title: Accelerated epigenetic aging and inflammatory/immunological profile (ipAGE) in patients with chronic kidney disease

abstract: Chronic kidney disease (CKD) is defined by reduced estimated glomerular filtration rate (eGFR). This failure can be related to a phenotype of accelerated aging. In this work we considered 76 subjects with end-stage renal disease (ESRD) and 83 healthy controls. We evaluated two measures that can be informative of the rate of aging, i.e. whole blood DNA methylation using the Illumina Infinium EPIC array and plasma levels of a selection of inflammatory/immunological proteins using Multiplex Immunoassays. We demonstrated accelerated aging in terms of the most common epigenetic age estimators in CKD patients. We developed a new predictor of age based on inflammatory/immunological profile (ipAGE) and confirmed age acceleration in CKD patients. Finally, we evaluated the relationship between epigenetic age predictors and ipAGE and further identified the inflammatory/immunological biomarkers differentially expressed between cases and controls. In summary, our data show an accelerated aging phenotype in CKD patients sustained by inflammatory processes.

category: bioinformatics


title: Towards an integrative multi-omics workflow

abstract:
The advent of high-throughput techniques has greatly enhanced biological discovery. Last years, analysis of multi-omics data has taken the front seat to improve physiological understanding. Handling functional enrichment results from various biological data raises practical questions.

We propose an integrative workflow to better interpret biological process insights in a multi-omics approach applied to breast cancer data from The Cancer Genome Atlas (TCGA) related to Invasive Ductal Carcinoma (IDC) and Invasive Lobular Carcinoma (ILC). Pathway enrichment by Over Representation Analysis (ORA) and Gene Set Enrichment Analysis (GSEA) has been conducted with both features information from differential expression analysis or selected features from multi-block sPLS-DA methods. Then, comprehensive comparisons of enrichment results have been carried out by looking at classical enrichment analysis, probabilities pooling by Stouffers Z scores method and pathways clustering in biological themes.

Our work shows that ORA enrichment with selected sPLS-DA features and pathways probabilities pooling by Stouffers method lead to enrichment maps highly associated to physiological knowledge of IDC or ILC phenotypes, better than ORA and GSEA with differential expression driven features.


title: Impact of neurite alignment on organelle motion.

abstract: Intracellular transport is pivotal for cell growth and survival. Malfunctions in this process have been associated with devastating neurodegenerative diseases, posing a need for deeper understanding of the involved mechanisms. Here, we used an experimental methodology that lead neurites of differentiated PC12 cells in either of two configurations: an one-dimensional, where the neurites align along lines, or a two-dimensional configuration, where the neurites adopt a random orientation and shape on a fiat substrate. We subsequently monitored the motion of functional organelles, the lysosomes, inside the neurites. Implementing a time-resolved analysis of the mean-squared displacement, we quantitatively characterized distinct motion modes of the lysosomes. Our results indicate that neurite alignment gives rise to faster diffusive and super-diffusive lysosomal motion in comparison to the situation where the neurites are randomly oriented. After inducing lysosome swelling through an osmotic challenge by sucrose, we confirmed the predicted slowdown in diffusive mobility. Surprisingly we found that the swelling-induced mobility change affected each of the (sub-/super-) diffusive motion modes differently and depended on the alignment configuration of the neurites. Our findings imply that intracellular transport is significantly and robustly dependent on cell morphology, which might be in part controlled by the extracellular matrix.

category: biophysics


title: Delineating Medical Education: Bibliometric Research Approach(es)

abstract:
BackgroundThe field of medical education remains poorly delineated such that there is no broad consensus of the articles and journals that comprise "the field." This lack of consensus has implications for conducting bibliometric studies and other research designs (e.g., systematic reviews); it also challenges the field to compare citation scores in the field and across others and for an individual to identify themselves as "a medical education researcher." Other fields have utilized bibliometric field delineation, which is the assigning of articles or journals to a certain field in an effort to define that field.

ProcessIn this Research Approach, three bibliometric field delineation approaches -- information retrieval, core journals, and journal co-citation -- are introduced. For each approach, the authors describe their attempt to apply it in the medical education context and identify related strengths and weaknesses. Based on co-citation, the authors propose the Medical Education Journal List 24 (MEJ-24), as a starting point for delineating medical education and invite the community to collaborate on improving and potentially expanding this list.

PearlsAs a research approach, field delineation is complicated, and there is no clear best way to delineate the field of medical education. However, recent advances in information and computer science provide potentially more fruitful approaches to deal with the complexity of the field. When considering these emerging approaches, researchers should consider collaborating with bibliometricians.

Bibliometric approaches rely on available metadata for articles and journals, which necessitates that researchers examine the metadata prior to analysis to understand its strengths and weaknesses, and to assess how this might affect their data interpretation. While using bibliometric approaches for field delineation is valuable, it is important to remember that these techniques are only as good as the research teams interpretation of the data, which suggests that an expanded research approach is needed to better delineate medical education, an approach that includes active discussion within the medical education community.

category: scientific communication and education


title: Resiniferatoxin hampers the nocifensive response of Caenorhabditis elegans to noxious heat, and pathway analysis revealed that the Wnt signaling pathway is involved

abstract: Resiniferatoxin (RTX) is a metabolite extracted from Euphorbia resinifera. RTX is a potent capsaicin analog with specific biological activities resulting from its agonist activity with the transient receptor potential channel vanilloid subfamily member 1 (TRPV1). RTX has been examined as a pain reliever, and more recently, investigated for its ability to desensitize cardiac sensory fibers expressing TRPV1 to improve chronic heart failure (CHF) outcomes using validated animal models. Caenorhabditis elegans (C. elegans) expresses orthologs of vanilloid receptors activated by capsaicin, producing antinociceptive effects. Thus, we used C. elegans to characterize the antinociceptive properties and performed proteomic profiling to uncover specific signaling networks. After exposure to RTX, wild-type (N2) and mutant C. elegans were placed on petri dishes divided into quadrants for heat stimulation. The thermal avoidance index was used to phenotype each tested C. elegans experimental group. The data revealed for the first time that RTX can hamper the nocifensive response of C. elegans to noxious heat (32{degrees}C - 35{degrees}C). The effect was reversed 6 h after RTX exposure. Additionally, we identified the RTX target, the C. elegans transient receptor potential channel OCR-3. The proteomics and pathway enrichment analysis results suggest that Wnt signaling is triggered by the agonistic effects of RTX on C. elegans vanilloid receptors.

category: neuroscience


title: EXPRESSION OF A SECRETABLE, CELL-PENETRATING CDKL5 PROTEIN ENHANCES THE EFFICACY OF AAV VECTOR-MEDIATED GENE THERAPY FOR CDKL5 DEFICIENCY DISORDER.

abstract: No therapy is currently available for CDKL5 (cyclin-dependent kinase-like 5) deficiency disorder (CDD), a severe neurodevelopmental disorder caused by mutations in the CDKL5 gene. Although delivery of a wild-type copy of the mutated gene to cells represents the most curative approach for a monogenic disease, proof-of-concept studies highlight significant efficacy caveats for brain gene therapy. Herein, we used a secretable TATk-CDKL5 protein to enhance the efficiency of a gene therapy for CDD. We found that, although AAVPHP.B_Igk-TATk-CDKL5 and AAVPHP.B_CDKL5 vectors had similar brain infection efficiency, the AAVPHP.B_Igk-TATk-CDKL5 vector led to a higher CDKL5 protein replacement due to secretion and transduction of the TATk-CDKL5 protein into the neighboring cells. Importantly, Cdkl5 KO mice treated with the AAVPHP.B_Igk-TATk-CDKL5 vector showed a behavioral and neuroanatomical improvement in comparison with vehicle-treated Cdkl5 KO mice or Cdkl5 KO mice treated with the AAVPHP.B_CDKL5 vector, indicating that a gene therapy based on a secretable recombinant TATk-CDKL5 protein is more effective at compensating Cdkl5-null brain defects than gene therapy based on the expression of the native CDKL5.


title: Understanding associative false memories in aging using multivariate analyses

abstract: Age-related declines in associative memory are ubiquitous, having been observed across a wide array of stimuli and experimental paradigms. Research further shows that such decreases in behavioral discriminability arise from increases in false memories for recombined lures. The current study examined the underlying neural basis of associative false memories by using representational similarity analyses to examine both age differences in the reactivation of encoded representations during retrieval and the neural overlap in the similarity of neural patterns underlying targets and related lures during retrieval. Behaviorally, we observed an age-related reduction in d, which was shown to be driven by increased false alarms in the older adults. While we found no age difference in the relationship between patterns of neural activity underlying hits across memory phases (as measured by ERS), the similarity of neural patterns underlying targets and lures was affected by age. Specifically, while younger and older adults exhibited overall higher pattern similarity between hits and CRs compared to hits and FAs (as measured by RSA), this difference was reduced in older adults within occipital cortices. Additionally, greater Hit-FA representational similarity correlated with increases in associative FAs across occipital, frontal, and parietal cortices. Results suggest that while neural representations underlying targets may not differ across age, greater pattern similarity between the neural representation of targets and lures may reflect reduced distinctiveness of the information encoded in memory, such that old and new items are more difficult to discriminate, leading to more false alarms.


title: A highly conserved lipase deacylates oxidized phospholipids and ameliorates acute lung injury

abstract: Oxidized phospholipids have diverse biological activities, many of which can be pathological, yet how they are inactivated in vivo is not fully understood. Here we present evidence that a previously unsuspected host lipase, acyloxyacyl hydrolase (AOAH), can play a significant role in reducing the pro-inflammatory activities of two prominent products of phospholipid oxidation, 1-palmitoyl-2-glutaryl-sn-glycero-3- phosphocholine (PGPC) and 1-palmitoyl-2-(5-oxovaleroyl)-sn-glycero-3-phosphocholine (POVPC). AOAH removed the sn-2 and sn-1 acyl chains from both lipids and reduced their ability to induce macrophage inflammasome activation and cell death in vitro and acute lung injury in vivo. In addition to transforming Gram-negative bacterial lipopolysaccharide from stimulus to inhibitor, its most studied activity, AOAH can inactivate these important danger-associated molecular pattern (DAMP) molecules and reduce tissue inflammation and cell death.

