title: Cover crop residue diversity enhances microbial activity and biomass with additive effects on microbial structure

abstract: Cover crops have been widely used in agroecosystems to improve soil fertility and environmental sustainability. The decomposition of cover crop residues can have further effects on belowground communities and their activity, which is important for a series of soil functions (e.g., nutrient cycling and organic matter decomposition). We tested the effect of plant residues from a range of cover crop species on soil microbial activity and community assemblage. We predicted that cover crop residues would alter the soil microbial community and that a greater diversity of residues would enhance microbial decomposition. In an incubation study, we assessed the effect of crop residue diversity on microbial activity (soil respiration) and its consequent effects on microbial community composition (PLFA). We used either a biodiverse mixture of four cover crop residues (buckwheat, clover, sunflower and radish) or an equal mass of the residues of each of the individual species. The diverse mixture of cover crop residues had a significantly (P < 0.05) greater soil respiration rate, by 57.61 {micro}g C g-1 h-1, than the average of the four individual residues, but did not have a significantly different soil microbial biomass or microbial community structure. This finding could be attributed to a greater diversity of organic resources increasing the number biochemical niches, and hence activating dormant microbial communities to increase microbial activity without affecting microbial biomass or community composition. Greater respiration from similar microbial biomasses suggests that microbial activity might be more efficient after a more diverse substrate input. This study confirms the positive impact of cover crop residues on soil microbial biomass and activity and highlights that mixtures of cover crop residues may deliver enhanced soil functions beyond the sum of individual cover crop residues.


title: Recognition and Differentiation of Dural Puncture Click Sensation: A Subjective and Objective Prospective Study of Dural Puncture Forces Using Fine-Gauge Spinal Needles

abstract:
BackgroundWe hypothesized that the click perceived when puncturing the dura-arachnoid with fine-gauge spinal needles can be subjectively identified, and investigated whether it may be distinguishable among different needle types.

MethodsSubjective and objective evaluations were performed. First, physicians punctured the polyamide film or porcine dura mater (n=70 and n=20, respectively) with seven types of spinal needles and numerically evaluated the perceived click sensations. Using an 11-point numerical rating scale (from "0" for "no click sensation" to "10" for "the strongest click perceived") data, subjective differentiation among needle types was assessed. Second, in the objective part of the study, total forces elicited by polyamide film or porcine dura mater punctures with each needle were measured using a biomechanical testing device, and load-displacement curves evaluated. Third, the results of subjective and objective evaluations were compared.

ResultsAll participants recognized the click and could discriminate among needles of different tip shape. The load-displacement curves for polyamide film and porcine dura mater were similar and needle-specific. The subjective numerical rating scale values corresponded well with the objectively measured changes in total forces (R2 = 0.862 and R2 = 0.881 for polyamide film and porcine dura mater, respectively), indicating that an increase in the largest drop in total force value of 0.30 N or 0.21 N would produce an increase of numerical rating scale value of 1 for polyamide film and porcine dura mater, respectively.

ConclusionsWe provide an objective proof of the click sensation felt upon dural puncture using different fine-gauge spinal needles. Click recognition could be used as an additional indicator of successful spinal puncture.

category: biophysics


title: Automatic Detection of Larval Zebrafish ECG: Computational Tool for High-throughput Cardiac Activity Analysis

abstract: Automatic analysis of larval zebrafish electrocardiographs (ECG) is essential for high-throughput measurements in environmental toxicity assays, cardiotoxicity measurements and drug screening. We have developed a MATLAB based software is built on methods that have previously been used to analyze human ECG, such as the Pan-Tompkins algorithm and Wavelet. For the first time these sophisticated tools have been applied to larval zebrafish ECG to automatically characterize the heart-beat waveforms. The ability of the automated algorithm to detect the QT interval for both normal and pharmacologically altered larval ECG is found and compared to previously used software that is built into LabChart(R) (AD Instruments). Using wavelet transforms it is shown that the typical larval ECG features are within the frequency range of 1 to 31 Hz. It is also shown that the automated software is capable of detecting QTc (heartrate corrected heartbeat interval) changes within pharmacologically altered zebrafish larval ECG. The automated process is a significant improvement on the approaches that were previously applied to the zebrafish ECG. The automated process described here that is based on established techniques of analyzing ECG can sensitively measure pharmacologically induced changes in the ECG. The novel, automated software is faster, more sensitive at detecting ECG changes and less dependent on user involvement, thus minimizing user error and human bias. The automated process can also be applied to human ECG.

category: physiology


title: Acoustic context modulates natural sound discrimination in auditory cortex through frequency specific adaptation

abstract: Vocal communication is essential to coordinate social interactions in mammals and it requires a fine discrimination of communication sounds. Auditory neurons can exhibit selectivity for specific calls, but how it is affected by preceding sounds is still debated. We tackled this using ethologically relevant vocalizations in a highly vocal mammalian species: Sebas short-tailed bat. We show that cortical neurons present several degrees of selectivity for echolocation and distress calls. Embedding vocalizations within natural acoustic streams leads to stimulus-specific suppression of neuronal responses that changes sound selectivity in disparate manners: increases in neurons with poor discriminability in silence and decreases in neurons selective in silent settings. A computational model indicates that the observed effects arise from two forms of adaptation: presynaptic frequency specific adaptation acting in cortical inputs and stimulus unspecific postsynaptic adaptation. These results shed light into how acoustic context modulates natural sound discriminability in the mammalian cortex.

category: neuroscience


title: Enhanced habit formation in Tourette syndrome : dopamine release and striatal disinhibition modulate shortcut connections in a hierarchical model of cortico-basal ganglia loops

abstract: In the Gilles de la Tourette syndrome, tics are often considered as habitual responses towards unwanted premonitory urges. Support for the relationship between tics and habits comes from devaluation protocols, which reveal that unmedicated Tourette patients show an increased tendency towards responses to devalued outcomes. We use a neuro-computational model of hierarchically organized cortico-basal ganglia-thalamo-cortical loops to shed more light on enhanced habit formation of Tourette patients. In our model, habitual behavior emerges from cortico-thalamic shortcut connections, where enhanced habit formation can be linked to faster plasticity in the shortcut or to a stronger feedback from the shortcut to the basal ganglia. Irregular activity in such shortcut connections may have different pathophysiological origins. Based on our model, we explore decreased local striatal inhibition, which may correspond to a loss of inhibitory interneurons, and increased dopaminergic modulation of striatal medium spiny neurons as causes for irregular shortcut plasticity or activation. Both lead to higher rates of response towards devalued outcomes in our model, similar to what is observed in Tourette patients. Our results support the view of tics in Tourette syndrome as maladaptive habits. We suggest to reveal more shortcuts between cortico-basal ganglia-thalamo-cortical loops in the human brain and study their potential role in the development of the Tourette syndrome.


