bluuebunny/crossref_metadata_2025 huggingface dataset:
Intracellular Expression of a Fluorogenic DNA Aptamer Using Retron Eco2
Authors: Vibhute Mahesh A. , Machatzke Corbin , Bigler Katrin , Krümpel Saskia , Summerer Daniel , Mutschler Hannes
Published: May 2024

DNA aptamers are short, single-stranded DNA molecules that bind specifically to a range of targets such as proteins, cells, and small molecules. Typically, they are utilized in the development of therapeutic agents, diagnostics, drug delivery systems, and biosensors. Although aptamers perform well in controlled extracellular environments, their intracellular use has been less explored due to challenges of expressing them in vivo. In this study, we employed the bacterial retron system Eco2, to express a DNA light-up aptamer inEscherichia coli. Both in vitro and in vivo assays confirm that structure-guided insertion of the aptamer domain into the non-coding region of the retron enables reverse transcription and folding of functional aptamer constructs in vivo. Notably, we find only a limited correlation between in vitro and in vivo aptamer performance, suggesting marked folding differences between the two environments. Our findings demonstrate that retrons can be used to effectively express short DNA aptamers within living cells, potentially broadening and optimizing their application in intracellular settings.
Calmodulin variants in schizophrenia patients display gain-of-function or loss-of-function effects
Authors: Jensen Helene Halkjær , Brohus Malene , Hussey John W. , Busuioc Ana-Octavia , Iversen Emil Drivsholm , Darki Faezeh , Nikolova Gabriela Dobromirova , Baisgaard Amalie Elton , Rohde Palle Duun , Holm Ida Elisabeth Gad , McQuillin Andrew , Moos Torben , Dick Ivy E. , Overgaard Michael Toft , Nyegaard Mette
Published: May 2024

Calmodulin acts as a vital calcium sensor in cells, crucial for relaying calcium signals to different protein partners. While rare missense variants in calmodulin are linked to cardiac arrhythmia, particularly long QT syndrome (LQTS), their role in schizophrenia remains unexplored. We investigated missense variants in the calmodulin-encoding genesCALM1-3in a large-scale sequencing effort involving 24,248 schizophrenia patients and 97,322 controls. Seven carriers were found among cases and twenty among controls. Notably, all schizophrenia variants affected the C-terminal lobe of the protein, compared to only five in controls, linking calmodulin C-lobe missense variants and schizophrenia risk (odds ratio 5.62,P=0.043). Functional analyses revealed two classes of calmodulin variants in schizophrenia: 1) loss-of-function variants that reduce calcium affinity and impair the interaction with voltage-gated calcium channel 1.2 (CaV1.2), akin to LQTS variants but with smaller effect size, and 2) gain-of-function variants that unexpectedly enhance calcium affinity with no impact on CaV1.2 gating. This study for the first time statistically and functionally links calmodulin missense variants to a neurological disorder, expanding the phenotypic spectrum of calmodulinopathies to include schizophrenia.

Significance

Neurons use calmodulin to monitor calcium signals and modulate hundreds of target proteins, thereby regulating key processes such as neuronal firing and memory and learning. Here, we link genetic variants in calmodulin to schizophrenia risk. Moreover, these genetic variants have divergent consequences for calmodulin protein function. Our results expand the current understanding of calmodulin variants, which are primarily reported in cardiac arrhythmia patients, and generally have a strong loss-of-function effect on the protein. We here provide the first identification and characterization of calmodulin variants in non-cardiac patients. This broadens our view of the physiological and functional consequences of human calmodulin variants and presents novel mechanistic entries to understanding the molecular mechanisms of schizophrenia.
The selective pressure induced by malaria vaccines onPlasmodium falciparum
Authors: Masserey Thiery , Lee Tamsin , Cavelan Aurélien , Neafsey Daniel E. , Malinga Josephine , Penny Melissa A
Published: May 2024

Antigens ofPlasmodium falciparumtargeted by most current and future vaccines are generally not conserved. There are limited studies estimating the risk of parasite selection with vaccines. We adapted an individual-based model of malaria to assess which conditions favour genotypes having some degree of vaccine resistance and estimate the impact of resistance spread on vaccine effectiveness. Even parasite genotypes with low degrees of vaccine resistance are likely to spread relatively quickly if vaccines are implemented in children and even faster in a broader population. However, only highly resistant genotypes could strongly reduce vaccine effectiveness. These results highlight that it is essential to understand the degree to which genotypes exhibit reduced sensitivity to vaccines and monitor genotype frequency and vaccine effectiveness along with vaccine deployment. If some genotypes exhibit reduced efficacy to vaccines, our results further suggest that polyvalent or combination vaccines should be considered to limit resistance spread.
Approximating R1 and R2: a quantitative approach to clinical weighted MRI
Authors: Moskovich Shachar , Shtangel Oshrat , Mezer Aviv A.
Published: May 2024

Weighted MRI images are widely used in clinical as well as open-source neuroimaging databases. Weighted images such as T1-weighted, T2-weighted, and proton density-weighted (T1w, T2w, and PDw, respectively) are used for evaluating the brain’s macrostructure; however, their values cannot be used for microstructural analysis, since they lack physical meaning. Quantitative MRI (qMRI) relaxation rate parameters (e.g., R1 and R2), and related relaxivity coefficients, do contain microstructural physical meaning.

