bluuebunny/crossref_metadata_2025 huggingface dataset:
Automated speech detection in eco-acoustic data enables privacy protection and human disturbance quantification
Authors: Cretois Benjamin , Rosten Carolyn , Sethi Sarab S.
Published: February 2022

Eco-acoustic monitoring is increasingly being used to map biodiversity across huge scales, yet little thought is given to the privacy concerns and potential scientific value of inadvertently recorded human speech. Automated speech detection is possible using Voice Activity Detection (VAD) models, but existing approaches have been developed for indoor or urban use cases, rather than diverse natural soundscapes. In this study we used a data augmentation approach to create ecoVAD, a convolutional neural network designed for robust voice detection in eco-acoustic data. We performed playback experiments using speech samples from a woman, man, and child in two ecosystems in Børsa, Norway, and showed that ecoVAD was able to accurately detect voices at distances of up to 20m – at which point the speech was unintelligible. We compared ecoVAD with two existing VAD models and found that ecoVAD consistently outperformed the state-of-the-art (mean F1 scores: ecoVAD, 0.917; pyannote, 0.890; WebRTC VAD, 0.876). Using long-term passive recordings from a popular hiking location in Bymarka, Norway, we found that the frequency of speech detections was linked closely to peak traffic hours (using bus timings) demonstrating how VAD models can be used to quantify human activity with a fine temporal resolution. Anonymising audio data effectively using VAD models will allow eco-acoustic monitoring to continue to deliver invaluable ecological insight at scale, whilst minimising the risk of data misuse. Furthermore, using speech detections as a fine scale measure of human disturbance opens new possibilities for studying subtle human-wildlife interactions on the vast scales made possible by eco-acoustic monitoring technology.

Author Summary

Eco-acoustic monitoring (i.e., recording and analysing the sounds of an ecosystem) allows us to monitor biodiversity and ecological health on vast spatial and temporal scales. However, as networks of audio recorders are increasingly being deployed across the world by scientists and land managers, little thought has been given to the implications of inadvertently recorded human speech in these datasets. We developed a neural-network model to detect and remove speech from audio data, and showed it performed more accurately than existing state-of-the-art models when evaluated on eco-acoustic datasets. Furthermore, we showed frequency of voice detections could be used to monitor human pressure on an ecosystem. Our work paves the way for ethical and responsible processing of eco-acoustic data and provides a novel approach to studying human wildlife interactions on a large scale with high temporal resolution.
Rapid design and prototyping of biocatalytic virus-like particle nanoreactors
Authors: Esquirol Lygie , McNeale Donna , Douglas Trevor , Vickers Claudia E , Sainsbury Frank
Published: February 2022

Protein cages are attractive as molecular scaffolds for the fundamental study of enzymes and metabolons, and for the creation of biocatalytic nanoreactors for in vitro and in vivo use. Virus-like particles (VLPs) such as those derived from the P22 bacteriophage capsid protein make versatile self-assembling protein cages and can be used to encapsulate a broad range of protein cargos. In vivo encapsulation of enzymes within VLPs requires fusion to the coat protein or a scaffold protein. However, the expression level, stability and activity of cargo proteins can vary upon fusion. Moreover, it has been shown that molecular crowding of enzymes inside virus-like particles can affect their catalytic properties. Consequently, testing of numerous parameters is required for production of the most efficient nanoreactor for a given cargo enzyme. Here we present a set of acceptor vectors that provide a quick and efficient way to build, test and optimise cargo loading inside P22 virus-like particles. We prototyped the system using yellow fluorescent protein then applied it to mevalonate kinases, a key enzyme class in the industrially important terpene (isoprenoid) synthesis pathway. Different mevalonate kinases required considerably different approaches to deliver maximal encapsulation as well as optimal kinetic parameters, demonstrating the value of being able to rapidly access a variety of encapsulation strategies. The vector system described here provides an approach to optimise cargo enzyme behaviour in bespoke P22 nanoreactors. This will facilitate industrial applications as well as basic research on nanoreactor-cargo behaviour.

Abstract Figure
Sequence-dependent mechanochemical coupling of helicase translocation and unwinding at single-nucleotide resolution
Authors: Laszlo Andrew H. , Craig Jonathan M. , Gavrilov Momčilo , Tippana Ramreddy , Nova Ian C. , Huang Jesse R. , Kim Hwanhee C. , Abell Sarah J. , deCampos-Stairiker Mallory , Mount Jonathan W. , Bowman Jasmine L. , Baker Katherine S. , Higinbotham Hugh , Bobrovnikov Dmitriy , Ha Taekjip , Gundlach Jens H.
Published: February 2022

We used single-molecule nanopore tweezers (SPRNT) to resolve the millisecond single-nucleotide steps of Superfamily 1 helicase PcrA as it translocates on, or unwinds, several kb-long DNA molecules. We recorded over 2 million enzyme steps under various assisting and opposing forces in diverse ATP and ADP conditions to comprehensively explore the mechanochemistry of PcrA motion. Forces applied in SPRNT mimic forces and physical barriers PcrA experiences in vivo, such as when the helicase encounters bound proteins or duplex DNA; we show how PcrA’s kinetics change with such stimuli. SPRNT allows for direct association of the underlying DNA sequence with observed enzyme kinetics. Our data reveal that the underlying DNA sequence passing through the helicase strongly influences the kinetics during translocation and unwinding. Surprisingly, unwinding kinetics are not solely dominated by the base-pairs being unwound. Instead, the sequence of the single stranded DNA on which the PcrA walks determines much of the kinetics of unwinding.
Restriction of dietary protein in rats increases progressive-ratio motivation for protein
Authors: Chiacchierini Giulia , Naneix Fabien , Apergis-Schoute John , McCutcheon James E.
Published: February 2022

