bluuebunny/crossref_metadata_2025 huggingface dataset:
Early transfusion of a large cohort of COVID-19 patients with high titer anti-SARS-CoV-2 spike protein IgG convalescent plasma confirms a signal of significantly decreased mortality
Authors: Salazar Eric , Christensen Paul A. , Graviss Edward A. , Nguyen Duc T. , Castillo Brian , Chen Jian , Lopez Bevin Valdez , Eagar Todd N. , Yi Xin , Zhao Picheng , Rogers John , Shehabeldin Ahmed , Joseph David , Masud Faisal , Leveque Christopher , Olsen Randall J. , Bernard David W. , Gollihar Jimmy , Musser James M.
Published: October 2020

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 remains a global threat with few proven efficacious treatments. Transfusion of convalescent plasma collected from donors who have recovered from COVID-19 disease has emerged as a promising therapy and has been granted emergency use authorization by the U.S. Food and Drug Administration (FDA). We recently reported results from interim analysis of a propensity-score matched study suggesting that early treatment of COVID-19 patients with convalescent plasma containing high titer anti-spike protein receptor binding domain (RBD) IgG significantly decreases mortality. We here present results from 60-day follow up of our cohort of 351 transfused hospitalized patients. Prospective determination of ELISA anti-RBD IgG titer facilitated selection and transfusion of the highest titer units available. Retrospective analysis by the Ortho VITROS IgG assay revealed a median signal/cutoff (S/C) ratio of 24.0 for transfused units, a value far exceeding the recently FDA-required cutoff of 12.0 for designation of high titer convalescent plasma. With respect to altering mortality, our analysis identified an optimal window of 44 hours post-hospitalization for transfusing COVID-19 patients with high titer convalescent plasma. In the aggregate, the analysis confirms and extends our previous preliminary finding that transfusion of COVID-19 patients soon after hospitalization with high titer anti-spike protein RBD IgG present in convalescent plasma significantly reduces mortality.
CD8+T-cell landscape in Indigenous and non-Indigenous people restricted by influenza mortality-associated HLA-A*24:02 allomorph
Authors: Hensen Luca , Illing Patricia T. , Clemens E. Bridie , Nguyen Thi H.O. , Koutsakos Marios , van de Sandt Carolien E. , Mifsud Nicole A. , Nguyen Andrea , Szeto Christopher , Chua Brendon Y. , Halim Hanim , Rizzetto Simone , Luciani Fabio , Loh Liyen , Grant Emma J. , Saunders Phillipa M. , Brooks Andrew G , Rockman Steve , Kotsimbos Tom C. , Cheng Allen C. , Richards Michael , Westall Glen P. , Wakim Linda M. , Loudovaris Thomas , Mannering Stuart I. , Elliott Michael , Tangye Stuart G. , Jackson David C , Flanagan Katie L , Rossjohn Jamie , Gras Stephanie , Davies Jane , Miller Adrian , Tong Steven Y.C. , Purcell Anthony W. , Kedzierska Katherine
Published: October 2020

Indigenous people worldwide are at high-risk of developing severe influenza disease. HLA-A*24:02 allele, highly prevalent in Indigenous populations, is associated with influenza-induced mortality, although the basis for this association is unclear. We defined CD8+T-cell immune landscapes against influenza A (IAV) and B (IBV) viruses in HLA-A*24:02-expressing Indigenous and non-Indigenous individuals, human tissues, influenza-infected patients and HLA-A*24:02-transgenic mice. We identified immunodominant protective CD8+T-cell epitopes, one towards IAV and six towards IBV, with A24/PB2550-558-specific CD8+T-cells cells being cross-reactive between IAV and IBV. Memory CD8+T-cells towards these specificities were present in blood (CD27+CD45RA-phenotype) and tissues (CD103+CD69+phenotype) of healthy subjects, and effector CD27-CD45RA-PD-1+CD38+CD8+T-cells in IAV/IBV patients. Our data present the first evidence of influenza-specific CD8+T-cell responses in Indigenous Australians, and advocate for T-cell-mediated vaccines that target and boost the breadth of IAV/IBV-specific CD8+T-cells to protect high-risk HLA-A*24:02-expressing Indigenous and non-Indigenous populations from severe influenza disease.

One Sentence Summary

Influenza-specific CD8+T-cell specificities restricted by HLA-A*24:02.
Validation of self-collected buccal swab and saliva as a diagnostic tool for COVID-19
Authors: Ku Chee Wai , Shivani Durai , Kwan Jacqueline Q T , Loy See Ling , Erwin Christina , Ko Karrie K K , Ng Xiang Wen , Oon Lynette , Thoon Koh Cheng , Kalimuddin Shirin , Chan Jerry KY
Published: October 2020

Background

Effective management of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) requires large-scale testing. Collection of nasopharyngeal swab (NPS) by healthcare workers (HCW) is currently used to diagnose SARS-CoV-2, which increases the risk of transmission to HCWs. Self-administered saliva and buccal swabs are convenient, painless and safe alternative sample collection methods.

Methods

A cross-sectional single centre study was conducted on 42 participants who were tested positive for SARS-CoV-2 via NPS within the past 7 days. A self-collected saliva and buccal swab and a HCW-collected NPS were obtained. Real-time polymerase chain reaction (RT-PCR) was performed and cycle threshold (CT) values were obtained. Positive percent agreement (PPA), negative percent agreement (NPA) and overall agreement (OA) were calculated for saliva and buccal swabs, as compared with NPS.

