bluuebunny/crossref_metadata_2025 huggingface dataset:
Quantifying transmissibility of COVID-19 and impact of intervention within long-term health care facilities
Authors: Stockdale Jessica E. , Anderson Sean C. , Edwards Andrew M. , Iyaniwura Sarafa A. , Mulberry Nicola , Otterstatter Michael C. , Janjua Naveed Z. , Coombs Daniel , Colijn Caroline , Irvine Michael A
Published: February 2021

Estimates of the basic reproduction number (R0) for Coronavirus disease 2019 (COVID-19) are particularly variable in the context of transmission within locations such as long-term health care (LTHC) facilities. We sought to characterise the heterogeneity ofR0across known outbreaks within these facilities. We used a unique comprehensive dataset of all outbreaks that have occurred within LTHC facilities in British Columbia, Canada. We estimatedR0with a Bayesian hierarchical dynamic model of susceptible, exposed, infected, and recovered individuals, that incorporates heterogeneity ofR0between facilities. We further compared these estimates to those obtained with standard methods that utilize the exponential growth rate and maximum likelihood. The total size of an outbreak varied dramatically, with a range of attack rates of 2%–86%. The Bayesian analysis provides more constrained overall estimates ofR0= 2.19 (90% CrI [credible interval] 0.19–6.69) than standard methods, with a range within facilities of 0.48–10.08. We further estimated that intervention led to 57% (47%–66%) of all cases being averted within the LTHC facilities, or 73% (63%–78%) when using a model with multi-level intervention effect. Understanding the risks and impact of intervention are essential in planning during the ongoing global pandemic, particularly in high-risk environments such as LTHC facilities.
Antibiotic stewardship in Norway: are strategies for rational use of antibiotics followed in Norwegian healthcare system?
Authors: Gutema Girma , Hakonsen Helle , Engidawork Ephrem , Toverud Else-Lydia
Published: February 2021

This manuscript has been withdrawn by medRxiv as it was submitted and made public without the full consent of all the authors. If you have any questions, please contact the corresponding author.
Protocol for a nationwide Internet-based health survey in workers during the COVID-19 pandemic in 2020
Authors: Fujino Yoshihisa , Ishimaru Tomohiro , Eguchi Hisashi , Tsuji Mayumi , Tateishi Seiichiro , Ogami Akira , Mori Koji , Matsuda Shinya ,   for the CORoNaWork project
Published: February 2021

The ever-changing social implications of the COVID-19 pandemic have resulted in an urgent need to understand the working environments and health status of workers. We conducted a nationwide Internet-based health survey in Japanese workers in December 2020, in the midst the country’s “third wave” of COVID-19 infection. Of 33,087 surveys collected, 6,051 were determined to have invalid responses. The 27,036 surveys included in the study were balanced in terms of geographical area, participant sex, and type of work, according to the sampling plan. Men were more likely than women to have telecommuted, while women were more likely to have resigned since April 2020. Moreover, 40% and 9.1% of respondents had a K6 score of 5 or higher and 13 or higher, respectively, they did not exhibit extremely poor health. The present study describes the protocol used to conduct an Internet-based health survey in workers and a summary of its results during a period when COVID-19 was spreading rapidly in Japan. In the future, we plan to use this survey to examine the impact of COVID-19 on workers’ work styles and health.
Estimating the Population Benefits of Blood Pressure Lowering: A Wide-Angled Mendelian Randomization Study in UK Biobank
Authors: Higgins Hannah , Mason Amy M. , Larsson Susanna C. , Gill Dipender , Langenberg Claudia , Burgess Stephen
Published: February 2021

Background

The causal relevance of elevated blood pressure for several cardiovascular diseases is uncertain, as is the population impact of blood pressure lowering on risk of cardiovascular diseases more broadly. This study systematically assesses evidence of causality for various cardiovascular diseases in a two-sample Mendelian randomization framework, and estimates the potential reduction in the prevalence of these diseases attributable to long-term population shifts in the distribution of systolic blood pressure (SBP).

Methods and Results

We investigated associations of genetically-predicted SBP as predicted by 256 genetic variants with 21 cardiovascular diseases in UK Biobank, a population-based cohort of UK residents. The sample consisted of 376,703 participants of European ancestry aged 40-69 years at baseline. Genetically-predicted SBP was positively associated with 14 of the outcomes (p<0.002), including dilated cardiomyopathy, endocarditis, peripheral vascular disease, and rheumatic heart disease. Using genetic variation to estimate the long-term impact of blood pressure lowering on disease, population reductions in SBP were predicted to result in an overall 16.9% (95% confidence interval (CI): 12.2-21.3%) decrease in morbidity for a 5 mmHg decrease from a population mean of 137.7 mmHg, 30.8% (95% CI: 22.8-38.0%) for a 10 mmHg decrease, and 56.2% (95% CI: 43.7-65.9%) decrease for a 22.7 mmHg decrease in SBP (22.7 mmHg represents a shift from the current mean SBP to 115 mmHg).

Conclusions

Risk of many cardiovascular diseases is influenced by long-term differences in SBP. The burden of a broad range of CVDs could be substantially reduced by long-term population-wide reductions in the distribution of blood pressure.
Exponential increase in neutralizing and spike specific antibodies following vaccination of COVID-19 convalescent plasma donors
Authors: Vickers Molly A. , Sariol Alan , Leon Judith , Ehlers Alexandra , Locher Aaron V. , Dubay Kerry A. , Collins Laura , Voss Dena , Odle Abby E. , Holida Myrl , Merrill Anna E. , Perlman Stanley , Knudson C. Michael
Published: February 2021

Background

With the recent approval of COVID-19 vaccines, recovered COVID-19 subjects who are vaccinated may be ideal candidates to donate COVID-19 convalescent plasma (CCP).

