bluuebunny/crossref_metadata_2025 huggingface dataset:
Detection and Recognition of Ultrasound Breast Nodules Based on Semi-supervised deep learning
Authors: Gao Yanhua , Liu Bo , Zhu Yuan , Chen Lin , Tan Miao , Xiao Xiaozhou , Guo Youmin
Published: April 2020

Background

The successful application of deep learning in medical images requires a large amount of annotation data for supervised training. However, massive labeling of medical data is expensive and time consuming. This paper proposes a semi-supervised deep learning method for the detection and classification of benign and malignant breast nodules in ultrasound images, which include two phases.

Methods

The nodule position in the ultrasound image is firstly detected using the faster RCNN network. Second, the recognition network is used to identify the benign and malignant types of nodules. The method in this paper uses a semi-supervised learning strategy, using 800 labeled nodules and 4396 unlabeled nodules.

Results

Based on mean teacher training strategy, the proposed semi-supervised network has obtained excellent results, which is similar to currently used with supervised training networks. On the two test data sets, the AUC of semi-supervised learning and supervised learning were: 93.7% vs 94.2% and 92% vs 92.3%.

Conclusions

The paper proves that semi-supervised learning strategies have good application potential in medical images. Based on a special learning strategy, the result of semi-supervised learning is expected to achieve close or even achieve similar result of supervised deep learning, which only need a small number of labeled samples and a large number of unlabeled samples. It means deep learning analysis of breast lesion will be more feasible and more efficient.
Cell-free DNA Profiling Informs Major Complications of Hematopoietic Cell Transplantation
Authors: Cheng Alexandre Pellan , Cheng Matthew Pellan , Loy Conor James , Lenz Joan Sesing , Chen Kaiwen , Smalling Sami , Burnham Philip , Timblin Kaitlyn Marie , Orejas José Luis , Silverman Emily , Polak Paz , Marty Francisco M. , Ritz Jerome , De Vlaminck Iwijn
Published: April 2020

Allogeneic hematopoietic cell transplantation (HCT) provides effective treatment for hematologic malignancies and immune disorders. Monitoring of post-transplant complications is critical, yet current diagnostic options are limited. Here, we show that cell-free DNA (cfDNA) in blood is a highly versatile analyte for monitoring of the most important complications that occur after HCT: graft-versus-host disease (GVHD), a frequent immune complication of HCT; infection; relapse of underlying disease; and graft failure. We demonstrate that these different therapeutic complications can be informed from a single assay, low-coverage bisulfite sequencing of cfDNA, followed by disease-specific bioinformatic analyses. To inform GVHD, we profile cfDNA methylation marks to trace the cfDNA tissues-of-origin and to quantify tissue-specific injury. To inform on infections, we implement metagenomic cfDNA profiling. To inform cancer relapse, we implement analyses of tumor-specific genomic aberrations. Finally, to detect graft failure we quantify the proportion of donor and recipient specific cfDNA. We applied this assay to 170 plasma samples collected from 27 HCT recipients at predetermined time points before and after allogeneic HCT. We found that the abundance of solid-organ derived cfDNA in the blood at one-month after HCT is an early predictor of acute graft-versus-host disease (area under the curve, 0.88). Metagenomic profiling of cfDNA revealed the frequent occurrence of viral reactivation in this patient population. The fraction of donor specific cfDNA was indicative of cell chimerism, relapse and remission, and the fraction of tumor specific cfDNA was informative of cancer relapse. This proof-of-principle study shows that cfDNA has the potential to improve the care of allogeneic HCT recipients by enabling earlier detection and better prediction of the complex array of complications that occur after HCT.
Mathematical Analysis of a COVID-19 Epidemic Model by using Data Driven Epidemiological Parameters of Diseases Spread in India
Authors: Pal D. , Ghosh D. , Santra P.K. , Mahapatra G.S.
Published: April 2020

This paper attempts to describe the outbreak of Severe Acute Respiratory Syndrome Coronavirus 2 (COVID-19) via an epidemic model. This virus has dissimilar effects in different countries. The number of new active coronavirus cases is increasing gradually across the globe. India is now in the second stage of COVID-19 spreading, it will be an epidemic very quickly if proper protection is not undertaken based on the database of the transmission of the disease. This paper is using the current data of COVID-19 for the mathematical modeling and its dynamical analysis. We bring in a new representation to appraise and manage the outbreak of infectious disease COVID-19 through SEQIR pandemic model, which is based on the supposition that the infected but undetected by testing individuals are send to quarantine during the incubation period. During the incubation period if any individual be infected by COVID-19, then that confirmed infected individuals are isolated and the necessary treatments are arranged so that they cannot taint the other residents in the community. Dynamics of the SEQIR model is presented by basic reproduction number Ro and the comprehensive stability analysis. Numerical results are depicted through apt graphical appearances using the data of five states and India.
A fractional-order SEIHDR model for COVID-19 with inter-city networked coupling effects
Authors: Lu Zhenzhen , Yu Yongguang , Chen YangQuan , Ren Guojian , Xu Conghui , Wang Shuhui , Yin Zhe
Published: April 2020