Results

Among the 42 participants, 73.8% (31/42) tested positive via any one of the 3 tests. With reference to NPS, the saliva test had PPA 66.7%, NPA 91.7% and OA 69.0%. The buccal swab had PPA 56.7%, NPA 100% and OA 73.8%. Presence of symptoms improved diagnostic accuracy. There was no statistically significant association between CT values and duration of symptom onset within the first 12 days of symptoms for all three modalities.

Conclusion

Self-collected saliva tests and buccal swabs have only moderate agreement with HCW-collected NPS swabs. Primary screening for SARS-CoV-2 may be performed with a saliva test or buccal swab, with a negative test warranting a confirmatory NPS to avoid false negatives. This combined strategy minimizes discomfort and reduces the risk of spread to the community and HCWs.
The Impact of COVID-19 on the Management of Heart Failure -A United Kingdom Patient Questionnaire Study
Authors: Sankaranarayanan R , Hartshorne-Evans N , Redmond-Lyon S , Wilson J , Essa H , Gray A , Clayton L , Barton C , Ahmed FZ , Cunnington C , Satchithananda DK , Murphy C
Published: October 2020

Aim

The coronavirus disease 2019 (COVID-19) pandemic has created significant challenges to healthcare globally, necessitating rapid restructuring of service provision. This questionnaire survey was conducted amongst adult heart failure (HF) patients in the United Kingdom (UK), to understand the impact of COVID-19 upon HF services.

Methods and Results

The survey was conducted by the Pumping Marvellous Foundation (PMF), a UK HF patient charity. “Survey Monkey” was used to disseminate the questionnaire in the PMF’s online patient group and in 10 UK hospitals (out-patient hospital and community HF clinics). 1050 responses were collected (693/1050-66% women); 55% (579/1050) were aged over 60 years. Anxiety level was significantly higher regarding COVID19 (mean 7±2.5 on anxiety scale of 0 to 10) compared to anxiety regarding HF (6.1±2.4; p<0.001). Anxiety was higher amongst patients aged ≤60 years about HF (6.3±2.2 versus 5.9±2.5 in those aged >60 years; p=0.005) and COVID-19 (7.3±2.3 versus 6.7±2.6 those aged >60 years; p<0.001). 65% respondents (686/1050) reported disruption to HF appointments (cancellation or postponement) during the lockdown period. 37% reported disruption to medication prescription services and 34% reported inability to access their HF teams promptly. 32% expressed reluctance to attend hospital (25% stated they would only attend hospital if there was no alternative and 7% stated that they would not attend hospital at all).

Conclusions

The COVID-19 pandemic has caused significant anxiety amongst HF patients regarding COVID-19 and HF. Cancellation or postponement of scheduled clinic appointments, investigations, procedures, prescription and monitoring services were implicated as sources of anxiety.
Diagnostic performance of the combined nasal and throat swab in patients admitted to hospital with suspected COVID-19
Authors: Lee Kuan Ken , Doudesis Dimitrios , Ross Daniella A. , Bularga Anda , MacKintosh Claire L. , Koch Oliver , Johannessen Ingolfur , Templeton Kate , Jenks Sara , Chapman Andrew R. , Shah Anoop S.V. , Anand Atul , Perry Meghan R. , Mills Nicholas L. ,   on behalf of the DataLoch COVID-19 Collaboration
Published: October 2020

Background

Accurate diagnosis in patients with suspected coronavirus disease 2019 (COVID-19) is essential to guide treatment and limit spread of the virus. The combined nasal and throat swab is used widely, but its diagnostic performance is uncertain.

Methods

In a prospective, multi-centre, cohort study conducted in secondary and tertiary care hospitals in Scotland, we evaluated the combined nasal and throat swab with reverse transcriptase-polymerase chain reaction (RT-PCR) for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in consecutive patients admitted to hospital with suspected COVID-19. Diagnostic performance of the index and serial tests was evaluated for a primary outcome of confirmed or probable COVID-19, and a secondary outcome of confirmed COVID-19 on serial testing. The diagnosis was adjudicated by a panel, who recorded clinical, laboratory and radiological features blinded to the test results.

Results

We enrolled 1,369 consecutive patients (68 [53-80] years, 47% women) who underwent a total of 3,822 tests (median 2 [1-3] tests per patient). The primary outcome occurred in 36% (496/1,369), of whom 65% (323/496) and 35% (173/496) had confirmed and probable COVID-19, respectively. The index test was positive in 255/496 (51%) patients with the primary outcome, giving a sensitivity and specificity of 51.4% (95% confidence interval [CI] 48.8 to 54.1%) and 99.5% (95% CI 99.0 to 99.8%). Sensitivity increased in those undergoing 2, 3 or 4 tests to 60.1% (95% CI 56.7 to 63.4%), 68.3% (95% CI 64.0 to 72.3%) and 77.6% (95% CI 72.7 to 81.9%), respectively. The sensitivity of the index test was 78.9% (95% CI 74.4 to 83.2%) for the secondary outcome of confirmed COVID-19 on serial testing.

Conclusions

In patients admitted to hospital, a single combined nasal and throat swab with RT-PCR for SARS-CoV-2 has excellent specificity, but limited diagnostic sensitivity for COVID-19. Diagnostic performance is significantly improved by repeated testing.
A systematic review of the knowledge, attitudes, and practices of physicians, health workers, and the general population about Coronavirus disease 2019 (COVID-19)
Authors: Saadatjoo Saeede , Miri Maryam , Hassanipour Soheil , Ameri Hosein , Arab-Zozani Morteza
Published: October 2020

Background

Understanding people, physicians, and healthcare workers’ knowledge, attitude, and practices (KAPs) can help to achieve the outcomes of planned behavior. The aim of this study was to investigate and synthesize the current evidence on KAPs regarding COVID-19.