Case Series

Three recovered COVID-19 patients were screened to donate CCP. All had molecularly confirmed COVID-19, and all were antibody positive by chemiluminescence immunoassay (DiaSorin) prior to vaccination. All were tested again for antibodies 11 to 21 days after they received the first dose of the vaccine (Pfizer). All showed dramatic increases (∼50 fold) in spike-specific antibody levels and had at least a 20-fold increase in the IC50 neutralizing antibody titer based on plaque reduction neutralization testing (PRNT). The spike-specific antibody levels following vaccination were significantly higher than those seen in any non-vaccinated COVID-19 subjects tested to date at our facility.

Conclusion

Spike-specific and neutralizing antibodies demonstrated dramatic increases following a single vaccination post COVID-19 infection which significantly exceeded values seen with COVID-19 infection alone. Recovered COVID-19 subjects who are vaccinated may make ideal candidates for CCP donation.
Mental Health, Substance Use, and Suicidal Ideation Among Unpaid Caregivers in the United States During the COVID-19 Pandemic: Relationships to Age, Race/Ethnicity, Employment, and Caregiver Intensity
Authors: Czeisler Mark É , Drane Alexandra , Winnay Sarah S , Capodilupo Emily R , Czeisler Charles A , Rajaratnam Shantha MW , Howard Mark E
Published: February 2021

Objectives

To estimate the prevalence of unpaid caregiving during the coronavirus disease 2019 (COVID-19) pandemic, and to identify factors associated with adverse mental health symptoms, substance use, and suicidal ideation in this population, which provides critical support in health care systems by providing care to older adults and those with chronic conditions.

Methods

In June 2020, Internet-based surveys with questions about demographics, caregiving responsibilities, and mental health were administered to US adults aged ≥18 years. Demographic quota sampling and survey weighting to improve cross-sectional sample representativeness of age, gender, and race/ethnicity. Prevalence ratios for adverse mental health symptoms were estimated using multivariable Poisson regressions.

Results

Of 9,896 eligible invited adults, 5,412 (54.7%) completed surveys; 5,011 (92.6%) respondents met screening criteria and were analysed, including 1,362 (27.2%) caregivers. Caregivers had higher prevalences of adverse mental health symptoms than non-caregivers, including anxiety or depressive disorder symptoms (57.6% vs 21.5%, respectively, p<0.0001) having recently seriously considered suicide (33.4% vs 3.7%, p<0.0001). Symptoms were more common among caregivers who were young vs older adults (e.g., aged 18–24 vs ≥65 years, aPR 2.75, 95% CI 1.95–3.88, p<0.0001), Hispanic or Latino vs non-Hispanic White (1.14, 1.04–1.25, p=0.0044), living with vs without disabilities (1.18, 1.10–1.26, p<0.0001), and with moderate and high vs low Caregiver Intensity Index scores (2.31, 1.65–3.23; 2.81, 2.00–3.94; both p<0.0001). Suicidal ideation was more prevalent among non-Hispanic Black vs non-Hispanic White caregivers (1.48, 1.15–1.90, p=0.0022).

Conclusions

Caregivers, who accounted for one in four US adult respondents in this nationally representative sample, more commonly reported adverse mental health symptoms than non-caregivers. Increased visibility of and access to mental health care resources are urgently needed to address mental health challenges of caregiving.
Recruitment location influences bias and uncertainty in SARS-CoV-2 seroprevalence estimates
Authors: Brown Tyler S. , de Salazar Munoz Pablo Martinez , Bhatia Abhishek , Bunda Bridget , Williams Ellen K. , Bor David , Miller James S. , Mohareb Amir M. , Thierauf Julia , Yang Wenxin , Villalba Julian , Naranbai Vivek , Beltran Wilfredo Garcia , Miller Tyler E. , Kress Doug , Stelljes Kristen , Johnson Keith , Larremore Daniel B. , Lennerz Jochen , Iafrate A. John , Balsari Satchit , Buckee Caroline O. , Grad Yonatan H.
Published: February 2021

The initial phase of the COVID-19 pandemic in the US was marked by limited diagnostic testing, resulting in the need for seroprevalence studies to estimate cumulative incidence and define epidemic dynamics. In lieu of systematic representational surveillance, venue-based sampling was often used to rapidly estimate a community’s seroprevalence. However, biases and uncertainty due to site selection and use of convenience samples are poorly understood. Using data from a SARS-CoV-2 serosurveillance study we performed in Somerville, Massachusetts, we found that the uncertainty in seroprevalence estimates depends on how well sampling intensity matches the known or expected geographic distribution of seropositive individuals in the study area. We use GPS-estimated foot traffic to measure and account for these sources of bias. Our results demonstrated that study-site selection informed by mobility patterns can markedly improve seroprevalence estimates. Such data should be used in the design and interpretation of venue-based serosurveillance studies.
Age-related heterogeneity in immune responses to SARS-CoV-2 vaccine BNT162b2
Authors: Collier Dami A. , Ferreira Isabella A.T.M. , Kotagiri Prasanti , Datir Rawlings , Lim Eleanor , Touzier Emma , Meng Bo , Abdullahi Adam ,   The CITIID-NIHR BioResource COVID-19 Collaboration, Elmer Anne , Kingston Nathalie , Graves Barbara , Gresley Emma Le , Caputo Daniela , Smith Kenneth GC , Bradley John R. , Ceron-Gutierrez Lourdes , Cortes-Acevedo Paulina , Barcenas-Morales Gabriela , Linterman Michelle , McCoy Laura , Davis Chris , Thomson Emma , Lyons Paul A. , McKinney Eoin , Doffinger Rainer , Wills Mark , Gupta Ravindra K.
Published: February 2021

