bluuebunny/crossref_metadata_2025 huggingface dataset:
A mammalian methylation array for profiling methylation levels at conserved sequences
Authors: Arneson Adriana , Haghani Amin , Thompson Michael J. , Pellegrini Matteo , Kwon Soo Bin , Vu Ha , Yao Mingjia , Li Caesar Z. , Lu Ake T. , Barnes Bret , Hansen Kasper D. , Zhou Wanding , Breeze Charles E. , Ernst Jason , Horvath Steve
Published: January 2021

Infinium methylation arrays are not available for the vast majority of non-human mammals. Moreover, even if species-specific arrays were available, probe differences between them would confound cross-species comparisons. To address these challenges, we developed the mammalian methylation array, a single custom array that measures 36k CpGs that are well conserved across mammalian species. We designed a set of probes on the array that can tolerate specific cross-species mutations. We annotate the array in over 200 species and report CpG island status and chromatin states in select species. Calibration experiments demonstrate the high fidelity in humans, rats, and mice. The mammalian methylation array has several strengths: it applies to all mammalian species even those that have not yet been sequenced, it provides deep coverage of conserved cytosines facilitating the development epigenetic biomarkers, and it increases the probability that biological insights gained in one species will translate to others.
The impact of PNPLA3 and TM6SF2 in cirrhosis related complications
Authors: Shao Xue , Uojima Haruki , Arai Taeang , Ogawa Yuji , Setsu Toru , Atsukawa Masanori , Furuichi Yoshihiro , Arase Yoshitaka , Horio Kazue , Hidaka Hisashi , Nakazawa Takahide , Kako Makoto , Kagawa Tatehiro , Iwakiri Katsuhiko , Nakajima Atsushi , Terai Shuji , Tanaka Yasuhito , Koizumi Wasaburo
Published: January 2021

Patatin-like phospholipase domain-containing 3 (PNPLA3) and transmembrane 6-superfamily member 2 (TM6SF2) polymorphisms have major impact for fibrosis due to steatohepatitis. However, there are scant data about correlations between cirrhosis-related complications and the polymorphisms of these genes. Therefore, we aimed to determine the role of the PNPLA3 and TM6SF2 polymorphisms in fibrosis progression for patients with liver cirrhosis. A multicenter study was performed at six hospitals in Japan enrolling 400 patients with liver cirrhosis caused by virus (n = 157), alcohol (n = 104), nonalcoholic fatty liver disease (NAFLD) (n = 106), or autoimmune disease (n = 33). These cirrhotic patients included those with complications of variceal bleeding, hepatic ascites, and/or hepatic encephalopathy and those without. To assess the role of the PNPLA3 and TM6SF2 polymorphisms in patients with cirrhosis related complications, we calculated the odds ratio and relative risk for the rs738409 and rs58542926 polymorphisms. We also accessed whether or not the interaction between these two polymorphisms contributed to cirrhosis related complications. As a result, the odds ratio for complications in the NAFLD group significantly increased in the presence of the rs738409 GG genotype when the CC genotype was used as the reference. There were no significant risks between complications and the presence of the rs738409 G allele in the virus or alcohol groups. There were no significant risks of complications in the frequency of the rs58542926 T polymorphism regardless of the etiology of liver cirrhosis. The interaction between the rs738409 and rs58542926 polymorphisms had the highest odds ratio of 2.415 for complications in the rs738409 GG + rs58542926 (CT+TT) group when rs738409 (CC+CG) + TM6SF2 CC was used as the reference in the NAFLD group although there was no statistically interaction between these gene polymorphisms (P=0.085).
Concentration-dependent duality of bFGF in regulation of barrier properties of human brain endothelial cells
Authors: Kriaučiūnaitė Karolina , Pociūtė Agnė , Kaušylė Aida , Pajarskienė Justina , Verkhratsky Alexei , Pivoriūnas Augustas
Published: January 2021

Multiple paracrine factors regulate barrier properties of human brain capillary endothelial cells (BCECs). Understanding precise mode of action of these factors remains a challenging task because of the limited availability of functionally competent BCECs and use of serum-containing medium. In the present study we employed defined protocol for producing BCECs from human inducible pluripotent stem cells. We found that autocrine secretion of basic fibroblast growth factor (bFGF) is necessary for the establishment a tight BCECs barrier, as revealed by measurements of trans-endothelial electric resistance (TEER). In contrast, exogenous bFGF in concentrations exceeding 4 ng/ml inhibited TEER and proliferation of BCECs in a concentration-dependent manner. Exogenous bFGF did not significantly affect expression and distribution of tight junction proteins claudin-5, occludin and ZO-1. Treatment with FGF receptor blocker PD173074 (15 μM) suppressed inhibitory effects of bFGF and induced nuclear translocation of protein ZO-1. Inhibition of phosphoinositide 3-Kinase (PI-3K) with LY294002 (25 μM) significantly potentiated inhibitory effect of bFGF on TEER indicating that PI-3K signalling pathway partially suppress inhibitory effects of bFGF on TEER. In conclusion we show that autocrine bFGF secretion is necessary for the proper barrier function of BCECs, whereas exogenous bFGF suppresses barrier resistance in a concentration-dependent manner. Our findings demonstrate a dual role for bFGF in the regulation of BCEC barrier function.
Transcriptomic profiles ofPlasmodium falciparumandPlasmodium vivax-infected individuals in Indonesia
Authors: Bobowik Katalina , Syafruddin Din , Darusallam Chelzie Crenna , Sudoyo Herawati , Wells Christine , Romero Irene Gallego
Published: January 2021