A novel coronavirus, designated as COVID-19, emerged in Wuhan, China, at the end of 2019. In this paper, a mathematical model is proposed to analyze the dynamic behavior of COVID-19. Based on inter-city networked coupling effects, a fractional-order SEIHDR system with the real-data from 23 January to 18 March, 2020 of COVID19 is discussed. Meanwhile, hospitalized individuals and the mortality rates of three types of individuals (exposed, infected and hospitalized) are firstly taken into account in the proposed model. And infectivity of individuals during incubation is also considered in this paper. By applying least squares method and predictor-correctors scheme, the numerical solutions of the proposed system in the absence of the inter-city network and with the inter-city network are stimulated by using the real-data from 23 January to 18 −mMarch, 2020 wheremis equal to the number of prediction days. Compared with integer-order system (α= 0), the fractional-order model without network is validated to have a better fitting of the data on Beijing, Shanghai, Wuhan, Huanggang and other cities. In contrast to the case without network, the results indicate that the inter-city network system may be not a significant case to virus spreading for China because of the lock down and quarantine measures, however, it may have an impact on cities that have not adopted city closure. Meanwhile, the proposed model better fits the data from 24 February to 31, March in Italy, and the peak number of confirmed people is also predicted by this fraction-order model. Furthermore, the existence and uniqueness of a bounded solution under the initial condition are considered in the proposed system. Afterwards, the basic reproduction numberR0is analyzed and it is found to hold a threshold: the disease-free equilibrium point is locally asymptotically stable whenR0≤ 1, which provides a theoretical basis for whether COVID-19 will become a pandemic in the future.
Volatile allosteric antagonists of mosquito odorant receptors inhibit normal odor-dependent behaviors
Authors: Kythreoti Georgia , Sdralia Nadia , Tsitoura Panagiota , Papachristos Dimitrios P. , Michaelakis Antonios , Karras Vasileios , Ruel David M. , Yakir Esther , Bohbot Jonathan D. , Schulz Stefan , Iatrou Kostas
Published: April 2020

Odorant-dependent behaviors in insects are triggered by the binding of odorant ligands to the variable subunits of heteromeric olfactory receptors. Previous studies have shown, however, that specific odor binding to ORco, the common subunit of odorant receptor heteromers, may alter allosterically olfactory receptor function and affect profoundly subsequent behavioral responses. Here we report on the identification of several antagonists of the odorant receptor co-receptor of the African malaria vectorAnopheles gambiae, AgamORco, in a small collection of natural volatile organic compounds (VOCs) using a relevant insect cell-based screening platform. Because some of the identified antagonists were previously shown to strongly repelAnophelesandCulexmosquitoes, here we examined the bioactivities of the identified antagonists againstAedes, the third major genus of the Culicidae family. The tested antagonists were found to inhibit the function ofAe. aegyptiORcoex vivoand repel Asian tiger,Ae. albopictus, adult mosquitoes. Specific antagonist binary mixtures elicited higher repellency than single antagonists. Binding competition assays suggested antagonist binding to distinct ORco sites as a likely cause for the enhanced repellence of the blends. These findings demonstrate that a simple screening assay may be used for the identification of allosteric modifiers of olfactory-driven behaviors capable of providing enhanced indoor and outdoor protection against multiple mosquito borne infectious diseases.
Integrated analysis of bulk multi omic and single-cell sequencing data confirms the molecular origin of hemodynamic changes in Covid-19 infection explaining coagulopathy and higher geriatric mortality
Authors: Johri Shreya , Jain Deepali , Gupta Ishaan
Published: April 2020

Besides severe respiratory distress, recent reports in Covid-19 patients have found a strong association between platelet counts and patient survival. Along with hemodynamic changes such as prolonged clotting time, high fibrin degradation products and D-dimers, increased levels of monocytes with disturbed morphology have also been identified. In this study, through an integrated analysis of bulk RNA-sequencing data from Covid-19 patients with data from single-cell sequencing studies on lung tissues, we found that most of the cell-types that contributed to the altered gene expression were of hematopoietic origin. We also found that differentially expressed genes in Covid-19 patients formed a significant pool of the expressing genes in phagocytic cells such as Monocytes and platelets. Interestingly, while we observed a general enrichment for Monocytes in Covid-19 patients, we found that the signal for FCGRA3+ Monocytes was depleted. Further, we found evidence that age-associated gene expression changes in Monocytes and platelets, associated with inflammation, mirror gene expression changes in Covid-19 patients suggesting that pro-inflammatory signalling during aging may worsen the infection in older patients. We identified more than 20 genes that change in the same direction between Covid-19 infection and aging cells that may act as potential therapeutic targets. Of particular interest were IL2RG, GNLY and GMZA expressed in platelets, which facilitates cytokine signalling in Monocytes through an interaction with platelets. To understand whether infection can directly manipulate the biology of Monocytes and platelets, we hypothesize that these non-ACE2 expressing cells may be infected by the virus through the phagocytic route. We observed that phagocytic cells such as Monocytes, T-cells, and platelets have a significantly higher expression of genes that are a part of the Covid-19 viral interactome. Hence these cell-types may have an active rather than a reactive role in viral pathogenesis to manifest clinical symptoms such as coagulopathy. Therefore, our results present molecular evidence for pursuing both anti-inflammatory and anticoagulation therapy for better patient management especially in older patients.
Biomarkers to distinguish bacterial from viral pediatric clinical pneumonia in a malaria endemic setting
Authors: Gillette Michael A. , Mani D. R. , Uschnig Christopher , Pellé Karell G. , Madrid Lola , Acácio Sozinho , Lanaspa Miguel , Alonso Pedro , Valim Clarissa , Carr Steven A. , Schaffner Stephen F. , MacInnis Bronwyn , Milner Danny A. , Bassat Quique , Wirth Dyann F.
Published: April 2020

BACKGROUND

Differentiating the etiology of acute febrile respiratory illness in children is a challenge in low-income, malaria-endemic settings because the main pathogens responsible (viruses, bacteria, and malaria parasites) overlap in clinical presentation and frequently occur together as mixed infections. The critical task is to rapidly identify bacterial pneumonia to enable appropriate antibiotic treatment, ideally at point of care. Current diagnostic tests are insufficient and there is a need for the discovery and development of new tools. Here we report the identification of a unique biomarker signature that can be identified in blood samples.

