bluuebunny/crossref_metadata_2025 huggingface dataset:
High impact of COVID-19 outbreak in a nursing home in the Nouvelle-Aquitaine region, France, March to April 2020
Authors: Bernadou A. , Bouges S. , Catroux M. , Rigaux J. C. , Laland C. , Levêque N. , Noury U. , Larrieu S. , Acef S. , Habold D. , Cazenave-Roblot F. , Filleul L.
Published: February 2021

Background

Elderly people in nursing homes are particularly vulnerable to COVID-19 due to their age, the presence of comorbidities, and community living. On March 14, 2020, at the beginning of the first epidemic wave of COVID-19 in France, a cluster was reported in a nursing home in the Nouvelle-Aquitaine region. We monitored the outbreak as well as the infection prevention and control (IPC) measures implemented.

Methods

A confirmed case was defined as laboratory-confirmed COVID-19 in a resident or staff member present in the nursing home between March 7 and May 1, 2020; and a probable case as a person presenting an acute respiratory illness after contact with a confirmed case. Symptomatic inpatient residents and symptomatic staff members were systematically tested for SARS-CoV-2. In addition, two screening sessions were held on site.

Results

We identified 109 cases (98 confirmed, 11 probable). The attack rate was 66% among residents and 45% among staff. Half of all cases were identified during the screening sessions. One-quarter of cases had minor symptoms or were asymptomatic. The case fatality rate among residents was 29%. IPC measures were rapidly implemented such as the quarantine of residents, the reinforcement of staff personal protective equipment, and home quarantine of staff testing positive, which were supplemented in April by systematic controls at the entrance of the nursing home and the creation of additional staff break rooms.

Conclusions

This outbreak confirmed the considerable health impact of SARS-CoV-2 transmission in a nursing home. In addition to the implementation of IPC measures, the early detection of cases through the screening of residents and staff is essential to identify asymptomatic and pre-symptomatic cases and limit the spread of the virus.
A prospectus of plant growth promoting endophytic bacterium from orchid (Vanda cristata)
Authors: Shah Sujit , Chand Krishna , Rekadwad Bhagwan , Shouche Yogesh S. , Sharma Jyotsna , Pant Bijaya
Published: February 2021

Background

A plant growth-promoting endophytic bacterium PVL1 isolated from the leaf of Vanda cristata has the ability to colonize with roots of plants and protect the plant. PVL1 was isolated using laboratory synthetic media. 16S rRNA gene sequencing method has been employed for identification before and after root colonization ability.

Results

Original isolated and remunerated strain from colonized roots were identified as Bacillus spp. as per EzBiocloud database. The presence of bacteria in the root section of the plantlet was confirmed through Epifluorescence microscopy of colonized roots. The in-vitro plantlet colonized by PVL1 as well as DLMB attained higher growth than the control. PVL1 capable of producing plant beneficial phytohormone under in vitro cultivation. HPLC and GC-MS analysis suggest that colonized plants contain Indole Acetic Acid (IAA). The methanol extract of Bacillus spp., contains 0.015 μg in 1 μl concentration of IAA. PVL1 has the ability to produce antimicrobial compounds such as ethyl iso-allocholate, which exhibits immune restoring property. One-way ANOVA shows that results were statistically significant at P ≤ 0.05 level.

Conclusions

Hence, it has been concluded that Bacillus spp. PVL1 can promote plant growth through secretion of IAA during root colonization and ethyl iso-allocholate to protect plants from foreign infections. Thus, this study supports to support Koch’s postulates of bacteria establishment.
Predictors of performance on the pediatric board certification examination
Authors:   Japan Pediatric Society Steering Committee of Board Examination, Nomura Osamu , Onishi Hirotaka , Park Yoon Soo , Michihata Nobuaki , Kobayashi Tohru , Kaneko Kazunari , Yoshikawa Tetsushi , Ishiguro Akira
Published: February 2021

Background

Examining the predictors of summative assessment performance is important for improving educational programs and structuring appropriate learning environments for trainees. However, predictors of certification examination performance in pediatric postgraduate education have not been comprehensively investigated in Japan.

Methods

The Pediatric Board Examination database in Japan, which includes 1578 postgraduate trainees from 2015 to 2016, was analyzed. The examinations included multiple-choice questions (MCQs), case summary reports, and an interview, and the predictors for each of these components were investigated by multiple regression analysis.

Results

The number of examination attempts and the training duration were significant negative predictors of the scores for the MCQ, case summary, and interview. Employment at a community hospital or private university hospital were negative predictors of the MCQ and case summary score, respectively. Female sex and the number of academic presentations positively predicted the case summary and interview scores. The number of research publications was a positive predictor of the MCQ score, and employment at a community hospital was a positive predictor of the case summary score.

Conclusion

This study found that delayed and repeated examination taking were negative predictors, while the scholarly activity of trainees was a positive predictor, of pediatric board certification examination performance.
Correction to: Intellectual capital-based performance improvement: a study in healthcare sector
Authors: Alfiero Simona , Brescia Valerio , Bert Fabrizio
Published: February 2021

An amendment to this paper has been published and can be accessed via the original article.
Distribution and utilization of homestead windbreak Fukugi (Garcinia subelliptica Merr.) trees: an ethnobotanical approach
Authors: Chen Bixia , Akamine Hikaru
Published: February 2021

Background

Garcinia subelliptica(Fukugi in Japanese) is an evergreen tropical tree, first identified in Batanes, the Philippines, which has been planted as a homestead windbreak and in coastal forests extensively on the Ryukyu Archipelago, Japan. This article focuses on the traditional uses and cultural values of Fukugi trees and provides ethnobotanical information for the conservation scheme of this important tree species.