title: Sensory experience modulates the reorganisation of temporal auditory regions for executive processing

abstract: Crossmodal plasticity refers to the reorganisation of sensory cortices in the absence of their main sensory input. Understanding this phenomenon provides insights into brain function and its potential for change and enhancement. Using fMRI, we investigated how early deafness influences crossmodal plasticity and the organisation of executive functions in the adult human brain. Results from a range of visual executive function tasks (working memory, task switching, planning, inhibition) show that deaf individuals specifically recruit "auditory" regions during task switching. Neural activity in superior temporal regions, most significantly in the right hemisphere, are good predictors of behavioural performance during task switching in the deaf group, highlighting the functional relevance of the observed cortical reorganisation. Our results show executive processing in typically sensory regions, suggesting that the development and ultimate role of brain regions are influenced by perceptual environmental experience.


title: Rare long-range cortical connections enhance information processing

abstract: What are the key topological features of connectivity critically relevant for generating the dynamics underlying efficient cortical function? A candidate feature that has recently emerged is that the connectivity of the mammalian cortex follows an exponential distance rule, which includes a small proportion of long-range high-weight anatomical exceptions to this rule. Whole-brain modelling of large-scale human neuroimaging data in 1003 participants offers the unique opportunity to create two models with and without long-range exceptions and explicitly study their functional consequences. We found that rare long-range exceptions are crucial for significantly improving information processing. Furthermore, modelling in a simplified ring architecture shows that this improvement is greatly enhanced by the turbulent regime found in empirical neuroimaging data. Overall, the results provide strong empirical evidence for the immense functional benefits of long-range exceptions combined with turbulence for information processing.


title: Simultaneous functional MRI of socially interacting marmosets

abstract: Social cognition is a dynamic process that requires the perception and integration of a complex set of idiosyncratic features between interacting conspecifics. Here we present a method for simultaneously measuring the whole-brain activation of two socially interacting marmoset monkeys using functional magnetic resonance imaging. MRI hardware (a radiofrequency coil and peripheral devices) and image-processing pipelines were developed to assess brain responses to socialization, both on an intra-brain and inter-brain level. Notably, brain-activation maps acquired during constant interaction demonstrated neuronal synchrony between marmosets in regions of the brain responsible for processing social interaction. This method enables a wide range of possibilities for studying social function and dysfunction in a non-human primate model, including using transgenic models of neuropsychiatric disorders.


title: Unexpected distribution of the 4-formylaminooxyvinylglycine (FVG) biosynthetic pathway in Pseudomonas and beyond

abstract: The biological herbicide and antibiotic 4-formylaminooxyvinylglycine (FVG) was originally isolated from several rhizosphere-associated strains of Pseudomonas fluorescens. Biosynthesis of FVG is dependent on the gvg biosynthetic gene cluster in P. fluorescens. In this investigation, we used comparative genomics to identify strains with the genetic potential to produce FVG due to presence of a gvg gene cluster. These strains primarily belong to two groups of Pseudomonas, P. fluorescens and P. syringae, however, a few strains with the gvg cluster were found outside of Pseudomonas. Mass spectrometry confirmed that all tested strains of the P. fluorescens species group produced FVG. However, P. syringae strains did not produce FVG under standard conditions. Several lines of evidence regarding the transmission of the gvg cluster including a robust phylogenetic analysis suggest that it was introduced multiple times through horizontal gene transfer within the Pseudomonas lineage as well as in select lineages of Thiomonas, Burkholderia and Pantoea. Together, these data broaden our understanding of the evolution and diversity of FVG biosynthesis. In the course of this investigation, additional gene clusters containing only a subset of the genes required to produce FVG were identified in a broad range of bacteria, including many non-pseudomonads.

category: microbiology


title: Calculating likelihoods and likelihood ratios at SNPs-based mixtures. A reappraisal of the binomial inference, as applied to forensic identity tests

abstract: Single nucleotide polymorphisms (SNPs) are useful forensic markers. When a SNPs-based forensic protocol targets a body fluid stain, it returns elementary evidence regardless of the number of individuals that might have contributed to the stain deposition. Therefore, drawing inference from a mixed stain with SNPs is different than drawing it while using multinomial polymorphisms. We here revisit this subject, with a view to contribute to a fresher insight into it. First, we manage to model conditional semi-continuous likelihoods in terms of matrices of genotype permutations vs number of contributors (NTZsc). Secondly, we redefine some algebraic formulas to approach the semi-continuous calculation. To address allelic dropouts, we introduce a peak height ratio index ( h, or: the minor read divided by the major read at any NGS-based typing result) into the semi-continuous formulas, for they to act as an acceptable proxy of the  split drop (Haned et al, 2012) model of calculation. Secondly, we introduce a new, empirical method to deduct the expected quantitative ratio at which the contributors of a mixture have originally mixed and the observed ratio generated by each genotype combination at each locus. Compliance between observed and expected quantity ratios is measured in terms of (1-{chi}2) values at each state of a locus deconvolution. These probability values are multiplied, along with the h index, to the relevant population probabilities to weigh the overall plausibility of each combination according to the quantitative perspective. We compare calculation performances of our empirical procedure (NITZq) with those of the EUROFORMIX software ver. 3.0.3. NITZq generates LR values a few orders of magnitude lower than EUROFORMIX when true contributors are used as POIs, but much lower LR values when false contributors are used as POIs. NITZ calculation routines may be useful, especially in combination with mass genomics typing protocols.

category: genetics


title: Whole genome 6-methyladenosine sequencing (6-mA-Seq) enables bacterial epigenomics studies.

