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Dpto. Medio Ambiente y Agronomía (INIA), The current environmental hazard assessment is based on the testing of the pristine substance. However, it cannot be excluded that (nano)pharmaceuticals are excreted into sewage during the use phase followed by entry into wastewater treatment plants (WWTPs). Sorption to sewage sludge or release via effluent can result in modified ecotoxicological effects which possibly can only be detected with a modified test approach. The objective of our study was to investigate a realistic exposure scenario for metallic nanoparticles (NPs) in pharmaceutical products, excreted into effluent, and released into the environment after treatment in WWTPs. The test approach was illustrated by using gold (Au) NPs. Effluent from model WWTPs were investigated in aquatic tests (Daphnia magna, fish cell lines). Sewage sludge was used as a sole food source (Eisenia fetida) or mixed with soil and used as test medium (soil microorganisms, Folsomia candida, Enchytraeus crypticus). To cover the aspect of regulation, the test systems described in OECD-test guidelines (OECD TG 201, 211, 220, 232, 249, 317) were applied. Modifications and additional test approaches were included to meet the needs arising out of the testing of nanomaterials and of the exposure scenarios. The results were assessed regarding the suitability of the test design and the toxicity of Au-NPs. Except for activated sludge as a sole food source for E.fetida, the selected test approach is suitable for the testing of nanomaterials. Additional information can be gained when compared to the common testing of the pristine nanomaterials in the standardized test systems. Effects of Au-NPs were observed in concentrations exceeding the modeled environmental., This project has received funding from the European Union’s Horizon 2020 research and innovation programme under BIORIMA grant agreement No 760928 for the testing and under NANORIGO grant agreement No 814530 regarding risk governance. Further, to FCT (Fundação para a Ciência e Tecnologia) / MCTES (Ministério da Ciência e Tecnologia do Ensino Superior) for the financial support to CESAM (UIDP/50017/2020+UIDB/50017/2020). S.I.L. Gomes is funded by National Funds (OE), through FCT, I.P., in the scope of the framework contract foreseen in the numbers 4, 5 and 6 of the article 23, of the Decree-Law 57/2016, of August 29, changed by Law 57/2017, of July 19.

Our earlier in vitro and in vivo studies have revealed that the phytosterol, pentalinonsterol (cholest-4,20,24-trien-3-one) (PEN), isolated from the roots of Pentalinon andrieuxii, possesss immunomodulatory properties in macrophages and dendritic cells. Leishmaniasis, caused by the infection of Leishmania spp. (a protozoan parasite), is emerging as the second-leading cause of mortality among the tropical diseases and there is an unmet need for a pharmacological intervention of leishmaniasis. Given the beneficial immunomodulatory actions and lipophilic properties of PEN, the objective of this study was to elucidate the mechanism(s) of action of the immunomodulatory action(s) of PEN in macrophages through the modulation of phospholipase A2 (PLA2) activity that might be crucial in the antileishmanial action of PEN. Therefore, in this study, we investigated whether PEN would modulate the activity of PLA2 in RAW 264.7 macrophages and mouse bone marrow-derived primary macrophages (BMDMs) in vitro and further determined how the upstream PLA2 activation would regulate the downstream cytokine release in the macrophages. Our current results demonstrated that (i) PEN induced PLA2 activation (arachidonic acid release) in a dose- and time-dependent manner that was regulated upstream by the mitogen-activated protein kinases (MAPKs); (ii) the PEN-induced activation of PLA2 was attenuated by the cPLA2-specific pharmacological inhibitors; and (iii) the cPLA2-specific pharmacological inhibitors attenuated the release of inflammatory cytokines from the macrophages. For the first time, our current study demonstrated that PEN exhibited its immunomodulatory actions through the activation of cPLA2 in the macrophages, which potentially could be used in the development of a pharmacological intervention against leishmaniasis.

Rationale: Coronary artery disease (CAD) is a pervasive and critical health care problem. Elevated high-density lipoprotein-associated cholesterol (HDL-C) is associated with improved atherosclerotic cardiovascular disease outcomes on a population level, but clinical trials aimed at HDL-C elevation have not succeeded in improving atherosclerotic cardiovascular disease event risk. Nevertheless, human variants in the HDL receptor, encoded by SCARB1 , are associated with dyslipidemia, suggesting that HDL metabolism, not HDL-C, is a suitable target for therapy. However, variants in SCARB1 have never been directly attributed to CAD by Mendelian inheritance. Objective: To determine if compound heterozygous variants in SCARB1 cause disease in 2 brothers with severe, early-onset CAD. Methods and Results: Using whole exome sequencing, we have identified rare, compound heterozygous variants in SCARB1 that segregate with severe, premature CAD, following patterns of Mendelian inheritance. Using induced pluripotent stem cell–derived hepatocyte-like cells from the proband, we discovered the maternal variant (c.754_755delinsC) to be the first identified SCARB1 null allele, characterized by the absence of RNA and protein expression. Further, we demonstrate that the variant on the paternal allele (c.956G>T [p.G319V]) results in decreased cholesterol uptake, decreased SR-BI:HDL binding, and increased affinity for SR-BI dimerization. Finally, we generated a p.G319V knock-in mouse model that displays nearly 100% homozygous lethality and elevated plasma cholesterol in heterozygous animals, confirming pathogenicity of this variant. Conclusions: In summary, our data provide the first molecular mechanism to show the Mendelian inheritance of CAD as a result of human SCARB1 variants. The rarity of these variants supports pathogenicity in this family. Furthermore, SR-BI p.G319V, which has previously been reported benign in the context of heterozygosity, was uniquely presented alongside a null allele, demonstrating the disease-contributing capability of loss-of-function SCARB1 variants within the population.

There are currently few suitable test systems for the chronic toxicity testing of aquatic macroinvertebrates under stream conditions. Therefore, a new test system mimicking running water conditions was developed for testing with lotic insects. This system uses small test cages, with 10 of these suspended inside each 25-L container and rotating at 0.1 m/s, to create a water flow for the individual organism inside each cage. To test the performance of the new exposure system, chronic effects (21 d) of the neonicotinoid imidacloprid were investigated with field-collected larvae of the stonefly Protonemura sp. Endpoints were survival, growth, and/or emergence (depending on the developmental stage of the larvae at the start of the exposure). Two experiments conducted 1 yr apart showed good reproducibility: growth 10% effect concentration (EC10) values were 15.3 and 18.5 mg/L and no-observed-effect concentration (NOEC) values were 30.3 and 21.5 mg/L. A third experiment, performed with further-developed larval instars, showed a significant effect of imidacloprid on emergence (with EC10 of 5.97 mg/L and NOEC of 2.89 mg/L) and a significant effect on survival (with median lethal concentration of 44.7 µg/L). The results of the present study show that the newly developed test system provides a suitable approach for toxicity testing with stonefly larvae and potentially for other lotic macroinvertebrate species. Environ Toxicol Chem 2021;40:2229-2239. © 2021 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Sirtuins have been shown to regulate the aging process. We have previously demonstrated that Sirt6 blocks the pressure overload-induced cardiac hypertrophy in mice. Here, we show that Sirt6 can also mitigate aging-induced cardiomyocyte senescence and cardiac hypertrophy. We found that aging is associated with altered Sirt6 activity along with development of cardiac hypertrophy and fibrosis. Compared to young mice (4-months), the hearts of aged mice (24-months) showed increased levels of mitochondrial DNA damage, shortened telomere length, and increased accumulation of 8-oxo-dG adducts, which are hallmarks of aging. The aged hearts also showed reduced levels of NAD+ and altered levels of mitochondrial fusion-fission proteins. Similar characteristics were observed in the hearts of Sirt6 deficient mice. Additionally, we found that doxorubicin (Dox) induced cardiomyocyte senescence, as measured by expression of p16INK4a, p53, and β-galactosidase, was associated with loss of Sirt6. However, Sirt6 overexpression protected cardiomyocytes from developing Dox-induced senescence. Further, compared to wild-type mice, the hearts of Sirt6.Tg mice showed reduced expression of aging markers, and the development of aging-associated cardiac hypertrophy and fibrosis. Our data suggest that Sirt6 is a critical anti-aging molecule that regulates various cellular processes associated with aging and protects the heart from developing aging-induced cardiac hypertrophy and fibrosis.

