title: Membrane voltage fluctuations in human breast cancer cells

abstract: Cancer cells feature a resting membrane potential (Vm) that is depolarized compared to normal cells, and express active ionic conductances, which factor directly in their pathophysiological behavior. Despite similarities to  excitable tissues, relatively little is known about cancer cell Vm dynamics. With high-throughput, cellular-resolution Vm imaging, we characterized Vm fluctuations of hundreds of human triple-negative breast cancer MDA-MB-231 cells and compared to non-cancerous breast epithelial MCF-10A cells. By quantifying their Dynamic Electrical Signatures (DESs) through an unsupervised machine-learning protocol, we identified four classes ranging from "noisy" to "blinking/waving". The Vm of MDA-MB-231 cells exhibited spontaneous, transient hyperpolarizations that were inhibited by the voltage-gated sodium channel blocker tetrodotoxin. The Vm of MCF-10A cells was comparatively static, but fluctuations increased following treatment with transforming growth factor-{beta}1, a canonical inducer of the epithelial-to-mesenchymal transition. These data suggest that the ability to generate Vm fluctuations is acquired during transformation and may participate in oncogenesis.


title: Transcriptional signature in microglia isolated from an Alzheimers disease mouse model treated with scanning ultrasound

abstract:
RationaleIntracranial scanning ultrasound combined with intravenously injected microbubbles (SUS+MB) has been shown to transiently open the blood-brain barrier and reduce amyloid-{beta} (A{beta}) pathology in the APP23 mouse model of Alzheimers disease (AD). This has been accomplished, at least in part, through the activation of microglial cells; however, their response to the SUS treatment is only incompletely understood.

MethodsWild-type (WT) and APP23 mice were subjected to SUS+MB, using non-SUS+MB-treated mice as sham controls. After 48 hours, the APP23 mice were injected with methoxy-XO4 to label A{beta} aggregates, followed by microglial isolation into XO4+ and XO4- populations using flow cytometry. Both XO4+ and XO4- cells were subjected to RNA sequencing and their transcriptome was analyzed through a bioinformatics pipeline.

ResultsThe transcriptomic analysis of the microglial cells revealed a clear segregation depending on genotype (AD model versus WT mice), as well as treatment (SUS+MB versus sham) and A{beta} internalization (XO4+ versus XO4- microglia). Differential gene expression analysis detected 278 genes that were significantly changed by SUS+MB in the XO4+ cells (248 up/30 down) and 242 in XO- cells (225 up/17 down). Not surprisingly given previous findings of increased phagocytosis of plaques following SUS+MB, the pathway analysis highlighted that the treatment induced an enrichment in genes related to the phagosome pathway in XO4+ microglia; however, when comparing SUS+MB to sham, the analysis revealed an enrichment in genes involved in the cell cycle in both the XO4+ and XO4- microglial population.

ConclusionOur data provide a comprehensive analysis of microglia in an AD mouse model subjected to ultrasound treatment as a function of A{beta} internalization, one of the defining hallmarks of AD. Several differentially expressed genes are highlighted, pointing to an ultrasound-induced activation of cell cycle mechanisms in microglial cells isolated from APP23 mice treated with SUS+MB.

Graphical abstract

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category: neuroscience


title: The adolescent pattern of sleep spindle development revealed by HD-EEG polisomnography

abstract: Sleep spindles are developmentally relevant cortical oscillatory patterns; however, they have mostly been studied by considering the entire spindle frequency range (11 to 15 Hz) without a distinction between the functionally and topographically different slow and fast spindles, using relatively few electrodes and analysing wide age-ranges. Here, we employ HD-EEG polysomnography in three age-groups between 12 to 20 years of age, with an equal distribution between the two genders, and analyse the adolescent developmental pattern of the four major parameters of slow and fast sleep spindles. Most of our findings corroborate those very few previous studies that also make a distinction between slow and fast spindles in their developmental analysis. We find spindle frequency increasing with age, although spindle density change is not obvious in our study. We confirm the declining tendencies for amplitude and duration, although within narrower, more specific age-windows than previously. Spindle frequency seems to be higher in females in the oldest age-group. Based on the pattern of our findings, we suggest that HD-EEG, specifically targeting slow and fast spindle ranges and relatively narrow age-ranges would advance the understanding of both adolescent development and the functional relevance of sleep spindles in general.


title: The adolescent pattern of sleep spindle development revealed by HD-EEG polysomnography


title: Dichotomous Activity and Function of Neurons with Low- and High-Frequency Discharge in the External Globus Pallidus of Non-Human Primates

abstract: To date, there is a consensus that there are at least two neuronal populations in the non-human primate (NHP) external globus pallidus (GPe): the low- and high-frequency discharge (LFD and HFD) neurons. Nevertheless, almost all NHP physiological studies have neglected the functional importance of LFD neurons. This study examined the discharge features of these two GPe neuronal subpopulations recorded in four NHPs engaged in a classical conditioning task with cues predicting reward, neutral and aversive outcomes. The results show that LFD neurons tended to burst, encoded the salience of behavioral cues, and exhibited correlated spiking activity. By contrast, the HFD neurons tended to pause, encoded cue valence, and exhibited uncorrelated spiking activity. Overall, these findings point to the dichotomic organization of the NHP GPe which is likely to be critical to the implementation of normal basal ganglia functions and computations.


title: Interpersonal brain synchronization during face-to-face economic exchange between acquainted dyads

abstract: Interpersonal brain synchronization (IBS) has been observed during social interactions and involves various factors, such as familiarity with the partner and type of social activity. A previous study has shown that face-to-face interactions in pairs of strangers increase IBS. However, it is unclear whether this can be observed when the nature of the interacting partners is different. Herein, we aimed to extend these findings to pairs of acquaintances. Neural activity in the frontal and temporal regions was recorded using functional near-infrared spectroscopy hyperscanning. Participants played an ultimatum game that required virtual economic exchange in two experimental settings: the face-to-face and face-blocked conditions. Random pair analysis confirmed whether IBS was induced by social interaction. Contrary to the aforementioned study, our results did not show any cooperative behavior or task-induced IBS increase. Conversely, the random pair analysis results revealed that the pair-specific IBS was significant only in the task condition at the left and right superior frontal, middle frontal, orbital superior frontal, right superior temporal, precentral, and postcentral gyri. Our results revealed that face-to-face interaction in acquainted pairs did not increase IBS and supported the idea that IBS is affected by "with whom we interact and how."


title: Ultra-deep through-skull mouse brain imaging via the union of skull optical clearing and three-photon microscopy

abstract: Optical microscopy has enabled in vivo monitoring of brain structures and functions with high spatial resolution. However, the strong optical scattering in turbid brain tissue and skull impedes the observation of microvasculature and neuronal structures at a large depth. Herein, we proposed a strategy to overcome the influence induced by the high scattering effect of both skull and brain tissue via the combination of skull optical clearing (SOC) technique and thee-photon fluorescence microscopy (3PM). The Visible-NIR-II compatible Skull Optical Clearing Agents (VNSOCA) we applied reduced the skull scattering and water absorption in long wavelength by refractive index matching and H2O replacement to D2O respectively. 3PM with the excitation in the 1300-nm window reached 1.5 mm cerebrovascular imaging depth in cranial window. Combining the two advanced technologies together, we achieved so far the largest cerebrovascular imaging depth of 1.0 mm and neuronal imaging depth of >700 m through intact mouse skull. Dual-channel through-skull imaging of both brain vessels and neurons was also successfully realized, giving an opportunity of non-invasively monitoring the deep brain structures and functions at single-cell level simultaneously.


title: Ultra-deep through-skull mouse brain imaging via the combination of skull optical clearing and three-photon microscopy


title: Unsupervised functional neurocartography of the Aplysia buccal ganglion

abstract: A major limitation of large-scale neuronal recordings is the difficulty in locating homologous neurons in different subjects, referred to as the "correspondence" issue. This issue stems, at least in part, from the lack of a unique feature that unequivocally identifies each neuron. One promising approach to this problem is the functional neurocartography framework developed by Frady et al. (2016), in which neurons are identified by a semisupervised machine learning algorithm using a combination of multiple selected features. Here, the framework was adapted to the buccal ganglia of Aplysia. Multiple features were derived from neuronal activity during motor pattern generation, responses to peripheral nerve stimulation, and the spatial properties of each cell. The feature set was optimized based on its potential usefulness in discriminating neurons from each other, and then used to match putatively homologous neurons across subjects with the functional neurocartography software. An alternative matching method based on a cyclic matching algorithm was also developed, which allows for unsupervised extraction of groups of neurons and automated selection of high-quality matches. This improvement enabled unsupervised implementation of the machine learning algorithm, thereby enhancing scalability of the analysis. This study paves the way for investigating the roles of both well-characterized and previously uncharacterized neurons in Aplysia, and helps to adapt the neurocartography framework to other systems.


title: Profiles and Dynamics of the Transcriptome of Microglial Cells Reveal their Inflammatory Status

abstract:
The primary role of microglia in the maintenance of brain homeostasis is to respond to disturbances in the microenvironment. In this study, we cultured murine neonatal microglia and activated them with lipopolysaccharide (LPS) and benzoyl ATP (bzATP) to characterize changes in the transcriptome in response to various in vivo stimuli caused by pathogens, injury, or toxins. Activation by bzATP, an agonist of purinergic receptors, induces a transient wave of transcriptional changes. However, a long-lasting transcriptional profile affecting thousands of genes occurs only following a combination of bzATP and LPS. This profile is dominated by a coordinated induction of cytokines (e.g., IL1- and IL1-{beta}), chemokines, and their direct regulators. Many of these inflammatory-related genes are up-regulated by several orders of magnitude. We identified the TNF- and NF-{kappa}B pathways as the principal hubs for signaling of interleukin and chemokine induction in this cell system. We propose that primary microglia under controlled activation paradigms can be used for testing reagents that could attenuate their activated state. Such a microglial system could serve as a model for changes occurring in the aging brain and neurodegenerative diseases.