abstract: Methylation sequencing using bisulfite treatment has revolutionized the field of molecular biology for eukaryotic systems, unveiling levels of intricacy in regulation of gene expression in response to different environmental conditions. While bacteria also utilize methylation to regulate gene expression, bisulfite sequencing does not work as well in bacteria as in eukaryotes, because bacteria methylate adenosine instead of cytosine. Therefore, global bacterial methylation patterns cannot be studied using common Illumina sequencers. In this work, we demonstrate 6mA-seq, a method that can be used to identify patterns in bacteria that methylate adenosines at GATC sites. Furthermore, this method was used on Escherichia coli cultured on four different carbon sources to demonstrate different methylation patterns due to carbon utilization. 6mA-seq can increase the speed in which epigenetic research is conducted in bacteria.

category: genomics


title: 5-Azacytidine: Effects on the Expression of alpha-Cardiac Actin in Pericytes from Human Adipose Tissue

abstract: DNA methylation patterns are closely related to the chromatin structure, and its remodeling is considered an important mechanism in the control of gene transcription during cell differentiation. In rodent, several studies have related the possibility that multipotent mesenchymal stromal cells (MSCs) undergo cardiomyogenesis. However, it has not been completely elucidated if human adult stem cell exhibits true differentiation potential for a cardiac lineage. In this study, the action of the DNA methylation inhibitor 5-azacytidine (5-aza) was examined in human adipose tissue pericytes (hATPCs: 3G5+) regarding their possible capacity to induce myocytes in vitro. Real-Time PCR revealed that cells treated with 5-aza presented time-dependent decrease in the mRNA expression of -cardiac actin (-CA). At 24 h, this diminution was statistically significant; however, there was not a correlation with the highest level of DNA demethylation at the same period using Methylation-Sensitive High Resolution Melting-PCR (MS-HRM-PCR). An evident increase in the -CA protein expression was observed by Western blotting in hATPCs treated with 5-aza at 24 h. The mRNA expression of -SMA (-smooth actin) also showed a time-dependent decrease after the treatment, however, it was not significant. The ultrastructural analysis showed similar structures such as like-cell junctions, caveolae, and actin myofilaments, which aligned in parallel. These phenotypic alterations were found only after the treatment; however, the hTAPCs after 5-aza treatment were not able to form thick myofilaments and consequently sarcomeres. These results indicated that a terminal cardiac differentiation of hTAPCs was not achieved and that the cardiomyogenesis failure could be related to the non-muscle origin of the adipose tissue.

category: cell biology


title: Investigating receptor-mediated antibody transcytosis using blood-brain barrier organoid arrays

abstract:
BackgroundThe pathways that control protein transport across the Blood-Brain Barrier (BBB) remain poorly characterized. Despite great advances in recapitulating the human BBB in vitro, current models are not suitable for systematic analysis of the molecular mechanisms of antibody transport. The gaps in our mechanistic understanding of antibody transcytosis hinder new therapeutic delivery strategy development.

MethodsWe applied a novel bioengineering approach to generate human BBB organoids by the self-assembly of astrocytes, pericytes and brain endothelial cells with unprecedented throughput and reproducibility using micro patterned hydrogels. We designed a semi-automated and scalable imaging assay to measure receptor-mediated transcytosis of antibodies. Finally, we developed a workflow to use CRISPR/Cas9 gene editing in BBB organoid arrays to knock out regulators of endocytosis specifically in brain endothelial cells in order to dissect the molecular mechanisms of receptor-mediated transcytosis.

ResultsBBB organoid arrays allowed the simultaneous growth of more than 5000 homogenous organoids per individual experiment in a highly reproducible manner. BBB organoid arrays showed low permeability to macromolecules and prevented transport of human non-targeting antibodies. In contrast, a monovalent antibody targeting the human transferrin receptor underwent dose- and time-dependent transcytosis in organoids. Using CRISPR/Cas9 gene editing in BBB organoid arrays, we showed that clathrin, but not caveolin, is required for transferrin receptor-dependent transcytosis.

ConclusionsHuman BBB organoid arrays are a robust high-throughput platform that can be used to discover new mechanisms of receptor-mediated antibody transcytosis. The implementation of this platform during early stages of drug discovery can accelerate the development of new brain delivery technologies.


title: Pkd1 mutation has no apparent effects on peroxisome structure or lipid metabolism

abstract:
BackgroundMultiple studies of tissue and cell samples from patients and pre-clinical models of autosomal dominant polycystic kidney disease report abnormal mitochondrial function and morphology and suggest metabolic reprogramming is an intrinsic feature of this disease. Peroxisomes interact with mitochondria physically and functionally, and congenital peroxisome biogenesis disorders can cause various phenotypes, including mitochondrial defects, metabolic abnormalities and renal cysts. We hypothesized that a peroxisomal defect might contribute to the metabolic and mitochondrial impairments observed in autosomal dominant polycystic kidney disease.

MethodsUsing control and Pkd1-/- kidney epithelial cells, we investigated peroxisome abundance, biogenesis and morphology by immunoblotting, immunofluorescent and live cell imaging of peroxisome-related proteins and assayed peroxisomal specific {beta}-oxidation. We further analyzed fatty acid composition by mass spectrometry in kidneys of Pkd1fl/fl; Ksp-Cre mice. We also evaluated peroxisome lipid metabolism in published metabolomics datasets of Pkd1 mutant cells and kidneys. Lastly, we investigated if the C-terminus or full-length polycystin-1 co-localize with peroxisome markers by imaging studies.

ResultsPeroxisome abundance, morphology and peroxisome-related protein expression in Pkd1-/- cells were normal, suggesting preserved peroxisome biogenesis. Peroxisomal {beta}-oxidation was not impaired in Pkd1-/- cells, and there was no obvious accumulation of very long chain fatty acids in kidneys of mutant mice. Re-analysis of published datasets provide little evidence of peroxisomal abnormalities in independent sets of Pkd1 mutant cells and cystic kidneys, while providing further evidence of mitochondrial fatty acid oxidation defects. Imaging studies with either full length polycystin-1 or its C-terminus, a fragment previously shown to go to the mitochondria, showed minimal co-localization with peroxisome markers.

ConclusionsOur studies showed that loss of Pkd1 does not disrupt peroxisome biogenesis nor peroxisomedependent fatty acid metabolism.