category: immunology


title: Identification of a new family of "megaphages" that are abundant in the marine environment

abstract: Megaphages - bacteriophages harbouring extremely large genomes - have recently been found to be ubiquitous, being described from a variety of microbiomes ranging from the animal gut to soil and freshwater systems. However, no complete marine megaphage has been identified to date. Here, using both short and long read sequencing, we assembled >900 high-quality draft viral genomes from water in the English Channel. One of these genomes included a novel megaphage, Mar_Mega_1 at >650 Kb, making it one of the largest phage genomes assembled to date. Utilising phylogenetic and network approaches, we found this phage represents a new family of bacteriophages. Genomic analysis showed Mar_Mega_1 shares relatively few homologues with its closest relatives, but, as with other mega-phages Mar_Mega_1 contained a variety of auxiliary metabolic genes responsible for carbon metabolism and nucleotide biosynthesis, including isocitrate dehydrogenase [NADP] and nicotinamide-nucleotide amidohydrolase [PncC] which have not previously been identified in megaphages. The results of this study indicate that phages containing extremely large genomes can be found in abundance in the marine environment and augment host metabolism by mechanisms not previously described.

category: microbiology


title: The robustness of a simple dynamic model of island biodiversity to geological and eustatic change

abstract:
AimBiodiversity on islands is affected by various geo-physical processes and sea-level fluctuations. Oceanic islands (never connected to a landmass) are initially vacant with diversity accumulating via colonisation and speciation, followed by a decline as islands shrink. Continental islands have species upon formation (when disconnected from the mainland) and may have transient land-bridge connections. Theoretical predictions for the effects of these geo-processes on rates of colonisation, speciation and extinction have been proposed, but methods of phylogenetic inference assume only oceanic island scenarios without accounting for island ontogeny, sea-level changes or past landmass connections. Here, we analyse to what extent ignoring geodynamics affects the inference performance of a phylogenetic island model, DAISIE, when confronted with simulated data that violate its assumptions.

LocationSimulation of oceanic and continental islands.

MethodsWe extend the DAISIE simulation model to include: area-dependent rates of colonisation and diversification associated with island ontogeny and sea-level fluctuations, and continental islands with biota present upon separation from the mainland, and shifts in rates to mimic temporary land-bridges. We quantify the error made when geo-processes are not accounted for by applying DAISIEs inference method to geodynamic simulations.

ResultsWe find that the robustness of the model to dynamic island area is high (error is small) for oceanic islands and for continental islands that have been separated for a long time, suggesting that, for these island types, it is possible to obtain reliable results when ignoring geodynamics. However, for continental islands that have been recently or frequently connected, robustness of DAISIE is low, and inference results should not be trusted.

Main conclusions: This study highlights that under a large proportion of island biogeographic geo-scenarios (oceanic islands and ancient continental fragments) a simple phylogenetic model ignoring geodynamics is empirically applicable and informative. However, recent connection to the continent cannot be ignored, requiring development of a new inference model.

category: evolutionary biology


title: Barcoded RH-seq illuminates the complex genetic basis of yeast thermotolerance

abstract: Decades of successes in statistical genetics have revealed the molecular underpinnings of traits as they vary across individuals of a given species. But standard methods in the field cant be applied to divergences between reproductively isolated taxa. Genome-wide reciprocal hemizygosity mapping (RH-seq), a mutagenesis screen in an inter-species hybrid background, holds promise as a method to accelerate the progress of interspecies genetics research. Here we describe an improvement to RH-seq in which mutants harbor barcodes for cheap and straightforward sequencing after selection in a condition of interest. As a proof of concept for the new tool, we carried out genetic dissection of the difference in thermotolerance between two reproductively isolated budding yeast species. Experimental screening identified dozens of candidate loci at which variation between the species contributed to the thermotolerance trait. Hits were enriched for mitosis genes and other housekeeping factors, and among them were multiple loci with robust sequence signatures of positive selection. Together, these results shed new light on the mechanisms by which evolution solved the problems of cell survival and division at high temperature in the yeast clade, and they illustrate the power of the barcoded RH-seq approach.


title: Barcoded reciprocal hemizygosity analysis via sequencing illuminates the complex genetic basis of yeast thermotolerance


title: Structure and function of the ROR2 cysteine-rich domain in vertebrate noncanonical WNT5A signaling

abstract: The receptor tyrosine kinase ROR2 mediates noncanonical WNT5A signaling to orchestrate tissue morphogenetic processes, and dysfunction of the pathway causes Robinow syndrome, Brachydactyly B and metastatic diseases. The domain(s) and mechanisms required for ROR2 function, however, remain unclear. We solved the crystal structure of the extracellular cysteine-rich (CRD) and Kringle (Kr) domains of ROR2 and found that, unlike other CRDs, the ROR2 CRD lacks the signature hydrophobic pocket that binds lipids/lipid-modified proteins, such as WNTs, suggesting a novel mechanism of receptor action. Functionally, we showed that the ROR2 CRD, but not other domains, is required and minimally sufficient to promote WNT5A signaling, and Robinow mutations in the CRD and the adjacent Kr alter ROR2 function. Moreover, we demonstrated that the activity of the ROR2 CRD requires Frizzled receptors. Thus, ROR2 acts via its CRD to potentiate the function of a receptor supercomplex that includes Frizzleds to transduce WNT5A signals.

category: biochemistry


title: Hoslundia opposita and other Nigerian plants inhibit sickle hemoglobin polymerization and prevent erythrocyte sickling: a virtual screening-guided identification of bioactive plants.

abstract: In sickle cell disease, a hereditary hemoglobinopathy, clinically observed disease presentations are the endpoint of a point mutation involving the substitution of glutamic acid with valine at the position 6 of the beta globin chain. With about 4.4 million people globally being affected, and another 43 million people bearing the sickle cell trait, several research efforts have been made to discover new and affordable treatment and possibly cure for the disease. Africa is endowed with a large flora population and traditional healers and citizens have over time depended on the use of herbs in folkloric medicine for different ailments including sickle cell disease (SCD). Such native knowledge has often formed the basis for different research exploration into the anti-sickling activities of selected African plants. These plants that have been so far investigated for their anti-sickling properties represent about 0.05 % of the 45,000 plant species enriching the flora landscape in Sub-Saharan Africa. Some of these have yielded potent anti-sickling profiles. In the current work we seek to achieve a more extensive search of the African plant diversity with anti-sickling properties: for this we have adopted a hybrid computational-cum-experimental protocol that employed computer-aided drug design (CADD) means for identifying plants with at least one constituent capable of interacting with the sickle hemoglobin, followed by extractive procedures and anti-sickling experiments for validating the predictions. Over two thousand (or 2,000) African natural products, representing over 200 plant species, were first virtually screened against the crystal structure of the dimerized human sickle hemoglobin. The natural products with the best computed sickle hemoglobin interaction energetics were found to belong to five plant species including Catharanthus roseus, Rauvolfia vomitoria, Hoslundia opposita, Lantana camara and Euphorbia hirta. The leaves of these plants were each collected subsequently and subjected to standard processing and extraction procedures. Using both HbSS polymerization inhibition and sickling reversal tests significant reductions in polymerization of erythrocyte hemolysate of the HbSS genotype were observed with the methanolic extracts of the plants, as well as sickling reversal levels of up to 68.50 % (H. opposita) was observed.


title: Molecular Learning of a Soft-Disks Fluid

abstract: This work is based on the Equivalence between Molecular Dynamics and Neural Network. It provides learning proofs in a Lennard-Jones (LJ) fluid, presented as a network of particles having non-bonded interactions. I describe the fluids learning as the property of an order that emerges as an adaptation in establishing equilibrium with energy and thermal conservation. The experimental section demonstrates the fluid can be trained with logic-gates patterns. The work goes beyond Molecular Computings application, explaining how this model uses its intrinsic minimizing properties in learning and predicting outputs. Finally, it gives hints for a theory on real chemistrys computational universality.


title: EMDA: A Python package for Electron Microscopy Data Analysis

abstract: An open-source Python library EMDA for cryo-EM map and model manipulation is presented with a specific focus on validation. The use of several functionalities in the library is presented through several examples. The utility of local correlation as a metric for identifying map-model differences and unmodeled regions in maps, and how it is used as a metric of map-model validation is demonstrated. The mapping of local correlation to individual atoms, and its use to draw insights on local signal variations are discussed. EMDAs likelihood-based map overlay is demonstrated by carrying out a superposition of two domains in two related structures. The overlay is carried out first to bring both maps into the same coordinate frame and then to estimate the relative movement of domains. Finally, the map magnification refinement in EMDA is presented with an example to highlight the importance of adjusting the map magnification in structural comparison studies.


title: Global protein responses of multi-drug resistant plasmid containing Escherichia coli to ampicillin, cefotaxime, imipenem and ciprofloxacin.

abstract: Antimicrobial resistance (AMR) and multi-drug resistance (MDR) in pathogenic bacteria are frequently mediated by plasmids. However, plasmids do not exist in isolation but rather require the bacterial host interaction in order to produce the AMR phenotype. This study aimed to utilise mass spectrometry-based proteomics to reveal the plasmid and chromosomally derived protein profile of Escherichia coli under antimicrobial stress. This was achieved by comparing the proteomes of E. coli containing the MDR pEK499 plasmid, under ampicillin, cefotaxime, imipenem or ciprofloxacin stress with the proteomes of these bacteria grown in the absence of antimicrobial. Our analysis identified statistically significant differentially abundant proteins common to groups exposed to the {beta}-lactam antimicrobials but not ciprofloxacin, indicating a {beta}-lactam stress response to exposure from this class of drugs, irrespective of {beta}-lactam resistance or susceptibility. These include ecotin and free methionine-R-sulfoxide reductase. These data also identified distinct differences in the cellular response to each {beta}-lactam. Data arising from comparisons of the proteomes of ciprofloxacin-treated E. coli and controls detected an increase in the relative abundance of proteins associated with ribosomes, translation, the TCA-cycle and several proteins associated with detoxification and a decrease in the relative abundances of proteins associated with stress response, including oxidative stress. We identified changes in proteins associated with persister formation in the presence of ciprofloxacin but not the {beta}-lactams. The plasmid proteome differed across each treatment and did not follow the pattern of antimicrobial - AMR protein associations. For example, a relative increase in the amount of blaCTX-M-15 in the presence of cefotaxime and ciprofloxacin but not the other {beta}-lactams, suggesting regulation of the blaCTX-M-15 protein production. The proteomic data from the this study provided novel insights into the proteins produced from the chromosome and plasmid under different antimicrobial stresses. These data also identified novel proteins not previously associated with AMR or antimicrobials responses in pathogens, which may well represent potential targets of AMR inhibition.


title: Interstitial and cerebrospinal fluid exchanging process revealed by phase alternate labeling with null recovery MRI

abstract:
PurposeTo develop Phase Alternate LAbeling with Null recovery (PALAN) MRI methods for the quantification of interstitial to cerebrospinal fluid flow (ICF) and cerebrospinal to interstitial fluid flow (CIF) in the brain.