Sex chromosomes often bear distinct patterns of genetic variation due to unique patterns of inheritance and demography. The processes of mutation, recombination, genetic drift and selection also influence rates of evolution on sex chromosomes differently than autosomes. Measuring such differences provides information about how these processes shape genomic variation and their roles in the origin of species. To test hypotheses and predictions about patterns of autosomal and sex-linked genomic diversity and differentiation, we measured population genetic statistics within and between populations and subspecies of the barn swallow (Hirundo rustica) and performed explicit comparisons between autosomal and Z-linked genomic regions. We first tested for evidence of low Z-linked genetic diversity and high Z-linked population differentiation relative to autosomes, then for evidence that the Z chromosome bears greater ancestry information due to faster lineage sorting. Finally, we investigated geographical clines across hybrid zones for evidence that the Z chromosome is resistant to introgression due to selection against hybrids. We found evidence that the barn swallow mating system, demographic history and linked selection each contribute to low Z-linked diversity and high Z-linked differentiation. While incomplete lineage sorting is rampant across the genome, our results indicate faster sorting of ancestral polymorphism on the Z. Finally, hybrid zone analyses indicate barriers to introgression on the Z chromosome, suggesting that sex-linked traits are important in reproductive isolation, especially in migratory divide regions. Our study highlights how selection, gene flow and demography shape sex-linked genetic diversity and underlines the relevance of the Z chromosome in speciation.

Rationale: Cardiac hypertrophy, a major risk factor for heart failure, occurs when cardiomyocytes remodel in response to complex signaling induced by injury or cell stress. Although cardiomyocytes are the ultimate effectors of cardiac hypertrophy, nonmyocyte populations play a large yet understudied role in determining how cardiomyocytes respond to stress. Objective: To identify novel paracrine regulators of cardiomyocyte hypertrophic remodeling. Methods and Results: We have identified a novel role for a nonmyocyte-derived and TGFβ1 (transforming growth factor β1)–induced extracellular matrix protein MFAP4 (microfibrillar-associated protein 4) in the pathophysiology of cardiac remodeling. We have determined that nonmyocyte cells are the primary sources of MFAP4 in the heart in response to TGFβ1 stimulation. Furthermore, we have demonstrated a crucial role of MFAP4 in the cardiac adaptation to stress. Global knockout of MFAP4 led to increased cardiac hypertrophy and worsened cardiac function following chronic pressure overload. Also, one week of angiotensin-mediated neurohumoral stimulation was sufficient to exacerbate cardiomyocyte hypertrophy in MFAP4 null mice. In contrast, administration of exogenous MFAP4 to isolated cardiomyocytes blunted their phenylephrine-induced hypertrophic growth through an integrin-dependent mechanism. Finally, MFAP4 deficiency leads to dysregulated integration of G protein-coupled receptor and integrin signaling in the heart. Conclusions: Altogether, our results demonstrate a critical paracrine role of MFAP4 in the development of cardiac hypertrophy and could inform future treatment options for patients with heart failure.

Background: The Tumor Proportion Score (TPS) and Combined Positive Score (CPS) scoring algorithms are used in conjunction with PD-L1 IHC 22C3 pharmDx for the immunohistochemical evaluation of PD-L1 in certain human cancer tissues; both algorithms include PD-L1 staining tumor cells (TC) and have been correlated with response to pembrolizumab therapy (KEYTRUDA®) at specific expression levels in specific tumor indications. This study aims to evaluate whether PD-L1 expression on TC could be important in the evaluation of TNBC with PD-L1 IHC 22C3 pharmDx and the impact on reaching a diagnostic cut-off, as current PD-L1 testing (SP142) for TNBC relies solely on scoring immune cells (IC). TNBC clinical data from KEYNOTE-119 (ClinicalTrials.gov, NCT02555657) and Agilent’s internal TNBC tumor bank data were evaluated by TPS, CPS, and Quantitative Immune Cell Density (QID) to determine the significance of including PD-L1 expressing TC in the scoring algorithms. PD-L1 IHC 22C3 pharmDx is not currently approved for use with TNBC specimens.Methods: TNBC specimens from clinical trial KEYNOTE-119 and Agilent’s internal tumor bank were stained with PD-L1 IHC 22C3 pharmDx and scored using the TPS, CPS, and QID algorithms. The TPS algorithm is defined as the number of PD-L1 staining tumor cells divided by the total number of viable TC, multiplied by 100. The CPS algorithm includes TC and IC and is defined as the number of PD-L1 staining cells (TC, lymphocytes and macrophages) divided by the total number of viable TC, multiplied by 100. In addition to TPS and CPS, QID was also calculated to quantify the contribution from PD-L1 expressing IC. QID is defined as the CPS minus the TPS (QID = CPS - TPS). The data was analyzed to show the percent of specimens that fell below the diagnostic PD-L1 cut-offs of ≥1 and ≥10 when evaluated using QID as compared to CPS. Both scores measure countable PD-L1 staining cells per 100 viable tumor cells. Results: A total of 120 PD-L1 IHC 22C3 pharmDx stained TNBC specimens from Agilent’s internal tumor bank were analyzed. When evaluated using CPS, 71 specimens were positive for the PD-L1 ≥1 cut-off and 49 were positive for the PD-L1 ≥10 cut-off. When analyzed with QID, 8.5% of specimens fell below the PD-L1 ≥1 cut-off and 26.5% fell below the PD-L1 ≥10 cut-off (Table 1). A total of 964 PD-L1 IHC 22C3 pharmDx stained TNBC specimens from KEYNOTE-119 were analyzed. When evaluated using CPS, 604 specimens were positive for the PD-L1 ≥1 diagnostic cut-off and 284 were positive for the PD-L1 ≥10 diagnostic cut-off. When analyzed with QID, 9.3% of patients fell below the PD-L1 ≥1 diagnostic cut-off and 33.5% fell below the PD-L1 ≥10 diagnostic cut-off (Table 1). Conclusion: PD-L1 IHC 22C3 pharmDx stains both TC and IC in TNBC. Removal of the PD-L1 staining TC from the CPS algorithm (QID score) reduces the number of specimens scored as positive for the PD-L1 ≥1 and PD-L1 ≥10 diagnostic cut-offs. This data highlights that a significant portion of TC express PD-L1 in TNBC and inclusion of TC in the scoring algorithm should be considered when evaluating the diagnostic status of a specimen. Table 1. Agilent Tumor Bank and KEYNOTE -119: CPS and QIDDiagnostic Cut-OffCPS Positive SpecimensQID (CPS – TPS) Positive SpecimensSpecimens Flipped at Diagnostic Cut-offDifference (%)Agilent Tumor BankPD-L1 ≥ 1716568.5PD-L1 ≥ 1049361326.5KEYNOTE-119PD-L1 ≥ 1604548569.3PD-L1 ≥ 102841899533.5 Citation Format: Tiffany Evans, Stephanie Hund, Darlene Krohn, Kenneth Emancipator, Jonathon Juco, Bryce Portier, Siena Tabuena-Frolli, Karina Kulangara. Significance of PD-L1 expressing tumor cells in the combined positive score with triple negative breast cancer [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS4-15.

Associations between parasite loads and sexual signal expression have long been the focus of research. However, our understanding of how sexual selection operates in the context of multiple parasite infections within the same host is still quite limited. We examined the expression of plumage coloration, which is sexually selected in North American barn swallows, Hirundo rustica erythrogaster, and tail streamer length, which is sexually selected in other barn swallow subspecies, in the context of eight different parasite infections. We found that two parasites, nest mites and Plasmodium, were associated with colour expression, but in opposite directions. Attractive males were less likely to have mites in their nests, but more likely to be infected with Plasmodium. We found that different underlying mechanisms were generating the links between these different parasites and the same colour signal. Males that invested more in colour expression were less likely to survive when infected with Plasmodium, suggesting a physiological mechanism linking colour to Plasmodium infections. However, we found no clear cost to males when examining nest mites, which primarily infect offspring. Instead, using experimental mite manipulations, we provide evidence for a behavioural mechanism, where territorial behaviour and male–male competition likely drive the association between colour expression and nest mite infections. We did not find associations between parasites and streamer length in this subspecies, suggesting that the links between divergent sexual signals and parasites may be quite flexible. These results suggest that the evolution of a single sexual signal can be shaped by several, even opposing, links with different parasite infections.

