Search

Abstract Innate antiviral factors are essential for effective defense against viral pathogens. However, the identity of major restriction mechanisms remains elusive. Current approaches to discover antiviral factors usually focus on the initial steps of viral replication and are limited to a single round of infection. Here, we engineered libraries of >1500 replication-competent HIV-1 constructs each expressing a single gRNAs to target >500 cellular genes for virus-driven discovery of antiviral factors. Passaging in CD4+ T cells robustly enriched HIV-1 encoding sgRNAs against GRN, CIITA, EHMT2, CEACAM3, CC2D1B and RHOA by >50-fold. Using an HIV-1 library lacking the accessory nef gene, we identified IFI16 as a Nef target. Functional analyses in cell lines and primary CD4+ T cells support that the HIV-driven CRISPR screen identified restriction factors targeting virus entry, transcription, release and infectivity. Our HIV-guided CRISPR technique enables sensitive discovery of physiologically relevant cellular defense factors throughout the entire viral replication cycle.

Abstract Under-reporting of COVID-19 and the limited information about circulating SARS-CoV-2 variants remain major challenges for many African countries. We analyzed SARS-CoV-2 infection dynamics in Addis Ababa and Jimma, Ethiopia, focusing on reinfection, immunity, and vaccination effects. We conducted an antibody serology study spanning August 2020 to July 2022 with five rounds of data collection across a population of 4723, sequenced PCR-test positive samples, used available test positivity rates, and constructed two mathematical models integrating this data. A multivariant model explores variant dynamics identifying wildtype, alpha, delta, and omicron BA.4/5 as key variants in the study population, and cross-immunity between variants, revealing risk reductions between 24% and 69%. An antibody-level model predicts slow decay leading to sustained high antibody levels. Retrospectively, increased early vaccination might have substantially reduced infections during the delta and omicron waves in the considered group of individuals, though further vaccination now seems less impactful.

Abstract The SARS-CoV-2 pandemic has highlighted the need to better define in-hospital transmissions, a need that extends to all other common infectious diseases encountered in clinical settings. To evaluate how whole viral genome sequencing can contribute to deciphering nosocomial SARS-CoV-2 transmission 926 SARS-CoV-2 viral genomes from 622 staff members and patients were collected between February 2020 and January 2021 at a university hospital in Munich, Germany, and analysed along with the place of work, duration of hospital stay, and ward transfers. Bioinformatically defined transmission clusters inferred from viral genome sequencing were compared to those inferred from interview-based contact tracing. An additional dataset collected at the same time at another university hospital in the same city was used to account for multiple independent introductions. Clustering analysis of 619 viral genomes generated 19 clusters ranging from 3 to 31 individuals. Sequencing-based transmission clusters showed little overlap with those based on contact tracing data. The viral genomes were significantly more closely related to each other than comparable genomes collected simultaneously at other hospitals in the same city (n = 829), suggesting nosocomial transmission. Longitudinal sampling from individual patients suggested possible cross-infection events during the hospital stay in 19.2% of individuals (14 of 73 individuals). Clustering analysis of SARS-CoV-2 whole genome sequences can reveal cryptic transmission events missed by classical, interview-based contact tracing, helping to decipher in-hospital transmissions. These results, in line with other studies, advocate for viral genome sequencing as a pathogen transmission surveillance tool in hospitals.

Abstract Both carbon dioxide uptake and albedo of the land surface affect global climate. However, climate change mitigation by increasing carbon uptake can cause a warming trade-off by decreasing albedo, with most research focusing on afforestation and its interaction with snow. Here, we present carbon uptake and albedo observations from 176 globally distributed flux stations. We demonstrate a gradual decline in maximum achievable annual albedo as carbon uptake increases, even within subgroups of non-forest and snow-free ecosystems. Based on a paired-site permutation approach, we quantify the likely impact of land use on carbon uptake and albedo. Shifting to the maximum attainable carbon uptake at each site would likely cause moderate net global warming for the first approximately 20 years, followed by a strong cooling effect. A balanced policy co-optimizing carbon uptake and albedo is possible that avoids warming on any timescale, but results in a weaker long-term cooling effect.

Abstract Despite previous reports on European growing seasons lengthening due to global warming, evidence shows that this trend has been reversing in the past decade due to increased transpiration needs. To asses this, we used an innovative method along with space-based observations to determine the timing of greening and dormancy and then to determine existing trends of them and causes. Early greening still occurs, albeit at slower rates than before. However, a recent (2011–2020) shift in the timing of dormancy has caused the season length to decrease back to 1980s levels. This shortening of season length is attributed primarily to higher atmospheric water demand in summer that suppresses transpiration even for soil moisture levels as of previous years. Transpiration suppression implies that vegetation is unable to meet the high transpiration needs. Our results have implications for future management of European ecosystems (e.g., net carbon balance and water and energy exchange with atmosphere) in a warmer world.

Abstract Coccolithophores are globally abundant, calcifying microalgae that have profound effects on marine biogeochemical cycles, the climate, and life in the oceans. They are characterized by a cell wall of CaCO3 scales called coccoliths, which may contribute to their ecological success. The intricate morphologies of coccoliths are of interest for biomimetic materials synthesis. Despite the global impact of coccolithophore calcification, we know little about the molecular machinery underpinning coccolithophore biology. Working on the model Emiliania huxleyi, a globally distributed bloom-former, we deploy a range of proteomic strategies to identify coccolithogenesis-related proteins. These analyses are supported by a new genome, with gene models derived from long-read transcriptome sequencing, which revealed many novel proteins specific to the calcifying haptophytes. Our experiments provide insights into proteins involved in various aspects of coccolithogenesis. Our improved genome, complemented with transcriptomic and proteomic data, constitutes a new resource for investigating fundamental aspects of coccolithophore biology.