METHODS

Blood samples from 195 pediatric Mozambican patients with clinical pneumonia were analyzed with an aptamer-based high dynamic range assay to quantify ∼1200 proteins. For discovery of new biomarkers, we identified a training set of patient samples in which the underlying etiology of the pneumonia was established as bacterial, viral or malaria. Proteins whose abundances varied significantly between patients with verified etiologies (FDR<0.01) formed the basis for predictive diagnostic models that were created using machine learning techniques (Random Forest, Elastic Net). These models were validated on a dedicated test set of samples.

RESULTS

219 proteins had significantly different abundances between bacterial and viral infections, and 151 differed between bacterial infections and a mixed pool of viral and malaria infections. Predictive diagnostic models achieved >90% sensitivity and >80% specificity, regardless of whether one or two pathogen classes were present. Bacterial pneumonia was strongly associated with markers of neutrophil activity, in particular neutrophil degranulation. Degranulation markers included HP, LCN2, LTF, MPO, MMP8, PGLYRP1, RETN, SERPINA1, S100A9, and SLPI.

CONCLUSION

Blood protein signatures highly associated with neutrophil biology reliably differentiated bacterial pneumonia from other causes. With appropriate technology, these markers could provide the basis for a rapid diagnostic for field-based triage for antibiotic treatment of pediatric pneumonia.
Bitter Melon (Momordica Charantia) Supplementation Has no Effect on Hypercholesterolemia and Atherosclerosis in Mice
Authors: Mohammadmoradi Shayan , Howatt Deborah A. , Lu Hong S. , Daugherty Alan , Saha Sibu P.
Published: April 2020

Momordica Charantia,commonly known as bitter melon, has been reported to ameliorate diet-induced obesity and dyslipidemia. However, the effects ofM. Charantiaon atherosclerosis have not been determined. This study investigated the effects ofM. Charantiaon diet-induced atherosclerosis in LDL receptor deficient mice. Female mice (6-8 weeks old) were fed a saturated fat-enriched diet. In group 1, mice were fed this diet alone, while mice in groups 2 and 3 were fed the diet supplemented withM. Charantiaeither 0.1% or 1% by weight, for 12 weeks. No significant differences in body weights were observed among the 3 groups. There were also no significant differences in plasma cholesterol concentrations among the 3 groups. To determine the effects on atherosclerosis, lesion areas were measured on aortic intima by anen facetechnique. Neither dose ofM. Charantiasupplementation changed atherosclerosis development.
Early-life stress facilitates the development of Alzheimer’s disease pathology via angiopathy
Authors: Tanaka Tomoko , Hirai Shinobu , Hosokawa Masato , Saito Takashi , Sakuma Hiroshi , Saido Takaomi , Hasegawa Masato , Okado Haruo
Published: April 2020

Background

Alzheimer’s disease (AD), a progressive neurodegenerative disorder, is a serious social problem. Recently, several early-life factors have been associated with an increased risk of a clinical diagnosis of AD.

Methods

We investigated the involvement of early-life stress in AD pathogenesis using heterozygous the amyloid precursor protein (APP) mutant mice (AppNL-G-F/wt) and wild-type (Appwt/wt) mice. Maternal separation was used as an animal paradigm for early-life stress. Object location and fear conditioning tests were performed to measure cognitive functions, in addition to biochemical tests. Immunohistochemical analyses were performed after the behavioral tests.

Results

We found that maternal-separatedAppwt/wtmice showed narrowing of vessels and decreased pericyte coverage of capillaries in prefrontal cortex, while maternal-separatedAppNL-G-F/wtmice additionally showed impairment of cognitive function, and earlier formation of Aβ plaques and disruption of the blood–brain barrier. Severe activation of microglia was detected in the maternal-separatedAppNL-G-F/wtmice and maternal-separatedAppwt/wtmice. At the early stage, morphological changes and inflammatory responses were observed in the microglia of the maternal-separatedAppNL-G-F/wtmice and maternal-separatedAppwt/wtmice, as well as morphological changes in the microglia of the non-maternal-separatedAppNL-G-F/wtmice.