Two dose mRNA vaccination provides excellent protection against SARS-CoV-2. However, there are few data on vaccine efficacy in elderly individuals above the age of 801. Additionally, new variants of concern (VOC) with reduced sensitivity to neutralising antibodies have raised fears for vulnerable groups. Here we assessed humoral and cellular immune responses following vaccination with mRNA vaccine BNT162b22 in elderly participants prospectively recruited from the community and younger health care workers. Median age was 72 years and 51% were females amongst 140 participants. Neutralising antibody responses after the first vaccine dose diminished with increasing age, with a marked drop in participants over 80 years old. Sera from participants below and above 80 showed significantly lower neutralisation potency against B.1.1.7, B.1.351 and P.1. variants of concern as compared to wild type. Those over 80 were more likely to lack any neutralisation against VOC compared to younger participants following first dose. The adjusted odds ratio for inadequate neutralisation activity against the B.1.1.7, P.1 and B.1.351 variant in the older versus younger age group was 4.3 (95% CI 2.0-9.3, p<0.001), 6.7 (95% CI 1.7-26.3, p=0.008) and 1.7 (95% CI 0.5-5.7, p=0.41). Binding IgG and IgA antibodies were lower in the elderly, as was the frequency of SARS-CoV-2 Spike specific B-memory cells. We observed a trend towards lower somatic hypermutation in participants with suboptimal neutralisation, and elderly participants demonstrated clear reduction in class switched somatic hypermutation, driven by the IgA1/2 isotype. SARS-CoV-2 Spike specific T-cell IFNγ and IL-2 responses fell with increasing age, and both cytokines were secreted primarily by CD4 T cells. We conclude that the elderly are a high risk population that warrant specific measures in order to mitigate against vaccine failure, particularly where variants of concern are circulating.
Affective neural signatures do not distinguish women with emotion dysregulation from healthy controls: A mega-analysis across three task-based fMRI studies
Authors: Sicorello M. , Herzog J. , Wager T.D. , Ende G. , Müller-Engelmann M. , Herpertz S.C. , Bohus M. , Schmahl C. , Paret C. , Niedtfeld I.
Published: February 2021

Pathophysiological models are urgently needed for personalized treatments of mental disorders. However, most potential neural markers for psychopathology are limited by low interpretability, prohibiting reverse inference from brain measures to clinical symptoms and traits. Neural signatures—i.e. multivariate brain-patterns trained to be both sensitive and specific to a construct of interest—might alleviate this problem, but are rarely applied to mental disorders. We tested whether previously developed neural signatures for negative affect and discrete emotions distinguish between healthy individuals and those with mental disorders characterized by emotion dysregulation, i.e. Borderline Personality Disorder (BPD) and complex Post-traumatic Stress Disorder (cPTSD). In three different fMRI studies, a total sample of 192 women (49 BPD, 62 cPTSD, 81 healthy controls) were shown pictures of scenes with negative or neutral content. Based on pathophysiological models, we hypothesized higher negative and lower positive reactivity of neural emotion signatures in participants with emotion dysregulation. The expression of neural signatures differed strongly between neutral and negative pictures (average Cohen’s d = 1.17). Nevertheless, a mega-analysis on individual participant data showed no differences in the reactivity of neural signatures between participants with and without emotion dysregulation. Confidence intervals ruled out even small effect sizes in the hypothesized direction and were further supported by Bayes factors. Overall, these results support the validity of neural signatures for emotional states during fMRI tasks, but raise important questions concerning their link to individual differences in emotion dysregulation.
Deep Learning Prediction of Biomarkers from Echocardiogram Videos
Authors: Hughes J Weston , Yuan Neal , He Bryan , Ouyang Jiahong , Ebinger Joseph , Botting Patrick , Lee Jasper , Theurer John , Tooley James E. , Neiman Koen , Lungren Matthew P. , Liang David , Schnittger Ingela , Harrington Bob , Chen Jonathan H. , Ashley Euan A. , Cheng Susan , Ouyang David , Zou James Y.
Published: February 2021

Laboratory blood testing is routinely used to assay biomarkers to provide information on physiologic state beyond what clinicians can evaluate from interpreting medical imaging. We hypothesized that deep learning interpretation of echocardiogram videos can provide additional value in understanding disease states and can predict common biomarkers results. Using 70,066 echocardiograms and associated biomarker results from 39,460 patients, we developed EchoNet-Labs, a video-based deep learning algorithm to predict anemia, elevated B-type natriuretic peptide (BNP), troponin I, and blood urea nitrogen (BUN), and abnormal levels in ten additional lab tests. On held-out test data across different healthcare systems, EchoNet-Labs achieved an area under the curve (AUC) of 0.80 in predicting anemia, 0.82 in predicting elevated BNP, 0.75 in predicting elevated troponin I, and 0.69 in predicting elevated BUN. We further demonstrate the utility of the model in predicting abnormalities in 10 additional lab tests. We investigate the features necessary for EchoNet-Labs to make successful predictions and identify potential prediction mechanisms for each biomarker using well-known and novel explainability techniques. These results show that deep learning applied to diagnostic imaging can provide additional clinical value and identify phenotypic information beyond current imaging interpretation methods.
Age-related susceptibility to insulin resistance is due to a combination of CPT1B decline and lipid overload
Authors: Vieira-Lara Marcel A. , Dommerholt Marleen B. , Zhang Wenxuan , Blankestijn Maaike , Wolters Justina C. , Abegaz Fentaw , Gerding Albert , van der Veen Ydwine , van Dijk Theo H. , van Os Ronald , Reijngoud Dirk-Jan , Jonker Johan W. , Kruit Janine K. , Bakker Barbara M.
Published: February 2021