The required energy savings can be achieved in all automotive domains through weight savings and the merging of manufacturing processes in production. This fact is taken into account through functional integration in lightweight materials and manufacturing in a process close to large-scale production. In previous work, separate steps of a process chain for manufacturing a center console cover utilizing a sensoric hybrid laminate have been developed and evaluated. This includes the process steps of joining, forming and inline polarization as well as connecting to an embedded system. This work continues the research process by evaluating impact localization methods to use the center console as a haptic input device. For this purpose, different deep learning methods are derived from the state of the art and analyzed for their applicability in two consecutive studies. The results show that MLPs, LSTMs, GRUs and CNNs are suitable to localize impacts on the novel laminate with high localization rates of up to 99%, and thus the usability of the developed laminate as a haptic input device has been proven.

IntroductionTo explore whether the reported lower pathogenicity in infected individuals of variant of concern (VoC) Omicron and its current subvariants compared to VoC Delta may be related to fundamental differences in the initial virus-tissue interaction, we assessed their ability to penetrate, replicate and cause damage in a human 3D respiratory model.MethodsFor this, we used TEER measurements, real-time PCR, LDH, cytokine and complex confocal imaging analyses.Results and discussionWe observed that Delta readily penetrated deep into the respiratory epithelium and this was associated with major tissue destruction, high LDH activity, high viral loads and pronounced innate immune activation as observed by intrinsic C3 activation and IL-6 release at infection sites. In contrast, Omicron subvariants BA.5, BQ.1.1 and BF7 remained superficially in the mucosal layer resulting merely in outward-directed destruction of cells, maintenance of epithelial integrity, minimal LDH activity and low basolateral release of virus at infection sites, as well as significantly smaller areas of complement activation and lower IL-6 secretion. Interestingly, also within Omicron subvariants differences were observed with newer Omicron subvariants BQ.1.1 and BF.7 illustrating significantly reduced viral loads, IL-6 release and LDH activity compared to BA.5. Our data indicate that earliest interaction events after SARS-CoV-2 transmission may have a role in shaping disease severity.

ABSTRACT Currently, SARS-CoV-2 Omicron BA.5 subvariants BF.7 and BQ.1.1 are rapidly emerging worldwide. To evaluate the SARS-CoV-2-neutralizing capacity of sera and saliva from triple vaccinated individuals, either boosted with an adapted bivalent COVID-19 vaccine or recovered from BA.4/BA.5 infection, we analyzed the sensitivity of replication-competent SARS-CoV-2 Omicron subvariants BA.4/5, BQ.1.1 and BF.7 to neutralization. Analysis of SARS-CoV-2-specific IgGs and IgAs showed increased serum IgG titers in the vaccinated group, while the serum and salivary IgA levels were comparable. Similar and efficient serum neutralization against the ancestral strain of SARS-CoV-2 and Omicron BA.4/BA.5 was detected in both cohorts, but critically reduced for BQ.1.1 and BF.7. In contrast, salivary neutralization against BA.4/BA.5 was increased in the convalescent compared to the vaccinated group, while salivary neutralizing capacity against BQ.1.1 and BF.7 was comparable in these groups. Further, personalized protective effects studied in a human 3D respiratory model revealed the importance of salivary protection against different Omicron subvariants. IMPORTANCE In BA.4/BA.5-convalescent versus vaccinated groups, salivary neutralization capacity increased against SARS-CoV-2 Omicron BA.4/BA.5. In contrast, it neutralized novel Omicron subvariants BQ.1.1 and BF.7 similarly. Salivary protection against various Omicron subvariants was even more evident when tested in a personalized approach using highly differentiated respiratory human 3D models.

European grasslands increase their water-use efficiency in summer through increased gross primary production and regulated transpiration, according to an analysis of three indices derived from remote sensing over the period 1995-2018

Cloud computing facilities can provide crucial computing support for processing the time series of satellite data and exploiting their spatio-temporal information content. However, dedicated efforts are still required to develop workflows, executable on cloud-based platforms, for ingesting the satellite data, performing the targeted processes, and generating the desired products. In this study, an operational workflow is proposed, based on monthly Evaporative Stress Index (ESI) anomaly, and implemented in cloud-based online Virtual Earth Laboratory (VLab) platform, as a demonstration, to monitor European agricultural water stress. To this end, daily time-series of actual and reference evapotranspiration (ETa and ET0), from the Spinning Enhanced Visible and Infrared Imager (SEVIRI) sensor, were used to execute the proposed workflow successfully on VLab. The execution of the workflow resulted in obtaining one decade (2011–2020) of European monthly agricultural water stress maps at 0.04˚ spatial resolution and corresponding stress reports for each country. To support open science, all the workflow outputs are stored in GeoServer, documented in GeoNetwork, and made available through MapStore. This enables creating a dashboard for better visualization of the results for end-users. The results from this study demonstrate the capability of VLab platform for water stress detection from time series of SEVIRI-ET data.

SARS-CoV-2 mutations associated with the escape from antibody-mediated neutralization have been widely reported. Here, in a patient with defective antibody responses, the authors find a potential association between SARS-CoV-2 mutations and CD8 T alterations to implicate possible contributions of CD8 T cells in evasion of SARS-CoV-2 from host immunity.

