bluuebunny/crossref_metadata_2025 huggingface dataset:
Knowledge, Attitude, and Practice among the Healthcare Professionals regarding the myths on COVID-19 vaccination - Demystified
Authors: Kumar S Lokesh , Naik Zameera , Panwar Arun , M Sridhar , Keluskar Vaishali , K Ram Surath Kumar
Published: August 2021

Background

COVID-19 vaccine is the mighty weapon opted by all the countries across the globe in an attempt to eradicate the fatal COVID-19 pandemic. The myths on the COVID-19 vaccine are spreading widely, causing a hindrance to this noble preventive measure. The prevalence of such myths among the healthcare professionals may be toxic and deadly.

Aim & Objectives

To assess the knowledge, attitude, and practice of the healthcare professionals regarding the myths on COVID-19 vaccination and to demystify them.

Materials and Methods

An 18-item questionnaire evaluating knowledge, attitude, and practice based on the existing myths on COVID-19 vaccination was circulated through Google Forms® among the 412 healthcare professionals of six disciplines belonging to a private University. The responses obtained were subjected to statistical analysis using SPSS® 20 software package.

Results

A total of 385 health professionals participated in this study. The majority of them had medium knowledge (165) and positive attitude (273) with the mean knowledge and attitude scores of 3.82 ± 1.55 out of 6 and 4.3 ± 1.58 out of 7 respectively. Even though 312 participants got vaccinated, 72 of them failed to receive it. The knowledge scores showed a high statistically significant difference among the participants of different designations (p=0.001), but not with gender, field, and staff with different years of experience (p>0.05). The attitude scores were statistically different among participants of fields and designation (p<0.05) but not among genders (p=0.31) and staff with different years of experience (p=0.87). Knowledge and attitude scores showed a positive linear correlation and a high statistically significant difference (p<0.001).

Conclusion

This study recommends more enhanced education programs on COVID-19 vaccination for the health professionals and demands an improved knowledge, attitude, and practice among them to achieve the goal of 100% vaccination so as to completely eradicate the COVID-19 pandemic.
COVID-19 Vaccine Uptake among U.S. Child Care Providers
Authors: Patel Kavin M. , Malik Amyn A. , Lee Aiden , Klotz Madeline , Humphries John Eric , Murray Thomas , Wilkinson David , Shafiq Mehr , Yildirim Inci , Elharake Jad A. , Diaz Rachel , Reyes Chin , Omer Saad B. , Gilliam Walter S.
Published: August 2021

STRUCUTRED ABSTRACT

Objectives

Ensuring a high COVID-19 vaccine uptake among U.S. child care providers is crucial to mitigating the public health implications of child-to-staff and staff-to-child transmission of SARS-CoV-2; however, the vaccination rate among this group is unknown.

Methods

To characterize the vaccine uptake among U.S. child care providers, we conducted a cross-sectional survey of the child care workforce. Providers were identified through various national databases and state registries. A link to the survey was sent via email between May 26 and June 23, 2021. Out of 44,771 potential respondents, 21,663 responded (48.4%).

Results

Overall COVID-19 vaccine uptake among U.S. child care providers (78.1%, 95% CI [77.3% to 78.9%]) was higher than that of the U.S. adult population (65%). Vaccination rates varied from 53.5% to 89.4% between states. Vaccine uptake differed significantly (p < .01) based on respondent age (70.0% for ages 25-34, 91.5% for ages 75-84), race (70.0% for Black or African Americans, 92.5% for Asian-Americans), annual household income (70.7% for <$35,000, 85.0% for>$75,000), and childcare setting (72.9% for home-based, 79.7% for center-based).

Conclusions

COVID-19 vaccine uptake among U.S. child care providers was higher than that of the general U.S. adult population. Those who were younger, lower income, Black or African American, resided in states either in the Mountain West or the South, and/or worked in home-based childcare programs reported the lowest rates of vaccination; state public health leaders and lawmakers should prioritize these subgroups for placement on the policy agenda to realize the largest gains in vaccine uptake among providers.

Tables of Contents Summary

This article describes the results of a national survey of childcare providers to determine the overall COVID-19 vaccine uptake and the gaps in vaccine coverage.

What’s Known on This Subject

Ensuring a high COVID-19 vaccine uptake among U.S. child care providers is crucial to mitigating the public health implications of child-to-staff and staff-to-child transmission of SARS-CoV-2; however, the vaccination rate among this group is unknown.

What This Study Adds

While the vaccine uptake among U.S. child care providers was higher than that of U.S. adults, certain subgroups continue to warrant focused attention for outreach and/or placement on the policy agenda.
Subcellular and regional localization of mRNA translation in midbrain dopamine neurons
Authors: Hobson Benjamin D. , Kong Linghao , Angelo Maria Florencia , Lieberman Ori J. , Mosharov Eugene V. , Herzog Etienne , Sulzer David , Sims Peter A.
Published: August 2021