Methods

A combination of ecological and ethnobotanical approaches was applied in this study. Extensive field surveys were conducted to collect dimensions of relatively large trees, and in-depth interviews with the village leaders and knowledgeable persons were conducted to collect ethnobotanical data.

Results

Fukugi trees have been primarily planted as homestead or farmland windbreaks. Timber was harvested during difficult times, for example, after WWII, and used for recreational purposes for children or farmers. The fruits were also eaten on some remote islands. Old-growth Fukugi trees are widely found in sacred sites, within cities, and as symbolic trees. The older generations respect Fukugi trees; however, the cultural significance valued by older generation seems to be lacking in the younger generation. We argue that Fukugi is a cultural keystone species in Okinawa, which underpins Ryukyu culture and has transformed islands into a pleasant land, a unique place, and shared identity for the community.

Conclusions

Fukugi windbreaks provide significant ecosystem services, such as biodiversity in the forest, reducing soil erosion, and spiritual and cultural values. A combination of biophysical environment, as well as tradition and custom, has played an essential role in tree species selection for windbreaks. The positive impacts that anthropogenic activities have had on the sustainability of woody species, namely, the active utilization of tree species, may have enabled the species to sustain. Strategies for protecting old-growth Fukugi trees, in addition to restoration of damaged trees, are needed to improve the sustainable management of Fukugi trees in Okinawa.
scSorter: assigning cells to known cell types according to marker genes
Authors: Guo Hongyu , Li Jun
Published: February 2021

On single-cell RNA-sequencing data, we consider the problem of assigning cells to known cell types, assuming that the identities of cell-type-specific marker genes are given but their exact expression levels are unavailable, that is, without using a reference dataset. Based on an observation that the expected over-expression of marker genes is often absent in a nonnegligible proportion of cells, we develop a method called scSorter. scSorter allows marker genes to express at a low level and borrows information from the expression of non-marker genes. On both simulated and real data, scSorter shows much higher power compared to existing methods.
Relaxometric learning: a pattern recognition method for T2 relaxation curves based on machine learning supported by an analytical framework
Authors: Date Yasuhiro , Wei Feifei , Tsuboi Yuuri , Ito Kengo , Sakata Kenji , Kikuchi Jun
Published: February 2021

Nuclear magnetic resonance (NMR)-based relaxometry is widely used in various fields of research because of its advantages such as simple sample preparation, easy handling, and relatively low cost compared with metabolomics approaches. However, there have been no reports on the application of the T2 relaxation curves in metabolomics studies involving the evaluation of metabolic mixtures, such as geographical origin determination and feature extraction by pattern recognition and data mining. In this study, we describe a data mining method for relaxometric data (i.e., relaxometric learning). This method is based on a machine learning algorithm supported by the analytical framework optimized for the relaxation curve analyses. In the analytical framework, we incorporated a variable optimization approach and bootstrap resampling-based matrixing to enhance the classification performance and balance the sample size between groups, respectively. The relaxometric learning enabled the extraction of features related to the physical properties of fish muscle and the determination of the geographical origin of the fish by improving the classification performance. Our results suggest that relaxometric learning is a powerful and versatile alternative to conventional metabolomics approaches for evaluating fleshiness of chemical mixtures in food and for other biological and chemical research requiring a nondestructive, cost-effective, and time-saving method.
Development and implementation of a point of care ultrasound curriculum at a multi-site institution
Authors: Russell Frances M. , Herbert Audrey , Ferre Robinson M. , Zakeri Bita , Echeverria Valerie , Peterson Dina , Wallach Paul
Published: February 2021

In 2014, over 60% of medical schools were incorporating point of care ultrasound (POCUS) into their curriculum. Today, over 6 years later, many more schools are teaching POCUS or are in the planning stages of implementing a POCUS curriculum. In 2019, the AAMC reported that 53 schools or over one-third of US medical schools have multi-site campuses for undergraduate medical education. Implementation of a POCUS educational initiative at a multi-site campus presents unique challenges for teaching a uniform curriculum statewide. This article will discuss the POCUS curriculum and implementation process at a large multi-site institution.
Are hospitalized patients with inflammatory bowel disease at increased risk of invasive bacterial infections? Results from POLIBD 3-year cohort study
Authors: Gruszecka Jolanta , Filip Rafał
Published: February 2021

The aim of this study was to determine the dominant species of bacteria found in blood cultures collected from patients under treatment in the tertiary inflammatory bowel disease (IBD) center in Poland. The dominant pathogen isolated from blood in patients with IBD was Staphylococcus epidermidis MRCNS (MRCNS—methicillin-resistant coagulase-negative Staphylococcus), a strain resistant to all beta-lactam antibiotics (penicillins, penicillins with B-lactamase inhibitor, cephalosporins and carbapenems). The second most commonly isolated pathogen found in the blood samples was Escherichia coli. Blood cultures were found to be positive for these pathogens more frequently in male patients (90.0%). An increased risk of bacteremia in IBD patients was associated with prolonged hospitalization.
Menarche in primary ovarian insufficiency after a month of hormone replacement therapy: a case report
Authors: Cheng Biwen
Published: February 2021