Low-protein diets can impact food intake and appetite, but it is not known if motivation for food is changed. In the present study, we used an operant behavioral task – the progressive ratio test – to assess whether motivation for different foods was affected when rats were maintained on a protein-restricted diet (REST, 5% protein diet) compared to non-restricted control rats (CON, 18% protein). Rats were tested either with nutritionally-balanced pellets (18.7% protein, Experiment 1) or protein-rich pellets (35% protein, Experiment 2) as reinforcers. Protein restriction increased breakpoint for protein-rich pellets, relative to CON rats, whereas no difference in breakpoint for nutritionally-balanced pellets was observed between groups. When given free access to either nutritionally-balanced pellets or protein-rich pellets, REST and CON rats did not differ in their intake. We also tested whether a previous history of protein restriction might affect present motivation for different types of food, by assessing breakpoint of previously REST animals that were subsequently put on standard maintenance chow (protein-repleted rats, REPL, Experiment 2). REPL rats did not show increased breakpoint, relative to their initial encounter with protein-rich pellets while they were protein-restricted. This study demonstrates that restriction of dietary protein induces a selective increased motivation for protein-rich food, a behavior that disappears once rats are not in need of protein.
Aging and white matter microstructure and macrostructure: a longitudinal multi-site diffusion MRI study of 1,184 participants
Authors: Schilling Kurt G , Archer Derek , Yeh Fang-Cheng , Rheault Francois , Cai Leon Y , Hansen Colin , Yang Qi , Ramdass Karthik , Shafer Andrea , Resnick Susan , Pechman Kimberly R. , Gifford Katherine A. , Hohman Timothy J. , Jefferson Angela , Anderson Adam W , Kang Hakmook , Landman Bennett A
Published: February 2022

Quantifying the microstructural and macrostructural geometrical features of the human brain’s connections is necessary for understanding normal aging and disease. Here, we examine brain white matter diffusion magnetic resonance imaging data from one cross-sectional and two longitudinal datasets totaling in 1184 subjects and 2236 sessions of people aged 50-97 years. Data was drawn from well-established cohorts, including the Baltimore Longitudinal Study of Aging dataset, Cambridge Centre for Ageing Neuroscience dataset, and the Vanderbilt Memory & Aging Project. Quantifying 4 microstructural features and, for the first time, 11 macrostructure-based features of volume, area, and length across 120 white matter pathways, we apply linear mixed effect modeling to investigate changes in pathway-specific features over time, and document large age associations within white matter. Conventional diffusion tensor microstructure indices are the most age-sensitive measures, with positive age associations for diffusivities and negative age associations with anisotropies, with similar patterns observed across all pathways. Similarly, pathway shape measures also change with age, with negative age associations for most length, surface area, and volume-based features. A particularly novel finding of this study is that while trends were homogeneous throughout the brain for microstructure features, macrostructural features demonstrated heterogeneity across pathways, whereby several projection, thalamic, and commissural tracts exhibited more decline with age compared to association and limbic tracts. The findings from this large-scale study provide a comprehensive overview of the age-related decline in white matter and demonstrate that macrostructural features may be more sensitive to heterogeneous white matter decline. Therefore, leveraging macrostructural features may be useful for studying aging and could have widespread implications for a variety of neurodegenerative disorders.
Dissecting Light Sensing and Metabolic Pathways on the Millimeter Scale in High-Altitude Modern Stromatolites
Authors: Alonso-Reyes Daniel Gonzalo , Galván Silvina , Irazoqui José Matías , Amadio Ariel , Tschoeke Diogo , Thompson Fabiano , Albarracín Virginia Helena , Farias María Eugenia
Published: February 2022

Modern non-lithifying stromatolites (STs) on the shore of the volcanic lake Socompa in the Puna are affected by several extreme conditions. Although STs were proposed as ecologic models for understanding stress response and resilience in microbial ecosystems constituting a window into the past, our knowledge of ST function is still nascent. The present study assesses for the first time light utilization and functional metabolic stratification of STs on a millimeter scale through shotgun metagenomics. In addition, a scanning-electron-microscopy approach was used to explore the community. Our results demonstrated that Bacteroidetes and Cyanobacteria play major roles as ST builders and primary producers to sustain a diverse community of heterotrophs. STs manifest a high occurrence of genes for the synthesis of UV-protecting pigments, the cryptochrome-photolyase family (CPF), and rhodopsins in the surface layers. Three different ecologic niches involving the use of light in energy production were defined. Calvin-Benson and Wood-Ljungdahl pathways were proposed as the main mechanisms for carbon fixation. Several genes account for the microelectrode chemical data and pigment measurements performed in previous publications. We also provide here an explanation for the vertical microbial mobility within the ST described previously. Finally, our study points to STs as ideal modern analogues of ancient STs.
A network-based approach for isolating the chronic inflammation gene signatures underlying complex diseases towards finding new treatment opportunities
Authors: Hickey Stephanie L , McKim Alexander , Mancuso Christopher A , Krishnan Arjun
Published: February 2022