Local translation within excitatory and inhibitory neurons is involved in neuronal development and synaptic plasticity. Despite the extensive dendritic and axonal arborizations of central monoaminergic neurons, the subcellular localization of protein synthesis is not well-characterized in these populations. Here, we investigated mRNA localization and translation in midbrain dopaminergic (mDA) neurons, cells with enormous axonal and dendritic projections, both of which exhibit stimulation-evoked dopamine (DA) release. Using highly-sensitive ribosome-bound RNA-sequencing and imaging approaches in mDA axons, we found no evidence for axonal mRNA localization or translation. In contrast, mDA neuronal dendritic projections into the substantia nigra reticulata (SNr) contain ribosomes and mRNAs encoding the major components of DA synthesis, release, and reuptake machinery. Surprisingly, we also observed dendritic localization of mRNAs encoding synaptic vesicle-related proteins, including those involved in vesicular exocytic fusion. Our results are consistent with a role for local translation in the regulation of DA release from dendrites, but not from axons. Our translatome data further defined a molecular signature of the sparse mDA neurons resident in the SNr, including enrichment of Atp2a3/SERCA3, an ER calcium pump previously undescribed in mDA neurons.
Ontology-aware deep learning for antibiotic resistance gene prediction: novel function discovery and comprehensive profiling from metagenomic data
Authors: Zha Yuguo , Chen Cheng , Jiao Qihong , Zeng Xiaomei , Cui Xuefeng , Ning Kang
Published: August 2021

Antibiotic resistance genes (ARGs) have emerged in pathogens and arousing a worldwide concern, which is estimated to cause millions of deaths each year globally. Accurately identifying and classifying ARGs is a formidable challenge in studying the generation and spread of antibiotic resistance. Current methods could identify close homologous ARGs, have limited utility for discovery of novel ARGs, thus rendering the profiling of ARGs incomprehensive. Here, an ontology-aware neural network (ONN) approach, ONN4ARG, is proposed for comprehensive ARG discovery. Systematic evaluation shows ONN4ARG is advanced than previous methods such as DeepARG in efficiency, accuracy, and comprehensiveness. Experiments using 200 million candidate microbial genes collected from 815 microbial community samples from diverse environments or hosts have resulted in 120,726 candidate ARGs, out of which more than 20% are not yet present in public databases. These comprehensive set of ARGs have clarified the environment-specific and host-specific patterns. The wet-experimental functional validation, together with structural investigation of docking sites, have also validated a novel streptomycin resistance gene from oral microbiome samples, confirming ONN4ARG’s ability for novel ARGs identification. In summary, ONN4ARG is superior to existing methods in efficiency, accuracy, and comprehensiveness. It enables comprehensive ARG discovery, which is helpful towards a grand view of ARGs worldwide. ONN4ARG is available athttps://github.com/HUST-NingKang-Lab/ONN4ARG, and online web service is available athttp://onn4arg.xfcui.com/.
Ecological specialisation, rather than the island effect, explains morphological diversification in an ancient radiation of geckos
Authors: Tejero-Cicuéndez Héctor , Simó-Riudalbas Marc , Menéndez Iris , Carranza Salvador
Published: August 2021

Island colonists are often assumed to experience higher levels of phenotypic diversification than continental taxa. However, empirical evidence has uncovered exceptions to this “island effect”. Here, we tested this pattern using the geckos of the genusPristurusfrom continental Arabia and Africa and the Socotra Archipelago. Using a recently published phylogeny and an extensive morphological dataset, we explore the differences in phenotypic evolution between Socotran and continental taxa. Moreover, we reconstructed ancestral habitat occupancy to examine if ecological specialisation is correlated with morphological change, comparing phenotypic disparity and trait evolution between habitats. We found a heterogeneous outcome of island colonisation. Namely, only one of the three colonisation events resulted in a body size increase. However, in general, Socotran species do not present higher levels or rates of morphological diversification than continental groups. Instead, habitat specialisation explains better the body size and shape evolution inPristurus. Particularly, the colonisation of ground habitats appears as the main driver of morphological change, producing the highest disparity and evolutionary rates. Additionally, arboreal species show very similar body size and head proportions. These results reveal a determinant role of ecological mechanisms in morphological evolution and corroborate the complexity of ecomorphological dynamics in continent-island systems.
Exploring the miRNA-mediated response to combined stresses in melon plants
Authors: Villalba-Bermell Pascual , Marquez-Molins Joan , Marques María-Carmen , Hernandez-Azurdia Andrea G. , Corell-Sierra Julia , Picó Belén , Monforte Antonio J. , Elena Santiago F. , Gomez Gustavo G.
Published: August 2021

Climate change has been associated with a higher incidence of combined adverse environmental conditions that can promote a significant decrease in crop productivity. However, knowledge on how a combination of stresses might affect plant development is still scarce. MicroRNAs (miRNAs) have been proposed as potential targets for improving crop-productivity. Here, we have combined deep-sequencing, computational characterization of responsive miRNAs and validation of their regulatory role in a comprehensive analysis of melon’s response to several combinations of four stresses (cold, salinity, short day, and infection with a fungus). Twenty-two miRNA families responding to double and/or triple stresses were identified. The regulatory role of the differentially expressed miRNAs was validated by quantitative measurements of the expression of the corresponding target genes. A high proportion (ca. 60%) of these families (mainly highly conserved miRNAs targeting transcription factors) showed a non-additive response to multiple stresses in comparison with that observed under each one of the stresses individually. Among those miRNAs showing non-additive response to stress-combinations, most interactions were negative suggesting the existence of functional convergence in the miRNA-mediated response to combined stresses. Taken together, our results provide compelling evidences that the response to combined stresses cannot be easily predicted from the study individual stresses.
Visuo-vestibular gaze control – conserved subcortical processing
Authors: Wibble Tobias , Pansell Tony , Grillner Sten , Pérez-Fernández Juan
Published: August 2021

