Phases of cortical actomyosin dynamics coupled to the neuroblast polarity cycle The Par complex dynamically polarizes to the apical cortex of asymmetrically dividing Drosophila neuroblasts where it directs fate determinant segregation. Previously we showed that apically directed cortical movements that polarize the Par complex require F-actin (Oon and Prehoda, 2019). Here we report the discovery of cortical actomyosin dynamics that begin in interphase when the Par complex is cytoplasmic but ultimately become tightly coupled to cortical Par dynamics. Interphase cortical actomyosin dynamics are unoriented and pulsatile but rapidly become sustained and apically-directed in early mitosis when the Par protein aPKC accumulates on the cortex. Apical actomyosin flows drive the coalescence of aPKC into an apical cap that is depolarized in anaphase when the flow reverses direction. Together with the previously characterized role of anaphase flows in specifying daughter cell size asymmetry, our results indicate that multiple phases of cortical actomyosin dynamics regulate asymmetric cell division.
Plasma membrane damage limits replicative lifespan in yeast and induces premature senescence in human fibroblasts Plasma membrane damage (PMD) occurs in all cell types due to environmental perturbation and cell-autonomous activities. However, cellular outcomes of PMD remain largely unknown except for recovery or death. Here, using yeast and human fibroblasts, we show that cellular senescence, irreversible cell cycle arrest contributing to organismal aging, is the third outcome of PMD. To identify the genes essential for PMD response, we performed a systematic yeast genome-wide screen. The screen identified 48 genes. The top hits in the screen are the endosomal sorting complexes required for transport (ESCRT) genes. Strikingly, the replicative lifespan regulator genes are enriched in our 48 hits, and indeed, PMD limits the replicative lifespan in budding yeast; the ESCRT activator AAA-ATPase VPS4-overexpression extends it. In normal human fibroblasts, PMD induces premature senescence via p53. Our study demonstrates that PMD limits replicative lifespan in two different eukaryotic cell types and highlights an underappreciated but ubiquitous senescent cell subtype, namely PMD-dependent senescent cells.
Single amino-acid mutation in a Drosophila melanogaster ribosomal protein: an insight in uL11 transcriptional activity The ribosomal protein uL11 is located at the basis of the ribosome P-stalk and plays a paramount role in translational efficiency. In addition, no mutant for uL11 is available suggesting that this gene is haplo-insufficient as many other Ribosomal Protein Genes (RPGs). We have previously shown that overexpression of Drosophila melanogaster uL11 enhances the transcription of many RPGs and Ribosomal Biogenesis genes (RiBis) suggesting that uL11 might globally regulate the level of translation through its transcriptional activity. Moreover, uL11 trimethylated on lysine 3 (uL11K3me3) interacts with the chromodomain of the Enhancer of Polycomb and Trithorax Corto, and both proteins co- localize with RNA Polymerase II at many sites on polytene chromosomes. These data have led to the hypothesis that the N-terminal end of uL11, and more particularly the trimethylation of lysine 3, supports the extra-ribosomal activity of uL11 in transcription. To address this question, we mutated the lysine 3 codon using a CRISPR/Cas9 strategy and obtained several lysine 3 mutants. We describe here the first mutants of D. melanogaster uL11. Unexpectedly, the uL11K3A mutant, in which the lysine 3 codon is replaced by an alanine, displays a genuine Minute phenotype known to be characteristic of RPG deletions (longer development, low fertility, high lethality, thin and short bristles) whereas the uL11K3Y mutant, in which the lysine 3 codon is replaced by a tyrosine, is unaffected. In agreement, the rate of translation decreases in uL11K3A but not in uL11K3Y. Co-immunoprecipitation experiments show that the interaction between uL11 and the Corto chromodomain is impaired by both mutations. RNA- seq analysis from wing imaginal discs shows enrichment in the GO categories "glutathione metabolism" for up-regulated genes in both uL11K3A and uL11K3Y mutants and "regulation of transcription" for down-regulated genes in uL11K3A only. Analysis of the cis-regulatory sequences of these genes suggests that uL11 might regulate transcription of target genes in concert with the couple of transcription factors Mad/Med that mediate response to the Bone Morphogenetic Protein (BMP) signaling pathway. Transcription disrupts DNA-scaffolded bacteriophage repressor complexes DNA can act as a scaffold for the cooperative binding of protein oligomers. For example, the phage 186 CI repressor forms a wheel of seven dimers wrapped in DNA carrying specific binding sites, while the phage {lambda} CI repressor binds in units of dimers to two well-separated sets of operators, forming a DNA loop. Atomic force microscopy was used to measure transcription elongation by E. coli RNA polymerase through these protein complexes. 