Complex diseases are associated with a wide range of cellular, physiological, and clinical phenotypes. To advance our understanding of disease mechanisms and our ability to treat these diseases, it is critical to delineate the molecular basis and therapeutic avenues of specific disease phenotypes, especially those that are associated with multiple diseases. Inflammatory processes constitute one such prominent phenotype, being involved in a wide range of health problems including ischemic heart disease, stroke, cancer, diabetes mellitus, chronic kidney disease, non-alcoholic fatty liver disease, and autoimmune and neurodegenerative conditions. While hundreds of genes might play a role in the etiology of each of these diseases, isolating the genes involved in the specific phenotype (e.g., inflammation “component”) could help us understand the genes and pathways underlying this phenotype across diseases and predict potential drugs to target the phenotype. Here, we present a computational approach that integrates gene interaction networks, disease-/trait-gene associations, and drug-target information to accomplish this goal. We apply this approach to isolate gene signatures of complex diseases that correspond to chronic inflammation and prioritize drugs to reveal new therapeutic opportunities.
Clostridia isolated from helminth-colonized humans promote hatching of Trichuris muris
Authors: Sargsian Shushan , Chen Ze , Lee Soo Ching , Robertson Amicha , Sproch Julia , Devlin Joseph C. , Tee Mian Zi , Er Yi Xian , Copin Richard , Heguy Adriana , Pironti Alejandro , Torres Victor J. , Ruggles Kelly V. , Lim Yvonne A.L. , Loke P’ng , Cadwell Ken
Published: February 2022

Soil transmitted intestinal worms known as helminths colonize over 1.5 billion people worldwide. Although helminth colonization has been associated with altered composition of the gut microbiota, such as increases in Clostridia, individual species have not been isolated and characterized. Here, we isolated and sequenced the genome of 13 Clostridia from the Orang Asli, an indigenous population in Malaysia with high prevalence of helminth infections. Metagenomic analysis of 650 fecal samples from urban and rural Malaysians revealed higher prevalence and abundance of these isolates compared to individuals in the United States, with Peptostreptococcaceae family members displaying a specific association with helminth colonization. Remarkably, Peptostreptococcaceae isolated from the Orang Asli displayed superior capacity to induce hatching of eggs from the murine helminth Trichuris muris. These findings support a model in which helminths select for gut colonization of microbes that in turn support their life cycle by promoting egg hatching.
Temporal and sequential transcriptional dynamics define lineage shifts in corticogenesis
Authors: Mukhtar Tanzila , Breda Jeremie , Boareto Marcelo , Grobecker Pascal , Karimaddini Zahra , Grison Alice , Eschbach Katja , Chandrasekhar Ramakrishnan , Vermeul Swen , Okoniewski Michal , Pachkov Mikhail , Atanasoski Suzana , Beisel Christian , Iber Dagmar , van Nimwegen Erik , Taylor Verdon
Published: February 2022

The cerebral cortex contains billions of neurons, and their disorganization or misspecification leads to neurodevelopmental disorders. Understanding how the plethora of projection neuron subtypes are generated by cortical neural stem cells (NSCs) is a major challenge. Here, we focused on elucidating the transcriptional landscape of murine embryonic NSCs, basal progenitors (BPs) and newborn neurons (NBNs) throughout cortical development. We uncover dynamic shifts in transcriptional space over time, and heterogeneity within each progenitor population. We identified signature hallmarks of NSC, BP and NBN clusters, and predict active transcriptional nodes and networks that contribute to neural fate specification. We find that the expression of receptors, ligands and downstream pathway components is highly dynamic over time and throughout the lineage implying differential responsiveness to signals. Thus, we provide an expansive compendium of gene expression during cortical development that will be an invaluable resource for studying neural developmental processes and neurodevelopmental disorders.
TP53 knockout in MCF7 breast cancer cells induces sensitivity to fluoropyrimidine drugs
Authors: Warrington JM , Mehta S , Moy VF , Patel JR , Liu C , Rago AR , McTighe C , Delgado I , Banister CE , Gmeiner WH , Buckhaults PJ
Published: February 2022

TP53 mutations are present in all molecular subtypes of breast cancer and often correlate with decreased survival; however, few therapeutic options exist for patients with TP53-mutant breast cancers. To discover therapeutic strategies for these patients, we investigated the sensitivity of 129 FDA-approved chemotherapies to TP53-KO and TP53-WT MCF7 breast adenocarcinoma cells and found p53 loss to confer sensitivity to 5-fluorouracil (5-FU). We then treated the p53-null cells and isogenic controls with F10, a second-generation polymeric fluoropyrimidine, and found this preferential cytotoxicity of TP53-KO cells to be significantly magnified. F10 killing could only minimally be rescued by addition of exogenous uridine, whereas it was completely abrogated by addition of exogenous thymidine, suggesting DNA incorporation to be central to the cytotoxic mechanism of action. Furthermore, F10 killing of p53-null cells was persistent even in heterogeneous cellular mixtures more reflective of natural tumor evolution. Together, our results suggest F10 may widen the therapeutic window for TP53-mutant breast cancer by enhancing genotoxicity in cells resistant to apoptosis.
CaSee: A lightning transfer-learning model directly used to discriminate cancer/normal cells from scRNA-seq
Authors: Sh Yuan , Guo Caixia , Shi Fanghao , Jia Fei , Hu Zhiyuan , Zhang Xiuli
Published: February 2022

