bluuebunny/crossref_metadata_2025 huggingface dataset:
Limited within-host diversity and tight transmission bottlenecks limit SARS-CoV-2 evolution in acutely infected individuals
Authors: Braun Katarina , Moreno Gage , Wagner Cassia , Accola Molly A. , Rehrauer William M. , Baker David , Koelle Katia , O’Connor David H. , Bedford Trevor , Friedrich Thomas C. , Moncla Louise H.
Published: May 2021

The recent emergence of divergent SARS-CoV-2 lineages has raised concerns about the role of selection within individual hosts in propagating novel variants. Of particular concern are variants associated with immune escape and/or enhanced transmissibility. Though growing evidence suggests that novel variants can arise during prolonged infections, most infections are acute. Understanding the extent to which variants emerge and transmit among acutely infected hosts is therefore critical for predicting the pace at which variants resistant to vaccines or conferring increased transmissibility might emerge in the majority of SARS-CoV-2 infections. To characterize how within-host diversity is generated and propagated, we combine extensive laboratory and bioinformatic controls with metrics of within- and between-host diversity to 133 SARS-CoV-2 genomes from acutely infected individuals. We find that within-host diversity during acute infection is low and transmission bottlenecks are narrow, with very few viruses founding most infections. Within-host variants are rarely transmitted, even among individuals within the same household. Accordingly, we also find that within-host variants are rarely detected along phylogenetically linked infections in the broader community. Together, these findings suggest that efficient selection and transmission of novel SARS-CoV-2 variants is unlikely during typical, acute infection.

One Sentence Summary

Patterns of SARS-CoV-2 within hosts suggest efficient selection and transmission of novel variants is unlikely during typical, acute infection.
RelA-SpoT Homologue toxins pyrophosphorylate the CCA end of tRNA to inhibit protein synthesis
Authors: Kurata Tatsuaki , Brodiazhenko Tetiana , Oliveira Sofia Raquel Alves , Roghanian Mohammad , Turnbull Kathryn Jane , Bulvas Ondřej , Takada Hiraku , Taman Hedvig , Ainelo Andres , Pohl Radek , Rejman Dominik , Tenson Tanel , Garcia-Pino Abel , Atkinson Gemma C. , Hauryliuk Vasili
Published: May 2021

RelA-SpoT Homolog (RSH) enzymes control bacterial physiology through synthesis and degradation of the nucleotide alarmone (p)ppGpp. We recently discovered multiple families of Small Alarmone Synthetase (SAS) RSH acting as toxins of toxin-antitoxin (TA) modules, with the FaRel subfamily of toxSAS abrogating bacterial growth by producing an analogue of (p)ppGpp, (pp)pApp. Here we probe the mechanism of growth arrest employed by four experimentally unexplored subfamilies of toxSAS: FaRel2, PhRel, PhRel2 and CapRel. Surprisingly, all these toxins specifically inhibit protein synthesis. To do so, they transfer a pyrophosphate moiety from ATP to the tRNA 3′ CCA. The modification inhibits both tRNA aminoacylation and the sensing of cellular amino acid starvation by the ribosome-associated RSH RelA. Conversely, we show that some Small Alarmone Hydrolase (SAH) RSH enzymes can reverse the pyrophosphorylation of tRNA to counter the growth inhibition by toxSAS. Collectively, we establish RSHs as a novel class of RNA-modifying enzymes.
Covariance-based information processing in reservoir computing systems
Authors: Lawrie Sofía , Moreno-Bote Rubén , Gilson Matthieu
Published: May 2021

In biological neuronal networks, information representation and processing are achieved through plasticity learning rules that have been empirically characterized as sensitive to second and higher-order statistics in spike trains. However, most models in both computational neuroscience and machine learning aim to convert diverse statistical properties in inputs into first-order statistics in outputs, like in modern deep learning networks. In the context of classification, such schemes have merit for inputs like static images, but they are not well suited to capture the temporal structure in time series. In contrast, the recently developed covariance perceptron uses second-order statistics by mapping input covariances to output covariances in a consistent fashion. Here, we explore the applicability of covariance-based perceptron readouts in reservoir computing networks to classify synthetic multivariate time series structured at different statistical orders (first and second). We show that the second-order framework outperforms or matches the classical mean paradigm in terms of accuracy. Our results highlight a nontrivial relationship between input and reservoir properties in generating the output reservoir activity, which suggests an important role for recurrent connectivity in transforming information representations in biologically inspired architectures. Finally, we solve a speech recognition task for the classification of spoken digits to further demonstrate the potential of covariance-based decoding for real data.
Low-cost, handheld near-infrared spectroscopy for root dry matter content prediction in cassava
Authors: Hershberger Jenna , Mbanjo Edwige Gaby Nkouaya , Peteti Prasad , Ikpan Andrew Smith , Ogunpaimo Kayode , Nafiu Kehinde , Rabbi Ismail Y. , Gore Michael A.
Published: May 2021