Nevertheless, qMRI is rarely done in large-scale clinical databases.

Currently, the weighted images ratio T1w/T2w is used as a quantifier to approximate the brain’s microstructure. In this paper, we propose three additional quantifiers that approximate quantitative maps, which can help bring quantitative MRI to the clinic for easy use.

Following the signal equations and using simple mathematical operations, we combine the T1w, T2w, and PDw images to estimate the R1 and R2.

We find that two of these quantifiers (T1w/PDw and T1w/ln(T2w)) can serve as a semi-quantitative proxy for R1, and that (ln(T2w/PDw)) can approximate R2.

We find that this approach also can be applied to T2w scans taken from widely available DTI datasets. We tested these quantifiers on bothin vitrophantom andin vivohuman datasets. We found that the quantifiers accurately represent the quantitative parameters across datasets. Finally, we tested the quantifiers within a clinical context, and found that they retain tissue information across datasets. Our work provides a simple pipeline to enhance the usability and quantitative accuracy of MRI weighted images.
Geospatial patterns and predictors of neonatal mortality among HIV infected and non-infected mothers of rural Zambia: A comparative analysis of the 2018 Zambia Demographic and Health Survey
Authors: Shumba Samson , Tembo Deborah , Simwaka Miyanda , Musonda Nedah Chikonde , Nkwemu Chipo , Mayimbo Sebean
Published: May 2024

Neonatal mortality is the death of a live-born infant within the first 28 completed days of life. Neonatal mortality remains a major public health concern in most African countries, with the Sub-Saharan region being the highest contributor at 27 deaths per 1000 live births, accounting for 43% of the total number of deaths. Zambia still fares poorly in terms of neonatal mortality, ranking 162 out of 195 countries globally. The study aimed to investigate the spatial patterns and predictors of neonatal mortality in rural Zambia. This study utilized the national-level data from the Zambia Demographic and Health Survey (ZDHS) program, utilizing the survey dataset from 2018. Statistical analyses were conducted using the Rao – Scott Chi-square test to assess associations between dependent and independent variables. Additionally, a multilevel mixed effect logistic regression model was used to examine predictors of neonatal mortality. Geospatial patterns of neonatal mortality across Zambia’s ten provinces were investigated using Quantum Geographical Information System (QGIS) version 3.34.1 to generate a univariate choropleth map. Data analysis was performed using Stata version 14.2. The study findings indicate a higher mortality rate among HIV-infected mothers aged 45 to 49 (100%) compared to 95.21% among non-HIV-infected mothers. Neonatal mortality was notably elevated among neonates born to mothers with no education (40.78%) and those with HIV infection (54.56%). Employment status also influenced mortality rates, with employed non-HIV-infected mothers showing 36.35% mortality compared to 49.39% among unemployed HIV-infected mothers. Higher birth weights, particularly 4000 grams or higher, were associated with increased mortality for both HIV-infected (81.15%) and non-infected (21.00%) mothers. Multilevel logistic regression identified predictors of mortality, including maternal age (40 to 44 years), neonate’s sex (female), and maternal HIV status. Geospatial analysis revealed Eastern and North-Western provinces as hotspots for neonatal mortality among HIV-infected mothers, while Muchinga was a hotspot for non-HIV-infected mothers. This study examined neonatal mortality among HIV-infected and non-infected mothers in rural Zambia, uncovering predictors such as maternal age, neonate sex, birthweight, maternal HIV status, and community desired number of children. Regional variations in mortality underscore the necessity for targeted interventions. Strengthening healthcare infrastructure, community outreach, healthcare worker training, maternal education, and addressing regional differences are crucial for improving maternal and child health and meeting Sustainable Development Goals targets.
Accuracy and reliability of data extraction for systematic reviews using large language models: A protocol for a prospective study
Authors: Oami Takehiko , Okada Yohei , Nakada Taka-aki
Published: May 2024

Background

Systematic reviews require extensive time and effort to manually extract and synthesize data from numerous screened studies. This study aims to investigate the ability of large language models (LLMs) to automate data extraction with high accuracy and minimal bias, using clinical questions (CQs) of the Japanese Clinical Practice Guidelines for Management of Sepsis and Septic Shock (J-SSCG) 2024. the study will evaluate the accuracy of three LLMs and optimize their command prompts to enhance accuracy.

Methods

This prospective study will objectively evaluate the accuracy and reliability of the extracted data from selected literature in the systematic review process in J-SSCG 2024 using three LLMs (GPT-4 Turbo, Claude 3, and Gemini 1.5 Pro). Detailed assessment of errors will be determined according to the predefined criteria for further improvement. Additionally, the time to complete each task will be measured and compared among the three LLMs. Following the primary analysis, we will optimize the original command with integration of prompt engineering techniques in the secondary analysis.

Trial registration

This research is submitted with the University hospital medical information network clinical trial registry (UMIN-CTR) [UMIN000054461].