Key points- While mitochondrial abnormalities and fatty acid oxidation impairment have been reported in ADPKD, no studies have investigated if peroxisome dysfunction contributes to these defects.
- We investigated peroxisome morphology, biogenesis and function in cell and animal models of ADPKD and investigated whether polycystin-1 co-localized with peroxisome proteins.
- Our studies show that loss of Pkd1 does not disrupt peroxisome biogenesis nor peroxisome-dependent fatty acid metabolism.


title: The anterior Hox gene ceh-13 and elt-1/GATA activate the posterior Hox genes nob-1 and php-3 to specify posterior lineages in the C. elegans embryo

abstract:
Hox transcription factors play a conserved role in specifying positional identity during animal development, with posterior Hox genes typically repressing the expression of more anterior Hox genes. Here, we dissect the regulation of the posterior Hox genes nob-1 and php-3 in the nematode C. elegans. We show that nob-1 and php-3 are co-expressed in gastrulation-stage embryos in cells that previously expressed the anterior Hox gene ceh-13. This expression is controlled by several partially redundant transcriptional enhancers. These enhancers require ceh-13 for expression, providing a striking example of an anterior Hox gene positively regulating a posterior Hox gene. Several other regulators also act positively through nob-1/php-3 enhancers, including elt-1/GATA, ceh-20/ceh-40/Pbx, unc-62/Meis, pop-1/TCF, ceh-36/Otx and unc-30/Pitx. We identified defects in both cell position and cell division patterns in ceh-13 and nob-1;php-3 mutants, suggesting that these factors regulate lineage identity in addition to positional identity. Together, our results highlight the complexity and flexibility of Hox gene regulation and function and the ability of developmental transcription factors to regulate different targets in different stages of development.

Author SummaryHox genes are critical for head-to-tail patterning during embryonic development in all animals. Here we examine the factors that are necessary to turn on two posterior Hox genes, nob-1 and php-3, in the nematode worm, C. elegans. We identified six new transcription factors and three enhancer regions of DNA that can activate expression of nob-1/php-3. Unexpectedly, these activating transcription factors included ceh-13, an anterior Hox gene, and elt-1, a regulator of skin development that is briefly expressed in many cells that do not adopt skin fates, including the cells that express nob-1. Furthermore, the cellular defects we observed in ceh-13 and nob- 1;php-3 mutant embryos indicate that the early embryonic functions of these Hox genes help determine the identity of cells as well as their position within the embryo. Our findings identify new roles for Hox genes in C. elegans and emphasize the ability of transcription factors to contribute to the diversification of cell types and the adoption of specific cell types at different phases of embryonic development.


title: On the Gestural Origin of Language Lateralisation: Manual Communication reflects Broca's Asymmetry in Monkeys

abstract:
Manual gestures and speech recruit a common neural network, involving Brocas area in the left hemisphere. Such speech-gesture integration gave rise to theories on the critical role of manual gesturing in the origin of language.

Within this evolutionary framework, research on gestural communication in our closer primate relatives has received renewed attention for investigating its potential language-like features.

Here, using in-vivo anatomical MRI in 50 baboons, we found that communicative gesturing is related to Broca homologues marker in monkeys, namely the ventral portion of the Inferior Arcuate sulcus (IA sulcus). In fact, both direction and degree of gestural communications handedness - but not handedness for object manipulation - are associated and correlated with contralateral depth asymmetry at this exact IA sulcus portion. In other words, baboons that prefer to communicate with their right hand have a deeper left-than-right IA sulcus, than those preferring to communicate with their left hand and vice versa. Interestingly, in contrast to handedness for object manipulation, gestural communications lateralisation is not associated to the Central sulcus depth asymmetry, suggesting a double dissociation of handedness types between manipulative action and gestural communication.

It is thus not excluded that this specific gestural lateralisation signature within the baboons frontal cortex might reflect a phylogenetical continuity with language-related Broca lateralisation in humans.


title: Broca areas' asymmetry reflects gestural communication lateralisation in monkeys (Papio anubis)


title: Finger somatotopy is preserved after tetraplegia but deteriorates over time

abstract:
Following spinal cord injury (SCI), the motor output flow to the limb(s) and sensory input to the brain is largely lost. While attempted movements with the paralysed and sensory deprived body part can still evoke signals in the sensorimotor system, this task-related "net" brain activity of SCI patients differs substantially from healthy controls. Such reorganised and/or altered activity is thought to reflect abnormal processing. It is however possible that this altered  net sensorimotor activity in SCI patients conceals preserved somatotopically-specific representations of the paralysed and sensory deprived body parts that could be exploited in a functionally meaningful manner (e.g. via neuroprosthetics).

In this cross-sectional study, we investigated whether a functional connection between the periphery and the brain is necessary to maintain somatosensory representations. We used functional MRI and an (attempted) finger movement task to characterise the somatotopic hand layout in the primary somatosensory cortex and structural MRI to assess spared spinal tissue bridges. We tested 14 tetraplegic SCI patients (mean age {+/-} s.e.m.=55 {+/-} 3.6; 1 female) who differed in terms of lesion completeness, retained sensorimotor functioning, and time since injury, as well as 18 healthy control participants (mean age {+/-} s.e.m.=56 {+/-} 3.6 years; 1 female).

Our results revealed somatotopically organised representations of patients hands in which neighbouring clusters showed selectivity for neighbouring fingers in contralateral S1, qualitatively similar to those observed in healthy controls. To quantify whether these representations were normal in tetraplegic SCI patients we correlated each participants intricate representational distance pattern across all fingers (revealed using representational similarity analysis) with a canonical inter-finger distance pattern obtained from an independent sample. The resulting hand representation typicality scores were not significantly different between patients and controls. This was even true when considering two individual patients with no sensory hand functioning, no hand motor functioning, and no spared spinal tissue bridges. However, a correlational analysis revealed that over years since SCI the hand representation typicality in primary somatosensory cortex deteriorates.