Over 800 million people across the tropics rely on cassava as a major source of calories. While the root dry matter content (RDMC) of this starchy root crop is important for both producers and consumers, characterization of RDMC by traditional methods is time-consuming and laborious for breeding programs. Alternate phenotyping methods have been proposed but lack the accuracy, cost, or speed ultimately needed for cassava breeding programs. For this reason, we investigated the use of a low-cost, handheld NIR spectrometer for field-based RDMC prediction in cassava. Oven-dried measurements of RDMC were paired with 21,044 scans of roots of 376 diverse clones from 10 field trials in Nigeria and grouped into training and test sets based on cross-validation schemes relevant to plant breeding programs. Mean partial least squares regression model performance ranged from R2p= 0.62 - 0.89 for within-trial predictions, which is within the range achieved with laboratory-grade spectrometers in previous studies. Relative to other factors, model performance was highly impacted by the inclusion of samples from the same environment in both the training and test sets. Random forest variable importance analysis of root spectra revealed increased importance in a region previously identified as predictive of water content in plants (~950 - 990 nm). With appropriate model calibration, the tested spectrometer will allow for field-based collection of spectral data with a smartphone for accurate RDMC prediction and potentially other quality traits, a step that could be easily integrated into existing harvesting workflows of cassava breeding programs.

CORE IDEAS

A low-cost, handheld near-infrared spectrometer was tested for phenotyping of cassava roots

Plant breeding-relevant cross-validation schemes were used for predictions

High prediction accuracies were achieved for cassava root dry matter content

A spectral region predictive of plant water content was identified as important
Multifunctional optrode for opsin delivery, optical stimulation, and electrophysiological recordings in freely moving rats
Authors: Sharma Kirti , Jaeckel Zoe , Schneider Artur , Paul Oliver , Diester Ilka , Ruther Patrick
Published: May 2021

Objective

Optogenetics involves delivery of light-sensitive opsins to the target brain region, as well as introduction of optical and electrical devices to manipulate and record neural activity, respectively, from the targeted neural population. Combining these functionalities in a single implantable device is of great importance for a precise investigation of neural networks while minimizing tissue damage.

Approach

We report on the development, characterization, and in vivo validation of a multifunctional optrode that combines a silicon-based neural probe with an integrated microfluidic channel, and an optical glass fiber in a compact assembly. The silicon probe comprises an 11-μm-wide fluidic channel and 32 recording electrodes (diameter 30 μm) on a tapered probe shank with a length, thickness, and maximum width of 7.5 mm, 50 μm, and 150 μm, respectively. The size and position of fluidic channels, electrodes, and optical fiber can be precisely tuned according to the in vivo application.

Main results

With a total system weight of 0.97 g, our multifunctional optrode is suitable for chronic in vivo experiments requiring simultaneous drug delivery, optical stimulation, and neural recording. We demonstrate the utility of our device in optogenetics by injecting a viral vector carrying a ChR2-construct in the prefrontal cortex and subsequent photostimulation of the transfected neurons while recording neural activity from both the target and adjacent regions in a freely moving rat. Additionally, we demonstrate a pharmacological application of our device by injecting GABA antagonist bicuculline in an anesthetized rat brain and simultaneously recording the electrophysiological response.

Significance

Our triple-modality device enables a single-step optogenetic surgery. In comparison to conventional multi-step surgeries, our approach achieves higher spatial specificity while minimizing tissue damage.

Graphical abstract
Natural tolerance to transposition is associated with double-strand break repair and germ-cell differentiation
Authors: Lama Jyoti , Srivastav Satyam , Tasnim Sadia , Hubbard Donald , Hadjipanteli Savana , Kelleher Erin S.
Published: May 2021

Transposable elements (TE) are mobile genetic parasites whose unregulated activity in the germline causes DNA damage and sterility. While the regulation of TE mobilization by hosts is studied extensively, little is known about mechanisms that could allow germline cells to persist in the face of genotoxic stress imposed by active transposition. Such tolerance mechanisms are predicted to be beneficial when new TEs invade and host repression has not yet evolved. Here we use hybrid dysgenesis—a sterility syndrome ofDrosophilacaused by transposition of invading DNA transposons—to uncover genetic variants that confer tolerance to transposition. Using a panel of highly recombinant inbred lines ofDrosophila melanogaster, we identified two linked quantitative trait loci (QTL), that determine tolerance in young and old females, respectively. Through transcriptomic and phenotypic comparisons, we provide evidence that young tolerant females exhibit enhanced repair of double-stranded breaks, explaining their ability to withstand high germline transposition rates. We furthermore identify the germline differentiation factorbratas an independent tolerance factor, whose activity may promote germline maintenance in aging dysgenic females. Together, our work reveals the diversity of potential tolerance mechanisms across development, as well as tolerant variants that may be beneficial in the context ofP-element transposition.
Signatures of copy number alterations in human cancer
Authors: Steele Christopher D. , Abbasi Ammal , Islam S. M. Ashiqul , Khandekar Azhar , Haase Kerstin , Hames Shadi , Tarabichi Maxime , Lesluyes Tom , Flanagan Adrienne M. , Mertens Fredrik , Van Loo Peter , Alexandrov Ludmil B. , Pillay Nischalan
Published: May 2021