Malaria is one of the leading causes of illness and death globally. The vast majority of transcriptomic studies of the impact of malaria on human hosts have been conducted on populations of African ancestry suffering fromPlasmodium falciparuminfection, making it unclear whether biological responses observed in these studies can be generalised to other populations. Here, we perform differential expression analysis between healthy controls and malaria-infected patients within Indonesia, a country of over 260 million people which has substantial morbidity due to endemic malaria. We find that in samples infected withP. falciparumandP. vivax, there is an upregulation of genes involved in inflammation, the immediate early immune response, translation, and apoptosis. When comparing these findings to transcriptomic studies conducted in Africa (onP. falciparum) and South America (onP. vivax), we find that many pathways are shared. This is particularly apparent for receptor recognition and inflammation-related genes inP. falciparumand innate immune and chemokine-related genes inP. vivaxinfection. However, we also find that many genes are unique to the Indonesian population, particularly those involved in RNA processing, splicing, and cell surface receptor genes. This study provides a more comprehensive view of malaria infection outside of Africa and contributes to a better characterisation of malaria pathogenesis within humans across a range of genetic architectures.
Increasing calling accuracy, coverage, and read depth in sequence data by the use of haplotype blocks
Authors: Pook Torsten , Nemri Adnane , Segovia Eric Gerardo Gonzalez , Simianer Henner , Schoen Chris-Carolin
Published: January 2021

High-throughput genotyping of large numbers of lines remains a key challenge in plant genetics, requiring geneticists and breeders to find a balance between data quality and the number of genotyped lines under a variety of different existing technologies when resources are limited. In this work, we are proposing a new imputation pipeline (“HBimpute”) that can be used to generate high-quality genomic data from low read-depth whole-genome-sequence data. The key idea of the pipeline is the use of haplotype blocks from the software HaploBlocker to identify locally similar lines and merge their reads locally. The effectiveness of the pipeline is showcased on a dataset of 321 doubled haploid lines of a European maize landrace, which were sequenced with 0.5X read-depth. Overall imputing error rates are cut in half compared to the state-of-the-art software BEAGLE, while the average read-depth is increased to 83X, thus enabling the calling of structural variation. The usefulness of the obtained imputed data panel is further evaluated by comparing the performance in common breeding applications to that of genomic data from a 600k array. In particular for genome-wide association studies, the sequence data is shown to be performing slightly better. Furthermore, genomic prediction based on the overlapping markers from the array and sequence is leading to a slightly higher predictive ability for the imputed sequence data, thereby indicating that the data quality obtained from low read-depth sequencing is on par or even slightly higher than high-density array data. When including all markers for the sequence data, the predictive ability is slightly reduced indicating overall lower data quality in non-array markers.

Author summary

High-throughput genotyping of large numbers of lines remains a key challenge in plant genetics and breeding. Cost, precision, and throughput must be balanced to achieve optimal efficiencies given available technologies and finite resources. Although genotyping arrays are still considered the gold standard in high-throughput quantitative genetics, recent advances in sequencing provide new opportunities for this. Both the quality and cost of genomic data generated based on sequencing are highly dependent on the used read depth. In this work, we are proposing a new imputation pipeline (“HBimpute”) that uses haplotype blocks to detect individuals of the same genetic origin and subsequently uses all reads of those individuals in the variant calling. Thus, the obtained virtual read depth is artificially increased, leading to higher calling accuracy, coverage, and the ability to all copy number variation based on relatively cheap low-read depth sequencing data. Thus, our approach makes sequencing a cost-competitive alternative to genotyping arrays with the additional benefit of the potential use of structural variation.
Microwell array based opto-electrochemical detections revealing co-adaptation of rheological properties and oxygen metabolism in budding yeast
Authors: Vajrala Venkata Suresh , Alric Baptiste , Laborde Adrian , Colin Camille , Suraniti Emmanuel , Temple-Boyer Pierre , Arbault Stephane , Delarue Morgan , Launay Jérome
Published: January 2021

Microdevices composed of microwell arrays integrating nanoelectrodes (OptoElecWell) were developed to achieve dual high-resolution optical and electrochemical detections on singleSaccharomyces cerevisiaebudding yeast cells. Each array consists in 1.6 × 105microwells of 8 µm diameter and 5 µm height, with a platinum nanoring electrode for in-situ electrochemistry, all integrated on a transparent thin wafer for further high-resolution live-cell imaging. After optimizing the filling rate, 32% of cells were effectively trapped within microwells. This allowed to analyseS. cerevisiaemetabolisms associated with basal respiration while simultaneously measuring optically other cellular parameters. In this study, we focused on the impact of glucose concentration on respiration and intracellular rheology. We found that while oxygen uptake rate decreased with increasing glucose concentration, diffusion of tracer nanoparticles increased. Our OptoElecWell based respiration methodology provided similar results compared to the commercial gold-standard Seahorse XF analyser, while using 20 times lesser biological samples, paving the way to achieve single cell metabolomics. In addition, it facilitates an optical route to monitor the contents within single cells. The proposed device, in combination with the dual detection analysis, opens up new avenues for measuring cellular metabolism, and relating it to various cellular physiological and rheological indicators at single cell level.
Multiomics Integration Elucidates Metabolic Modulators of Drug Resistance in Lymphoma
Authors: Choueiry Fouad , Singh Satishkumar , Sun Xiaowei , Zhang Shiqi , Sircar Anuvrat , Amber Hart , Alinari Lapo , Narendranath Epperla , Baiocchi Robert , Zhu Jiangjiang , Sehgal Lalit
Published: January 2021

Background

Diffuse large B-cell lymphoma (DLBCL) is the most common non-Hodgkin lymphoma (NHL). B-cell NHLs rely on Bruton’s tyrosine kinase (BTK) mediated B-cell receptor signaling for survival and disease progression. However, they are often resistant to BTK inhibitors or soon acquire resistance after drug exposure resulting in the drug tolerant form. The drug tolerant clones proliferate faster, have increased metabolic activity, and shift to oxidative phosphorylation; however, how this metabolic programming occurs in the drug resistant tumor is poorly understood.