We show that somatosensory representations can be maintained for several years following SCI even in the absence of perhiperhal inputs. Such preserved cortical hand representations could therefore be exploited in a functionally meaningful way by rehabilitation approaches that attempt to establish new functional connections between the hand and the brain after an SCI (e.g. through neuroprosthetics). However, time since injury may critically influence the somatotopic representations of SCI patients and might thereby impact the success of such rehabilitation approaches.


title: Using Adaptive Psychophysics to Identify the Neural Network Reset Time in Sub-second Interval Timing

abstract:
State dependent network models of sub-second interval timing propose that duration is encoded in states of neuronal populations that need to reset prior to a novel timing operation in order to maintain optimal timing performance. Previous research has shown that the approximate boundary of this reset interval can be inferred by varying the interstimulus interval between two to-be-timed intervals. However, the estimated boundary of this reset interval is broad (250-500ms) and remains underspecified with implications for the characteristics of state dependent network dynamics subserving interval timing. Here we probed the interval specificity of this reset boundary by manipulating the interstimulus interval between standard and comparison intervals in two sub-second auditory duration discrimination tasks (100 and 200ms) and a control (pitch) discrimination task using adaptive psychophysics. We found that discrimination thresholds improved with the introduction of a 333ms interstimulus interval relative to a 250ms interstimulus interval in both duration discrimination tasks, but not in the control task. This effect corroborates previous findings of a breakpoint in the discrimination performance for sub-second stimulus interval pairs as a function of an incremental interstimulus delay but more precisely localizes the minimal interstimulus delay range. These results suggest that state dependent networks subserving sub-second timing require approximately 250-333ms for the network to reset in order to maintain optimal interval timing.

New & NoteworthyThe state-dependent-network model considers interval timing as an intrinsic ability of neuronal populations to track the temporal evolution of their collective state. However, the time-dependent nature of neuronal properties imposes constraints on a maximum encodable interval and on the processing of intervals that are presented before the network resets to its baseline state. Investigating temporal discrimination thresholds as a function of variable inter-stimulus-intervals, we showed that the network reset time is between 250 and 333ms.


title: No evidence for somatosensory attenuationduring action observation of self-touch

abstract: The discovery of mirror neurons in the macaque brain in the 1990s triggered investigations on putative human mirror neurons and their potential functionality. The leading proposed function has been action understanding: accordingly, we understand the actions of others by  simulating them in our own motor system through a direct matching of the visual information to our own motor programs. Furthermore, it has been proposed that this simulation involves the prediction of the sensory consequences of the observed action, similar to the prediction of the sensory consequences of our executed actions. Here, we tested this proposal by quantifying somatosensory attenuation behaviorally during action observation. Somatosensory attenuation manifests during voluntary action and refers to the perception of self-generated touches as less intense than identical externally generated touches because the self-generated touches are predicted from the motor command. Therefore, we reasoned that if an observer simulates the observed action and, thus, he/she predicts its somatosensory consequences, then he/she should attenuate tactile stimuli simultaneously delivered to his/her corresponding body part. In three separate experiments, we found a systematic attenuation of touches during executed self-touch actions, but we found no evidence for attenuation when such actions were observed. Failure to observe somatosensory attenuation during observation of self-touch is not compatible with the hypothesis that the putative human mirror neuron system automatically simulates the observed action. In contrast, our findings emphasize a sharp distinction between the motor representations of self and others.


title: Maternal obesity programs white and brown adipose tissue transcriptome and lipidome in offspring in a sex-dependent manner

abstract:
The prevalence of overweight and obesity among children has drastically increased during the last decades and maternal obesity has been demonstrated as one of the ultimate factors. Nutrition-stimulated transgenerational epigenetic regulation of key metabolic genes is fundamental to the developmental origins of the metabolic syndrome. Fetal nutrition may differently influence female and male offspring. In this work, we investigated the sex-dependent programming of maternal obesity in visceral, subcutaneous and brown adipose tissues of offspring using magnetic resonance imaging and spectroscopy and a lipidomic approach combined with a Smart-Seq2 differential sequencing analysis. We show that the triglyceride profile varies between adipose depots, sexes and maternal diet. Our results demonstrate for the first time that a sex-dependent gene programming exists in visceral, subcutaneous and brown adipose tissues. Maternal obesity differentially programs gene expression in adipose depots of female and male offspring, which may contribute to the sex-dependent metabolic complications later in life.

Graphical abstract

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title: Aerosphere occupancy ({psi}) model of lesser short nosed fruit bat (Cynopterus brachyotis Muller, 1838) related to tree species in Asia mountainous paddy fields

abstract: Bat is animal that occupies aerosphere, especially fruit bats that forage on the space around the trees. The fruit bats use whether narrow space below tree canopy or in edge space on the edge of canopy. Whereas the aerosphere occupancy of fruits bats related to the specific tree species is poorly understood. Here, this paper aims to assess and model the association of fruit bat Cynopterus brachyotis aerosphere occupancy ({Psi}) with tree species planted in mountainous paddy fields in West Java. The studied tree species including Alianthus altissima, Acacia sp., Cocos nucifera, Mangifera indica, Pinus sp., and Swietenia macrophylla. The result shows that the tree species diversity has significantly (x2= 27.67, P < 0.05) affected the C. brachyotis aerosphere occupancy. According to values of {Psi} and occupancy percentage, high occupancy of narrow space by C. brachyotis was observed in Swietenia macrophylla ({Psi} = 0.934, 78%), followed by Alianthus altissima ({Psi} = 0.803, 57%), and Mangifera indica ({Psi} = 913, 55%). While high occupancy of edge space was observed in Mangifera indica ({Psi} = 0.685, 41%), followed by Pinus sp. ({Psi} = 0.674, 38%), and Alianthus altissima sp. ({Psi} = 0.627, 36%). The best model for explaining C. brachyotis occupation in narrow space is the tree height with preferences on high tree ({Psi}~tree height, AIC = 1.574, R2= 0.5535, Adj. R = 0.4047). While for edge space occupant, the best model is also the tree height ({Psi}~tree height, AIC = -26.1510, R2= 0.7944, Adj. R = 0.7258).