BACKGROUND

Advanced age increases the susceptibility to diet-induced insulin resistance (IR). A key driver of this phenomenon is lipid accumulation in the skeletal muscle. It is debated, however, whether this is due to dietary lipid overload or decline of mitochondrial function. To address the interplay of diet and age in the flexibility of muscle lipid and glucose handling, we put young and aged mice on a low- or high-fat diet (HFD).

RESULTS

As expected, aged mice were more susceptible to IR when given a HFD than young mice. The HFD induced intramuscular lipid accumulation specifically in aged mice, including C18:0-containing ceramides and diacylglycerols. This was reflected by the mitochondrial β-oxidation capacity, which was upregulated by the HFD in young, but not in old mice. Conspicuously, most β-oxidation proteins were upregulated by the HFD in both groups, but carnitine palmitoyltransferase 1B (CPT1B) declined in aged animals. Computational modelling traced the flux control mostly to CPT1B, suggesting a CPT1B-driven loss of flexibility to the HFD with age. Finally, in old animals glycolytic protein levels were reduced and less flexible to the diet.

CONCLUSION

We conclude that intramuscular lipid accumulation and decreased insulin sensitivity are not due to age-related mitochondrial dysfunction or nutritional overload alone, but rather to their interaction. Moreover, we identify CPT1B as a potential target to counteract age-dependent intramuscular lipid accumulation and thereby IR.
The epigenetic landscape in purified myonuclei from fast and slow muscles
Authors: Bengtsen Mads , Winje Ivan Myhre , Eftestøl Einar , Landskron Johannes , Sun Chengyi , Domanska Diana , Millay Douglas P. , Meza-Zepeda Leonardo , Gundersen Kristian
Published: February 2021

Muscle cells have different phenotypes adapted to different usage and can be grossly divided into fast/glycolytic and slow/oxidative types. While most muscles contain a mixture of such fiber types, we aimed at providing a genome-wide analysis of chromatin environment by ChIP-Seq in two muscle extremes, the almost completely fast/glycolytic extensor digitorum longus (EDL) and slow/oxidative soleus muscles. Muscle is a heterogeneous tissue where less than 60% of the nuclei are inside muscle fibers. Since cellular homogeneity is critical in epigenome-wide association studies we devised a new method for purifying skeletal muscle nuclei from whole tissue based on the nuclear envelope protein Pericentriolar material 1 (PCM1) being a specific marker for myonuclei. Using antibody labeling and a magnetic-assisted sorting approach we were able to sort out myonuclei with 95% purity. The sorting eliminated influence from other cell types in the tissue and improved the myo-specific signal. A genome-wide comparison of the epigenetic landscape in EDL and soleus reflected the functional properties of the two muscles each with a distinct regulatory program involving distal enhancers, including a glycolytic super-enhancer in the EDL. The two muscles are also regulated by different sets of transcription factors; e.g. in soleus binding sites for MEF2C, NFATC2 and PPARA were enriched, while in EDL MYOD1 and SOX1 binding sites were found to be overrepresented. In addition, novel factors for muscle regulation such as MAF, ZFX and ZBTB14 were identified.
Habitat complexity in organic olive orchards modulates the abundance of natural enemies but not the attraction to plant species
Authors: Álvarez Hugo Alejandro , Morente Marina , Ruano Francisca
Published: February 2021

Semi-natural habitat complexity and organic management could affect the abundance and diversity of natural enemies and pollinators in olive orchards. Nonetheless, in such agroecosystems the effect of plant structure, plant richness, and plant attraction on the arthropod fauna has been poorly documented. Here we evaluate the influence of those effects jointly as an expression of arthropod abundance and richness in olive trees, ground cover, and adjacent vegetation within organic olive orchards. For this, we used generalized linear models and non-metric multidimensional scaling (NMDS) integrating generalized additive models. Our results suggest that natural enemies and pollinators are mainly attracted toA. radiatus,D. catholica, andL. longirrostriswithin ground cover andG. cinerea speciosa,Q. rotundifolia,R. officinalis,T. zygis gracilis, andU. parvifloruswithin adjacent vegetation. Accordingly, habitat complexity showed a positive relationship with the abundance of key families of natural enemies and pollinators but not with the number of taxa. NMDS showed that plant richness and plant arrangement and scattering affected the key families differently, suggesting that each key family responds to their individual needs for plant resources but forming groups modulated by complexity. This pattern was especially seeing in predators and omnivores. Our findings support that the higher the plant richness and structure of a semi natural-habitat within an olive orchard, the higher the abundance and richness of a given arthropod community (a pattern found in natural ecosystems). The information presented here can be used by producers and technicians to increase the presence and abundance of natural enemies and pollinators within organic olive orchards, and thus improve the ecosystem services provided by semi-natural habitats.