Conflicts of interest

All authors declare no conflicts of interest to have.
Influenza H5N1 and H1N1 viruses remain infectious in unpasteurized milk on milking machinery surfaces
Authors: Le Sage Valerie , Campbell A.J. , Reed Douglass , Duprex W Paul , Lakdawala Seema
Published: May 2024

Spillover of highly pathogenic avian H5N1 into the cattle population poses a risk to humans through the close contact with farm workers. High viral loads of influenza viruses in the unpasteurized milk of infected lactating cows has the potential to contaminate equipment within milking parlors and create fomites for transmission to dairy workers. Cattle H5N1 and human 2009 H1N1 pandemic influenza viruses were found to remain infectious on surfaces commonly found in milking equipment materials for a few hours. The data presented here provide a compelling case for the risk of contaminated surfaces generated during milking to facilitate transmission of H5N1 from cattle-to-cattle and to dairy farm workers.
Subclinical SARS-CoV-2 Infections and Endemic Human Coronavirus Immunity Shape SARS-CoV-2 Saliva Antibody Responses
Authors: Conner Tonia L. , Goguet Emilie , Haines-Hull Hannah , Segard Allison , Darcey Emily S. , Kobi Priscilla , Balogun Bolatito , Olsen Cara , Esposito Dominic , Jones Milissa , Burgess Timothy H. , O’Connell Robert J. , Broder Christopher C. , Saunders David , Pollett Simon , Laing Eric D. , Mitre Edward
Published: May 2024

This study characterized antibody responses induced by COVID-19 mRNA vaccination and SARS-CoV-2 infection in saliva. Utilizing multiplex microsphere-based immunoassays, we measured saliva anti-SARS-CoV-2 spike IgG, IgA, and secretory IgA in 1,224 saliva samples collected from healthcare workers in the Prospective Assessment of SARS-CoV-2 Seroconversion study between August of 2020 through December of 2022. By spring of 2022, most individuals had detectable spike-specific antibodies in saliva. Longitudinal measurements of saliva anti-SARS-CoV-2 nucleocapsid IgG revealed that most spike-specific IgA and secretory IgA detected in saliva was driven by subclinical and clinically-evident infections, rather than by vaccination alone. In contrast, saliva anti-SARS-CoV-2 spike IgG was strongly induced by vaccination and exhibited improved durability with hybrid immunity. Baseline levels of saliva antibodies to the endemic human coronaviruses positively correlated with post-vaccination anti-SARS-CoV-2 spike IgG levels. This study provides insights for development of vaccines that generate mucosal antibodies to respiratory pathogens.

HIGHLIGHTS

Saliva anti-spike antibodies were present in > 90% of participants by spring 2022

Saliva anti-spike IgA was driven by subclinical and clinically evident infections

COVID-19 mRNA vaccination alone was a weak inducer of saliva IgA antibodies

HCoV immunity correlates with post-vaccine anti-spike saliva antibody levels
Discordance between a deep learning model and clinical-grade variant pathogenicity classification in a rare disease cohort
Authors: Kong Sek Won , Lee In-Hee , Collen Lauren V. , Manrai Arjun K. , Snapper Scott B. , Mandl Kenneth D.
Published: May 2024

Genetic testing has become an essential component in the diagnosis and management of a wide range of clinical conditions, from cancer to developmental disorders, especially in rare Mendelian diseases. Efforts to identify rare phenotype-associated variants have predominantly focused on protein-truncating variants, while the interpretation of missense variants presents a considerable challenge. Deep learning algorithms excel in various applications across biomedical tasks1,2, yet accurately distinguishing between pathogenic and benign genetic variants remains an elusive goal3-5. Specifically, even the most sophisticated models encounter difficulties in accurately assessing the pathogenicity of missense variants of uncertain significance (VUS). Our investigation of AlphaMissense (AM)5, the latest iteration of deep learning methods for predicting the potential functional impact of missense variants and assessing gene essentiality, reveals important limitations in its ability to identify pathogenic missense variants within a rare disease cohort. Indeed, AM struggles to accurately assess the pathogenicity of variants in intrinsically disordered regions (IDRs), leading to unreliable gene-level essentiality scores for certain genes containing IDRs. This limitation highlights the challenges in applying AM faces in the context of clinical genetics6.
Optimized Time-segmented Acquisition Expands Peptide and Protein Identification in TIMS-TOF Pro Mass Spectrometry
Authors: Zhang Huoming , Bensaddek Dalila
Published: May 2024

We introduce here a novel approach, termed time-segmented acquisition (Seg), to enhance the identification of peptides and proteins in trapped ion mobility spectrometry (TIMS)-Time of flight (TOF) mass spectrometry. Our method exploits the positive correlation between ion mobility values and liquid chromatography (LC) retention time to improve ion separation and resolution. By dividing the LC retention time into multiple segments and applying a segment-specific narrower ion mobility range within the TIMS tunnel, we achieved better separation and higher resolution of ion mobility. This resulted in a substantial increase in peptide identification. In comparison to conventional TIMS methods, which typically scan a static ion mobility range (either from 0.6 to 1.6 or from 0.85 to 1.3), the Seg method significantly enhances the identification of peptides, proteins and average sequence coverage per protein. These findings highlight the potential of the Seg method in expanding the capabilities of TIMS-TOF mass spectrometry, especially for peptide-focused analysis such as post-translational modifications and peptidomics.
reCRAC: A Stringent Method for Precise Mapping of Protein-RNA Interactions in Yeast
Authors: Ristova Michaela , Shchepachev Vadim , Tollervey David
Published: May 2024