Highlight points* Primary murine microglia cultures release cytokines following activation with bzATP and LPS
* The wave of changes in gene expression by bzATP is transient.
* bzATP+LPS causes a transcription program dominated by the induction of interleukins and chemokines.


title: IL-23 signaling prevents ferroptosis-driven renal immunopathology during candidiasis

abstract: During infection the host relies on pattern-recognition receptors to sense invading fungal pathogens to launch immune defense mechanisms. While fungal recognition and immune effector responses are organ and cell type specific, during disseminated candidiasis myeloid cells exacerbate collateral tissue damage. However, the complex interplay between protective antifungal immunity and immunopathology remains incompletely understood. The {beta}-glucan receptor ephrin type-A 2 receptor (EphA2) is required to initiate mucosal inflammatory responses during oral Candida infection. Here we report that Epha2 promotes renal immunopathology during disseminated candidiasis. EphA2 deficiency leads to reduced renal inflammation and injury. Comprehensive analyses reveal that EphA2 limits IL-23 secretion in dendritic cells, while IL-23 signaling prevents ferroptotic myeloid cell death during infection. Further, ferroptosis aggravates inflammation during infection, while at the same time reducing the fungal killing capacity of macrophages. Thus, we identify ferroptotic cell death as a critical pathway of Candida-mediated renal immunopathology that opens a new avenue to tackle Candida infection and inflammation.

category: immunology


title: Transcriptomes and metabolism define mouse and human MAIT cell heterogeneity

abstract: Mucosal-associated invariant T (MAIT) cells are a subpopulation of T lymphocytes that respond to microbial metabolites. We performed single-cell RNA sequencing and metabolic analyses of MAIT cell subsets in thymus and peripheral tissues from mice and humans to define the heterogeneity and developmental pathway of these innate-like lymphocytes. We show that the predominant mouse subset, which produces IL-17 (MAIT17), and the subset that produces IFN{gamma} (MAIT1), have greatly different transcriptomes and metabolic states in the thymus and periphery. A splenic MAIT subset has a transcriptome similar to circulating lymphocytes, and in mice these also are found in recent thymic emigrants, suggesting partially mature cells emigrate from the thymus. Human MAIT cells are predominantly MAIT1 cells, but have a different metabolism from their mouse counterparts with increased fatty acid uptake and storage. Although mouse and human subsets are similar in thymus, in the periphery they diverge, likely reflecting environmental influences.


title: Stitchr: stitching coding TCR nucleotide sequences from V/J/CDR3 information

abstract: The study and manipulation of T cell receptors (TCRs) is central to multiple fields across basic and translational immunology research. Produced by V(D)J recombination, TCRs are often only recorded in the literature and data repositories as a combination of their V and J gene symbols, plus their hypervariable CDR3 amino acid sequence. However, numerous applications require full-length coding nucleotide sequences. Here we present Stitchr, a software tool developed to specifically address this limitation. Given minimal V/J/CDR3 information, Stitchr produces complete coding sequences representing a fully spliced TCR cDNA. Due to its modular design, Stitchr can be used for TCR engineering using either published germline or novel/modified variable and constant region sequences. Sequences produced by Stitchr were validated by synthesizing and transducing TCR sequences into Jurkat cells, recapitulating the expected antigen specificity of the parental TCR. Using a companion script, Thimble, we demonstrate that Stitchr can process a million TCRs in under ten minutes using a standard desktop personal computer. By systemizing the production and modification of TCR sequences, we propose that Stitchr will increase the speed, repeatability, and reproducibility of TCR research. Stitchr is available on GitHub.


title: Effect of Human Mesenchymal Stem Cells on Freund's adjuvant-induced Rheumatoid Arthritis in Sprague Dawley Rats

abstract:
IntroductionMesenchymal stem cells (MSC) therapy is a new approach to treat RA. Studies evaluating anti-inflammatory effects of MSCs per RA severity are scarce. Our primary objective was to evaluate anti-inflammatory effects, change in cytokine levels and cartilage regeneration of two different MSC preparations delivered through two different routes of administration in three RA stages: mild, moderate and severe.

MethodsHuman-derived umbilical cord tissue MSCs (hUCT-MSCs) and human bone marrow-derived MSCs (hBM-MSCs) delivered via intra-plantar and intravenous routes were tested in Freunds adjuvant-induced arthritis in rats. Arthritis severity was based on the arthritis score (<3=mild, 3=moderate and 4=severe). Assessments included changes in arthritis scoring, paw swelling, haematology parameters, biomarkers (TNF and IL-10) and histopathology analysis.

ResultsMSC treatment significantly reduced arthritis scores in all treatment groups. IL-10 levels increased 30 days after treatment with (IP)hUCT-MSCs (P=0.0241), (IV)hUCT-MSCs (P=0.0095) and (IP)hBM-MSCs (P=0.0002). TNF- levels reduced compared to positive control at 30 days: (IP)hUCT-MSCs (P=0.0060), (IV)hUCT-MSCs (P=0.0003), (IP)hBM-MSCs (P=0.0005), (IV)hBM-MSCs (P<0.0001) and continued through 30-60 days. Microscopic examination showed regenerative changes in animal joints treated with both intra-plantar or intravenous MSCs. Arthritis scores reduced in all RA severity groups while benefits (changes in IL-10 and TNF-alpha) were more pronounced in moderate and severe RA. Haematology parameters remained similar among all animal groups at baseline, 30 days and 60 days indicating safety of MSCs.

ConclusionTreatment with hUCT MSCs and hBM MSCs were safe, well-tolerated and effectively reduced joint inflammation, synovial cellularity and pro-inflammatory cytokine levels in CFA-induced RA rat model.

category: cell biology


title: Assessing the impact of fire on spider abundance through a global comparative analysis

abstract: Fire is an important ecological disturbance; however, in many regions fire regimes are changing due to anthropogenic factors. Understanding the responses of species and ecosystem to fire can help to develop predictive models and inform fire management decisions. Despite their massive diversity and abundance, relatively little is known about the impacts of fire on invertebrates. Spiders are a diverse and ubiquitous group, with variation in ecological, behavioural and life history traits, and can offer important insights into the impacts of fire on invertebrates and whether these depend on environmental factors, phylogenetic history, or species traits. We conducted phylogenetic comparative analyses of published data from studies investigating the impacts of fire on spiders. We investigated whether fire affects family level abundance and whether this effect depends on ecologically relevant traits or site-specific factors (vegetation, latitude, fire type, time since fire). We found that time since fire had a significant effect on the abundance of some spider families (Lycosidae, Linyphiidae, Agelenidae), and within forest vegetation types. There was no relationship between family level traits and response to fire, suggesting that a trait-based approach does not increase predictive power in this group at this taxonomic level. Overall, it is difficult to make broad generalisations about the impacts of fire on spider abundance due to variation in site- and fire-specific factors. We did, however, find evidence that short fire intervals may be a threat to some spider families, and in forest ecosystems, where abundance initially decreased after fire, which may have important implications for fire management strategies. Finally, we show that analyses of published data can be used to detect broad scale ecological patterns and provide an alternative to traditional meta-analytical approaches.

category: ecology


title: Background selection under evolving recombination rates

abstract: Background selection (BGS), the effect that purifying selection exerts on sites linked to deleterious alleles, is expected to be ubiquitous across eukaryotic genomes. The effects of BGS reflect the interplay of the rates and fitness effects of deleterious mutations with recombination. A fundamental assumption of BGS models is that recombination rates are invariant over time. However, in some lineages recombination rates evolve rapidly, violating this central assumption. Here, we investigate how recombination rate evolution affects genetic variation under BGS. We show that recombination rate evolution modifies the effects of BGS in a manner similar to a localised change in the effective population size, potentially leading to an underestimation of the genome-wide effects of selection. Furthermore, we find evidence that recombination rate evolution in the ancestors of modern house mice may have impacted inferences of the genome-wide effects of selection in that species.

category: evolutionary biology


title: The origin and evolution of loqs2: a gene encoding an antiviral dsRNA binding protein in Aedes mosquitoes

abstract: Mosquito borne viruses such as dengue, Zika, yellow fever and Chikungunya cause millions of infections every year. These viruses are mostly transmitted by two urban-adapted mosquito species, Aedes aegypti and Aedes albopictus, that appear to be more permissive to arbovirus infections compared to closely related species, although mechanistic understanding remains unknown. Aedes mosquitoes may have evolved specialized antiviral mechanisms that potentially contribute to the low impact of viral infection. Recently, we reported the identification of an Aedes specific double-stranded RNA binding protein (dsRBP), named Loqs2, that is involved in the control of infection by dengue and Zika viruses in Ae. aegypti. Loqs2 interacts with two important co-factors of the RNA interference (RNAi) pathway, Loquacious (Loqs) and R2D2, and seems to be a strong regulator of the antiviral defense. However, the origin and evolution of loqs2 remains unclear. Here, we describe that loqs2 likely originated from two independent duplications of the first dsRNA binding domain (dsRBD) of loquacious that occurred before the radiation of the Aedes Stegomyia subgenus. After its origin, our analyses suggest that loqs2 evolved by relaxed positive selection towards neofunctionalization. In fact, loqs2 is evolving at a faster pace compared to other RNAi components such as loquacious, r2d2 and Dicer-2 in Aedes mosquitoes. Unlike loquacious, transcriptomic analysis showed that loqs2 expression is tightly regulated, almost restricted to reproductive tissues in Ae. aegypti and Ae. albopictus. Transgenic mosquitoes engineered to ubiquitously express loqs2 show massive dysregulation of stress response genes and undergo developmental arrest at larval stages. Overall, our results uncover the possible origin and neofunctionalization of a novel antiviral gene, loqs2, in Aedes mosquitoes that ultimately may contribute to their effectiveness as vectors for arboviruses.


title: Dynamic collateral sensitivity profiles highlight challenges and opportunities for optimizing antibiotic sequences

abstract: As traditional antimicrobial therapies fail at escalating rates, recent focus has shifted to evolution-based therapies to slow resistance. Collateral sensitivity-the increased susceptibility to one drug associated with evolved resistance to a different drug-offers a potentially exploitable evolutionary constraint, but the manner in which collateral effects emerge over time is not well understood. Here, we use laboratory evolution in the opportunistic pathogen E. faecalis to phenotypically characterize collateral profiles through evolutionary time. Specifically, we measure collateral profiles for 400 strain-antibiotic combinations over the course of 4 evolutionary time points as strains are selected in increasing concentrations of antibiotic. We find that collateral resistance dominates during early phases of adaptation, whereas a diverse set of collateral profiles are accessible with further selection. Using simple numerical simulations, we illustrate how these temporally dynamic profiles potentially impact sequential drug therapies. Finally, we show experimentally how dynamic collateral sensitivity relationships can create optimal dosing windows that depend on finely timed switching between drugs.