186 CI or {lambda} CI bound along unlooped DNA negligibly interfered with transcription by RNAP. More complex topologies induced by scaffolded, cooperatively bound repressor oligomers did not form significantly better roadblocks to transcription. Thus, despite binding with rather high affinity, these repressors are not effective roadblocks to transcription. Microvesicles Transfer Mitochondria and Increase Mitochondrial Function in Brain Endothelial Cells We have demonstrated, for the first time that microvesicles, a sub-type of extracellular vesicles (EVs) derived from hCMEC/D3: a human brain endothelial cell (BEC) line transfer polarized mitochondria to recipient BECs in culture and to neurons in mice acute brain cortical and hippocampal slices. This mitochondrial transfer increased ATP levels by 100 to 200-fold (relative to untreated cells) in the recipient BECs exposed to oxygen-glucose deprivation, an in vitro model of cerebral ischemia. We have also demonstrated that transfer of microvesicles, the larger EV fraction, but not exosomes resulted in increased mitochondrial function in hypoxic endothelial cultures. Gene ontology and pathway enrichment analysis of EVs revealed a very high association to glycolysis-related processes. In comparison to heterotypic macrophage- derived EVs, BEC-derived EVs demonstrated a greater selectivity to transfer mitochondria and increase endothelial cell survival under ischemic conditions.

HighlightsO_LIMicrovesicles transfer mitochondria to endothelial cells and brain slice neurons
C_LIO_LIMitochondrial transfer increased ATP in ischemic brain endothelial cells (BECs)
C_LIO_LITransfer of microvesicles increased mitochondrial function in hypoxic BECs
C_LIO_LITransfer of exosomes did not affect mitochondrial function in hypoxic BECs
C_LIO_LIHomotypic BEC-derived EVs result in greater ATP levels in the recipient BECs
C_LI Balancing sensitivity and specificity in preclinical research The success of scientific discovery in preclinical drug development is based on the different roles of exploration and confirmation. Exploration requires sensitivity while confirmation needs to be specific. Based on published empirical data, we simulated two preclinical research trajectories to examine their effectiveness to detect true effects while keeping animal numbers low. We compare a conventional p-value based approach with a method based on an a priori determined smallest effect size of interest (SESOI). Using a SESOI increases transition rates from exploration to confirmation and results in higher detection rates across the trajectory. Specificity in the SESOI trajectory increases if the prior probability of true effects is low. We conclude that employing a SESOI is superior to a p-value based approach. Based on our findings, we propose a reconsideration of planning and conducting preclinical experiments, especially when the prior probability of true hypotheses is low. Synthetic periphyton as a model system to understand species dynamics in complex microbial freshwater communities Phototrophic biofilms, also known as periphyton, are microbial freshwater communities that drive crucial ecological processes in streams and lakes. Gaining a deep mechanistic understanding of the biological processes occurring in natural periphyton remains challenging due to the high complexity and variability of such communities. To address this challenge, we rationally developed a workflow to construct a synthetic community by co-culturing 26 phototrophic species (i.e., diatoms, green algae and cyanobacteria) that were inoculated in a successional sequence to create a periphytic biofilm on glass slides. We show that this community is diverse, stable and highly reproducible in terms of microbial composition, function and 3D spatial structure of the biofilm. We also demonstrate the ability to monitor microbial dynamics at the single species level during periphyton development and how their abundances are impacted by stressors such as increased temperature and a herbicide, singly and in combination. Overall, such a synthetic periphyton, grown under controlled conditions, can be used as a model system for theory testing through targeted manipulation. Cellular feedback to organotelluranes displays modulation of antioxidant proteins gene expression Organotelluranes RT3 and RT4 are thiol reagents that induce mitochondrial transition pore (MTP) opening in a sensitive and insensitive manner to cyclosporin A. Although RT3 and RT4 promote glutathione depletion, paradoxically, they are also an efficient antioxidant for membrane lipids. These compounds antagonistic effects elicited the challenging question of how the gene expression of antioxidant enzymes would respond to treatment with these compounds. The influence of RT3 and RT4 on antioxidant enzyme expression was investigated in cultured aortic smooth muscle cells (ASMC). RT3 and RT4 promoted disruption of ionic calcium homeostasis, mitochondrial transmembrane potential ({Delta}{Psi}), and cell death in a dose-dependent manner. The cell death mechanisms responded qualitatively to the increase of the organotellurane concentration and changed from apoptosis to necrosis. RT3 and RT4 increased the expression of thioredoxin significantly. RT3 also increased the expression of glutaredoxin and glutathione peroxidase, slightly the catalase expression without significant effects on SOD expression. The results are consistent with GSH and protein thiol depletion and discussed based on the cell toxicity mechanism exhibited by these compounds.