Graphical abstract
Inhibitory synaptic transmission is impaired at higher extracellular Ca2+concentrations inScn1a+/−mouse model of Dravet syndrome
Authors: Uchino Kouya , Kawano Hiroyuki , Tanaka Yasuyoshi , Adaniya Yuna , Asahara Ai , Deshimaru Masanobu , Kubota Kaori , Watanabe Takuya , Katsurabayashi Shutaro , Iwasaki Katsunori , Hirose Shinichi
Published: February 2021

Dravet syndrome (DS) is an intractable form of childhood epilepsy that occurs in infancy. More than 80% of all patients have a heterozygous abnormality in theSCN1Agene, which encodes a subunit of Na+channels in the brain. However, the detailed pathogenesis of DS remains unclear. This study investigated the synaptic pathogenesis of this disease in terms of excitatory/inhibitory balance using a mouse model of DS. We show that excitatory postsynaptic currents were similar betweenScn1aknock-in neurons (Scn1a+/–neurons) and wild-type neurons, but inhibitory postsynaptic currents were significantly lower inScn1a+/–neurons. Moreover, both the vesicular release probability and the number of inhibitory synapses were significantly lower inScn1a+/–neurons compared with wild-type neurons. There was no proportional increase in inhibitory postsynaptic current amplitude in response to increased extracellular Ca2+concentrations. Our study revealed that the number of inhibitory synapses is significantly reduced inScn1a+/–neurons, while the sensitivity of inhibitory synapses to extracellular Ca2+concentrations is markedly increased. These data suggest that Ca2+tethering in inhibitory nerve terminals may be disturbed following the synaptic burst, likely leading to epileptic symptoms.
Extensive characterization of HIV-1 reservoirs reveals links to plasma viremia before and during analytical treatment interruption
Authors: Cole Basiel , Lambrechts Laurens , Boyer Zoe , Noppe Ytse , De Scheerder Marie-Angélique , Eden John-Sebastian , Vrancken Bram , Schlub Timothy E. , McLaughlin Sherry , Frenkel Lisa M. , Palmer Sarah , Vandekerckhove Linos
Published: February 2021

The HIV-1 reservoir is composed of cells harboring latent proviruses that are capable of contributing to viremia upon antiretroviral treatment (ART)interruption. Although this reservoir is known to be maintained by clonal expansion, the contribution of large, infected cell clones to residual viremia and viral rebound remains underexplored. Here, we conducted an extensive analysis on four ART-treated individuals who underwent an analytical treatment interruption (ATI). We performed subgenomic (V1-V3env), near full-length proviral and integration site sequencing, and used multiple displacement amplification to sequence both the integration site and provirus from single HIV-infected cells. We found eight proviruses that could phylogenetically be linked to plasma virus obtained before or during the ATI. This study highlights a role for HIV-infected cell clones in the maintenance of the replication-competent reservoir and suggests that infected cell clones can directly contribute to rebound viremia upon ATI.
Analytical Performance of a Standardized Kit for Mass Spectrometry-based Measurements of Human Glycosaminoglycans
Authors: Tamburro Davide , Bratulic Sinisa , Shameh Souad Abou , Soni Nikul K , Bacconi Andrea , Maccari Francesca , Galeotti Fabio , Mattsson Karin , Volpi Nicola , Nielsen Jens , Gatto Francesco
Published: February 2021

Glycosaminoglycans (GAGs) are long linear sulfated polysaccharides implicated in processes linked to disease development such as mucopolysaccharidosis, respiratory failure, cancer, and viral infections, thereby serving as potential biomarkers. A successful clinical translation of GAGs as biomarkers depends on the availability of standardized GAG measurements. However, owing to the analytical complexity associated with the quantification of GAG concentration and structural composition, a standardized method to simultaneously measure multiple GAGs is missing. In this study, we sought to characterize the analytical performance of a ultra-high-performance liquid chromatography coupled with triple-quadrupole tandem mass spectrometry (UHPLC-MS/MS)-based kit for the quantification of 17 GAG disaccharides. The kit showed acceptable linearity, selectivity and specificity, accuracy and precision, and analyte stability in the absolute quantification of 15 GAG disaccharides. In native human samples, here using urine as a reference matrix, the analytical performance of the kit was acceptable for the quantification of CS disaccharides. Intra- and inter-laboratory tests performed in an external laboratory demonstrated robust reproducibility of GAG measurements showing that the kit was acceptably standardized. In conclusion, these results indicated that the UHPLC-MS/MS kit was standardized for the simultaneous measurement of GAG disaccharides allowing for comparability of measurements and enabling translational research.

Analytical performance of a kit for standardized GAG measurements, based on an established UHPLC-MS/MS method
Selection on growth rate and local adaptation drive genomic adaptation during experimental range expansions in the protistTetrahymena thermophila
Authors: Moerman Felix , Fronhofer Emanuel A. , Altermatt Florian , Wagner Andreas
Published: February 2021

Populations that expand their range can undergo rapid evolutionary adaptation of life-history traits, dispersal behaviour, and adaptation to the local environment. Such adaptation may be aided or hindered by sexual reproduction, depending on the context.

However, few studies have investigated the genomic causes and consequences or genetic architecture of such adaptation during range expansions.

We here studied genomic adaptation during experimental range expansions of the protistTetrahymena thermophilain landscapes with a uniform environment or a pH-gradient. Specifically, we investigated two aspects of genomic adaptation during range expansion. Firstly, we investigated the genetic architecture of adaptation in terms of the underlying numbers of allele frequency changes from standing genetic variation andde novovariants. We focused on how sexual reproduction may alter this genetic architecture. Secondly, identified genes subject to selection caused by the expanding range itself, and directional selection due to the presence or absence of the pH-gradient. We focused this analysis on alleles with large frequency changes that occurred in parallel in more than one population to identify the most likely candidate targets of selection.