Single-cell RNA sequencing (scRNA-seq) is one of the most efficient technologies for human tumor research. However, data analysis is still faced with some technical challenges, especially the difficulty in efficiently and accurately discriminate cancer/normal cells in the scRNA-seq expression matrix. In this study, we developed a cancer/normal cell discrimination pipeline called pan-cancer seeker (CaSee) devoted to scRNA-seq expression matrix, which is based on the traditional high-quality pan-cancer bulk sequencing data using transfer learning. It is compatible with mainstream sequencings technology platforms, 10x Genomics Chromium, Smart-seq2, and Microwell-seq. Here, CaSee pipeline exhibited excellent performance in the multicenter data evaluation of 11 retrospective cohorts and one independent dataset, with an average discrimination accuracy of 96.69%. In general, the development of a deep-learning based, pan-cancer cell discrimination model, CaSee, to distinguish cancer cells from normal cells will be compelling to researchers working in the genomics, cancer, and single-cell fields.
PP1 inhibitor-2 promotes PP1γ positive regulation of synaptic transmission
Authors: Foley Karl , Altimimi Haider , Hou Hailong , Zhang Yu , McKee Cody , Papasergi-Scott Makaía M. , Yang Hongtian , Mayer Abigail , Ward Nancy , MacLean David M. , Nairn Angus C. , Stellwagen David , Xia Houhui
Published: February 2022

Inhibitor-2 (I-2) is a prototypic inhibitor of protein phosphatase-1 (PP1), a major serine-threonine phosphatase that regulates synaptic plasticity and learning and memory. Although I-2 is a potent inhibitor of PP1 in vitro, our previous work has elucidated that, in vivo, I-2 may act as a positive regulator of PP1. Here we show that I-2 and PP1γ, but not PP1α, positively regulate synaptic transmission in hippocampal neurons. Moreover, we demonstrated that I-2 enhances PP1γ interaction with its major synaptic scaffold, neurabin, by Förster resonance energy transfer (FRET)/Fluorescence lifetime imaging microscopy (FLIM) studies, while having a limited effect on PP1 autoinhibitory phosphorylation. Furthermore, our study indicates that the effect of I-2 on PP1 activity in vivo is dictated by I-2 threonine-72 phosphorylation. Our work thus demonstrates a molecular mechanism by which I-2 positively regulates PP1 function in synaptic transmission.
Structure, dynamics and assembly of the ankyrin complex on human red blood cell membrane
Authors: Xia Xian , Liu Shiheng , Zhou Z. Hong
Published: February 2022

The cytoskeleton of red blood cell (RBC) is anchored to cell membrane by the ankyrin complex. This complex is assembled during RBC genesis and comprises primarily band 3, protein 4.2 and ankyrin, whose mutations contribute to numerous human inherited diseases. High-resolution structures of the ankyrin complex have been long sought-after to understand its assembly and disease-causing mutations. Here, we analyzed native complexes on human RBC membrane by stepwise fractionation. Cryo-electron microscopy structures of nine band 3-associated complexes reveal that protein 4.2 stabilizes the cytoplasmic domain of band 3 dimer. In turn, the superhelix-shaped ankyrin binds to this protein 4.2 via ankyrin repeats (ARs) 6-13 and to another band 3 dimer via ARs 17-20, bridging two band 3 dimers in the ankyrin complex. Integration of these structures with both prior and our biochemical data supports a model of ankyrin complex assembly during erythropoiesis and identifies interactions essential for mechanical stability of RBC.
Low-cost sample preservation methods for high-throughput processing of rumen microbiomes
Authors: Budel Juliana C. C. , Hess Melanie K. , Bilton Timothy P. , Henry Hannah , Dodds Ken G. , Janssen Peter H. , McEwan John C. , Rowe Suzanne J.
Published: February 2022

Background

The use of rumen microbial community (RMC) profiles to predict methane emissions has driven interest in ruminal DNA preservation and extraction protocols that can be processed cheaply while also maintaining or improving DNA quality for RMC profiling. Our standard approach for preserving rumen samples, as defined in the Global Rumen Census (GRC), requires time-consuming pre-processing steps of freeze drying and grinding prior to international transportation and DNA extraction. This impedes researchers unable to access sufficient funding or infrastructure. To circumvent these pre-processing steps, we investigated three methods of preserving rumen samples for subsequent DNA extraction, based on existing lysis buffers Tris-NaCl-EDTA-SDS (TNx2) and guanidine hydrochloride (GHx2), or 100% ethanol.

Results

Rumen samples were collected via stomach intubation from 151 sheep at two time-points two weeks apart. Each sample was separated into four subsamples and preserved using the three preservation methods and the GRC method (n = 4×302). DNA was extracted and sequenced using Restriction Enzyme-Reduced Representation Sequencing to generate RMC profiles. Differences in DNA yield, quality and integrity, and sequencing metrics were observed across the methods (p < 0.0001). Ethanol exhibited poorer quality DNA (A260/A230 < 2) and more failed samples compared to the other methods. Samples preserved using the GRC method had smaller relative abundances in gram-negative genera Anaerovibrio, Bacteroides, Prevotella, Selenomonas, and Succiniclasticum, but larger relative abundances in the majority of 56 additional genera compared to TNx2 and GHx2. However, log10 relative abundances across all genera and time-points for TNx2 and GHx2 were on average consistent (R2 > 0.99) but slightly more variable compared to the GRC method. Relative abundances were moderately to highly correlated (0.68 ± 0.13) between methods for samples collected within a time-point, which was greater than the average correlation (0.17 ± 0.11) between time-points within a preservation method.