title: Multiscale selection in spatially structured populations

abstract: The spatial structure of natural populations is key to many of their evolutionary processes. Formal theories analysing the interplay between natural selection and spatial structure have mostly focused on populations divided into distinct, non-overlapping groups. Most populations, however, are not structured in this way, but rather (self-)organise into dynamic patterns unfolding at various spatial scales. Here, we present a mathematical framework that quantifies how patterns and processes at different spatial scales contribute to natural selection in such populations. To that end, we define the Local Selection Differential (LSD): a measure of the selection acting on a trait within a given local environment. Based on the LSD, natural selection in a population can be decomposed into two parts: the contribution of local selection, acting within local environments, and the contribution of interlocal selection, acting among them. Varying the size of the local environments subsequently allows one to measure the contribution of each length scale. To illustrate the use of this new multiscale selection framework, we apply it to two simulation models of the evolution of traits known to be affected by spatial population structure: altruism and pathogen transmissibility. In both models, the spatial decomposition of selection reveals that local and interlocal selection can have opposite signs, thus providing a mathematically rigorous underpinning to intuitive explanations of how processes at different spatial scales may compete. It furthermore identifies which length scales--and hence which patterns--are relevant for natural selection. The multiscale selection framework can thus be used to address complex questions on evolution in spatially structured populations.


title: Identification of novel genetic regions associated with resistance to European canker in apple

abstract: Resistance to Neonectria ditissima, the fungus causing European canker in apple, was studied in a multiparental population of apple scions using several phenotyping methods. The studied population consists of individuals from multiple families connected through a common pedigree. The degree of disease of each individual in the population was assessed in three experiments: artificial inoculations of detached dormant shoots, potted trees in a glasshouse and in a replicated field experiment. The genetic basis of the differences in disease was studied using a pedigree-based analysis (PBA). Three quantitative trait loci (QTL), on linkage groups (LG) 6, 8 and 10 were identified in more than one of the phenotyping strategies. An additional four QTL, on LG 2, 5, 15 and 16 were only identified in the field experiment. The QTL on LG2 and 16 were further validated in a biparental population. QTL effect sizes were small to moderate with 4.3 to 19 % of variance explained by a single QTL. A subsequent analysis of QTL haplotypes revealed a dynamic response to this disease, in which the estimated effect of a haplotype varied over the field time-points. Two groups of QTL-haplotypes could be distinguished, one that displayed increased effect and one with a constant effect across time-points. These results suggest that there are different modes of control of N. ditissima in the early stages of infection compared to later time-points of disease development. It also shows that multiple QTL will need to be considered to improve resistance to European canker in apple breeding germplasm.

category: genetics


title: An exploration of linkage fine-mapping on sequences from case-control studies

abstract: Linkage analysis maps genetic loci for a heritable trait by identifying genomic regions with excess relatedness among individuals with similar trait values. Analysis may be conducted on related individuals from families, or on samples of unrelated individuals from a population. For allelically heterogeneous traits, population-based linkage analysis can be more powerful than genotypic-association analysis. Here, we focus on linkage analysis in a population sample, but use sequences rather than individuals as our unit of observation. Earlier investigations of sequence-based linkage mapping relied on known sequence relatedness, whereas we infer relatedness from the sequence data. We propose two ways to associate similarity in relatedness of sequences with similarity in their trait values and compare the resulting linkage methods to two genotypic-association methods. We also introduce a procedure to label case sequences as potential carriers or non-carriers of causal variants after an association has been found. This post-hoc labeling of case sequences is based on inferred relatedness to other case sequences. Our simulation results indicate that methods based on sequence-relatedness improve localization and perform as well as genotypic-association methods for detecting rare causal variants. Sequence-based linkage analysis therefore has potential to fine-map allelically heterogeneous disease traits.


title: hapCon: Estimating contamination of ancient genomes by copying from reference haplotypes

abstract:
MotivationHuman ancient DNA (aDNA) studies have surged in recent years, revolutionizing the study of the human past. Typically, aDNA is preserved poorly, making such data prone to contamination from other human DNA. Therefore, it is important to rule out substantial contamination before proceeding to downstream analysis. As most aDNA samples can only be sequenced to low coverages (<1x average depth), computational methods that can robustly estimate contamination in the low coverage regime are needed. However, the ultra low-coverage regime (0.1x and below) remains a challenging task for existing approaches.

ResultsWe present a new method to estimate contamination in aDNA for male individuals. It utilizes a Li&Stephens haplotype copying model for haploid X chromosomes, with mismatches modelled as genotyping error or contamination. We assessed an implementation of this new approach, hapCon, on simulated and down-sampled empirical aDNA data. Our results demonstrate that hapCon outperforms a commonly used tool for estimating male X contamination (ANGSD), with substantially lower variance and narrower confidence intervals, especially in the low coverage regime. We found that hapCon provides useful contamination estimates for coverages as low as 0.1x for SNP capture data (1240k) and 0.02x for whole genome sequencing data (WGS), substantially extending the coverage limit of previous male X chromosome based contamination estimation methods.

Availability and ImplementationA implementation of our software (hapCON) using Python and C has been deposited at https://github.com/hyl317/hapROH. We make hapCon available as part of a python package (hapROH), which is available at the Python Package Index (https://pypi.org/project/hapROH) and can be installed via pip. The documentation provides example use cases as blueprints for custom applications (https://haproh.readthedocs.io).


title: A Cartography of Differential Gene Methylation in Myalgic Encephalomyelitis/Chronic Fatigue Syndrome: Different Network Roles in the Protein-Protein Interactions Network Play Different, Biologically Relevant, Roles

abstract: Myalgic Encephalomyelitis, or Chronic Fatigue Syndrome (ME/CFS), is characterised by severe fatigue and associated with immune dysfunction. Previous studies of DNA methylation have found evidence of changes in immune cells for ME/CFS. However these studies have been limited by their small sample size. Here, we aggregate three comparable datasets to achieve a larger sample size and detect small changes to DNA methylation. We find 10,824 differentially methylated genes, with a small average change. Next, from the currently known interactions of relevant proteins, we build a Protein-Protein interaction network and,localising the network cartography analysis, we identify 184 hub genes. We find that different hub types play different, and meaningful, biological roles. Finally, we perform Gene ontology enrichment analysis, and we find that these hubs are involved in immune system processes, including response to TGF-{beta} and LPS, as well as mitochondrial functioning, supporting previous theories about ME/CFS. We also show that dopaminergic signalling may potentially contribute to immune pathology in ME/CFS, suggesting a possible interplay with Long Covid. Our results demonstrate the potentiality of network analysis in shedding light on the epigenetic contribution to the immune dysregulation of ME/CFS.

category: genomics


title: BISoN: A Bayesian Framework for Inference of Social Networks

abstract: Social networks are often constructed from point estimates of edge weights. In many contexts, edge weights are inferred from observational data, and the uncertainty around point estimates can be affected by various factors. Though this has been acknowledged in previous work, methods that explicitly quantify uncertainty in edge weights have not yet been widely adopted, and remain undeveloped for common types of data. Furthermore, existing methods are unable to cope with some of the complexities often found in observational data, and do not propagate uncertainty in edge weights to subsequent analyses. We introduce a unified Bayesian framework for modelling social networks based on observational data. This framework, which we call BISoN, can accommodate many common types of observational social data, can capture confounds and model effects at the level of observations, and is fully compatible with popular methods of social network analysis. We show how the framework can be applied to common types of data and how various types of downstream analyses can be performed, including non-random association tests and regressions on network properties. Our framework opens up the opportunity to test new types of hypotheses, make full use of observational datasets, and increase the reliability of scientific inferences. We have made example R code available to enable adoption of the framework.

category: animal behavior and cognition


title: Informeasure: an R/Bioconductor package to quantify nonlinear dependence between variables in biological networks from an information theory perspective

abstract: Using information measures to infer biological regulatory networks can observe nonlinear relationship between variables, but it is computationally challenging and there is currently no convenient tool available. We here describe an information theory R package named Informeasure that devotes to quantifying nonlinear dependence between variables in biological regulatory networks from an information theory perspective. This package compiles most of the information measures currently available: mutual information, conditional mutual information, interaction information, partial information decomposition and part mutual information. The first estimator is used to infer bivariate networks while the last four estimators are dedicated to analysis of trivariate networks. The base installation of this turn-key package allows users to approach these information measures out of the box. Informeasure is implemented in R program and is available as an R/Bioconductor package at https://bioconductor.org/packages/Informeasure.

category: bioinformatics


title: Inductive inference of novel protein-molecule interactions using Heterogeneous Graph Transformer (HGT) AutoEncoder

abstract: 1Protein-molecule interactions are promoted by the physicochemical characteristics of the actors involved, but structural information alone does not capture expression patterns, localization and pharmacokinetics. In this work we propose an integrative strategy for protein-molecule interaction discovery that combines different layers of information through the use of convolutional operators on graph, and frame the problem as missing link prediction task on an heterogeneous graph constituted by three node types: 1) molecules 2) proteins 3) diseases. Physicochemical information of the actors are encoded using shallow embedding techniques (SeqVec, Mol2Vec, Doc2Vec respectively) and are supplied as feature vectors to a Graph AutoEncoer (GAE) that uses a Heterogeneous Graph Transformer (HGT) in the encoder module. We show in this work that HGT Autoencoder can be used to accurately recapitulate the proteinmolecule interactions set and propose novel relationships in inductive settings that are grounded in biological and functional information extracted from the graph.