Methods

In this study, we explored for the first time the metabolic regulators of ibrutinib-resistant activated B-cell (ABC) DLBCL using a ‘multi-omics’ analysis that integrated metabolomics (using high-resolution mass spectrometry) and transcriptomic (gene expression analysis). Overlay of the unbiased statistical analyses, genetic perturbation and pharmaceutical inhibition, were further used to identify the key players that contribute to the metabolic reprograming of the drug resistant clone.

Results

Gene-metabolite integration revealed interleukin 4 induced 1 (IL4I1) at the crosstalk of two significantly altered metabolic pathways involved in the production of various amino acids. We showed for the first time that drug resistant clones undergo metabolic reprogramming from glycolysis towards oxidative phosphorylation & is activated via the BTK-PI3K-AKT-IL4I1 axis and can be targeted therapeutically.

Conclusions

Our report shows how these cells become dependent on PI3K/AKT signaling for survival after acquiring ibrutinib resistance and shift to sustained Oxidative phosphorylation, additionally we outline the compensatory, pathway that regulates this metabolic reprogramming in the drug resistant cells. These findings from our unbiased analyses highlight the role of metabolic reprogramming during drug resistance development. Furthermore, our work demonstrates that a multi-omics approach can be a powerful and unbiased strategy to uncover genes and pathways that drive metabolic dysregulation in cancer cells.
Antibiotic treatment increases yellowness of carotenoid feather coloration in greenfinches (Chloris chloris)
Authors: Lind Mari-Ann , Sepp Tuul , Štšeglova Kristiina , Hõrak Peeter
Published: January 2021

Carotenoid plumage coloration is an important signal of quality, and plays an important role in mate choice in many bird species. However, it remains unclear what mechanism makes carotenoids an honest signal. Here, we test the hypothesis that carotenoid plumage coloration might indicate gut health. Parasitic and microbial infections can affect nutrient absorption due to decreased gut surface or by altered gut microbiome. We took an advantage of a naturally occurring coinfection of parasites inhabiting the upper and lower portions of the digestive track to distinguish between the direct and indirect effects of parasites on carotenoid acquisition. Protozoan coccidian intestinal parasites are widespread in greenfinches (Chloris chloris) and the majority of greenfinches are infected in nature. Trichomonosis is an emerging disease of the upper digestive track that causes high mortality among greenfinches. We captured wild greenfinches (N=71) and administered anticoccidial medication toltrazuril (TOLTRA) to one group, antibiotic metronidazole (METRO) that is also effective for treatingTrichomonas gallinea, to the second group, and third group received no medication. In the METRO group, feathers grown during the experiment had significantly higher chroma of yellow parts, but there was no effect of TOLTRA on feather chroma. These results suggest that METRO increased the efficiency of carotenoid modification or deposition to the feathers rather than nutrient acquisition, and/or freed energy resources that could be invested in coloration. Alternatively, in accordance with shared pathway hypothesis, increase in efficiency of vital cellular processes might have occurred, as many microbial metabolites can modulate mitochondrial and immune function.
Sexually Transmitted Infections Select for Different Levels of Immunocompetence And Reproductive Efforts in The Two Sexes
Authors: Aldakak Lafi , Rühli Frank , Bender Nicole
Published: January 2021

1.

Sex differences in immunity have been described in humans and other mammal species. Females have a lower incidence of infections and non-reproductive malignancies and exhibit higher antibody levels after vaccination. Existing evolutionary explanations are based on differences in reproductive strategies and reaction to extrinsic differences in susceptibility and virulence between the sexes. Here, we test the hypothesis that known differences in the probability of transmission and outcome of sexually transmitted infections contribute to sex differences in immunocompetence. We modelled reproductive and immune investments against a fertility limiting Sexually Transmitted Infection (STI). We show that, in line with previous findings, increased susceptibility selects for tolerance to the parasite while increased virulence selects for resistance against it. Differences in reproductive strategies between the sexes lead to sex differences in immunocompetence, mostly with higher competence in females. Extrinsic differences in susceptibility and virulence between the sexes can augment or alleviate the evolutionary consequences of intrinsic differences depending on their direction and magnitude. This indicates that the selection of sex-specific immune strategies is less predictable than thought before and explains why sex differences in immunity have been found to be not universal and pervasive across animal species.
GLS-driven glutamine catabolism contributes to prostate cancer radiosensitivity by regulating the redox state, stemness and ATG5-mediated autophagy
Authors: Mukha Anna , Kahya Uğur , Linge Annett , Chen Oleg , Löck Steffen , Lukiyanchuk Vasyl , Richter Susan , Alves Tiago C. , Peitzsch Mirko , Telychko Vladyslav , Skvortsov Sergej , Negro Giulia , Aschenbrenner Bertram , Skvortsova Ira-Ida , Mirtschink Peter , Lohaus Fabian , Hölscher Tobias , Neubauer Hans , Rivandi Mahdi , Franken André , Behrens Bianca , Stoecklein Nikolas H. , Toma Marieta , Sommer Ulrich , Zschaeck Sebastian , Rehm Maximilian , Eisenhofer Graeme , Schwager Christian , Abdollahi Amir , Groeben Christer , Kunz-Schughart Leoni A. , Baretton Gustavo B. , Baumann Michael , Krause Mechthild , Peitzsch Claudia , Dubrovska Anna
Published: January 2021