Conclusions

The two modified lysis buffers solutions (TNx2 and GHx2) proposed in this study were shown to be viable alternatives to the GRC method for RMC profiling in sheep. Use of these preservative solutions reduces cost and improves throughput associated with processing and sequencing ruminal samples. This development could significantly advance implementation of RMC profiles as a tool for breeding ruminant livestock.
Barrier-free liquid condensates of nanocatalysts as effective concentrators of catalysis
Authors: Bedi Silky , Kumar Gaurav , Rose S M , Rakshit Sabyasachi , Sinha Sharmistha
Published: February 2022

Molecular confinement of catalysts enhances the catalytic activity significantly. However, physicochemical barriers in traditional confinements restrict the free-passage of substrates/products. To obtain a barrier-free confinement of catalysts, here we explored the liquid-liquid phase separation. Using favourable ionic strength and crowding agents, we recruit the protein-inorganic-composites in phase separated liquid condensates from a solution. The phase separation propensity of these nanocomposites was seen to be independent of the native conformation of the component protein. Using standard catalytic oxidation-reduction reactions, we show that the close-proximity yet diffusive nature of catalysts in solution amplifies the homogeneous catalytic-efficiency of metal particles significantly. Overall, our work demonstrates the roadmap of using inorganic catalysts in homogeneous homogenous solution phase with amplified efficiency and longevity.
Sequence- and chemical specificity define the functional landscape of intrinsically disordered regions
Authors: Langstein-Skora Iris , Schmid Andrea , Huth Frauke , Shabani Drin , Spechtenhauser Lorenz , Likhodeeva Mariia , Kunert Franziska , Metzner Felix J. , Emenecker Ryan J. , Richardson Mary O.G. , Aftab Wasim , Götz Maximilian J. , Payer Sarah K. , Pietrantoni Niccoló , Sjeničić Valentina , Ravichandran Sakthi K. , Bartke Till , Hopfner Karl-Peter , Gerland Ulrich , Korber Philipp , Holehouse Alex S.
Published: February 2022

Intrinsically disordered protein regions (IDRs) pervasively engage in essential molecular functions, yet they are often poorly conserved as assessed by sequence alignment. To understand the seeming paradox of how sequence variability is compatible with persistent function, we examined the functional determinants for a poorly conserved but essential IDR. We show that IDR function depends on two distinct but related properties: sequence- and chemical specificity. While sequence-specificity works via linear binding motifs, chemical specificity reflects the sequence-encoded chemistry of multivalent interactions through amino acids across an IDR. Unexpectedly, a binding motif that is essential in the wild-type IDR can be removed if compensatory changes to the sequence chemistry are made, highlighting the orthogonality and interoperability of both properties and providing a much deeper sequence space compatible with function. Our results provide a general framework to understand the functional constraints on IDR sequence evolution.
Ecological Observations Based on Functional Gene Sequencing Are Sensitive to the Amplicon Processing Method
Authors: Cholet Fabien , Lisik Agata , Agogué Hélène , Ijaz Umer Z , Pineau Philippe , Lachaussée Nicolas , Smith Cindy J
Published: February 2022

Until recently, the de-facto method for short read-based amplicons reconstruction is a sequence similarity threshold approach (Operational taxonomic Units OTUs). This assumption was relaxed by shifting to Amplicon Sequencing Variants (ASVs) where distributions are fitted to abundance profiles of individual genes using a noise-error model. Whilst OTUs-based approach is still useful for 16SrRNA/18S rRNA regions, where typically 97-99% thresholds are used, their utility to functional genes is still debatable as there is no consensus on how to cluster the sequences together. Here, we compare OTUs- and ASVs-based reconstruction approaches as well as taxonomy assignment methods, Naïve Bayesian Classifier (NBC) and Bayesian Lowest Common Ancestor Algorithm (BLCA), using functional genes dataset from the microbial nitrogen-cycling community in the Brouage mudflat (France). A range of OTU similarity thresholds and ASV were used to compare amoA (AOA and AOB), nxrB, nirS, nirK and nrfA communities between differing sedimentary structures. We show that for AOA-amoA and nrfA, the use of ASV led to differences in the communities between sedimentary structures whereas the use of OTUs didn’t. Conversely, significant differences were detected when using OTU (97%) for AOB-amoA but not with ASV or OTUs at other similarity thresholds. Interestingly, conclusions drawn from the other three functional genes were consistent between amplicon reconstruction methods. We also show that, when the sequences in the reference-database are related to the environment in question, BLCA leads to more phylogenetically relevant classifications. However, when the reference database contains sequences more dissimilar to the ones retrieved, NBC helps obtain more information.