Conclusions

Microglia activation induced by maternal separation in combination with the APP mutation may impairs the vascular system, leading to AD progression. These findings therefore suggest that maternal separation causes early induction of AD pathology via angiopathy.
Evolutionary analysis of the LORELEI gene family in angiosperms reveals regulatory subfunctionalization
Authors: Noble Jennifer A. , Bielski Nicholas V. , Liu Ming-Che James , DeFalco Thomas A. , Stegmann Martin , Nelson Andrew D.L. , McNamara Kara , Sullivan Brooke , Dinh Khanhlinh K. , Khuu Nicholas , Hancock Sarah , Shiu Shin-Han , Zipfel Cyril , Cheung Alice Y. , Beilstein Mark A. , Palanivelu Ravishankar
Published: April 2020

A signaling complex comprising members of the LORELEI (LRE)-LIKE GPI-anchored protein (LLG) andCatharanthus roseusRECEPTOR-LIKE KINASE 1-LIKE (CrRLK1L) families perceive RAPID ALKALINIZATION FACTOR (RALF) peptides and regulate growth, reproduction, immunity, and stress responses in Arabidopsis. Genes encoding these proteins are members of multi-gene families in most angiosperms and could generate thousands of signaling complex variants. However, the link(s) between expansion of these gene families and the functional diversification of this critical signaling complex as well as the evolutionary factors underlying the maintenance of gene duplicates remain unknown. Here, we investigatedLLGgene family evolution, function, and expression in angiosperms. We found thatLLGsin monocots and eudicots are descendants of a duplication early in angiosperm evolution and that both ancient and recentLLGduplicates are retained. Complementation and expression analysis showed that expression divergence ofLLGs(regulatory subfunctionalization), rather than functional divergence, explains the retention of paralogs in Brassicales. All but one extant monocot and eudicot species examined maintained anLLGcopy with preferential expression in male reproductive tissues, with the other duplicate copies showed highest levels of expression in female or vegetative tissues. Interestingly, the singleLLGcopy in Amborella (sister to all other angiosperms) is expressed vastly higher in male compared to female reproductive or vegetative tissues. Reconstruction of expression evolution showed that the highest inferred expression levels for the single copy ancestral angiospermLLGwas in male reproductive tissues. We propose that expression divergence played an important role in maintenance ofLLGduplicates in angiosperms.

One Sentence Summary

Expression divergence played an important role in maintenance of two sub-groups ofLLGduplicates in angiosperms
Site-specific effects of neurosteroids on GABAAreceptor activation and desensitization
Authors: Sugasawa Yusuke , Cheng Wayland W. L. , Bracamontes John R. , Chen Zi-Wei , Wang Lei , Germann Allison L. , Pierce Spencer R. , Senneff Thomas C. , Krishnan Kathiresan , Reichert David E. , Covey Douglas F. , Akk Gustav , Evers Alex S.
Published: April 2020

This study examines how site-specific binding to the three identified neurosteroid binding sites in the α1β3GABAAreceptor (GABAAR) contributes to neurosteroid allosteric modulation. We found that the potentiating neurosteroid, allopregnanolone, but not its inhibitory 3β-epimer epi-allopregnanolone, binds to the canonical β3(+)–α1(-) intersubunit site that mediates receptor activation by neurosteroids. In contrast, both allopregnanolone and epi-allopregnanolone bind to intrasubunit sites in the β3subunit, promoting receptor desensitization and the α1subunit promoting ligand-specific effects. Two neurosteroid analogues with diazirine moieties replacing the 3-hydroxyl (KK148 and KK150) bind to all three sites, but do not potentiate GABAAR currents. KK148 is a desensitizing agent, whereas KK150 is devoid of allosteric activity. These compounds provide potential chemical scaffolds for site-specific and general neurosteroid antagonists. Collectively, these data show that differential occupancy and efficacy at three discrete neurosteroid binding sites determine whether a neurosteroid has potentiating, inhibitory, or competitive antagonist activity on GABAARs.
Optical TrkB activation in Parvalbumin interneurons regulates intrinsic states to orchestrate cortical plasticity
Authors: Winkel Frederike , Voigt Mathias B. , Didio Giuliano , Matéo Salomé , Jetsonen Elias , Pou Maria Llach , Steinzeig Anna , Ryazantseva Maria , Harkki Juliana , Englund Jonas , Khirug Stanislav , Rivera Claudio , Palva Satu , Taira Tomi , Lauri Sari E. , Umemori Juzoh , Castrén Eero
Published: April 2020

Activation state of Parvalbumin (PV) interneurons regulates neuronal plasticity, driving the closure of developmental critical periods and alternating between high and low plasticity states in response to experience in adulthood. We now show that PV plasticity states are regulated through the activation of TrkB neurotrophin receptors. Activation of an optically activatable TrkB (optoTrkB) specifically in PV interneurons switches adult cortical networks into a state of elevated plasticity within minutes by decreasing excitability of PV neurons. OptoTrkB activation induces changes in gene expression related to neuronal plasticity and excitability, and increases the phosphorylation of Kv3.1 channels. OptoTrkB activation shifted cortical networks towards a low PV configuration, promoting oscillatory synchrony and ocular dominance plasticity. Visual plasticity induced by fluoxetine was lost in mice lacking TrkB in PV neurons. Our data suggest a novel mechanism that dynamically regulates PV interneurons configuration state and orchestrates cortical networks during adulthood.