category: ecology


title: An LR framework incorporating sensitivity analysis to model multiple direct and secondary transfer events on skin surface

abstract: Bayesian logistic regression is used to model the probability of DNA recovery following direct and secondary transfer and persistence over a 24 hour period between deposition and sample collection. Sub-source level likelihood ratios provided the raw data for activity-level analysis. Probabilities of secondary transfer are typically low, and there are challenges with small data-sets with low numbers of positive observations. However, the persistence of DNA over time can be modelled by a single logistic regression for both direct and secondary transfer, except that the time since deposition must be compensated by an offset value for the latter. This simplifies the analysis. Probabilities are used to inform an activity-level Bayesian Network that takes account of alternative propositions e.g. time of assault and time of social activities. The model is extended in order to take account of multiple contacts between person of interest and   victim. Variables taken into account include probabilities of direct and secondary transfer, along with background DNA from unknown individuals. The logistic regression analysis is Bayesian -for each analysis, 4000 separate simulations were carried out. Quantile assignments enable calculation of a plausible range of probabilities and sensitivity analysis is used to describe the corresponding variation of LRs that occur when modelled by the Bayesian network. It is noted that there is need for consistent experimental design, and analysis, to facilitate inter-laboratory comparisons. Appropriate recommendations are made. The open-source program written in R-code ALTRaP (Activity Level, Transfer, Recovery and Persistence) enables analysis of complex multiple transfer propositions that are commonplace in cases-work e.g. between those who cohabit. A number of case examples are provided. ALTRaP can be used to replicate the results and can easily be modified to incorporate different sets of data and variables.


title: Copy number variation profile-based genomic subtyping of premenstrual dysphoric disorder in Chinese

abstract: Premenstrual dysphoric disorder (PMDD) affects nearly 5% women of reproductive age. The symptomatic heterogeneity, along with largely unknown genetics, of PMDD have greatly hindered its effective treatment. In the present study, 127 Chinese PMDD patients of the  invasion and  depression subtypes clinically differentiated by us earlier were analyzed together with 108 non-PMDD controls for genome-wide copy number variations (CNVs). Germline genomic DNA samples from white blood cells were subjected to AluScan sequencing-based CNV profiling, which enabled clustering of patient samples readily into the V and D groups, dominated by the "invasion" and "depression" clinical subtypes, respectively; the CNVs obtained with 100-kb windows yielded two clusters that were correlated with these subtypes with a consistency of up to 89.8%. Diagnostic correlation- and frequency-based CNV features of either CNV-gain (CNVG) or CNV-loss (CNVL) that could differentiate between V and D subtypes were selected and analyzed. CNVG features located preferentially in S2-phase replicating regions and enriched with steroid hormone biosynthesis pathway of genes were found protective against PMDD. Moreover, machine learning employing the correlation-based CNV features could predict with >80% accuracy whether a genomic sample was D-type, V-type or control. In terms of their CNV profiles, the D- and V-types differed more from one another than from the controls, thereby providing a genomic basis for the clinical D-V subtyping of PMDD. Genome-wide profiling of CNVs, as a new approach to complex disease genetics, has revealed recurrent CNVs and genomic features beyond individual genes and mutations underlying PMDD clinical diversity.


title: RNA fragment assembly with experimental restraints

abstract:
We present RNA Masonry, a computer program and a web service for a fully automated assembly of RNA fragments into geometrically plausible models fulfilling user-provided secondary structure constraints and restraints on tertiary contacts and Small Angle X-ray Scattering (SAXS) data. We illustrate the method description with its recent application to structural studies of viral RNAs with SAXS restraints. The program web server is available at http://iimcb.genesilico.pl/rnamasonry.

Contactgchojnowski@embl-hamburg.de

category: bioinformatics


title: Germaphobia! Does our Relationship with, and Knowledge of Biodiversity Affect our Attitudes Towards Microbes?

abstract: Germaphobia -- a pathological aversion to microorganisms -- could be contributing to an explosion in human immune-related disorders via mass sterilisation of surfaces and reduced exposure to biodiversity. Loss of biodiversity and our connectedness to nature, along with poor microbial literacy may be augmenting the negative consequences of germaphobia on ecosystem health. In this study, we created an online questionnaire to acquire data on attitudes towards, and knowledge of microbes. We collected data on nature connectedness and interactions with nature and explored the relationships between these variables. We found a significant association between attitudes towards microbes and both duration and frequency of visits to natural environments. A higher frequency of visits to nature per week, and a longer duration spent in nature per visit, significantly associated with positive attitudes towards microbes. We found no association between nature connectedness and attitudes towards microbes. We found a significant relationship between knowledge of  lesser known microbial groups (e.g., identifying that fungi, algae, protozoa, and archaea are microbes) and positive attitudes towards microbes. However, we also found that people who correctly identified viruses as being microbes expressed less positive views of microbes overall -- this could potentially be attributed to a  COVID-19 effect. Our results suggest that basic microbial literacy and nature engagement may be important in reducing/preventing germaphobia. The results also suggest that a virus-centric phenomenon (e.g., COVID-19) could increase broader germaphobia. As the rise of immune-related disorders and mental health conditions have been linked to germaphobia, reduced biodiversity, and non-targeted sterilisation, our findings point to a feasible strategy to potentially help ameliorate these negative consequences. A greater emphasis on microbial literacy and promoting time spent in nature could be useful in promoting resilience in human health and more positive/constructive attitudes towards the foundations of our ecosystems - the microorganisms.

category: scientific communication and education


title: Modeling features of addiction with an oral oxycodone self-administration paradigm

abstract: Prescription opioid use is an initiating factor driving the current opioid epidemic. There are several challenges with modeling prescription opioid addiction. First, prescription opioids such as oxycodone are orally self-administered and have different pharmacokinetics and dynamics than morphine or fentanyl. This oral route of administration determines the pharmacokinetic profile, which is critical for establishing reliable drug-reinforcement associations in animals. Moreover, the pattern of intake and environment in which addictive drugs are self-administered intake are critical determinants of the levels of drug intake, sensitization and relapse behavior. This is an important consideration with prescription opioid use, which is characterized by continuous drug access in familiar environments. Thus, to model features of prescription opioid use and the transition to abuse, we present an oral oxycodone self-administration paradigm that is administered in the home cage. Mice voluntarily self-administer oxycodone in this paradigm without any taste modification such as sweeteners, and exhibit preference for oxycodone, escalation of intake, physical signs of dependence, reinstatement of seeking after withdrawal, and a subset of animals demonstrate drug taking that is resistant to aversive consequences. This model could be useful for studying the neurobiological substrates specifically relevant to prescription opioid abuse.


title: Methylomic alteration in peripheral blood lymphocytes of prodromal stage and first-episode Chinese Han schizophrenia patients

abstract:
BackgroundAlthough epigenetic dysregulation has long been proposed to promote the onset of schizophrenia, the landscape of the methylomic changes across the whole genome is yet established.