Radiotherapy is one of the curative treatment options for localized prostate cancer (PCa). The curative potential of radiotherapy is mediated by irradiation-induced oxidative stress and DNA damage in tumor cells. However, PCa radiocurability can be impeded by tumor resistance mechanisms and normal tissue toxicity. Metabolic reprogramming is one of the major hallmarks of tumor progression and therapy resistance. Here, we found that radioresistant PCa cells and prostate cancer stem cells (CSCs) have a high glutamine demand. Glutaminase (GLS)-driven catabolism of glutamine serves not only for energy production but also for the maintenance of the redox state. Consequently, glutamine depletion or inhibition of critical regulators of glutamine utilization, such as glutaminase (GLS) and the transcription factor MYC results in PCa radiosensitization. On the contrary, we found that a combination of glutamine metabolism inhibitors with irradiation does not cause toxic effects on nonmalignant prostate cells. Glutamine catabolism contributes to the maintenance of CSCs through regulation of the alpha-ketoglutarate (α-KG)-dependent chromatin-modifying dioxygenase. The lack of glutamine results in the inhibition of CSCs with a high aldehyde dehydrogenase (ALDH) activity, decreases the frequency of the CSC populationsin vivoand reduces tumor formation in xenograft mouse models. Moreover, this study shows that activation of the ATG5-mediated autophagy in response to a lack of glutamine is a tumor survival strategy to withstand radiation-mediated cell damage. In combination with autophagy inhibition, the blockade of glutamine metabolism might be a promising strategy for PCa radiosensitization. High blood levels of glutamine in PCa patients significantly correlate with a shorter prostate-specific antigen (PSA) doubling time. Furthermore, high expression of critical regulators of glutamine metabolism, GLS1 and MYC, is significantly associated with a decreased progression-free survival in PCa patients treated with radiotherapy. Our findings demonstrate that GLS-driven glutaminolysis is a prognostic biomarker and therapeutic target for PCa radiosensitization.
A Self-Supervised Machine Learning Approach for Objective Live Cell Segmentation and Analysis
Authors: Robitaille Michael C. , Byers Jeff M. , Christodoulides Joseph A. , Raphael Marc P.
Published: January 2021

Machine learning algorithms hold the promise of greatly improving live cell image analysis by way of (1) analyzing far more imagery than can be achieved by more traditional manual approaches and (2) by eliminating the subjective nature of researchers and diagnosticians selecting the cells or cell features to be included in the analyzed data set. Currently, however, even the most sophisticated model based or machine learning algorithms require user supervision, meaning the subjectivity problem is not removed but rather incorporated into the algorithm’s initial training steps and then repeatedly applied to the imagery. To address this roadblock, we have developed a self-supervised machine learning algorithm that recursively trains itself directly from the live cell imagery data, thus providing objective segmentation and quantification. The approach incorporates an optical flow algorithm component to self-label cell and background pixels for training, followed by the extraction of additional feature vectors for the automated generation of a cell/background classification model. Because it is self-trained, the software has no user-adjustable parameters and does not require curated training imagery. The algorithm was applied to automatically segment cells from their background for a variety of cell types and five commonly used imaging modalities - fluorescence, phase contrast, differential interference contrast (DIC), transmitted light and interference reflection microscopy (IRM). The approach is broadly applicable in that it enables completely automated cell segmentation for long-term live cell phenotyping applications, regardless of the input imagery’s optical modality, magnification or cell type.
dynUGENE: an R package for uncertainty-aware gene regulatory network inference, simulation, and visualization
Authors: Lu Tianyu , Silva Anjali
Published: January 2021

Methods for gene regulatory network inference focus on network architecture identification but neglect model selection and simulation. We implement an extension to the dynGENIE3 algorithm that accounts for model uncertainty as an R package, providing users with an easy to use interface for model selection and gene expression profile simulation. Source code is available athttps://github.com/tianyu-lu/dynUGENEwith a detailed user guide. A webserver with interactive controls is available athttps://tianyulu.shinyapps.io/dynUGENE/.
Priming anodal transcranial direct current stimulation attenuates short interval intracortical inhibition and increases time to task failure of a constant workload cycling exercise
Authors: Sidhu Simranjit K
Published: January 2021

Transcranial direct current stimulation (tDCS), a non-invasive neuromodulatory technique has been shown to increase the excitability of targeted brain area and influence endurance exercise performance. However, tDCS-mediated interaction between corticospinal excitability, GABAAmediated intracortical inhibition and endurance exercise performance remains understudied. In two separate sessions, twelve subjects performed fatigue cycling exercise (80% peak power output) sustained to task failure in a double-blinded design, following either ten minutes of anodal tDCS (atDCS) or sham. Corticospinal excitability and short interval intracortical inhibition (SICI) were measured at baseline, post neuromodulation and post-exercise using paired-pulse transcranial magnetic stimulation (TMS) in a resting hand muscle. There was a greater a decrease in SICI (P< 0.05) post fatigue cycling with atDCS priming compared to sham. Time to task failure (TTF) was significantly increased following atDCS compared to sham (P< 0.05). These findings suggest that atDCS applied over the motor cortex can augment cycling exercise performance; and this outcome may be mediated via a decrease in the excitability of GABAAinhibitory interneurons.
Testing for phylogenetic signal in single-cell RNA-seq data
Authors: Moravec Jiří C. , Lanfear Rob , Spector David L. , Diermeier Sarah D. , Gavryushkin Alex
Published: January 2021