Background

Gynecologic anomalies, including uterine agenesis and ovarian dysgenesis, are some of the several differential diagnoses in adolescent females with primary amenorrhea and delayed puberty. Primary ovarian insufficiency is reported in the clinical practice of reproductive endocrinology can be determined by conducting sex hormone tests to evaluate the hypothalamic-pituitary-ovarian axis. However, confirmation of Mullerian agenesis by image modalities can be extremely challenging. Once the diagnosis is established, breakthrough bleeding usually occurs 2 to 3 years after hormonal replacement therapy.

Case presentation

We report a case of a seventeen year old Taiwanese female, 46 XX karyotype, with ovarian dysgenesis and an initial tentative diagnosis of uterine agenesis who experienced a breakthrough bleeding after a month of hormonal replacement therapy.

Conclusions

The breakthrough bleeding after a month of estrogen therapy in primary ovarian insufficiency is uncommon, and the diagnosis of the absent uterus can have an extensive psychological impact on patients and their families.
Biliary type-II sphincter of Oddi dysfunction with a pancreatic duct dilation: a case report and review of the literature
Authors: Al-Khoury Naseem , Mohamad Okbah , Mazloum Abd Al-Jawad , Madi Maher
Published: February 2021

Background

The double-duct sign is defined as dilation of both the common bile duct and pancreatic duct, which usually indicates pancreatic malignancy. However, benign causes have also been reported to cause a double-duct sign.

Case presentation

We present the case of a 59-year-old Caucasian female patient admitted to the Gastroenterology Department with intermittent right epigastric abdominal pain and an intact gallbladder. A double-duct sign was seen on endoscopic ultrasound. The suspicion of pancreatic malignancy was excluded through follow-up investigations. Biliary type II sphincter of Oddi dysfunction was diagnosed with an association of the double-duct sign. Sphincterotomy was performed to reduce pain, and there was no recurrence of symptoms during follow-up.

Conclusions

This is the third reported case in the literature of the double-duct sign associated with sphincter of Oddi dysfunction. This case emphasizes that the double-duct sign is not always caused by a local malignancy. The literature review of the reported cases has been summarized to help in the diagnosis of future similar cases.
Streptococcus suis serotype 2 enolase interaction with host brain microvascular endothelial cells and RPSA-induced apoptosis lead to loss of BBB integrity
Authors: Liu Hongtao , Lei Siyu , Jia Li , Xia Xiaojing , Sun Yingying , Jiang Hexiang , Zhu Rining , Li Shuguang , Qu Guanggang , Gu Jingmin , Sun Changjiang , Feng Xin , Han Wenyu , Langford Paul R. , Lei Liancheng
Published: February 2021

Host proteins interacting with pathogens are receiving more attention as potential therapeutic targets in molecular medicine. Streptococcus suis serotype 2 (SS2) is an important cause of meningitis in both humans and pigs worldwide. SS2 Enolase (Eno) has previously been identified as a virulence factor with a role in altering blood brain barrier (BBB) integrity, but the host cell membrane receptor of Eno and The mechanism(s) involved are unclear. This study identified that SS2 Eno binds to 40S ribosomal protein SA (RPSA) on the surface of porcine brain microvascular endothelial cells leading to activation of intracellular p38/ERK-eIF4E signalling, which promotes intracellular expression of HSPD1 (heat-shock protein family D member 1), and initiation of host-cell apoptosis, and increased BBB permeability facilitating bacterial invasion. This study reveals novel functions for the host-interactional molecules RPSA and HSPD1 in BBB integrity, and provides insight for new therapeutic strategies in meningitis.
Efficient genome editing with CRISPR/Cas9 in Pleurotus ostreatus
Authors: Boontawon Tatpong , Nakazawa Takehito , Inoue Chikako , Osakabe Keishi , Kawauchi Moriyuki , Sakamoto Masahiro , Honda Yoichi
Published: February 2021

Pleurotus ostreatus is one of the most commercially produced edible mushrooms worldwide. Improved cultivated strains with more useful traits have been obtained using classical breeding, which is laborious and time-consuming. Here, we attempted efficient gene mutagenesis using plasmid-based CRISPR/Cas9 as the first step for non-genetically modified (non-GM) P. ostreatus generation. Plasmids harboring expression cassettes of Cas9 and different single guide RNAs targeting fcy1 and pyrG were individually transferred into fungal protoplasts of the PC9 strain, which generated some strains exhibiting resistance to 5-fluorocytosine and 5-fluoroorotic acid, respectively. Genomic PCR followed by sequencing revealed small insertions/deletions or insertion of a fragment from the plasmid at the target site in some of the drug-resistant strains. The results demonstrated efficient CRISPR/Cas9-assisted genome editing in P. ostreatus, which could contribute to the molecular breeding of non-GM cultivated strains in the future. Furthermore, a mutation in fcy1 via homology-directed repair using this CRISPR/Cas9 system was also efficiently introduced, which could be applied not only for precise gene disruption, but also for insertions leading to heterologous gene expression in this fungus.
One-step electropolymerization of methylene blue films on highly flexible carbon fiber electrode as supercapacitors
Authors: Samyn Leandro Marques , Suresh Babu Rajendran , Devendiran Mani , de Barros Ana Lucia Ferreira
Published: February 2021