Methods

We conducted a systematic search on PubMed/LitCovid, Scopus, and Web of Sciences databases for papers in the English language only, up to 1 Jul 2020. We used the Joanna Briggs Institute (JBI) checklist developed for cross-sectional studies to appraise the quality of the included studies. All stages of the review conducted by two independent reviewers and potential discrepancies solved with a consultation with a third reviewer. We reported the result as number and percentage. PROSPERO registration code: (CRD42020186755).

Results

Fifty-two studies encompassing 49786 participants were included in this review. 45.76% of the participants were male. The mean age of the participants was 32.6 years. 44.2% of the included studies were scored as good quality, 46.2% as fair quality, and remaining (9.6%) as low quality. 30.76% examined all three components of the KAPs model. The knowledge component was reported as good, fair, and poor in 59%, 34%, and 7%, respectively. Of the studies that examined the attitude component, 82% reported a positive attitude, 11% a fairly positive attitude, and 7% a negative attitude. For the practice component, 52% reported good practice, 44% fair practice, and 4% poor practice.

Conclusion

This systematic review showed that the overall KAP components in the included studies were at an acceptable level. In general, knowledge was at a good level, the attitude was positive and practice was at a fairly good level. Using an integrated international system can help better evaluate these components and compare them between countries.
Strategies for infection control and prevalence of anti-SARS-CoV-2 IgG in 4,554 employees of a university hospital in Munich, Germany
Authors: Erber Johanna , Kappler Verena , Haller Bernhard , Mijočević Hrvoje , Galhoz Ana , da Costa Clarissa Prazeres , Gebhardt Friedemann , Graf Natalia , Hoffmann Dieter , Thaler Markus , Lorenz Elke , Roggendorf Hedwig , Kohlmayer Florian , Henkel Andreas , Menden Michael P , Ruland Jürgen , Spinner Christoph D , Protzer Ulrike , Knolle Percy , Lingor Paul ,   on behalf of the SeCoMRI Study Group
Published: October 2020

Background

Hospital staff are at high risk of infection during the coronavirus disease (COVID-19) pandemic. We analysed the exposure characteristics, efficacy of protective measures, and transmission dynamics in this hospital-wide prospective seroprevalence study.

Methods and Findings

Overall, 4554 individuals were tested for anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) IgG antibodies using a chemiluminescent immunoassay. Individual risk factors, use of personal protective equipment (PPE), occupational exposure, previous SARS-CoV-2 infection, and symptoms were assessed using a questionnaire and correlated to anti-SARS-CoV-2 IgG antibody titres and PCR testing results. Odds ratios with corresponding exact 95% confidence intervals were used to evaluate associations between individual factors and seropositivity. Spatio-temporal trajectories of SARS-CoV-2-infected patients and staff mobility within the hospital were visualised to identify local hotspots of virus transmission.

The overall seroprevalence of anti-SARS-CoV-2-IgG antibody was 2.4% [95% CI 1.9–2.9]. Patient-facing staff, including those working in COVID-19 areas, had a similar probability of being seropositive as non-patient-facing staff. Prior interaction with SARS-CoV-2-infected co-workers or private contacts and unprotected exposure to COVID-19 patients increased the probability of seropositivity. Loss of smell and taste had the highest positive predictive value for seropositivity. The rate of asymptomatic SARS-CoV-2 infections was 25.9%, and higher anti-SARS-CoV-2 IgG antibody titres were observed in symptomatic individuals. Spatio-temporal hotspots of SARS-CoV-2-positive staff and patients only showed partial overlap.

Conclusions

Patient-facing work in a healthcare facility during the SARS-CoV-2 pandemic may be safe if adequate PPE and hygiene measures are applied. The high numbers of asymptomatic SARS-CoV-2 infections that escaped detection by symptomatic testing underline the value of cross-sectional seroprevalence studies. Unprotected contact is a major risk factor for infection and argues for the rigorous implementation of hygiene measures.
Artificial intelligence-driven real-time 3D surface quantification of Barrett’s oesophagus for risk stratification and therapeutic response monitoring
Authors: Ali Sharib , Bailey Adam , East James E. , Leedham Simon J. , Haghighat Maryam ,   TGU Investigators, Lu Xin , Rittscher Jens , Braden Barbara
Published: October 2020

BACKGROUND & AIMS

Barrett’s epithelium measurement using widely accepted Prague C&M criteria is highly operator dependent. By reconstructing the surface of the Barrett’s area in 3D from endoscopy video, we propose a novel methodology for measuring the C&M score automatically. This 3D reconstruction provides an extended field of view and also allows to precisely quantify the Barrett’s area including islands. We aim to assess the accuracy of the extracted measurements from phantom and demonstrate their clinical usability.

METHODS

Advanced deep learning techniques are utilised to design estimators for depth and camera pose required to map standard endoscopy video to a 3D surface model. By segmenting the Barrett’s area and locating the position of the gastro-oesophageal junction (GEJ) we measure C&M scores and the Barrett’s oesophagus areas (BOA). Experiments using a purpose-built 3D printed oesophagus phantom and high-definition video from 98 patients scored by an expert endoscopist are used for validation.