Graphical Abstract
Improved freshwater macroinvertebrate detection from eDNA through minimized non-target amplification
Authors: Leese Florian , Sander Mandy , Buchner Dominik , Elbrecht Vasco , Haase Peter , Zizka Vera M.A.
Published: April 2020

DNA metabarcoding of freshwater communities typically relies on PCR amplification of a fragment of the mitochondrial cytochrome c oxidase (COI) gene with degenerate primers. The advantage of COI is its taxonomic resolution and the availability of an extensive reference database. However, when universal primers are used on environmental DNA (eDNA) isolated from stream water, macroinvertebrate read and OTU numbers are typically “watered down”, i.e. diluted, compared to whole specimen ‘bulk samples’ due to greater co-amplification of abundant non-target taxa such as algae and bacteria. Because stream macroinvertebrate taxa are of prime importance for regulatory biomonitoring, more effective ways to capture their diversity via eDNA isolated from water are important. In this study, we aimed to improve macroinvertebrate assessment from eDNA by minimizing non-target amplification. Therefore, we generated data using universal primers BF2/BR2 throughout 15 months from a German Long-Term Ecological Research (LTER) site, the River Kinzig, to identify most abundant non-target taxa. Based on these data, we designed a new reverse primer (EPTDr2n) with 3’-specificity towards macrozoobenthic taxa and validated its specificityin silicotogether with universal forward primer fwhF2 using available data from GenBank and BOLD. We then performedin vitrotests using 20 eDNA samples taken in the Kinzig catchment. We found that the percentage of target reads was much higher for the new primer combination compared to two universal macrozoobenthic primer pairs, BF2/BR2 and fwhF2/fwhR2n (>99 % vs. 21.4 % and 41.25 %, respectively). Likewise, number of detected macroinvertebrate taxa was substantially higher (351 vs. 46 and 170, respectively) and exceeded the number of 257 taxa identified by expert taxonomists at nearby sites across two decades of sampling. While few taxa such as Turbellaria were not detected, we show that the optimized primer avoids the dilution problem and thus significantly improves macroinvertebrate detection for bioassessment and -monitoring.
TCGA Pan-Cancer genomic analysis of Alternative Lengthening of Telomeres (ALT) related genes
Authors: Armendáriz-Castillo Isaac , López-Cortés Andrés , García-Cárdenas Jennyfer , Guevara-Ramírez Patricia , Leone Paola E. , Pérez-Villa Andy , Yumiceba Verónica , Zambrano Ana K. , Guerrero Santiago , Paz-y-Miño César
Published: April 2020

Telomere maintenance mechanisms (TMM) are used by cancer cells to avoid apoptosis, 85-90% reactivate telomerase, while 10-15% use the alternative lengthening of telomeres (ALT). Due to anti-telomerase-based treatments, some tumors have the ability to switch from a telomerase-dependent mechanism to ALT, in fact, the co-existence between telomerase and the ALT pathway have been observed in a variety of cancer types. Despite different elements in the ALT pathway have been uncovered, the molecular mechanism and other factors are still poorly understood, which difficult the detection and treatment of ALT-positive cells, which are known to present poor prognosis. Therefore, with the aim to identify potential molecular markers to be used in the study of ALT, we combined simplisticin silicoapproaches in 411 telomere maintenance (TM) genes which have been previously validated or predicted to be involved in the ALT pathway. In consequence, we conducted a genomic analysis of these genes in 31 Pan-Cancer Atlas studies (n=9,282) from The Cancer Genome Atlas in the cBioPortal and found 325,936 genomic alterations, being mRNA high and low the top alterations with 65,.8% and 10.7% respectively. Moreover, we analyzed the highest frequency means of genomics alterations, identified and proposed 20 genes, which are highly mutated and up and down regulated in the cancer studies and could be used for future analysis in the study of ALT. Finally, we made a protein-protein interaction network and enrichment analysis to obtain an insight into the main pathways these genes are involved. We could observe their role in main processes related to the ALT mechanism like homologous recombination, homology directed repair (HDR), HDR through homologous recombination and telomere maintenance and organization.. Overall, due to the lack of understanding of the molecular mechanisms and detection of ALT-positive cancers, we identified and proposed more molecular targets that can be used for expression analysis and additionalex vivoassays to validate them as new potential therapeutic markers in the study of the ALT mechanism.
The preservation of ancient DNA in archaeological fish bone
Authors: Ferrari Giada , Cuevas Angelica , Gondek-Wyrozemska Agata T. , Ballantyne Rachel , Kersten Oliver , Pálsdóttir Albína H. , van der Jagt Inge , Hufthammer Anne-Karin , Ystgaard Ingrid , Wickler Stephen , Bigelow Gerald F. , Harland Jennifer , Nicholson Rebecca , Orton David , Clavel Benoît , Boessenkool Sanne , Barrett James H. , Star Bastiaan
Published: April 2020

The field of ancient DNA is taxonomically dominated by studies focusing on mammals. This taxonomic bias limits our understanding of endogenous DNA preservation for vertebrate taxa with different bone physiology, such as teleost fish. In contrast to most mammalian bone, teleost bone is typically brittle, porous, lightweight and is characterized by a lack of bone remodeling during growth. Using high-throughput shotgun sequencing, we here investigate the preservation of DNA in a range of different bone elements from over 200 archaeological Atlantic cod (Gadus morhua) specimens from 38 sites in northern Europe, dating up to 8000 years before present. We observe that the majority of archaeological sites (79%) yield endogenous DNA, with 40% of sites providing samples that contain high levels (> 20%). Library preparation success and levels of endogenous DNA depend mainly on excavation site and pre-extraction laboratory treatment. The use of pre-extraction treatments lowers the rate of library success, although — if successful — the fraction of endogenous DNA can be improved by several orders of magnitude. This trade-off between library preparation success and levels of endogenous DNA allows for alternative extraction strategies depending on the requirements of down-stream analyses and research questions. Finally, we find that — in contrast to mammalian bones — different fish bone elements yield similar levels of endogenous DNA. Our results highlight the overall suitability of archaeological fish bone as a source for ancient DNA and provide novel evidence for a possible role of bone remodeling in the preservation of endogenous DNA across different classes of vertebrates.
Conformational stability and dynamics in crystals recapitulate protein behaviour in solution
Authors: Sala BM , Le Marchand T , Pintacuda G , Camilloni C , Natalello A , Ricagno S
Published: April 2020

A growing body of evidences has established that in many cases proteins may preserve most of their function and flexibility in a crystalline environment, and several techniques are today capable to detect transiently-populated states of macromolecules in tightly packed lattices. Intriguingly, in the case of amyloidogenic precursors, the presence of these conformations (hidden to conventional crystallographic studies) can be correlated to the pathological fate of the native fold.