Energy crisis and environmental pollution have been one of the major global issues. In this regard, the search for new energy storage materials is cheap, flexible and high-performance supercapacitors electrode which has become intensive. Also, reducing the amount of organic dyes polluting in water is a great significance. Herein, one-step electropolymerization of methylene blue on carbon fiber and the resulting films were applied to the supercapacitor. The high performance is associated to the outstanding conductivity, electrochemical stability and superior mechanical flexibility of carbon fiber. A new flexible electrode for supercapacitors was successfully fabricated by demonstrating with a good electrochemical performance and a promising alternative to reduce the water pollution.
Socket seal surgery techniques in the esthetic zone: a systematic review with meta-analysis and trial sequential analysis of randomized clinical trials
Authors: López-Pacheco Andrea , Soto-Peñaloza David , Gómez Mayra , Peñarrocha-Oltra David , Alarcón Marco Antonio
Published: February 2021

Background

The socket seal surgery (SSS) technique is a common alternative for the management of the post-extraction sockets that requires a primary closure of the wound to promote proper regeneration and ridge preservation.

Objective

To learn about the effect of different SSS techniques on alveolar ridge preservation

Material and methods

Two independent and calibrated reviewers conducted an electronic search in PubMed, Cochrane, and Web of Science for randomized clinical trials (RCT) published up to June 2020. The evaluation of the risk of bias in the included studies was carried out following the Cochrane manual for interventions of systematic reviews, version 5.1.0. A meta-analysis of ridge width changes at − 1, − 3, and − 5 mm cutoff points from bone crest was conducted using a random-effects model. The risk of types I and II errors against accrued data was appraised obtaining the required information size using a trial sequential analysis package (TSA).

Results

A total of 135 sockets located in the esthetic zone were evaluated with a minimum of a 3-month follow-up after tooth extraction in 6 RCTs. The evaluated SSS techniques were free gingival graft (FGG), collagen matrix (CM), collagen sponge (CS), acellular dermal matrix (ADM), and polytetrafluoroethylene membrane (PTFEm). The FGG in sockets without bone filling showed significant results in preserving both buccal and lingual bone height (− 1.42 mm in the experimental group versus − 0.01 in the control group). The comparison of CM and FGG with bone filling did not show clinical differences in terms of dimensional bone changes. No clinical differences were found in either width or gingival thickness when comparing CM and CS. The meta-analyses of RW changes comparing CM versus FGG showed no significant differences, but a trend for lessening horizontal reduction at − 1, − 3, and − 5 mm in favor of FGG. The TSA showed that accrued data did not reach the required information size, and more evidence is required for clinical significance inferences.

Conclusions

There are several predictable SSS techniques to improve clinical results in ridge preservation. More clinical studies in the form of clinical trials are required to demonstrate the superiority of one technique over another.
Hydrocephalus with lateral ventricular lesions: case series and review of literature
Authors: Alselisly Ahmed Mohamed Ahmed , Ashry Ahmed Hamdy , Mahmoud Ayman Tarek
Published: February 2021

Background

Lateral ventricular lesions associated with hydrocephalus are considered a challenge to neurosurgeons. Hydrocephalus after surgery of such lesions and its pathogenesis and how to deal with it is a big question facing neurosurgeons.

Objectives

In this study, we tried to discuss the pathogenesis and different forms of presentation of hydrocephalus in lateral ventricular lesions and how to deal with it.

Methods

Eleven patients with lateral ventricular lesions associated with hydrocephalus either preoperative or postoperative presenting to our hospital were managed by excision of the lesion. A prospective study was done for these cases including their clinical data, radiological data, the presence, or absence of hydrocephalus either preoperative or postoperative and how we managed it.

Results

This study included 11 cases. The mean patient age at surgery was 25 years old. Nine cases were presented with radiological signs of hydrocephalus preoperatively. Two cases developed new onset hydrocephalus after lesion excision. Six cases ended with permanent CSF diversion.

Conclusion

Management of cases with lateral ventricular lesions does not stand on only excision of the lesion. Hydrocephalus should be kept into consideration perioperatively. We should try to avoid events that could lead to ventriculitis. Prolonged follow-up of the patients postoperative is very important as hydrocephalus may develop later after surgery.
Educational Assortative Mating in Sub-Saharan Africa: Compositional Changes and Implications for Household Wealth Inequality
Authors: Pesando Luca Maria
Published: February 2021