title: fastMSA: Accelerating Multiple Sequence Alignment with Dense Retrieval on Protein Language

abstract: Evolutionarily related sequences provide information for the protein structure and function. Multiple sequence alignment, which includes homolog searching from large databases and sequence alignment, is efficient to dig out the information and assist protein structure and function prediction, whose efficiency has been proved by AlphaFold. Despite the existing tools for multiple sequence alignment, searching homologs from the entire UniProt is still time-consuming. Considering the success of AlphaFold, foreseeably, large-scale multiple sequence alignments against massive databases will be a trend in the field. It is very desirable to accelerate this step. Here, we propose a novel method, fastMSA, to improve the speed significantly. Our idea is orthogonal to all the previous accelerating methods. Taking advantage of the protein language model based on BERT, we propose a novel dual encoder architecture that can embed the protein sequences into a low-dimension space and filter the unrelated sequences efficiently before running BLAST. Extensive experimental results suggest that we can recall most of the homologs with a 34-fold speed-up. Moreover, our method is compatible with the downstream tasks, such as structure prediction using AlphaFold. Using multiple sequence alignments generated from our method, we have little performance compromise on the protein structure prediction with much less running time. fastMSA will effectively assist protein sequence, structure, and function analysis based on homologs and multiple sequence alignment.


title: Towards mechanistic models of mutational effects: Deep Learning on Alzheimer's Aβ peptide

abstract: Alzheimers Disease (AD) is a debilitating form of dementia with a high prevalence in the global population and a large burden on the community and health care systems. ADs complex pathobiology consists of extracellular {beta}-amyloid deposition and intracellular hyperphosphorylated tau. Comprehensive mutational analyses can generate a wealth of knowledge about protein properties and enable crucial insights into molecular mechanisms of disease. Deep Mutational Scanning (DMS) has enabled multiplexed measurement of mutational effects on protein properties, including kinematics and self-organization, with unprecedented resolution. However, potential bottlenecks of DMS characterization include experimental design, data quality, and the depth of mutational coverage. Here, we apply Deep Learning to comprehensively model the mutational effect of the AD-associated peptide A{beta}42 on aggregation-related biochemical traits from DMS measurements. Among tested neural network architectures, Convolutional Neural Networks (ConvNets) and Recurrent Neural Networks (RNN) are found to be the most cost-effective models with robust high performance even under insufficiently-sampled DMS studies. While sequence features are essential for satisfactory prediction from neural networks, geometric-structural features further enhance the prediction performance. Notably, we demonstrate how mechanistic insights into phenotype may be extracted from the neural networks themselves suitably designed. This methodological benefit is particularly relevant for biochemical systems displaying a strong coupling between structure and phenotype such as the conformation of A{beta}42 aggregate and nucleation, as shown here using a Graph Convolutional Neural Network (GCN) developed from the protein atomic structure input. In addition to accurate imputation of missing values (which here ranged up to 55% of all phenotype values at key residues), the mutationally-defined nucleation phenotype generated from a GCN shows improved resolution for identifying known disease-causing mutations relative to the original DMS phenotype. Our study suggests that neural network derived sequence-phenotype mapping can be exploited not only to provide direct support for protein engineering or genome editing but also to facilitate therapeutic design with the gained perspectives from biological modeling.


title: Isotropic Three-Dimensional Dual-Color Super-Resolution Microscopy with Metal-Induced Energy Transfer

abstract: Over the last two decades, super-resolution microscopy has seen a tremendous development in speed and resolution, but for most of its methods, there exists a remarkable gap between lateral and axial resolution. Similar to conventional optical microscopy, the axial resolution is by a factor three to five worse than the lateral resolution. One recently developed method to close this gap is metal-induced energy transfer (MIET) imaging which achieves an axial resolution down to nanometers. It exploits the distance dependent quenching of fluorescence when a fluorescent molecule is brought close to a metal surface. In the present manuscript, we combine the extreme axial resolution of MIET imaging with the extraordinary lateral resolution of single-molecule localization microscopy, in particular with direct stochastic optical reconstruction microscopy (dSTORM). This combination allows us to achieve isotropic three-dimensional super-resolution imaging of sub-cellular structures. Moreover, we employed spectral demixing for implementing dualcolor MIET-dSTORM that allows us to image and co-localize, in three dimensions, two different cellular structures simultaneously.

category: biophysics


title: Recurrence of cancer cell states across diverse tumors and their interactions with the microenvironment

abstract: While genetic tumor heterogeneity has long been recognized, recent work has revealed significant variation among cancer cells at the epigenetic and transcriptional levels. Profiling tumors at the single-cell level in individual cancer types has shown that transcriptional heterogeneity is organized into cancer cell states, implying that diverse cell states may represent stable and functional units with complementary roles in tumor maintenance and progression. However, it remains unclear to what extent these states span tumor types, constituting general features of cancer. Furthermore, the role of cancer cell states in tumor progression and their specific interactions with cells of the tumor microenvironment remain to be elucidated. Here, we perform a pan-cancer single-cell RNA-Seq analysis across 15 cancer types and identify a catalog of 16 gene modules whose expression defines recurrent cancer cell states, including  stress,  interferon response,  epithelial-mesenchymal transition,  metal response,  basal and  ciliated. Using mouse models, we find that induction of the interferon response module varies by tumor location and is diminished upon elimination of lymphocytes. Moreover, spatial transcriptomic analysis further links the interferon response in cancer cells to T cells and macrophages in the tumor microenvironment. Our work provides a framework for studying how cancer cell states interact with the tumor microenvironment to form organized systems capable of immune evasion, drug resistance, and metastasis.

category: cancer biology


title: Striatal dopamine explains novelty-induced behavioral dynamics and individual variability in threat prediction

abstract:
Animals exhibit diverse behavioral responses, such as exploration and avoidance, to novel cues in the environment. However, it remains unclear how dopamine neuron-related novelty responses influence behavior. Here, we characterized dynamics of novelty exploration using multi-point tracking (DeepLabCut) and behavioral segmentation (MoSeq). Novelty elicits a characteristic sequence of behavior, starting with investigatory approach and culminating in object engagement or avoidance. Dopamine in the tail of striatum (TS) suppresses engagement, and dopamine responses were predictive of individual variability in behavior. Behavioral dynamics and individual variability were explained by a novel reinforcement learning (RL) model of threat prediction, in which behavior arises from a novelty-induced initial threat prediction (akin to "shaping bonus"), and a threat prediction that is learned through dopamine-mediated threat prediction errors. These results uncover an algorithmic similarity between reward- and threat-related dopamine sub-systems.

HighlightsO_LINovelty-induced behaviors are analyzed using modern machine-learning methods
C_LIO_LINovelty induces risk assessment which develops into engagement or avoidance
C_LIO_LIDopamine in the tail of striatum correlates with individual behavioral variability
C_LIO_LIReinforcement learning with shaping bonus and uncertainty explains the data
C_LI


title: Neuronal primary cilia regulate pyramidal cell positioning to the deep and superficial sublayers in the hippocampal CA1

abstract:
It is well-recognized that primary cilia regulate embryonic neurodevelopment, but little is known about their roles in postnatal neurodevelopment. The striatum pyramidal (SP) of hippocampal CA1 consists of superficial and deep sublayers, however, it is not well understood how early- and late-born pyramidal neurons position to two sublayers postnatally. Here we show that the axonemes of primary cilia of early-born pyramidal neurons point to the stratum oriens (SO), whereas late-born neuronal cilia orient toward the stratum radiatum (SR), reflecting an inside-out lamination pattern. Neuronal primary cilia in SP undergo marked changes in morphology and orientation from postnatal day 5 (P5) to P14, concurrent with pyramidal cell positioning to the deep and superficial sublayers and with neuronal maturation. Transgenic overexpression of Arl13B, a protein regulating ciliogenesis, not only elongates primary cilia and promotes earlier cilia protrusion, but also affects centriole positioning and cilia orientation in SP. The centrioles of late-born neurons migrate excessively to cluster at SP bottom before primary cilia protrusion and a reverse movement back to the main SP. Similarly, this backward movement of centriole/cilia is also manifested on late-born cortical pyramidal neurons, although early- and late-born cortical neurons display the same cilia orientation. Together, this study provides the first evidence demonstrating that late-born pyramidal neurons exhibit a reverse movement for cell positioning, and primary cilia regulate pyramidal neuronal positioning to the deep and superficial sublayers in the hippocampus.

Significance Statement1. Because the axonemes of primary cilia of early-born neurons point to the stratum oriens (SO), whereas late-born neuronal cilia orient toward the opposite direction, neuronal primary cilia orientation can be used as a marker to distinguish early- and late-born pyramidal neurons in the CA1.
2. This work is the first to show that neuronal primary cilia modulate pyramidal neuronal positioning to the deep and superficial sublayers of the hippocampal CA1.
3. Transgenic overexpression of Arl13B not only elongates primary cilia and promotes earlier cilia protrusion, but also affects centriole positioning and cilia orientation in the CA1.
4. We have identified a novel "reverse movement" of late-born CA1 and cortical pyramidal neurons for cell positioning, which may affect the maturation and formation of synaptic connectivity of late-born neurons.


title: The N-linker region of hERG1a upregulates hERG1b potassium channels

abstract:
A major physiological role of hERG1 (human Ether-a-go-go-Related Gene) potassium channels is to repolarize cardiac action potentials. Two isoforms, hERG1a and hERG1b, associate to form the native cardiac IKr current in vivo. Inherited mutations in hERG1a or hERG1b cause prolonged cardiac repolarization, Long QT Syndrome and sudden death arrhythmia. hERG1a subunits assemble with and enhance the number of hERG1b subunits at the plasma membrane, but the mechanism for the increase in hERG1b by hERG1a is not well understood. Here, we report that the hERG1a N-terminal PAS (Per-Arnt-Sim) domain-N-linker region expressed in trans with hERG1b markedly increased hERG1b currents and increased biotin-labelled hERG1b protein at the membrane surface. hERG1b channels with a deletion of the 1b domain did not have a measurable increase in current or biotinylated protein when co-expressed with hERG1a PAS domain-N-linker regions indicating that the 1b domain was required for the increase in hERG1b. Using a biochemical pull-down interaction assay and a FRET hybridization experiment, we detected a direct interaction between the hERG1a PAS domain-N-linker region and the hERG1b N-terminal 1b domain. Using engineered deletions and alanine mutagenesis, we identified a short span of amino acids at positions 216-220 within the hERG1a N-linker region that were necessary for the upregulation of hERG1b. Taken together, we propose that direct structural interactions between the hERG1a N-linker region and the hERG1b N-terminal 1b domain increase hERG1b at the plasma membrane. Mechanisms that enhance hERG1b current would be anticipated to shorten action potentials, which could be anti-arrhythmic, and may point toward hERG1b or the hERG1a N-linker as molecular targets for therapy for Long QT syndrome.