MethodIn both T1-PALAN and apparent diffusion coefficient (ADC)-PALAN MRI methods, the cerebrospinal fluid (CSF) signal was nulled, while the residual interstitial fluid (ISF) was labeled by alternating the phase of pulses. ICF was extracted from the difference between the recovery curves of CSF with and without labeling. Similarly, CIF was measured by the T2-PALAN MRI method by labeling CSF, which took advance of the significant T2 difference between CSF and parenchyma.

ResultsBoth T1-PALAN and ADC-PALAN observed a rapid occurrence of ICF at 67{+/-}56 ms and 13{+/-}2 ms interstitial fluid transit times, respectively. ICF signal peaked at 1.5 s for both methods. ICF was 1153{+/-}270 ml/100ml/min with T1-PALAN in the third and lateral ventricles, which was higher than 891{+/-}60 ml/100ml/min obtained by ADC-PALAN. The results of the T2-PALAN suggested the ISF exchanging from ependymal layer to the parenchyma was extremely slow.

ConclusionThe PALAN methods are suitable tools to study ISF and CSF flow kinetics in the brain.

category: bioengineering


title: ABlooper: Fast accurate antibody CDR loop structure prediction with accuracy estimation

abstract: Antibodies are a key component of the immune system and have been extensively used as biotherapeutics. Accurate knowledge of their structure is central to understanding their antigen binding function. The key area for antigen binding and the main area of structural variation in antibodies is concentrated in the six complementarity determining regions (CDRs), with the most important for binding and most variable being the CDR-H3 loop. The sequence and structural variability of CDR-H3 make it particularly challenging to model. Recently deep learning methods have offered a step change in our ability to predict protein structures. In this work we present ABlooper, an end-to-end equivariant deep-learning based CDR loop structure prediction tool. ABlooper rapidly predicts the structure of CDR loops with high accuracy and provides a confidence estimate for each of its predictions. On the models of the Rosetta Antibody Benchmark, ABlooper makes predictions with an average CDR-H3 RMSD of 2.49[A], which drops to 2.05[A] when considering only its 76% most confident predictions.


title: Crosstalk with keratinocytes causes GNAQ oncogene specificity in melanoma

abstract: Different melanoma subtypes exhibit specific and non-overlapping sets of oncogene and tumor suppressor mutations, despite a common cell of origin in melanocytes. For example, activation of the Gq/11 signaling pathway is a characteristic initiating event in primary melanomas that arise in the dermis, uveal tract or central nervous system. It is rare in melanomas arising in the epidermis. Here, we present evidence that in the mouse, crosstalk with the epidermal microenvironment actively impairs the survival of melanocytes expressing the GNAQQ209L oncogene, providing a new model for oncogene specificity in cancer. The presence of epidermal cells inhibited cell division and fragmented dendrites of melanocytes expressing GNAQQ209L in culture, while they promoted the growth of normal melanocytes. Differential gene expression analysis of FACS sorted epidermal melanocytes showed that cells expressing GNAQQ209L exhibit an oxidative stress and apoptosis signature previously linked to vitiligo. Furthermore, PLCB4, the direct downstream effector of Gq/11 signaling, is frequently mutated in cutaneous melanoma alongside P53 and NF1. Our results suggest that a deficiency of PLCB4 promotes cutaneous melanomagenesis by reducing GNAQ driven signaling. Hence, our studies reveal the flip side of the GNAQ/PLCB4 signaling pathway, which was hitherto unsuspected. In the future, understanding how epidermal crosstalk restrains the GNAQQ209L oncogene could suggest novel melanoma therapies.

category: cancer biology


title: The Role of Biofilms, Bacterial Phenotypes, and Innate Immune Response in Mycobacterium avium Colonization to Infection

abstract:
Mycobacterium avium complex (MAC), is known for colonizing and infecting humans following inhalation of the bacteria. MAC pulmonary disease is notoriously difficult to treat and prone to recurrence. Both the incidence and prevalence MAC pulmonary disease have been increasing globally.

MAC is well known to form biofilms in the environment, and in vitro, these biofilms have been shown to aid MAC in epithelial cell invasion, protect MAC from phagocytosis, and cause premature apoptosis in macrophages. In vivo, the system of interactions between MAC, biofilms and host macrophages is complex, difficult to replicate in vitro and in animal models, has not been fully characterized. Here we present a three-dimensional agent-based model of a lung airway to help understand how these interactions evolve in the first 14 days post-bacterial inhalation. We parameterized the model using published data and performed uncertainty analysis to characterize outcomes and parameters effects on those outcomes.

Model results show diverse outcomes, including wide ranges of macrophage recruitment levels, and bacterial loads and phenotype distribution. Though most bacteria are phagocytosed by macrophages and remain intracellular, there are also many simulations in which extracellular bacteria continue to drive the colonization and infection. Initial parameters dictating host immune levels, bacterial loads introduced to the airway, and biofilm conditions have significant and lasting impacts on the course of these results. Additionally, though macrophage recruitment is key for suppressing bacterial loads, there is evidence of significant excess recruitment that fail to impact bacterial numbers. These results highlight a need and identify a path for further exploration into the inhalation events in MAC infection. Early infection dynamics could have lasting impacts on the development of nodular bronchiectatic or fibrocavitary disease as well as inform possible preventative and treatment intervention targeting biofilm-macrophage interactions.

category: systems biology


title: Synaptic Mechanisms of Top-Down Control by The Auditory Cortico-Collicular Pathway

abstract: Corticofugal projections to evolutionarily ancient, sub-cortical structures are ubiquitous across mammalian sensory systems. These "descending" pathways enable the neocortex to control ascending sensory representations in a predictive or feedback manner, but the underlying cellular mechanisms are poorly understood. Here we combine optogenetic approaches with in vivo and in vitro patch-clamp electrophysiology to study the projection from auditory cortex to the inferior colliculus (IC), a major descending auditory pathway that controls IC neuron feature selectivity, plasticity and auditory perceptual learning. Although individual auditory cortico-collicular synapses were generally weak, IC neurons often integrated inputs from multiple corticofugal axons that generated reliable, tonic depolarizations even during prolonged presynaptic activity. Latency measurements in vivo showed that descending signals reach the IC within 30 ms of sound onset, which in IC neurons corresponded to the peak of synaptic depolarizations evoked by short sounds. Activating ascending and descending pathways at latencies expected in vivo caused a NMDA receptor dependent, supra-linear EPSP summation, indicating that descending signals can non-linearly amplify IC neurons moment-to-moment acoustic responses. Our results shed light upon the synaptic bases of descending sensory control, and imply that heterosynaptic cooperativity contributes to the auditory cortico-collicular pathways role in plasticity and perceptual learning.


title: Synaptic Mechanisms of Top-Down Control in the Non-Lemniscal Inferior Colliculus


title: Plasmid supercoiling decreases during the dark phase in cyanobacteria: a clarification of the interpretation of chloroquine-agarose gels.

abstract: In cyanobacteria DNA supercoiling varies over the diurnal light/dark cycle and is integrated with the circadian transcription program and (Woelfle et al. [2007], Vijayan et al. [2009], PNAS). Specifically, Woelfle et al. have reported that DNA supercoiling of an endogenous plasmid became progressively higher during prolonged dark phases in Synechococcus elongatus PCC 7942. This is counterintuitive, since higher levels of negative DNA supercoiling are commonly associated with exponential growth and high metabolic flux. Vijayan et al. then have reverted the interpretation of plasmid mobility on agarose gels supplemented with chloroquine diphosphate (CQ), but not further discussed the differences. Here, we set out to clarify this open issue in cyanobacterial DNA supercoiling dynamics. We first re-capitulate Kellers band counting method (1975, PNAS) using CQ instead of ethidium bromide as the intercalating agent. A 500x-1000x higher CQ concentration is required in the DNA relaxation reaction (topoisomerase I) than in the agarose gel buffer to quench all negative supercoiling of pUC19 extracted from Escherichia coli. This is likely due to the dependence of both, the DNA binding affinity of CQ and the induced DNA unwinding angle, on the ionic strength of the buffer. Lower levels of CQ were required to fully relax in vivo pUC19 supercoiling than were used by Woelfle et al. Next, we analyzed the in vivo supercoiling of endogenous plasmids of Synechocystis sp. PCC 6803, at two different CQ concentrations. These experiments indicate that negative supercoiling of plasmids does not increase but decreases in the dark phase, and progressively decreases further in prolonged darkness.


title: A bacterial dynamin-like protein confers a novel phage resistance strategy on the population level in Bacillus subtilis

abstract: Bacillus subtilis DynA is a member of the dynamin superfamily, involved in membrane remodeling processes. DynA was shown to catalyze full membrane fusion and it plays a role in membrane surveillance against antibiotics. We show here that DynA also provides a novel resistance mechanism against phage infection. Cells lacking DynA are efficiently lysed after phage infection and virus replication. DynA does not prevent phage infection and replication in individual cells, but significantly delays host cell lysis, thereby slowing down the release of phage progeny from the host cells. During the process, DynA forms large, almost immobile clusters on the cell membrane that seem to support membrane integrity. Single molecule tracking revealed a shift of freely diffusive molecules within the cytosol towards extended, confined motion at the cell membrane following phage induction. Thus, the bacterial dynamins are the first anti-phage system reported to delay host cell lysis and the last line of defense of a multilayered antiviral defense. DynA is therefore providing protective effects on the population, but not on single cell level.