Background Atrial fibrillation (AF) is the most common sustained arrhythmia, with growing evidence identifying obesity as an important risk factor for the development of AF. Although defective atrial myocyte excitability due to stress-induced remodeling of ion channels is commonly observed in the setting of AF, little is known about the mechanistic link between obesity and AF. Recent studies have identified increased cardiac late sodium current (INa,L) downstream of calmodulin-dependent kinase II (CaMKII) activation as an important driver of AF susceptibility. Methods Here, we investigated a possible role for CaMKII-dependent INa,L in obesity-induced AF using wild-type (WT) and whole-body knock-in mice that ablates phosphorylation of the Nav1.5 sodium channel and prevents augmentation of the late sodium current (S571A; SA mice). Results A high-fat diet (HFD) increased susceptibility to arrhythmias in WT mice, while SA mice were protected from this effect. Unexpectedly, SA mice had improved glucose homeostasis and decreased body weight compared to WT mice. However, SA mice also had reduced food consumption compared to WT mice. Controlling for food consumption through pair feeding of WT and SA mice abrogated differences in weight gain and AF inducibility, but not atrial fibrosis, premature atrial contractions or metabolic capacity. Conclusions These data demonstrate a novel role for CaMKII-dependent regulation of Nav1.5 in mediating susceptibility to arrhythmias and whole-body metabolism under conditions of diet-induced obesity.

Temperature response of stem elongation in wheat grown under field conditions indicated that temperature response is highly heritable and linked to the flowering pathway., In wheat, temperature affects the timing and intensity of stem elongation. Genetic variation for this process is therefore important for adaptation. This study investigates the genetic response to temperature fluctuations during stem elongation and its relationship to phenology and height. Canopy height of 315 wheat genotypes (GABI wheat panel) was scanned twice weekly in the field phenotyping platform (FIP) of ETH Zurich using a LIDAR. Temperature response was modelled using linear regressions between stem elongation and mean temperature in each measurement interval. This led to a temperature-responsive (slope) and a temperature-irresponsive (intercept) component. The temperature response was highly heritable (H2=0.81) and positively related to a later start and end of stem elongation as well as final height. Genome-wide association mapping revealed three temperature-responsive and four temperature-irresponsive quantitative trait loci (QTLs). Furthermore, putative candidate genes for temperature-responsive QTLs were frequently related to the flowering pathway in Arabidopsis thaliana, whereas temperature-irresponsive QTLs corresponded to growth and reduced height genes. In combination with Rht and Ppd alleles, these loci, together with the loci for the timing of stem elongation, accounted for 71% of the variability in height. This demonstrates how high-throughput field phenotyping combined with environmental covariates can contribute to a smarter selection of climate-resilient crops.

Oxidant stress can contribute to health and disease. Here we show that invertebrates and vertebrates share a common stereospecific redox pathway that protects against pathological responses to stress, at the cost of reduced physiological performance, by constraining Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) activity. MICAL1, a methionine monooxygenase thought to exclusively target actin, and MSRB, a methionine reductase, control the stereospecific redox status of M308, a highly conserved residue in the calmodulin-binding (CaM-binding) domain of CaMKII. Oxidized or mutant M308 (M308V) decreased CaM binding and CaMKII activity, while absence of MICAL1 in mice caused cardiac arrhythmias and premature death due to CaMKII hyperactivation. Mimicking the effects of M308 oxidation decreased fight-or-flight responses in mice, strikingly impaired heart function in Drosophila melanogaster, and caused disease protection in human induced pluripotent stem cell–derived cardiomyocytes with catecholaminergic polymorphic ventricular tachycardia, a CaMKII-sensitive genetic arrhythmia syndrome. Our studies identify a stereospecific redox pathway that regulates cardiac physiological and pathological responses to stress across species.

Hirundo is the most species-rich genus of the passerine swallow family (Hirundinidae) and has a cosmopolitan distribution. Here we report the complete, annotated mitochondrial genomes for 25 indivi...

Many closely related populations are distinguished by variation in sexual signals and this variation is hypothesized to play an important role in reproductive isolation and speciation. Within populations, there is considerable evidence that sexual signals provide information about the incidence and severity of parasite infections, but it remains unclear if variation in parasite communities across space could play a role in initiating or maintaining sexual trait divergence. To test for variation in parasite-associated selection, we compared three barn swallow subspecies with divergent sexual signals. We found that parasite community structure and host tolerance to ecologically similar parasites varied between subspecies. Across subspecies we also found that different parasites were costly in terms of male survival and reproductive success. For each subspecies, the preferred sexual signal(s) were associated with the most costly local parasite(s), indicating that divergent signals are providing relevant information to females about local parasite communities. Across subspecies, the same traits were often associated with different parasites, indicating that parasite-sexual signal links are quite flexible and may evolve relatively quickly. This study provides evidence for (1) variation in parasite communities and (2) different parasite-sexual signal links among three closely related subspecies with divergent sexual signal traits, suggesting that parasites may play an important role in initiating and/or maintaining the divergence of sexual signals among these closely related, yet geographically isolated populations.

Objective To quantify peripheral nerve lesions in symptomatic and asymptomatic hereditary transthyretin amyloidosis with polyneuropathy (ATTRv‐PNP) by analyzing the magnetization transfer ratio (MTR) of the sciatic nerve, and to test its potential as a novel biomarker for macromolecular changes. Methods Twenty‐five patients with symptomatic ATTRv‐PNP, 30 asymptomatic carriers of the mutant transthyretin gene (mutTTR), and 20 age‐/sex‐matched healthy controls prospectively underwent magnetization transfer contrast imaging at 3 Tesla. Two axial three‐dimensional gradient echo sequences with and without an off‐resonance saturation rapid frequency pulse were conducted at the right distal thigh. Sciatic nerve regions of interest were manually drawn on 10 consecutive axial slices in the images without off‐resonance saturation, and then transferred to the corresponding slices that were generated by the sequence with the off‐resonance saturation pulse. Subsequently, the MTR and cross‐sectional area (CSA) of the sciatic nerve were evaluated. Detailed neurologic and electrophysiologic examinations were conducted in all ATTRv‐PNP patients and mutTTR‐carriers. Results Sciatic nerve MTR and CSA reliably differentiated between ATTRv‐PNP, mutTTR‐carriers, and controls. MTR was lower in ATTRv‐PNP (26.4 ± 0.7; P

The widespread integration of engineered nanomaterials into consumer and industrial products creates new challenges and requires innovative approaches in terms of design, testing, reliability, and safety of nanotechnology. The aim of this review article is to give an overview of different product groups in which nanomaterials are present and outline their safety aspects for consumers. Here, release of nanomaterials and related analytical challenges and solutions as well as toxicological considerations, such as dose-metrics, are discussed. Additionally, the utilization of engineered nanomaterials as pharmaceuticals or nutraceuticals to deliver and release cargo molecules is covered. Furthermore, critical pathways for human exposure to nanomaterials, namely inhalation and ingestion, are discussed in the context of risk assessment. Analysis of NMs in food, innovative medicine or food contact materials is discussed. Specific focus is on the presence and release of nanomaterials, including whether nanomaterials can migrate from polymer nanocomposites used in food contact materials. With regard to the toxicology and toxicokinetics of nanomaterials, aspects of dose metrics of inhalation toxicity as well as ingestion toxicology and comparison between in vitro and in vivo conclusions are considered. The definition of dose descriptors to be applied in toxicological testing is emphasized. In relation to potential exposure from different products, opportunities arising from the use of advanced analytical techniques in more unique scenarios such as release of nanomaterials from medical devices such as orthopedic implants are addressed. Alongside higher product performance and complexity, further challenges regarding material characterization and safety, as well as acceptance by the general public are expected.

This article reviews the current status of nanotechnology with emphasis on application and related environmental considerations as well as legislation. Application and analysis of nanomaterials in infrastructure (construction, building coatings, and water treatment) is discussed, and in particular nanomaterial release during the lifecycle of these applications. Moreover, possible grouping approaches with regard to ecotoxicological and toxicological properties, and the fate of nanomaterials in the environment are evaluated. In terms of potential exposure, the opportunities that arise from leveraging advances in several key areas, such as water treatment and construction are addressed. Additionally, this review describes challenges with regard to the European Commission’s definition of ‘nanomaterial’. The revised REACH information requirements, intended to enable a comprehensive risk assessment of nanomaterials, are outlined.