O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=150 SRC="FIGDIR/small/425411v4_ufig1.gif" ALT="Figure 1">
View larger version (73K):
org.highwire.dtl.DTLVardef@52bb82org.highwire.dtl.DTLVardef@19476d9org.highwire.dtl.DTLVardef@6aa99org.highwire.dtl.DTLVardef@1aa605e_HPS_FORMAT_FIGEXP  M_FIG C_FIG Efficient and Economical Targeted Insertion in Plant Genomes via Protoplast Regeneration Versatile genome editing can be facilitated by the insertion of DNA sequences into specific locations. Current protocols involving clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) proteins rely on low efficiency homology-directed repair or non-homologous end joining with modified double-stranded DNA oligonucleotides as donors. Our simple protocol eliminates the need for expensive equipment, chemical and enzymatic donor DNA modification or plasmid construction by using polyethylene glycol-calcium to deliver non-modified single-stranded DNA oligonucleotides and CRISPR-Cas9 ribonucleoprotein into protoplasts. Plants regenerated via edited protoplasts achieved targeted insertion frequencies of up to 50.0% in Nicotiana benthamiana and 13.6% in rapid cycling Brassica oleracea without antibiotic selection. Using a 60-nt donor containing 27 nt in each homologous arm, 6 of 22 regenerated N. benthamiana plants showed targeted insertions, and one contained a precise insertion of a 6-bp HindIII site. The inserted sequences were transmitted to the next generation and invite the possibility of future exploration of versatile genome editing by targeted DNA insertion in plants. Single cell profiling of functionally cured Chronic Hepatitis B patients reveals the emergence of activated innate and an altered adaptive immune response in the intra-hepatic environment Hepatitis B surface antigen (HBsAg) loss or functional cure (FC), is considered the desirable therapeutic outcome for chronic hepatitis B (CHB) patients. However, the immuno-pathological biomarkers and underlying mechanisms remain unclear. Here we present a comprehensive single cell-transcriptomic atlas together with immune-phenotyping of disease-associated cell states (DACS) isolated from intra-hepatic tissue and matched PBMCs of either CHB or FC patients. We find that the intra-hepatic environment displays specific cell identities and molecular signatures that are distinct from PBMCs. FC is associated with emergence of an altered adaptive immune response marked by CD4 cytotoxic T lymphocytes (CD4-CTLs), and an activated innate response represented by liver-resident natural killer (LR-NK) cells. Overall, these findings provide novel insights into immuno-pathological cell states associated with FC that could serve as prognostic biomarkers. State-dependent network interactions differentially gate sensory input at the motor and command neuron level in Caenorhabditis elegans Neural responses are influenced by both external stimuli and internal network states. While network states have been linked to behavioral and stimulus states, little is known about how sensory inputs are filtered by whole-brain activity to affect motor and command neurons. Here, we recorded whole-brain activity of Caenorhabditis elegans experiencing bacterial food stimuli, and modeled how sensory inputs affect motor and command neurons in a network state-dependent manner. First, we classified active neurons into six functional clusters: two sensory neuron clusters (ON, OFF), and four motor/command neuron clusters (AVA, RME, SMDD, SMDV). Using encoding models, we found that ON and OFF sensory neurons that respond to onset and removal of bacteria, respectively, employ different adaptation strategies. Next, we used decoding models to show that bacterial onset and removal differentially drive AVA and RME cluster activity. To explore state-dependent effects on AVA and RME clusters, we developed a model that identified network states and fitted submodels for each state to predict how each of the six functional clusters drive AVA and RME cluster activity. We also identified network states in which AVA and RME clusters were either largely unperturbed by or receptive to bacterial sensory input. Furthermore, this model allowed us to disentangle the state-dependent contributions of stimulus timescales and bacterial content to neural activity. Collectively, we present an interpretable approach for modeling network dynamics that goes beyond implication of neurons in particular states, and moves toward explicitly dissecting how neural populations work together to produce state dependence.