The gains and losses of DNA that emerge as a consequence of mitotic errors and chromosomal instability are prevalent in cancer. These copy number alterations contribute to cancer initiaition, progression and therapeutic resistance. Here, we present a conceptual framework for examining the patterns of copy number alterations in human cancer using whole-genome sequencing, whole-exome sequencing, and SNP6 microarray data making it widely applicable to diverse datasets. Deploying this framework to 9,873 cancers representing 33 human cancer types from the TCGA project revealed a set of 19 copy number signatures that explain the copy number patterns of 93% of TCGA samples. 15 copy number signatures were attributed to biological processes of whole-genome doubling, aneuploidy, loss of heterozygosity, homologous recombination deficiency, and chromothripsis. The aetiology of four copy number signatures are unexplained and some cancer types have unique patterns of amplicon signatures associated with extrachromosomal DNA, disease-specific survival, and gains of proto-oncogenes such asMDM2. In contrast to base-scale mutational signatures, no copy number signature associated with known cancer risk factors. The results provide a foundation for exploring patterns of copy number changes in cancer genomes and synthesise the global landscape of copy number alterations in human cancer by revealing a diversity of mutational processes giving rise to copy number changes.
Maternal Piwi Regulates Primordial Germ Cell Development to Ensure the Fertility of Female Progeny inDrosophila
Authors: Gonzalez Lauren E , Tang Xiongzhuo , Lin Haifan
Published: May 2021

In many animals, germline development is initiated by proteins and RNAs that are expressed maternally. PIWI proteins and their associated small noncodingPIWI-interacting RNAs (piRNAs), which guide PIWI to target RNAs by base-pairing, are among the maternal components deposited into the germline of the early embryo inDrosophila. Piwi has been extensively studied in the adult ovary and testis, where it is required for transposon suppression, germline stem cell self-renewal, and fertility. Consequently, loss of Piwi in the adult ovary usingpiwi-null alleles or knockdown from early oogenesis results in complete sterility, limiting investigation into possible embryonic functions of maternal Piwi. In this study, we show that the maternal Piwi protein persists in the embryonic germline through gonad coalescence, suggesting that maternal Piwi can regulate germline development beyond early embryogenesis. Using a maternal knockdown strategy, we find that maternal Piwi is required for the fertility and normal gonad morphology of female, but not male, progeny. Following maternalpiwiknockdown, transposons were mildly derepressed in the early embryo but were fully repressed in the ovaries of adult progeny. Furthermore, the maternal piRNA pool was diminished, reducing the capacity of the PIWI/piRNA complex to target zygotic genes during embryogenesis. Examination of embryonic germ cell proliferation and ovarian gene expression showed that the germline of female progeny was partially masculinized by maternalpiwiknockdown. Our study reveals a novel role for maternal Piwi in the germline development of female progeny and suggests that the PIWI/piRNA pathway is involved in germline sex determination inDrosophila.
Single-molecule imaging of microRNA-mediated gene silencing in cells
Authors: Kobayashi Hotaka , Singer Robert H.
Published: May 2021

MicroRNAs (miRNAs) are small non-coding RNAs, which regulate the expression of thousands of genes; miRNAs silence gene expression from complementary mRNAs through translational repression and mRNA decay. For decades, the function of miRNAs has been studied primarily by ensemble methods, where a bulk collection of molecules is measured outside cells. Thus, the behavior of individual molecules during miRNA-mediated gene silencing, as well as their spatiotemporal regulation inside cells, remains mostly unknown. Here we report single-molecule methods to visualize each step of miRNA-mediated gene silencingin situinside cells. Simultaneous visualization of single mRNAs, translation, and miRNA-binding revealed that miRNAs preferentially bind to translated mRNAs rather than untranslated mRNAs. Spatiotemporal analysis based on our methods uncovered that miRNAs bind to mRNAs immediately after nuclear export. Subsequently, miRNAs induced translational repression and mRNA decay within 30 and 60 min, respectively, after the binding to mRNAs. This methodology provides a framework for studying mRNA regulation at the single-molecule level with spatiotemporal information inside cells.
AutoGGN: A Gene Graph Network AutoML Tool for Multi-Omics Research
Authors: Zhang Lei , Li Ping , Shen Wen , Xu Chi , Liu Denghui , He Wenjun , Xu Zhimeng , Zhang Chenyi , Qiao Nan
Published: May 2021

Omics data identifies biological characteristics from genetic to phenotypic levels during the life span. Molecular interaction networks have a fundamental impact on life activities. Integrating omics data and molecular interaction networks will help researchers delve into comprehensive information underlying the data. Here, we proposed a new multimodal method called AutoGGN to aggregate multi-omics data and molecular interaction networks based on graph convolutional neural networks. We evaluated AutoGGN using two different tasks: cancer type classification and single-cell stage classification. On both tasks, AutoGGN showed better performance compared to other methods, the trend is relevant to the ability of utilizing much more information from biological data. The phenomenon indicated AutoGGN has the potential to incorporate valuable information from molecular interaction networks and multi-omics data effectively. Furthermore, in order to provide a better understanding of the mechanism of prediction results, we assessed the explanation using SHAP module and identified the key genes contributing to the prediction of classification, which will provide insights for the downstream design of biological experiments.
Metagenomic investigation of the equine faecal microbiome reveals extensive taxonomic and functional diversity
Authors: Gilroy Rachel , Leng Joy , Ravi Anuradha , Adriaenssens Evelien M. , Oren Aharon , Baker David , La Ragione Roberto M. , Proudman Christopher , Pallen Mark J.
Published: May 2021

Background

The horse plays crucial roles across the globe, including in horseracing, as a working and companion animal and as a food animal. The horse hindgut microbiome makes a key contribution in turning a high fiber diet into body mass and horsepower. However, despite its importance, the horse hindgut microbiome remains largely undefined. Here, we applied culture-independent shotgun metagenomics to thoroughbred equine faecal samples to deliver novel insights into this complex microbial community.