Intricate interactions between RNA-binding proteins (RBPs) and RNA play pivotal roles in cellular homeostasis, impacting a spectrum of biological processes vital for survival. UV crosslinking methods to study protein-RNA interactions have been instrumental in elucidating their interactions but can be limited by degradation of target proteins during the process, low signal-to-noise ratios, and non-specific interactions. Addressing these limitations, we describe reCRAC (reverse CRAC), a novel adaptation of the CRAC (crosslinking and analysis of cDNA) technique, optimized for yeastSaccharomyces cerevisiae. Like CRAC, reCRAC applies tandem affinity purification to yield highly enriched protein preparations. However, reCRAC is redesigned to work with unstable proteins. This is achieved by lysing the cells directly into highly denaturing buffer conditions, followed by stringent purification steps. The reCRAC method was successfully applied to the easily degraded yeast protein Pin4, allowing identification of precise binding sites at base-pair resolution with greatly reduced target protein degradation and improved signal-to-noise ratios.
Loss of p21-activated kinase 4 (PAK4) suppresses pancreatic tumor progression and metastasis through regulating E-cadherin
Authors: Kim Kui-Jin , Nam Milang , Na Hee Young , Sung Ji Hea , Park Bo-Ram , Hwang Sung-Hyun , Park Woochan , Seo Jeongmin , Kang Minsu , Jung Eun Hee , Kim Sang-A , Suh Koung Jin , Lee Ji Yun , Kim Ji-Won , Kim Se Hyun , Lee Jeong-Ok , Kim Yu Jung , Lee Keun-Wook , Kim Jee Hyun , Bang Soo-Mee , Wainberg Zev , Kim Jin Won
Published: May 2024

Pancreatic ductal adenocarcinoma (PDAC) is characterized by a poor prognosis with early and frequent metastasis. While p21-activated kinase 4 (PAK4) has been implicated in cell migration, and invasion, the molecular mechanisms in PDAC remain unknown. In this study, we found that PAK4 overexpression was correlated with poor survival in PDAC patients through analysis of TCGA data. PAK4-amplified PDAC cells showed enhanced mobility in contrast with wild-type. PAK4 knockdown in PAK4 amplified cells inhibited cell migration, invasion, and displacement by increased and stabilized E-cadherin, which was attributed to decreased activity of Cdc42. PAK4 knock-in in PAK4 wild-type models enhanced cell migration, invasion, and displacement by reduced E-cadherin through elevated Cdc42 activity. PAK4 bounded to E-cadherin, Cdc42, and p120ctn in immunoprecipitation. In confocal imaging, the colocalization of PAK4, E-cadherin, p120ctn, and Cdc42 was also identified. In an orthotopic PDAC mouse model, PAK4 knockdown decreased primary tumor size and occurrence of malignant ascites by activation of E-cadherin. Notably, in patients tissue specimens, inverse correlation on expression of PAK4 and E-cadherin were also shown. In conclusion, our study highlights that PAK4 promotes invasive and metastatic behavior by regulating E-cadherin in PDAC. PAK4 could be a potential therapeutic target for PDAC patients.
Spatial Profiling of Metals through Matrix-Assisted Laser Desorption Ionization Mass Spectrometry Imaging
Authors: Stopka Sylwia A. , Bodineau Clément , Baquer Gerard , Lin Jia-Ren , Hossain Md Amin , Regan Michael S. , Ruiz Daniela , Fletcher Stecia-Marie , Pourquié Olivier , Lutsenko Svetlana , Payne Connor N. , Agar Jeffrey N. , Mazitschek Ralph , McDannold Nathan J. , Sorger Peter K. , Santagata Sandro , Agar Nathalie Y. R.
Published: May 2024

The spatial-omic analysis of biomolecules such as nucleic acids, lipids, metabolites, and proteins is advancing the study of biological systems and processes in a physio-pathological context. Here, we describe an innovative matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI MSI) method to detect metals within biological tissues using instrumentation that is widely available in research and clinical laboratories. We characterize the spatial distribution of metals in diverse settings including mouse embryogenesis, genetic disorders leading to abnormal metal accumulation, and preclinical testing for improved platinum-based chemotherapy delivery through focused ultrasound across the blood-brain barrier. Spatial metal profiling will advance research studies and the clinical analysis of metal-related diseases, enabling more precise use of metal-based therapies and advances in diverse scientific fields beyond biomedicine.

One-Sentence Summary

Spatial metallomic profiling maps native metals or those coordinated to xenobiotics, antibodies, and biomolecules in tissues.
Tau pathology leads to lonely non-traveling slow waves that mediate human memory impairment
Authors: Sharon Omer , Chen Xi , Dude Jason , Shah Vyoma D. , Ju Yo-El S. , Jagust Willam J. , Walker Matthew P
Published: May 2024