Phylogenetic methods are emerging as a useful tool to understand cancer evolutionary dynamics, including tumor structure, heterogeneity, and progression. Most currently used approaches utilize either bulk whole genome sequencing (WGS) or single-cell DNA sequencing (scDNA-seq) and are based on calling copy number alterations and single nucleotide variants (SNVs). scRNA-seq is commonly applied to explore differential gene expression of cancer cells throughout tumor progression. The method exacerbates the single-cell sequencing problem of low yield per cell with uneven expression levels. This accounts for low and uneven sequencing coverage and makes SNV detection and phylogenetic analysis challenging. In this paper, we demonstrate for the first time that scRNA-seq data contains sufficient evolutionary signal and can also be utilized in phylogenetic analyses. We explore and compare results of such analyses based on both expression levels and SNVs called from scRNA-seq data. Both techniques are shown to be useful for reconstructing phylogenetic relationships between cells, reflecting the clonal composition of a tumor. Both standardized expression values and SNVs appear to be equally capable of reconstructing a similar pattern of phylogenetic relationship. This pattern is stable even when phylogenetic uncertainty is taken in account. Our results open up a new direction of somatic phylogenetics based on scRNA-seq data. Further research is required to refine and improve these approaches to capture the full picture of somatic evolutionary dynamics in cancer.
Neutralizing antibodies targeting the SARS-CoV-2 receptor binding domain isolated from a naïve human antibody library
Authors: Bell Benjamin N. , Powell Abigail E. , Rodriguez Carlos , Cochran Jennifer R. , Kim Peter S.
Published: January 2021

Infection with SARS-CoV-2 elicits robust antibody responses in some patients, with a majority of the response directed at the receptor binding domain (RBD) of the spike surface glycoprotein. Remarkably, many patient-derived antibodies that potently inhibit viral infection harbor few to no mutations from the germline, suggesting that naïve antibody libraries are a viable means for discovery of novel SARS-CoV-2 neutralizing antibodies. Here, we used a yeast surface-display library of human naïve antibodies to isolate and characterize three novel neutralizing antibodies that target the RBD: one that blocks interaction with angiotensin-converting enzyme 2 (ACE2), the human receptor for SARS-CoV-2, and two that target other epitopes on the RBD. These three antibodies neutralized SARS-CoV-2 spike-pseudotyped lentivirus with IC50values as low as 60 ng/mLin vitro. Using a biolayer interferometry-based binding competition assay, we determined that these antibodies have distinct but overlapping epitopes with antibodies elicited during natural COVID-19 infection. Taken together, these analyses highlight howin vitroselection of naïve antibodies can mimic the humoral responsein vivo, yielding neutralizing antibodies and various epitopes that can be effectively targeted on the SARS-CoV-2 RBD.
Targeted sequence capture ofOrientia tsutsugamushiDNA from chiggers and humans
Authors: Elliott Ivo , Thangnimitchok Neeranuch , de Cesare Mariateresa , Linsuwanon Piyada , Paris Daniel H. , Day Nicholas PJ , Newton Paul N. , Bowden Rory , Batty Elizabeth M.
Published: January 2021

Scrub typhus is a febrile disease caused byOrientia tsutsugamushi, transmitted by larval stage Trombiculid mites (chiggers), whose primary hosts are small mammals. The phylogenomics ofO. tsutsugamushiin chiggers, small mammals and humans remains poorly understood. To combat the limitations imposed by the low relative quantities of pathogen DNA in typicalO. tsutsugamushiclinical and ecological samples, along with the technical, safety and cost limitations of cell culture, a novel probe-based target enrichment sequencing protocol was developed. The method was designed to capture variation among conserved genes and facilitate phylogenomic analysis at the scale of population samples. A whole-genome amplification step was incorporated to enhance the efficiency of sequencing by reducing duplication rates. This resulted in on-target capture rates of up to 93% for a diverse set of human, chigger, and rodent samples, with the greatest success rate in samples with real-time PCR Ctvalues below 35. Analysis of the best-performing samples revealed phylogeographic clustering at local, provincial and international scales. Applying the methodology to a comprehensive set of samples could yield a more complete understanding of the ecology, genomic evolution and population structure ofO. tsutsugamushiand other similarly challenging organisms, with potential benefits in the development of diagnostic tests and vaccines.
Therapeutic adenine base editing corrects nonsense mutation and improves visual function in a mouse model of Leber congenital amaurosis
Authors: Jo Dong Hyun , Jang Hyeon-Ki , Cho Chang Sik , Han Jun Hee , Ryu Gahee , Jung Youngri , Bae Sangsu , Kim Jeong Hun
Published: January 2021

