV-2.Fil: Hufsky, Franziska. Friedrich Schiller University Jena; AlemaniaFil: Lamkiewicz, Kevin. Friedrich Schiller University Jena; AlemaniaFil: Almeida, Alexandre. the Wellcome Sanger Institute; Reino UnidoFil: Aouacheria, Abdel. Centre National de la Recherche Scientifique; FranciaFil: Arighi, Cecilia. Biocuration and Literature Access at PIR; Estados UnidosFil: Bateman, Alex. European Bioinformatics Institute. Head of Protein Sequence Resources; Reino UnidoFil: Baumbach, Jan. Universitat Technical Zu Munich; AlemaniaFil: Beerenwinkel, Niko. Universitat Technical Zu Munich; AlemaniaFil: Brandt, Christian. Jena University Hospital; AlemaniaFil: Cacciabue, Marco Polo Domingo. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación En Ciencias Veterinarias y Agronómicas. Instituto de Agrobiotecnología y Biología Molecular. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Agrobiotecnología y Biología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Chuguransky, Sara Rocío. European Bioinformatics Institute; Reino Unido. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Drechsel, Oliver. Robert Koch-Institute; AlemaniaFil: Finn, Robert D.. Biocurator for Pfam and InterPro databases; Reino UnidoFil: Fritz, Adrian. Helmholtz Centre for Infection Research; AlemaniaFil: Fuchs, Stephan. Robert Koch-Institute; AlemaniaFil: Hattab, Georges. University Marburg; AlemaniaFil: Hauschild, Anne Christin. University Marburg; AlemaniaFil: Heider, Dominik. University Marburg; AlemaniaFil: Hoffmann, Marie. Freie Universität Berlin; AlemaniaFil: Hölzer, Martin. Friedrich Schiller University Jena; AlemaniaFil: Hoops, Stefan. University of Virginia; Estados UnidosFil: Kaderali, Lars. University Medicine Greifswald; AlemaniaFil: Kalvari, Ioanna. European Bioinformatics Institute; Reino UnidoFil: von Kleist, Max. Robert Koch-Institute; AlemaniaFil: Kmiecinski, Renó. Robert Koch-Institute; AlemaniaFil: Kühnert, Denise. Max Planck Institute for the Science of Human History; AlemaniaFil: Lasso, Gorka. Albert Einstein College of Medicine; Estados UnidosFil: Libin, Pieter. Hasselt University; BélgicaFil: List, Markus. Universitat Technical Zu Munich; AlemaniaFil: Löchel, Hannah F.. University Marburg; Alemani

Prototype ATLAS IBL Modules using the FE-I4A Front-End Readout Chip

The ATLAS Collaboration will upgrade its semiconductor pixel tracking detector with a new Insertable B-layer (IBL) between the existing pixel detector and the vacuum pipe of the Large Hadron Collider. The extreme operating conditions at this location have necessitated the development of new radiation hard pixel sensor technologies and a new front-end readout chip, called the FE-I4. Planar pixel sensors and 3D pixel sensors have been investigated to equip this new pixel layer, and prototype modules using the FE-I4A have been fabricated and characterized using 120 GeV pions at the CERN SPS and 4 GeV positrons at DESY, before and after module irradiation. Beam test results are presented, including charge collection efficiency, tracking efficiency and charge sharing.Comment: 45 pages, 30 figures, submitted to JINS

Neonatal presentation of ventricular tachycardia and a Reye-like syndrome episode associated with disturbed mitochondrial energy metabolism

BACKGROUND: Hyperammonemia, hypoglycemia, hepatopathy, and ventricular tachycardia are common presenting features of carnitine-acylcarnitine translocase deficiency (Mendelian Inheritance in Man database: *212138), a mitochondrial fatty acid oxidation disorder with a lethal prognosis. These features have not been identified as the presenting features of mitochondrial cytopathy in the neonatal period. CASE PRESENTATION: We describe an atypical presentation of mitochondrial cytopathy in a 2 day-old neonate. She presented with a Reye-like syndrome episode, premature ventricular contractions and ventricular tachycardia. Initial laboratory evaluation exhibited a large amount of 3-methylglutaconic acid on urine organic acid analysis, mild orotic aciduria and a nonspecific abnormal acylcarnitine profile. The evaluation for carnitine-acylcarnitine translocase deficiency and other fatty acid oxidation disorders was negative. The patient later developed a hypertrophic cardiomyopathy and continued to be affected by recurrent Reye-like syndrome episodes triggered by infections. A muscle biopsy exhibited signs of a mitochondrial cytopathy. During the course of her disease, her Reye-like syndrome episodes have subsided; however, cardiomyopathy has persisted along with fatigue and exercise intolerance. CONCLUSIONS: This case illustrates that, in the neonatal period, hyperammonemia and ventricular tachycardia may be the presenting features of a lethal carnitine-acylcarnitine translocase deficiency or of a mitochondrial cytopathy, associated with a milder clinical course. This association broadens the spectrum of presenting phenotypes observed in patients with disturbed mitochondrial energy metabolism. Also, the presence of 3-methylglutaconic aciduria suggests mitochondrial dysfunction and mild orotic aciduria could potentially be used as a marker of mitochondrial disease