We found that sexual reproduction altered genetic architecture both in terms ofde novovariants and standing genetic variation. However, sexual reproduction affected allele frequency changes in standing genetic variation only in the absence of long-distance gene flow. Adaptation to the range expansion affected genes involved in cell divisions and DNA repair, whereas adaptation to the pH-gradient additionally affected genes involved in ion balance, and oxidoreductase reactions. These genetic changes may result from selection on growth and adaptation to low pH.

Our results suggest that the evolution of life-history and the adaptation to the local environment has a genetic basis during our range expansion experiment. In the absence of gene flow, sexual reproduction may have aided genetic adaptation. Gene flow may have swamped expanding populations with maladapted alleles, thus reducing the extent of evolutionary adaptation during range expansion. Sexual reproduction also altered the genetic architecture of our evolving populations viade novovariants, possibly by purging deleterious mutations or by revealing fitness benefits of rare genetic variants.
Chronic effects of short-term aerobic and anaerobic physical training on the ventral prostate of adult rats
Authors: Borges Rodrigo Miranda Ramos , Rocha Emerson Souza da , de Andrade Edila Monteiro , da Silva Nazaré Nagaywer Edno , Ortega Araújo Paulo Afonso , Cembraneli Pedro Nogarotto , Rocha Ruiz Thalles Fernando , Colleta Simone Jacovaci , Carlos Carla Patricia , Marques Vanessa Belentani , Taboga Sebastião Roberto , de Campos Gomes Fabiana , de Melo-Neto João Simão
Published: February 2021

Aims

To analyze the chronic effects of short-term aerobic and anaerobic physical training on prostate compartments, extracellular matrix, microvascularization, TGFβ, cyclooxygenase-2, inducible nitric oxide synthase (iNOS) and macrophage immunostaining, and ventral prostate histopathology in adult rats.

Main methods

Fifteen male rats (90 days old) were divided into three groups (n = 5/group): sedentary, aerobic (AE) (swimming), and anaerobic physical training (AN) (jumping), performed three days per week, for 8-weeks. The animals were sacrificed within 21 weeks of age. The ventral prostate was collected, weighed, and processed for histological and immunohistochemical analyses.

Key findings

Our results show that AE increases blood capillaries and reduces the percentage and increases the thickness of smooth muscle. AN promotes increased iNOS levels in the prostatic tissue, and both modalities reduce TGFβ and elastic fibers, in addition to being protective against benign prostatic hyperplasia and atrophy.

Significance

Different physical training modalities can activate specific mechanisms that modify the prostate environment.
New loci and neuronal pathways for resilience to heat stress in animals
Authors: Cheruiyot Evans K. , Haile-Mariam Mekonnen , Cocks Benjamin G. , MacLeod Iona M. , Xiang Ruidong , Pryce Jennie E.
Published: February 2021

Climate change and resilience to warming climates have implications for humans, livestock, and wildlife. The genetic mechanisms that confer thermotolerance to mammals are still not well characterized. We used dairy cows as a model to study heat tolerance because they are lactating, and therefore often prone to thermal stress. The data comprised almost 0.5 million milk records (milk, fat, and proteins) of 29,107 Australian Holsteins, each having around 15 million imputed sequence variants. Dairy animals often reduce their milk production when temperature and humidity rise; thus, the phenotypes used to measure an individual’s heat tolerance were defined as the rate of milk production decline (slope traits) with a rising temperature-humidity index. With these slope traits, we performed a genome-wide association study (GWAS) using different approaches, including conditional analyses, to correct for the relationship between heat tolerance and level of milk production. The results revealed multiple novel loci for heat tolerance, including 61 potential functional variants at sites highly conserved across vertebrate species. Moreover, it was interesting that specific candidate variants and genes are related to the neuronal system (ITPR1, ITPR2,andGRIA4) and neuroactive ligand-receptor interaction functions for heat tolerance (NPFFR2, CALCR,andGHR), providing a novel insight that can help to develop genetic and management approaches to combat heat stress.

Author summary

While understanding the genetic basis of heat tolerance is crucial in the context of global warming’s effect on humans, livestock, and wildlife, the specific genetic variants and biological features that confer thermotolerance in animals are still not well characterized. The ability to tolerate heat varies across individuals, with substantial genetic control of this complex trait. Dairy cattle are excellent model in which to find genes associated with individual variations in heat tolerance since they significantly suffer from heat stress due to the metabolic heat of lactation. By genome-wide association studies of more than 29,000 cows with 15 million sequence variants and controlled phenotype measurements, we identify many new loci associated with heat tolerance. The biological functions of these loci are linked to the neuronal system and neuroactive ligand-receptor interaction functions. Also, several putative causal mutations for heat tolerance are at genomic sites that are otherwise evolutionarily conserved across 100 vertebrate species. Overall, our findings provide new insight into the molecular and biological basis of heat tolerance that can help to develop genetic and management approaches to combat heat stress.
Type-I interferon signatures in SARS-CoV-2 infected Huh7 cells
Authors: Chen Xi , Saccon Elisa , Appelberg K. Sofia , Mikaeloff Flora , Rodriguez Jimmy Esneider , Vinhas Beatriz Sá , Frisan Teresa , Végvári Ákos , Mirazimi Ali , Neogi Ujjwal , Gupta Soham
Published: February 2021