RESULTS

Endoscopic phantom video data demonstrated a 95 % accuracy with a marginal ± 1.8 mm average deviation for C&M and island measurements, while for BOA we achieved nearly 93 % accuracy with only ± 1.1 cm2 average deviation compared to the ground-truth measurements. On patient data, the C&M measurements provided by our system concord with the reference provided by expert upper GI endoscopists.

CONCLUSIONS

The proposed methodology is suitable for extracting Prague C&M scores automatically with a high degree of accuracy. Providing an accurate measurement of the entire Barrett’s area provides new opportunities for risk stratification and the assessment of therapy response.
Fixation beats – a novel marker for reaching the left bundle branch area during deep septal lead implantation
Authors: Jastrzębski Marek , Kiełbasa Grzegorz , Moskal Paweł , Bednarek Agnieszka , Kusiak Aleksander , Sondej Tomasz , Bednarski Adam , Rajzer Marek , Vijayaraman Pugazhendhi
Published: October 2020

Introduction

One of the challenges of left bundle branch (LBB) pacing is to place the pacing lead deep enough in the septum to obtain capture of the LBB, yet not too deep to avoid perforation. We hypothesized that the occurrence of the ectopic beats of qR/rsR’ morphology in V1 lead (fixation beats) during the lead fixation would predict that the final desired intraseptal lead depth was just reached, while the lack of fixation beats would indicate too shallow position, and need for more lead rotations.

Methods

Consecutive patients during LBB pacing device implantation were analyzed retrospectively and then prospectively with respect to the occurrence of the fixation beats during each episode of lead rotation. We compared the presence of fixation beats during the lead rotation event directly before the LBB capture area depth was reached versus during the events before intermediate/unsuccessful positions.

Results

A total of 339 patients and 1278 lead rotation events were analyzed. In the retrospective phase, the fixation beats were observed in 327/339 of final lead positions and in 9/939 of intermediate lead positions (p<0.001). The sensitivity, specificity, positive and negative predictive value of the LBB area fixation beats as a marker for reaching the LBB capture area was 96.4%, 99.0%, 97.3% and 99.0%, respectively. In the prospective, fixation-beats-guided, implantation phase the fixation beats were observed in all patients and only at the LBB capture depth.

Conclusions

Monitoring fixation beats during deep septal lead deployment can facilitate the procedure and possibly increase the safety of lead implantation.
Serological testing in addition to PCR screening for the re-opening of American colleges and universities: potential for cost-savings without compromising pandemic mitigation
Authors: Jo Youngji , Singh Ruby , Rao Gabriella , Galea Sandro , Nichols Brooke
Published: October 2020

Importance

The addition of a serological testing could reduce the overall testing costs of a PCR-based SARS-CoV-2 testing reopening plan for colleges/universities in the United States, without compromising the efficacy of the testing plan.

Objectives

To determine whether a college/university reopening SARS-CoV-2 testing plan that includes serological testing can be cost-saving compared to a PCR-only testing.

Design, Setting, and Participants

We assessed costs of serological testing in addition to PCR testing under various scenarios of university sizes (2000, 10,000, and 40,000) and epidemic conditions (initial antibody prevalence 2.5-15%; cumulative SARS-CoV-2 incidence during the school year 5-30%) of SARS-CoV-2 in the United States. We estimated total testing costs and relative percentage of cost-savings of different screening (i.e. targeted/ universal) and testing (i.e. in-sourcing/out-sourcing) scenarios between September 2020-May 2021.

Main Outcomes and Measures

Testing costs of serological testing and PCR testing, Relative percentage of cost saving by including serology testing in addition to PCR testing.

Results

Including baseline serology testing alongside routine regular PCR testing can reduce total test volumes and related costs throughout the school year. While the total testing cost is likely much lower if regular PCR testing is insourced compared to outsourced ($5 million vs $34 million for university size 10,000), including serologic testing could achieve the up to 20% cost-savings relative to PCR testing alone. The insourcing of serological testing when PCR testing is insourced can achieve greater cost-savings under high initial antibody prevalence (>5%) and cumulative incidence throughout the school year (>10%) at medium and large sized universities. If PCR testing is outsourced, however, the inclusion of serological testing becomes always preferred in most university sizes and epidemic conditions.

Conclusions and Relevance

While regular PCR testing alone is the preferred strategy for containing epidemics, including serology testing may help achieve cost-savings if outbreaks are anticipated, or if baseline seropositivity is high.

Key Points (96/100)

Question

Can the addition of a serological testing reduce the overall testing costs of a PCR-based SARS-CoV-2 testing reopening plan for universities in the United States?

Findings

This costing study suggested that inclusion of serological testing in addition to outsourced PCR testing as part of a university re-opening strategy could achieve cost savings of up to 20%. The amount of savings, or additional costs, is dependent on insourcing or outsourcing of testing, epidemic conditions and university size.