BackgroundThe Combined Positive Score (CPS)1 algorithm includes tumor and immune cells for determination of Programed Death-Ligand 1 (PD-L1) protein expression in tumor tissues and has been analytically and clinically validated for use with PD-L1 IHC 22C3 pharmDx across multiple indications and cutoffs.2 PD-L1 22C3 IHC pharmDx is a qualitative immunohistochemical assay using anti-PD-L1, Clone 22C3 to detect PD-L1 in formalin-fixed, paraffin-embedded (FFPE) tumor tissues using Autostainer Link 48. PD-L1 IHC 22C3 pharmDx is FDA-approved as an aid in identifying patients for treatment with KEYTRUDA® for six tumor indications at clinically validated CPS diagnostic cutoffs2: gastric or gastroesophageal junction (GC/GEJ) adenocarcinoma (CPS ≥ 1), cervical cancer (CPS ≥ 1), urothelial carcinoma (CPS ≥ 10), head and neck squamous cell carcinoma (HNSCC) (CPS ≥ 1), esophageal squamous cell carcinoma (ESCC) (CPS ≥ 10)3, and triple-negative breast cancer (TNBC) (CPS ≥ 10).MethodsPrecision of PD-L1 IHC 22C3 pharmDx using CPS was assessed for all six indications at the corresponding clinically validated diagnostic cutoffs and at additional exploratory cutoffs under normal, day-to-day testing conditions. Precision testing included Combined Precision (inter-instrument/operator/run (day)), Intra-Run Repeatability, and Observer (inter-/intra-) Scoring Reproducibility studies. FFPE specimens were stained with PD-L1 IHC 22C3 pharmDx and scored using CPS as described in the package insert.2 Four CPS cutoffs were evaluated: CPS ≥ 1 (GC/GEJ, urothelial carcinoma, ESCC, cervical cancer, HNSCC, TNBC), CPS ≥ 10 (GC/GEJ, urothelial carcinoma, ESCC, TNBC), CPS ≥ 20 (HNSCC), and CPS ≥ 50 (HNSCC). Data were analyzed using negative percent agreement (NPA), positive percent agreement (PPA), and overall agreement (OA) with two-sided 95% percentile bootstrap confidence intervals (CIs) based on PD-L1 binary status at the applicable cutoff(s). For each study, data from each CPS cutoff-indication pair were individually analyzed. Meta-analyses were also performed by pooling data from all indications per (i) study and cutoff, and (ii) per study for all tested cutoffs.ResultsNearly all agreement analyses (142/144) for each CPS cutoff-indication pair showed NPA/PPA/OA point estimates (PE) ≥ 90% and CI lower bounds (CILB) ≥ 85%. Meta-analyses showed PE ≥ 90% for NPA/PPA/OA and CILB ≥ 85% per study and cutoff, and per study for all tested cutoffs. Discordant comparisons accounted for ConclusionsCPS used with PD-L1 IHC 22C3 pharmDx provides precise evaluation of PD-L1 expression across multiple tumor indications and cutoffs under normal, day-to-day testing conditions.AcknowledgementsWe thank the IUSCC Cancer Center at Indiana University School of Medicine, for the use of the Tissue Procurement and Distribution Core, which provided Dako North America, Inc. service.The data and biospecimens used in this project were provided by US Biolab (Gaithersburg, MD, USA), Sofia Bio LLC (New York, NY, USA), Contract Research Ltd (Charlestown, Nevis), and Centre Hospitalier Universitaire (CHU) de Nice (Nice, France) with appropriate ethics approval and through Trans-Hit Biomarkers Inc. Tissue samples were provided by the Cooperative Human Tissue Network which is funded by the National Cancer Institute. Other investigators may have received specimens from the same subjects. Tissue samples supplied by BioIVT (Hicksville, NY, USA).Trial RegistrationN/AReferencesCPS = (# PD-L1 staining cells (tumor cells, lymphotcytes, macrophages))/(Total # viable tumor cells )×100PD-L1 IHC 22C3 pharmDx [Instructions for Use]. Available at: www.agilent.com/library/eifu. Code SK006. Accessed July 2, 2021ESCC was analytically validated as a subtype of esophageal cancer [2].Ethics ApprovalN/AConsentN/A

Neurological research and practice 3, 57 (2021). doi:10.1186/s42466-021-00155-8, Published by BioMed Central, [London]

Simple Summary Bovine parafilariosis is a disease caused by the helminth Parafilaria bovicola (Filariidae, Nematoda). Flies transmit the parasite, which grows to adulthood in an unknown location in the affected animals. The adult female worms are located in nodules under the skin, which they penetrate and lay their eggs in the fluid leaking from the site. There is virtually no information about Parafilaria bovicola in Austria. In this study, these parasites were documented in the provinces of Lower Austria, Upper Austria, Styria, Salzburg, Carinthia, and Tyrol. With a high number of cases during the 2020 study period, it can be assumed that the number of reports will increase in the near future. Abstract Veterinarians reported cases of cutaneous bleeding in cattle in Austria in the spring and summer of 2020. It was our goal to confirm the tentative diagnosis of parafilariosis by identifying Parafilaria bovicola in exudate samples using molecular methods for the first time in Austria. We asked veterinarians in the field to collect exudate from typical lesions on cattle. We performed polymerase chain reactions (PCRs) and sequenced a 674-bp section of the mitochondrial cytochrome oxidase subunit I in all positive samples. Overall, in 57 of 86 samples, P. bovicola was confirmed by PCR in cattle from Lower Austria, Upper Austria, Styria, Salzburg, Carinthia, and Tyrol. Sequencing detected four different haplotypes or genotypes, respectively, indicating multiple routes of introduction. We conclude that parafilariosis has spread in Austria and we expect that the number of reports of clinical signs and losses due to carcass damage will increase in the future.

High throughput phenotyping (HTP) platforms and devices are increasingly used for the characterization of growth and developmental processes for large sets of plant genotypes. Such HTP data require challenging statistical analyses in which longitudinal genetic signals need to be estimated against a background of spatio-temporal noise processes. We propose a two-stage approach for the analysis of such longitudinal HTP data. In a first stage, we correct for design features and spatial trends per time point. In a second stage, we focus on the longitudinal modelling of the spatially corrected data, thereby taking advantage of shared longitudinal features between genotypes and plants within genotypes. We propose a flexible hierarchical three-level P-spline growth curve model, with plants/plots nested in genotypes, and genotypes nested in populations. For selection of genotypes in a plant breeding context, we show how to extract new phenotypes, like growth rates, from the estimated genotypic growth curves and their first-order derivatives. We illustrate our approach on HTP data from the PhenoArch greenhouse platform at INRAE Montpellier and the outdoor Field Phenotyping platform at ETH Zürich.

Background and Purpose: Mechanical properties of thromboemboli play an important role in the efficacy of endovascular thrombectomy (EVT) for acute ischemic stroke. However, very limited data on mechanical properties of human stroke thrombi are available. We aimed to mechanically characterize thrombi retrieved with EVT, and to assess the relationship between thrombus composition and thrombus stiffness. Methods: Forty-one thrombi from 19 patients with acute stroke who underwent EVT between July and October 2019 were mechanically analyzed, directly after EVT. We performed unconfined compression experiments and determined tangent modulus at 75% strain (E t75 ) as a measure for thrombus stiffness. Thrombi were histologically analyzed for fibrin/platelets, erythrocytes, leukocytes, and platelets, and we assessed the relationship between histological components and E t75 with univariable and multivariable linear mixed regression. Results: Median E t75 was 560 (interquartile range, 393–1161) kPa. In the multivariable analysis, fibrin/platelets were associated with increased E t75 (aβ, 9 [95% CI, 5 to 13]) kPa, erythrocytes were associated with decreased E t75% (aβ, −9 [95% CI, −5 to −13]) kPa. We found no association between leukocytes and E t75 . High platelet values were strongly associated with increased E t75 (aβ, 56 [95% CI, 38–73]). Conclusions: Fibrin/platelet content of thrombi retrieved with EVT for acute ischemic stroke is strongly associated with increased thrombus stiffness. For thrombi with high platelet values, there was a very strong relationship with thrombus stiffness. Our data provide a basis for future research on the development of next-generation EVT devices tailored to thrombus composition.

We propose an application of network analysis to determine which traits and behaviors predict fertilizations within and between populations. This approach quantifies how reproductive behavior between individuals shapes patterns of selection and gene flow, filling an important gap in our understanding of the connection between evolutionary processes and emergent patterns.