title: SARS-CoV-2 Delta variant pathogenesis and host response in Syrian hamsters

abstract: B.1.617 lineage is becoming a dominant SARS-CoV-2 lineage worldwide and was the dominant lineage reported in second COVID-19 wave in India, which necessitated studying the properties of the variant. We evaluated the pathogenicity and virus shedding of B.1.617.2 (Delta) and B.1.617.3 lineage of SARS-CoV-2 and compared with that of B.1, an early virus isolate with D614G mutation in Syrian hamster model. Viral load, antibody response and lung disease were studied. No significant difference in the virus shedding pattern was observed among these variants studied. A significantly high SARS-CoV-2 sub genomic RNA could be detected in the respiratory tract of hamsters infected with Delta variant for 14 days. Delta variant induced lung disease of moderate severity in 40% of infected animals. The neutralizing capability of the B.1, Delta and B.1.617.3 variant infected animals were found significantly lower with the B.1.351 (Beta variant). The findings of the study support the attributed disease severity and the increased transmission potential of the Delta variant.

category: molecular biology


title: The extracellular matrix gene, Svep1, orchestrates airway patterning and the transition from lung branching morphogenesis to alveolar maturation in the mouse

abstract: Proper lung development and function requires two independent but interrelated processes: branching morphogenesis to form the airway tree, and alveolar cell differentiation for peripheral gas exchange. The disruption of either branching or differentiation results in severe respiratory deficiencies and often in neonatal death. The molecular mechanisms that control branching patterns and the transition to alveolar differentiation are not completely understood. Here we report on the in vitro and in vivo characterization of the lungs of mouse embryos lacking a functional Svep1 gene. Our data demonstrate that the SVEP1 extracellular matrix protein is critical for airway patterning and for the process of transitioning from branching to alveolar maturation. Svep1-/- embryos on a C57BL/6J genetic background are characterized by hypoplastic lungs and a disorganized increase in distal airway tips which disrupts airway architecture and lobe shape. The lungs of Svep1 knockout embryos also have defects in alveolar differentiation. In vitro lung explant experiments demonstrated that SVEP1 normally inhibits branching morphogenesis and that treatment with a SVEP1 peptide can rescue the branching defects observed in Svep1 knockouts. Our findings reveal for the first time that Svep1 is essential for constructing the basic airway architecture and for the transition from lung branching to alveolar differentiation. Our results suggest therapeutic strategies to enhance lung development in patients with life-threatening respiratory disorders such as the lung hypoplasia and prematurity observed in neonates with congenital diaphragmatic hernia (CDH).

category: developmental biology


title: Unraveling three-dimensional chromatin structural dynamics during spermatogonial differentiation

abstract: Spermatogonial stem cells (SSCs) are able to undergo self-renewal and differentiation. Unlike the self-renewal that replenishes the SSC and progenitor pool, the differentiation is an irreversible process committed to meiosis. While the preparations for meiotic events in differentiating spermatogonia (Di-SG) are likely to be accompanied by alterations in chromatin structure, the three-dimensional (3D) chromatin architectural difference between SSCs and Di-SG, and the higher-order chromatin dynamics during spermatogonial differentiation, have not been systematically investigated. Here, we performed in situ high-throughput chromosome conformation capture (Hi-C), RNA-sequencing (RNA-seq) and chromatin immunoprecipitation-sequencing (ChIP-seq) analyses on porcine undifferentiated spermatogonia (Un-SG, which consist of SSCs and progenitors) and Di-SG. By integrating and analyzing these data, we identified that Di-SG exhibited increased disorder but weakened compartmentalization and topologically associating domains (TADs) in comparison with Un-SG, suggesting that diminished higher-order chromatin architecture in meiotic cells, as shown by recent reports, is preprogramed in Di-SG. Our data also revealed that A/B compartments and TADs were related to dynamic gene expression during spermatogonial differentiation. We further unraveled the contribution of promoter-enhancer interactions (PEIs) to pre-meiotic transcriptional regulation, which has not been accomplished in previous studies due to limited cell input and resolution. Together, our study uncovered the 3D chromatin structure of SSCs/progenitors and Di-SG, as well as the interplay between higher-order chromatin architecture and dynamic gene expression during spermatogonial differentiation, providing novel insights into the mechanisms for SSC self-renewal and differentiation and having implications for diagnosis and treatment of male sub-/infertility.


title: A multispecies amplicon sequencing approach for genetic diversity assessment in grassland plant species

abstract:
Grasslands are widespread and economically relevant ecosystems at the basis of sustainable roughage production. Plant genetic diversity (PGD; i.e., within-species diversity) is related to many beneficial effects to the ecosystem functioning of grasslands. The monitoring of PGD in temperate grasslands is complicated by the multiplicity of species present and by a shortage of methods for large-scale assessment. However, the continuous advancement of high-throughput DNA sequencing approaches have improved the prospects of broad, multispecies PGD monitoring. Among them, amplicon sequencing stands out as a robust and cost-effective method.

Here we report a set of twelve multispecies primer pairs that can be used for high-throughput PGD assessment in multiple grassland plant species. The loci targeted by the amplicons were selected and tested in two phases: a "discovery phase" based on a sequence capture assay (611 target nuclear loci assessed in 16 grassland plant species), which resulted in the selection of eleven loci; and a "validation phase", in which the selected loci were targeted and sequenced using twelve multispecies primers in test populations of Dactylis glomerata L., Lolium perenne L., Festuca pratensis Huds., Trifolium pratense L. and T. repens L. The resulting multispecies amplicons had overall nucleotide diversities per species ranging from 5.19 x 10-3 to 1.29 x 10-2, which is in the range of flowering-related genes but slightly lower than pathogen resistance genes. We conclude that the methodology, the DNA sequence resources, and the amplicon-specific primer pairs reported in this study provide the basis for large-scale, multispecies PGD monitoring in grassland plants.

category: ecology


title: COPILOT: a Containerised wOrkflow for Processing ILlumina genOtyping daTa

abstract:
BackgroundThe Illumina genotyping microarrays generate data in image format, which is processed by the platform-specific software GenomeStudio, followed by an array of complex bioinformatics analyses. This process can be time-consuming, lead to reproducibility errors, and be a daunting task for novice bioinformaticians.

ResultsHere we introduce the COPILOT (Containerised wOrkflow for Processing ILlumina genOtyping daTa) protocol, which provides an in-depth and clear guide to process raw Illumina genotype data in GenomeStudio, followed by a containerised workflow to automate an array of complex bioinformatics analyses involved in a GWAS quality control (QC). The COPILOT protocol was applied to two independent cohorts consisting of 2791 and 479 samples genotyped on the Infinium Global Screening (GSA) array with Multi-disease (MD) drop-in (~750,000 markers) and the Infinium H3Africa consortium array (~2,200,000 markers) respectively. Following the COPILOT protocol, an average sample quality improvement of 1.24% was observed across sample call rates, with notable improvement for low-quality samples. For example, from the 3270 samples processed, 141 samples had an initial sample call rate below 98%, averaging 96.6% (95% CI 95.6-97.7%), which is considered below the acceptable sample call rate threshold for a typical GWAS analysis. However, following the COPILOT protocol, all 141 samples had a call rate above 98% after QC and averaged 99.6% (95% CI 99.5-99.7%). In addition, the COPILOT pipeline automatically identified potential data issues, including gender discrepancies, heterozygosity outliers, related individuals, and population outliers through ancestry estimation.

ConclusionsThe COPILOT protocol makes processing Illumina genotyping data transparent, effortless and reproducible. The container is deployable on multiple platforms, improves data quality, and the end product is analysis-ready PLINK formatted data, with a comprehensive and interactive summary report to guide the user for further data analyses.

category: genomics


title: NMR spectroscopy of a single mammalian embryo

abstract: The resolving power, chemical sensitivity and non-invasive nature of NMR has made it an established technique for in vivo studies of large organisms both for research and clinical applications. These features would clearly be beneficial at the nanoliter scale (nL), typical of early development of mammalian embryos, microtissues and organoids, the scale where the building blocks of complex organisms could be observed. However, the handling of such small samples (about 100 micrometers) and sensitivity issues have prevented the widespread adoption of NMR. Recently we have shown how these limitations can be overcome with ultra-compact single-chip probes. In this article we show that such probes have sufficient sensitivity to detect and resolve NMR signals from individual bovine pre-implantation embryos. In less than 1 hour these spherical samples of just 130-190 micrometers produce distinct spectral peaks, largely originating from lipids contained inside them. We further observe how the spectral features, namely the peak intensities, vary from one sample to another despite their optical and morphological similarities.


title: N-glycosylation profiles of the SARS-CoV-2 spike D614G mutant and its ancestral protein characterized by advanced mass spectrometry

abstract: N-glycosylation plays an important role in the structure and function of membrane and secreted proteins. The spike protein on the surface of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus that causes COVID-19, is heavily glycosylated and the major target for developing vaccines, therapeutic drugs and diagnostic tests. The first major SARS-CoV-2 variant carries a D614G substitution in the spike (S-D614G) that has been associated with altered conformation, enhanced ACE2 binding, and increased infectivity and transmission. In this report, we used mass spectrometry techniques to characterize and compare the N-glycosylation of the wild type (S-614D) or variant (S-614G) SARS-CoV-2 spike glycoproteins prepared under identical conditions. The data showed that half of the N-glycosylation sequons changed their distribution of glycans in the S-614G variant. The S-614G variant showed a decrease in the relative abundance of complex-type glycans (up to 45%) and an increase in oligomannose glycans (up to 33%) on all altered sequons. These changes led to a reduction in the overall complexity of the total N-glycosylation profile. All the glycosylation sites with altered patterns were in the spike head while the glycosylation of three sites in the stalk remained unchanged between S-614G and S-614D proteins.