Meaning

The relative cost-savings depend strongly on whether PCR and/or serology are being insourced or outsourced, university sizes and cumulative incidence.
Reconstructing the diet, trophic level, and migration pattern of Mysticete whales based on baleen isotopic composition
Authors: Riekenberg Philip M. , Camalich Jaime , Svensson Elisabeth , IJsseldijk Lonneke L. , Brasseur Sophie M.J.M. , Witbaard Rob , Leopold Mardik F. , Rebolledo Elisa Bravo , Middelburg Jack J. , van der Meer Marcel T.J. , Damsté Jaap S. Sinninghe , Schouten Stefan
Published: October 2020

Baleen from mysticete whales is a well-preserved proteinaceous material that can be used to identify migrations and feeding habits for species whose migration pathways are unknown. Analysis of δ13C and δ15N from bulk baleen has been used to infer migration patterns for individuals. However, this approach has fallen short of identifying migrations between regions as it is difficult to determine variations in isotopic shifts without temporal sampling of prey items. Here we apply analysis of δ15N values of amino acids to five baleen plates belonging to three species, revealing novel insights on trophic position, metabolic state, and migration between regions. Humpback and minke whales had higher reconstructed trophic levels than fin whales (3.4-3.5 versus 2.7-2.9, respectively) as expected due to different feeding specialization. Isotopic niche areas between baleen minima and maxima were well separated, indicating regional resource use for individuals during migration that aligned with isotopic gradients in Atlantic Ocean particulate organic matter. δ15N values from phenylalanine confirmed regional separation between the niche areas for two fin whales as migrations occurred and elevated glycine and threonine δ15N values revealed physiological changes due to fasting. Simultaneous resolution of trophic level and physiological changes allow for identification of regional migrations in mysticetes.
Force-dependent remodeling of cytoplasmic ZO-1 condensates contributes to robust cell-cell adhesion
Authors: Kinoshita Noriyuki , Yamamoto Takamasa S. , Yasue Naoko , Takagi Chiyo , Fujimori Toshihiko , Ueno Naoto
Published: October 2020

Although the physiological importance of biomolecular condensates is widely recognized, how it is controlled in time and space during development is largely unknown. Here we show that a tight junction protein ZO-1 forms cytoplasmic condensates in the trophectoderm (TE) of the mouse embryo before E4.0. These disappear via dissolution, and ZO-1 accumulates at the cell junction as the blastocyst cavity grows and internal pressure on TE cells increases. In contrast, this dissolution was less evident in TE cells attached to the inner cell mass, as they receive weaker tensile forces. Furthermore, analyses using MDCK cells have demonstrated that the ZO-1 condensates are generated and maintained by liquid-liquid phase separation. Our study also highlights that the dynamics of these condensates depends on the physical environment via an interaction between ZO-1 and F-actin. We propose that the force-dependent regulation of ZO-1 condensation contributes to establishing robust cell-cell adhesion during early development.
Increased functional coupling of the mu opioid receptor in the anterior insula of depressed individuals
Authors: Lutz Pierre-Eric , Almeida Daniel , Filliol Dominique , Jollant Fabrice , Kieffer Brigitte L. , Turecki Gustavo
Published: October 2020

The mu opioid receptor (MOR) is a G protein-coupled receptor that plays an essential role in reward and hedonic processes, and that has been implicated in disorders such as depression and addiction. Over the last decade, several brain imaging studies in depressed patients have consistently found that dysregulation of MOR function occurs in particular in the anterior insular cortex, an important brain site for the perception of internal states and emotional regulation. To investigate molecular mechanisms that may underlie these effects, here we assessed genetic polymorphisms, expression, and functional G-protein coupling of MOR in a large post-mortem cohort (N=95) composed of depressed individuals who died by suicide, and healthy controls. Results indicated that depression, but not comorbid substance use disorder or acute opiate consumption, was associated with increased MOR activity. This effect was partly explained by a specific increase in expression of the inhibitory alpha G-protein subunit GNAI2. Consistent with previous neuroimaging studies, our findings support the notion that enhanced endogenous opioidergic tone in the anterior insula may buffer negative affective states in depressed individuals, a mechanism that could potentially contribute to the antidepressant efficacy of emerging opioid-based medications.
Protlego: A Python package for the analysis and design of chimeric proteins
Authors: Ferruz Noelia , Noske Jakob , Höcker Birte
Published: October 2020

Motivation

Gene duplication and recombination of protein fragments have led to the highly diverse protein space that we observe today. By mimicking this natural process, the design of protein chimeras via fragment recombination has proven experimentally successful and has opened a new era for the design of customizable proteins. Thein-silicobuilding of structural models for these chimeric proteins, however, remains a manual task that requires a considerable degree of expertise and is not amenable for high-throughput studies. Energetic and structural analysis of the designed proteins often require the use of several tools, each with their unique technical difficulties and available in different programming languages or web servers.

Results

We have implemented a Python package that enables automated, high-throughput design of chimeras and their structural analysis. First, it is possible to fetch evolutionarily conserved fragments from a built-in database (also available at fuzzle.uni-bayreuth.de). These relationships can then be represented via networks or further selected for chimera construction via recombination. Designed chimeras or natural proteins are then scored and minimised with the Charmm and Amber forcefields and their diverse structural features can be analysed at ease. Here, we showcase Protlego’s pipeline by exploring the relationships between the P-loop and Rossmann superfolds and building and characterising their offspring chimeras. We believe that Protlego provides a powerful new tool for the protein design community.

Availability and implementation

Protlego is freely available at (https://hoecker-lab.github.io/protlego/) with tutorials and documentation.
High-throughput generation and phenotypic characterization of zebrafish CRISPR mutants of DNA repair genes
Authors: Shin Unbeom , Nakhro Khriezhanuo , Oh Chang-Kyu , Carrington Blake , Song Hayne , Varshney Gaurav , Kim Youngjae , Song Hyemin , Jeon Sangeun , Robbins Gabrielle , Kim Sangin , Yoon Suhyeon , Choi Yongjun , Park Suhyung , Kim Yoo Jung , Burgess Shawn , Kang Sukhyun , Sood Raman , Lee Yoonsung , Myung Kyungjae
Published: October 2020