Importance

Several analysis pipelines are available to microbial ecologists to process amplicon sequencing data yet to-date, there is no consensus as to the most appropriate method, and it becomes more difficult for genes that encode a specific function (functional genes). Standardised approaches need to be adopted to increase reliability and reproducibility of environmental amplicon sequencing-based datasets. In this paper, we argue that the recently developed ASV approach offers a better opportunity to achieve such standardisation compared to OTUs for functional genes. We also propose a comprehensive framework for quality filtering of the sequencing reads based on protein sequence verification and merging.
Global Force-of-Infection Trends for Human Taenia solium Taeniasis/Cysticercosis
Authors: Dixon Matthew A. , Winskill Peter , Harrison Wendy E. , Whittaker Charles , Schmidt Veronika , Flórez Sánchez Astrid Carolina , Cucunubá Zulma M. , Edia-Asuke Agnes U. , Walker Martin , Basáñez Maria-Gloria
Published: February 2022

Infection by Taenia solium poses a major burden across endemic countries. The World Health Organization (WHO) 2021–2030 Neglected Tropical Diseases roadmap has proposed that 30% of endemic countries achieve intensified T. solium control in hyperendemic areas by 2030. Understanding geographical variation in age-prevalence profiles and force-of-infection (FoI) estimates will inform intervention designs across settings.

Human taeniasis (HTT) and human cysticercosis (HCC) age-prevalence data from 16 studies in Latin America, Africa and Asia were extracted through a systematic review. Catalytic models, incorporating diagnostic performance uncertainty, were fitted to the data using Bayesian methods, to estimate rates of antibody (Ab)-seroconversion, infection acquisition and Ab-seroreversion or infection loss. HCC FoI and Ab-seroreversion rates were also estimated across 23 departments in Colombia from 28,100 individuals.

Across settings, there was extensive variation in all-ages seroprevalence. Evidence for Ab- seroreversion or infection loss was found in most settings for both HTT and HCC and for HCC Ab- seroreversion in Colombia. The average duration until humans became Ab-seropositive/infected decreased as all-age (sero)prevalence increased. There was no clear relationship between the average duration humans remain Ab-seropositive and all-age seroprevalence.

Marked geographical heterogeneity in T. solium transmission rates indicate the need for setting- specific intervention strategies to achieve the WHO goals.
Predictive Machine Learning for Personalised Medicine in Major Depressive Disorder
Authors: Gountouna Viktoria-Eleni , Bermingham Mairead L. , Kuznetsova Ksenia , Muñoz Daniel Urda , Agakov Felix , Robson Siân E. , Meijsen Joeri J. , Campbell Archie , Hayward Caroline , Wigmore Eleanor M. , Clarke Toni-Kim , Fernandez Ana Maria , MacIntyre Donald J. , McKeigue Paul , Porteous David J. , Nicodemus Kristin K.
Published: February 2022

Depression is a common psychiatric disorder with substantial recurrence risk. Accurate prediction from easily collected data would aid in diagnosis, treatment and prevention. We used machine learning in the Generation Scotland cohort to predict lifetime risk of depression and, among cases, recurrent depression. Rank aggregation was used to combine results across ten different algorithms and identify highly predictive variables. The model containing all but the cardiometabolic predictors had the highest predictive ability on independent data. Rank aggregation produced a reduced set of predictors without decreasing predictive performance (lifetime: 20 out of 154 predictors and Receiver Operating Characteristic area under the curve (AUC)=0·84, recurrent: 10 out of 180 predictors and AUC=0·76). Here we develop a pipeline which leads to a small set of highly predictive variables. This information can be easily collected with a smartphone ‘application’ to help diagnosis and treatment, while longitudinal tracking may help patients in self-management.

Significance

Depression is the most common psychiatric disorder and a leading cause of disability worldwide. Patients are often diagnosed and treated by non-specialist clinicians who have limited time available to assess them. We present a novel methodology which allowed us to identify a small set of highly predictive variables for a diagnosis of depression, or recurrent depression in patients. This information can easily be collected using a tablet or smartphone application in the clinic to aid diagnosis.
Whole genome functional characterization of RE1 silencers using a modified massively parallel reporter assay
Authors: Mouri Kousuke , Dewey Hannah B , Castro Rodrigo , Berenzy Daniel , Kales Susan , Tewhey Ryan
Published: February 2022

Both upregulation and downregulation by cis-regulatory elements help establish precise gene expression. Our understanding of how elements repress transcriptional activity is far more limited than activating elements. To address this gap, we characterized RE1, a group of transcriptional silencers bound by REST, on a genome-wide scale using an modified massively parallel reporter assay (MPRAduo). MPRAduo empirically defined a minimal binding strength of REST required for silencing (REST m-value), above which multiple cofactors colocalize and act to directly silence transcription. We identified 1,500 human variants that alter RE1 silencing and found their effect sizes are predictable when they overlap with REST binding sites above the m-value. In addition, we demonstrate that non-canonical REST binding motifs exhibit silencer function only if they precisely align two half sites with specific spacer length. Our results show mechanistic insights into RE1 silencer which allows us to predict its activity and effect of variants on RE1, providing a paradigm for performing genome-wide functional characterization of transcription factors binding sites.
Antisense oligonucleotides targeting exon 11 are able to partially rescue the Neurofibromatosis Type 2 phenotype in vitro
Authors: Catasús N. , Rosas I. , Bonache S. , Negro A. , Torres-Martin M. , Plana A. , Salvador H. , Serra E. , Blanco I. , Castellanos E.
Published: February 2022