Significance StatementPotassium (K) ion channels help to control the membrane voltage in cardiac cells and are responsible for maintaining the normal rhythm of the heart. A decrease in the number of K channels causes cardiac arrhythmia and sudden death. One K channel (KCNH2b, hERG1b) weakly forms channels at the cell membrane. Here we describe a new mechanism for upregulation of hERG1b. An N-terminal domain of a related K channel (KCNH2a, hERG1a), expressed in trans as a separate piece, interacts with hERG1b and increases the number of hERG1b channels at the membrane. A loss of hERG channels is pro-arrhythmic and an increase in hERG channels is considered anti-arrhythmic, thus the hERG1a N-terminal domain is a potential therapeutic for cardiac arrhythmias.

category: physiology


title: Pattern similarity in the frontoparietal control network reflects an "off-veridical" template that optimizes target-match decisions during visual search

abstract: Theories of attention hypothesize the existence of an "attentional" or "target" template that contains task-relevant information in memory when searching for an object. The target template contributes to visual search by directing visual attention towards potential targets and serving as a decisional boundary for target identification. However, debate still exists regarding how template information is stored in the human brain. Here, we conducted a pattern-based fMRI study to assess how template information is encoded to optimize target-match decisions during visual search. To ensure that match decisions reflect visual search demands, we used a visual search paradigm in which all distractors were linearly separable but highly similar to the target and were known to shift the target representation away from the distractor features (Yu & Geng, 2019). In a separate match-to-sample probe task, we measured the target representation used for match decisions across two resting state networks that have long been hypothesized to maintain and control target information: the frontoparietal control network (FPCN) and the visual network (VisN). Our results showed that lateral prefrontal cortex in FPCN maintained the context-dependent "off-veridical" template; in contrast, VisN encoded a veridical copy of the target feature during match decisions. By using behavioral drift diffusion modeling, we verified that the decision criterion during visual search and the probe task relied on a common biased target template. Taken together, our results suggest that sensory-veridical information is transformed in lateral prefrontal cortex into an adaptive code of target-relevant information that optimizes decision processes during visual search.


title: Genetic and transcriptomic characteristics of RhlR-dependent quorum sensing in cystic fibrosis isolates of Pseudomonas aeruginosa

abstract: In people with the genetic disease cystic fibrosis (CF), bacterial infections involving the opportunistic pathogen Pseudomonas aeruginosa are a significant cause of morbidity and mortality. P. aeruginosa uses a cell-cell signaling mechanism called quorum sensing (QS) to regulate many virulence functions. One type of QS consists of acyl-homoserine lactone (AHL) signals produced by LuxI-type signal synthases, which bind a cognate LuxR-type transcription factor. In laboratory strains and conditions, P. aeruginosa employs two AHL synthase/receptor pairs arranged in a hierarchy, with the LasI/R system controlling the RhlI/R system and many downstream virulence factors. However, P. aeruginosa isolates with inactivating mutations in lasR are frequently isolated from chronic CF infections. We and others have shown that these isolates frequently use RhlR as the primary QS regulator. RhlR is rarely mutated in CF and environmental settings. We were interested if there were reproducible genetic characteristics of these isolates and if there was a central group of genes regulated by RhlR in all isolates. We examined five isolates and found signatures of adaptation common to CF isolates. We did not identify a common genetic mechanism to explain the switch from Las-to Rhl-dominated QS. We describe a core RhlR regulon encompassing 20 genes encoding 7 products. These results suggest a key group of QS-regulated factors important for pathogenesis of chronic infection, and position RhlR as a target for anti-QS therapeutics. Our work underscores the need to sample a diversity of isolates to understanding QS beyond what has been described in laboratory strains.

category: microbiology


title: 5-aminolevulinic acid-photodynamic therapy ameliorates cutaneous granuloma by killing drug-resistant Mycobacterium marinum

abstract:
BackgroundAlthough 5-aminolevulinic acid photodynamic therapy (ALA-PDT) has been extensively used to treat to various skin disease, the application of ALA-PDT on cutaneous infection caused by Mycobacterium marinum (M. marinum), especially by drug-resistant M. marinum is still not clear.

ObjectivesWe evaluated the therapeutic efficacy of ALA-PDT on M. marinum in a mouse infection model and tested its killing effect on M. marinum in vitro. Finally, we investigated the clinical effect of ALA-PDT in treating cutaneous granuloma caused by drug-resistant M. marinum.

Materials and methodsWe isolated total 9 strains of M. marinum from patients and confirmed by morphological and molecular approaches. The strains were identified by anti-mycobacterial susceptibility. Then we evaluated the killing effect of ALA-PDT on M. marinum in vitro and in a mouse model to observe the antimycobacterial effect of ALA-PDT. Therapeutic efficacy was further assessed in patients with cutaneous granuloma caused by drug-resistant M. marinum.

ResultsWe demonstrated that the ALA-PDT directly killed M. marinum in vitro. The paws cutaneous lesions of mice caused by M. marinum were fully recovered 2 weeks after ALA-PDT treatment. However, there was no significant difference for immune cells in peripheral blood before and after ALA-PDT therapy. Finally, ALA-PDT proved to be effective in treat two patients with cutaneous infection caused by drug-resistant M. marinum.

ConclusionsThe results suggest that ALA-PDT is effective in treating M. marinum cutaneous infections by killing M. marinum directly, independent of systemic immune responses. The data highlight the ALA-PDT as a promising therapeutic choice for M. marinum infection, especially for drug-resistant strains.


title: SHOT-R: A next generation algorithm for particle kinematics analysis

abstract: Analysis of live-imaging experiments is crucial to decipher a plethora of cellular mechanisms within physiological and pathological contexts. Kymograph, i.e. graphical representations of particle spatial position over time, and single particle tracking (SPT) are the currently available tools to extract information on particle transport and velocity. However, the spatiotemporal approximation applied in particle trajectory reconstruction with those methods intrinsically prevents an accurate analysis of particle kinematics and of instantaneous behaviours. Here, we present SHOT-R, a novel numerical method based on polynomial reconstruction of 4D (3D+time) particle trajectories. SHOT-R, contrary to other tools, computes bona fide instantaneous and directional velocity, and acceleration. Thanks to its high order continuous reconstruction it allows, for the first time, kinematics analysis of co-trafficked particles. Overall, SHOT-R is a novel, versatile, and physically reliable numerical method that achieves all-encompassing particle kinematics studies at unprecedented accuracy on any live-imaging experiment where the spatiotemporal coordinates can be retrieved.


title: HSP70-binding motifs function as protein quality control degrons

abstract: Protein quality control (PQC) degrons are short protein segments that target misfolded proteins for degradation through the ubiquitin-proteasome system (UPS). To uncover how PQC degrons function, we performed a screen in Saccharomyces cerevisiae by fusing a library of flexible tetrapeptides to the C-terminus of the Ura3-HA-GFP reporter. The identified degrons exhibited high sequence variation but with marked hydrophobicity. Notably, the best scoring degrons constitute predicted Hsp70-binding motifs. When directly tested, a canonical Hsp70 binding motif (RLLL) functioned as a dose-dependent PQC degron that was targeted by Hsp70, Hsp110, Fes1, several Hsp40 J-domain co-chaperones and the PQC E3 ligase Ubr1. Our results suggest that multiple PQC degrons overlap with chaperone-binding sites and that PQC-linked degradation achieves specificity via chaperone binding. Thus, the PQC system has evolved to exploit the intrinsic capacity of chaperones to recognize misfolded proteins, thereby placing them at the nexus of protein folding and degradation.

category: biochemistry


title: Geminivirus C4 proteins inhibit GA signaling via prevention of NbGAI degradation, to promote viral infection and symptom development in N. benthamiana

abstract:
Phytohormone gibberellin (GA) is an important plant signaling molecule that regulates plant growth and defense against abiotic and biotic stresses. To date, the molecular mechanism of the plant responses to viral infection mediated by GA is still undetermined. DELLA is a repressor of GA signaling and is recognized by the F-box protein, a component of the SCFSLY1/GID2 complex. The recognized DELLA is degraded by the ubiquitin-26S proteasome, leading to the activation of the GA signaling. Here, we report that ageratum leaf curl Sichuan virus (ALCScV)-infected N. benthamiana plants showed dwarfing symptom and abnormal flower development. The infection of ALCScV alters the expressions of GA pathway-related genes and decreases the content of endogenous GA significantly in N. benthamiana. Further, ALCScV-encoded C4 protein interacts with the DELLA protein NbGAI, and interferes with the interaction between NbGAI and NbGID2 to prevent the degradation of NbGAI, leading to the inhibition of the GA signaling pathway. Silencing of NbGAI or exogenous GA3 treatment significantly reduces viral accumulation and disease symptoms in N. benthamiana plants. The same results were proved by the experiments with C4 protein encoded by tobacco curly shoot virus (TbCSV). Therefore, we propose a novel mechanism of geminivirus C4 proteins controling virus infection and disease symptom development through interfering GA signaling pathway.