It remains unclear, however, to which extent these minor conformations reflect the protein behaviour that is more commonly studied in solution. Here, we address this question by investigating some biophysical properties of a prototypical amyloidogenic system, β2-microglobulin (β2m) in solution and in microcrystalline state.

By combining NMR chemical shifts with Molecular Dynamics (MD) simulations, we confirmed that conformational dynamics of β2m native state in the crystal lattice is in keeping with what observed in solution.

A comparative study of protein stability in solution andin crystallois then carried out, monitoring the change in protein secondary structure at increasing temperature by Fourier transform infrared (FTIR) spectroscopy. The increased structural order of the crystalline state contributes to provide better resolved spectral components compared to those collected in solution and crucially, the crystalline samples display thermal stabilities in good agreement with the trend observed in solution.

Overall, this work shows that protein stability and occurrence of pathological hidden states in crystals parallel their solution counterpart, confirming the interest of crystals as a platform for the biophysical characterisation of processes such as unfolding and aggregation.
Red light corrects neonicotinoid induced immunodeficiency and impaired respiration in poisoned bumblebees
Authors: Powner Michael B , Priestley Graham , Hogg Chris , Jeffery Glen
Published: April 2020

Neonicotinoid pesticides undermine mitochondrial function in insects including bumblebees, reducing ATP, mobility and leading to death. They also reduce bumblebee immunocompetency leaving them vulnerable to pathogen attack. This undermines key pollinators critical in the agricultural economy. However, 670nm light exposure improves mitochondrial function undermined by age or disease, increasing respiratory chain efficiency, improving ATP production, mobility and survival in bumblebees and fruit flies. Here we show that 670nm restores immunocompetence, improving hemocyte counts and hemolymph anti-microbial action. Additionally, we measure whole body respirationin vivoin individual bumblebees revealing that it is a functional metric of both neonicotinoid impact and light induced mitochondrial protection. Critically we show that only 1 min 670nm exposure is sufficient to correct respiratory deficits induced by pesticide and restore normal immune ability. Longer exposures are not more effective. Further, single 1 min exposure protects respiration and immunity for approximately 3-6 days. Hence, 670nm impact is not dose dependent but switch like.

These data provide a compelling rational for 670nm application to protect pollinators on which a major part of the agro economy is based and who are being challenged by aggressive pesticide application.
Fast and furious: host aggression modulates behaviour of brood parasites
Authors: Jelínek Václav , Šulc Michal , Štětková Gabriela , Honza Marcel
Published: April 2020

Avian brood parasites pose a serious threat for hosts, substantially reducing their fitness which selects for the evolution of host defences. A classic example of a host frontline defence is mobbing which frequently includes contact attacking of brood parasites. Here, we investigated how the nest defence of a very aggressive great reed warbler (Acrocephalus arundinaceus) host influences the speed of egg-laying and egg-removing behaviour of its brood parasite – the common cuckoo (Cuculus canorus). We video-recorded 168 brood parasitic events at 102 active host nests and found that cuckoos avoided host mobbing in only 62% of cases. If hosts spotted the cuckoo at their nests, they almost always attacked it (in 91 of 104 cases), however, such attacks only rarely and temporarily prevented cuckoos from parasitizing (11 additional cases). When attacked, cuckoos parasitized host nests significantly faster and left them immediately. However, when not attacked, cuckoos frequently stayed at or near the nest suggesting that host aggression, rather than the risk of being spotted, influences the speed of brood parasitism in this species. Further, we found that cuckoos performed egg-removing behaviour in all parasitic events without regard to host aggression. As a result, cuckoos removed at least one egg during all brood parasitism events except those when an egg slipped from their beaks and fell back into the nest (in 9 of 73 cases). This indicates that egg-removing behaviour is not costly for the common cuckoo and is an essential part of its parasitism strategy, widening understanding of this currently unexplained behaviour.
Identification of essential genes and fluconazole resistance genes inCandida glabrataby profiling ofHermestransposon insertions
Authors: Gale Andrew N. , Sakhawala Rima M. , Levitan Anton , Sharan Roded , Berman Judith , Timp Winston , Cunningham Kyle W.
Published: April 2020

Within the budding yeasts, the opportunistic pathogenCandida glabrataand other members of theNakaseomycesclade have developed virulence traits independently from the CTG clade that includesCandida albicans. To begin exploring the genetic basis ofC. glabratavirulence and its innate resistance to antifungals, we launched theHermestransposon from a plasmid and obtained more than 500,000 different semi-random insertions throughout the genome. Using machine learning, we identify up to 1278 protein-encoding genes (25% of total) that cannot tolerate transposon insertions and are thus essential forC. glabratafitness in vitro. Interestingly, genes involved in mRNA splicing were less likely to be essential inC. glabratathan their orthologs inS. cerevisiae, whereas the opposite is true for genes involved in kinetochore function and chromosome segregation. Insertions in several known genes (e.g.PDR1,CDR1,PDR16,PDR17,UPC2A,DAP1) caused hypersensitivity to the first-line antifungal fluconazole, and we identify 12 additional genes that also contribute to innate fluconazole resistance (KGD1,KGD2,YHR045W, etc). Insertions in 200 other genes conferred significant resistance to fluconazole, two-thirds of which function in mitochondria and likely down-regulate Pdr1 expression or function. These findings show the utility of transposon insertion profiling in genome-wide forward-genetic investigations of fungal pathogens.