Neurofibromatosis type 2 (NF2) is an autosomal dominant condition caused by loss of function variants in the NF2 gene, which codes for the protein Merlin, and characterized by the development of multiple tumours of the nervous system. The clinical presentation of the disease is variable and related to the type of the inherited germline variant. Here, we tested if PMOs could be used to correct the splice signalling caused by variants at +/-13 within the intron-exon boundary region. Here we show that the PMOs designed for these variants do not constitute a therapeutic approach. Furthermore, we evaluated the use of phosphorodiamidate morpholino oligomers (PMOs) to reduce the severity of the effects of NF2 truncating variants with the aim of generating milder hypomorphic isoforms in vitro through the induction of the in-frame deletion of the exon-carrying variant. We were able to specifically induce the skipping of exons 4, 8 and 11 maintaining the NF2 gene reading frame at cDNA level. Only the skipping of exon 11 produced a hypomorphic Merlin (Merlin-e11), able to partially rescue the observed phenotype in primary fibroblast cultures from NF2 patients, being encouraging for the treatment of patients harbouring truncating variants located in exon 11.
Delayed viral vector mediated delivery of neurotrophin-3 improves skilled hindlimb function and stability after thoracic contusion in rats
Authors: Sydney-Smith Jared D. , Koltchev Alice M. , Moon Lawrence D. F. , Warren Philippa M.
Published: February 2022

It has been reported that intramuscular injection of an Adeno-associated viral vector serotype 1 (AAV1) encoding Neurotrophin-3 (NT3) into hindlimb muscles 24 hours after a severe T9 contusion in rats induced lumbar spinal neuroplasticity, partially restored locomotive function and reduced spasms during swimming. Here we investigated whether a targeted delivery of NT3 to lumbar and thoracic motor neurons 48 hours following a severe contusive injury aids locomotive recovery in rats. AAV1-NT3 was injected into the tibialis anterior, gastrocnemius and rectus abdominus muscles 48-hours following trauma, persistently elevating serum levels of the neurotrophin. NT3 improved trunk stability, accuracy of stepping during skilled locomotive tasks, and alternation of the hindlimbs during swimming, but it had no effect on gross locomotion function in the open field. The number of vGlut1+ (likely proprioceptive afferent) boutons on gastrocnemius α-motor neurons was increased after injury but normalised following NT3 treatment suggestive of a mechanism in which the functional effects may be mediated through proprioceptive feedback. Ex vivo MRI revealed substantial loss of grey and white matter at the lesion epicentre but no effect of delayed NT3 treatment to induce neuroprotection or prevent secondary damage. Spasms and hyperreflexia were not reliably induced in this severe injury model suggesting a more complex anatomical or physiological cause to their induction. We have shown that delayed intramuscular AAV-NT3 treatment can promote recovery in skilled stepping and coordinated swimming supporting a role for NT3 as a therapeutic strategy for spinal injuries potentially through modulation of somatosensory feedback.

Key Points

Targeted delivery of NT3 to hindlimb and trunk muscles at a clinically relevant 48h following a severe thoracic contusion aids fine locomotor control and synchronised movement.

NT3 mediated improvements in trunk stability, accuracy of stepping during skilled locomotive tasks, and alternation of the hindlimbs during swimming through the normalisation of vGlut1+ boutons on presumptive proprioceptive afferents innervating these muscles.

250kDyn thoracic contusion does not reliably result in measurable signs of spasticity.
Exponential magnetophoretic gradient for the direct isolation of basophils from whole blood in a microfluidic system
Authors: Castaño Nicolas , Kim Sungu , Martin Adrian M. , Galli Stephen J. , Nadeau Kari C. , Tang Sindy K.Y.
Published: February 2022

Despite their rarity in peripheral blood, basophils play important roles in allergic disorders and other diseases including sepsis and COVID-19. Existing basophil isolation methods require many manual steps and suffer from significant variability in purity and recovery. We report an integrated basophil isolation device (i-BID) in microfluidics for negative immunomagnetic selection of basophils directly from 100 μL of whole blood within 10 minutes. We use a simulation-driven pipeline to design a magnetic separation module to apply an exponentially increasing magnetic force to capture magnetically tagged non-basophils flowing through a microtubing sandwiched between magnetic flux concentrators sweeping across a Halbach array. The exponential profile captures non-basophils effectively while preventing their excessive initial buildup causing clogging. The i-BID isolates basophils with a mean purity of 93.9%±3.6% and recovery of 95.6%±3.4% without causing basophil degradation or unintentional activation. Our i-BID has the potential to enable basophil-based point-of-care diagnostics such as rapid allergy assessment.
Aspirin Synergizes with Regorafenib to Reduce Growth of Colorectal Cancer
Authors: Su Chang , Fennell Lochlan J , Bond Catherine E , Kane Alexandra M , Kerr Genevieve , Jardé Thierry , Micati Diana , Engel Rebekah M , Chan Wing Hei , Hlavca Sara , Archer Stuart , McMurrick Paul J , Hammerlindl Heinz , Lim Fayth , Salik Basit , McKeone Diane M , Hartel Gunter , Borowsky Jennifer , Ladwa Rahul , Leggett Barbara A , Schaider Helmut , Abud Helen E , Boyle Glen M , Burge Matthew E , Whitehall Vicki LJ
Published: February 2022

Purpose

Regorafenib is a multi-kinase inhibitor approved for refractory metastatic colorectal cancer. Previous studies have suggested that combining kinase inhibitors with aspirin may improve patient outcomes. We aimed to determine the effects of aspirin and regorafenib combination treatment in preclinical models of colorectal cancer.