Results

We performed metagenomic sequencing on five equine faecal samples to construct 123 high- or medium-quality metagenome-assembled genomes from Bacteria and Archaea. In addition, we recovered nearly 200 bacteriophage genomes. We document surprising taxonomic and functional diversity, encompassing dozens of novel or unnamed bacterial genera and species, to which we have assigned newCandidatusnames. Many of these genera are conserved across a range of mammalian gut microbiomes.

Conclusions

Our metagenomic analyses provide new insights into the bacterial, archaeal and bacteriophage components of the horse gut microbiome. The resulting datasets provide a key resource for future high-resolution taxonomic and functional studies on the equine gut microbiome.
Diverse mechanisms for epigenetic imprinting in mammals
Authors: Andergassen Daniel , Smith Zachary D. , Rinn John L. , Meissner Alexander
Published: May 2021

Genomic imprinting and X chromosome inactivation (XCI) require epigenetic mechanisms to direct allele-specific expression. Despite their critical roles in embryonic development, how universal epigenetic regulators coordinate these specific tasks from single locus to chromosome-scale remains understudied. Here, we systematically disrupted multiple essential epigenetic pathways within polymorphic F1 zygotes to examine postimplantation effects on canonical and non-canonical genomic imprinting as well as X chromosome inactivation. We find that DNA methylation and Polycomb group repressors are both indispensable for autosomal imprinting, albeit at distinct gene sets. Moreover, the extraembryonic ectoderm relies on a broader spectrum of unique imprinting mechanisms, including non-canonical targeting of maternal endogenous retrovirus (ERV) driven promoters by G9a. We further utilize our data to identify Polycomb dependent and independent gene clusters on the imprinted X chromosome, which appears to reflect distinct domains of Xist-mediated suppression. Our data has allowed us to assemble a comprehensive inventory of the epigenetic mechanisms utilized in eutherian mammals to maintain parent-specific imprinting, including an expanded view of the placental lineage that comprises multiple unique pathways.
Neurodegenerative and functional signatures of the cerebellar cortex in m.3243A>G patients
Authors: Haast Roy AM. , De Coo Irenaeus FM. , Ivanov Dimo , Khan Ali R , Jansen Jacobus FA. , Smeets Hubert JM. , Uludağ Kâmil
Published: May 2021

Mutations of the mitochondrial DNA are an important cause of inherited diseases that can severely affect the tissue’s homeostasis and integrity. The m.3243A>G mutation is the most commonly observed across mitochondrial disorders and is linked to multisystemic complications, including cognitive deficits. In line with in vitro experiments demonstrating the m.3243A>G’s negative impact on neuronal energy production and integrity, m.3243A>G patients show cerebral gray matter tissue changes. However, its impact on the most neuron-dense, and therefore energy-consuming brain structure – the cerebellum – remains elusive.

In this work, we used high resolution structural and functional data acquired using 7 Tesla MRI to characterize the neurodegenerative and functional signatures of the cerebellar cortex in m.3243A>G patients. Our results reveal altered tissue integrity within distinct clusters across the cerebellar cortex, apparent by their significantly reduced volume and longitudinal relaxation rate compared to healthy controls, indicating macroscopic atrophy and microstructural pathology. Spatial characterization reveals that these changes occur especially in regions related to the frontoparietal brain network that is involved in information processing and selective attention. In addition, based on resting-state fMRI data, these clusters exhibit reduced functional connectivity to frontal and parietal cortical regions, especially in patients characterized by (i) a severe disease phenotype and (ii) reduced information processing speed and attention control.

Combined with our previous work, these results provide insight into the neuropathological changes and a solid base to guide longitudinal studies aimed to track disease progression.
Multi-animal pose estimation and tracking with DeepLabCut
Authors: Lauer Jessy , Zhou Mu , Ye Shaokai , Menegas William , Nath Tanmay , Rahman Mohammed Mostafizur , Di Santo Valentina , Soberanes Daniel , Feng Guoping , Murthy Venkatesh N. , Lauder George , Dulac Catherine , Mathis Mackenzie W. , Mathis Alexander
Published: May 2021

Estimating the pose of multiple animals is a challenging computer vision problem: frequent interactions cause occlusions and complicate the association of detected keypoints to the correct individuals, as well as having extremely similar looking animals that interact more closely than in typical multi-human scenarios. To take up this challenge, we build on DeepLabCut, a popular open source pose estimation toolbox, and provide high-performance animal assembly and tracking—features required for robust multi-animal scenarios. Furthermore, we integrate the ability to predict an animal’s identity directly to assist tracking (in case of occlusions). We illustrate the power of this framework with four datasets varying in complexity, which we release to serve as a benchmark for future algorithm development.
The impact of multiple species invasion on soil and plant communities increases with invader diversity
Authors: Dušanka Vujanović , Gianalberto Losapio , Stanko Milić , Dubravka Milić
Published: May 2021