IMPORTANCE

Pathogenic yeasts cause mucosal and systemic infections in millions of people each year. The innate resistance ofCandida glabratato fluconazole and its ability to acquire resistance to 2 other antifungals are contributing to its rise in incidence. Our understanding ofC. glabratabiology has been hampered by inefficient genetic and genomic tools. This study addresses those deficiencies by developing powerful transposon mutagenesis strategies for the first time in this pathogen. We identify nearly all essential genes ofC. glabratathat could be targeted for development of new antifungals. We generate large pools of random insertion mutants that can be easily monitoreden massewith deep sequencing, thus enabling identification of genes involved in any number of biological processes. We identify dozens of new genes that increase or decrease innate resistance of clinical isolate BG2 to fluconazole and provide resources for further exploration ofC. glabratagenetics and genomics.
Functional characterization of Neurofilament Light b splicing andmisbalance in zebrafish
Authors: Demy DL , Campanari ML , Munoz-Ruiz R , Durham HD , Gentil BJ , Kabashi E
Published: April 2020

Neurofilaments (NFs), a major cytoskeletal component of motor neurons, play a key role in their differentiation, establishment and maintenance of their morphology and mechanical strength. Thede novoassembly of these neuronal intermediate filaments requires the presence of the neurofilament light subunit, NEFL, which expression is reduced in motor neurons in Amyotrophic Lateral Sclerosis (ALS). This study used zebrafish as a model to characterize the NEFL homologueneflb, which encodes two different isoforms via splicing of the primary transcript (neflbE4andneflbE3).In vivoimaging showed thatneflbis crucial for proper neuronal development, and that disrupting the balance between its two isoforms specifically affects NF assembly and motor axon growth, with resulting motor deficits. This equilibrium is also disrupted upon partial depletion of TDP-43, a RNA binding protein that is mislocalized into cytoplasmic inclusions in ALS. The study supports interaction of NEFL expression and splicing with TDP-43 in a common pathway, both biologically and pathogenetically.
The draft genome sequence of herbaceous diploid bambooRaddia distichophylla
Authors: Li Wei , Shi Cong , Li Kui , Zhang Qun-jie , Tong Yan , Zhang Yun , Wang Jun , Clark Lynn , Gao Li-zhi
Published: April 2020

Bamboos are important non-timber forest plants widely distributed in the tropical and subtropical regions of Asia, Africa, America, and Pacific islands. They comprise the Bambusoideae in the grass family (Poaceae), including approximately 1,700 described species in 127 genera. In spite of the widespread uses of bamboo for food, construction and bioenergy, the gene repertoire of bamboo still remains largely unexplored.Raddia distichophylla(Schrad. ex Nees) Chase, belonging to the tribe Olyreae (Bambusoideae, Poaceae), is diploid herbaceous bamboo with only slightly lignified stems. In this study, we report a draft genome assembly of the approximately ∼589 Mb whole-genome sequence ofR. distichophyllawith a contig N50 length of 86.36 Kb. Repeated sequences account for ∼49.08% of the genome, of which LTR retrotransposons occupy ∼35.99% of whole genome. A total of 30,763 protein-coding genes were annotated in theR. distichophyllagenome with an average transcript size of 2,887 bp. Access to this herbaceous bamboo genome sequence will provide novel insights into biochemistry, molecular-assisted breeding programs and germplasm conservation for bamboo species world-wide.
A versatile system to introduce clusters of genomic double-strand breaks in large cell populations
Authors: Kolb Thorsten , Khalid Umar , Simović Milena , Ratnaparkhe Manasi , Wong John , Jauch Anna , Schmezer Peter , Rode Agata , Sebban Shulamit , Haag Daniel , Hergt Michaela , Devens Frauke , Buganim Yosef , Zapatka Marc , Lichter Peter , Ernst Aurélie
Published: April 2020

In vitroassays for clustered DNA lesions will facilitate the analysis of the mechanisms underlying complex genome rearrangements such as chromothripsis, including the recruitment of repair factors to sites of DNA double-strand breaks. We present a novel method generating localized DNA double-strand breaks using UV-irradiation with photomasks. The size of the damage foci and the spacing between lesions are fully adjustable, making the assay suitable for different cell types and targeted areas. We validated this set-up with genomically stable epithelial cells, normal fibroblasts, pluripotent stem cells and patient-derived primary cultures. Our method does not require a specialized device such as a laser, making it accessible to a broad range of users. Sensitization by BrdU incorporation is not required, which enables analyzing the DNA damage response in post-mitotic cells. Irradiated cells can be cultivated further, followed by time-lapse imaging or used for downstream biochemical analyses, thanks to the high-throughput of the system. Importantly, we showed genome rearrangements in the irradiated cells, providing a proof of principle for the induction of structural variants by localized DNA lesions.
Roles for the RNA polymerase III regulator MAFR-1 in regulating sperm quality inCaenorhabditis elegans
Authors: Hammerquist Amy M. , Curran Sean P.
Published: April 2020