Leber congenital amaurosis (LCA) is an inherited retinal degeneration that causes severe visual dysfunction in children and adolescents. In patients with LCA, pathogenic variants are evident in specific genes, such as RPE65, which are related to the functions of retinal pigment epithelium and photoreceptors. Base editing confers a way to correct pathogenic substitutions without double-stranded breaks in contrast to the original Cas9. In this study, we prepared dual adeno-associated virus vectors containing the split adenine base editors with trans-splicing intein (AAV-ABE) for in vivo adenine base editing in retinal degeneration 12 (rd12) mice, an animal model of LCA, which possess a nonsense mutation of C to T transition in the Rpe65 gene (p.R44X). AAV-ABE induced an A to G transition in retinal pigment epithelial cells of rd12 mice when injected into the subretinal space. The on-target editing was sufficient to recover wild-type mRNA, RPE65 protein, and light-induced electrical responses of retinal tissues. We suggest adenine base editing to correct pathogenic variants in the treatment of LCA.
Surveillance of Shiga toxin-producingEscherichia coliassociated bloody diarrhea in Argentina
Authors: Rivas Marta , Pichel Mariana , Colonna Mariana , Casanello Adrián López , Alconcher Laura F. , Galavotti Jimena , Principi Iliana , Araujo Sofía Pérez , Ramírez Flavia B. , González Gladys , Pianciola Luis A. , Mazzeo Melina , Suarez Ángela , Oderiz Sebastián , Ghezzi Lidia F. R. , Arrigo Diego J. , Paladini José H. , Baroni María R. , Pérez Susana , Tamborini Ana , Chinen Isabel , Miliwebsky Elizabeth S. , Goldbaum Fernando , Muñoz Luciana , Spatz Linus , Sanguineti Santiago ,   the EPI-HUS Investigation Team
Published: January 2021

In Argentina, the hemolytic uremic syndrome associated with Shiga toxin-producingEscherichia coli(STEC-HUS) infection is endemic, and reliable data about prevalence and risk factors are available since 2000. However, information about STEC-associated bloody diarrhea (BD) cases is limited. A prospective study was carried out in seven tertiary-hospitals and 18 Referral Units from different regions, aiming to determine (i) STEC-positive BD cases frequency in 714 children aged 1 to 9 years old; and (ii) rate of progression to HUS. The number and regional distribution of STEC-HUS cases assisted in the same hospitals and period was also assessed. A total of 29 (4.1%) STEC-positive BD cases were confirmed by Shiga Toxin Quik Chek (STQC) and/or mPCR. The highest frequencies were found in the Southern region (Neuquén, 8.7%; Bahía Blanca, 7.9%), in children between 12 and 23 month of age (8.8%), during summertime. Four (13.8%) cases progressed to HUS, three to five days after BD onset. Twenty-seven STEC-HUS children mainly under 5 years old (77.8%) were enrolled, 51.9% were female; 44% were Stx-positive by STQC and all by mPCR. The most common serotypes were O157:H7 and O145:H28 and prevalent genotypes werestx2a-only or associated, both among BD and HUS cases. Considering the endemic behavior of HUS and its impact on public health, it is important to have updated information about the epidemiology of the diarrheal disease for early recognition of infected patients and initiation of supportive treatment. Finally, it also gives the opportunity to respond to outbreak situations effectively and in timely manner.
Hunting Eagles with glass mice: revisiting the inoculum effect forStreptococcus pyogeneswith a hollow fibre infection model
Authors: Marum Darcy , Manning Laurens , Raby Edward
Published: January 2021

Background

Severe, invasiveStreptococcus pyogenes(Strep A) infections result in greater than 500,000 deaths annually. First line treatment for such infections is combination benzylpenicillin and clindamycin, but treatment failure can occur with this regimen. This failure has been partially attributed to the inoculum effect, which presents as reduced antibiotic susceptibility during high bacterial density and plateau-phase growth. Hollow fibre infection models (HFIM) have been proposed as an alternative toin vivoresearch to study these effects.

Objectives

To re-evaluate the inoculum effect for benzylpenicillin, clindamycin, linezolid and trimethoprim-sulfamethoxazole using a Strep A HFIM.

Methods

Differential antibiotic susceptibility of Strep A was measured in a HFIM starting from low- and high-density inocula. Dynamic antibiotic concentrations were delivered over 48 hours to simulate human pharmacokinetics. Differences in antibiotic susceptibility were determined at 24 and 48 hours by plate count of remaining viable colony-forming units.

Results

Inoculum effects were seen in benzylpenicillin and linezolid at 24 hours, and benzylpenicillin, linezolid and clindamycin at 48 hours. The effect size was greatest for continuously infused benzylpenicillin. No inoculum effect was seen in trimethoprim-sulfamethoxazole.

Conclusions

Inoculum effects were seen in the HFIM model using benzylpenicillin, linezolid and clindamycin, which may predict reduced clinical efficacy following treatment delay. The model has proven robust and largely in agreeance with published data, recommending it for further Strep A study.
EdU incorporation to assess cell proliferation and drug susceptibility for the brain-eating amoeba,Naegleria fowleri
Authors: Troth Emma V. , Kyle Dennis E.
Published: January 2021