Despite increasing evidence indicating that invasive species are harming ecological systems and processes, impacts of multiple invasions, and the linkages between these events and changes in vegetation and soil are inadequately documented and remain poorly understood. Addressing multiple invasions would help to highlight high priority invaders and would aid in designing more effective control strategies, contributing to environmental restoration and sustainability. In this work, we tested the impact of three concurring invasive plant species,Amorpha fruticosa, Fraxinus pennsylvanicaandAcer negundo, on soil conditions and native plant diversity. The research was conducted in riparian ecosystem and included the following treatments: (1) co-occurrence of the three invasive plant species, (2) occurrence of a single invasive species, and (3) control, i.e., absence of invasive species. Our findings revealed that the impact of invasive plants on soil properties and native plant diversity is magnified by their co-occurrence. Soil in mixed plots (those populated with all three invaders) contained much higher levels of nitrifying bacteria (NB), organic matter (Om), nitrogen (N), and carbon (C) as well as lower carbon to nitrogen ratio (C:N) levels, compared to single species invaded plots and control plots. Mixed plots were also characterized by reduced native plant diversity compared to single species invaded and control plots. Differences in soil conditions and native plant diversity revealed the interactive potential of invasive plants in depleting biodiversity, and thus in affecting ecological and biogeochemical processes. Our results highlight the need to study the impact of multispecies invasion and suggest that sites in riparian areas affected by co-occurring invaders, should be prioritized for ecosystem restoration.
Genetic diversity and connectivity of the Ostreid herpesvirus 1 populations in France: a first attempt to phylogeographic inference for a marine mollusc disease
Authors: Delmotte Jean , Pelletier Camille , Morga Benjamin , Galinier Richard , Petton Bruno , Lamy Jean-Baptiste , Kaltz Oliver , Avarre Jean-Christophe , Jacquot Maude , Montagnani Caroline , Escoubas Jean-Michel
Published: May 2021

The genetic diversity of viral populations is a key driver of the spatial and temporal diffusion of viruses; yet, studying the diversity of whole genomes from natural populations still remains a challenge. Phylodynamic approaches are commonly used for RNA viruses harboring small genomes, but have only rarely been applied to DNA viruses with larger genomes. Here, we used the Pacific Oyster Mortality Syndrome (POMS, a disease that affects oyster farms around the world) as a model to study the genetic diversity of its causative agent, the Ostreid herpesvirus 1 (OsHV-1) in the three main French oyster-farming areas. Using ultra-deep sequencing on individual moribund oysters and an innovative combination of bioinformatics tools, wede novoassembled 21 OsHV-1 new genomes. Combining quantification of major and minor genetic variations, phylogenetic analysis and ancestral state reconstruction of discrete traits approaches; we assessed the connectivity of OsHV-1 viral populations between the three oyster-farming areas. Our results suggest that the Marennes- Oléron Bay represents the main source of OsHV-1 diversity, from where the virus has dispersed to other farming areas, a scenario consistent with current practices of oyster transfers in France. We demonstrate that phylodynamic approaches can be applied to aquatic DNA viruses to determine how epidemiological, immunological, and evolutionary processes act and potentially interact to shape their diversity patterns.

Importance

Phylogeography is a field of research that attempts to reconstruct the relationships between individual genotypes within a species and then correlate these genealogical relationships with their geographic and temporal origin. This field of research has become an essential step in the understanding of pandemics, in particular to determine the origin, spread and evolution of a pathogen as currently illustrated in studies on viral pandemics. However, because phylogeographic analyses are based on genome variation, stable genomes yield less information than labile genomes. Accordingly, viruses with double-stranded DNA (dsDNA) genomes generally have lower nucleotide diversity than RNA viruses. In this study, by combining the use of both major and minor genetic variations with phylogeographic analyses of the oyster herpesvirus OsHV-1, we highlight genealogical relationships that are not depicted in phylogenetic trees based on consensus viral genomes only. These data offer a plausible scenario reflecting the origin and spread of OsHV-1 populations between oyster- farming sites.
A comparison of tools for copy-number variation detection in germline whole exome and whole genome sequencing data
Authors: Gabrielaite Migle , Torp Mathias Husted , Rasmussen Malthe Sebro , Andreu-Sánchez Sergio , Vieira Filipe Garrett , Pedersen Christina Bligaard , Kinalis Savvas , Madsen Majbritt Busk , Yde Christina Westmose , Rønn Olsen Lars , Marvig Rasmus L. , Østrup Olga , Rossing Maria , Nielsen Finn Cilius , Winther Ole , Bagger Frederik Otzen
Published: May 2021

Background

Copy-number variations (CNVs) have important clinical implications for several diseases and cancers. The clinically relevant CNVs are hard to detect because CNVs are common structural variations that define large parts of the normal human genome. CNV calling from short-read sequencing data has the potential to leverage available cohort studies and allow full genomic profiling in the clinic without the need for additional data modalities. Questions regarding performance of CNV calling tools for clinical use and suitable sequencing protocols remain poorly addressed, mainly because of the lack of good reference data sets.

Methods

We reviewed 50 popular CNV calling tools and included 11 tools for benchmarking in a unique reference cohort encompassing 39 whole genome sequencing (WGS) samples paired with analysis by the current clinical standard —SNP-array based CNV calling. Additionally, for nine of these samples we performed whole exome sequencing (WES) performed, in order to address the effect of sequencing protocol on CNV calling. Furthermore, we included Gold Standard reference sample NA12878, and tested 12 samples with CNVs confirmed by multiplex ligation-dependent probe amplification (MLPA).