Sub-Saharan Africa (SSA) is undergoing rapid transformations in the realm of union formation in tandem with significant educational expansion and rising labor force participation rates. Concurrently, the region remains the least developed and most unequal along multiple dimensions of human and social development. In spite of this unique scenario, never has the social stratification literature examined patterns and implications of educational assortative mating for inequality in SSA. Using 126 Demographic and Health Surveys from 39 SSA countries between 1986 and 2016, this study is the first to document changing patterns of educational assortative mating by marriage cohort, subregion, and household location of residence and relate them to prevailing sociological theories on mating and development. Results show that net of shifts in educational distributions, mating has increased over marriage cohorts in all subregions except for Southern Africa, with increases driven mostly by rural areas. Trends in rural areas align with the status attainment hypothesis, whereas trends in urban areas are consistent with the inverted U-curve framework and the increasing applicability of the general openness hypothesis. The inequality analysis conducted through a combination of variance decomposition and counterfactual approaches reveals that mating accounts for a nonnegligible share (3% to 12%) of the cohort-specific inequality in household wealth, yet changes in mating over time hardly move time trends in wealth inequality, which is in line with findings from high-income societies.
BoneJ2 - refactoring established research software
Authors: Domander Richard , Felder Alessandro A , Doube Michael
Published: February 2021

Research software is often developed with expedience as a core development objective because experimental results, but not the software, are specified and resourced as a project output. While such code can help find answers to specific research questions, it may lack longevity and flexibility to make it reusable. We reimplemented BoneJ, our software for skeletal biology image analysis, to address design limitations that put it at risk of becoming unusable. We improved the quality of BoneJ code by following contemporary best programming practices. These include separation of concerns, dependency management, thorough testing, continuous integration and deployment, source code management, code reviews, issue and task ticketing, and user and developer documentation. The resulting BoneJ2 represents a generational shift in development technology and integrates with the ImageJ2 plugin ecosystem.
La cuestión probatoria de la falta de disposición cuando el Estado ha tenido el propósito de sustraer o blindar al individuo de su responsabilidad por los crímenes internacionales
Authors: Vera Sánchez Juan Sebastián
Published: February 2021

Este trabajo intenta ofrecer una mirada “probatoria” de la falta de disposición cuando el Estado ha tenido el propósito de sustraer o blindar al individuo de su responsabilidad por los crímenes internacionales, como hipótesis excepcional que permite la admisión del caso para ser conocido por la CPI en contrapartida al principio de complementariedad. Se intentará aplicar la visión epistemológica fundaherentista de Susan Haack. Ello, con la idea de generar una visión de análisis más objetivo a la hora de aplicar indicadores que tanto la doctrina como jurisprudencia han elaborado para saber cuándo un procedimiento judicial no es genuino.
Social Network Positions, Peer Effects, and Evaluation Updating: An Experimental Test in the Entrepreneurial Context
Authors: Greenberg Jason
Published: February 2021

In many facets of life, individuals make evaluations that they may update after consulting with others in their networks. But not all individuals have the same positional opportunities for social interaction in a given network or the ability and desire to make use of those opportunities that are available to them. The configuration of a person’s network can also alter how information is spread or interpreted. To complicate matters further, scant research has considered how positions in social networks and the valence of network content interact because of the difficulty of (a) separating the “player” from the position in networks and (b) measuring all germane content in a particular network. This research develops a novel experimental platform that addresses these issues. Participants viewed and evaluated an entrepreneurial video pitch and were then randomly assigned to different networks, and positions within networks, and thus various opportunities for peer influence that were orthogonal to their network history, inclinations, attributes, or capabilities. Furthermore, all the content of social interaction, including its valence, was recorded to test underlying assumptions. Results reveal that those assigned to a position with brokerage opportunities in a network updated their evaluations of the entrepreneurial video considerably more negatively.
Selective internal radiation therapy in the management of primary and metastatic disease in the liver
Authors: Preston Emma , Shaida Nadeem
Published: February 2021

Selective internal radiation therapy is a type of brachytherapy used to provide targeted radiotherapy, most commonly to treat primary or metastatic disease within the liver. This review outlines current clinical practice, dosimetric considerations, the pre-treatment workup and safety considerations before treatment. It also examines the clinical evidence for its use in patients with both primary and metastatic disease within the liver.
Trinocular vision and spatial prior based method for ground clearance measurement of transmission lines
Authors: Zhou Yaqin , Li Qingwu , Wu Yi , Ma Yunpeng , Wang Chunkuan
Published: February 2021

It is an essential task to inspect ground clearance of transmission lines in time. However, the weak texture of transmission lines and high complexity of the background make it difficult to balance efficiency and accuracy. To solve the problem, a trinocular vision and spatial prior based method is proposed, which is specifically designed for ground clearance measurement of transmission lines with unmanned aerial vehicles (UAVs). In this novel method, a perpendicular double-baseline trinocular vision module is applied to improve the accuracy of transmission line reconstruction. Then the spatial prior information of geometric models under different shooting attitudes is analyzed in detail, and it is adopted to determine the ground crossing points and compute ground clearance efficiently. Also, an interactive software is developed and tested in the simulation environment of UAV inspection. Experimental results verify the feasibility of the measurement method. Finally, we discuss in detail how to apply the method effectively in practice and give a set of recommended camera parameters.
Quantitative profiling of protease specificity
Authors: Ratnikov Boris I. , Cieplak Piotr , Remacle Albert G. , Nguyen Elise , Smith Jeffrey W.
Published: February 2021