MethodsUsing Infinium Human Methylation 850 BeadChip Array and MethylTarget sequencing method, we investigated the genome-wide methylation profiles and further validated methylation profiles of target genes in peripheral blood lymphocytes between individuals with psychosis risk syndrome (PRS), patients with first-episode schizophrenia (FES) and healthy controls (HC) in Chinese Han population.

ResultsWe detected 372 sites between psychosis risk syndrome (PRS) and healthy controls (HC), which increased to 460 sites in first-episode schizophrenia (FES) with 207 sites shared. Both PRS and FES featured profound hypomethylation within gene body. Gene ontology and network annotation merged on loci enriched in disease associated signaling pathways (MAPK(Mitogen Activated Protein Kinases), Glutamatergic, GABAergic etc.).

ConclusionsOur study implicated characteristic hypomethylation in both the discovery and validation cohorts in SYNGAP1, one of the frequently studied genes in neurodevelopmental disorders. This is the first methylome-wide association study between PRS and FES in Chinese Han population. Our findings provide potential biomarkers that can be used for future development of disease therapy and management.


title: pyconsFold: A fast and easy tool for modelling and docking using distance predictions.

abstract:
MotivationContact predictions within a protein has recently become a viable method for accurate prediction of protein structure. Using predicted distance distributions has been shown in many cases to be superior to only using a binary contact annotation. Using predicted inter-protein distances has also been shown to be able to dock some protein dimers.

ResultsHere we present pyconsFold. Using CNS as its underlying folding mechanism and predicted contact distance it outperforms regular contact prediction based modelling on our dataset of 210 proteins. It performs marginally worse than the state of the art pyRosetta folding pipeline but is on average about 20 times faster per model. More importantly pyconsFold can also be used as a fold-and-dock protocol by using predicted inter-protein contacts to simultaneously fold and dock two protein chains.

Availability and implementationpyconsFold is implemented in Python 3 with a strong focus on using as few dependencies as possible for longevity. It is available both as a pip package in Python 3 and as source code on GitHub and is published under the GPLv3 license.

Contactarne@bioinfo.se

Supplemental materialInstall instructions, examples and parameters can be found in the supplemental notes.

Availability of dataThe data underlying this article together with source code are available on github, at https://github.com/johnlamb/pyconsfold.


title: Recognition of N6-methyladenosine by the YTHDC1 YTH domain studied by molecular dynamics and NMR spectroscopy: The role of hydration

abstract: The YTH domain of YTHDC1 belongs to a class of protein "readers", recognizing the N6-methyladenosine (m6A) chemical modification in mRNA. Static ensemble-averaged structures revealed details of N6-methyl recognition via a conserved aromatic cage. Here, we performed molecular dynamics (MD) simulations along with nuclear magnetic resonance (NMR) and isothermal titration calorimetry (ITC) to examine how dynamics and solvent interactions contribute to the m6A recognition and negative selectivity towards unmethylated substrate. The structured water molecules surrounding the bound RNA and the methylated substrates ability to exclude bulk water molecules contribute to the YTH domains preference for m6A. Intrusions of bulk water deep into the binding pocket disrupt binding of unmethylated adenosine. The YTHDC1s preference for the 5'-Gm6A-3' motif is partially facilitated by a network of water-mediated interactions between the 2-amino group of the guanosine and residues in the m6A binding pocket. The 5'-Im6A-3' (where I is inosine) motif can be recognized too but disruption of the water network lowers affinity. The D479A mutant also disrupts the water network and destabilizes m6A binding. Our interdisciplinary study of YTHDC1 protein/RNA complex reveals an unusual physical mechanism by which solvent interactions contributes towards m6A recognition.


title: Male pheromones modulate synaptic transmission at the C. elegans neuromuscular junction in a sexually dimorphic manner

abstract: The development of functional synapses in the nervous system is important for animal physiology and behaviors. The synaptic transmission efficacy can be modulated by the environment to accommodate external changes, which is crucial for animal reproduction and survival. However, the underlying plasticity of synaptic transmission remains poorly understood. Here we show that in C. elegans, the male pheromone increases the hermaphrodite cholinergic transmission at the neuromuscular junction (NMJ), which alters hermaphrodites locomotion velocity and mating efficiency in a developmental stage-dependent manner. Dissection of the sensory circuits reveals that the AWB chemosensory neurons sense those male pheromones and further transduce the information to NMJ using cGMP signaling. Exposure of hermaphrodites to male pheromones specifically increases the accumulation of presynaptic CaV2 calcium channels and clustering of postsynaptic receptors at cholinergic synapses of NMJ, which potentiates cholinergic synaptic transmission. Thus, our study demonstrates a circuit mechanism for synaptic modulation by sexual dimorphic pheromones.


title: Repetitive Mild Head Trauma Induces Activity-Mediated Lifelong Brain Deficits in a Novel Drosophila Model

abstract: Mild head trauma, including concussion, can lead to chronic brain dysfunction and degeneration but the underlying mechanisms remain poorly understood. Here, we developed a novel head impact system to investigate the long-term effects of mild head trauma on brain structure and function, as well as the underlying mechanisms in Drosophila melanogaster. We find that Drosophila subjected to repetitive head impacts develop long-term deficits, including impaired startle-induced climbing, progressive brain degeneration, and shortened lifespan, all of which are substantially exacerbated in female flies. Interestingly, head impacts elicit an elevation in neuronal activity and its acute suppression abrogates the detrimental effects in female flies. Together, our findings validate Drosophila as a suitable model system for investigating the long-term effects of mild head trauma, suggest an increased vulnerability in brain injury in female flies, and indicate that early altered neuronal excitability may be a key mechanism linking mild brain trauma to chronic degeneration.