Results

Tool performance varied greatly in the number of called CNVs and bias for CNV lengths. Some tools had near-perfect recall of CNVs from arrays for some samples, but poor precision. Filtering output by CNV ranks from tools did not salvage precision. Several tools had better performance patterns for NA12878, and we hypothesize that this is the result of overfitting during the tool development.

Conclusions

We suggest combining tools with the best recall: GATK gCNV, Lumpy, DELLY, and cn.MOPS. These tools also capture different CNVs. Further improvements in precision requires additional development of tools, reference data sets, and annotation of CNVs, potentially assisted by the use of background panels for filtering of frequently called variants.
Nanopore sequencing reveals endogenous NMD-targeted isoforms in human cells
Authors: Karousis Evangelos D. , Gypas Foivos , Zavolan Mihaela , Mühlemann Oliver
Published: May 2021

Background

Nonsense-mediated mRNA decay (NMD) is a eukaryotic, translation-dependent degradation pathway that targets mRNAs with premature termination codons and also regulates the expression of some mRNAs that encode full-length proteins. Although many genes express NMD-sensitive transcripts, identifying them based on short-read sequencing data remains a challenge.

Results

To identify and analyze endogenous targets of NMD, we applied cDNA Nanopore sequencing and short-read sequencing to human cells with varying expression levels of NMD factors. Our approach detects full-length NMD substrates that are highly unstable and increase in levels or even only appear when NMD is inhibited. Among the many new NMD-targeted isoforms that our analysis identified, most derive from alternative exon usage. The isoform-aware analysis revealed many genes with significant changes in splicing but no significant changes in overall expression levels upon NMD knockdown. NMD-sensitive mRNAs have more exons in the 3΄UTR and, for those mRNAs with a termination codon in the last exon, the length of the 3΄UTRper sedoes not correlate with NMD sensitivity. Analysis of splicing signals reveals isoforms where NMD has been co-opted in the regulation of gene expression, though the main function of NMD seems to be ridding the transcriptome of isoforms resulting from spurious splicing events.

Conclusions

Long-read sequencing enabled the identification of many novel NMD-sensitive mRNAs and revealed both known and unexpected features concerning their biogenesis and their biological role. Our data provide a highly valuable resource of human NMD transcript targets for future genomic and transcriptomic applications.
New App knock-in mice that accumulate wild-type human Aβ as rapidly as AppNL-G-F mice exhibit intensive cored plaque pathology and neuroinflammation
Authors: Sato Kaori , Watamura Naoto , Fujioka Ryo , Mihira Naomi , Sekiguchi Misaki , Nagata Kenichi , Ohshima Toshio , Saito Takashi , Saido Takaomi C. , Sasaguri Hiroki
Published: May 2021

We previously developed single App knock-in mouse models of Alzheimer’s disease (AD), harboring the Swedish and Beyreuther/Iberian mutations with or without the Arctic mutation (AppNL-G-F and AppNL-F mice). These models showed amyloid β peptide (Aβ) pathology, neuroinflammation and cognitive impairment in an age-dependent manner. The former line exhibits extensive pathology as early as 6 months but is unsuitable for investigating Aβ metabolism and clearance because the Arctic mutation renders Aβ resistant to proteolytic degradation and prone to aggregation. In particular, it is inapplicable to preclinical immunotherapy studies due to its discrete affinity for anti-Aβ antibodies. The weakness of the latter model is that it may take as long as 18 months for the pathology to become prominent. We have thus generated a new model that exhibits early deposition of wild-type human Aβ by crossbreeding the AppNL-F line with the Psen1P117L/WT line. We show that the effects of the pathogenic mutations in the App and Psen1 genes are additive or synergistic. This new mouse model showed more cored plaque pathology and neuroinflammation than AppNL-G-F mice and will help accelerate the development of disease-modifying therapies to treat preclinical AD.
The adaptive potential of non-heritable somatic mutations
Authors: Majic Paco , Yagmur Erten E. , Payne Joshua L.
Published: May 2021

Non-heritable somatic mutations are typically associated with deleterious effects such as in cancer and senescence, so their role in adaptive evolution has received little attention. However, most somatic mutations are harmless and some even confer a fitness advantage to the organism carrying them. We hypothesized that heritable, germline genotypes that are likely to express an advantageous phenotype via non-heritable somatic mutation will have a selective advantage over other germline genotypes, and this advantage will channel evolving populations toward more fit germline genotypes, thus promoting adaptation. We tested this hypothesis by simulating evolving populations of developing organisms with an impermeable germline-soma separation navigating a minimal fitness landscape. The simulations revealed the conditions under which non-heritable somatic mutations promote adaptation. Specifically, this can occur when the somatic mutation supply is high, when only very few cells with the advantageous somatic mutation are required to increase organismal fitness, and when the somatic mutation also confers a selective advantage to cells with that mutation. We therefore provide proof-of-principle that non-heritable somatic mutations can promote adaptive evolution via a process we call somatic genotypic exploration. We discuss the biological plausibility of this phenomenon, as well as its evolutionary implications.