The molecular consequences of the metabolic stress caused by milk production of dairy cows in the early embryo are largely unknown. The objective was to determine the impact of dam metabolic status or in vitro culture during embryonic genome activation (EGA) on the transcriptomic profiles of bovine 16-cell stage embryos. Two days after synchronized oestrus, in vitro produced 2- to 4-cell stage embryos were endoscopically transferred in pools of 50 into the oviduct ipsilateral to the corpus luteum of lactating (LACT, n = 3) or nonlactating (i.e. dried off immediately at calving; DRY, n = 3) dairy cows. On Day 4, the oviducts were flushed to recover the embryos. Pools of five Day-2 embryos (n = 5) and Day-4 16-cell stage embryos obtained in vitro (n = 3) or from LACT or DRY cows were subjected to RNAseq. Temporally differentially expressed genes (DEG; FDR

Deforestation has a wide range of effects on hydrological and geochemical processes. Dissolved organic carbon (DOC) dynamics, a sensitive environmental change indicator, is expected to be affected by deforestation, with changes in atmospheric sulfur (S) deposition compounding this. However, how precisely anthropogenic disturbance (deforestation) under a declining atmospheric S input scenario affects the underlying spatiotemporal dynamics and relationships of river DOC and sulfate with hydro-climatological variables e.g., stream water temperature, runoff, pH, total dissolved iron (Fetot), and calcium (Ca2+) remains unclear. We, therefore, examined this issue within the TERENO Wüstebach catchment (Eifel, Germany), where partial deforestation had taken place in 2013. Wavelet transform coherence (WTC) analysis was applied based on a 10-year time series (2010–2020) from three sampling stations, whose (sub) catchment areas have different proportions of deforested area (W10: 31%, W14: 25%, W17: 3%). We found that water temperature and DOC, sulfate, and Fetot concentrations showed distinct seasonal patterns, with DOC averaging concentrations ranging from 2.23 (W17) to 4.56 (W10) mg L–1 and sulfate concentration ranging from 8.04 (W10) to 10.58 (W17) mg L–1. After clear-cut, DOC significantly increased by 59, 58% in the mainstream (W10, W14), but only 26% in the reference stream. WTC results indicated that DOC was negatively correlated with runoff and sulfate, but positively correlated with temperature, Ca2+, and Fetot. The negative correlation between DOC with runoff and sulfate was apparent over the whole examined 10-year period in W17 but did end in W10 and W14 after the deforestation. Sulfate (SO4) was highly correlated with stream water temperature, runoff, and Fetot in W10 and W14 and with a longer lag time than W17. Additionally, pH was stronger correlated (higher R2) with sulfate and DOC in W17 than in W10 and W14. In conclusion, WTC analysis indicates that within this low mountainous forest catchment deforestation levels over 25% (W10 and W14) affected the coupling of S and C cycling substantially more strongly than “natural” environmental changes as observed in W17.

Given the use of high-strength steels to achieve lightweight construction goals, conventional shear-cutting processes are reaching their limits. Therefore, so-called high-speed impact cutting (HSIC) is used to achieve the required cut surface qualities. A new machine concept consisting of linear motors and an impact mass is presented to investigate HSIC. It allows all relevant parameters to be flexibly adjusted and measured. The design and construction of the test bench, as well as the mechanism for coupling the impact mass, are described. To validate the theoretically determined process speeds, the cutting process was recorded with high-speed cameras, and HSIC with a mild deep-drawing steel sheet was performed. It was discovered that very good cutting edges could be produced, which showed a significantly lower hardening depth than slowly cut reference samples. In addition, HSIC was numerically modelled in LS-DYNA, and the calculated cutting edges were compared with the real ones. With the help of adaptive meshing, a very good agreement for the cutting edges could be achieved. The results show the great potential of using a linear motor in HSIC.

Shearing high-strength steels often leads to a subpar cut quality and excessive stress on the tool components. To enhance the quality of the cut surface, intricate techniques like fine blanking are commonly employed. However, for applications with lower quality requirements, precision shear cutting offers an alternative solution. This research paper introduces a novel approach to directly superimpose radial stress on a workpiece during the precision shear cutting process and showcases for the first time how the application of direct stress superimposition can impact the cut surface by concurrently modifying the shear cutting edge and punch surface. A statistical experimental design is employed to investigate the interrelationships between the parameters and their effects. The results indicate that the overall cut quality, including cylindricity, clean-cut angle, rollover height, and tool stress, defined by punch force and retraction force, is influenced by the superimposed stress. Regarding the clean-cut zone, the statistical significance of direct radially superimposed stress was not observed, except when interacting with sheet thickness and clearance. Additionally, the sheet thickness and cutting gap emerged as significant parameters affecting the overall quality of the cut surface.

In this study, a range of miniaturized Ag/AgCl reference electrodes with various layouts were successfully fabricated on wafer-level silicon-based substrates with metallic intermediate layers by precisely controlling the electrochemical deposition of Ag, followed by electrochemical chlorination of the deposited Ag layer. The structure, as well as the chemical composition of the electrode, were characterized with SEM & EDS. The results showed that the chlorination is very sensitive to the applied electric field and background solution. Potentiostatic chlorination, in combination with an adjusted mushroom-shaped Ag sealing deposition, enabled the formation of electrochemical usable Ag/AgCl layers. The stability of the electrodes was tested using open circuit potential (OCP) measurement. The results showed that the reference electrodes stayed stable for 300 s under 3 M KCl solution. The first stage study showed that the stability of the Ag/AgCl reference electrode in a chip highly depends on chip size design, chlorination conditions, and a further protection layer.