Spectrins are cytoskeletal proteins essential for membrane biogenesis and regulation and serve critical roles in protein targeting and cellular signaling. αII spectrin (SPTAN1) is one of two α spectrin genes and αII spectrin dysfunction is linked to alterations in axon initial segment formation, cortical lamination, and neuronal excitability. Furthermore, human αII spectrin loss-of-function variants cause neurological disease. As global αII spectrin knockout mice are embryonic lethal, the in vivo roles of αII spectrin in adult heart are unknown and untested. Here, based on pronounced alterations in αII spectrin regulation in human heart failure we tested the in vivo roles of αII spectrin in the vertebrate heart. We created a mouse model of cardiomyocyte-selective αII spectrin-deficiency (cKO) and used this model to define the roles of αII spectrin in cardiac function. αII spectrin cKO mice displayed significant structural, cellular, and electrical phenotypes that resulted in accelerated structural remodeling, fibrosis, arrhythmia, and mortality in response to stress. At the molecular level, we demonstrate that αII spectrin plays a nodal role for global cardiac spectrin regulation, as αII spectrin cKO hearts exhibited remodeling of αI spectrin and altered β-spectrin expression and localization. At the cellular level, αII spectrin deficiency resulted in altered expression, targeting, and regulation of cardiac ion channels Na(V)1.5 and K(V)4.3. In summary, our findings define critical and unexpected roles for the multifunctional αII spectrin protein in the heart. Furthermore, our work provides a new in vivo animal model to study the roles of αII spectrin in the cardiomyocyte.

Research question: What concentration of anti-Mullerian hormone (AMH) corresponds to an antral follicle count (AFC) >15 for determination of ovarian reserve? Design: A prospective study conducted at 13 US fertility clinics in women aged 21–44 years who presented for AFC evaluation by transvaginal ultrasound. Serum samples were collected at the time of AFC evaluation (menstrual cycle day 2–4). AMH concentrations were measured by the Elecsys® AMH immunoassay; oestradiol and follicle-stimulating hormone (FSH) concentrations were also measured. The serum AMH cut-off able to detect AFC >15 with high sensitivity was determined (derivation cohort). Clinical performance of the AMH assay at the derived cut-off was evaluated (validation cohort). Receiver operating characteristic (ROC) analyses were also performed. Results: In the derivation cohort (n = 306), an optimal serum AMH cut-off value of 1.77 ng/ml was determined to correspond to AFC >15 with 89.63% sensitivity and 69.01% specificity, using the Elecsys AMH assay. In the validation cohort (n = 856), this 1.77 ng/ml cut-off could identify women with an AFC >15 with a sensitivity of 88.34% and a specificity of 68.29%; corresponding positive predictive and negative predictive values were 75.19% and 84.34%, respectively. ROC analyses demonstrated that AMH performed better than oestradiol or FSH in predicting AFC, with area under the curves of 85.7%, 57.1% and 69.7%, respectively, in the validation cohort. Conclusion: The Elecsys AMH immunoassay provides a robust and fully automated method to measure serum AMH levels. Women with AMH values below the cut-off of 1.77 ng/ml are unlikely to have AFC >15.

Hemophilia type A (HA) is the most common type of blood coagulation disorder. While the vast majority of cases are inherited and caused by mutations in the F8 gene, recent data raises new questions regarding the non-heritability of this disease, as well as how other molecular mechanisms might lead to the development of HA or increase the severity of the disease. Some data suggest that miRNAs may affect the severity of HA, but for some patients, miRNA-based interference might cause HA, in the absence of an F8 mutation. A mechanism in HA installation that is also worth investigating and which could be identified in the future is the epigenetic silencing of the F8 gene that might be only temporarily. Acquired HA is increasingly reported and as more cases are identified, the description of the disease might become challenging, as cases without FVIII autoantibodies might be identified.

Climate change-induced heat waves represent a severe threat to future global crop production. The extent of heat stress for agricultural crops depends on the co-occurrence of heat periods and heat sensitive phenological stages. Using the Wang and Engel phenology model and the most recent climate change projections for four sites across the Swiss Central Plateau, we estimated future heat stress exposure in Swiss wheat production and tested the potential of winter wheat genotypes with differing phenological characteristics to escape future heat periods. Across all genotypes, heat stress days (Tmax ≥ 30 °C) during the temperature sensitive stages of flowering and early grain filling increased from an average of 1.5 heat days in 1982−2006 to 2.1 by 2075−2099 with RCP2.6 (with climate change mitigation) and to 3.6 by 2075−2099 with RCP8.5 (without climate change mitigation), respectively. Across all genotypes and locations, a considerable escape from future heat periods was modelled due to a mainly temperature-driven advancement in the phenological development. Under both RCP scenarios, we predicted lower exposure to heat stress for early varieties than for late varieties. However, under the RCP8.5 scenario, for each location, heat stress exposure for early varieties was still projected to be higher by 2075−2099 than for late varieties under current conditions. Further, heat stress exposure was considerably increased at locations with cooler spring conditions, slowing down the early season phenological development and resulting in late heading dates. Our findings imply needs for a regionally adequate cultivar selection as well as for phenological adaptions and heat tolerance traits in Swiss wheat breeding to adapt to future climate change and regional climatic differences. Different strategies of breeding adaptations and their trade-offs are discussed., European Journal of Agronomy, 131, ISSN:1161-0301, ISSN:1873-7331

Background PD-L1 IHC 22C3 pharmDx uses Tumor Proportion Score (TPS) and Combined Positive Score (CPS) scoring algorithms for the immunohistochemical (IHC) evaluation of PD-L1 protein expression in human cancer tissues; both algorithms include PD-L1 staining tumor cells (TC) in scoring and CPS also includes scoring of PD-L1 staining mononuclear inflammatory cells to aid in the identification of patients for treatment with pembrolizumab (KEYTRUDA®) using indication-specific diagnostic cut-offs. This study evaluated contribution of TC in determining specimen diagnostic status based on the CPS scoring algorithm by looking into four tumor indications approved for use with KEYTRUDA®: gastric or gastroesophageal junction (GEJ) adenocarcinoma (GC/GEJ), urothelial carcinoma (UC), head and neck squamous cell carcinoma (HNSCC), and esophageal squamous cell carcinoma (ESCC). Detection of specimens expressing PD-L1 is significantly dependent on the PD-L1 staining TC component. Methods A retrospective analysis was done looking at Dako’s internal tumor bank of the mentioned indications that were all stained with PD-L1 IHC 22C3 pharmDx and scored using the TPS, CPS and Quantitative Immune Cell Density (QID) methods described in figure 1. Statistical analysis encompassed looking at the scores generated that met the following criteria: CPS>0, TPS>0 and CPS≠TPS and then evaluating the percentage of those samples that changed from positive to negative diagnostic status upon removal of the TC component from the scoring. Results A noticeable downward trend was observed in all four indications in the total number of positives with the removal of the TC component. Table 1 aptly captures this by showing the number of specimens for each indication that had changed from positive to negative around each indication’s diagnostic cut-off(s). The three indications that showed the highest percentages of diagnostic status change were HNSCC (CPS ≥20) with a remarkable 83.3% (130) followed by UC (CPS ≥10) at 46.3% (57) and ESCC (CPS ≥10) at 36.6% (45) of the specimens reclassified as negative. Conclusions PD-L1 IHC 22C3 pharmDx (Dako, USA) stains both TC and immune cells. Removal of the PD-L1 staining TC from the CPS algorithm reduces the number of specimens scored as positive for each indication’s respective diagnostic cut-off(s). Scoring only IC reduces the number of specimens scored as positive for each indication’s respective cutoff.