Significance

The immensity of non-heritable genetic diversity arising in the soma has been largely disregarded in evolutionary theory as a source of adaptation. Here, we introduce a model in which non-heritable somatic mutations arising during development confer an organismal fitness advantage. Analysis of this model shows how such mutations channel evolving populations toward adaptive germline genotypes. This is most likely to occur when somatic mutations confer a fitness benefit at both the cellular and organismal levels, evoking a synergistic form of multi-level selection that contrasts with the antagonistic forms typically associated with somatic mutations in cancer and senescence. As such, our study invites a new view of somatic genetic diversity in evolutionary theory as a potential source of adaptation.
Long-term sensitization of rat spinal neurons induced by adolescent psychophysical stress is further enhanced by a mild-nociceptive lumbar input
Authors: Singaravelu Sathish Kumar , Goitom Alexander Dawit , Surakka Akseli , Moerz Handan , Schilder Andreas , Hansson Anita C , Spanagel Rainer , Treede Rolf-Detlef
Published: May 2021

Background

Non-specific low back pain (LBP) is one of the most common chronic pain conditions and adverse childhood experiences (ACEs) are known mediators for chronicity of LBP. Sensitization of dorsal horn neurons (DHNs) is a significant element that contributes to chronic LBP. Repeated restraint stress in adult animals is known to cause manifest DHN sensitization when combined with a short-lasting nociceptive input.

Objective

In this study, we investigated whether repeated restraint stress in early adolescence leads to a long-term sensitization of DHNs and if an additional mild-nociceptive input by intramuscular nerve growth factor (NGF) leads to further sensitization.

Methods

Adolescent Wistar rats were stressed repeatedly in a narrow plastic restrainer, 1 hour per day for 12 consecutive days. Control animals were handled but not restrained. In adulthood, rats were treated with intramuscular injections of saline or NGF (short-lasting mild-nociceptive input) into the lumbar multifidus muscle (L5). Behavioral tests for pain sensitivity were performed before, after stress and inconjunction with intramuscular injections. Rats were transcardially perfused and immunohistochemistry was performed on lumbar (L2) spinal segments.

Results

Adolescent restraint stress significantly lowered the low back pressure pain threshold (PPT) immediately after the stress (p<0.0001) and was maintained throughout adulthood (p<0.05). Additionally, paw withdrawal threshold (PWT) was significantly lowered by stress (p<0.0001) but normalized towards adulthood. An NGF injection in adulthood in previously stressed animals lowered the PPT (Cohen’sd=0.87) and increased microglia marker (Iba-1) immunoreactive area in the superficial DHN (p<0.01) with a trend in increased feret’s diameter of the immunoreactive cell (p=0.05).

Conclusions

Our adolescence stress model induced behavioral signs of sensitization and enhanced sensitivity to further sensitization and dorsal horn microglia activation by subsequent mild nociceptive input (NGF injection). These findings help to understand certain aspects of how adolescent stress might predispose to exacerbation of pain with an additional insult.
Benchmarking metagenomics classifiers on ancient viral DNA: a simulation study
Authors: Arizmendi Cárdenas Yami Ommar , Neuenschwander Samuel , Malaspinas Anna-Sapfo
Published: May 2021

Owing to technological advances in ancient DNA, it is now possible to sequence viruses from the past to track down their origin and evolution. However, ancient DNA data is considerably more degraded and contaminated than modern data making the identification of ancient viral genomes particularly challenging. Several methods to characterise the modern microbiome (and, within this, the virome) have been developed; in particular, tools that assign sequenced reads to specific taxa in order to characterise the organisms present in a sample of interest. While these existing tools are routinely used in modern data, their performance when applied to ancient microbiome data to screen for ancient viruses remains unknown.

In this work, we conducted an extensive simulation study using public viral sequences to establish which tool is the most suitable to screen ancient samples for human DNA viruses. We compared the performance of four widely used classifiers, namely Centrifuge, Kraken2, DIAMOND and MetaPhlAn2, in correctly assigning sequencing reads to the corresponding viruses. To do so, we simulated reads by adding noise typical of ancient DNA to a set of publicly available human DNA viral sequences and to the human genome. We fragmented the DNA into different lengths, added sequencing error and C to T and G to A deamination substitutions at the read termini. Then we measured the resulting sensitivity and precision for all classifiers.

Across most simulations, more than 228 out of the 233 simulated viruses are recovered by Centrifuge, Kraken2 and DIAMOND, in contrast to MetaPhlAn2 which recovers only around one third. Overall, Centrifuge and Kraken2 have the best performance with the highest values of sensitivity and precision. We found that deamination damage has little impact on the performance of the classifiers, less than the sequencing error and the length of the reads. Since Centrifuge can handle short reads (in contrast to DIAMOND and Kraken2 with default settings) and since it achieves the highest sensitivity and precision at the species level across all the simulations performed, it is our recommended tool. Regardless of the tool used, our simulations indicate that, for ancient human studies, users should use strict filters to remove all reads of potential human origin. Finally, we recommend to verify which species are present in the database used, as it might happen that default databases lack sequences for viruses of interest.
Spiking activity in the visual thalamus is coupled to pupil dynamics across temporal scales
Authors: Crombie Davide , Spacek Martin A. , Leibold Christian , Busse Laura
Published: May 2021