Significance StatementA major function of the brain is to transform sensory information into behavior. As the first receiver of sensory input, sensory neuron activity is often most correlated with stimulus features. However, this high-fidelity representation of sensory input becomes diluted as it travels to downstream neurons, where sensory information is integrated with network activity. By the time sensory information reaches motor neurons, it is often difficult to dissociate the influence of sensory input from the influence of network activity. Here, we describe a method that is fully interpretable such that we can show how neural populations on a whole-brain scale interact to produce network states. From there, we can attribute motor neuron activity to network history and sensory input. Water levels primarily drive variation in photosynthesis and nutrient use of scrub Red Mangroves in the southeastern Florida Everglades We investigated how mangrove-island micro-elevation (i.e., habitat: center vs. edge) affects tree physiology in a scrub mangrove forest of the southeastern Everglades. We measured leaf gas exchange rates of scrub Rhizophora mangle L. trees monthly during 2019, hypothesizing that CO2 assimilation (Anet) and stomatal conductance (gsw) would decline with increasing water levels and salinity, expecting more-considerable differences at mangrove-island edges than centers, where physiological stress is greatest. Water levels varied between 0 and 60 cm from the soil surface, rising during the wet season (May-October) relative to the dry season (November-April). Porewater salinity ranged from 15 to 30 ppt, being higher at mangrove-island edges than centers. Anet maximized at 15.1 {micro}mol m-2 s-1, and gsw was typically <0.2 mol m-2 s-1, both of which were greater in the dry than the wet season and greater at island centers than edges, with seasonal variability being roughly equal to variation between habitats. After accounting for season and habitat, water level positively affected Anet in both seasons but did not affect gsw. Our findings suggest that inundation stress (i.e., water level) is the primary driver of variation in leaf gas exchange rates of scrub mangroves in the Florida Everglades, while also constraining Anet more than gsw. The interaction between inundation stress due to permanent flooding and habitat varies with season as physiological stress is alleviated at higher-elevation mangrove-island center habitats during the dry season. Freshwater inflows during the wet season, increase water levels and inundation stress at higher-elevation mangrove-island centers, but also potentially alleviate salt and sulfide stress in soils. Thus, habitat heterogeneity leads to differences in nutrient and water acquisition and use between trees growing in island centers versus edges, creating distinct physiological controls on photosynthesis, which likely affect carbon flux dynamics of scrub mangroves in the Everglades. Effect of Adolescent Stress on Adult Morphine-Induced Behavioral Sensitization is Dependent Upon Genetic Background 1Deaths related to opioid use have skyrocketed in the United States, leading to a public health epidemic. Research has shown that both biological (genes) and environmental (stress) precursors are linked to opioid use. In particular, stress during adolescence - a critical period of frontal lobe development - influences the likelihood of abusing drugs. However, little is known about the biological mechanisms through which adolescent stress leads to long-term risk of opioid use, or whether genetic background moderates this response. Male and female C57BL/6J and BALB/cJ mice were exposed to chronic variable social stress (CVSS) or control conditions throughout adolescence and then tested for morphine locomotor sensitization or morphine consumption in adulthood. To examine possible mechanisms that underlie stress-induced changes in morphine behaviors, we assessed physiological changes in response to acute stress exposure and prefrontal cortex miRNA gene expression. Adolescent stress did not influence morphine sensitization or consumption in BALB/cJ animals, and there was limited evidence of stress effects in female C57BL/6J mice. In contrast, male C57BL/6J mice exposed to adolescent CVSS had blunted morphine sensitization compared to control animals; no differences were observed in the acute locomotor response to morphine administration or morphine consumption. Physiologically, C57BL/6J mice exposed to CVSS had an attenuated corticosterone recovery following an acute stressor and downregulation of twelve miRNA in the prefrontal cortex compared to control mice. The specificity of the effects for C57BL/6J versus BALB/cJ mice provides evidence of a gene by environmental interaction influencing opioid behaviors. Long-term differences in stress reactivity or miRNA expression suggests two possible biological mechanisms to evaluate in future research. An automated workflow for multiplexed single-cell proteomics sample preparation at unprecedented sensitivity. The analysis of single-cell proteomes has recently become a viable complement to transcriptomics and genomics studies. Proteins are the primary driver of cellular functionality