title: Mutations in two SARS-CoV-2 variants of concern reflect two distinct strategies of antibody escape

abstract: Understanding the factors that contribute to antibody escape of SARS-CoV-2 and its variants is key for the development of drugs and vaccines that provide broad protection against a variety of virus variants. Using microfluidic diffusional sizing, we determined the dissociation constant (KD) for the interaction between receptor binding domains (RBDs) of SARS-CoV-2 in its original version (WT) as well as alpha and beta variants with the host-cell receptor angiotensin converting enzyme 2 (ACE2). For RBD-alpha, the ACE2-binding affinity was increased by a factor of ten when compared with RBD-WT, while ACE2-binding of RBD-beta was largely unaffected. However, when challenged with a neutralizing antibody that binds to both RBD-WT and RBD-alpha with low nanomolar KD values, RBD-beta displayed no binding, suggesting a substantial epitope change. In SARS-CoV-2 convalescent sera, RBD-binding antibodies showed low nanomolar affinities to both wild-type and variant RBD proteins--strikingly, the concentration of antibodies binding to RBD-beta was half that of RBD-WT and RBD-alpha, again indicating considerable epitope changes in the beta variant. Our data therefore suggests that one factor contributing to the higher transmissibility and antibody evasion of SARS-CoV-2 alpha and beta is a larger fraction of viruses that can form a complex with ACE2. However, the two variants employ different mechanisms to achieve this goal. While SARS-CoV-2 alpha RBD binds with greater affinity to ACE2 and is thus more difficult to displace from the receptor by neutralizing antibodies, RBD-beta is less accessible to antibodies due to epitope changes which increases the chances of ACE2-binding and infection.


title: The structural role of bacterial eDNA in the formation of biofilm streamers

abstract: Across diverse habitats, bacteria are mainly found as biofilms, surface-attached communities embedded in a self-secreted matrix of extracellular polymeric substances (EPS), which enhances bacterial resistance to antimicrobial treatment and mechanical stresses. In the presence of flow and geometric constraints such as corners or constrictions, biofilms take the form of long, suspended threads known as streamers, which bear important consequences in industrial and clinical settings by causing clogging and fouling. The formation of streamers is thought to be driven by the viscoelastic nature of the biofilm matrix. Yet, little is known about the structural composition of streamers and how it affects their mechanical properties. Here, using a microfluidic platform that allows growing and precisely examining biofilm streamers, we show that extracellular DNA (eDNA) constitutes the backbone and is essential for the mechanical stability of Pseudomonas aeruginosa s streamers. This finding is supported by the observations that DNA-degrading enzymes prevent the formation of streamers and clear already formed ones, and that the antibiotic ciprofloxacin promotes their formation by increasing the release of eDNA. Furthermore, using mutants for production of the exopolysaccharide Pel, an important component of P. aeruginosa s EPS, we reveal a new, although indirect role of Pel, in tuning the mechanical properties of the streamers. Taken together, these results highlight the importance of eDNA and of its interplay with Pel in determining the mechanical properties of P. aeruginosa streamers, and suggest that targeting the composition of streamers can be an effective approach to control the formation of these biofilm structures.


title: Differential Interactions Between Human ACE2 and Spike RBD of SARS-CoV-2 Variants of Concern

abstract:
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the current coronavirus disease 2019 (COVID-19) pandemic. It is known that the receptor-binding domain (RBD) of the spike protein of SARS-CoV-2 interacts with the human angiotensin-converting enzyme 2 (ACE2) receptor, initiating the entry of SARS-CoV-2. Since its emergence, a number of SARS-CoV-2 variants have been reported, and the variants that show high infectivity are classified as the variants of concern according to the US CDC. In this study, we performed both all-atom steered molecular dynamics (SMD) simulations and microscale thermophoresis (MST) experiments to characterize the binding interactions between ACE2 and RBD of all current variants of concern (Alpha, Beta, Gamma, and Delta) and two variants of interest (Epsilon and Kappa). We report that the RBD of the Alpha (N501Y) variant requires the highest amount of force initially to be detached from ACE2 due to the N501Y mutation in addition to the role of N90-glycan, followed by Beta/Gamma (K417N/T, E484K, and N501Y) or Delta (L452R and T478K) variant. Among all variants investigated in this work, the RBD of the Epsilon (L452R) variant is relatively easily detached from ACE2. Our results combined SMD simulations and MST experiments indicate what makes each variant more contagious in terms of RBD and ACE2 interactions. This study could help develop new drugs to inhibit SARS-CoV-2 entry effectively.

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title: A machine learning based analysis to probe the relationship between odorant structure and olfactory behaviour in C. elegans

abstract: The chemical basis of smell remains an unsolved problem, with ongoing studies mapping perceptual descriptor data from human participants to the chemical structures using computational methods. These approaches are, however, limited by linguistic capabilities and inter-individual differences in participants. We use olfactory behaviour data from the nematode C. elegans, which has isogenic populations in a laboratory setting, and employ machine learning approaches for a binary classification task predicting whether or not the worm will be attracted to a given monomolecular odorant. Among others, we use architectures based on Natural Language Processing methods on the SMILES representation of chemicals for molecular descriptor generation and show that machine learning algorithms trained on the descriptors give robust prediction results. We further show, by data augmentation, that increasing the number of samples increases the accuracy of the models. From this detailed analysis, we are able to achieve accuracies comparable to that in human studies and infer that there exists a non trivial relationship between the features of chemical structures and the nematodes behaviour.


title: Evaluating the feasibility and efficacy of a dual-modality nanoparticle contrast agent (Nanotrast-CF800) for image-guided sentinel lymph node mapping in the oral cavity of healthy dogs

abstract: A combination of pre- and intraoperative sentinel lymph node (SLN) mapping techniques has been suggested to optimize SLN detection. A novel liposomal nanoparticle, Nanotrast-CF800 (CF800), utilizes computed tomography lymphography (CTL) and near infrared fluorescence imaging (NIRF) for image-guided surgery and SLN mapping. This novel tracer agent has not been evaluated in companion animals. The objective of this study was to evaluate the feasibility and efficacy of CF800 for SLN mapping in the oral cavity of healthy dogs and to report any local adverse effects. Six healthy adult purpose-bred research dogs randomly received either 1 mL (group 1) or 2 mL (group 2) of CF800 injected into the submucosa at the level of the right canine maxillary tooth. CTL and percutaneous NIRF were performed at 1, 3, and 10 minutes, then 1, 2, 4, 7, and 10 days post-injection (p.i). Overall, both CTL and NIRF identified SLNs in all dogs. The overall peak mean contrast enhancement of the SLNs was 73.98 HU (range 63.45-86.27 HU) at 2 days p.i. Peak fluorescence of the SLN occurred at 1 day p.i. The agent was retained within the SLN for at least 7 days for CTL and 4 days for NIRF. No adverse effects were observed. Local administration of CF800 was simple and feasible for the detection of SLNs using CTL+NIRF in the head and neck of healthy dogs and was not associated with significant local adverse events.


title: The peroxisomal transporter ABCD3 plays a major role in dicarboxylic fatty acid metabolism

abstract: Peroxisomes metabolize a specific subset of fatty acids, which include dicarboxylic fatty acids (DCAs) generated by {omega}-oxidation. Data obtained in vitro suggest that the peroxisomal transporter ABCD3 (also known as PMP70) mediates the transport of DCAs into the peroxisome, but in vivo evidence to support this role is lacking. In this study, we studied an Abcd3 KO mouse model generated by CRISPR-Cas9 technology using targeted and untargeted metabolomics, histology, immunoblotting, and stable isotope tracing technology. We show that ABCD3 functions in DCA metabolism and uncover a novel role for this peroxisomal transporter in lipid metabolic homeostasis. The Abcd3 KO mouse presents with lipodystrophy, increased circulating free fatty acids, decreased ketone bodies, enhanced hepatic cholesterol synthesis and decreased hepatic de novo lipogenesis. Moreover, our study suggests that DCAs are metabolized by mitochondrial {beta}-oxidation when ABCD3 is not functional, reflecting the importance of the metabolic compartmentalization and communication between peroxisomes and mitochondria. In summary, this study provides data on the role of the peroxisomal transporter ABCD3 in hepatic lipid homeostasis and DCA metabolism, and the consequences of peroxisomal dysfunction for the liver.

category: physiology


title: Model predicts the impact of root system architecture on soil water infiltration.

abstract: There is strong experimental evidence that root systems substantially change the saturated hydraulic conductivity of soil. However, the mechanisms by which roots affect soil hydraulic properties remain largely unknown. In this work, we made the hypothesis that preferential soil moisture transport occurs along the axes of roots, and that this is what changes a soils saturated hydraulic conductivity. We modified Richards equation to incorporate the preferential flow of soil moisture along the axes of roots. Using the finite element method and Bayesian optimisation, we developed a pipeline to calibrate our model with respect to a given root system. When applied to simulated root systems, the pipeline successfully predicted the pore-water pressure profiles corresponding to saturated hydraulic conductivity values, observed by Leung et al. (2018), for soils vegetated with willow and grass. Prediction accuracy improved for root systems with more realistic architectures, therefore suggesting that changes in saturated hydraulic conductivity are a result of roots enabling preferential soil moisture transport along their axes. The model proposed in this work improves our ability to predict moisture transport through vegetated soil and could help optimise irrigation, forecast flood events and plan landslide prevention strategies.


title: Regulatory start-stop elements in 5' untranslated regions pervasively modulate translation

abstract:
Translation includes initiation, elongation, and termination, followed by ribosome recycling. We characterize a new sequence element in 5 untranslated regions that consists of an adjacent start and stop codon and thereby excludes elongation. In these start-stop elements, an initiating ribosome is simultaneously positioned for termination without having translocated. At the example of activating transcription factor 4 (ATF4), we demonstrate that start-stops modify downstream re-initiation, thereby repressing translation of upstream open reading frames and enhancing ATF4s inducibility under stress. Start-stop elements are abundant in both mammals and yeast and affect key regulators such as DROSHA and the oncogenic transcription factor NFIA. They provide a unique regulatory layer that impedes ribosome scanning without the energy-expensive peptide production that accompanies upstream open reading frames.