The processing of sensory information, even at early processing stages, is influenced by the internal state of the animal. Internal states, such as arousal, are often characterized by relating neural activity to a single “level” of arousal, defined by a behavioral indicator such as pupil size. In this study, we expand the understanding of arousal-related modulations in sensory systems by uncovering multiple timescales of pupil dynamics and their relationship to neural activity. Specifically, we observed coupling between spiking activity in the mouse dorsal lateral geniculate nucleus (dLGN) of the thalamus and pupil dynamics across timescales spanning three orders of magnitude, from seconds to minutes. Throughout all of these timescales, two distinct spiking patterns – tonic spikes and bursts – preferred opposing phases of pupil dynamics. This multi-scale coupling captures modulations distinct from those captured by pupil sizeper se, transitions between locomotion and quiescence, or saccadic eye movements. Furthermore, coupling persisted even during viewing of a naturalistic movie, where it contributed to differences in how visual information was encoded. We conclude that dLGN spiking activity is influenced by arousal processes associated with pupil dynamics occurring simultaneously across a broad range of timescales, with implications for the transfer of sensory information to the cortex.
Imaging translational control by Argonaute with single-molecule resolution in live cells
Authors: Cialek Charlotte A. , Morisaki Tatsuya , Zhao Ning , Montgomery Taiowa A. , Stasevich Timothy J.
Published: May 2021

A major challenge to our understanding of translational control has been deconvolving the individual impact specific regulatory factors have on the complex dynamics of mRNA translation. MicroRNAs (miRNAs), for example, guide Argonaute and associated proteins to target mRNAs, where they direct gene silencing in multiple ways that are not well understood. To better deconvolve these dynamics, we have developed technology to directly visualize and quantify the impact of human Argonaute2 (Ago2) on the translation and subcellular localization of individual reporter mRNAs in living cells. We show that our combined translation and Ago2 tethering system reflects endogenous miRNA-mediated gene silencing. Using the system, we find that Ago2 association leads to progressive silencing of translation at individual mRNA. The timescale of silencing was similar to that of translation, consistent with a role for Ago2 in blocking translation initiation, leading to ribosome runoff. At early time points, we observed occasional brief bursts of translational activity at Ago2-tethered mRNAs undergoing silencing, suggesting that translational repression may initially be reversible. At later time points, Ago2-tethered mRNAs cluster and coalesce with endogenous P-bodies, where a translationally silent state is maintained. These results provide a framework for exploring miRNA-mediated gene regulation in live cells at the single-molecule level. Furthermore, our tethering-based, single-molecule reporter system will likely have wide-ranging application in studying general RNA-protein interactions.
Identification and characterization of human CD34+and CD34dim/-neutrophil-committed progenitors
Authors: Calzetti Federica , Finotti Giulia , Tamassia Nicola , Bianchetto-Aguilera Francisco , Castellucci Monica , Cavallini Chiara , Mattè Alessandro , Gasperini Sara , Benedetti Fabio , Bonifacio Massimiliano , Tecchio Cristina , Scapini Patrizia , Cassatella Marco A.
Published: May 2021

We report the identification of human CD66b−CD64dimCD115−neutrophil-committed progenitors within SSClowCD45dimCD34+and CD34dim/−bone marrow cells, that we named neutrophil myeloblast (NMs). CD34+and CD34dim/−NMs resulted as either CD45RA+or CD45RA−, with CD34+CD45RA−NMs found as selectively expanded in chronic-phase chronic myeloid leukemia patients. By scRNA-seq experiments, CD34+and CD34dim/−NMs were found to consist of combinations of four cell clusters, characterized by different maturation stages and distributed along two differentiation routes. Cell clusters were identified by neutrophil-specific gene profiles, one of them associated to an interferon-stimulated gene (ISG) signature, hence supporting recently identified expansions of mature neutrophil subsets expressing ISGs in blood of diseased individuals. Altogether, our data shed light on the very early phases of neutrophil ontogeny.
Prediction of Whole-Cell Transcriptional Response with Machine Learning
Authors: Eslami Mohammed , Borujeni Amin Espah , Doosthosseini Hamid , Vaughn Matthew , Eramian Hamed , Clowers Katie , Gordon D. Benjamin , Gaffney Niall , Weston Mark , Becker Diveena , Dorfan Yuval , Fonner John , Urrutia Joshua , Corbet Carolyn , Zheng George , Stubbs Joe , Cristofaro Alexander , Maschhoff Paul , Singer Jedediah , Voigt Christopher A , Yeung Enoch
Published: May 2021

Applications in synthetic and systems biology can benefit from measuring whole-cell response to biochemical perturbations. Execution of experiments to cover all possible combinations of perturbations is infeasible. In this paper, we present the host response model (HRM), a machine learning approach that takes the cell response to single perturbations as the input and predicts the whole cell transcriptional response to the combination of inducers. We find that the HRM is able to qualitatively predict the directionality of dysregulation to a combination of inducers with an accuracy of >90% using data from single inducers. We further find that the use of known prior, known cell regulatory networks doubles the predictive performance of the HRM (an R2from 0.3 to 0.65). This tool will significantly reduce the number of high-throughput sequencing experiments that need to be run to characterize the transcriptional impact of the combination of perturbations on the host.