and mRNA levels are often an unreliable proxy of such. Therefore, the global analysis of the proteome is essential to study cellular identities. Multiplexed and label-free mass spectrometry-based approaches with single-cell resolution have lately attributed surprising heterogeneity to presumed homogenous cell populations. Even though specialized experimental designs and instrumentation have demonstrated remarkable advances, the efficient sample preparation of single cells still lag. Here, we introduce the proteoCHIP, a universal option for single-cell proteomics sample preparation at surprising sensitivity and throughput. The automated processing using a commercial system combining single-cell isolation and picoliter dispensing, the cellenONE(R), reduces final sample volumes to low nanoliters submerged in a hexadecane layer simultaneously eliminating error-prone manual sample handling and overcoming evaporation. The specialized proteoCHIP design allows direct injection of single cells via a standard autosampler resulting in around 1,500 protein groups per analytical run at remarkable reporter ion signal to noise while reducing or eliminating the carrier proteome. We identified close to 2,600 proteins across 170 multiplexed single cells from two highly similar human cell types. This dedicated loss-less workflow allows distinguishing in vitro co-differentiated cell types of self-organizing cardiac organoids based on indicative markers across 150 single cells. In-depth characterization revealed enhanced cellular motility of cardiac endothelial cells and sarcomere organization in cardiomyocytes. Our versatile and automated sample preparation has not only proven to be easily adaptable but is also sufficiently sensitive to drive biological applications of single-cell proteomics. Identification of age and ethnicity specific gene expression biomarkers for immune aging Human immune system functions over an entire lifetime, yet how and why the immune system becomes less effective with age are not well understood. Here, we characterize peripheral blood mononuclear cells transcriptome from 172 healthy adults with 21~90 years of age using RNA-seq and the weighted gene correlation network analyses (WGCNA). These data reveal a set of insightful gene expression modules and representative gene biomarkers for human immune system aging from Asian and Caucasian ancestry, respectively. Among them, the aging-specific modules show an age-related gene expression variation spike around early-seventies. In addition, it is not known whether Asian and Caucasian immune systems go through similar gene expression changes throughout their lifespan, and to what extent these aging-associated changes are shared among ethnicities. We find the top hub genes including NUDT7, CLPB, OXNAD1 and MLLT3 are shared between Asian and Caucasian aging related modules and further validated in human PBMCs from different age groups. Overall, the impact of age and race on transcriptional variation elucidated from this study provide insights into the transcriptional driver of immune aging. Diverse stem-chondrichthyan oral structures and evidence for an independently acquired acanthodid dentition The teeth of sharks famously form a series of transversely-organised files with conveyor-belt replacement that are borne directly on the jaw cartilages, in contrast to the dermal plate-borne dentition of bony fishes that undergoes site-specific replacement. A major obstacle in understanding how this system evolved is the poorly understood relationships of the earliest chondrichthyans and the profusion of morphologically and terminologically diverse bones, cartilages, splints and whorls that they possess. Here we use tomographic methods to investigate mandibular structures in several early branching  acanthodian-grade stem-chondrichthyans. We show that the dentigerous jaw bones of disparate genera of ischnacanthids are united by a common construction, being growing bones with non-shedding dentition. Mandibular splints, which support the ventro-lateral edge of the Meckels cartilage in some taxa, are formed from dermal bone and may be an acanthodid synapomorphy. We demonstrate that the teeth of Acanthodopsis are borne directly on the mandibular cartilage and that this taxon is deeply nested within an edentulous radiation, representing an unexpected independent origin of teeth. Many or even all of the range of unusual oral structures may be apomorphic, but they should nonetheless be considered when building hypotheses of tooth and jaw evolution, both in chondrichthyans and more broadly. A nickase Cas9 gene-drive system promotes super-Mendelian inheritance in Drosophila CRISPR-based gene-drive systems have been proposed for managing insect populations, including disease-transmitting mosquitoes, due to their ability to bias their inheritance towards super-Mendelian rates (>50%). Current technologies employ a Cas9 that introduces DNA double-strand breaks into the opposing wildtype allele to replace it with a copy of the gene drive allele via DNA homology-directed repair. Yet, the use of different Cas9s versions is unexplored, and alternative approaches could increase the available toolkit for gene-drive designs. Here, we report a