One-Sentence SummaryRegulatory start-stop elements in the 5UTR are hitherto unappreciated contributors to 5 UTR code and alter the translatome.


title: Immunization with synthetic SARS-CoV-2 S glycoprotein virus-like particles protects Macaques from infection

abstract: The SARS-CoV-2 pandemic causes an ongoing global health crisis, which requires efficient and safe vaccination programs. Here, we present synthetic SARS-CoV2 S glycoprotein-coated liposomes that resemble in size and surface structure virus-like particles. Soluble S glycoprotein trimers were stabilized by formaldehyde cross-linking and coated onto lipid vesicles (S-VLP). Immunization of cynomolgus macaques with S-VLPs induced high antibody titers and TH1 CD4+ biased T cell responses. Although antibody responses were initially dominated by RBD specificity, the third immunization boosted non-RBD antibody titers. Antibodies showed potent neutralization against the vaccine strain and the Alpha variant after two immunizations and robust neutralization of Beta and Gamma strains. Challenge of animals with SARS-CoV-2 protected all vaccinated animals by sterilizing immunity. Thus, the S-VLP approach is an efficient and safe vaccine candidate based on a proven classical approach for further development and clinical testing.


title: Affinity maturation for an optimal balance between long-term immune coverage and short-term resource constraints

abstract: In order to target threatening pathogens, the adaptive immune system performs a continuous reorganization of its lymphocyte repertoire. Following an immune challenge, the B cell repertoire can evolve cells of increased specificity for the encountered strain. This process of affinity maturation generates a memory pool whose diversity and size remain difficult to predict. We assume that the immune system follows a strategy that maximizes the long-term immune coverage and minimizes the short-term metabolic costs associated with affinity maturation. This strategy is defined as an optimal decision process on a finite dimensional phenotypic space, where a pre-existing population of naive cells is sequentially challenged with a neutrally evolving strain. We unveil a trade-off between immune protection against future strains and the necessary reorganization of the repertoire. This plasticity of the repertoire drives the emergence of distinct regimes for the size and diversity of the memory pool, depending on the density of naive cells and on the mutation rate of the strain. The model predicts power-law distributions of clonotype sizes observed in data, and rationalizes antigenic imprinting as a strategy to minimize metabolic costs while keeping good immune protection against future strains.


title: Multivariate mining of an alpaca immune repertoire identifies potent cross-neutralising SARS-CoV-2 nanobodies

abstract: Conventional approaches to isolate and characterize nanobodies are laborious and cumbersome. Here we combine phage display, multivariate enrichment, and novel sequence analysis techniques to annotate an entire nanobody repertoire from an immunized alpaca. We combine this approach with a streamlined screening strategy to identify numerous anti-SARS-CoV-2 nanobodies, and use neutralization assays and Hydrogen/Deuterium exchange coupled to mass spectrometry (HDX-MS) epitope mapping to characterize their potency and specificity. Epitope mapping revealed that the binding site is a key determinant of neutralization potency, rather than affinity alone. The most potent nanobodies bind to the receptor binding motif of the RBD, directly preventing interaction with the host cell receptor ACE2, and we identify two exceptionally potent members of this category (with monomeric IC50s around 13 and 16 ng/ml). Other nanobodies bind to a more conserved epitope on the side of the RBD, and are able to potently neutralize the SARS-CoV-2 founder virus (42 ng/ml), the beta variant (B.1.351/501Y.V2) (35 ng/ml), and also cross-neutralize the more distantly related SARS-CoV-1 (0.46 g/ml). The approach presented here is well suited for the screening of phage libraries to identify functional nanobodies for various biomedical and biochemical applications.


title: Temporal Dynamics of HCMV Gene Expression in Lytic and Latent Infections

abstract: Primary infection with Human cytomegalovirus (HCMV) results in a persistent lifelong infection due to its ability to establish latent infection. During productive HCMV infection, viral genes are expressed in a coordinated cascade that is characteristic of all herpesviruses and traditionally relies on the dependencies of viral genes on protein synthesis and viral DNA replication. In contrast, the transcriptional landscape associated with HCMV latency is still disputed and poorly understood. Here, we examine viral transcriptomic dynamics during the establishment of both productive and latent HCMV infections. Our temporal measurements reveal that viral gene expression dynamics along productive infection and their dependencies on protein synthesis and viral DNA replication, do not fully align. This illustrates that the regulation of herpesvirus genes does not represent a simple sequential transcriptional cascade, and surprisingly, many viral genes are regulated by multiple independent modules. Using our improved classification of viral gene expression kinetics in conjunction with transcriptome-wide measurements of the effects of a wide array of chromatin modifiers, we unbiasedly show that a defining characteristic of latent cells is the unique repression of immediate early (IE) genes. Altogether, our findings provide an elaborate definition of HCMV gene expression patterns and reveal novel principles that govern viral gene expression in lytic and latent infection states.


title: Characterizing enterotypes in human metagenomics: a viral perspective

abstract: The diversity and high genomic mutation rates of viral species hinder our understanding of viruses and their contributions to human health. Here we investigated the human fecal virome using previously published sequencing data of 2,690 metagenomes from seven countries. We found that the virome was dominated by double-stranded DNA viruses, and young children and adults showed dramatic differences in their fecal enterovirus composition. Beta diversity showed there were significantly higher distances to centroids in individuals with severe phenotypes, such as cirrhosis. In contrast, there were no significant differences in lengths to centroids or viral components between patients with mild phenotypes, such as hypertension. Enterotypes showed the same specific viruses and enrichment direction after independent determination of enterotypes in various projects. Confounding factors, such as different sequencing platforms and library construction, did not result in a batch effect to confuse enterotype assignment. The gut virome composition pattern could be described by two viral enterotypes, which supported a discrete, rather than a gradient, distribution. Compared with enterotype 2, enterotype 1 had a higher viral count and Shannon index, but a lower beta diversity, indicating more resistance to the external environments harmful effects. Disease was usually accompanied by a viral enterotype disorder. However, a sample outside of the enterotyping mathematical space of enterotype database did not necessarily indicate sickness. Therefore, the background context must be carefully considered when using a viral enterotype as a biomarker for disease prediction. The disease, second only to the enterotype, explains significant variation in viral community composition, implying that double-stranded DNA is relevant to human health. Our results of investigating a baseline viral database highlight important insights into the virome composition of human ecosystems, and provide an alternate biomarker for early disease screening.


title: Increased aerosol transmission for B.1.1.7 (alpha variant) over lineage A variant of SARS-CoV-2

abstract: Airborne transmission, a term combining both large droplet and aerosol transmission, is thought to be the main transmission route of SARS-CoV-2. Here we investigated the relative efficiency of aerosol transmission of two variants of SARS-CoV-2, B.1.1.7 (alpha) and lineage A, in the Syrian hamster. A novel transmission caging setup was designed and validated, which allowed the assessment of transmission efficiency at various distances. At 2 meters distance, only particles <5 {micro}m traversed between cages. In this setup, aerosol transmission was confirmed in 8 out of 8 (N = 4 for each variant) sentinels after 24 hours of exposure as demonstrated by respiratory shedding and seroconversion. Successful transmission occurred even when exposure time was limited to one hour, highlighting the efficiency of this transmission route. Interestingly, the B.1.1.7 variant outcompeted the lineage A variant in an airborne transmission chain after mixed infection of donors. Combined, this data indicates that the infectious dose of B.1.1.7 required for successful transmission may be lower than that of lineage A virus. The experimental proof for true aerosol transmission and the increase in the aerosol transmission potential of B.1.1.7 underscore the continuous need for assessment of novel variants and the development or preemptive transmission mitigation strategies.


title: FH loss in RPE cells causes retinal degeneration in a human RPE-porcine retinal explant co-culture model

abstract: Age-related Macular degeneration (AMD) is a degenerative disease of the macula affecting the elderly population. Treatment options are limited, partly due to the lack of understanding of AMD pathology and the sparse availability of research models, that replicate the complexity of the human macula and the intricate interplay of the genetic, aging and life-style risk factors contributing to AMD. One of the main genetic risks associated with AMD is located on Complement Factor H (CFH) gene, leading to an amino acid substitution in the FH protein (Y402H). However, the mechanism of how this FH variant promotes the onset of AMD remains unclear. Previously, we have shown that FH deprivation in RPE cells, via CFH silencing, leads to increased inflammation, metabolic impairment and vulnerability towards oxidative stress. In this study, we established a novel co-culture model comprised of CFH silenced RPE cells and porcine retinal explants derived from the visual streak of the porcine eyes, closely resembling the human macula. We show that retinae exposed to FH-deprived RPE cells show signs of retinal degeneration, with rod cells being the first cells to undergo degeneration. Moreover, via Raman analyses, we observe that the main changes involve the mitochondria and lipid composition of the co-cultured retinae upon FH loss. Interestingly, the detrimental effects of FH loss in RPE cells on the neuroretina were independent of glial cell activation and external complement sources. Moreover, we show that the co-culture model is also suitable for human retinal explants, and we observed a similar trend when RPE cells deprived of FH were co-cultured with human retinal explants from a single donor eye. Our findings highlight the importance of RPE derived FH for retinal homeostasis and provide a valuable model for AMD research.

category: cell biology


title: The generation of a Bcl11a lineage tracing mouse model

abstract: The transcription factor B-cell lymphoma/leukaemia 11A (BCL11A) has essential functions in physiological processes as well as haematological and solid malignancies, however, its contribution to tissue development and tumour progression is poorly understood. Here we show the generation of a Bcl11a lineage tracing mouse model to allow for in vivo tracking of Bcl11a-expressing cells and their progeny. We validate the model in the mammary gland by using flow cytometry and whole-tissue 3D imaging to locate labelled cells after induction of tracing in early development. We show that Bcl11a is predominantly expressed in long-lived luminal progenitors which populate mammary alveoli upon pregnancy, confirming bona fide labelling of Bcl11a cells. The Bcl11a lineage tracing mouse model therefore provides a powerful resource to study Bcl11a cells in development, homeostasis, and cancer.