Species of Alternaria (phylum Ascomycota, family Pleosporaceae) are known as serious plant pathogens, causing major losses on a wide range of crops. Alternaria atra (previously known as Ulocladium atrum) can grow as a saprophyte on many hosts and causes Ulocladium blight on potato. It has been reported that it can also be used as a biocontrol agent against Botrytis cinerea. Here, we present a scaffold-level reference genome assembly for A. atra. The assembly contains 43 scaffolds with a total length of 39.62 Mbp, with scaffold N50 of 3,893,166 bp, L50 of 4, and the longest 10 scaffolds containing 89.9% of the assembled data. RNA-sequencing-guided gene prediction using BRAKER resulted in 12,173 protein-coding genes with their functional annotation. This first high-quality reference genome assembly and annotation for A. atra can be used as a resource for studying evolution in the highly complicated Alternaria genus and might help in understanding the mechanisms defining its role as pathogen or biocontrol agent.[Graphic: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

Moderate heat stress primes plants to acquire tolerance to subsequent, more severe heat stress. Here the authors show that the HSFA3 transcription factor forms a heteromeric complex with HSFA2 to sustain activated transcription of genes required for acquired thermotolerance by promoting H3K4 hyper-methylation.

Immunocompromised individuals are at higher risk of developing protracted and severe COVID-19, and understanding individual disease courses and SARS-CoV-2 immune responses in these individuals is of the utmost importance. For more than two years, we followed an immunocompromised individual with a protracted SARS-CoV-2 infection that was eventually cleared in the absence of a humoral neutralizing SARS-CoV-2 antibody response. By conducting an in-depth examination of this individual’s immune response and comparing it to a large cohort of convalescents who spontaneously cleared a SARS-CoV-2 infection, we shed light on the interplay between B- and T-cell immunity and how they interact in clearing SARS-CoV-2 infection.

Protein-protein interactions are thought to channel substrates between consecutive enzymes during glycolysis. Here the authors show that Arabidopsis phosphoglycerate mutase and enolase can form a substrate-channelling metabolon and also play a moonlighting role in promoting colocalization of chloroplasts and mitochondria.

Mutations in CLRN1 cause Usher syndrome (USH) type III (USH3A), a disease characterized by progressive hearing impairment, retinitis pigmentosa, and vestibular dysfunction. Due to the lack of appropriate disease models, no efficient therapy for retinitis pigmentosa in USH patients exists so far. In addition, given the yet undefined functional role and expression of the different CLRN1 splice isoforms in the retina, non-causative therapies such as gene supplementation are unsuitable at this stage. In this study, we focused on the recently identified deep intronic c.254-649T>G CLRN1 splicing mutation and aimed to establish two causative treatment approaches: CRISPR-Cas9-mediated excision of the mutated intronic region and antisense oligonucleotide (AON)-mediated correction of mRNA splicing. The therapeutic potential of these approaches was validated in different cell types transiently or stably expressing CLRN1 minigenes. Both approaches led to substantial correction of the splice defect. Surprisingly, however, no synergistic effect was detected when combining both methods. Finally, the injection of naked AONs into mice expressing the mutant CLRN1 minigene in the retina also led to a significant splice rescue. We propose that both AONs and CRISPR-Cas9 are suitable strategies to initiate advanced preclinical studies for treatment of USH3A patients.

Abstract Riparian zones as the transition zone between terrestrial and aquatic ecosystems play an important role in C and N cycling and greenhouse gas (GHG) emissions. As such, they may help to mitigate climate change but could also accelerate it, depending on the particular processes affected by changes in the hydrologic regime. Hydrological observations indicated frequent shallow groundwater in the riparian zone, especially near the stream and during the wet winter and spring seasons with consequently frequent occurrence of soil water saturation. The redox potential was mainly governed by the soil water regime: under water saturation conditions, the redox potential of the soil decreased and returned to the oxic state after soil drainage. We found that soil temperature and soil water content were the main drivers of the variations in CO2 fluxes, with highest CO2 emission during summer and the lowest emissions in the winter period (162.2–5.4 mg CO2–C m−2 h−1). The annual average daily N2O emission rate was low (2.3 μg N2O‐N m−2 h−1), with the highest average daily N2O emission in March as a result of low temperature and partial soil saturation after heavy precipitation events (37.5 μg N2O‐N m−2 h−1). Our study showed that continuous measurement of redox potential, soil temperature, and soil water content can improve the understanding of GHG emissions in riparian zones.

Abstract The C9orf72 repeat expansion is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and/or frontotemporal dementia (FTD). Non‐canonical translation of the expanded repeat results in abundant poly‐GA inclusion pathology throughout the CNS. (GA)149‐CFP expression in mice triggers motor deficits and neuroinflammation. Since poly‐GA is transmitted between cells, we investigated the therapeutic potential of anti‐GA antibodies by vaccinating (GA)149‐CFP mice. To overcome poor immunogenicity, we compared the antibody response of multivalent ovalbumin‐(GA)10 conjugates and pre‐aggregated carrier‐free (GA)15. Only ovalbumin‐(GA)10 immunization induced a strong anti‐GA response. The resulting antisera detected poly‐GA aggregates in cell culture and patient tissue. Ovalbumin‐(GA)10 immunization largely rescued the motor function in (GA)149‐CFP transgenic mice and reduced poly‐GA inclusions. Transcriptome analysis showed less neuroinflammation in ovalbumin‐(GA)10‐immunized poly‐GA mice, which was corroborated by semiquantitative and morphological analysis of microglia/macrophages. Moreover, cytoplasmic TDP‐43 mislocalization and levels of the neurofilament light chain in the CSF were reduced, suggesting neuroaxonal damage is reduced. Our data suggest that immunotherapy may be a viable primary prevention strategy for ALS/FTD in C9orf72 mutation carriers.