Memory markedly declines with age, exaggerated in those with Alzheimer’s disease, yet the mechanisms are still not resolved. Here, we show that frontal lobe tau pathology in humans leads to impaired en masse unity and cortical traveling propagation of NREM slow waves, consequentially impairing memory retention. We elucidate these findings using PET tau brain imaging, and then replicate and extend them using AD pathology markers derived from lumbar puncture CSF in an independent clinical cohort. Thus, tau-associated memory deficits are not wholly direct, but indirectly mediated through consequential “lonely”, non-traveling slow-wave events.
Gene deletion as a possible strategy adopted by New WorldLeishmania infantumto maximize geographic dispersion
Authors: Florêncio Monique , Chagas Marne Coimbra , Guimarães-Costa Anderson , Oliveira Jullyanna , Waclawiak Ingrid , Oliveira Thamara K. F. , Saraiva Elvira Maria , Freitas-Mesquita Anita Leocadio , Meyer-Fernandes José Roberto , Aragão-Farias Laura , Trindade Camilly Enes , Escobar Patricia Cuervo , Nascimento Renata Azevedo do , Moreira Otacilio C. , Ribeiro-Gomes Flávia Lima , Traub-Csekö Yara M. , Telleria Erich Loza , Vaselek Slavica , Leštinová Tereza , Volf Petr , Späth Gerald F. , Cupolillo Elisa , Boité Mariana C.
Published: May 2024

Background

The present study investigates implications of a sub-chromosomal deletion inLeishmania infantumstrains, the causative agent of American Visceral Leishmaniasis (AVL). Primarily found in New World strains, the deletion leads to the absence of the ecto-3’-nucleotidase/nuclease enzyme (3’NU/NT), impacting parasite virulence, pathogenicity, and drug susceptibility. The potential factors favoring prevalence and the widespread geographic distribution of these deleted mutant parasites (DEL) in the New World (NW) are discussed under the generated data.

Methods

We conducted phenotypic analyses of the parasites showing the sub- chromosomal deletion by applyingin vitroassays of 3’NU/NT activity, metacyclic enrichment, and relative quantitation of transcripts abundance on axenic parasites. We further performed experimental infections in bothin vitroandin vivomodels of vertebrate and invertebrate hosts using geographically diverse mutant field isolates.

Results

Virulence assays, poorer ability to survive neutrophil traps (NETs) and murine model infection revealed reduced pathogenicity in vertebrate hosts by the DEL strains. Conversely, these parasites exhibit enhanced metacyclogenesis and colonization rates in sand flies, potentially facilitating transmission. This combination may represent a more efficient way to maintain and disperse the transmission cycle of DEL strains.

Conclusions

Phenotypic assessments reveal altered parasite fitness, with enhanced transmissibility at the population level. Reduced susceptibility of DEL strains to miltefosine, a key drug in VL treatment, further complicates control efforts. Our study underscores the importance of typing parasite genomes for surveillance and control and proposes the sub-chromosomal deletion as a molecular marker in AVL management.
Cell-type-specific effects of autism-associated chromosome 15q11.2-13.1 duplications in human brain
Authors: Dias Caroline , Mo Alisa , Cai Chunhui , Sun Liang , Cabral Kristen , Brownstein Catherine A. , Rockowitz Shira , Walsh Christopher A.
Published: May 2024

Recurrent copy number variation represents one of the most well-established genetic drivers in neurodevelopmental disorders, including autism spectrum disorder (ASD). Duplication of 15q11.2-13.1 (dup15q) is a well-described neurodevelopmental syndrome that increases the risk of ASD by over 40-fold. However, the effects of this duplication on gene expression and chromatin accessibility in specific cell types in the human brain remain unknown. To identify the cell-type-specific transcriptional and epigenetic effects of dup15q in the human frontal cortex we conducted single-nucleus RNA-sequencing and multi-omic sequencing on dup15q cases (n=6) as well as non-dup15q ASD (n=7) and neurotypical controls (n=7). Cell-type-specific differential expression analysis identified significantly regulated genes, critical biological pathways, and differentially accessible genomic regions. Although there was overall increased gene expression across the duplicated genomic region, cellular identity represented an important factor mediating gene expression changes. Neuronal subtypes, showed greater upregulation of gene expression across a critical region within the duplication as compared to other cell types. Genes within the duplicated region that had high baseline expression in control individuals showed only modest changes in dup15q, regardless of cell type. Of note, dup15q and ASD had largely distinct signatures of chromatin accessibility, but shared the majority of transcriptional regulatory motifs, suggesting convergent biological pathways. However, the transcriptional binding factor motifs implicated in each condition implicated distinct biological mechanisms; neuronal JUN/FOS networks in ASD vs. an inflammatory transcriptional network in dup15q microglia. This work provides a cell-type-specific analysis of how dup15q changes gene expression and chromatin accessibility in the human brain and finds evidence of marked cell-type-specific effects of this genetic driver. These findings have implications for guiding therapeutic development in dup15q syndrome, as well as understanding the functional effects CNVs more broadly in neurodevelopmental disorders.
VesselBoost: A Python Toolbox for Small Blood Vessel Segmentation in Human Magnetic Resonance Angiography Data
Authors: Xu Marshall , Ribeiro Fernanda L. , Barth Markus , Bernier Michaël , Bollmann Steffen , Chatterjee Soumick , Cognolato Francesco , Gulban Omer Faruk , Itkyal Vaibhavi , Liu Siyu , Mattern Hendrik , Polimeni Jonathan R. , Shaw Thomas B. , Speck Oliver , Bollmann Saskia
Published: May 2024