The negative regulator of RNA polymerase (pol) IIImafr-1has been shown to affect RNA pol III transcript abundance, lipid biosynthesis and storage, progeny output, and lifespan. We deletedmafr-1from theCaenorhabditis elegansgenome and found that animals lackingmafr-1replicated many phenotypes from previous RNAi-based studies, but strikingly not the oocyte-related reproductive phenotypes. Utilizing a yeast two-hybrid assay, we discovered several novel interactors of MAFR-1 that are expressed in a sperm- and germline-enriched manner. In support of a role for MAFR-1 in the male germline, we foundmafr-1null males have smaller spermatids that are less capable in competition for fertilization; a phenotype that was dependent on RNA pol III activity. Restoration of MAFR-1 expression specifically in the germline rescued the spermatid-related phenotypes, suggesting a cell autonomous role for MAFR-1 in nematode male fertility. Based on the high degree of conservation of Maf1 activity across species, our study may inform similar roles for Maf1 and RNA pol III in mammalian male fertility.
Visualizing drug inhibitor binding interactions using microcrystal electron diffraction
Authors: Clabbers Max T.B. , Fisher S. Zoë , Coinçon Mathieu , Zou Xiaodong , Xu Hongyi
Published: April 2020

Visualizing drug inhibitor binding interactions at the atomic level is important for both structure-based drug design and fragment-based screening methods. Rapid and uniform soaking with potentially less lattice defects make small macromolecular crystals attractive targets for studying ligand biding using 3D microcrystal electron diffraction (MicroED). However, so far no drug inhibitor binding interactions could unambiguously be resolved by electron diffraction. Here, we use MicroED to study the binding of a sulfonamide inhibitor to human carbonic anhydrase isoform II (HCA II). We show that MicroED data can efficiently be collected in-house on a conventional TEM from thin hydrated microcrystals after a brief soaking with the clinical drug inhibitor acetazolamide (AZM). The data are of high enough quality to unequivocally fit and resolve the inhibitor bound to the active site of the protein. We anticipate MicroED can play an important role in future drug discovery experiments, complementing existing methods in structural biology such as x-ray and neutron diffraction.
Expression ofBrassica napusGLO1 is sufficient to breakdown artificial self-incompatibility inArabidopsis thaliana
Authors: Kenney Patrick , Sankaranarayanan Subramanian , Balogh Michael , Indriolo Emily
Published: April 2020

Members of the Brassicaceae family have the ability to regulate pollination events occurring on the stigma surface. InBrassicaspecies, self-pollination leads to an allele specific interaction between the pollen small cysteine-rich peptide ligand (SCR/SP11) and the stigmatic S-receptor kinase (SRK) that activates the E3 ubiquitin ligase ARC1 (Armadillo repeat-containing 1), resulting in proteasomal degradation of various compatibility factors including Glyoxalase I (GLO1) which is necessary for successful pollination. Suppression of GLO1 was sufficient to reduce compatibility, and overexpression of GLO1 in self-incompatibleBrassica napusstigmas resulted in partial breakdown of the self-incompatibility response. Here, we verified ifBnGLO1 could function as a compatibility factor in the artificial self-incompatibility system ofArabidopsis thalianaexpressingAlSCRb,AlSRKbandAlARC1 proteins fromA. lyrata. Overexpression ofBnGLO1 is sufficient to breakdown self-incompatibility response inA. thalianastigmas, suggesting that GLO1 functions as an inter-species compatibility factor. Therefore, GLO1 has an indisputable role as a compatibility factor in the stigma in regulating pollen attachment and pollen tube growth. Lastly, this study demonstrates the usefulness of an artificial self-incompatibility system inA. thalianafor interspecific self-incompatibility studies.
Serum levels and tumour expression of leptin and leptin receptor as promising clinical biomarkers of specific feline mammary carcinoma subtypes
Authors: Gameiro Andreia , Nascimento Catarina , Urbano Ana Catarina , Correia Jorge , Ferreira Fernando
Published: April 2020

Obesity is a risk factor for human breast cancer, being associated with increased serum levels of leptin. In cat, although obesity is a common nutritional disorder, the role of leptin and its receptor in mammary carcinoma is unknown. In this study, serum levels of leptin and leptin receptor (ObR) were evaluated in 58 cats with mammary carcinoma and compared with healthy controls by ELISA, as tumour expression by immunohistochemistry. Results showed that the free leptin index is decreased in cats with mammary carcinoma (p=0.0006), particularly for those with luminal B and HER2-positive disease that showed significantly lower serum leptin levels (p<0.0001 and p<0.005, respectively). Serum leptin levels above 4.17 pg/mL were associated with ulcerating tumours (p=0.0005) and shorter DFS (p=0.0217). Elevated serum ObR levels were found in all cats with mammary carcinoma (p<0.0001), with levels above 16.89 ng/mL being associated with smaller tumours (p=0.0118), ER-negative status in HER2-positive tumours (p=0.0291) and increased serum levels of CTLA-4 (p=0.0056), TNF-α (p=0.0025), PD-1 (p=0.0023) and PD-L1 (p=0.0002). In tumour samples, leptin was overexpressed in luminal B and triple-negative carcinomas (p=0.0046), whereas ObR was found overexpressed in luminal B samples (p=0.0425). Altogether, our results reinforce the importance of feline mammary carcinoma in comparative oncology.