novel gene-drive approach that relies on Cas9 nickases that generate staggered paired nicks in DNA to propagate the engineered gene-drive cassette. We show that generating 5 overhangs in the system yields efficient allelic conversion. The nickase gene-drive arrangement produces large, stereotyped deletions that are advantageous for targeting essential genes. Indeed, our nickase approach should expand the repertoire for gene-drive designs aimed at applications in mosquitoes and beyond. Fatal COVID-19 and non-COVID-19 Acute Respiratory Distress Syndrome is Associated with Incomplete Alveolar Type 1 Epithelial Cell Differentiation from the Transitional State Without Fibrosis ARDS due to COVID-19 and other etiologies results from injury to the alveolar epithelial cell (AEC) barrier resulting in noncardiogenic pulmonary edema, which causes acute respiratory failure; clinical recovery requires epithelial regeneration. During physiologic regeneration in mice, AEC2s proliferate, exit the cell cycle, and transiently assume a transitional state before differentiating into AEC1s; persistence of the transitional state is associated with pulmonary fibrosis in humans. It is unknown whether transitional cells emerge and differentiate into AEC1s without fibrosis in human ARDS and why transitional cells differentiate into AEC1s during physiologic regeneration but persist in fibrosis. We hypothesized that incomplete but ongoing AEC1 differentiation from transitional cells without fibrosis may underlie persistent barrier permeability and fatal acute respiratory failure in ARDS. Immunostaining of postmortem ARDS lungs revealed abundant transitional cells in organized monolayers on alveolar septa without fibrosis. They were typically cuboidal or partially spread, sometimes flat, and occasionally expressed AEC1 markers. Immunostaining and/or interrogation of scRNAseq datasets revealed that transitional cells in mouse models of physiologic regeneration, ARDS, and fibrosis express markers of cell cycle exit but only in fibrosis express a specific senescence marker. Thus, in severe, fatal early ARDS, AEC1 differentiation from transitional cells is incomplete, underlying persistent barrier permeability and respiratory failure, but ongoing without fibrosis; senescence of transitional cells may be associated with pulmonary fibrosis. Accommodating site variation in neuroimaging data using normative and hierarchical Bayesian models AO_SCPLOWBSTRACTC_SCPLOWThe potential of normative modeling to make individualized predictions from neuroimaging data has enabled inferences that go beyond the case-control approach. However, site effects are often confounded with variables of interest in a complex manner and can bias estimates of normative models, which has impeded the application of normative models to large multi-site neuroimaging data sets. In this study, we suggest accommodating for these site effects by including them as random effects in a hierarchical Bayesian model. We compared the performance of a linear and a non-linear hierarchical Bayesian model in modeling the effect of age on cortical thickness. We used data of 570 healthy individuals from the ABIDE (autism brain imaging data exchange) data set in our experiments. In addition, we used data from individuals with autism to test whether our models are able to retain clinically useful information while removing site effects. We compared the proposed single stage hierarchical Bayesian method to several harmonization techniques commonly used to deal with additive and multiplicative site effects using a two stage regression, including regressing out site and harmonizing for site with ComBat, both with and without explicitly preserving variance related to age and sex as biological variation of interest. In addition, we made predictions from raw data, in which site has not been accommodated for. The proposed hierarchical Bayesian method showed the best predictive performance according to multiple metrics. Beyond that, the resulting z-scores showed little to no residual site effects, yet still retained clinically useful information. In contrast, performance was particularly poor for the regression model and the ComBat model in which age and sex were not explicitly modeled. In all two stage harmonization models, predictions were poorly scaled, suffering from a loss of more than 90 % of the original variance. Our results show the value of hierarchical Bayesian regression methods for accommodating site variation in neuroimaging data, which provides an alternative to harmonization techniques. While the approach we propose may have broad utility, our approach is particularly well suited to normative modelling where the primary interest is in accurate modelling of inter-subject variation and statistical quantification of deviations from a reference model.

1 HighlightsO_LIDevelopment and presentation of normative modeling approach based on hierarchical Bayesian modeling that can be applied to large multi-site neuroimaging data sets.
C_LIO_LIComparison of performance of Hierarchical Bayesian model including site as predictor to several common ways to harmonize for multi-site effects.
C_LIO_LIPresentation of normative modeling as site correction tool.