Magnetic resonance angiography (MRA) performed at ultra-high magnetic field provides a unique opportunity to study the arteries of the living human brain at the mesoscopic level. From this, we can gain new insights into the brain’s blood supply and vascular disease affecting small vessels. However, for quantitative characterization and precise representation of human angioarchitecture to, for example, inform blood-flow simulations, detailed segmentations of the smallest vessels are required. Given the success of deep learning-based methods in many segmentation tasks, we here explore their application to high-resolution MRA data, and address the difficulty of obtaining large data sets of correctly and comprehensively labelled data. We introduceVesselBoost, a vessel segmentation package, which utilizes deep learning and imperfect training labels for accurate vasculature segmentation. Combined with an innovative data augmentation technique, which leverages the resemblance of vascular structures,VesselBoostenables detailed vascular segmentations.
A single amino acid substitution in the Borna disease virus glycoprotein enhances the infectivity titer of vesicular stomatitis virus pseudotyped virus by altering membrane fusion activity
Authors: Akiba Yusa , Matsugo Hiromichi , Kanda Takehiro , Sakai Modoka , Makino Akiko , Tomonaga Keizo
Published: May 2024

Borna disease virus 1 (BoDV-1) causes acute fatal encephalitis in mammals, including humans. Despite its importance, research on BoDV-1 cell entry has been hindered by low viral particle production in cells and the lack of cytopathic effects. To address these issues, we developed a method to efficiently produce vesicular stomatitis virus (VSV) pseudotyped with glycoprotein (G) of the genusOrthobornavirus, including BoDV-1. We discovered that optimal G expression is required to obtain a high infectivity titer of the VSV pseudotyped virus. Remarkably, the infectivity of the VSV pseudotyped virus with G from the BoDV-1 strain huP2br was significantly higher than that of the VSV pseudotyped virus with G from the He/80 strain. Mutational analysis demonstrated that the BoDV-1-G residue 307 determines the infectivity titer of VSV pseudotyped with BoDV-1-G (VSV-BoDV-1-G). A cell‒cell fusion assay indicated that this residue plays a pivotal role in membrane fusion, thus suggesting that high membrane fusion activity and a broad pH range for membrane fusion are crucial for achieving a high infectivity titer of VSV-BoDV-1-G. This finding may be extended to increase the infectivity titer of VSV pseudotyped virus with other orthobornavirus G. Our study also contributes to identifying functional domains of BoDV-1-G and provides insight into G-mediated cell entry.
Deciphering functional tumor-immune crosstalk through highly multiplexed imaging and deep visual proteomics
Authors: Zheng Xiang , Mund Andreas , Mann Matthias
Published: May 2024

Deciphering the intricate tumor-immune interactions within the microenvironment is crucial for advancing cancer immunotherapy. Here, we developed a novel approach integrating highly multiplexed imaging, laser microdissection, and deep visual proteomics (DVP) to spatially profile the proteomes of 21 distinct cell populations in human colorectal and tonsil cancers with high sensitivity. We selected colorectal tumor as an example of a cold tumor, uncovering an immunosuppressive macrophage barrier impeding T cell infiltration and regionally altering lymphocyte proteomes. Spatial proteomic analysis revealed distinct functional states of T cells in different tumor compartments. In tonsil cancer - a hot tumor, we identified significant heterogeneity of cell proteomes influenced by proximity to cytotoxic T cell subtypes. Tumor-infiltrating T cells exhibited metabolic adaptations and enhanced stress resilience, enabling migration into hypoxic regions. Our spatially-resolved, highly multiplexed strategy deciphers the complex cellular interplay within the tumor microenvironment, with implications for identifying new immunotherapy targets and signatures.

Highlights

Novel approach integrates multiplexed imaging and deep proteomics to profile the tumor microenvironment

Identified 21 distinct cell populations and thousands of proteins from single cell types

Macrophage barrier impedes T cell tumor infiltration and T cells adapt to hypoxia

Spatial insights may guide new immunotherapy targets and personalized oncology
Bread and hummus: trait connectance and correlation pleiades in grain crops
Authors: Sadras Victor O.
Published: May 2024

Phenotypic integration has been investigated from multiple perspectives. From a developmental perspective, connectance has been defined as the level of linkage between traits. Correlation pleiades,i.e., correlations between some traits and, simultaneously, lack of correlations between these and other traits have been interpreted as the independence of certain developmental processes with respect to other processes within the organism, and as the outcome from the discrepancy between the agencies participating in the formation of the trait and the selective forces influencing its function. Here, I use two published data sets to test the variation in connectance with both trait and genotype and the existence and meaning of correlation pleiades in wheat and chickpea. Connectance varied from 0.09 to 4.2 in wheat and from 0.06 to 22.8 in chickpea, and cluster analyses revealed correlation pleiades. The frequency distribution of connectance conformed to a power law with similar slopes = −1.665 ± 0.222 for wheat and −1.555 ± 0.126 for chickpea, consistent with developmental self-organisation. Connectance was lower for traits with higher heritability such as seed weight, which together with the negative association between heritability and phenotypic plasticity completes a relational triangle: high connectance ⇔ low heritability ⇔ high phenotypic plasticity.
Ca2+Regulation of Myosin II and Myosin VI during Rupture of theShigella-Containing Vacuole
Authors: Bonnet Mariette , Sun Chun Hui , Enninga Jost , Houdusse Anne , Tran Van Nhieu Guy
Published: May 2024