Experimental Design

SW480, RKO and LIM1215 colorectal cancer cell lines were treated with aspirin and regorafenib to determine effects on proliferation and cytotoxicity. RNA sequencing and Western blotting were performed to explore underlying molecular effects. Aspirin and regorafenib combination treatment was also tested using organoids derived from three human colorectal cancer tissue specimens. For the in vivo study, SW480-derived tumors were established in athymic mice. Tumor volume was measured during treatment with aspirin and regorafenib, followed by immunohistochemical staining for markers of proliferation and apoptosis.

Results

Aspirin and regorafenib synergistically inhibited proliferation of colorectal cancer cell lines and patient-derived organoids, irrespective of KRAS or BRAF mutation status. This was associated with inhibition of the PI3K-Akt-mTOR pathway and activation of the AMPK pathway. Aspirin and regorafenib effectively inhibited growth of microsatellite stable KRAS-mutant SW480-derived tumors in vivo. Immunohistochemical staining for Ki67 and cleaved caspase 3 showed that combination treatment elicited a synergistic anti-proliferative effect, in addition to a pro-apoptotic effect that was driven by regorafenib.

Conclusions

Aspirin and regorafenib demonstrate synergistic anti-proliferative effects in preclinical models of colorectal cancer. This suggests that combining regorafenib with aspirin may be an improved treatment strategy for patients with refractory metastatic colorectal cancer.
Functional assessment of the ionotropic receptors IR25a and IR93a in predator perception in the freshwater crustaceanDaphniausing RNA interference
Authors: Graeve A , Huster J , Mayweg J , Fiedler R , Plaßmann J , Wahle P , Weiss LC
Published: February 2022

Acquiring environmental information is vital for organisms as it informs about the location of resources, mating partners, and predators. The freshwater crustaceanDaphniadetects predator specific chemical cues released by its predators and subsequently develops defensive morphological features that reduce the predation risk. The detection of such chemical information is generally processed via distinct chemoreceptors that are located on chemoreceptor cells. Lately an ancestral type of ionotropic receptors (IRs) has been identified in crustaceans. IRs and the putative co-receptors IR25a and IR93a are postulated to be involved in chemoreception However, functional roles have not been assessed. Here, using threeDaphniaspecies as model, we report that the two co-receptors are expressed within the chemosensory antennules and gene expression is increased with predator perception. Importantly, RNA interference mediated knock-down of the two IRs impedes species-specific defense expression in the threeDaphniaspecies. Our results suggest that (albeit not testing the enigmatic receptor protein directly), the reduction of two associated proteins has impaired the functional aggregation of the postulated chemoreceptor complex. This in turn has hampered the perception of environmentally relevant chemical cues resulting in a substantial reduction of defensive morphological features.
Structure of the vasopressin hormone-V2 receptor-β-arrestin1 ternary complex
Authors: Bous Julien , Fouillen Aurélien , Orcel Hélène , Trapani Stefano , Cong Xiaojing , Fontanel Simon , Saint-Paul Julie , Lai-Kee-Him Joséphine , Urbach Serge , Sibille Nathalie , Sounier Rémy , Granier Sébastien , Mouillac Bernard , Bron Patrick
Published: February 2022

Arrestins interact with G protein-coupled receptors (GPCRs) to stop G protein activation and to initiate key signaling pathways. Recent structural studies shed light on the molecular mechanisms involved in GPCR-arrestin coupling, but whether this process is conserved among GPCRs is poorly understood. Here, we report the cryo-electron microscopy active structure of the wild-type arginine-vasopressin V2 receptor (V2R) in complex with β-arrestin1. It reveals an atypical position of β-arrestin1 compared to previously described GPCR-arrestin assemblies, associated with an original V2R/β-arrestin1 interface involving all receptor intracellular loops. Phosphorylated sites of the V2R C-terminus are clearly identified and interact extensively with the β-arrestin1 N-lobe, in agreement with structural data obtained with chimeric or synthetic systems. Overall, these findings highlight a striking structural variability among GPCR-arrestin signaling complexes.
RpoN is required for a functional type III secretion system in Yersinia pseudotuberculosis
Authors: Mahmud A.K.M.F. , Navais R. , Nilsson K. , Choudhury R. , Avican K. , Fallman M.
Published: February 2022

Pathogenic bacteria use a broad range of virulence factors to thrive within their host. Yersinia pseudotuberculosis, a gram-negative enteropathogen in humans, utilises a Type III secretion system to overcome the host’s innate immune response. A global regulator previously shown to be essential for virulence in Y. pseudotuberculosis is RpoN. We show here that a strain lacking RpoN has a severely reduced capacity to secrete Yop effectors. This strain has a substantially reduced expression of genes encoding structural components of the secretion apparatus, the ysc operons, while expression of genes encoding effectors and translocators is less affected. We show that RpoN regulates a complex network, where one part is suggested to contribute to inducing Type III secretion via the enhancer-binding protein GlrR/RpoN regulon. By analogy, during inducement of Type III secretion, RpoN has a positive effect on expression of the sigma factor RpoE, which also is known to act downstream of GlrR. Interestingly, we found putative RpoE binding sites upstream of the ysc operons, and also a partial rescue of Yop secretion in the rpoN mutant strain by overexpression of RpoE. Our findings suggest that the RpoN-mediated effect on expression of type III genes involves a sigma factor hierarchy, where RpoN via RpoE contributes to inducing Type III secretion.