Gaze stabilization compensates for movements of the head or external environment to minimize image blurring, which is critical for visually-guided behaviors. Multisensory information is used to stabilize the visual scene on the retina via the vestibulo-ocular (VOR) and optokinetic (OKR) reflexes. While the organization of neuronal circuits underlying VOR is well described across vertebrates, less is known about the contribution and evolutionary origin of the OKR circuits. Moreover, the integration of these two sensory modalities is still poorly understood. Here, we developed a novel experimental model, the isolated lamprey eye-brain-labyrinth preparation, to analyze the neuronal pathways underlying visuo-vestibular integration which allowed electrophysiological recordings while applying vestibular stimulation using a moving platform, coordinated with visual stimulation via two screens. We show that lampreys exhibit robust visuo-vestibular integration, with optokinetic information processed in the pretectum and integrated with vestibular inputs at several subcortical levels. The enhanced eye movement response to multimodal stimulation favored the vestibular response at increased velocities. The optokinetic signals can be downregulated from tectum. Additionally, saccades are present in the form of nystagmus. The lamprey represents the oldest living group of vertebrates, thus all basic components of the visuo-vestibular control of gaze were present already at the dawn of vertebrate evolution.
Xenotransplantation of porcine progenitor cells in an epileptic California sea lion (Zalophus californianus)
Authors: Simeone Claire A. , Andrews John P. , Johnson Shawn P. , Casalia Mariana , Kochanski Ryan , Chang Edward F. , Cameron Dianne , Dennison Sophie , Inglis Ben , Scott Gregory , Kruse-Elliott Kris , Okonski F. Fabian , Calvo Eric , Goulet Kelly , Robles Dawn , Griffin-Stence Ashley , Kuiper Erin , Krasovec Laura , Field Cara L. , Hoard Vanessa F. , Baraban Scott C.
Published: August 2021

Background

Domoic acid (DA) is a naturally occurring neurotoxin harmful to marine animals and humans. California sea lions exposed to DA in prey during algal blooms along the Pacific coast exhibit significant neurological symptoms, including epilepsy with hippocampal atrophy.

Observations

Here we describe a xenotransplantation procedure to deliver interneuron progenitor cells into the damaged hippocampus of an epileptic sea lion with suspected DA toxicosis. The sea lion has had no evidence of seizures following the procedure, and clinical measures of well-being including weight and feeding habits have stabilized.

Lessons

These preliminary results suggest xenotransplantation has improved the quality-of-life (QOL) for this animal and holds tremendous therapeutic promise.
FACT regulates pluripotency through distal regulation of gene expression in murine embryonic stem cells
Authors: Klein David C. , Lardo Santana M. , McCannell Kurtis N. , Hainer Sarah J.
Published: August 2021

The FACT complex is a conserved histone chaperone with essential roles in transcription and histone deposition. FACT is essential in pluripotent and cancer cells, but otherwise dispensable for most mammalian cell types. FACT deletion or inhibition can block induction of pluripotent stem cells, yet the mechanism through which FACT regulates cell fate decisions remains unclear. To determine this mechanism, we used inducible depletion of FACT subunit SPT16 in murine embryonic stem cells paired with genomic factor localization, nascent transcription, and chromatin accessibility analyses. Over a timecourse of SPT16 depletion, nucleosomes invade loci bound by master pluripotency factors and gene-distal DNaseI hypersensitive sites. Simultaneously, transcription of Pou5f1 (OCT4), Sox2, Nanog, and enhancer RNAs produced at the genes’ associated enhancers are downregulated, suggesting that FACT regulates expression of the pluripotency factors themselves. We find that FACT maintains cellular pluripotency through a precise nucleosome-based regulatory mechanism for appropriate expression of both coding and non-coding transcripts associated with pluripotency.
Length Biases in Single-Cell RNA Sequencing of pre-mRNA
Authors: Gorin Gennady , Pachter Lior
Published: August 2021

Single-molecule pre-mRNA and mRNA sequencing data can be modeled and analyzed using the Markov chain formalism to yield genome-wide insights into transcription. However, quantitative inference with such data requires careful assessment and understanding of noise sources. We find that long pre-mRNA transcripts are over-represented in sequencing data, and explore the mechanistic implications. A biological explanation for this phenomenon within our modeling framework requires unrealistic transcriptional parameters, leading us to posit a length-based model of capture bias. We provide solutions for this model, and use them to find concordant and mechanistically plausible parameter trends across data from multiple single-cell RNA-seq experiments in several species.
Modular Fluorescent Nanoparticle DNA Probes for Detection of Peptides and Proteins
Authors: Stawicki Cassandra M. , Rinker Torri E. , Burns Markus , Tonapi Sonal S. , Galimidi Rachel P. , Anumala Deepthi , Robinson Julia K. , Klein Joshua S. , Mallick Parag
Published: August 2021