Статья прослеживает динамику использования и роль иностранных языков в русской литературе в XVIII–XX веках. Обсуждаются факторы, регулирующие отношение к иностранным языкам, в диапазоне от индивидуальных до культурно-исторических. В центре внимания оказываются изменения вектора культурной ориентации как совокупность разнородных явлений.

Abstract The majority of cellular processes are carried out by protein complexes. Various size fractionation methods have previously been combined with mass spectrometry to identify protein complexes. However, most of these approaches lack the quantitative information which is required to understand how changes of protein complex abundance and composition affect metabolic fluxes. In this paper we present a proof of concept approach to quantitatively study the complexome in the model plant Arabidopsis thaliana at the end of the day (ED) and the end of the night (EN). We show that size-fractionation of native protein complexes by Clear-Native-PAGE (CN-PAGE), coupled with mass spectrometry can be used to establish abundance profiles along the molecular weight gradient. Furthermore, by deconvoluting complex protein abundance profiles, we were able to drastically improve the clustering of protein profiles. To identify putative interaction partners, and ultimately protein complexes, our approach calculates the Euclidian distance between protein profile pairs. Acceptable threshold values are based on a cut-off that is optimized by a receiver-operator characteristic (ROC) curve analysis. Our approach shows low technical variation and can easily be adapted to study in the complexome in any biological system.

Abstract Background Lactation and associated metabolic stresses during the post-partum period have been shown to impair fertility in dairy cows. The oviduct plays key roles in embryo development and the establishment of pregnancy in cattle. The aim of this study was to investigate the effects of lactation and location relative to the corpus luteum (CL) on the transcriptome of the bovine oviduct epithelium. Results An original animal model was used. At 60 days post-partum, Holstein lactating (n = 4) and non-lactating (i.e. never milked after calving; n = 5) cows, as well as control nulliparous heifers (n = 5), were slaughtered on Day 3 following induced estrus, and epithelial samples from the oviductal ampulla and isthmus ipsilateral and contralateral to the corpus luteum (CL) were recovered for RNA sequencing. In the oviduct ipsilateral to the CL, differentially expressed genes (DEGs) were identified between heifers compared with both postpartum cow groups. However, only 15 DEGs were identified between post-partum lactating and non-lactating cows in the ipsilateral isthmus and none were identified in the ipsilateral ampulla. In contrast, 192 and 2583 DEGs were identified between ipsilateral and contralateral ampulla and isthmus, respectively. In both regions, more DEGs were identified between ipsilateral and contralateral oviducts in non-lactating cows and heifers than in lactating cows. Functional annotation of the DEGs associated with comparisons between metabolic groups highlighted a number of over-represented biological functions and cell pathways including immune response and cholesterol/steroid biosynthesis. Conclusions Gene expression in the oviduct epithelium, particularly in the isthmus, was more affected by the location relative to the CL than by lactation at Day 3 post-estrus. Furthermore, the effect of the proximity to the CL was modulated by the metabolic status of the cow.

Identification of fusion genes in clinical routine is mostly based on cytogenetics and targeted molecular genetics, such as metaphase karyotyping, fluorescence in situ hybridization and reverse-transcriptase polymerase chain reaction. However, sequencing technologies are becoming more important in clinical routine as processing time and costs per sample decrease. To evaluate the performance of fusion gene detection by RNAsequencing compared to standard diagnostic techniques, we analyzed 806 RNA-sequencing samples from patients with acute myeloid leukemia using two state-of-the-art software tools, namely Arriba and FusionCatcher. RNA-sequencing detected 90% of fusion events that were reported by routine with high evidence, while samples in which RNA-sequencing failed to detect fusion genes had overall lower and inhomogeneous sequence coverage. Based on properties of known and unknown fusion events, we developed a workflow with integrated filtering strategies for the identification of robust fusion gene candidates by RNA-sequencing. Thereby, we detected known recurrent fusion events in 26 cases that were not reported by routine and found discrepancies in evidence for known fusion events between routine and RNA-sequencing in three cases. Moreover, we identified 157 fusion genes as novel robust candidates and comparison to entries from ChimerDB or Mitelman Database showed novel recurrence of fusion genes in 14 cases. Finally, we detected the novel recurrent fusion gene NRIP1- MIR99AHG resulting from inv(21)(q11.2;q21.1) in nine patients (1.1%) and LTN1-MX1 resulting from inv(21)(q21.3;q22.3) in two patients (0.25%). We demonstrated that NRIP1-MIR99AHG results in overexpression of the 3' region of MIR99AHG and the disruption of the tricistronic miRNA cluster miR-99a/let-7c/miR-125b-2. Interestingly, upregulation of MIR99AHG and deregulation of the miRNA cluster, residing in the MIR99AHG locus, are known mechanisms of leukemogenesis in acute megakaryoblastic leukemia. Our findings demonstrate that RNA-sequencing has a strong potential to improve the systematic detection of fusion genes in clinical applications and provides a valuable tool for fusion discovery.