Author SummaryGibberellins (GAs) are plant hormones that are essential for many developmental processes in plants. It has indicated that plant virus infection can induce abnormal flower development and influence GA pathway resulting the plant dwarfing symptom, but the underlying mechanisms is still not well described. Here, we demonstrate that geminivirus - encoded C4 protein regulates the GA signaling pathway to promote viral accumulation and disease symptom development. Through directly interacting with NbGAI, the C4 protein interferes with the interaction between NbGAI and NbGID2, which inhibits the degradation of NbGAI. As a result, the GA signaling pathway is blocked, and the infected plants display symptoms of typical dwarfing and delayed flowering. Our results reveal a novel mechanism by which geminivirus C4 proteins influence viral pathogenicity via interfering the GA signaling pathway, and provide new insights into the interaction between virus and host.

category: pathology


title: Applying computer vision to digitised natural history collections for climate change research: temperature-size responses in British butterflies

abstract:
O_LINatural history collections (NHCs) are invaluable resources for understanding biotic response to global change. Museums around the world are currently imaging specimens, capturing specimen data, and making them freely available online. In parallel to the digitisation effort, there have been great advancements in computer vision (CV): the computer trained automated recognition/detection, and measurement of features in digital images. Applying CV to digitised NHCs has the potential to greatly accelerate the use of NHCs for biotic response to global change research. In this paper, we apply CV to a very large, digitised collection to test hypotheses in an established area of biotic response to climate change research: temperature-size responses.
C_LIO_LIWe develop a CV pipeline (Mothra) and apply it to the NHM iCollections of British butterflies (>180,000 specimens). Mothra automatically detects the specimen in the image, sets the scale, measures wing features (e.g., forewing length), determines the orientation of the specimen (pinned ventrally or dorsally), and identifies the sex. We pair these measurements and meta-data with temperature records to test how adult size varies with temperature during the immature stages of species and to assess patterns of sexual-size dimorphism across species and families.
C_LIO_LIMothra accurately measures the forewing lengths of butterfly specimens and compared to manual baseline measurements, Mothra accurately determines sex and forewing lengths of butterfly specimens. Females are the larger sex in most species and an increase in adult body size with warm monthly temperatures during the late larval stages is the most common temperature size response. These results confirm suspected patterns and support hypotheses based on recent studies using a smaller dataset of manually measured specimens.
C_LIO_LIWe show that CV can be a powerful tool to efficiently and accurately extract phenotypic data from a very large collection of digital NHCs. In the future, CV will become widely applied to digital NHC collections to advance ecological and evolutionary research and to accelerate the use of NHCs for biotic response to global change research.
C_LI


title: Assessment of the Finnish wolf population combines DNA captures, citizen observations and mortality data using a Bayesian state-space model

abstract: Assessment of the Finnish wolf population relies on multiple sources of information. This paper describes how Bayesian inference is used to pool the information contained in different kind of data sets (point observations, non-invasive genetics, known mortalities) for the estimation of the number of territories occupied by family packs and pairs. The output of the assessment model is a joint probability distribution, which describes current knowledge about the number of wolves within each territory. The joint distribution can be used to derive probability distributions for the total number of wolves in all territories and for the pack status within each territory. Most of the data set comprises of both voluntary-provided point observations and DNA samples provided by volunteers and research personnel. The new method reduces the role of expert judgement in the assessment process, providing increased transparency and repeatability.


title: Social wasps repair emergent topological defects

abstract: Social insects have evolved a variety of architectural formations. Bees and wasps are well known for their ability to achieve compact structures by building hexagonal cells. Polistes wattii, an open nesting paper wasp species, builds planar hexagonal structures. Here, using the pair correlation function approach, we show that their nests exhibit short-range hexagonal order but no long-range order akin to amorphous materials. Hexagonal orientational order was well preserved globally. We also show the presence of emergent topological defects such as disclination pairs (pentagon-heptagon dipoles), Stone-Wales quadrupoles, and other higher-order defects and discuss how these defects were fixed in the nest, thereby restoring order. Furthermore, we suggest the possible role of such defects in shaping nesting architectures of other social insect species.


title: Prenatal thyroid hormones accelerate postnatal growth and telomere shortening in wild great tits

abstract: Early-life environment is known to affect later-life health and disease, which could be mediated by the early-life programming of telomere length, a key hallmark of ageing. According to the fetal programming of telomere biology hypothesis, variation in prenatal exposure to hormones is likely to influence telomere length. Yet the contribution of key metabolic hormones, i.e. thyroid hormones (THs), has been largely ignored. We recently showed that in contrast to predictions, exposure to elevated prenatal THs increased postnatal telomere length in wild collared flycatchers, but the generality of such effect, its underlying proximate mechanisms and consequences on survival have not been investigated. We therefore conducted a comprehensive study evaluating the impact of THs on potential drivers of telomere dynamics (growth, post-natal THs, mitochondria and oxidative stress), telomere length and medium-term survival using wild great tits as a model system. While prenatal THs did not significantly affect telomere length after hatching (i.e. day 7), they influenced postnatal telomere shortening (i.e. shorter telomeres at day 14 and the following winter) but not apparent survival. Circulating THs, mitochondrial density or oxidative stress biomarkers were not significantly influenced, whereas TH-supplemented group showed accelerated growth, which may explain the observed delayed effect on telomeres. We discuss several alternative hypotheses that may explain the contrast with our previous findings in flycatchers. Given that shorter telomeres in early life tend to be carried until adulthood and are often associated with decreased survival prospects, the effects of prenatal THs on telomeres may have long-lasting effects on senescence.


title: The marine gastropod Conomurex luhuanus (Strombidae) has high-resolution spatial vision and eyes with complex retinas

abstract:
All species within the conch snail family Strombidae possess large camera-type eyes that are surprisingly well-developed compared to those found in most other gastropods. Although these eyes are known to be structurally complex, very little research on their visual function has been conducted. Here, we use isoluminant expanding visual stimuli to measure the spatial resolution and contrast sensitivity of a strombid, Conomurex luhuanus. Using these stimuli, we show that this species responds to objects as small as 1.06{degrees} in its visual field. We also show that C. luhuanus responds to Michelson contrasts of 0.07, a low contrast sensitivity between object and background. The defensive withdrawal response elicited by visual stimuli of such small angular size and low contrast suggests that conch snails may use spatial vision for the early detection of potential predators. We support these findings with morphological estimations of spatial resolution of 1.04 {+/-} 0.14{degrees}. These anatomical data therefore agree with the behavioural measures and highlight the benefits of integrating morphological and behavioural approaches in animal vision studies. Furthermore, using contemporary imaging techniques including serial block-face scanning electron microscopy (SBF-SEM), in conjunction with transmission electron microscopy (TEM), we found that C. luhuanus have more complex retinas, in terms of cell type diversity, than previous studies of the group have discovered using TEM alone. We found the C. luhuanus retina is comprised of six cell types, including a newly identified ganglion cell and accessory photoreceptor, rather than the previously described four cell types.

Summary statementBehavioural trials indicate the eyes of conch snail species Conomurex luhuanus provide high-resolution spatial vision, and morphological examination reveals the retina contains more cell types than those of other gastropods.


title: dynamAedes: a unified modelling framework for invasive Aedes mosquitoes

abstract:
O_LIMosquito species belonging to the genus Aedes have attracted the interest of scientists and public health officers for their invasive species traits and efficient capacity of transmitting viruses affecting humans. Some of these species were brought outside their native range by human activities such as trade and tourism, and colonised new regions thanks to a unique combination of eco-physiological traits.
C_LIO_LIConsidering mosquito physiological and behavioural traits to understand and predict the spatial and temporal population dynamics is thus a crucial step to develop strategies to mitigate the local densities of invasive Aedes populations.
C_LIO_LIHere, we synthesised the life cycle of four invasive Aedes species (Ae. aegypti, Ae. albopictus, Ae. japonicus and Ae. koreicus) in a single multi-scale stochastic modelling framework which we coded in the R package dynamAedes. We designed a stage-based and time-discrete stochastic model driven by temperature, photo-period and inter-specific larval competition that can be applied to three different spatial scales: punctual, local and regional. These spatial scales consider different degrees of spatial complexity and data availability, by accounting for both active and passive dispersal of mosquito species as well as for the heterogeneity of the input temperature data.
C_LIO_LIOur overarching aim was to provide a flexible, open-source and user-friendly tool rooted in the most updated knowledge on species biology which could be applied to the management of invasive Aedes populations as well as for more theoretical ecological inquiries.
C_LI


title: Expanded FLP toolbox for spatiotemporal protein degradation and transcriptomic profiling in C. elegans

abstract: Control of gene expression in specific tissues and/or at certain stages of development allows the study and manipulation of gene function with high precision. Site-specific genome recombination by the Flippase (FLP) and Cre enzymes has proven particularly relevant. Joint efforts of many research groups have led to the creation of efficient FLP and Cre drivers to regulate gene expression in a variety of tissues in Caenorhabditis elegans. Here, we extend this toolkit by the addition of FLP lines that drive recombination specifically in distal tip cells, the somatic gonad, coelomocytes and the epithelial P lineage. In some cases, recombination-mediated gene knockouts do not completely deplete protein levels due to persistence of long-lived proteins. To overcome this, we developed a spatiotemporally regulated degradation system for GFP fusion proteins (GFPdeg) based on FLP-mediated recombination. Using two stable nuclear pore proteins, MEL-28/ELYS and NPP-2/NUP85 as examples, we report the benefit of combining tissue-specific gene knockout and protein degradation to achieve complete protein depletion. We also demonstrate that FLP-mediated recombination can be utilized to identify nascent transcripts in a tissue of interest. We have adapted thiol(SH)-linked alkylation for the metabolic sequencing of RNA in tissue (SLAM-ITseq) for C. elegans. By focusing on a well-characterized tissue, the hypodermis, we show that the vast majority of genes identified by SLAM-ITseq are known to be expressed in this tissue, but with the added value of temporal resolution. These tools allow combining FLP activity for simultaneous gene inactivation and transcriptomic profiling, thus enabling the inquiry of gene function in various complex biological processes.


title: Confidence as a noisy decision reliability estimate

abstract: Decisions vary in difficulty. Humans know this and typically report more confidence in easy than in difficult decisions. However, confidence reports do not perfectly track decision accuracy, but also reflect response biases and difficulty misjudgments. To isolate the quality of confidence reports, we developed a model of the decision-making process underlying choice-confidence data. In this model, confidence reflects a subjects estimate of the reliability of their decision. The quality of this estimate is limited by the subjects uncertainty about the uncertainty of the variable that informs their decision ("meta-uncertainty"). This model provides an accurate account of choice-confidence data across a broad range of perceptual and cognitive tasks, revealing that meta-uncertainty varies across subjects, is stable over time, generalizes across some domains, and can be manipulated experimentally. The model offers a parsimonious explanation for the computational processes that underlie and constrain the sense of confidence.


title: From mass spectral features to molecules in molecular networks: a novel workflow for untargeted metabolomics.

abstract:
In the context of untargeted metabolomics, molecular networking is a popular and efficient tool which organizes and simplifies mass spectrometry fragmentation data (LC-MS/MS), by clustering ions based on a cosine similarity score. However, the nature of the ion species is rarely taken into account, causing redundancy as a single compound may be present in different forms throughout the network. Taking advantage of the presence of such redundant ions, we developed a new method named MolNotator. Using the different ion species produced by a molecule during ionization (adducts, dimers, trimers, in-source fragments), a predicted molecule node (or neutral node) is created by triangulation, and ultimately computing the associated molecules calculated mass. These neutral nodes provide researchers with several advantages. Firstly, each molecule is then represented in its ionization context, connected to all produced ions and indirectly to some coeluted compounds, thereby also highlighting unexpected widely present adduct species. Secondly, the predicted neutrals serve as anchors to merge the complementary positive and negative ionization modes into a single network. Lastly, the dereplication is improved by the use of all available ions connected to the neutral nodes, and the computed molecular masses can be used for exact mass dereplication. MolNotator is available as a Python library and was validated using the lichen database spectra acquired on an Orbitrap, computing neutral molecules for >90% of the 156 molecules in the dataset. By focusing on actual molecules instead of ions, MolNotator greatly facilitates the selection of molecules of interest.