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that causes Coronavirus disease 2019 (COVID-19) has caused a global health emergency. A key feature of COVID-19 is dysregulated interferon-response. Type-I interferon (IFN-I) is one of the earliest antiviral innate immune responses following viral infection and plays a significant role in the pathogenesis of SARS-CoV-2. In this study, using a proteomics-based approach, we identified that SARS-CoV-2 infection induces delayed and dysregulated IFN-I signaling in Huh7 cells. We demonstrate that SARS-CoV-2 is able to inhibit RIG-I mediated IFN-β production. Our results also confirm the recent findings that IFN-I pretreatment is able to reduce susceptibility of Huh7 cells to SARS-CoV-2, but not post-treatment. Moreover, senescent Huh7 cells, in spite of showing accentuated IFN-I response were more susceptible to SARS-CoV-2 infection, and the virus effectively inhibited IFIT1 in these cells. Finally, proteomic comparison between SARS-CoV-2, SARS-CoV and MERS-CoV revealed a distinct differential regulatory signature of interferon-related proteins emphasizing that therapeutic strategies based on observations in SARS-CoV and MERS-CoV should be used with caution. Our findings provide a better understanding of SARS-CoV-2 regulation of cellular interferon response and a perspective on its use as a treatment. Investigation of different interferon stimulated genes and their role in inhibition of SARS-CoV-2 pathogenesis may direct novel antiviral strategies.
Targetedin situcross-linking mass spectrometry and integrative modeling reveal the architectures of Nsp1, Nsp2, and Nucleocapsid proteins from SARS-CoV-2
Authors: Slavin Moriya , Zamel Joanna , Zohar Keren , Eliyahu Siona , Braitbard Merav , Brielle Esther , Baraz Leah , Stolovich-Rain Miri , Friedman Ahuva , Wolf Dana G , Rouvinski Alexander , Linial Michal , Schneidman-Duhovny Dina , Kalisman Nir
Published: February 2021

Atomic structures of several proteins from the coronavirus family are still partial or unavailable. A possible reason for this gap is the instability of these proteins outside of the cellular context, thereby prompting the use of in-cell approaches.In situcross-linking and mass spectrometry (in situCLMS) can provide information on the structures of such proteins as they occur in the intact cell. Here, we applied targetedin situCLMS to structurally probe Nsp1, Nsp2, and Nucleocapsid (N) proteins from SARS-CoV-2, and obtained cross-link sets with an average density of one cross-link per twenty residues. We then employed integrative modeling that computationally combined the cross-linking data with domain structures to determine full-length atomic models. For the Nsp2, the cross-links report on a complex topology with long-range interactions. Integrative modeling with structural prediction of individual domains by the AlphaFold2 system allowed us to generate a single consistent all-atom model of the full-length Nsp2. The model reveals three putative metal binding sites, and suggests a role for Nsp2 in zinc regulation within the replication-transcription complex. For the N protein, we identified multiple intra- and inter-domain cross-links. Our integrative model of the N dimer demonstrates that it can accommodate three single RNA strands simultaneously, both stereochemically and electrostatically. For the Nsp1, cross-links with the 40S ribosome were highly consistent with recent cryo-EM structures. These results highlight the importance of cellular context for the structural probing of recalcitrant proteins and demonstrate the effectiveness of targetedin situCLMS and integrative modeling.
Speciation of theMycobacterium tuberculosiscomplex using mass spectrometry: a proof-of-concept
Authors: Robinne Simon , Saad Jamal , Morsli Madjid , Harouna Hamidou Zelika , Tazerart Fatah , Drancourt Michel , Baron Sophie Alexandra
Published: February 2021

Mycobacteria that form theMycobacterium tuberculosiscomplex are responsible for deadly tuberculosis in animals and patients. Identification of these pathogens at the species level is of primary importance for treatment and source tracing, and currently relies on DNA analysis, including whole genome sequencing (WGS), which takes a whole day. In this study, we report on the unprecedented identification of theM. tuberculosiscomplex species using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF-MS), with WGS as the comparative gold standard. In a first step, an optimised peptide extraction applied to 24 isolates otherwise identified in three of the 11M. tuberculosiscomplex species by WGS, yielded 94 MALDI-TOF spectra which clustered according to WGS identification. In a second step, 70/74 (95%) other isolates were correctly identified at the species level by this clustering method. This study is the first to report a MALDI-TOF-MS method of identification ofM. tuberculosiscomplex mycobacteria at the species level and is easily implantable in clinical microbiology laboratories.
Perturbation of BRMS1 interactome reveals pathways that impact cell migration
Authors: Zimmermann Rosalyn , Sardiu Mihaela E. , Manton Christa A. , Miah Md. Sayem , Banks Charles A.S. , Adams Mark K. , Koestler Devin C. , Hurst Douglas R. , Edmonds Mick D. , Washburn Michael P. , Welch Danny R.
Published: February 2021

Breast Cancer Metastasis Suppressor 1 (BRMS1) expression is associated with longer patient survival in multiple cancer types. Understanding BRMS1 functionality will provide insights into both mechanism of action and will enhance potential therapeutic development. In this study, we confirmed that the C-terminus of BRMS1 is critical for metastasis suppression and hypothesized that critical protein interactions in this region would explain its function. Phosphorylation status at S237 regulates BRMS1 protein interactions related to a variety of biological processes, phenotypes [cell cycle (e.g., CDKN2A), DNA repair (e.g., BRCA1)], and metastasis [(e.g., TCF2 and POLE2)]. Presence of S237 also directly decreased MDA-MB-231 breast carcinoma migrationin vitroand metastasesin vivo. The results add significantly to our understanding of how BRMS1 interactions with Sin3/HDAC complexes regulate metastasis and expand insights into BRMS1’s molecular role, as they demonstrate that BRMS1 C-terminus involvement in distinct direct protein-protein interactions.
Escherichia coliS2P family intramembrane protease RseP is engaged in the regulated sequential cleavages of FecR in the ferric citrate signaling
Authors: Yokoyama Tatsuhiko , Niinae Tomoya , Tsumagari Kazuya , Imami Koshi , Ishihama Yasushi , Hizukuri Yohei , Akiyama Yoshinori
Published: February 2021