A systematic knowledge of the roles of DNA repair genes at the level of the organism has been limited due to the lack of appropriate experimental techniques. Here, we generated zebrafish loss-of-function mutants for 32 DNA repair and replication genes through multiplexed CRISPR/Cas9-mediated mutagenesis. High-throughput phenotypic characterization of our mutant collection revealed that three genes (atad5a,ddb1, pcna) are essential for proper embryonic development and hematopoiesis; seven genes (apex1,atrip,ino80,mre11a,shfm1,telo2,wrn) are required for growth and development during juvenile stage and six genes (blm,brca2,fanci, rad51,rad54l,rtel1) play critical roles in sex development. Furthermore, mutation in six genes (atad5a,brca2,polk,rad51,shfm1,xrcc1) displayed hypersensitivity to DNA damage agents. Further characterization ofatad5a−/−mutants demonstrate that Atad5a is required for normal brain development and hematopoiesis. Our zebrafish mutant collection provides a unique resource for understanding of the roles of DNA repair genes at the organismal level.
Biogeographical and ecological factors associated with kill rates of an apex predator
Authors: Cristescu Bogdan , Elbroch L. Mark , Dellinger Justin A. , Binder Wesley , Wilmers Christopher C. , Wittmer Heiko U.
Published: October 2020

Abstract

Kill rates and functional responses are fundamental to the study of predator ecology and the understanding of predatory-prey dynamics. As the most widely distributed apex predator in the western hemisphere pumas (Puma concolor) have been widely studied yet a biogeographical synthesis of their kill rates is currently lacking. We reviewed the literature and compiled data on sex- and age-specific kill rate estimates of pumas on ungulates, and conducted analyses aimed at understanding ecological factors explaining the observed variation across their range. Kill rate studies on pumas, while numerous, were primarily conducted in Temperate Conifer Forests (< 10% of puma range), revealing a dearth of knowledge across much of their range, especially from tropical and subtropical habitats. Across studies, kill rates in ungulates/week were highest for adult females with kitten(s) (1.24 ± 0.41 ungulates/week) but did not vary significantly between adult males (0.84 ± 0.18) and solitary adult females (0.99 ± 0.26). Kill rates in kg/day did not differ significantly among reproductive classes. Kill rates of adult pumas increased with ungulate density. Ungulate species richness had a weak negative association with adult male kill rates. Neither scavenger richness, the proportion of non-ungulate prey in the diet, nor regional human population density had a significant effect on ungulate kill rates. Our results had a strong temperate-ecosystem bias highlighting the need for further research across the diverse biomes pumas occupy in order to make species level inferences. Data from more populations would also allow for multivariate analyses providing deeper inference into the ecological and behavioural factors driving kill rates and functional responses of pumas, and apex predators in general.
Whole Genome Sequencing identifies novel structural variant in a large Indian family affected with X - linked agammaglobulinemia
Authors: Jain Abhinav , Govindaraj Geeta Madathil , Edavazhippurath Athulya , Faisal Nabeel , Bhoyar Rahul C , Gupta Vishu , Uppuluri Ramya , Manakkad Shiny Padinjare , Kashyap Atul , Kumar Anoop , Divakar Mohit Kumar , Imran Mohamed , Sawant Sneha , Dalvi Aparna , Chakyar Krishnan , Madkaikar Manisha , Raj Revathi , Sivasubbu Sridhar , Scaria Vinod
Published: October 2020

Background

X - linked agammaglobulinemia (XLA, OMIM #300755) is a primary immunodeficiency disorder caused by pathogenic variations in theBTKgene, characterized by failure of development and maturation of B lymphocytes. The estimated prevalence worldwide is 1 in 190,000 male births. Recently, genome sequencing has been widely used in difficult to diagnose and familial cases. We report a large Indian family suffering from XLA with five affected individuals.

Methods

We performed complete blood count, immunoglobulin assay, and lymphocyte subset analysis for all patients and analyzed Btk expression for one patient and his mother. Whole exome sequencing (WES) for four patients, and whole genome sequencing (WGS) for two patients have been performed. Carrier screening was done for 17 family members using Multiplex Ligation-dependent Probe Amplification (MLPA) and haplotype ancestry mapping using fineSTRUCTURE was performed.

Results

All patients had hypogammaglobulinemia and low CD19+ B cells. One patient who underwent Btk estimation had low expression and his mother showed a mosaic pattern. On structural variant analysis of WGS data, we found a novel large deletion of 5,296 bp at loci chrX:100,624,323-100,629,619 encompassing exons 3-5 of theBTKgene. Family screening revealed seven carriers for the deletion. Two patients had a successful HSCT. Haplotype mapping revealed mainly South Asian ancestry.

Conclusion

Whole genome sequencing led to identification of the accurate genetic mutation which could help in early diagnosis leading to improved outcomes, prevention of permanent organ damage and improved quality of life, as well as enabling prenatal diagnosis.
Development of a customised data management system for a COVID-19-adapted colorectal cancer pathway
Authors: Gunn Frances , Miller Janice , Dunlop Malcolm G , Din Farhat V N , Maeda Yasuko
Published: October 2020

Purpose

The COVID-19 pandemic posed an unprecedented challenge to healthcare systems around the world. To mitigate the risks of those referred with possible colorectal cancer during the pandemic we implemented a clinical pathway which required a customised data management system for robust operation. Here, we describe the principal concepts and evaluation of the performance of a spreadsheet-based data management system.

Methods

A system was developed using Microsoft Excel® 2007 aiming to retain the spreadsheets inherent intuitiveness of direct data entry. Data was itemised limiting entry errors. Visual Basic for Applications (VBA) was used to construct a user-friendly interface to enhance efficiency of data entry and segregate the data required for operational tasks. This was done with built-in loop-back data entry. Finally data derivation and analysis was performed to facilitate pathway monitoring.