Closely related populations often differ in resistance to a given parasite, as measured by infection success or failure. Yet, the immunological mechanisms of these evolved differences are rarely specified. Does resistance evolve via changes to the host’s ability to recognize that an infection exists, actuate an effective immune response, or attenuate that response? We tested whether each of these phases of the host response contributed to threespine sticklebacks’ recently evolved resistance to their tapeworm Schistocephalus solidus. While marine stickleback and some susceptible lake fish permit fast-growing tapeworms, other lake populations are resistant and suppress tapeworm growth via a fibrosis response. We subjected lab-raised fish from three populations (susceptible marine ‘ancestors’, a susceptible lake, a resistant lake), to a novel immune challenge (injection of: 1) a saline control, 2) alum, a generalized pro-inflammatory adjuvant that causes fibrosis, 3) a tapeworm protein extract, and 4) a combination of alum and tapeworm protein). All three populations were capable of a robust fibrosis response to the alum treatments (but not the saline control). Yet, only the resistant population exhibited a fibrosis response to the tapeworm protein alone. Thus, these populations differed in their ability to recognize the tapeworm but shared an intact fibrosis pathway. However, the resistant population also initiated fibrosis faster, and was able to attenuate fibrosis, unlike the susceptible populations slow but longer-lasting response to alum. As fibrosis has presumed pathological side-effects, this difference may reflect adaptions to mitigate costs of immunity in the resistant population. Broadly, our results confirm that parasite detection, activation speed, and immune attenuation simultaneously contribute to adaptations to parasite infection in natural populations.IMPACT SUMMARYDramatic variation in parasite resistance is common in nature, even to the same parasite, yet we are still working to understand the mechanisms of how such differences evolve. Many evolution studies focus on the broad outcomes of infection (infected or not) when studying this question, without specifying what part of the immune response has evolved. Here, we experimentally partition different sequential stages in the host immune response (recognition, actuation, attenuation), to evaluate which stage(s) underly the evolution of host resistance to infection. This study compares three populations of threespine stickleback that naturally differ in their exposure and their ability to resist infections of a freshwater tapeworm. These include a “resistant” lake population, a “susceptible” lake population, and an ancestral marine population that is rarely exposed to the tapeworm in nature, but is susceptible when exposed in the lab. The resistant population exhibits a fibrosis immune response to infection, which has previously been linked to suppressed tapeworm growth and viability. We injected different immune challenges directly into the site of infection (peritoneal cavity) and measured the subsequent fibrosis response through time. We found that all populations were capable of producing fibrosis in response to a general immune stimulant (alum). But, only the resistant population was able to recognize and respond to tapeworm protein alone. This population also responded faster than the others, within 24 hours, and attenuated its fibrosis by 90 days post-injections whereas the other populations exhibited a slower response that did not attenuate in the study time-frame. We concluded that variation in parasite recognition, an early phase in the host response, shapes the evolution of the initiation and resolution of the physical response to infection. Broadly, our results support that parasite detection mechanisms could play a key role in the rapid evolution of parasite resistance.

Journal of neurology 268(10), 3610-3625 (2021). doi:10.1007/s00415-020-09962-6, Published by Steinkopff, [Darmstadt]

From biofilms to whale pods, organisms have repeatedly converged on sociality as a strategy to improve individual fitness. Yet, it remains challenging to identify the most important drivers—and by extension, the evolutionary mechanisms—of sociality for particular species. Here, we present a conceptual framework, literature review, and model demonstrating that the direction and magnitude of the response of group size to sudden resource shifts provides a strong indication of the underlying drivers of sociality. We catalog six functionally distinct mechanisms related to the acquisition of resources, and we model these mechanisms’ effects on the survival of individuals foraging in groups. We find that whether, and to what degree, optimal group size increases, decreases, or remains constant when resource abundance declines depends strongly on the dominant mechanism. Existing empirical data support our model predictions, and we demonstrate how our framework can be used to predict the dominant social benefit for particular species. Together, our framework and results show that a single easily measurable characteristic, namely, group size under different resource abundances, can illuminate the potential drivers of sociality across the tree of life.

Introduction: Cardiac fibrosis contributes to the development of cardiovascular disease (CVD) and arrhythmia. Cardiac fibroblasts (CFs) are collagen-producing cells that regulate extracellular matrix (ECM) homeostasis. A complex signaling network has been defined linking environmental stress to changes in CF function and fibrosis. Signal Transducer and Activator of Transcription 3 (STAT3) has emerged as a critical integrator of pro-fibrotic signals in CFs downstream of several established signaling networks. Areas covered: This article provides an overview of STAT3 function in CFs and its involvement in coordinating a vast web of intracellular pro-fibrotic signaling molecules and transcription factors. We highlight recent work elucidating a critical role for the fibroblast cytoskeleton in maintaining spatial and temporal control of STAT3-related signaling . Finally, we discuss potential opportunities and obstacles for therapeutic targeting of STAT3 to modulate cardiac fibrosis and arrhythmias. Relevant publications on the topic were identified through Pubmed. Expert opinion: Therapeutic targeting of STAT3 for CVD and arrhythmias presents unique challenges and opportunities. Thus, it is critical to consider the multimodal and dynamic nature of STAT3 signaling. Going forward, it will be beneficial to consider ways to maintain balanced STAT3 function, rather than large-scale perturbations in STAT3 function.

Hereditary transthyretin amyloidosis is caused by pathogenic variants in the TTR gene and typically manifests, alongside cardiac and other organ dysfunctions, with a rapidly progressive sensorimotor and autonomic polyneuropathy (ATTRv-PN) leading to severe disability. While most prospective studies have focussed on endemic ATTRv-PN, real-world data on non-endemic, mostly late-onset ATTRv-PN are limited. This retrospective study investigated ATTRv-PN patients treated at the Amyloidosis Centre of Heidelberg University Hospital between November 1999 and July 2020. Clinical symptoms, survival, prognostic factors and efficacy of treatment with tafamidis were analysed. Neurologic outcome was assessed using the Coutinho ATTRv-PN stages, and the Peripheral Neuropathy Disability (PND) score. Of 346 subjects with genetic TTR variants, 168 patients had symptomatic ATTRv-PN with 32 different TTR variants identified. Of these, 81.6% had the late-onset type of ATTRv-PN. Within a mean follow-up period of 4.1 ± 2.8 years, 40.5% of patients died. Baseline plasma N-terminal prohormone of brain natriuretic peptide (NT-proBNP) ≥900 ng/l (HR 3.259 [1.421–7.476]; p = .005) was the main predictor of mortality in multivariable analysis. 64 patients were treated with tafamidis and presented for regular follow-up examinations. The therapeutic benefit of tafamidis was more pronounced when treatment was started early in ATTRv-PN stage 1 (PND scores II vs. I; HR 2.718 [1.258–5.873]; p = .011). In non-endemic, mostly late-onset ATTRv-PN, cardiac involvement assessed by NT-proBNP is a strong prognosticator for overall survival. Long-term treatment with tafamidis is safe and efficacious. Neurologic disease severity at the start of treatment is the main predictor for ATTRv-PN progression on tafamidis.

Der Klimawandel hat unsere Aufmerksamkeit vermehrt auf Veränderungen in unserer Umwelt gelenkt. Extreme Ereignisse wie Stürme, Überschwemmungen, Trocken- und Hitzeperioden haben direkte Auswirkungen auf unsere Lebenswelt und unseren Alltag. Weniger offensichtlich aber von grosser Tragweite sind die Einflüsse, die der Klimawandel auf die Jahreszeiten ausübt. Die Verschiebung von Blüh- und Ernteterminen, der Rückgang von Schneetagen in höheren Lagen und gar das Ausbleiben von Winterschnee im Mittelland zeigt vor allem mit Blick über mehrere Jahre und Jahrzehnte die gravierenden Folgen für die Umwelt aber auch für Landwirtschaft, Tourismus und Raumplanung. In phänologischen Beobachtungen – breit gefasst definiert als jahreszeitlich wiederkehrende Erscheinungsformen in der Umwelt – lassen sich sinnlich und alltäglich die Veränderungen erfahren, die der Klimawandel mit sich bringt. Darüber hinaus bilden die Überlieferungen dieser Beobachtungen von Generation zu Generation wichtige Brücken in Familiengeschichten und wichtigen Ortsbeschreibungen. Vor allem aber bringen uns die Beobachtungen näher, wie eng Mensch und Natur schon immer verbunden waren. Am Geographischen Institut der Universität Bern hat phänologische Forschung eine lange Tradition. Was in den späten 1960er-Jahren als Beobachtungsnetz und Datengrundlage für die Raum- und Agrareignungsplanung begann, mündete 1970 in die erste komplette Saison des Beobachtungsprogramms BernClim und bildet heute zusammen mit dem Datenschatz des Schweizer Phänologie Beobachtungsnetzes von MeteoSchweiz das Rückgrat für raum-zeitliche Beschreibungen seit Mitte des 20. Jahrhunderts. Beobachtungen seit dem Spätmittelalter bieten darüber hinaus die einmalige Möglichkeit, auch langfristige Veränderungen des Klimas zu zeigen. Zum 50-jährigen Bestehen des Beobachtungsprogramms BernClim entstand die vorliegende Broschüre. Über die Aktivitäten in Bern hinaus kommen auch Forschende zu Wort, die sich mit ebenso viel Herzblut und Ausdauer für die phänologischen Beobachtungen und Auswertungen an anderen Institutionen engagieren. Sie forschen mit Pflanzen- und Tierbeobachtungen, im Wald und auf dem Feld und ziehen Schlüsse aus Wetterdaten, Schnee- und Gewässermessungen. Die Grundlage für viele der hier präsentierten Jahreszeiten-Geschichten bilden Beobachtungen, die zum grossen Teil von Freiwilligen und oft über Jahre und Jahrzehnte gemacht wurden. Phänologie ist ohne dieses Engagement nicht möglich und bringt die Herausforderung mit sich, aus vielen individuellen Beobachtungen ein systematisches, grösseres Bild zu zeichnen. Die Publikation dieser Broschüre wurde durch die Sebastiana-Stiftung, das Oeschger-Zentrum für Klimaforschung (OCCR) und die Kommission für Phänologie und Saisonalität (KPS) der Akademie der Naturwissenschaften Schweiz (SCNAT) unterstützt. Gewidmet ist sie allen, die sich dafür einsetzen, das Zusammenspiel zwischen Mensch und Natur aus jahreszeitlicher Perspektive zu dokumentieren und besser zu verstehen., Geographica Bernensia, G97