Abstract. Introduction:. Civilian ballistic injuries are commonly associated with fracture, vascular injury, and soft tissue trauma. Posttraumatic pseudoaneurysms represent an extremely rare subset of vascular injuries following ballistic fractures. Case:. We present the rare case of a posttraumatic pseudoaneurysm that occurred after retrograde femoral nailing of a ballistic distal femur fracture. The patient presented in clinic postoperatively with a pulsatile popliteal mass. Distal pulses were intact but subsequent ultrasound and angiography revealed a pseudoaneurysm of the distal superficial femoral artery. The pseudoaneurysm was subsequently treated with a covered stent and the patient's recovery was uncomplicated. Conclusion:. Due to the potential life and limb-threating complications from pseudoaneurysm rupture, this case report emphasizes the early recognition and expeditious management of vascular complications following ballistic fractures in the civilian population.

Raman spectroscopy is an analytical technology for the simultaneous measurement of important process parameters, such as concentrations of nutrients, metabolites, and product titer in mammalian cell culture. The majority of published Raman studies have concentrated on using the technique for the monitoring and control of bioreactors at pilot and manufacturing scales. This research presents a novel approach to generating Raman models using a high-throughput 250 mL mini bioreactor system with the following two integrated analysis modules: a prototype flow cell enabling on-line Raman measurements and a bioanalyzer to generate reference measurements without a significant time-shift, compared to the corresponding Raman measurement. Therefore, spectral variations could directly be correlated with the actual analyte concentrations to build reliable models. Using a design of experiments (DoE) approach and additional spiked samples, the optimized workflow resulted in robust Raman models for glucose, lactate, glutamine, glutamate and titer in Chinese hamster ovary (CHO) cell cultures producing monoclonal antibodies (mAb). The setup presented in this paper enables the generation of reliable Raman models that can be deployed to predict analyte concentrations, thereby facilitating real-time monitoring and control of biologics manufacturing.

Microbial-Induced Calcite Precipitation (MICP) stimulates soil microbiota to induce a cementation of the soil matrix. Urea, calcium and simple carbon nutrients are supplied to produce carbonates via urea hydrolysis and induce the precipitation of the mineral calcite. Calcium chloride (CaCl2) is typically used as a source for calcium, but basic silicate rocks and other materials have been investigated as alternatives. Weathering of calcium-rich silicate rocks (e.g., basalt and dolerite) releases calcium, magnesium and iron; this process is associated with sequestration of atmospheric CO2 and formation of pedogenic carbonates. We investigated atmospheric carbon fluxes of a MICP treated sandy soil using CaCl2 and dolerite fines applied on the soil surface as sources for calcium. Soil-atmosphere carbon fluxes were monitored over 2 months and determined with an infrared gas analyser connected to a soil chamber. Soil inorganic carbon content and isotopic composition were determined with isotope-ratio mass spectrometry. In addition, soil-atmosphere CO2 fluxes during chemical weathering of dolerite fines were investigated in incubation experiments with gas chromatography. Larger CO2 emissions resulted from the application of dolerite fines (116 g CO2-C m–2) compared to CaCl2 (79 g CO2-C m–2) but larger inorganic carbon precipitation also occurred (172.8 and 76.9 g C m–2, respectively). Normalising to the emitted carbon to precipitated carbon, the environmental carbon cost was reduced with dolerite fines (0.67) compared to the traditional MICP treatment (1.01). The carbon isotopic signature indicated pedogenic carbonates (δ13Cav = −8.2 ± 5.0‰) formed when dolerite was applied and carbon originating from urea (δ13Cav = −46.4 ± 1.0‰) precipitated when CaCl2 was used. Dolerite fines had a large but short-lived (

Introduction: Hip fractures are one of the most common indications for hospitalization and orthopedic intervention. Fragility hip fractures are frequently associated with multiple comorbidities and thus may benefit from a structured multidisciplinary approach for treatment. The purpose of this article was to retrospectively analyze patient outcomes after the implementation of a multidisciplinary hip fracture pathway at a level I trauma center. Materials and Methods: A retrospective review of 263 patients over the age of 65 with fragility hip fracture was performed. Time to surgery, hospital length of stay, Charlson Comorbidity Index (CCI), American Society of Anesthesiologists, complication rates, and other clinical outcomes were compared between patients treated in the year before and after implementation of a multidisciplinary hip fracture pathway. Results: Timing to OR, hospital length of stay, and complication rates did not differ between pre- and postpathway groups. The postpathway group had a greater CCI score (pre: 3.10 ± 3.11 and post: 3.80 ± 3.18). Fewer total blood products were administered in the postpathway group (pre: 1.5 ± 1.8 and post: 0.8 ± 1.5). Discussion: The maintenance of clinical outcomes in the postpathway cohort, while having a greater CCI, indicates the same quality of care was provided for a more medically complex patient population. With a decrease in total blood products in the postpathway group, this highlights the economic importance of perioperative optimization that can be obtained in a multidisciplinary pathway. Conclusion: Implementation of a multidisciplinary hip fracture pathway is an effective strategy for maintaining care standards for fragility hip fracture management, particularly in the setting of complex medical comorbidities.