Shigella, the causative agent of bacillary dysentery, invades epithelial cells to colonize the intestinal mucosa. Following invasion,Shigellais enclosed in a vacuole that needs to rupture for bacterial intra-cytosolic replication. We show here that rupture of theShigellavacuole requires Ca2+influx leading to long lasting local Ca2+increases that regulate actin dynamics affecting theShigellavacuole integrity. These Ca2+increases promote vacuolar rupture by activating myosin II associated with actin filaments in membrane ruffles distant from the vacuole, while tethering myosin VI at the actin coat-surrounded vacuole. Ca2+depletion and myosin II inhibition impair formation of the actin coat and vacuole rupture. Inhibition of myosin VI also delays rupture of vacuoles but lead to their tumbling. These findings highlight a role for Ca2+in coordinating actin–based forces and constraints during early rupture steps of bacterial vacuole, that pull on vacuolar membranes and tether them to the actin cortex via myosin II and VI, respectively, a process relevant to intracellular pathogen and endomembrane trafficking.
Influenza A virus circumvents the innate immune response through the sequestration of double-stranded RNA
Authors: Nakano Masahiro , Miyamoto Sho , Ohnishi Chiho , Nogami Chiharu , Hirose Nanami , Fujita-Fujiharu Yoko , Muramoto Yukiko , Noda Takeshi
Published: May 2024

Double-stranded RNA (dsRNA), which induces an innate immune response against viral infections, is rarely detected in influenza A virus (IAV)-infected cells. Nevertheless, we previously reported that the IAV ribonucleoprotein complex (vRNP) generates looped dsRNAs during RNA synthesisin vitro. This finding suggests that IAV possesses a specific mechanism for sequestering dsRNA within infected cells, thereby enabling viral evasion of the innate immune response. Here, we found that dsRNAs were produced in infected cells lacking the expression of viral non-structural protein 1 (NS1) and nuclear export protein (NEP), both encoded by the same RNA segment. Interestingly, NS1 molecules masked the entire looped dsRNA generated by vRNP, implying a potential role for NS1 in segregating viral dsRNA from cytoplasmic dsRNA sensors, including retinoic acid inducible gene-I (RIG-I). Furthermore, dsRNAs were sequestered within the nucleus of infected cells due to the absence of NEP, while their translocation to the cytoplasm occurred only upon NEP expression. Notably, the cytoplasmic translocation of dsRNA triggered the innate immune response in an RIG-I-dependent manner. These findings highlight IAV’s distinctive strategy for circumventing innate immunity by sequestration of dsRNAs.

Author summary

It is widely recognized that double-stranded RNA (dsRNA) produced during viral infection triggers an innate immune response. However, the influenza A virus (IAV) has been thought to rarely produce dsRNA within infected cells. Here, we confirmed the limited dsRNA production in the nucleus of IAV-infected cells and found that the cells lacked expressions of viral non-structural protein 1 (NS1) and nuclear export protein (NEP), both derived from a single RNA segment. High-speed atomic force microscopy demonstrated that NS1 entirely concealed dsRNA produced by the viral ribonucleoprotein complexes, thereby segregating it from cytoplasmic dsRNA sensors that trigger the innate immune response. Interestingly, NEP expression caused cytoplasmic translocation of dsRNA, resulting in the nuclear translocation of interferon regulatory factor 3, which initiates an innate immune response. Collectively, our findings suggest that IAV employs a sophisticated strategy to circumvent the innate immune system, wherein the expressions of NS1 and NEP exert considerable influence.
Neural tracking of the speech envelope predicts binaural unmasking
Authors: Dieudonné Benjamin , Decruy Lien , Vanthornhout Jonas
Published: May 2024

Binaural unmasking is the remarkable phenomenon that it is substantially easier to detect a signal in noise, when the interaural parameters of the signal are different from those of the noise – a mechanism that comes in handy in so-called cocktail party scenarios. In this study, we investigated the effect of binaural unmasking on neural tracking of the speech envelope. We measured EEG in 8 participants who listened to speech in noise at a fixed signal-to-noise ratio (−12 dB or −9 dB, depending on the speech material), in two conditions: one where speech and noise had the same interaural phase difference (both speech and noise having an opposite waveform across ears,SπNπ), and one where the interaural phase difference of the speech was different from that of the noise (only the speech having an opposite waveform across ears,SπN0). We measured a clear benefit of binaural unmasking in behavioral speech understanding scores, accompanied with increased neural tracking of the speech envelope. Moreover, analyzing the temporal response functions revealed that binaural unmasking also resulted in decreased peak latencies and increased peak amplitudes. Our results are consistent with previous research using auditory evoked potentials and steady-state responses to quantify binaural unmasking at cortical levels. Moreover, they confirm that neural tracking of speech is modulated by speech understanding, even if the acoustic signal-to-noise ratio is kept constant.