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org.highwire.dtl.DTLVardef@730ecaorg.highwire.dtl.DTLVardef@1d012e1org.highwire.dtl.DTLVardef@187a1e5org.highwire.dtl.DTLVardef@195c53f_HPS_FORMAT_FIGEXP  M_FIG C_FIG


title: Engineered High-Affinity ACE2 Peptide Mitigates ARDS and Death Induced by Multiple SARS-CoV-2 Variants

abstract: Vaccine hesitancy and continuing emergence of SARS-CoV-2 variants of concern that may escape vaccine-induced immune responses highlight the urgent need for effective COVID-19 therapeutics. Monoclonal antibodies used in the clinic have varying efficacies against distinct SARS-CoV-2 variants; thus, there is considerable interest in engineered ACE2 peptides with augmented binding affinities for SARS-CoV-2 Spike protein. These could have therapeutic benefit against multiple viral variants. Using molecular dynamics simulations, we show how three amino acid substitutions in an engineered soluble ACE2 peptide (sACE22.v2.4-IgG1) markedly increase affinity for the SARS-CoV-2 Spike (S) protein. We demonstrate high binding affinity to S protein of the early SARS-CoV-2 WA-1/2020 isolate and also to multiple variants of concern: B.1.1.7 (Alpha), B.1.351 (Beta), P.1 (Gamma), and B.1.617.2 (Delta) SARS-CoV-2 variants. In humanized K18-hACE2 mice, prophylactic and therapeutic administration of sACE22.v2.4-IgG1 peptide prevented acute lung vascular endothelial injury and lung edema (essential features of ARDS) and significantly improved survival after infection by SARS-CoV-2 WA-1/2020 as well as P.1 variant of concern. These studies demonstrate for the first time broad efficacy in vivo of an ACE2 decoy peptide against multiple SARS-CoV-2 variants and point to its therapeutic potential.

category: pharmacology and toxicology


title: From climate warming to accelerated cellular ageing: an experimental study in wild birds

abstract:
Climate change is increasing both the average ambient temperature and the frequency and severity of heat waves. While direct mortality induced by heat waves is increasingly reported, sub-lethal effects are also likely to impact wild populations. We hypothesized that accelerated ageing could be a cost of being exposed to higher ambient temperature, especially in early-life when thermoregulatory capacities are not fully developed. We tested this hypothesis in wild great tit (Parus major) by experimentally increasing nest box temperature by ca. 2{degrees}C during postnatal growth and measuring telomere length, a biomarker of cellular ageing predictive of survival prospects in many bird species. While increasing early-life temperature does not affect growth or survival to fledging, it accelerates telomere shortening and reduces medium-term survival from 34% to 19%. Heat-induced telomere shortening was not explained by oxidative stress, but more likely by an increase in energy demand (i.e. higher thyroid hormones levels, increased expression of glucocorticoid receptor, increased mitochondrial density) leading to a reduction in telomere maintenance mechanisms (i.e. decrease in the gene expression of telomerase and protective shelterin). Our results thus suggest that climate warming can affect ageing rate in wild birds, with potential impact on population dynamics and persistence.

Significance statementStressful environmental conditions are known to accelerate cellular ageing, especially when experienced early in life. One unexplored avenue through which climate warming might affect wild animal populations is accelerated ageing. Here we show that increasing nest temperature by ca. 2{degrees}C during postnatal growth in a wild bird species can impact numerous physiological pathways and medium-term survival. Notably, artificially warming nests accelerates the shortening of telomeres, which are the protective end-caps of chromosomes considered as a hallmark of ageing. We thus suggest that warm ambient temperatures might accelerate ageing in wild animals, which can potentially impact population dynamics and extinction risk in the face of climate change.


title: Longitudinal characterisation of phagocytic and neutralisation functions of anti-Spike antibodies in plasma of patients after SARS-CoV-2 infection.

abstract:
Phagocytic responses by effector cells to antibody or complement-opsonised viruses have been recognized to play a key role in anti-viral immunity. These include antibody dependent cellular phagocytosis mediated via Fc-receptors, phagocytosis mediated by classically activated complement-fixing IgM or IgG1 antibodies and antibody independent phagocytosis mediated via direct opsonisation of viruses by complement products activated via the mannose-binding lectin pathway. Limited data suggest these phagocytic responses by effector cells may contribute to the immunological and inflammatory responses in SARS-CoV-2 infection, however, their development and clinical significance remain to be fully elucidated. In this cohort of 62 patients, acutely ill individuals were shown to mount phagocytic responses to autologous plasma-opsonised SARS-CoV-2 Spike protein-coated microbeads as early as 10 days post symptom onset. Heat inactivation of the plasma prior to use as an opsonin caused 77-95% abrogation of the phagocytic response, and pre-blocking of Fc-receptors on the effector cells showed only 18-60% inhibition. These results suggest that SARS-CoV-2 can provoke early phagocytosis, which is primarily driven by heat labile components, likely activated complements, with variable contribution from anti-Spike antibodies. During convalescence, phagocytic responses correlated significantly with anti-Spike IgG titers. Older patients and patients with severe disease had significantly higher phagocytosis and neutralisation functions when compared to younger patients or patients with asymptomatic, mild, or moderate disease. A longitudinal study of a subset of these patients over 12 months showed preservation of phagocytic and neutralisation functions in all patients, despite a drop in the endpoint antibody titers by more than 90%. Interestingly, surface plasmon resonance showed a significant increase in the affinity of the anti-Spike antibodies over time correlating with the maintenance of both the phagocytic and neutralisation functions suggesting that improvement in the antibody quality over the 12 months contributed to the retention of effector functions.

Author SummaryLimited data suggest antibody dependent effector functions including phagocytosis may contribute to the immunological and inflammatory responses in SARS CoV-2 infection, however, their development, maintenance, and clinical significance remain unknown. In this study we show: O_LIPatients with acute SARS CoV-2 infection can mount phagocytic responses as early as 10 days post symptom onset and these responses were primarily driven by heat labile components of the autologous plasma. These results indicate that the current approach of studying phagocytosis using purified or monoclonal antibodies does not recapitulate contribution by all components in the plasma.
C_LIO_LIIn convalescent patients, high phagocytic responses significantly correlated with increasing age, increasing disease severity, high neutralisation functions and high anti-Spike antibody titers, particularly IgG1.
C_LIO_LILongitudinal study of convalescent patients over a 12-month period showed maintenance of phagocytic and neutralisation functions, despite a drop in the anti-Spike endpoint antibody titers by more than 90%. However, we found significant increase in the affinity of the anti-Spike antibodies over the 12-month period and these correlated with the maintenance of functions suggesting that improvement in the antibody quality over time contributed to the retention of effector functions. Clinically, measuring antibody titers in sera but not the quality of antibodies is considered a gold standard indicator of immune protection following SARS-CoV 2 infection or vaccination. Our results challenge this notion and recommends change in the current clinical practice.
C_LI


title: mRNA-1273 and BNT162b2 mRNA vaccines have reduced neutralizing activity against the SARS-CoV-2 Omicron variant

abstract: The BNT162b2 (Pfizer-BioNTech) and mRNA-1273 (Moderna) vaccines generate potent neutralizing antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, the global emergence of SARS-CoV-2 variants with mutations in the spike protein, the principal antigenic target of these vaccines, has raised concerns over the neutralizing activity of vaccine-induced antibody responses. The Omicron variant, which emerged in November 2021, consists of over 30 mutations within the spike protein. Here, we used an authentic live virus neutralization assay to examine the neutralizing activity of the SARS-CoV-2 Omicron variant against mRNA vaccine-induced antibody responses. Following the 2nd dose, we observed a 30-fold reduction in neutralizing activity against the omicron variant. Through six months after the 2nd dose, none of the sera from naive vaccinated subjects showed neutralizing activity against the Omicron variant. In contrast, recovered vaccinated individuals showed a 22-fold reduction with more than half of the subjects retaining neutralizing antibody responses. Following a booster shot (3rd dose), we observed a 14-fold reduction in neutralizing activity against the omicron variant and over 90% of boosted subjects showed neutralizing activity against the omicron variant. These findings show that a 3rd dose is required to provide robust neutralizing antibody responses against the Omicron variant.


title: Immune escape of SARS-CoV-2 Omicron variant from mRNA vaccination-elicited RBD-specific memory B cells.

abstract: Memory B cells (MBCs) represent a second layer of immune protection against SARS-CoV-2. Whether MBCs elicited by mRNA vaccines can recognize the Omicron variant is of major concern. We used bio-layer interferometry to assess the affinity against the receptor-binding-domain (RBD) of Omicron spike of 313 naturally expressed monoclonal IgG that were previously tested for affinity and neutralization against VOC prior to Omicron. We report here that Omicron evades recognition from a larger fraction of these antibodies than any of the previous VOCs. Additionally, whereas 30% of these antibodies retained high affinity against Omicron-RBD, our analysis suggest that Omicron specifically evades antibodies displaying potent neutralizing activity against the D614G and Beta variant viruses. Further studies are warranted to understand the consequences of a lower memory B cell potency on the overall protection associated with current vaccines.