Fluorescently labeled antibody and aptamer probes are used in biological studies to characterize binding interactions, measure concentrations of analytes, and sort cells. Fluorescent nanoparticle labels offer an excellent alternative to standard fluorescent labeling strategies due to their enhanced brightness, stability and multivalency; however, challenges in functionalization and characterization have impeded their use. This work introduces a straightforward approach for preparation of fluorescent nanoparticle probes using commercially available reagents and common laboratory equipment. Fluorescent polystyrene nanoparticles, Thermo Fisher Scientific FluoSpheres™, were used in proof-of-principle studies. Particle passivation was achieved by covalent attachment of amine-PEG-azide to carboxylated particles, neutralizing the surface charge from -43 to -15 mV. A conjugation-annealing handle and DNA aptamer probe was attached to the azide-PEG nanoparticle surface either through reaction of pre-annealed handle and probe or through a stepwise reaction of the nanoparticles with the handle followed by aptamer annealing. Nanoparticles functionalized with DNA aptamers targeting histidine tags and VEGF protein had high affinity (EC50s ranging from 3-12 nM) and specificity, and were more stable than conventional labels. This protocol for preparation of nanoparticle probes relies solely on commercially available reagents and common equipment, breaking down the barriers to use nanoparticles in biological experiments.
Emergence of distinct and heterogeneous strains of amyloid beta as Alzheimer’s disease progresses in Down syndrome
Authors: Maxwell Alison M. , Yuan Peng , Rivera Brianna M. , Schaaf Wilder , Mladinov Mihovil , Prasher Vee P. , Robinson Andrew C. , DeGrado William F. , Condello Carlo
Published: August 2021

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Amyloid beta (Aβ) is thought to play a critical role in the pathogenesis of Alzheimer’s disease (AD). Prion-like Aβ polymorphs, or “strains”, can have varying pathogenicity and may underlie the phenotypic heterogeneity of the disease. In order to develop effective AD therapies, it is critical to identify the strains of Aβ that might arise prior to the onset of clinical symptoms and understand how they may change with progressing disease. Down syndrome (DS), as the most common genetic cause of AD, presents promising opportunities to compare such features between early and advanced AD. In this work, we evaluate the neuropathology and Aβ strain profile in the post-mortem brain tissues of 210 DS, AD, and control individuals. We assayed the levels of various Aβ and tau species and used conformation-sensitive fluorescent probes to detect differences in Aβ strains among individuals and populations. We found that these cohorts have some common but also some distinct strains from one another, with the most heterogeneous populations of Aβ emerging in subjects with high levels of AD pathology. The emergence of distinct strains in DS at these later stages of disease suggests that the confluence of aging, pathology, and other DS-linked factors may favor conditions that generate strains that are unique from sAD.
Chronic ethanol drinking in non-human primates induces inflammatory cathepsin gene expression in alveolar macrophages accompanied by functional defects
Authors: Lewis Sloan A. , Doratt Brianna , Sureshchandra Suhas , Jankeel Allen , Newman Natali , Grant Kathleen A. , Messaoudi Ilhem
Published: August 2021

Chronic alcohol drinking is associated with increased susceptibility to viral and bacterial respiratory pathogens. Investigating the effects of alcohol on the lung is challenging in humans because of the complexity of human drinking behavior and the challenge of obtaining samples. In this study, we utilize a rhesus macaque model of voluntary ethanol self-administration to study the effects of alcohol on the lung in a physiologically and genetically relevant model. We report a heightened activation and inflammatory state in alveolar macrophages (AM) obtained from ethanol drinking animals that is accompanied by increased chromatin accessibility in intergenic regions that regulate inflammatory genes and contain binding motifs for transcription factors AP-1, IRF8, and NFKB p-65. In line with these transcriptional and epigenetic changes at basal state, AM from ethanol drinking animals generate elevated inflammatory mediator responses to LPS and respiratory syncytial virus (RSV). Analysis using scRNA-Seq revealed heterogeneity in lung-resident macrophage and monocyte populations, including increased abundance of activated and cathepsin-expressing clusters and accelerated differentiation with ethanol. Finally, functional assays show increased mitochondrial content in AM from ethanol drinking animals, which is associated with observed increased ROS and decreased phagocytosis capacity. This comprehensive epigenomic, transcriptional and functional profiling of lung macrophages after ethanol drinking in macaques provides previously unidentified mechanisms of ethanol induced infection susceptibility in patients with alcohol use disorders.
Close agreement between deterministic vs. stochastic modeling of first-passage time to vesicle fusion
Authors: Matveev Victor
Published: August 2021

Ca2+-dependent cell processes such as neurotransmitter or endocrine vesicle fusion are inherently stochastic due to large fluctuations in Ca2+ channel gating, Ca2+ diffusion and Ca2+ binding to buffers and target sensors. However, prior studies revealed closer-than-expected agreement between deterministic and stochastic simulations of Ca2+ diffusion, buffering and sensing, as long as Ca2+ channel gating is not Ca2+-dependent. To understand this result more fully, we present a comparative study complementing prior work, focusing on Ca2+ dynamics downstream of Ca2+ channel gating. Specifically, we compare deterministic (mean-field / mass-action) and stochastic simulations of vesicle exocytosis latency, quantified by the probability density of the first-passage time (FPT) to the Ca2+-bound state of a vesicle fusion sensor, following a brief Ca2+ current pulse. We show that under physiological constraints, the discrepancy between FPT densities obtained using the two approaches remains small even if as few as ∼50 Ca2+ ions enter per single channel-vesicle release unit. Using a reduced two-compartment model for ease of analysis, we illustrate how this close agreement arises from the smallness of correlations between fluctuations of the reactant molecule numbers, despite the large magnitude of the fluctuation amplitudes. This holds if all relevant reactions are heteroreaction between molecules of different species, as is the case for the bimolecular Ca2+ binding to buffers and downstream sensor targets. In this case diffusion and buffering effectively decorrelate the state of the Ca2+ sensor from local Ca2+ fluctuations. Thus, fluctuations in the Ca2+ sensor’s state underlying the FPT distribution are only weakly affected by the fluctuations in the local Ca2+ concentration around its average, deterministically computable value.