Significance Statement

Binaural unmasking in an important contributing factor to speech understanding in noisy environments. This is the first time that this benefit in speech understanding is measured objectively with EEG. From a clinical perspective, these results could enable the evalu-ation of binaural speech understanding mechanisms in populations for whom behavioral measures are difficult to obtain, such as young children or people with cognitive impair-ment. Moreover, behavioral research has shown that many pathologies (e.g., asymmetric hearing loss, auditory neuropathy, and age-related deficits) are more sensitive to tests that contain a binaural processing component. Our paradigm could enable the detection of such pathologies with an objective approach using neural tracking of speech.
SARS-CoV-2 sequencing artifacts associated with targeted PCR enrichment and read mapping
Authors: Ellegaard Kirsten Maren , Gunalan Vithiagaran , Sieber Raphael , Baig Sharmin Jamshid , Larsen Nicolai Balle , Bennedbæk Marc , Bybjerg-Grauholm Jonas , Escobar-Herrera Leandro Andrés , Hansen Tobias Nikolaj Gress , Thorsen Theis Hass , Krusager Anders , Aasbjerg Gitte Nygaard , Al-Tamimi Nour Saad , Westergaard Casper , Svarrer Christina Wiid , Rasmussen Morten , Stegger Marc
Published: May 2024

Protocols and pipelines for SARS-CoV-2 genome sequencing were rapidly established when the COVID-19 outbreak was declared a pandemic. The most widely used approach for sequencing SARS-CoV-2 includes targeted enrichment by PCR, followed by shotgun sequencing and reference-based genome assembly. As the continued surveillance of SARS-CoV-2 worldwide is transitioning towards a lower level of intensity, it is timely to re-visit the sequencing protocols and pipelines established during the acute phase of the pandemic. In the current study, we have investigated the impact of primer scheme and reference genome choice by sequencing samples with multiple primer schemes (Artic V3, V4.1 and V5.3.2) and re-processing reads with multiple reference genomes. We have also analysed the temporal development in ambiguous base calls during the emergence of the BA.2.86.x variant. We found that the primers used for targeted enrichment can result in recurrent ambiguous base calls, which can accumulate rapidly in response to the emergence of a new variant. We also found examples of consistent base calling errors, associated with PCR artifacts and amplicon drop-out. Similarly, misalignments and partially mapped reads on the reference genome resulted in ambiguous base calls, as well as defining mutations being omitted from the assembly. These findings highlight some key limitations of using targeted enrichment by PCR and reference-based genome assembly for sequencing SARS-CoV-2, and the importance of continuously monitoring and updating primer schemes and bioinformatic pipelines.
Short-Term Evolution and Dispersal Patterns of Fluconazole-Resistance inCandida aurisclade III
Authors: Cancino-Muñoz Irving , Mulet-Bayona Juan Vicente , Salvador-García Carme , Tormo-Palop Nuria , Guna Remedios , Gimeno-Cardona Concepción , González-Candelas Fernando
Published: May 2024

The rapid increase of infections caused by the emerging fungal pathogenCandida aurisis of global concern, and understanding its expansion is a priority. The phylogenetic diversity of the yeast is clustered in five major clades, among which clade III is particularly relevant, as most of its strains exhibit resistance to fluconazole, reducing the therapeutic alternatives and provoking outbreaks that are difficult to control. In this study, we have investigated the phylogenetic structure of clade III by analyzing a global collection of 566 genomes. We have identified three subgroups within clade III, among which two are genetically most closely related. Moreover, we have estimated the evolutionary rate of clade III to be 2.25e-7 s/s/y (2.87 changes per year). We found that one of these subgroups shows intrinsic resistance to fluconazole and is responsible for the majority of cases within this clade globally. We inferred that this subgroup may have originated around December 2010 (95% HPD: April 2010 - June 2011), and since then it has spread across continents, generating multiple large outbreaks, each with a unique pattern of transmission and dissemination. These results highlight the remarkable ability of the pathogen to adapt to its environment and its rapid global spread, underscoring the urgent need to effectively address this epidemiological challenge.

IMPORTANCE

The number of cases affected byCandida aurishas increased worryingly worldwide. Among the currently recognized clades, clade III has the highest proportion of fluconazole-resistant cases and is spreading very rapidly, causing large nosocomial outbreaks across the globe. By analyzing complete fungal genomes from around the world, we have confirmed the origin of this clade and unraveled its dispersal patterns in the early 2010s. This finding provides knowledge that may be helpful to the public health authorities for the control of the disease.
New drinking water genome catalog identifies a globally distributed bacterial genus adapted to disinfected drinking water systems
Authors: Sudarshan Ashwin S , Dai Zihan , Gabrielli Marco , Oosthuizen-Vosloo Solize , Konstantinidis Konstantinos T. , Pinto Ameet J
Published: May 2024

Genome-resolved insights into the structure and function of the drinking water microbiome can advance the effective management of drinking water quality. To enable this, we constructed and curated thousands of metagenome-assembled and isolate genomes from drinking water distribution systems globally to develop a Drinking Water Genome Catalog (DWGC). The current DWGC disproportionately represents disinfected drinking water systems due to a paucity of metagenomes from non-disinfected systems. Using the DWGC, we identify core genera of the drinking water microbiome including a genus (UBA4765) within the orderRhizobialesthat is frequently detected and highly abundant in disinfected drinking water systems. We demonstrate that this genus has been widely detected but incorrectly classified in previous amplicon sequencing-based investigations of the drinking water microbiome. Further, we show that a single genome variant (genomevar) within this genus is detected in >82% of drinking water systems included in this study. We propose a provisional name for this uncultured bacterium as “Raskinella chlorumaquaticus” and describe the genus as “Raskinella” (pending SeqCode approval). Metabolic annotation and modeling-based predictions indicate that this bacterium is capable of necrotrophic growth, able to metabolize halogenated compounds, proliferates in a biofilm-based environment and shows clear indications of disinfection-mediated selection.

Synopsis

Creation and analysis of a new curated drinking water genome catalog identifies uncharacterized bacterial species that is widely distributed in disinfected drinking water systems.