title: Blockade of TMPRSS2-mediated priming of SARS-CoV-2 by the N-terminal peptide of lactoferrin

abstract: In addition to vaccines, there is an urgent need for supplemental antiviral therapeutics to dampen the persistent COVID-19 pandemic caused by the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). The transmembrane protease serine 2 (TMPRSS2), which is responsible for the proteolytic processing of the SARS-CoV-2 spike protein as virus priming for cell entry, appears as a rational therapeutic target for the clearance of SARS-CoV-2 infection. Accordingly, selective inhibitors of TMPRSS2 represent potential tools for prevention and treatment of COVID-19. Here, we tested the inhibitory capacities of the human milk glycoprotein lactoferrin and its N-terminal peptide pLF1, which we identified as inhibitors of plasminogen, a serine protease homologous to TMPRSS2. In vitro proteolysis assays revealed that, unlike full-length lactoferrin, pLF1 significantly inhibited the proteolytic activity of TMPRSS2. pLF1 inhibited both the proteolytic processing of the SARS-CoV-2 spike protein and the SARS-CoV-2 infection of simian Vero cells. Because lactoferrin is a natural product and several biologically active peptides, such as the N-terminally derived lactoferricins, are produced naturally by pepsin-mediated digestion, natural or synthetic peptides from lactoferrin represent well-achievable candidates for supporting prevention and treatment of COVID-19.


title: Virus-like particles (VLPs) are efficient tools for boosting mRNA-induced antibodies

abstract: mRNA based vaccines against COVID-19 have proven most successful at keeping the SARS-CoV-2 pandemic at bay in many countries. Recently, there is an increased interest in heterologous prime-boost vaccination strategies for COVID-19 to maintain antibody response for the control of continuously emerging SARS-CoV-2 variants of concern (VoCs) and to overcome other obstacles such as supply shortage, costs and reduced safety issues or inadequate induced immune-response. In this study, we investigate the antibody responses induced by heterologous prime-boost with vaccines based on mRNA and virus-like particles (VLPs). The VLP-based mCuMVTT-RBM vaccine candidate and the approved mRNA-1273 vaccine were used for this purpose. We find that homologous prime boost regimens with either mRNA or VLP induced high levels of high avidity antibodies. Optimal antibody responses were, however, induced by heterologous regimens both for priming with mRNA and boosting with VLP and vice versa, priming with VLP and boosting with mRNA. Thus, heterologous prime boost strategies may be able to optimize efficacy and economics of novel vaccine strategies.


title: Evolution of Gram+ Streptococcus pyogenes has maximized efficiency of the Sortase A cleavage site

abstract:
Human plasminogen (hPg)-binding M-protein (PAM), a major virulence factor of Pattern D Streptococcus pyogenes (GAS), is the primary receptor responsible for binding and activating hPg. PAM is covalently bound to the cell wall (CW) through cell membrane (CM)-resident sortase A (SrtA)-catalyzed cleavage of the PAM-proximal C-terminal LPST{downarrow}-GEAA motif present immediately upstream of its transmembrane domain (TMD), and subsequent transpeptidation to the CW. These steps expose the N-terminus of PAM to the extracellular milieu (EM) to interact with PAM ligands, e.g., hPg. Previously, we found that inactivation of SrtA showed little reduction in functional binding of PAM to hPg, indicating that PAM retained in the cell membrane (CM) by the TMD nonetheless exposed its N-terminus to the EM. In the current study, we assessed the effects of mutating the Thr4 (P1) residue of the SrtA-cleavage site in PAM (Thr355 in PAM) to delay PAM in the CM in the presence of SrtA. Using rSrtA in vitro, LPSYGEAA and LPSWGEAA peptides were shown to have low activities, while LPSTGEAA had the highest activity. Isolated CM fractions of AP53/{Delta}SrtA cells showed that LPSYGEAA and LPSWGEAA peptides were cleaved at substantially faster rates than LPSTGEAA, even in CMs with an AP53/{Delta}SrtA/PAM[T355Y] double mutation, but the transpeptidation step did not occur. These results implicate another CM-resident enzyme that cleaves LPSYGEAA and LPSWGEAA motifs, most likely LPXTGase, but cannot catalyze the transpeptidation step. We conclude that the natural P1 (Thr) of the SrtA cleavage site has evolved to dampen PAM from nonfunctional cleavage by LPXTGase.

IMPORTANCEWe show in this study that functional cleavage of the sortase A (SrtA) cleavage signal for M-protein, LPST*GEAA, in the Gram+ cell membrane, which allows transpeptidation of M-protein to the cell wall, as opposed to non-functional cleavage by the highly active cell membrane nonribosomal enzyme, LPXTGase, at the downstream G-residue, is highly dependent on the presence of T at position 4. From our studies, we conclude that Streptococcus pyogenes has evolved in a manner that maximized T at this position so that SrtA preferentially cleaved the sorting signal in order that the virulence factor, M-protein, was stabilized on the cell surface through covalent attachment to the cell wall.


title: Genomic diversity and biosynthetic capabilities ofsponge-associated chlamydiae

abstract: Sponge microbiomes contribute to host health, nutrition, and defense through the production of secondary metabolites. Chlamydiae, a phylum of obligate intracellular bacteria ranging from animal pathogens to endosymbionts of microbial eukaryotes, are frequently found associated with sponges. However, sponge-associated chlamydial diversity has not yet been investigated at the genomic level and host-interactions remain thus far unexplored. Here, we sequenced the microbiomes of three sponge species and found high, though variable, Chlamydiae relative abundances of up to 21.2% of bacterial diversity. Using genome-resolved metagenomics 18 high-quality sponge-associated chlamydial genomes were reconstructed, covering four chlamydial families. Among these, Sorochlamydiaceae shares a common ancestor with Chlamydiaceae animal pathogens, suggesting long-term co-evolution with animals. Sponge-associated chlamydiae genomes mostly resembled environmental chlamydial endosymbionts, but not pathogens, and encoded genes for degrading diverse compounds associated with sponges, such as taurine. Unexpectedly, we identified widespread genetic potential for secondary metabolite biosynthesis across Chlamydiae, which may represent an explored reservoir of novel natural products. This finding suggests that chlamydiae may partake in defensive symbioses and that secondary metabolites play a wider role in mediating intracellular interactions. Furthermore, sponge-associated chlamydiae relatives were found in other marine invertebrates, pointing towards wider impacts of this phylum on marine ecosystems.


title: Plant growth promoting activity of bacteria isolated from Asian rice (Oryza sativa L.) are plant subspecies dependent

abstract: Asian rice is one of the most important crops because it is a staple food for almost half of the worlds population. Rice has two subspecies, JAPONICA and INDICA. To have production of rice keep pace with a growing world population, it is anticipated that the use of fertilizers will also need to increase, which may cause environmental damage through runoff impacts. An alternative strategy to increase crop yield is the use of plant growth promoting bacteria. Thousands of microbial species can exist in association with plant roots and shoots, and some are critical to the plants survival. We isolated 140 bacteria from rice and investigated whether JAPONICA and INDICA rice subspecies were positively influenced by these isolates. The bacterial isolates were screened for their ability to solubilize phosphate, a known plant growth promoting characteristic, and 25 isolates were selected for further analysis. These 25 phosphate solubilizing isolates were also able to produce other potentially growth-promoting factors. Five of the most promising bacterial isolates were chosen for whole genome sequencing. Four of these bacteria, isolates related to Pseudomonas mosselii, Microvirga sp., Paenibacillus rigui and Paenibacillus graminis, improved root and shoot growth, root to shoot ratio, and increased root dry weights of JAPONICA plants but had no effect on growth and development of INDICA plants. This indicates that while bacteria have several known plant growth promoting functions, their effects on growth parameters can be plant subspecies dependent and suggest close relationships between plants and their microbial partners.


title: The 'squalene route' to carotenoid biosynthesis is widespread in Bacteria.

abstract: Squalene is mostly associated with the biosynthesis of polycyclic triterpenes. Although there have been suggestions that squalene could be involved in the biosynthesis of carotenoids, functionally and evolutionarily related to polycyclic triterpenes, evidence of this  squalene route in nature was lacking. We demonstrate that planctomycetes synthesize C30 carotenoids via squalene and that this  squalene route is widely distributed in Bacteria. We also investigated the functional roles of hopanoids and carotenoids in Planctomycetes and show that their protective functions under stress conditions are complementary. Our evolutionary analyses suggest that the C30 carotenoid biosynthetic pathway is the most ancestral, with a potential origin in Firmicutes or Planctomycetes. In addition, we propose an evolutionary scenario to explain the diversification of the different carotenoid and squalene pathways. Together, these results improve the evolutionary contextualization of these molecules. Likewise, the widespread occurrence of the squalene route in bacteria increases the functional repertoire of squalene.


title: Neutralization against Omicron SARS-CoV-2 from previous non-Omicron infection

abstract: The explosive spread of the Omicron SARS-CoV-2 variant underscores the importance of analyzing the cross-protection from previous non-Omicron infection. We developed a high-throughput neutralization assay for Omicron SARS-CoV-2 by engineering the Omicron spike gene into an mNeonGreen USA-WA1/2020 SARS-CoV-2 (isolated in January 2020). Using this assay, we determined the neutralization titers of patient sera collected at 1- or 6-months after infection with non-Omicron SARS-CoV-2. From 1- to 6-month post-infection, the neutralization titers against USA-WA1/2020 decreased from 601 to 142 (a 4.2-fold reduction), while the neutralization titers against Omicron-spike SARS-CoV-2 remained low at 38 and 32, respectively. Thus, at 1- and 6-months after non-Omicron SARS-CoV-2 infection, the neutralization titers against Omicron were 15.8- and 4.4-fold lower than those against USA-WA1/2020, respectively. The low cross-neutralization against Omicron from previous non-Omicron infection supports vaccination of formerly infected individuals to mitigate the health impact of the ongoing Omicron surge.