Abstract Weighable lysimeters were used to study the relation between soil water content (SWC) and the actual evapotranspiration (ETa) of grassland under two different climate regimes of Rollesbroich and Selhausen but for an identical soil from Rollesbroich. All components of the water balance were determined from 2012 until 2018. Budyko analysis was used to characterize the hydrological status of the studied sites. Wavelet analysis was also applied to study the power spectrum of ETa, vegetation‐height‐adjusted reference evapotranspiration (ETcrop), and water stress index (WSI) defined as ETa/ETcrop, as well as SWC at three different depths and the coherence between SWC and ETa and WSI. The Budyko analysis showed that 2018 resulted in a shift of both locations towards more water‐limited conditions, although Rollesbroich remained an energy‐limited system. Based on the power spectrum analysis, the annual timescale is the dominant scale for the temporal variability of ETa, ETcrop, and SWC. The results also showed that increasing dryness at the energy‐limited site led to more temporal variability of SWC at all depths at the annual timescale. Wavelet coherence analysis showed a reduction of the phase shift between SWC and ETa at an annual scale caused by the increase in dryness during the measurement period. We found that phase shifts between SWC and ETa and SWC and WSI were stronger at the water‐limited site than at the energy‐limited site. The wavelet coherence analysis also showed that from 2014 to 2018, the control of ETa and WSI on SWC increased due to higher dryness of soil.

Abstract In-gel digestion has been used as a standard method for the preparation of protein samples for mass spectrometry analysis for over 25 years. Traditional in gel-digestion procedures require extensive sample handling, are prone to contamination and not compatible with high-throughput sample preparation. To address these shortcomings, we have modified the conventional in-gel digestion procedure for high-throughput proteomics studies. The modified method, termed “High Throughput in Gel digestion” (HiT-Gel), is based on a 96-well plate format which results in a drastic reduction in labour intensity and sample handling. Direct comparison revealed that HiT-Gel reduces technical variation and significantly decreases sample contamination over the conventional in-gel digestion method. HiT-Gel also produced superior results when a single protein band was excised from a gel and processed by in-gel digestion. Moreover, we applied Hit-Gel for a mass spectrometry analysis of Arabidopsis thaliana protein complexes separated by native PAGE in 24 fractions and four biological replicates. We show that the high throughput capacity of HiT-Gel facilitates large scale studies with high sample replication or detailed fractionation. Our method can easily be implemented as it does not require specialised laboratory equipment.

Abstract Plants respond to seasonal cues such as the photoperiod, to adapt to current conditions and to prepare for environmental changes in the season to come. To assess photoperiodic responses at the protein level, we quantified the proteome of the model plant Arabidopsis thaliana by mass spectrometry across four photoperiods. This revealed coordinated changes of abundance in proteins of photosynthesis, primary and secondary metabolism, including pigment biosynthesis, consistent with higher metabolic activity in long photoperiods. Higher translation rates in the day than the night likely contribute to these changes, via an interaction with rhythmic changes in RNA abundance. Photoperiodic control of protein levels might be greatest only if high translation rates coincide with high transcript levels in some photoperiods. We term this proposed mechanism “translational coincidence”, mathematically model its components, and demonstrate its effect on the Arabidopsis proteome. Datasets from a green alga and a cyanobacterium suggest that translational coincidence contributes to seasonal control of the proteome in many phototrophic organisms. This may explain why many transcripts but not their cognate proteins exhibit diurnal rhythms.

A Correction to this paper has been published: https://doi.org/10.1038/s41597-021-00851-9.

A metabolon is a complex of sequential metabolic enzymes that channels substrates directly between enzymes, thus optimizing metabolic flux. Here Zhanget al. provide protein interaction and isotope dilution data that support the existence of a metabolon that channels both citrate and fumarate in the plant TCA cycle.

Persistent sciatic arteries are a rare but clinically significant vascular etiology of lower extremity arterial aneurysms leading to distal embolization and limb ischemia. We present the case of a 60-year-old man with a symptomatic persistent left sciatic artery aneurysm with a hypoplastic superficial femoral artery who was successfully treated with a hybrid open and endovascular technique. This consisted of a surgical bypass from the common femoral artery to the above-knee popliteal artery with prosthetic graft combined with retrograde endovascular embolization of the sciatic artery aneurysm.

The t(8;21) translocation is often found in acute myeloid leukaemia but is not sufficient for development of the disease. In this study, the authors identify frequent mutations in the transcriptional repressor, ZBTB7A, in these patients and show that the mutations reduce DNA binding activity.

Panniculectomy performed in patients with morbid obesity before or after massive weight loss improves body contour, incidence of soft tissue infection and overall functional status. However, obtaining adequate surgical exposure remains one of the most critical and challenging aspects of the procedure. We present the case of a 75-year-old veteran who underwent successful panniculectomy using a novel technique of abdominal suspension that is easier, more effective and less costly than previously used alternatives.

This work investigates a new generation structural health monitoring (SHM) system for fibre metal laminates (FML) based on an embedded thermoplastic film with compounded piezoceramics, termed piezo-active fibre metal laminate (PFML). The PFML is manufactured using near-series processes and its potential as a passive SHM system is being investigated. A commercial Polyvinylidene fluoride (PVDF) sensor film is used for comparative evaluation of the sensor signals. Furthermore, thermoset and thermoplastic-based FML are equipped with the sensor films and evaluated. For this purpose, static and dynamic three-point bending tests are carried out and the data are recorded. The data obtained from the sensors and the testing machine are compared with the type and time of damage by means of intelligent signal processing. By using a smart sensor system, further investigations are planned which the differentiation between various failure modes, e.g., delamination or fibre breakage.