Proteases are an important class of enzymes, whose activity is central to many physiologic and pathologic processes. Detailed knowledge of protease specificity is key to understanding their function. Although many methods have been developed to profile specificities of proteases, few have the diversity and quantitative grasp necessary to fully define specificity of a protease, both in terms of substrate numbers and their catalytic efficiencies. We have developed a concept of “selectome”; the set of substrate amino acid sequences that uniquely represent the specificity of a protease. We applied it to two closely related members of the Matrixin family–MMP-2 and MMP-9 by using substrate phage display coupled with Next Generation Sequencing and information theory-based data analysis. We have also derived a quantitative measure of substrate specificity, which accounts for both the number of substrates and their relative catalytic efficiencies. Using these advances greatly facilitates elucidation of substrate selectivity between closely related members of a protease family. The study also provides insight into the degree to which the catalytic cleft defines substrate recognition, thus providing basis for overcoming two of the major challenges in the field of proteolysis: 1) development of highly selective activity probes for studying proteases with overlapping specificities, and 2) distinguishing targeted proteolysis from bystander proteolytic events.
A mathematical model of the role of aggregation in sonic hedgehog signalling
Authors: Derrick Daniel J. A. , Wolton Kathryn , Currie Richard A. , Tindall Marcus John
Published: February 2021

Effective regulation of the sonic hedgehog (Shh) signalling pathway is essential for normal development in a wide variety of species. Correct Shh signalling requires the formation of Shh aggregates on the surface of producing cells. Shh aggregates subsequently diffuse away and are recognised in receiving cells located elsewhere in the developing embryo. Various mechanisms have been postulated regarding how these aggregates form and what their precise role is in the overall signalling process. To understand the role of these mechanisms in the overall signalling process, we formulate and analyse a mathematical model of Shh aggregation using nonlinear ordinary differential equations. We consider Shh aggregate formation to comprise of multimerisation, association with heparan sulfate proteoglycans (HSPG) and binding with lipoproteins. We show that the size distribution of the Shh aggregates formed on the producing cell surface resembles an exponential distribution, a result in agreement with experimental data. A detailed sensitivity analysis of our model reveals that this exponential distribution is robust to parameter changes, and subsequently, also to variations in the processes by which Shh is recruited by HSPGs and lipoproteins. The work demonstrates the time taken for different sized Shh aggregates to form and the important role this likely plays in Shh diffusion.
Rapid 3D phenotypic analysis of neurons and organoids using data-driven cell segmentation-free machine learning
Authors: Mergenthaler Philipp , Hariharan Santosh , Pemberton James M. , Lourenco Corey , Penn Linda Z. , Andrews David W.
Published: February 2021

Phenotypic profiling of large three-dimensional microscopy data sets has not been widely adopted due to the challenges posed by cell segmentation and feature selection. The computational demands of automated processing further limit analysis of hard-to-segment images such as of neurons and organoids. Here we describe a comprehensive shallow-learning framework for automated quantitative phenotyping of three-dimensional (3D) image data using unsupervised data-driven voxel-based feature learning, which enables computationally facile classification, clustering and advanced data visualization. We demonstrate the analysis potential on complex 3D images by investigating the phenotypic alterations of: neurons in response to apoptosis-inducing treatments and morphogenesis for oncogene-expressing human mammary gland acinar organoids. Our novel implementation of image analysis algorithms called Phindr3D allowed rapid implementation of data-driven voxel-based feature learning into 3D high content analysis (HCA) operations and constitutes a major practical advance as the computed assignments represent the biology while preserving the heterogeneity of the underlying data. Phindr3D is provided as Matlab code and as a stand-alone program (https://github.com/DWALab/Phindr3D).
Calcium-vesicles perform active diffusion in the sea urchin embryo during larval biomineralization
Authors: Winter Mark R. , Morgulis Miri , Gildor Tsvia , Cohen Andrew R. , Ben-Tabou de-Leon Smadar
Published: February 2021

Biomineralization is the process by which organisms use minerals to harden their tissues and provide them with physical support. Biomineralizing cells concentrate the mineral in vesicles that they secret into a dedicated compartment where crystallization occurs. The dynamics of vesicle motion and the molecular mechanisms that control it, are not well understood. Sea urchin larval skeletogenesis provides an excellent platform for investigating the kinetics of mineral-bearing vesicles. Here we used lattice light-sheet microscopy to study the three-dimensional (3D) dynamics of calcium-bearing vesicles in the cells of normal sea urchin embryos and of embryos where skeletogenesis is blocked through the inhibition of Vascular Endothelial Growth Factor Receptor (VEGFR). We developed computational tools for displaying 3D-volumetric movies and for automatically quantifying vesicle dynamics. Our findings imply that calcium vesicles perform an active diffusion motion in both, calcifying (skeletogenic) and non-calcifying (ectodermal) cells of the embryo. The diffusion coefficient and vesicle speed are larger in the mesenchymal skeletogenic cells compared to the epithelial ectodermal cells. These differences are possibly due to the distinct mechanical properties of the two tissues, demonstrated by the enhanced f-actin accumulation and myosinII activity in the ectodermal cells compared to the skeletogenic cells. Vesicle motion is not directed toward the biomineralization compartment, but the vesicles slow down when they approach it, and probably bind for mineral deposition. VEGFR inhibition leads to an increase of vesicle volume but hardly changes vesicle kinetics and doesn’t affect f-actin accumulation and myosinII activity. Thus, calcium vesicles perform an active diffusion motion in the cells of the sea urchin embryo, with diffusion length and speed that inversely correlate with the strength of the actomyosin network. Overall, our studies provide an unprecedented view of calcium vesicle 3D-dynamics and point toward cytoskeleton remodeling as an important effector of the motion of mineral-bearing vesicles.
Ancient and recent introgression shape the evolutionary history of pollinator adaptation and speciation in a model monkeyflower radiation (Mimulus section Erythranthe)
Authors: Nelson Thomas C. , Stathos Angela M. , Vanderpool Daniel D. , Finseth Findley R. , Yuan Yao-wu , Fishman Lila
Published: February 2021