Statement of Significance

Many fundamental Ca2+-dependent cell processes are triggered by local Ca2+ elevations involving only a few hundred Ca2+ ions. Therefore, one expects large Ca2+ concentration fluctuations, which are ignored by deterministic reaction-diffusion modeling approaches. However, more accurate stochastic approaches require tracking trajectories of individual Ca2+ ions and its binding targets, which is very computationally expensive. This study reveals conditions under which Ca2+-dependent processes like secretory vesicle fusion can be modeled using efficient deterministic approaches, without sacrificing significant accuracy. We find that deterministic methods can accurately predict the delay to the fusion of a neurotransmitter-containing vesicle, as long as the number of Ca2+ ions is above about 50. We reveal factors that explain the limited impact of stochastic fluctuations in this case.
Uncovering perturbations in human hematopoiesis associated with healthy aging and myeloid malignancies at single cell resolution
Authors: Ainciburu Marina , Ezponda Teresa , Berastegui Nerea , Alfonso-Pierola Ana , Vilas-Zornoza Amaia , Martin-Uriz Patxi San , Alignani Diego , de Espinosa Jose Lamo , Julian Mikel San , Jimenez Tamara , López Félix , Muntion Sandra , Sanchez-Guijo Fermin , Molero Antonieta , Montoro Julia , Serrano Guillermo , Diaz-Mazkiaran Aintzane , Lasaga Miren , Gomez-Cabrero David , Díez-Campelo María , Valcarcel David , Hernaez Mikel , Romero Juan P. , Prosper Felipe
Published: August 2021

Early hematopoiesis is a continuous process in which hematopoietic stem and progenitor cells (HSPCs) gradually differentiate toward specific lineages. Aging and myeloid malignant transformation are characterized by changes in the composition and regulation of HSPCs. In this study, we used single cell RNA sequencing (scRNAseq) to characterize an enriched population of human hematopoietic stem and progenitor cells (HSPC) obtained from young and elderly healthy individuals.

Based on their transcriptional profile, we identified changes in the proportions of progenitor compartments during aging, and differences in their functionality, as evidenced by gene set enrichment analysis. Trajectory inference revealed that altered gene expression dynamics accompanied cell differentiation, which could explain age-associated changes in hematopoiesis. Next, we focused on key regulators of transcription by constructing gene regulatory networks and detected regulons that were specifically active in elderly individuals. Using previous findings in healthy cells as a reference, we analyzed scRNA-seq data obtained from patients with myelodysplastic syndrome (MDS) and detected specific alterations of the expression dynamics of genes involved in erythroid differentiation in all patients with MDS such as TRIB2. In addition, the comparison between transcriptional programs and GRN regulating normal HSPCs and MDS HSPCs allowed identification of regulons that were specifically active in MDS cases such as SMAD1, HOXA6, POU2F2 and RUNX1 suggesting a role of these TF in the pathogenesis of the disease.

In summary, we demonstrate that the combination of single cell technologies with computational analysis tools enable the study of a variety of cellular mechanisms involved in early hematopoiesis and can be used to dissect perturbed differentiation trajectories associated with aging and malignant transformation. Furthermore, the identification of abnormal regulatory mechanisms associated with myeloid malignancies could be exploited for personalized therapeutic approaches in individual patients.
Circadian Proteins Cry and Rev-erb Converge to Deepen Cellular Quiescence by Downregulating Cyclin D and Cdk4,6
Authors: Wang Xia , Liu Bi , Pan Qiong , Kwon Jungeun Sarah , Miller Matthew A. , Croce Kimiko Della , Yao Guang
Published: August 2021

The proper balance and transition between cellular quiescence and proliferation are critical to tissue homeostasis, and their deregulations are commonly found in many human diseases, including cancer and aging. Recent studies showed that the reentry of quiescent cells to the cell cycle is subjected to circadian regulation. However, the underlying mechanisms are largely unknown. Here, we report that two circadian proteins, Cryptochrome (Cry) and Rev-erb, deepen cellular quiescence in rat embryonic fibroblasts, resulting in stronger serum stimulation required for cells to exit quiescence and reenter the cell cycle. This finding was opposite from what we expected from the literature. By modeling a library of possible regulatory topologies linking Cry and Rev-erb to a bistable Rb-E2f gene network switch that controls the quiescence-to-proliferation transition and by experimentally testing model predictions, we found Cry and Rev-erb converge to downregulate Cyclin D/Cdk4,6 activity, leading to an ultrasensitive increase of the serum threshold to activate the Rb-E2f bistable switch. Our findings suggest a mechanistic role of circadian proteins in modulating the depth of cellular quiescence, which may have implications in the varying potentials of tissue repair and regeneration at different times of the day.
A single nucleotide change in thepolCDNA polymerase III inClostridium thermocellumis sufficient to create a hypermutator phenotype
Authors: Lanahan Anthony , Zakowicz Kamila , Tian Liang , Olson Daniel G. , Lynd Lee R.
Published: August 2021

Clostridium thermocellumis a thermophilic, anaerobic, bacterium that natively ferments cellulose to ethanol, and is a candidate for cellulosic biofuel production. Recently, we identified a hypermutator strain ofC. thermocellumwith a C669Y mutation in thepolCgene, which encodes a DNA polymerase III enzyme. Here we reintroduce this mutation using recently-developed CRISPR tools to demonstrate that this mutation is sufficient to recreate the hypermutator phenotype. The resulting strain shows an approximately 30-fold increase in the mutation rate. This mutation appears to function by interfering with metal ion coordination in the PHP domain responsible for proofreading. The ability to selectively increase the mutation rate inC. thermocellumis a useful tool for future directed evolution experiments.