C_LI A mixture of innate cryoprotectants is key for freeze tolerance and cryopreservation of a drosophilid fly larva. Insects that naturally tolerate internal freezing produce complex mixtures of multiple cryoprotectants (CPs). Better knowledge on composition of these mixtures, and on mechanisms of how the individual CPs interact, could inspire development of laboratory CP formulations optimized for cryopreservation of cells and other biological material. Here we identify and quantify (using high resolution mass spectrometry) a range of putative CPs in larval tissues of a subarctic fly, Chymomyza costata that survives long-term cryopreservation in liquid nitrogen. The CPs (proline, trehalose, glutamine, asparagine, glycine betaine, glycerophosphoethanolamine, glycerophosphocholine, and sarcosine) accumulate in hemolymph in a ratio of 313:108:55:26:6:4:3:0.5 mmol.L-1. Using calorimetry, we show that the artificial mixtures, mimicking the concentrations of major CPs in hemolymph of freeze-tolerant larvae, suppress the melting point of water and significantly reduce the ice fraction. We demonstrate in a bioassay that mixtures of CPs administered through the diet act synergistically rather than additively to enable cryopreservation of otherwise freeze-sensitive larvae. Using MALDI-MSI, we show that during slow extracellular freezing trehalose becomes concentrated in partially dehydrated hemolymph where it stimulates transition to the amorphous glass phase. In contrast, proline moves to the boundary between extracellular ice and dehydrated hemolymph and tissues where it likely forms a layer of dense viscoelastic liquid. We propose that amorphous glass and viscoelastic liquids may protect macromolecules and cells from thermomechanical shocks associated with freezing and transfer into and out of liquid nitrogen.

Summary statementThe composition of natural cryoprotectant mixture of the extremely freeze-tolerant insect is revealed. Components of the mixture work in synergy and behave differently during organismal freezing and cryopreservation. A parametric logistic equation with light flux and medium concentration for cultivation planning of microalgae Microalgae are considered to be promising producers of bioactive chemicals, feeds, and fuels from carbon dioxide by photosynthesis. Thus, the prediction of microalgal growth profiles is important for the planning of cost-effective and sustainable cultivation-harvest cycles. This paper proposes a mathematical model capable of predicting the effect of light flux into culture and medium concentration on the growth profiles of microalgae by incorporating these growth-limiting factors into the logistic equation. The specific form of the equation is derived based on the experimentally measured growth profiles of Monoraphidium sp., a microalgal strain isolated by the authors, under 16 conditions consisting of the combinations of incident light fluxes into culture and initial medium concentrations. Using a cross-validation method, it is shown that the proposed model has the ability to predict necessary incident light flux into culture and initial medium concentration for harvesting target biomass at target time. Finally, the model-guided cultivation planning is performed and is evaluated by comparing the result with the experimental data. Programmed knockout mutation of liver fluke granulin, Ov-grn-1, impedes malignant transformation during chronic opisthorchiasis Infection with the food-borne liver fluke Opisthorchis viverrini is the principal risk factor for cholangiocarcinoma in the Mekong Basin countries of Thailand, Lao PDR, Vietnam, Myanmar and Cambodia. Using a novel model of CCA, involving infection with gene-edited liver flukes in the hamster during concurrent exposure to a nitrosamine, we explored the role of the fluke granulin-like growth factor Ov-GRN-1 in malignancy. We produced programmed gene knockout flukes ({Delta}Ov-grn-1) by delivery of a CRISPR/Cas9/gRNA system by electroporation. Genome sequencing confirmed Cas9-catalyzed mutations in the targeted genes, which was accompanied by rapid depletion of transcripts and the cognate proteins. Whereas Ov-grn-1 gene-edited parasites colonized the biliary tract and developed into adult flukes, less hepatobiliary tract disease manifested during chronic infection with{Delta} Ov-grn-1 worms in comparison to hamsters infected with control parasites. Specifically, immunohistochemical analysis of thin sections of livers revealed markedly less periductal fibrosis surrounding the flukes and less liver fibrosis globally during infection with{Delta} Ov-grn-1 genotype worms, minimal biliary epithelial cell proliferation, and markedly fewer mutations of TP53 in biliary epithelial cells. Moreover, fewer hamsters developed high-grade cholangiocarcinoma when infected with the{Delta} Ov-grn-1 flukes compared to controls. The clinically-relevant, pathophysiological phenotype of the hepatobiliary tract confirmed a role for this secreted growth factor in malignancy and morbidity during opisthorchiasis. Pre- and post-oviposition behavioural strategies to protect eggs against extreme winter cold in an insect with maternal care AO_SCPLOWBSTRACTC_SCPLOWDepositing eggs in an area with adequate temperature is often crucial for mothers and their offspring, as the eggs are immobile and therefore cannot avoid exposure to sub-optimal temperatures. However, the importance of temperature on oviposition site selection is less clear when mothers can avoid these potential adverse effects by both moving their eggs after oviposition and providing other forms of egg care. In this study, we addressed this question in the European earwig, an insect in which mothers care for the eggs during several months in winter, frequently moving them during this period. We set up 60 females from two random