Early generation breeding nurseries with thousands of genotypes in single-row plots are well suited to capitalize on high throughput phenotyping. Nevertheless, methods to monitor the intrinsically hard-to-phenotype early development of wheat are yet rare. We aimed to develop proxy measures for the rate of plant emergence, the number of tillers, and the beginning of stem elongation using drone-based imagery. We used RGB images (ground sampling distance of 3 mm pixel-1) acquired by repeated flights (≥ 2 flights per week) to quantify temporal changes of visible leaf area. To exploit the information contained in the multitude of viewing angles within the RGB images, we processed them to multiview ground cover images showing plant pixel fractions. Based on these images, we trained a support vector machine for the beginning of stem elongation (GS30). Using the GS30 as key point, we subsequently extracted plant and tiller counts using a watershed algorithm and growth modeling, respectively. Our results show that determination coefficients of predictions are moderate for plant count (R 2 = 0:52), but strong for tiller count (R 2 = 0:86) and GS30 (R 2 = 0:77). Heritabilities are superior to manual measurements for plant count and tiller count, but inferior for GS30 measurements. Increasing the selection intensity due to throughput may overcome this limitation. Multiview image traits can replace hand measurements with high efficiency (85-223%). We therefore conclude that multiview images have a high potential to become a standard tool in plant phenomics., Plant Phenomics, 2020

Parasite populations are never evenly distributed among the hosts they infect. Avian nest ectoparasites, such as mites, are no exception, as their distribution across the landscape is highly aggregated. It remains unclear if this pattern is driven by differences in transmission events alone, or if the environment that parasites inhabit after transmission also plays a role. Here, we experimentally examined the influence of the post-transmission microclimate, nest characteristics, and host condition on ectoparasite population growth in a bird-ectoparasite system. We infested barn swallow (Hirundo rustica erythrogaster) nests with a standardized number of Northern Fowl Mites (Ornithonyssus sylvarium) and analyzed both biotic (nestling mass, wing length, number of other arthropods present in the nest, and brood size) and abiotic (temperature, humidity, nest lining, nest dimensions, and substrate upon which the nest was built) predictors of mite population growth. Our results suggest that mite populations were most successful, in terms of growth, in nests with higher temperatures, lower humidity, few other arthropods, and hosts in good condition. We also found that nests built on wooden substrates support larger populations of mites than those constructed on metal or concrete. These findings lend insight into the factors that drive large-scale patterns of ectoparasite distributions., Graphical abstract Image 1, Highlights • Experimental manipulations of nest mites controls for initial transmission. • Tested the role of microclimate and host factors on mite population growth. • Mite populations grow larger in warmer, drier nests with high body condition hosts. • Nests on wooden substrates had more stable microclimate and larger mite populations. • Presence of other arthropods was negatively correlated with mite population growth.

Animals use morphological signals such as ornamental traits or weaponry to mediate social interactions, and the extent of signal trait elaboration is often positively associated with reproductive success. By demonstrating relationships between signal traits and fitness, researchers often make inferences about how behaviour operates to shape those outcomes. However, detailed information about fine-scale individual behaviour, and its physiological basis, can be difficult to obtain. Here we show that experimental manipulations to exaggerate a signal trait (plumage colour) and concomitant changes in testosterone and stress-induced corticosterone levels altered social interactivity between manipulated males and their social mates. On average, darkened males did not have higher levels of interactivity than unmanipulated males; however, males who experienced a greater shift in colour (pale to dark), a larger, positive change in testosterone levels, and a dampened stress-induced corticosterone response had a larger increase in the number of interactions with their social mate post-manipulation compared to pre-manipulation. This work provides new insights into the integration and real-time flexibility of multivariate phenotypes and direct evidence for the role of social interactions in pair bond maintenance. University of Colorado Innovative Seed Grant; National Science Foundation Postdoctoral Research Fellowship [DBI-1306059] We thank J. Burt for assistance with the tags, S. Turbek, and T. Albrecht for assistance in the field, and Colorado Horse Rescue for access to their barn. A University of Colorado Innovative Seed Grant provided funding for this research and I. Levin was supported by a National Science Foundation Postdoctoral Research Fellowship (DBI-1306059).

Accounting for historical demographic features is vital for many types of evolutionary inferences, including the estimation of divergence times between closely related populations. In barn swallow, Hirundo rustica, inferring historical population sizes and subspecies divergence times can shed light on the recent co-evolution of this species with humans. Pairwise sequentially Markovian coalescent uncovered population growth beginning on the order of one million years ago-which may reflect the radiation of the broader Hirundo genus-and a more recent population decline. Additionally, we used approximate Bayesian computation to evaluate hypotheses about recent timescale barn swallow demography, including population growth due to human commensalism, and a potential founder event associated with the onset of nesting on human structures. We found signal for a bottleneck event approximately 7,700 years ago, near the time that humans began building substantial structures, although there was considerable uncertainty associated with this estimate. Subspecies differentiation and subsequent growth occurred after the bottleneck in the best-supported model, an order of magnitude more recently than previous estimates in this system. We also compared results obtained from whole-genome sequencing versus reduced representation sequencing, finding many similar results despite substantial allelic dropout in the reduced representation data, which may have affected estimates of some parameters. This study presents the first genetic evidence of a potential barn swallow founder effect and subspecies divergence coinciding with the Holocene, which is an important step in analysing the biogeographical history of a well-known human commensal species.

Nest-switching is an important breeding strategy for multiple-brooded bird species. When deciding whether or not to switch nests for subsequent breeding attempts, pairs must weigh the costs and benefits of various factors related to the number of fledglings of the first breeding attempt, the likelihood of nest predation, and qualities of the nest environment, such as nest ectoparasites and the age of the nest. In this study, we analyzed the predictors and consequences of nest-switching behavior at 6 breeding sites of North American Barn Swallows (Hirundo rustica erythrogaster), where 60% of pairs that raised 2 broods within a season switched nests for a second breeding attempt. Pairs often reused existing (old) nests constructed during previous years, and pairs that settled in old nests for their first breeding attempt were the most likely to switch nests for a second breeding attempt. Contrary to previous studies, nest predation and nest ectoparasitism had no influence on whether or not pairs sw...

Women with renal disease have menstrual and gonadal dysfunction manifesting as hormonal imbalance. Anti-Müllerian hormone (AMH) is a potential measure of ovarian reserve. We examined circulating AMH concentrations in young women with renal failure, determined associations with clinical characteristics, and compared AMH with age-matched healthy individuals. AMH was measured in 77 women: 26 had chronic kidney disease (CKD), 26 were on haemodialysis (HD), and 25 had a kidney transplant. Random AMH levels were highest in women on HD [HD 2.9 (1.1–5.2), CKD 1.6 (0.7–2.2), transplant 1.5 (1.0–4.2) ng/ml]. On multiple linear regression, AMH was 53% higher [95% CI 0.20–0.98, P = 0.002] in women on HD and decreased by 20% per 5-year increase in age (P < 0.001). AMH was 43% lower in women with renal failure compared with 600 age-matched controls [1.7 (0.9–3.8) versus 3.0 (1.9–5.0) ng/ml, P < 0.001]; however, we found no difference in AMH between those on HD and healthy individuals [2.9 (1.1–5.2) versus 3.0 (1.9–5.0) ng/ml]. AMH may be a useful biomarker in female renal patients with non-dialysis dependent renal disease pursuing pregnancy. In contrast, AMH levels are higher in HD but unlikely to reflect ovarian reserve.