Inferences about past processes of adaptation and speciation require a gene-scale and genome-wide understanding of the evolutionary history of diverging taxa. In this study, we use genome-wide capture of nuclear gene sequences, plus skimming of organellar sequences, to investigate the phylogenomics of monkeyflowers inMimulussectionErythranthe(27 accessions from seven species). Taxa withinErythranthe, particularly the parapatric and putatively sister speciesM.lewisii(bee-pollinated) andM.cardinalis(hummingbird-pollinated), have been a model system for investigating the ecological genetics of speciation and adaptation for over five decades. Across >8000 nuclear loci, multiple methods resolve a predominant species tree in whichM.cardinalisgroups with other hummingbird-pollinated taxa (37% of gene trees), rather than being sister toM.lewisii(32% of gene trees). We independently corroborate a single evolution of hummingbird pollination syndrome inErythrantheby demonstrating functional redundancy in genetic complementation tests of floral traits in hybrids; together, these analyses overturn a textbook case of pollination-syndrome convergence. Strong asymmetries in allele sharing (Patterson’s D-statistic and related tests) indicate that gene tree discordance reflects ancient and recent introgression rather than incomplete lineage sorting. Consistent with abundant introgression blurring the history of divergence, low-recombination and adaptation-associated regions support the new species tree, while high-recombination regions generate phylogenetic evidence for sister status forM.lewisiiandM.cardinalis. Population-level sampling of core taxa also revealed two instances of chloroplast capture, with SierranM.lewisiiand Southern CalifornianM.parishiieach carrying organelle genomes nested within respective sympatricM.cardinalisclades. A recent organellar transfer fromM.cardinalis, an outcrosser where selfish cytonuclear dynamics are more likely, may account for the unexpected cytoplasmic male sterility effects of selferM.parishiiorganelles in hybrids withM.lewisii. Overall, our phylogenomic results reveal extensive reticulation throughout the evolutionary history of a classic monkeyflower radiation, suggesting that natural selection (re-)assembles and maintains species-diagnostic traits and barriers in the face of gene flow. Our findings further underline the challenges, even in reproductively isolated species, in distinguishing re-use of adaptive alleles from true convergence and emphasize the value of a phylogenomic framework for reconstructing the evolutionary genetics of adaptation and speciation.
A multicenter non-randomized, uncontrolled single arm trial for evaluation of the efficacy and the safety of the treatment with favipiravir for patients with severe fever with thrombocytopenia syndrome
Authors: Suemori Koichiro , Saijo Masayuki , Yamanaka Atsushi , Himeji Daisuke , Kawamura Masafumi , Haku Takashi , Hidaka Michihiro , Kamikokuryo Chinatsu , Kakihana Yasuyuki , Azuma Taichi , Takenaka Katsuto , Takahashi Toru , Furumoto Akitsugu , Ishimaru Toshiyuki , Ishida Masayuki , Kaneko Masahiko , Kadowaki Norimitsu , Ikeda Kenichi , Sakabe Shigetoshi , Taniguchi Tomohiro , Ohge Hiroki , Kurosu Takeshi , Yoshikawa Tomoki , Shimojima Masayuki , Yasukawa Masaki
Published: February 2021

Severe fever with thrombocytopenia syndrome (SFTS) is a bunyavirus infection with high mortality. Favipiravir has shown effectiveness in preventing and treating SFTS virus (SFTSV) infection in animal models. A multicenter non-randomized, uncontrolled single arm trial was conducted to collect data on the safety and the effectiveness of favipiravir in treatment of SFTS patients. All participants received favipiravir orally (first-day loading dose of 1800 mg twice a day followed by 800 mg twice a day for 7–14 days in total). SFTSV RT-PCR and biochemistry tests were performed at designated time points. Outcomes were 28-day mortality, clinical improvement, viral load evolution, and adverse events (AEs). Twenty-six patients were enrolled, of whom 23 were analyzed. Four of these 23 patients died of multi-organ failure within one week (28-day mortality rate: 17.3%). Oral favipiravir was well tolerated in the surviving patients. AEs (abnormal hepatic function and insomnia) occurred in about 20% of the patients. Clinical symptoms improved in all patients who survived from a median of day 2 to day10. SFTSV RNA levels in the patients who died were significantly higher than those in the survivors (p = 0.0029). No viral genomes were detectable in the surviving patients a median of 8 days after favipiravir administration. The 28-day mortality rate in this study was lower than those of the previous studies in Japan. The high frequency of hepatic dysfunction as an AE was observed. However, it was unclear whether this was merely a side effect of favipiravir, because liver disorders are commonly seen in SFTS patients. The results of this trial support the effectiveness of favipiravir for patients with SFTS.
NLRP6-associated host microbiota composition impacts in the intestinal barrier to systemic dissemination of Brucella abortus
Authors: Rungue Marcella , Melo Victor , Martins David , Campos Priscila C. , Leles Gabriela , Galvão Izabela , Mendes Viviani , Aganetti Mariana , Pedersen Ágatha , Assis Natan R. G. , Santos Raiany , Cassali Geovanni D. , Godard Ana Lúcia B. , Martins Flaviano S. , Oliveira Sergio C. , Vieira Angélica T.
Published: February 2021