Importance

Cellulosic biofuels are a promising approach to decarbonize the heavy duty transportation sector. A longstanding barrier to cost-effective cellulosic biofuel production is the recalcitrance of the material to solubilization. Native cellulose-consuming organisms, such asClostridium thermocellum, are promising candidates for cellulosic biofuel production, however they often need to be genetically modified to improve product formation. One approach is adaptive laboratory evolution. Our findings demonstrate a way to increase the mutation rate in this industrially-relevant organism, which can reduce the time needed for adaptive evolution experiments.
Telmisartan restricts Chikungunya virus infection in vitro and in vivo through the AT1/ PPAR-γ/MAPKs pathways
Authors: De Saikat , Mamidi Prabhudutta , Ghosh Soumyajit , Keshry Supriya Suman , Mahish Chandan , Pani Sweta Smita , Laha Eshna , Ray Amrita , Datey Ankita , Chatterjee Sanchari , Singh Sharad , Mukherjee Tathagata , Khamaru Somlata , Chattopadhyay Subhasis , Subudhi Bharat Bhusan , Chattopadhyay Soma
Published: August 2021

Chikungunya virus (CHIKV) has re-emerged as a global public health threat. The inflammatory pathways of RAS and PPAR-γ are usually involved in viral infections. Thus, Telmisartan (TM) with known capacity to block AT1 receptor and activate PPAR-γ, was investigated against CHIKV. The anti-CHIKV effect of TM was investigated in vitro (Vero, RAW 264.7 cells and hPBMCs) and in vivo (C57BL/6 mice). TM was found to abrogate CHIKV infection efficiently (IC50 of 15.34-20.89µM in the Vero and RAW 264.7 cells respectively). Viral RNA and proteins were reduced remarkably with the TM driven modulation of host m-TOR signaling. Additionally, TM interfered in the early and late stages of CHIKV life cycle with efficacy in both pre and post-treatment assay. Moreover, the agonist of AT1 receptor and antagonist of PPAR-γ increased CHIKV infection suggesting TM’s anti-viral potential by modulating host factors. Besides, reduced activation of all major MAPKs, NF-κB (p65) and cytokines by TM through the inflammatory axis supported the fact that the anti-CHIKV efficacy of TM is partly mediated through the AT1/PPAR-γ/MAPKs pathways. Interestingly, at the human equivalent dose, TM abrogated CHIKV infection and inflammation significantly leading to reduced clinical score and complete survival of C57BL/6 mice. Additionally, TM reduced infection in hPBMC derived monocyte-macrophage populations in vitro. Hence, TM was found to reduce CHIKV infection by targeting both viral and host factors. Considering its safety and in vivo efficacy, it can be a suitable candidate in future for repurposing against CHIKV.
External Validation of a Mobile Clinical Decision Support System for Diarrhea Etiology Prediction in Children: A Multicenter Study in Bangladesh and Mali
Authors: Garbern Stephanie C , Nelson Eric J , Nasrin Sabiha , Keita Adama Mamby , Brintz Ben J , Gainey Monique , Badji Henry , Nasrin Dilruba , Howard Joel , Taniuchi Mami , Platts-Mills James A. , Kotloff Karen L , Haque Rashidul , Levine Adam C , Sow Samba O , Alam Nur H , Leung Daniel T
Published: August 2021

Background

Diarrheal illness is a leading cause of antibiotic use for children in low- and middle-income countries. Determination of diarrhea etiology at the point-of-care without reliance on laboratory testing has the potential to reduce inappropriate antibiotic use.

Methods

This prospective observational study aimed to develop and externally validate the accuracy of a mobile software application (“App”) for the prediction of viral-only etiology of acute diarrhea in children 0-59 months in Bangladesh and Mali. The App used previously derived and internally validated models using combinations of “patient-intrinsic” information (age, blood in stool, vomiting, breastfeeding status, and mid-upper arm circumference), pre-test odds using location-specific historical prevalence and recent patients, climate, and viral seasonality. Diarrhea etiology was determined with TaqMan Array Card using episode-specific attributable fraction (AFe) >0.5.

Results

Of 302 children with acute diarrhea enrolled, 199 had etiologies above the AFe threshold. Viral-only pathogens were detected in 22% of patients in Mali and 63% in Bangladesh. Rotavirus was the most common pathogen detected (16% Mali; 60% Bangladesh). The viral seasonality model had an AUC of 0.754 (0.665-0.843) for the sites combined, with calibration-in-the-large α=-0.393 (−0.455 – -0.331) and calibration slope β=1.287 (1.207 – 1.367). By site, the pre-test odds model performed best in Mali with an AUC of 0.783 (0.705 - 0.86); the viral seasonality model performed best in Bangladesh with AUC 0.710 (0.595 - 0.825).