Results

For a pathway which required rapid implementation and development of a customised data management system, the use of a spreadsheet was advantageous due to its user-friendly direct data entry capability. Its function was enhanced by UserForm and large data handling by data segregation using VBA macros. Data validation and conditional formatting minimised data entry errors. Computation by the COUNT function facilitated live data monitoring on a dashboard. During the three months the pathway ran for, the system processed 36 nodal data points for each of the included 837 patients. Data monitoring confirmed its accuracy.

Conclusion

Large volume data management using a spreadsheet system is possible with appropriate data definition and VBA programmed data segregation. Clinicians’ regular input and optimisation made the system adaptable for rapid implementation.
Comprehensive characterization of migration profiles of murine cerebral cortical neurons during development using FlashTag labeling
Authors: Yoshinaga Satoshi , Shin Minkyung , Kitazawa Ayako , Ishii Kazuhiro , Tanuma Masato , Kasai Atsushi , Hashimoto Hitoshi , Kubo Ken-ichiro , Nakajima Kazunori
Published: October 2020

In mammalian cerebral neocortex, different regions have different cytoarchitecture, neuronal birthdates and functions. In most regions, neuronal migratory profiles have been speculated similar to each other based on observations using thymidine analogues. Few reports investigated regional migratory differences from mitosis at the ventricular surface. Here, in mice, we applied FlashTag technology, in which dyes are injected intraventricularly, to describe migratory profiles. We revealed a mediolateral regional difference in migratory profiles of neurons that is dependent on the developmental stages, e.g., neurons labeled at E12.5-15.5 reached their destination earlier dorsomedially than dorsolaterally even where there were underlying ventricular surfaces, reflecting sojourning below the subplate. This difference was hardly recapitulated by thymidine analogues, which visualize neurogenic gradient, suggesting biological significance different from neurogenic gradient. These observations advance understanding of cortical development, portraying strength of FlashTag in studying migration, and are thus a resource for studies of normal and abnormal neurodevelopment.
A framework for comparing microbial networks reveals core associations
Authors: Röttjers Lisa , Vandeputte Doris , Raes Jeroen , Faust Karoline
Published: October 2020

Microbial network construction and analysis is an important tool in microbial ecology. As microbial interactions are challenging to infer experimentally, such networks are often constructed from statistically inferred associations and may not represent ecological interactions. Hence, microbial association networks contain a large number of errors and their derived properties do not necessarily reflect true community structure. Such errors can be identified with the use of appropriate null models. We have developed anuran, a toolbox for investigation of noisy networks with null models, for identification of non-random patterns in groups of association networks. This toolbox compares multiple networks to identify conserved subsets (core association networks, CANs) and other network properties that are shared across all networks. Such groups of networks can be generated from a collection of time series data or from cross-sectional sample sets. We use data from the Global Sponge Project to demonstrate that different orders of sponges have a larger CAN than expected at random.
Multiplexing Mutation Rate Assessment: Determining Pathogenicity of Msh2 Variants inS. cerevisiae
Authors: Ollodart Anja R. , Yeh Chiann-Ling C. , Miller Aaron W. , Shirts Brian H. , Gordon Adam S. , Dunham Maitreya J.
Published: October 2020

Despite the fundamental importance of mutation rate as a driving force in evolution and disease risk, common methods to assay mutation rate are time consuming and tedious. Established methods such as fluctuation tests and mutation accumulation experiments are low-throughput and often require significant optimization to ensure accuracy. We established a new method to determine the mutation rate of many strains simultaneously by tracking mutation events in a chemostat continuous culture device and applying deep sequencing to link mutations to alleles of a DNA-repair gene. We applied this method to assay the mutation rate of hundreds ofSaccharomyces cerevisiaestrains carrying mutations in the gene encoding Msh2, a DNA repair enzyme in the mismatch repair pathway (MMR). Loss-of-function (LOF) mutations inMSH2are associated with hereditary non-polyposis colorectal cancer (HNPCC), an inherited disorder that increases risk for many different cancers. However, the vast majority ofMSH2variants found in human populations have insufficient evidence to be classified as either pathogenic or benign. We first benchmarked our method against Luria-Delbrück fluctuation tests using a collection of publishedMSH2variants. Our pooled screen successfully identified previously-characterized non-functional alleles as high mutators. We then created an additional 185 human variants in the yeast ortholog, including both characterized and uncharacterized alleles curated from ClinVar and other clinical testing data. In a set of alleles of known pathogenicity, our assay recapitulated ClinVar’s classification; we then estimated pathogenicity for 157 variants classified as uncertain or conflicting reports of significance. This method is capable of studying the mutation rate of many microbial species and can be applied to problems ranging from the generation of high-fidelity polymerases to measuring the rate of antibiotic resistance emergence.
Trans-generational inheritance of centromere identity requires the CENP-A N-terminal tail in theC. elegansmaternal germ line
Authors: Prosée Reinier F. , Wenda Joanna M. , Gabus Caroline , Delaney Kamila , Schwager Francoise , Gotta Monica , Steiner Florian A.
Published: October 2020