Escherichia coliRseP, a member of the S2P family of intramembrane proteases, is involved in the activation of the σEextracytoplasmic stress response and elimination of remnant signal peptides. However, whether RseP has additional cellular functions is unclear. In this study, we attempted to identify new RseP substrates to explore still unknown physiological roles of this protease. Our mass spectrometry-based quantitative proteomic analysis revealed that the levels of several Fec system proteins encoded by thefecABCDEoperon (fecoperon) were significantly decreased in an RseP-deficient strain. The Fec system is responsible for the uptake of ferric citrate, and the transcription of thefecoperon is controlled by FecI, an alternative sigma factor, and its regulator FecR, a single-pass transmembrane protein. Assays with thefecoperon expression reporter demonstrated that the proteolytic activity of RseP is essential for the ferric citrate-dependent upregulation of thefecoperon. Analysis using the FecR protein and FecR-derived model proteins showed that FecR undergoes sequential processing at the membrane and that RseP participates in the last step of this sequential processing to generate the N-terminal cytoplasmic fragment of FecR that participates in the transcription of thefecoperon with FecI. Ferric citrate signal-dependent generation of this cleavage product is the essential and sufficient role of RseP in the transcriptional activation of thefecoperon. Our study unveiled thatE. coliRseP performs the intramembrane proteolysis of FecR, a novel physiological role that is essential for regulating iron uptake by the ferric citrate transport system.
In-planta Gene Targeting in Barley using Cas9, with and without Geminiviral Replicons
Authors: Lawrenson Tom , Hinchliffe Alison , Clarke Martha , Morgan Yvie , Harwood Wendy
Published: February 2021

Advances in the use of RNA-guided Cas9-based genome editing in plants have been rapid over the last few years. A desirable application of genome editing is gene targeting (GT), as it allows a wide range of precise modifications, however this remains inefficient especially in key crop species. Here we describe successful, heritable gene targeting in barley using anin-plantastrategy but fail to achieve the same using a wheat dwarf virus replicon to increase copy number of the repair template. Without the replicon, we were able to delete 150bp of the coding sequence of our target gene whilst simultaneously fusing in-frame mCherry in its place. Starting from 14 original transgenic plants, two plants appeared to have the required gene targeting event. From one of these T0 plants, three independent gene targeting events were identified, two of which were heritable. When the replicon was included, 39 T0 plants were produced and shown to have high copy numbers of the repair template. However, none of the 17 lines screened in T1 gave rise to significant or heritable gene targeting events despite screening twice the number of plants in T1 compared to the non-replicon strategy. Investigation indicated that high copy numbers of repair template created by the replicon approach cause false positive PCR results which are indistinguishable at the sequence level to true GT events in junction PCR screens widely used in GT studies. In the successful non-replicon approach, heritable gene targeting events were obtained in T1 and subsequently the T-DNA was found to be linked to the targeted locus. Thus, physical proximity of target and donor sites may be a factor in successful gene targeting.
Missing the sweet spot: one of the two N-glycans on human Gb3/CD77 synthase is expendable
Authors: Mikolajczyk Krzysztof , Bereznicka Anna , Szymczak-Kulus Katarzyna , Haczkiewicz-Lesniak Katarzyna , Szulc Bozena , Olczak Mariusz , Rossowska Joanna , Majorczyk Edyta , Kapczynska Katarzyna , Bovin Nicolai , Lisowska Marta , Kaczmarek Radoslaw , Miazek Arkadiusz , Czerwinski Marcin
Published: February 2021

N-glycosylation is a ubiquitous posttranslational modification that may influence folding, subcellular localization, secretion, solubility and oligomerization of proteins. In this study, we examined the effects of N-glycans on the activity of human Gb3/CD77 synthase, which catalyzes the synthesis of glycosphingolipids with terminal Galα1→4Gal (Gb3 and the P1 antigen) and Galα1→4GalNAc disaccharides (the NOR antigen). The human Gb3/CD77 synthase contains two occupied N-glycosylation sites at positions N121and N203. Intriguingly, we found that while the N-glycan at N203is essential for activity and correct subcellular localization, the N-glycan at N121is dispensable and its absence did not reduce, but, surprisingly, even increased the activity of the enzyme. The fully N-glycosylated human Gb3/CD77 synthase and its glycoform missing the N121glycan correctly localized in the Golgi, whereas a glycoform without the N203site partially mislocalized in the endoplasmic reticulum. A double mutein missing both N-glycans was inactive and accumulated in the endoplasmic reticulum. Our results suggest that the decreased specific activity of human Gb3/CD77 synthase glycovariants results from their improper subcellular localization and, to a smaller degree, a decrease in enzyme solubility. Taken together, our findings show that the two N-glycans of human Gb3/CD77 synthase have opposing effects on its properties, revealing a dual nature of N-glycosylation and potentially a novel regulatory mechanism controlling the biological activity of proteins.