Conventional preparation methods of plant ribosomes fail to resolve non-translating chloroplast or cytoplasmic ribosome subunits from translating fractions. We established preparation of these ribosome complexes from Arabidopsis thaliana leaf, root, and seed tissues by optimized sucrose density gradient centrifugation of protease protected plant extracts. The method co-purified non-translating 30S and 40S ribosome subunits separated non-translating 50S from 60S subunits, and resolved assembled monosomes from low oligomeric polysomes. Combining ribosome fractionation with microfluidic rRNA analysis and proteomics, we characterized the rRNA and ribosomal protein (RP) composition. The identity of cytoplasmic and chloroplast ribosome complexes and the presence of ribosome biogenesis factors in the 60S-80S sedimentation interval were verified. In vivo cross-linking of leaf tissue stabilized ribosome biogenesis complexes, but induced polysome run-off. Omitting cross-linking, the established paired fractionation and proteome analysis monitored relative abundances of plant chloroplast and cytoplasmic ribosome fractions and enabled analysis of RP composition and ribosome associated proteins including transiently associated biogenesis factors.

Boron-oxygen related light-induced degradation (BO-LID) of effective charge carrier lifetime is one of the major problems for photovoltaics based on oxygen-rich boron-doped wafer substrates. Within this contribution, the dependence of slow BO-related degradation rate on total hole concentration at 30°C is investigated. A widened high power 805 nm IR-laser is used to reach injection levels comparable with the doping level of the used 2 Ωcm material thus significantly impacting total hole concentration. It is found that slow BO-related degradation rate scales almost quadratically with total hole concentration in best agreement with results from other groups suggesting the involvement of two holes in the slow BO-related degradation mechanism.

A major challenge in biology is determining how evolutionarily novel characters originate; however, mechanistic explanations for the origin of new characters are almost completely unknown. The evolution of pregnancy is an excellent system in which to study the origin of novelties because mammals preserve stages in the transition from egg laying to live birth. To determine the molecular bases of this transition, we characterized the pregnant/gravid uterine transcriptome from tetrapods to trace the evolutionary history of uterine gene expression. We show that thousands of genes evolved endometrial expression during the origins of mammalian pregnancy, including genes that mediate maternal-fetal communication and immunotolerance. Furthermore, thousands of cis-regulatory elements that mediate decidualization and cell-type identity in decidualized stromal cells are derived from ancient mammalian transposable elements (TEs). Our results indicate that one of the defining mammalian novelties evolved from DNA sequences derived from ancient mammalian TEs co-opted into hormone-responsive regulatory elements distributed throughout the genome.

Heterozygous mutations in GATA2 underlie different syndromes, previously described as monocytopenia and mycobacterial avium complex infection (MonoMAC); dendritic cell, monocytes, B- and NK lymphocytes deficiency (DCML); lymphedema, deafness and myelodysplasia (Emberger syndrome) and familiar myelodysplastic syndrome/acute myeloid leukemia (MDS / AML). Onset and severity of clinical symptoms vary and preceding cytopenias are not always present. We describe a case of symptomatic DCML deficiency and rather discrete bone marrow findings due to GATA2 mutation. Exome sequencing revealed a somatic ASXL1 mutation and the patient underwent allogeneic stem cell transplantation successfully.

The circadian clock regulates physiological processes central to growth and survival. To date, most plant circadian clock studies have relied on diurnal transcriptome changes to elucidate molecular connections between the circadian clock and observable phenotypes in wild-type plants. Here, we have integrated RNA-sequencing and protein mass spectrometry data to comparatively analyse the lhycca1, prr7prr9, gi and toc1 circadian clock mutant rosette at the end of day and end of night. Each mutant affects specific sets of genes and proteins, suggesting that the circadian clock regulation is modular. Furthermore, each circadian clock mutant maintains its own dynamically fluctuating transcriptome and proteome profile specific to subcellular compartments. Most of the measured protein levels do not correlate with changes in their corresponding transcripts. Transcripts and proteins that have coordinated changes in abundance are enriched for carbohydrate- and cold-responsive genes. Transcriptome changes in all four circadian clock mutants also affect genes encoding starch degradation enzymes, transcription factors and protein kinases. The comprehensive transcriptome and proteome datasets demonstrate that future system-driven research of the circadian clock requires multi-level experimental approaches. Our work also shows that further work is needed to elucidate the roles of post-translational modifications and protein degradation in the regulation of clock-related processes.

Plants as sessile organisms can adapt to environmental stress to mitigate its adverse effects. As part of such adaptation they maintain an active memory of heat stress for several days that promotes a more efficient response to recurring stress. We show that this heat stress memory requires the activity of the FORGETTER1 (FGT1) locus, with fgt1 mutants displaying reduced maintenance of heat-induced gene expression. FGT1 encodes the Arabidopsis thaliana orthologue of Strawberry notch (Sno), and the protein globally associates with the promoter regions of actively expressed genes in a heat-dependent fashion. FGT1 interacts with chromatin remodelers of the SWI/SNF and ISWI families, which also display reduced heat stress memory. Genomic targets of the BRM remodeler overlap significantly with FGT1 targets. Accordingly, nucleosome dynamics at loci with altered maintenance of heat-induced expression are affected in fgt1. Together, our results suggest that by modulating nucleosome occupancy, FGT1 mediates stress-induced chromatin memory.