Brucella abortus is a Gram-negative bacterium responsible for a worldwide zoonotic infection—Brucellosis, which has been associated with high morbidity rate in humans and severe economic losses in infected livestock. The natural route of infection is through oral and nasal mucosa but the invasion process through host gut mucosa is yet to be understood. Studies have examined the role of NLRP6 (NOD-like receptor family pyrin domain-containing-6 protein) in gut homeostasis and defense against pathogens. Here, we investigated the impact of gut microbiota and NLRP6 in a murine model of Ba oral infection. Nlrp6-/- and wild-type (WT) mice were infected by oral gavage with Ba and tissues samples were collected at different time points. Our results suggest that Ba oral infection leads to significant alterations in gut microbiota. Moreover, Nlrp6-/- mice were more resistant to infection, with decreased CFU in the liver and reduction in gut permeability when compared to the control group. Fecal microbiota transplantation from WT and Nlrp6-/- into germ-free mice reflected the gut permeability phenotype from the donors. Additionally, depletion of gut microbiota by broad-spectrum-antibiotic treatment prevented Ba replication in WT while favoring bacterial growth in Nlrp6-/-. Finally, we observed higher eosinophils in the gut and leukocytes in the blood of infected Nlrp6-/- compared to WT-infected mice, which might be associated to the Nlrp6-/- resistance phenotype. Altogether, these results indicated that gut microbiota composition is the major factor involved in the initial stages of pathogen host replication and partially also by the resistance phenotype observed in Nlrp6 -/- mice regulating host inflammation against Ba infection.
The fungus Leptosphaerulina persists in Anopheles gambiae and induces melanization
Authors: Nattoh Godfrey , Bargul Joel L. , Magoma Gabriel , Mbaisi Lilian , Butungi Hellen , Mararo Enock , Teal Evan , Herren Jeremy Keith
Published: February 2021

Anopheles mosquitoes are colonized by diverse microorganisms that may impact on host biology and vectorial capacity. Eukaryotic symbionts such as fungi have been isolated from Anopheles, but whether they are stably associated with mosquitoes and transmitted transstadially across mosquito life stages or to subsequent generations remains largely unexplored. Here, we show that a Leptosphaerulina sp. fungus isolated from the midgut of An. gambiae can be stably associated with An. gambiae host and that it imposes low fitness cost when re-introduced through co-feeding. This fungus is transstadially transmitted across An. gambiae developmental stages and to their progeny. It is present in field-caught larvae and adult mosquitoes at moderate levels across geographical regions. We observed that Leptosphaerulina sp. induces a distinctive melanotic phenotype across the developmental stages of mosquito. As a eukaryotic symbiont that is stably associated with An. gambiae Leptosphaerulina sp. can be explored for paratransgenesis.
Biomarker-focused multi-drug combination therapy and repurposing trial in mdx mice
Authors: Ziemba Michael , Barkhouse Molly , Uaesoontrachoon Kitipong , Giri Mamta , Hathout Yetrib , Dang Utkarsh J. , Gordish-Dressman Heather , Nagaraju Kanneboyina , Hoffman Eric P.
Published: February 2021

Duchenne muscular dystrophy is initiated by dystrophin deficiency, but downstream pathophysiological pathways such as membrane instability, NFĸB activation, mitochondrial dysfunction, and induction of TGFβ fibrosis pathways are thought to drive the disability. Dystrophin replacement strategies are hopeful for addressing upstream dystrophin deficiency; however, all methods to date use semi-functional dystrophin proteins that are likely to trigger downstream pathways. Thus, combination therapies that can target multiple downstream pathways are important in treating DMD, even for dystrophin-replacement strategies. We sought to define blood pharmacodynamic biomarkers of drug response in the mdx mouse model of Duchenne muscular dystrophy using a series of repurposed drugs. Four-week-old mdx mice were treated for four weeks with four different drugs singly and in combination: vehicle, prednisolone, vamorolone, rituximab, β-aminoisobutyric acid (BAIBA) (11 treatment groups; n = 6/group). Blood was collected via cardiac puncture at study termination, and proteomic profiling was carried out using SOMAscan aptamer panels (1,310 proteins assayed). Prednisolone was tested alone and in combination with other drugs. It was found to have a good concordance of prednisolone-responsive biomarkers (56 increased by prednisolone, 39 decreased) focused on NFκB and TGFβ cascades. Vamorolone shared 45 (80%) of increased biomarkers and 13 (33%) of decreased biomarkers with prednisolone. Comparison of published human corticosteroid-responsive biomarkers to our mdx data showed 14% (3/22) concordance between mouse and human. Rituximab showed fewer drug-associated biomarkers, with the most significant being human IgG. On the other hand, BAIBA treatment (high and low dose) showed a drug-associated increase in 40 serum proteins and decreased 5 serum proteins. Our results suggest that a biomarker approach could be employed for assessing drug combinations in both mouse and human studies.