title: The benefits of merging passive and active tracking approaches: new insights into riverine migration by salmonid smolts

abstract: The process of smolting is a critical phase in the life-cycle of anadromous salmonids and it has been associated with substantial rates of mortality. Survival during freshwater and marine migration is known to have population level effects, thus an understanding of the patterns of mortality has the potential to yield important insights into population bottlenecks. Despite important advancements in tracking techniques, the specifics of mortality events in anadromous salmonids during their initial migration to sea remains somewhat elusive. Here, we develop a framework combining spatial and temporal detections of smolt riverine migration from two tracking techniques, which enable inferences to be made about mortality locations, causes, and rates. In this study, we demonstrate that during their initial riverine transitional phase, smolts were particularly vulnerable to predators. Specifically, avian predation appeared to be the main cause of mortality (42%), although piscine predation events were not trivial (14%). Our results suggested some direct and indirect tagging-induced mortality (e.g., through increased predation vulnerability), which highlights the importance of determining tagging mortality in a telemetry study to ensure adequate interpretation of migration success. Overall, by estimating migration loss and its variability, our study framework should help to guide management actions to mitigate the widespread population declines these species are currently facing.


title: Different contribution of the monkey prefrontal and premotor dorsal cortex in decision-making supported by inferential reasoning

abstract: Several studies have reported similar neural modulations between brain areas of the frontal cortex, such as the dorsolateral prefrontal (DLPFC) and the premotor dorsal (PMd) cortex, in tasks requiring encoding of the abstract rules for selecting the proper action. Here we compared the neuronal modulation of the DLPFC and PMd of monkeys trained to choose the higher rank from a pair of abstract images (target item), selected from an arbitrarily rank-ordered set (A>B>C>D>E>F) in the context of a transitive inference task. Once acquired by trial-and-error, the ordinal relationship between pairs of adjacent images (i.e., A>B; B>C; C>D; D>E; E>F), monkeys were tested in indicating the ordinal relation between items of the list not paired during learning. During these decisions, we observed that the choice accuracy increased and the reaction time decreased as the rank difference between the compared items enhanced. This result is in line with the hypothesis that after learning, the monkeys built an abstract mental representation of the ranked items, where rank comparisons correspond to the items position comparison on this representation. In both brain areas, we observed higher neuronal activity when the target item appeared in a specific location on the screen with respect to the opposite position and that this difference was particularly enhanced at lower degrees of difficulty. By comparing the time evolution of the activity of the two areas, we observed that the neural encoding of target item spatial position occurred earlier in the DLPFC than in the PMd.

category: animal behavior and cognition


title: Different contribution of the monkey prefrontal and premotor dorsal cortex in decision making during a transitive inference task


title: Correlational networking guides the discovery of cryptic proteases for lanthipeptide maturation

abstract:
Bacterial natural product biosynthetic genes, canonically clustered, have been increasingly found to rely on cryptic enzymes encoded elsewhere in the genome for completion of biosynthesis. The study and application of lanthipeptides are frequently hindered by cryptic proteases required for final maturation. Here, we establish a global correlation network bridging the gap between lanthipeptide precursors and cryptic proteases. Applying our analysis to 161,954 bacterial genomes, we establish 5,209 correlations between precursors and cryptic proteases, with 91 prioritized. We use network predictions and co-expression analysis to reveal a previously missing protease for the maturation of class I lanthipeptide paenilan. We further discover widely distributed bacterial M16B metallopeptidases of previously unclear biological function as a new family of lanthipeptide proteases. We show the involvement of a pair of bifunctional M16B proteases in the production of novel class III lanthipeptides with high substrate specificity. Together, these results demonstrate the strength of our correlational networking approach to the discovery of cryptic lanthipeptide proteases and potentially other missing enzymes for natural products biosynthesis.

Graphical abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY


title: Correlational networking guides the discovery of cryptic natural product biosynthetic enzymes


title: Using topic modeling to detect cellular crosstalk in scRNA-seq

abstract:
Cell-cell interactions are vital for numerous biological processes including development, differentiation, and response to inflammation. Currently most methods for studying interactions on scRNA-seq level are based on curated databases of ligands and receptors. While those methods are useful, they are limited to our current biological knowledge. Recent advances in single cell protocols have allowed for physically interacting cells to be captured and as such we have the potential to study interactions in a complimentary way without relying on prior knowledge. We introduce a new method for detecting genes that change as a result of interaction based on Latent Dirichlet Allocation (LDA). We apply our method to synthetic datasets to demonstrate its ability to detect genes that change in an interacting population compared to a reference population. Next, we apply our approach to two datasets of physically interacting cells and identify genes that change as a result of interaction, examples include adhesion and co-stimulatory molecules which confirm physical interaction between cells. For each dataset we produce a ranking of genes that are changing in subpopulations of the interacting cells. In addition to genes, discussed in the original publications we highlight further candidates for interaction in the top 100 and 300 ranked genes. Lastly, we apply our method to a dataset generated by a standard droplet based protocol, not designed to capture interacting cells and discuss its suitability for analysing interactions. We present a method that streamlines the detection of interactions and does not require prior clustering and generation of synthetic reference profiles to detect changes in expression.

Author summaryWhile scRNA-seq research is a dynamic area, progress is lacking when it comes to developing methods that allow analysis of interaction that is independent of curated resources of known interacting pairs. Recent advances of sequencing protocols have allowed for interacting cells to be captured. We propose a novel method based on LDA that captures changes in gene expression as a result of interaction. Our method does not require prior information in the form of clustering or generation of synthetic reference profiles. We demonstrate the suitability of our approach by applying it to synthetic and real datasets and manage to capture biologically interesting candidates of interaction.

category: bioinformatics


title: SIRV: Spatial inference of RNA velocity at the single-cell resolution

abstract: Studying cellular differentiation using single-cell RNA sequencing (scRNA-seq) rapidly expands our understanding of cellular development processes. Recently, RNA velocity has created new possibilities in studying these cellular differentiation processes, as differentiation dynamics can be obtained from measured spliced and unspliced mRNA expression. However, to map these differentiation processes to developments within a tissue, the spatial context of the tissue should be taken into account, which is not possible with current approaches as they start from dissociated cells. We present SIRV (Spatially Inferred RNA Velocity), a method to infer spatial differentiation trajectories within the spatial context of a tissue at the single-cell resolution. SIRV integrates spatial transcriptomics data with reference scRNA-seq data, to enrich the spatially measured genes with spliced and unspliced expressions from the scRNA-seq data. Next, SIRV calculates RNA velocity vectors for every spatially measured cell and maps these vectors to the spatial coordinates within the tissue. We tested SIRV on the Developing Mouse Brain Atlas data and obtained biologically relevant spatial differentiation trajectories. Additionally, SIRV annotates spatial cells with cellular identities and the region of origin which are transferred from the annotated reference scRNA-seq data. Altogether, with SIRV, we introduce a new tool to enrich spatial transcriptomics data that can assist in understanding how tissues develop.


title: AMaLa: analysis of Directed Evolution experiments via Annealed Mutational approximated Landscape

abstract:
We present Annealed Mutational approximated landscape (AMaLa), a new method to infer fitness landscapes from Directed Evolution experiment sequencing data. Directed Evolution experiments typically start from a single wild-type sequence, which undergoes Darwinian in vitro evolution acted via multiple rounds of mutation and selection with respect to a target phenotype. In the last years, Directed Evolution is emerging as a powerful instrument to probe fitness landscapes under controlled experimental condition and, thanks to the use of high-throughput sequencing of the different rounds, as a relevant testing ground to develop accurate statistical models and inference algorithms.

Fitness landscape modeling strategies, either use as input data the enrichment of variants abundances and hence require observing the same variants at different rounds, or they simply assume that the variants at the last sequenced round are the results of a sampling process at equilibrium. AMaLa aims at leveraging effectively the information encoded in the time evolution of all sequenced rounds. To do so, on the one hand we assume statistical sampling independence between sequenced rounds, and on the other we gauge all possible trajectories in sequence space with a time-dependent statistical weight consisting of two contributions: (i) a statistical energy term accounting for the selection process, (ii) a simple generalized Jukes-Cantor model to describe the purely mutational step.

This simple scheme allows us to accurately describe the Directed Evolution dynamics in a concrete experimental setup and to infer a fitness landscape that reproduces correctly the measures of the phenotype under selection (e.g. antibiotic drug resistance), notably outperforming widely used inference strategies. We assess the reliability of AMaLa by showing how the inferred statistical model could be used to predict relevant structural properties of the wild-type sequence, and to reproduce the mutational effects of large scale functional screening not used to train the model.

category: biophysics


title: ETAS(R)50 Attenuates SARS-CoV-2 Spike Protein-Induced IL-6 and IL-1β Production by Suppressing p44/42 MAPK and Akt Phosphorylation in Murine Primary Macrophages

abstract: Excessive host inflammation following infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is associated with severity and mortality in coronavirus disease 2019 (COVID-19). We recently reported that the SARS-CoV-2 spike protein S1 subunit (S1) induces pro-inflammatory responses by activating toll-like receptor 4 (TLR4) signaling in macrophages. ETAS(R)50, a standardized extract of Asparagus officinalis stem, is a unique functional food that elicits anti-photoaging effects by suppressing pro-inflammatory signaling in hydrogen peroxide- and ultraviolet B-exposed skin fibroblasts. To elucidate its potential in preventing excessive inflammation in COVID-19, we examined the effects of ETAS(R)50 on pro-inflammatory responses in S1-stimulated murine peritoneal exudate macrophages. Co-treatment of the cells with ETAS(R)50 significantly attenuated S1-induced secretion of interleukin (IL)-6 in a concentration-dependent manner without reducing cell viability. ETAS(R)50 also markedly suppressed the S1-induced transcription of IL-6 and IL-1{beta}. However, among the TLR4 signaling proteins, ETAS(R)50 did not affect the degradation of inhibitor {kappa}B, nuclear translocation of nuclear factor-{kappa}B p65 subunit, and phosphorylation of c-Jun N-terminal kinase p54 subunit after S1 exposure. In contrast, ETAS(R)50 significantly suppressed S1-induced phosphorylation of p44/42 mitogen-activated protein kinase (MAPK) and Akt. Attenuation of S1-induced transcription of IL-6 and IL-1{beta} by the MAPK kinase inhibitor U0126 was greater than that by the Akt inhibitor perifosine, and the effects were potentiated by simultaneous treatment with both inhibitors. These results suggest that ETAS(R)50 attenuates S1-induced IL-6 and IL-1{beta} production by suppressing p44/42 MAPK and Akt signaling in macrophages. Therefore, ETAS(R)50 may be beneficial in regulating excessive inflammation in patients with COVID-19.