The ability to avoid dehydration is a drought resistance mechanism becoming increasingly more important even in temperate regions. In wheat, dehydration avoidance can be associated with a maintained canopy cooling during dry periods. However, in an average year under temperate conditions, drought periods are rather short which makes it difficult to routinely screen for drought avoidance using canopy temperature (CT). Furthermore, confounding factors such as differences in height, shoot biomass, canopy structure and phenology complicate the interpretation of differences in CT. We aimed to use temporal trends in CT and canopy greenness during short-term drought and heat events in the early grain filling phase to circumvent these problems. During this phase, evaporative demand is high and phenology-driven senescence has yet little effect on CT. Diverse sets of 354 and 71 wheat genotypes where grown in the field phenotyping platform of ETH Zurich in 2018 and 2019, respectively. CT was repeatedly measured during early grain filling by means of drone-based imaging. The temporal trends in CT during early grain filling showed a moderate to high within-year heritability (h2 = 0.35 and h2 = 0.88 in 2018 and 2019, respectively). These trends were largely independent of confounding factors when compared to single time point measurements and likely represent genotype-specific reactions to decreasing water availability more directly than absolute CT. CT trends were also largely independent of the temporal trends in stay-green indices. Therefore, we used a combination of time-resolved CT and stay-green trends to identify genotypes combining both traits. Significant differences were observed in the combined time-resolved index among three replicated check varieties. We therefore propose to use the combined time-resolved index to identify genotypes with improved drought avoidance and functional stay green., Field Crops Research, 274, ISSN:0378-4290, ISSN:1872-6852

Fibrosis is a pronounced feature of heart disease and the result of dysregulated activation of resident cardiac fibroblasts (CFs). Recent work identified stress-induced degradation of the cytoskeletal protein βIV-spectrin as an important step in CF activation and cardiac fibrosis. Furthermore, loss of βIV-spectrin was found to depend on Ca2+/calmodulin-dependent kinase II (CaMKII). Therefore, we sought to determine the mechanism for CaMKII-dependent regulation of βIV-spectrin and CF activity. Computational screening and MS revealed a critical serine residue (S2250 in mouse and S2254 in human) in βIV-spectrin phosphorylated by CaMKII. Disruption of βIV-spectrin/CaMKII interaction or alanine substitution of βIV-spectrin Ser2250 (βIV-S2254A) prevented CaMKII-induced degradation, whereas a phosphomimetic construct (βIV-spectrin with glutamic acid substitution at serine 2254 [βIV-S2254E]) showed accelerated degradation in the absence of CaMKII. To assess the physiological significance of this phosphorylation event, we expressed exogenous βIV-S2254A and βIV-S2254E constructs in βIV-spectrin-deficient CFs, which have increased proliferation and fibrotic gene expression compared with WT CFs. βIV-S2254A but not βIV-S2254E normalized CF proliferation, gene expression, and contractility. Pathophysiological targeting of βIV-spectrin phosphorylation and subsequent degradation was identified in CFs activated with the profibrotic ligand angiotensin II, resulting in increased proliferation and signal transducer and activation of transcription 3 nuclear accumulation. While therapeutic delivery of exogenous WT βIV-spectrin partially reversed these trends, βIV-S2254A completely negated increased CF proliferation and signal transducer and activation of transcription 3 translocation. Moreover, we observed βIV-spectrin phosphorylation and associated loss in total protein within human heart tissue following heart failure. Together, these data illustrate a considerable role for the βIV-spectrin/CaMKII interaction in activating profibrotic signaling.

Atrial fibrillation (AF) affects more than three million people per year in the United States and is associated with high morbidity and mortality. Both electrical and structural remodeling contribute to AF, but the molecular pathways underlying AF pathogenesis are not well understood. Recently, a role for Ca2+/calmodulin-dependent protein kinase II (CaMKII) in the regulation of persistent “late” Na+ current ( INa,L) has been identified. Although INa,L inhibition is emerging as a potential antiarrhythmic strategy in patients with AF, little is known about the mechanism linking INa,L to atrial arrhythmogenesis. A computational approach was used to test the hypothesis that increased CaMKII-activated INa,L in atrial myocytes disrupts Ca2+ homeostasis, promoting arrhythmogenic afterdepolarizations. Dynamic CaMKII activity and regulation of multiple downstream targets [ INa,L, L-type Ca2+ current, phospholamban, and the ryanodine receptor sarcoplasmic reticulum Ca2+-release channel (RyR2)] were incorporated into an existing well-validated computational model of the human atrial action potential. Model simulations showed that constitutive CaMKII-dependent phosphorylation of Nav1.5 and the subsequent increase in INa,L effectively disrupt intracellular atrial myocyte ion homeostasis and CaMKII signaling. Specifically, increased INa,L promotes intracellular Ca2+ overload via forward-mode Na+/Ca2+ exchange activity, which greatly increases RyR2 open probability beyond that observed for CaMKII-dependent phosphorylation of RyR2 alone. Increased INa,L promotes atrial myocyte repolarization defects (afterdepolarizations and alternans) in the setting of acute β-adrenergic stimulation. We anticipate that our modeling efforts will help identify new mechanisms for atrial NaV1.5 regulation with direct relevance for human AF. NEW & NOTEWORTHY Here, we present a novel computational model to study the effects of late Na+ current ( INa,L) in human atrial myocytes. Simulations predict that INa,L promotes intracellular accumulation of Ca2+, with subsequent dysregulation of Ca2+/calmodulin-dependent protein kinase II (CaMKII) signaling and ryanodine receptor 2-mediated Ca2+ release. Although INa,L plays a small role in regulating atrial myocyte excitability at baseline, CaMKII-dependent enhancement of the current promoted arrhythmogenic dynamics. Listen to this article’s corresponding podcast at http://ajpheart.podbean.com/e/camkii-dependent-regulation-of-atrial-late-sodium-current-and-excitability/ .

Background: Biochemical DNA modification resembles a crucial regulatory layer among genetic information, environmental factors, and the transcriptome. To identify epigenetic susceptibility regions and novel biomarkers linked to myocardial dysfunction and heart failure, we performed the first multi-omics study in myocardial tissue and blood of patients with dilated cardiomyopathy and controls. Methods: Infinium human methylation 450 was used for high-density epigenome-wide mapping of DNA methylation in left-ventricular biopsies and whole peripheral blood of living probands. RNA deep sequencing was performed on the same samples in parallel. Whole-genome sequencing of all patients allowed exclusion of promiscuous genotype-induced methylation calls. Results: In the screening stage, we detected 59 epigenetic loci that are significantly associated with dilated cardiomyopathy (false discovery corrected P ≤0.05), with 3 of them reaching epigenome-wide significance at P ≤5×10 −8 . Twenty-seven (46%) of these loci could be replicated in independent cohorts, underlining the role of epigenetic regulation of key cardiac transcription regulators. Using a staged multi-omics study design, we link a subset of 517 epigenetic loci with dilated cardiomyopathy and cardiac gene expression. Furthermore, we identified distinct epigenetic methylation patterns that are conserved across tissues, rendering these CpGs novel epigenetic biomarkers for heart failure. Conclusions: The present study provides to our knowledge the first epigenome-wide association study in living patients with heart failure using a multi-omics approach.

Vaquero-Alba and colleagues published a study in The Auk: Ornithological Advances comparing objective color measurements of plumage taken in the field directly on a bird's body to those taken in the lab on collected feathers arranged to emulate the appearance of a bird's natural plumage. Although the field measures of plumage color were less repeatable than lab measures, the authors concluded that measurements taken in the field were more representative of a bird's “true color.” Accordingly, they recommend that researchers should bring spectrophotometers into the field to measure color on live birds. We question the assumption that their field measurements represent true color and highlight concerns regarding their experimental design and methodology. Because they did not measure color of live birds in the lab or the color of plucked feathers in the field, they cannot directly test whether the assessment of color in the field on a live bird is superior. Also, rather than assume field measures are...

BACKGROUND Whole-grain wheat flour is facing the quality challenge of lipid rancidity, which decreases its nutritional, sensory, and technological properties. One of the major causes of lipid rancidity is endogenous esterases and lipases. This study evaluated 66 European wheat varieties grown at a single site over three years (2014, 2015, and 2016). RESULTS The 66 wheat varieties showed up to threefold variance on esterase and lipase activities. Wheat varieties that are suitable for lipid-stable whole-grain products ('Julius', 'Lona', and 'Banquet') were selected according to their consistently low esterase and lipase activities. The 3-year mean-based broad-sense heritability of esterase and lipase was 0.75 and 0.44 respectively. CONCLUSIONS The findings indicate great genetic dependence of both esterase and lipase activities in wheat. The moderate to high heritability brings a new prospect of breeding selection of low-lipase-activity wheat for stable whole-grain products. This result will improve the use of wheat as raw material, benefit cultivation selection, and provide consumers with better quality products. © 2020 Society of Chemical Industry.