Conclusion

The App accurately identified children with high likelihood of viral-only diarrhea etiology. Further studies to evaluate the App’s potential use in diagnostic and antimicrobial stewardship are underway.
COVID-19 vaccine response in people with multiple sclerosis
Authors: Tallantyre Emma C , Vickaryous Nicola , Anderson Valerie , Asardag Aliye Nazli , Baker David , Bestwick Jonathan , Bramhall Kath , Chance Randy , Evangelou Nikos , George Katila , Giovannoni Gavin , Grant Leanne , Harding Katharine E , Hibbert Aimee , Ingram Gillian , Jones Meleri , Kang Angray S , Loveless Samantha , Moat Stuart J , Robertson Neil P , Schmierer Klaus , Shah Sita Navin , Simmons Jessica , Upcott Matthew , Willis Mark , Jolles Stephen , Dobson Ruth
Published: August 2021

Objective

To investigate the effect of disease modifying therapies on serological response to SARS-CoV2 vaccines in people with multiple sclerosis

Methods

473 people with multiple sclerosis from 5 centres provided one or more dried blood spot samples and questionnaires about COVID-19. Information about disease and drug history was extracted from their medical records. Dried blood spots were eluted and tested for antibodies to SARS-CoV2 receptor binding domain. Seropositivity was expressed according to validated cut-off indices. Antibody titers were partitioned into tertiles using data from people on no disease modifying therapy as a reference. We calculated the odds ratio of seroconversion (Univariate logistic regression) and compared quantitative vaccine response (Kruskal Wallis) following SARS-CoV2 vaccine according to disease modifying therapy. We used regression modelling to explore the effect of factors including vaccine timing, treatment duration, age, vaccine type and lymphocyte count on vaccine response.

Results

Compared to no disease modifying therapy, the use of anti-CD20 monoclonal antibodies (odds ratio 0.03; 95% confidence interval 0.01-0.06, p<0.001) and fingolimod (odds ratio 0.41; 95% confidence interval 0.01-0.12) were associated with lower seroconversion following SARS-CoV2 vaccine. All other drug groups did not differ significantly from the untreated cohort. Time since last anti-CD20 treatment and total time on treatment were significantly related with response to vaccination. Vaccine type significantly predicted seroconversion, but not in those on anti-CD20 medications.

Interpretation

Some disease modifying therapies carry a risk of attenuated response to SARS-CoV2 vaccination in people with MS. We provide recommendations for the practical management of this patient group.
Cognitive reserve in a sample of Cuban older adults with Mild Cognitive Impairment
Authors: Jiménez-Puig Elizabeth , Baute-Abreu Ana Cristina , Valdés-Pérez Daniela Liz , Santana-Domínguez Yeisel , Añón-Villegas Marina Jesús , Pérez-Leiva Brayan Deivi
Published: August 2021

Introduction

Aging is a natural process that occurs in all individuals. Due to the increase in aging figures worldwide, the diagnosis of mild cognitive impairment and other dementia syndromes has increased. An element of essential relevance in protecting against the risk of suffering from dementia is the cognitive reserve.

Objective

To indicate the levels of cognitive reserve in a sample of Cuban older adults with Mild Cognitive Impairment.

Methods

An exploratory study was carried out using questionnaires, with a cross-sectional, quantitative design. The sampling was non-probabilistic and intentional, obtaining a sample of 51 older adults with mild cognitive impairment. The information collection was carried out from the Cognitive Reserve Scale (CRS). Quantitative data was analyzed through frequency analysis.

Results

Most of the participants showed higher levels of cognitive reserve for carrying out activities of daily life and for relating to other people. The dimensions most affected were Formation / Information and Hobbies.

Conclusions

The research allowed a preliminary approach to the cognitive reserve of older adults. It would be interesting to conduct investigations of this nature in larger samples, including groups of healthy controls, which would allow comparison of results. In addition, it would be relevant to investigate the impact of institutionalization.
Development of an efficient single-cell cloning and expansion strategy for genome edited induced pluripotent stem cells
Authors: Bhargava Nupur , Thakur Priya , Muruganandam Thulasi Priyadharshini , Jaitly Shashank , Gupta Pragya , Lohani Neelam , Goswami Sangam Giri , Bhattacharya Saurabh Kumar , Jain Suman , Ramalingam Sivaprakash
Published: August 2021

Disease-specific human induced pluripotent stem cells (hiPSCs) can be generated directly from individuals with known disease characteristics or alternatively be modified using genome editing approaches to introduce disease causing genetic mutations to study the biological response of those mutations. The genome editing procedure in hiPSCs is still inefficient, particularly when it comes to homology directed repair (HDR) of genetic mutations or targeted transgene insertion in the genome and single cell cloning of edited cells. In addition, genome editing processes also involve additional cellular stresses such as trouble with cell viability and genetic stability of hiPSCs. Therefore, efficient workflows are desired to increase genome editing application to hiPSC disease models and therapeutic applications. Apart from genome editing efficiency, hiPSC survival following single-cell cloning has proved to be challenging and has thus restricted the capability to easily isolate homogeneous clones from edited hiPSCs. To this end, we demonstrate an efficient workflow for feeder-free single cell clone generation and expansion in both CRISPR-mediated knock-out (KO) and knock-in (KI) hiPSC lines. Using StemFlex medium and CloneR supplement in conjunction with Matrigel cell culture matrix, we show that cell viability and expansion during single-cell cloning in edited and unedited cells is significantly enhanced. Our reliable single-cell cloning and expansion workflow did not affect the biology of the hiPSCs as the cells retained their growth and morphology, expression of various pluripotency markers and normal karyotype. This simplified and efficient workflow will allow for a new level of sophistication in generating hiPSC-based disease models to promote rapid advancement in basic research and also the development of novel cellular therapeutics.
Histone H1 prevents non-CG methylation-mediated small RNA biogenesis in Arabidopsis heterochromatin
Authors: Choi Jaemyung , Lyons David B. , Zilberman Daniel
Published: August 2021