natural populations (as this species often exhibits population-specific life-history traits and behaviours) under controlled thermal gradients, and recorded the temperature at which they built their nests, tested whether they moved their eggs after an experimental temperature change, and measured the effects on egg development and hatching rate. Our results demonstrate that females indeed select oviposition sites according to temperature, and can move their eggs to reach warmer temperatures. We also show that these warmer temperatures are necessary to ensure egg hatching. Although this set of behavioural thermoregulations is present in the two tested populations, we found a population-specific modality of expression. These included the range of temperatures explored before oviposition, temperature selected at oviposition and dynamics of egg transport following a temperature change. Overall, our study sheds light on a new post-oviposition strategy in female insects that overwinter with their eggs for coping with temperature changes. More generally, it also reveals that egg care and/or egg transport do not prevent behavioural thermoregulation via oviposition site selection and highlights the diversity of behaviours that insects can adopt to enhance their tolerance to global climate change. Household transmission of SARS-CoV-2 from humans to pets in Washington and Idaho: burden and risk factors SARS-CoV-2 is believed to have emerged from an animal reservoir; however, the frequency of and risk factors for inter-species transmission remain unclear. We carried out a community-based study of pets in households with one or more confirmed SARS-CoV-2 infection in humans. Among 119 dogs and 57 cats with completed surveys, clinical signs consistent with SARS-CoV-2 were reported in 20 dogs (21%) and 19 cats (39%). Out of 81 dogs and 32 cats sampled for testing, 40% of dogs and 43% of cats were seropositive, and 5% of dogs and 8% of cats were PCR positive; this discordance may be due to delays in sampling. Respondents commonly reported close human-animal contact and willingness to take measures to prevent transmission to their pets. Reported preventative measures showed a slightly protective trend for both illness and seropositivity in pets, while sharing of beds and bowls had slight harmful effects. The joint evolution of animal movement and competition strategies Competition typically takes place in a spatial context, but eco-evolutionary models rarely address the joint evolution of movement and competition strategies. Here we investigate a spatially explicit producer-scrounger model where consumers can either forage on a heterogeneous resource landscape or steal resource items from conspecifics (kleptoparasitism). We consider three scenarios: (1) a population of foragers in the absence of kleptoparasites; (2) a population of consumers that are either specialized on foraging or on kleptoparasitism; and (3) a population of individuals that can fine-tune their behavior by switching between foraging and kleptoparasitism depending on local conditions. By means of individual-based simulations, we study the joint evolution of movement and competition strategies, and we investigate the implications on the resource landscape and the distribution of consumers over this landscape. In all scenarios and for all parameters considered, movement and competition strategies evolved rapidly and consistently across replicate simulations. The evolved movement and resource exploitation patterns differ considerably across the three scenarios. For example, foragers are attracted by conspecifics in scenario (1), while they are repelled by conspecifics in scenario (2). Generally the movement strategies of kleptoparasites differ markedly from those of foragers, but even within each class of consumers polymorphisms emerge, corresponding to pronounced differences in movement patterns. In all scenarios, the distribution of consumers over resources differs substantially from  ideal free predictions. We show that this is related to the intrinsic difficulty of moving effectively on a depleted landscape with few reliable cues for movement. Our study emphasises the advantages of a mechanistic approach when studying competition in a spatial context, and suggests how evolutionary modelling can be integrated with current work in animal movement ecology. A novel two-generation approach to the population dynamics of gregarious parasitoids Parasitoids, as important natural enemies, help maintain balance in natural ecosystems. Their population dynamics is generally predicted from the number of individuals. Here, using gregarious parasitoids as models, we show that this traditional approach omits one important parameter: the mothers manipulation of offspring fertility due to the clutch size-body size- fertility correlation. As a result of this correlation, when females deliberately adjust the number of offspring laid in a host, they determine not only the number but also the body sizes and reproductive potentials of those offspring. For the first time in the model species Anaphes flavipes, we determined the parasitoids offspring fertility from clutch size. Using this, we experimentally clarified the advantage of specific clutch size combinations and we show that identical fertility in the F1 generation can lead to distinctly different fertility values in the F2 generation. Even with the same number of hosts, lower fertility in the F1 generation can cause higher fertility in the F2 generation. Based on these results, we propose a novel two-generation approach which includes the clutch size-body size-fertility correlation. Our novel approach provides a new perspective for determining the individual fitness levels of gregarious parasitoids with new options for the modelling of parasitoid population dynamics.