Naegleria fowleriis a pathogenic free-living amoeba that is commonly found in warm, freshwater and can cause a rapidly fulminant disease known as primary amoebic meningoencephalitis (PAM). New drugs are urgently needed to treat PAM, as the fatality rate is >97%. Until recently, few advances have been made in the discovery of new drugs forN. fowleriand one drawback is the lack of validated tools and methods to enhance drug discovery and diagnostics research. In this study we aimed to validate alternative methods to assess cell proliferation that are commonly used for other cell types and develop a novel drug screening assay to evaluate drug efficacy onN. fowlerireplication. EdU (5-ethynyl-2′-deoxyuridine) is a pyrimidine analog of thymidine that can be used as a quantitative endpoint for cell proliferation. EdU incorporation is detected via a copper catalyzed click reaction with an Alexa Fluor linked azide. EdU incorporation in replicatingN. fowleriwas validated using fluorescence microscopy and quantitative methods for assessing EdU incorporation were developed by using an imaging flow cytometer. Currently used PAM therapeutics inhibitedN. fowlerireplication and EdU incorporationin vitro. EdA (5′ethynyl-2′-deoxyadenosine), an adenine analog, also was incorporated by N. fowleri, but was more cytotoxic than EdU. In summary, EdU incorporation could be used as a complimentary method for drug discovery for these neglected pathogens.
Quality of life and health-related utility after head&neck cancer surgery
Authors: Parimbelli Enea , Simon Christian , Soldati Federico , Duchoud Lorry , Armas Gian Luca , de Almeida John , Quaglini Silvana
Published: January 2021

Purpose

This work describes the methodology adopted and the results obtained in a utility elicitation task. The purpose was to elicit utility coefficients (UCs) needed to calculate quality-adjusted life years for a cost/utility analysis of TORS (Trans-Oral robotic Surgery) versus TLM (Trans-oral Laser Microsurgery), which are two minimally-invasive trans-oral surgery techniques for head & neck cancers.

Methods

Since the economic evaluation would be conducted from the point of view of the Swiss healthcare system, Swiss people (healthy volunteers) have been interviewed in order to tailor the model to that specific country. The utility elicitation was performed using a computerized tool (UceWeb). Standard gamble and rating scale methods were used.

Results

UCs have been elicited from 47 individuals, each one providing values for 18 health states, for a total of 1692 expected values. Health states, described using graphical factsheets, ranged from remission to palliative care. Elicited UCs were different among states, ranging from 0.980 to 0.213. Those values were comparable to previously published results from a Canadian population, except for states related to recurrent disease (local, regional, and distant), and palliation, where the Swiss population showed lower utility values.

Conclusion

From a methodological point of view, our study shows that the UceWeb tool can be profitably used for utility elicitation from healthy volunteers. From an application point of view, the study provides utility values that can be used not only for a specific cost-utility analysis, but for future studies involving health states following trans-oral head & neck surgery. Moreover, the study confirms that some UCs vary among countries, demanding for tailored elicitation tasks.
The incremental value of computed tomography of COVID-19 pneumonia in predicting ICU admission
Authors: Bartolucci Maurizio , Benelli Matteo , Betti Margherita , Bicchi Sara , Fedeli Luca , Giannelli Federico , Aquilini Donatella , Baldini Alessio , Consales Guglielmo , Di Natale Massimo Edoardo , Lotti Pamela , Vannucchi Letizia , Trezzi Michele , Mazzoni Lorenzo Nicola , Santini Sandro , Carpi Roberto , Matarrese Daniela , Bernardi Luca , Mascalchi Mario ,   COVID Working Group USL Toscana Centro and Azienda Ospedaliero-Universitaria Careggi,   members of the Working Group USL Toscana Centro and Azienda Ospedaliero-Universitaria Careggi, Allescia Germana , Busoni Simone , Bindi Alessandra , Cavigli Edoardo , Cozzi Diletta , Dedola Giovanni Luca , Mazzocchi Silvia , Miele Vittorio , Pozzessere Chiara , Taddeucci Adriana , Viviani Adriano , Zini Chiara
Published: January 2021

Rationale

Triage is crucial for patient’s management and estimation of the required Intensive Care Unit (ICU) beds is fundamental for Health Systems during the COVID-19 pandemic.

Objective

To assess whether chest Computed Tomography (CT) of COVID-19 pneumonia has an incremental role in predicting patient’s admission to ICU.

Methods

We performed volumetric and texture analysis of the areas of the affected lung in CT of 115 outpatients with COVID-19 infection presenting to the Emergency Room with dyspnea and unresponsive hypoxyemia. Admission blood laboratory including lymphocyte count, serum lactate dehydrogenase, D-dimer and C-Reactive Protein and the ratio between the arterial partial pressure of oxygen and inspired oxygen were collected. By calculating the areas under the receiver-operating characteristic curves (AUC), we compared the performance of blood laboratory-arterial gas analyses features alone and combined with the CT features in two hybrid models (Hybrid radiological and Hybrid radiomics)for predicting ICU admission. Following a machine learning approach, 63 patients were allocated to the training and 52 to the validation set.

Measurements and Main Results

Twenty-nine (25%) of patients were admitted to ICU. The Hybrid radiological model comprising the lung %consolidation performed significantly (p=0.04) better in predicting ICU admission in the validation (AUC=0.82; 95%Confidence Interval 0.68-0.95) set than the blood laboratory-arterial gas analyses features alone (AUC=0.71; 95%Confidence Interval 0.56-0.86). A risk calculator for ICU admission was derived and is available at:https://github.com/cgplab/covidapp

Conclusions

The volume of the consolidated lung in CT of patients with COVID-19 pneumonia has a mild but significant incremental value in predicting ICU admission.
Levels of SARS-CoV-2 population exposure are considerably higher than suggested by seroprevalence surveys
Authors: Chen Siyu , Flegg Jennifer A , White Lisa J , Aguas Ricardo
Published: January 2021