Flowering plants utilize small RNA molecules to guide DNA methyltransferases to genomic sequences. This RNA-directed DNA methylation (RdDM) pathway preferentially targets euchromatic transposable elements. However, RdDM is thought to be recruited by methylation of histone H3 at lysine 9 (H3K9me), a hallmark of heterochromatin. How RdDM is targeted to euchromatin despite an affinity for H3K9me is unclear. Here we show that loss of histone H1 enhances heterochromatic RdDM, preferentially at nucleosome linker DNA. Surprisingly, this does not require SHH1, the RdDM component that binds H3K9me. Furthermore, H3K9me is dispensable for RdDM, as is CG DNA methylation. Instead, we find that non-CG methylation is specifically required for small RNA biogenesis, and without H1 small RNA production quantitatively expands to non-CG methylated loci. Our results demonstrate that H1 enforces the separation of euchromatic and heterochromatic DNA methylation pathways by excluding the small RNA-generating branch of RdDM from non-CG methylated heterochromatin.
A pore-forming protein drives macropinocytosis to facilitate toad water maintaining
Authors: Zhao Zhong , Shi Zhi-Hong , Ye Chen-Jun , Zhang Yun
Published: August 2021

Maintaining water balance is a real challenge for amphibians in terrestrial environments. Our previous studies with toad Bombina maxima discovered a secretory aerolysin family pore-forming protein and trefoil factor complex βγ-CAT, which is assembled under tight regulation depending on environmental cues. Here we report an unexpected role for βγ-CAT in toad water maintaining. Deletion of toad skin secretions, in which βγ-CAT is a major component, increased animal mortality under hypertonic stress. βγ-CAT was constitutively expressed in toad osmoregulatory organs, which was inducible under the variation of osmotic conditions. The protein induced and participated in macropinocytosis in vivo and in vitro. During extracellular hyperosmosis, βγ-CAT stimulated macropinocytosis to facilitate water intake and enhanced exosomes release, which simultaneously regulated aquaporins distribution. Collectively, these findings uncovered that besides membrane integrated aquaporins, a secretory pore-forming protein can facilitate toad water maintaining via macropinocytosis induction and exocytosis modulation, especially in responses to osmotic stress.
The immediate early protein 1 of human herpesvirus 6B counteracts ATM activation in an NBS1-dependent manner
Authors: Collin Vanessa , Biquand Élise , Tremblay Vincent , Lavoie Élise G. , Dessapt Julien , Blondeau Andréanne , Gravel Annie , Flamand Louis , Fradet-Turcotte Amélie
Published: August 2021

Viral infection often trigger an ATM-dependent DNA damage response (DDR) in host cells that suppresses viral replication. To counteract this antiviral surveillance system, viruses evolved different strategies to induce the degradation of the MRE11/RAD50/NBS1 (MRN) complex and prevent subsequent DDR signaling. Here, we report that human herpesvirus 6B (HHV-6B) infection causes genomic instability by suppressing the host cell’s ability to induce ATM-dependent signaling pathways. Expression of immediate early protein 1 (IE1) phenocopies this phenotype and blocks further homology-directed double-strand break (DSB) repair. In contrast to other viruses, IE1 does not affect the stability of the MRN complex. Instead, it uses two distinct domains to inhibit ATM serine/threonine kinase (ATM) activation at DSBs. Structure-based analyses revealed that the N-terminal domain of IE1 interacts with the BRCA1 C-terminal domain 2 of nibrin (NBN, also known as NBS1), while ATM inhibition is attributable to on its C-terminal domain. Consistent with the role of the MRN complex in antiviral responses, NBS1 depletion resulted in increased HHV-6B replication in infected cells. However, in semi-permissive cells, viral integration of HHV-6B into the telomeres was not strictly dependent on NBS1, supporting models where this process occurs via telomere elongation rather than through DNA repair. Interestingly, as IE1 expression has been detected in cells of subjects with inherited chromosomally-integrated form of HHV-6B (iciHHV-6B), a condition associated with several health conditions, our results raise the possibility of a link between genomic instability and the development of iciHHV-6-associated diseases.

Significance Statement

Many viruses have evolved ways to inhibit DNA damage signaling, presumably to prevent infected cells from activating an antiviral response. Here, we show that this is also true for human herpesvirus 6B (HHV-6B), through its immediate early protein 1 (IE1). However, in contrast to adenovirus’ immediate early proteins, HHV-6B IE1 is recruited to double-strand breaks in an NBS1-dependent manner and inhibits ATM serine/threonine kinase activation. Characterizing this phenotype revealed a unique mechanism by which HHV-6B manipulates DNA damage signaling in infected cells. Consistently, viral replication is restricted by the MRN complex in HHV-6B infected cells. Viral integration of HHV-6B into the host’s telomeres is not strictly dependent on NBS1, challenging current models where integration occurs through homology-directed repair.