Centromere protein A (CENP-A) is a histone H3 variant that defines centromeric chromatin and is essential for centromere function. In most eukaryotes CENP-A-containing chromatin is epigenetically maintained, and centromere identity is inherited from one cell cycle to the next. In the germ line of the holocentric nematodeCaenorhabditis elegans, this inheritance cycle is disrupted. CENP-A is removed at the mitosis-to-meiosis transition and is establishedde novoon chromatin during diplotene of meiosis I. Here we show that the N-terminal tail of CENP-A is required for thede novoestablishment of centromeres, but dispensable for centromere maintenance during embryogenesis. Worms homozygous for a CENP-A tail deletion maintain a functional centromere during development, but give rise to inviable offspring because they fail to re-establish centromeres in the maternal germ line. We identify the N-terminal tail of CENP-A as a critical domain for the interaction with the conserved kinetochore protein KNL-2, and argue that this interaction plays an important role in setting centromere identity in the germ line. We conclude that centromere establishment and maintenance are functionally distinct inC. elegans.
Genetic Variance for Fusarium Crown Rot Tolerance in Durum Wheat
Authors: Kadkol Gururaj , Meza Jess , Simpfendorfer Steven , Harden Steve , Cullis Brian
Published: October 2020

Tolerance to the cereal disease Fusarium crown rot (FCR) was investigated in a set of 34 durum wheat genotypes, with Suntop, (bread wheat) and EGA Bellaroi (durum) as tolerant and intolerant checks, in a series of replicated field trials over four years with inoculated (FCR-i) and non-inoculated (FCR-n) plots of the genotypes. The genotypes included conventional durum lines and lines derived from crossing durum with 2-49, a bread wheat line with the highest level of partial resistance to FCR. A split plot trial design was chosen to optimize the efficiency for the prediction of FCR tolerance for each genotype. A multi-environment trial (MET) analysis was undertaken which indicated that there was good repeatability of FCR tolerance across years. Based on an FCR tolerance index, Suntop was the most tolerant genotype and EGA Bellaroi was very intolerant, but many durum wheats had FCR tolerance indices which were comparable to Suntop. These included some conventional durum lines, V101030, TD1702, V11TD013*3X-63 and DBA Bindaroi, as well as genotypes from crosses with 2-49 (V114916 and V114942). The correlation between FCR tolerance and FCR-n yield predictions was moderately negative indicating it could be somewhat difficult to develop high yielding FCR-tolerant genotypes. However, FCR tolerance showed a positive correlation with FCR-i yield predictions in seasons of high disease expression indicating it could be possible to screen for FCR tolerance using only FCR-i treatments. These results are the first demonstration of genetic diversity in durum germplasm for FCR tolerance and they provide a basis for breeding for this trait.
Evolutionary history of Carnivora (Mammalia, Laurasiatheria) inferred from mitochondrial genomes
Authors: Hassanin Alexandre , Véron Géraldine , Ropiquet Anne , Jansen van Vuuren Bettine , Lécu Alexis , Goodman Steven M. , Haider Jibran , Nguyen Trung Thanh
Published: October 2020

The order Carnivora, which currently includes 296 species classified into 16 families, is distributed across all continents. The phylogeny and the timing of diversifications are still a matter of debate.

Here, complete mitochondrial genomes were analysed to reconstruct the phylogenetic relationships and to estimate divergence times among species of Carnivora. We assembled 51 new mitogenomes from 13 families, and aligned them with available mitogenomes by selecting only those showing more than 1% of nucleotide divergence and excluding those suspected to be of low-quality or from misidentified taxa. Our final alignment included 220 taxa representing 2,442 mitogenomes. Our analyses led to a robust resolution of suprafamilial and intrafamilial relationships. We identified 22 fossil calibration points to estimate a molecular timescale for carnivorans. According to our divergence time estimates, crown carnivorans appeared during or just after the Early Eocene Climatic Optimum; all major groups of Caniformia (Cynoidea/Arctoidea; Ursidae; Musteloidea/Pinnipedia) diverged from each other during the Eocene, while all major groups of Feliformia (Nandiniidae; Feloidea; Viverroidea) diversified more recently during the Oligocene, with a basal divergence ofNandiniaat the Eocene/Oligocene transition; intrafamilial divergences occurred during the Miocene, except for the Procyonidae, asPotosseparated from other genera during the Oligocene.
CRISPR/Cas9-engineered inducible gametocyte producer lines as a novel tool for basic and applied research onPlasmodium falciparummalaria transmission stages
Authors: Boltryk Sylwia D. , Passecker Armin , Alder Arne , van de Vegte-Bolmer Marga , Sauerwein Robert W. , Brancucci Nicolas M. B. , Beck Hans-Peter , Gilberger Tim-Wolf , Voss Till S.
Published: October 2020

The malaria parasitePlasmodium falciparumreplicates inside erythrocytes in the blood of infected humans. During each replication cycle, a small proportion of parasites commits to sexual development and differentiates into gametocytes, which are essential for parasite transmission to other human hosts via the mosquito vector. Detailed molecular investigation of gametocyte biology and transmission has been hampered by difficulties in generating large numbers of these highly specialized cells. Here, we engineered marker-freeP. falciparuminducible gametocyte producer (iGP) lines for the routine mass production of synchronous gametocytes. Through targeted overexpression of the sexual commitment factor GDV1, iGP lines consistently achieve sexual commitment rates of 75% and produce gametocytes that are infectious to mosquitoes. Subsequent tagging of a nucleoporin allowed us to visualize marked nuclear transformations during gametocytogenesis and demonstrates that further genetic engineering of iGP lines is an invaluable tool for the targeted exploration of gametocyte biology. We believe the iGP approach developed here opens up unprecedented opportunities that will expedite future basic and applied research onP. falciparumtransmission stages.