Accurate knowledge of accurate levels of prior population exposure has critical ramifications for preparedness plans of subsequent SARS-CoV-2 epidemic waves and vaccine prioritization strategies. Serological studies can be used to estimate levels of past exposure and thus position populations in their epidemic timeline. To circumvent biases introduced by decaying antibody titers over time, population exposure estimation methods should account for seroreversion, to reflect that changes in seroprevalence measures over time are the net effect of increases due to recent transmission and decreases due to antibody waning. Here, we present a new method that combines multiple datasets (serology, mortality, and virus positivity ratios) to estimate seroreversion time and infection fatality ratios and simultaneously infer population exposure levels. The results indicate that the average time to seroreversion is six months, and that true exposure may be more than double the current seroprevalence levels reported for several regions of England.
COVID-19 seroprevalence among healthcare workers of a large COVID Hospital in Rome reveals strengths and limits of two different serological tests
Authors: Vetrugno Giuseppe , La Milia Daniele Ignazio , D’Ambrosio Floriana , Di Pumpo Marcello , Pastorino Roberta , Boccia Stefania , Ricci Rosalba , De-Giorgio Fabio , Cicconi Michela , Foti Federica , Pascucci Domenico , Castrini Francesco , Carini Elettra , Cambieri Andrea , D’Alfonso Maria Elena , Capalbo Gennaro , Fantoni Massimo , Moscato Umberto , Staiti Domenico , De Simone Francesco Maria , Berloco Filippo , Zega Maurizio , Cattani Paola , Posteraro Brunella , Sanguinetti Maurizio , Laurenti Patrizia
Published: January 2021

In several hospitals worldwide, healthcare workers are currently at the forefront against coronavirus disease 2019 (COVID-19). Since Fondazione Policlinico Universitario A. Gemelli (FPG) IRCCS has been enlisted as a COVID hospital, healthcare workers deployed to COVID wards were separated from those with limited or no exposure, whereas administrative staff was destined to work-from-home. Between June 4 and July 3 2020, an investigation was carried out to evaluate seroprevalence of SARS-CoV-2 IgG antibodies among employees of the FPG using point-of-care (POC) and venous blood tests. Sensitivity, specificity and predictive values were determined with reverse-transcription polymerase chain reaction (RT-PCR) on nasal/oropharyngeal swabs as gold standard.

Four thousand, seven hundred seventy-seven participants were enrolled. Seroprevalence was 3.66% using the POC test and 1.19% using venous blood test, with a significant difference between the two (p < 0.05).

POC sensitivity and specificity were, respectively, 63.64% (95% confidence interval (CI): 62.20% to 65.04%) and 96.64% (95% CI: 96.05% to 97.13%), while those of the venous blood test were, respectively, 78.79% (95% CI: 77.58% to 79.94%) and 99.36% (95% CI: 99.07% to 99.55%). Among low-risk population, point-of-care’s predictive values were 58.33% (positive) and 98.23% (negative) whereas venous blood test’s were 92.86% (positive) and 98.53% (negative). In conclusion, point-of-care tests have low diagnostic accuracy, while venous blood tests seem to show an overall poor reliability.
The Effect of Temperature on Covid-19 Confirmed Cases: Evidence from US Counties
Authors: NoghaniBehambari Hamid , Salari Mahmoud , Noghani Farzaneh , Tavassoli Nahid
Published: January 2021

This paper studies the effect of air temperature on the transmission of COVID-19 in the U.S. using daily observations across counties. This study uses various ordinary least squares (OLS) models with a comprehensive set of fixed effects to overcome unobserved heterogeneity issues across counties as well as the generalized method of moments (GMM) estimators as dynamic models to address endogeneity issue. Our main results indicate that an increase of one degree in temperature is associated with a reduction of 0.041 cases per 100,000 population at the county-level. We run several robustness tests and all the models confirm the impact of temperature on COVID-19 confirmed new cases. These results help policymakers and economists in optimizing decisions and investments to reduce COVID- 19 new cases.

JEL Codes

I10; Q51; Q54; H12
Whole-Cell Dissociated Suspension Analysis in Human Brain Neurodegenerative Disease: A Pilot Study
Authors: Serrano Geidy E. , Walker Jessica E. , Intorcia Anthony J. , Glass Michael J. , Arce Richard A. , Piras Ignazio S. , Talboom Joshua S. , Nelson Courtney M. , Cutler Brett D. , Sue Lucia I. , Lue Lih-Fen , Huentelman Matthew , Beach Thomas G.
Published: January 2021

Biochemical analysis of human brain tissue is typically done by homogenizing whole pieces of brain and separately characterizing the proteins, RNA, DNA, and other macromolecules within. While this has been sufficient to identify substantial changes, there is little ability to identify small changes or alterations that may occur in subsets of cells. To effectively investigate the biochemistry of disease in the brain, with its different cell types, we must first separate the cells and study them as phenotypically defined populations or even as individuals. In this project, we developed a new method for the generation of whole-cell-dissociated-suspensions (WCDS) in fresh human brain tissue that could be shared as a resource with scientists to study single human cells or populations. Characterization of WCDS was done in paraffin-embedded sections stained with H&E, and by phenotyping with antibodies using immunohistochemistry and fluorescence-activated cell sorting (FACS). Additionally, we compared extracted RNA from WCDS with RNA from adjacent intact cortical tissue, using RT-qPCR for cell-type-specific RNA for the same markers as well as whole transcriptome sequencing. More than 11,626 gene transcripts were successfully sequenced and classified using an external database either as being mainly expressed in neurons, astrocytes, microglia, oligodendrocytes, endothelial cells, or mixed (in two or more cell types). This demonstrates that we are currently capable of producing WCDS with a full representation of different brain cell types combined with RNA quality suitable for use in biochemical analysis.