category: pharmacology and toxicology


title: Inseparable color and form in mouse visual cortex described by random pooling from rods and cones

abstract: Spatial transitions in color can aid any visual perception task, and its neural representation - the "integration of color and form" - is thought to begin at primary visual cortex (V1). An integration of color and form is untested in mouse V1, yet studies show that the ventral retina provides the necessary substrate from green-sensitive rods and UV-sensitive cones. Here, we used two-photon imaging in V1 to measure spatial frequency (SF) tuning along four axes of rod and cone contrast space, including luminance and color. We first reveal that V1s sensitivity to color is similar to luminance, yet average SF tuning is significantly shifted lowpass for color. Next, guided by linear models, we used SF tuning along all four color axes to estimate the proportion of neurons that fall into classic models of color opponency - "single-", "double-", and "non-opponent". Few neurons (~6%) fit criteria for double-opponency, which are uniquely tuned for chromatic borders. Most of the population can be described as a unimodal distribution ranging from strongly single-opponent to non-opponent. Consistent with recent studies of the rodent and primate retina, our V1 data is well-described by a simple model in which ON and OFF channels to V1 sample the photoreceptor mosaic randomly.

category: neuroscience


title: Joint representations of color and form in mouse visual cortex described by random pooling from rods and cones


title: Caldendrin represses neurite regeneration via a sex-dependent mechanism in sensory neurons

abstract: Caldendrin is a calmodulin-like Ca2+ binding protein that is expressed primarily in neurons and regulates multiple effectors including Cav1 L-type Ca2+ channels. Here, we tested the hypothesis that caldendrin regulates Cav1-dependent pathways that repress neurite growth in dorsal root ganglion neurons (DRGNs). By immunofluorescence, caldendrin was localized in medium- and large-diameter DRGNs. Consistent with an inhibitory effect of caldendrin on neurite growth, neurite initiation and growth was enhanced in dissociated DRGNs from caldendrin knockout (KO) mice compared to those from wild type (WT) mice. In an in vitro axotomy assay, caldendrin KO DRGNs grew longer neurites via a mechanism that was more sensitive to inhibitors of transcription as compared to WT DRGNs. Strong depolarization, which normally represses neurite growth through activation of Cav1 channels, had no effect on neurite growth in DRGN cultures from female caldendrin KO mice. Remarkably, DRGNs from caldendrin KO males were no different from those of WT males in terms of depolarization-dependent neurite growth repression. We conclude that caldendrin opposes neurite regeneration and growth, and this involves coupling of Cav1 channels to growth-inhibitory pathways in DRGNs of females but not males. Our findings suggest that caldendrin KO mice represent an ideal model in which to interrogate the transcriptional pathways controlling neurite regeneration and how these pathways may differ in males and females.


title: Untangling Microbiota Diversity and Assembly Patterns in the World's Largest Water Diversion Canal

abstract: Large water diversion projects are important constructions for reallocation of human-essential water resources. Deciphering microbiota dynamics and assembly mechanisms underlying canal water ecosystem services especially during long-distance diversion is the prerequisite for water quality monitoring, biohazard warning and sustainable management. Using a 1432-km canal of the South-to-North Water Diversion Projects as a model system, we answer three central questions: how bacterial and micro-eukaryotic communities spatio-temporally develop, how much ecological stochasticity contributes to microbiota assembly, and which immigrating populations better survive and navigate across the canal. We applied quantitative ribosomal RNA gene sequence analyses to investigate canal water microbial communities sampled over a year, as well as null model- and neutral model-based approaches to disentangle the microbiota assembly processes. Our results showed clear microbiota dynamics in community composition driven by seasonality more than geographic location, and seasonally dependent influence of environmental parameters. Overall, bacterial community was largely shaped by deterministic processes, whereas stochasticity dominated micro-eukaryotic community assembly. We defined a local growth factor (LGF) and demonstrated its innovative use to quantitatively infer microbial proliferation, unraveling taxonomically dependent population response to local environmental selection across canal sections. Using LGF as a quantitative indicator of immigrating capacities, we also found that most micro-eukaryotic populations (82%) from the source lake water sustained growth in the canal and better acclimated to the hydrodynamical water environment than bacteria (67%). Taxa inferred to largely propagate include Limnohabitans sp. and Cryptophyceae, potentially contributing to water auto-purification. Combined, our work poses first and unique insights into the microbiota assembly patterns and dynamics in the worlds largest water diversion canal, providing important ecological knowledge for long-term sustainable water quality maintenance in such a giant engineered system.

category: microbiology


title: Protection of human ACE2 transgenic Syrian hamsters from SARS CoV-2 variants by human polyclonal IgG from hyper-immunized transchromosomic bovines

abstract: Pandemic SARS CoV-2 has been undergoing rapid evolution during spread throughout the world resulting in the emergence of many Spike protein variants, some of which appear to either evade antibody neutralization, transmit more efficiently, or potentially exhibit increased virulence. This raises significant concerns regarding the long-term efficacy of protection elicited after primary infection and/or from vaccines derived from single virus Spike (S) genotypes, as well as the efficacy of anti-S monoclonal antibody based therapeutics. Here, we used fully human polyclonal human IgG (SAB-185), derived from hyperimmunization of transchromosomic bovines with DNA plasmids encoding the SARS-CoV-2 Wa-1 strain S protein or purified ectodomain of S protein, to examine the neutralizing capacity of SAB-185 in vitro and the protective efficacy of passive SAB-185 antibody (Ab) transfer in vivo. The Ab preparation was tested for neutralization against five variant SARS-CoV-2 strains: Munich (Spike D614G), UK (B.1.1.7), Brazil (P.1) and SA (B.1.3.5) variants, and a variant isolated from a chronically infected immunocompromised patient (Spike {Delta}144-146). For the in vivo studies, we used a new human ACE2 (hACE2) transgenic Syrian hamster model that exhibits lethality after SARS-Cov-2 challenge and the Munich, UK, SA and {Delta}144-146 variants. SAB-185 neutralized each of the SARS-CoV-2 strains equivalently on Vero E6 cells, however, a control convalescent human serum sample was less effective at neutralizing the SA variant. In the hamster model, prophylactic SAB-185 treatment protected the hamsters from fatal disease and minimized clinical signs of infection. These results suggest that SAB-185 may be an effective treatment for patients infected with SARS CoV-2 variants.


title: Counting actin in contractile rings reveals novel contributions of cofilin and type II myosins to fission yeast cytokinesis

abstract: Cytokinesis by animals, fungi and amoebas depends on actomyosin contractile rings, which are stabilized by continuous turnover of actin filaments. Remarkably little is known about the amount of polymerized actin in contractile rings, so we used low concentration of GFP-Lifeact to count total polymerized actin molecules in the contractile rings of live fission yeast cells. Contractile rings of wild-type cells accumulated polymerized actin molecules at 4,900/min to a peak number of [~]198,000 followed by a loss of actin at 5,400/min throughout ring constriction. In adf1-M3 mutant cells with cofilin that severs actin filaments poorly, contractile rings accumulated polymerized actin at twice the normal rate and eventually had almost two-fold more actin along with a proportional increase in type II myosins Myo2, Myp2 and formin Cdc12. Although 30% of adf1-M3 mutant cells failed to constrict their rings fully, the rest lost actin from the rings at the wild-type rates. Mutations of type II myosins Myo2 and Myp2 reduced contractile ring actin filaments by half and slowed the rate of actin loss from the rings.


title: Correcting machine learning models using calibrated ensembles with "mlensemble"

abstract: Machine learning models in bioinformatics are often trained and used within the scope of a single project, but some models are also reused across projects and deployed in translational settings. Over time, trained models may turn out to be maladjusted to the properties of new data. This creates the need to improve their performance under various constraints. This work explores correcting models without retraining from scratch and without accessing the original training data. It uses a taxonomy of strategies to guide the development of a software package,  mlensemble. Key features include joining heterogeneous models into ensembles and calibrating ensembles to the properties of new data. These are well-established techniques but are often hidden within more complex tools. By exposing them to the application level, the package enables analysts to use expert knowledge to adjust models whenever needed. Calculations with imaging data show benefits when the noise characteristics of the training and the application datasets differ. An example using genomic single-cell data demonstrates model portability despite batch effects. The generality of the framework makes it applicable also in other subject domains.


title: Trehalose induces tomato defenses and increases drought tolerance and bacterial wilt disease resistance

abstract: Ralstonia solanacearum causes bacterial wilt disease, leading to severe crop losses. Xylem sap from R. solanacearum-infected tomato is enriched in the disaccharide trehalose. Water-stressed plants also accumulate trehalose, which increases drought tolerance via abscisic acid (ABA) signaling. Because R. solanacearum-infected plants suffer reduced water flow, we hypothesized that bacterial wilt physiologically mimics drought stress, which trehalose could mitigate. We found that R. solanacearum-infected plants differentially expressed drought-associated genes, including those involved in ABA and trehalose metabolism, and had more ABA in xylem sap. Consistent with this, treating tomato roots with ABA reduced both stomatal conductance and stem colonization by R. solanacearum. Treating roots with trehalose increased xylem sap ABA and reduced plant water use by lowering stomatal conductance and temporarily improving water use efficiency. Trehalose treatment also upregulated expression of salicylic acid (SA)-dependent tomato defense genes; increased xylem sap levels of SA and other antimicrobial compounds; and increased bacterial wilt resistance of SA-insensitive NahG tomato plants. Additionally, trehalose treatment increased xylem concentrations of jasmonic acid and related oxylipins. Finally, trehalose-treated plants were substantially more resistant to bacterial wilt disease. Together, these data show that exogenous trehalose reduced both water stress and bacterial wilt disease and triggered systemic disease resistance, possibly through a Damage Associated Molecular Pattern (DAMP) response pathway. This suite of responses revealed unexpected linkages between plant responses to biotic and abiotic stress and suggested that R. solanacearum-infected plants increase trehalose to improve water use efficiency and increase wilt disease resistance. The pathogen may degrade trehalose to counter these efforts. Together, these results suggest that treating tomatoes with exogenous trehalose could be a practical strategy for bacterial wilt management.

category: plant biology


title: Trehalose increases tomato drought tolerance, induces defenses, and increases resistance to bacterial wilt disease