title: A screen for sleep and starvation resistance identifies a wake-promoting role for the auxiliary channel unc79

abstract: The regulation of sleep and metabolism are highly interconnected, and dysregulation of sleep is linked to metabolic diseases that include obesity, diabetes, and heart disease. Further, both acute and long-term changes in diet potently impact sleep duration and quality. To identify novel factors that modulate interactions between sleep and metabolic state, we performed a genetic screen for their roles in regulating sleep duration, starvation resistance, and starvation-dependent modulation of sleep. This screen identified a number of genes with potential roles in regulating sleep, metabolism or both processes. One such gene encodes the auxiliary ion channel UNC79, which was implicated in both the regulation of sleep and starvation resistance. Genetic knockdown or mutation of unc79 results in flies with increased sleep duration, as well as increased starvation resistance. Previous findings have shown that unc79 is required in pacemaker for 24-hour circadian rhythms. Here, we find that unc79 functions in the mushroom body, but not pacemaker neurons, to regulate sleep duration and starvation resistance. Together, these findings reveal spatially localized separable functions of unc79 in the regulation of circadian behavior, sleep, and metabolic function.


title: Wavelength-dependent effects of artificial light at night on phytoplankton growth and community structure

abstract:
Artificial light at night (ALAN) is increasingly recognised as a disruptive form of environmental pollution, impacting many physiological and behavioural processes that may scale up to population and community-level effects. Mounting evidence from animal studies show that the severity and type of the impact depends on the wavelength and intensity of ALAN. This knowledge has been instrumental for informing policy-making and planning for wildlife-friendly illumination. However, most of this evidence comes from terrestrial habitats, while research testing alternative wavelength illumination in marine environments is lagging behind. In this study we investigated the effect of such alternative ALAN colours on marine primary producers. Specifically, we tested the effect of green, red, and natural white LED illumination at night, compared to a dark control, on the growth of a green microalgae as well as the biomass, diversity and composition of a phytoplankton assemblage. Our findings show that green ALAN boosted chlorophyll production at the exponential growth stage, resulting in higher biomass production in the green algae Tetraselmis suesica. All ALAN wavelengths affected the biomass and diversity of the assemblage with the red and green ALAN having the stronger effects, leading to higher overall abundance and selective dominance of specific diatom species compared to white ALAN and the dark control.

SynthesisOur work indicates that the wavelength of artificial light sources in marine areas should be carefully considered in management and conservation plans. In particular, green and red light should be used with caution in coastal areas, where there might be a need to strike a balance between the strong effects of green and red light on marine primary producers with the benefit they bring to other organisms.


title: How much does it take to be old? Modelling the time since the last harvesting to infer the distribution of mature forests in France

abstract:
AimThe distribution of overmature forests in metropolitan France is poorly known, with only a few well-studied prominent sites, and has never been evaluated countrywide. Here, we modelled French forest reserves time since the last harvesting operation - a proxy for forest maturity - then inferred the current statistical distribution of overmature forests (i.e. forests over 50 years without harvesting) in France.

LocationMetropolitan France

MethodsWe used inventories from forest reserves and managed forests to calibrate a generalised linear mixed model explaining the time since the last harvesting with selected structural attributes and environmental variables. We then projected this model on the independent National Forest Inventory dataset. We thus obtained an updated estimation of the proportion and a rough distribution of overmature forest stands in metropolitan France.

ResultsWe found that high basal area of very large trees, high volumes of standing and downed deadwood, high diversity of tree-related microhabitats and more marginally diversity of decay stages best characterized the time since the last harvesting. Volumes of stumps and high density of coppices translating legacy of past forest management also distinguished more overmature plots. Our projection yielded an estimated 3% of French forests over 50 years without harvesting mostly located in more inaccessible areas (i.e. mountainous areas) and a promising proportion of future overmature forests if left unharvested.

Main conclusionsOur study showed that the time since the last harvesting is a good proxy for a combination of stand structure attributes key in characterising overmature temperate forests. It gives the first robust statistical estimate of the proportion of overmature forests and may serve to report on their status in metropolitan France. Our method could be implemented at a larger spatial scale, notably in countries with accessible National Forest Inventory and calibration data, to produce indicators at international level.


title: How much does it take to be old? Modelling the time since the last harvesting to infer the distribution of overmature forests in France


title: Evolution of lower levels of inter-locus sexual conflict in D. melanogaster populations under strong selection for rapid development.

abstract: D. melanogaster laboratory populations subjected to selection for rapid development and early reproduction have been found to have evolved reduced adult body size and lower levels of inter-locus sexual conflict compared to their ancestral controls. We tested the contribution of a smaller body to the evolution of reduced sexual conflict in these populations, since body size differences are known to affect sexual conflict levels in this species. We cultured larvae from the control populations at high density to obtain flies as small as those from the selected populations. The effect of body size reduction on sexual conflict was asymmetric, with smaller body size resulting in reduced male manipulative ability but not female resistance to mating-induced harm. These results were not due to differences in behavioural patterns of smaller flies, such as differences in overall mating exposure of females to different types of males. We hypothesize that evolution for rapid development and the correlated reduction in body size has resulted in lower male manipulative ability, and sexually antagonistic co-evolution has lowered female resistance to such manipulations. These populations have also evolved incipient reproductive isolation from their controls, likely through sexual conflict (reported earlier), and our results support the view that this is an outcome of strong, directional selection for rapid development.

category: evolutionary biology


title: Development and evaluation of a new methodology for Soft Tissue Artifact compensation in the lower limb

abstract:
Skin Marker (SM) based motion capture is the most widespread technique used for motion analysis. Yet, the accuracy is often hindered by Soft Tissue Artifact (STA). This is a major issue in clinical gait analysis where kinematic results are used for decision-making. It also has a considerable influence on the results of rigid body and Finite Element (FE) musculoskeletal models that rely on SM-based kinematics to estimate muscle, contact and ligament forces. Current techniques designed to compensate for STA, in particular multi-body optimization methods, assume anatomical simplifications to define joint constraints. These methods, however, cannot adapt to subjects bone morphology, particularly for patients with joint lesions, nor easily can account for subject- and location-dependent STA. In this perspective, we propose to develop a conceptual FE based model of the lower limb for STA compensation and evaluate it for 66 healthy subjects under level walking motor task.

Both hip and knee joint kinematics were analyzed, considering both rotational and translational joint motion. Results showed that STA caused underestimation of the hip joint kinematics (up to 2.2{degrees}) for all rotational DoF, and overestimation of knee joint kinematics (up to 12{degrees}) except in flexion/extension. Joint kinematics, in particular the knee joint, appeared to be sensitive to soft tissue stiffness parameters (rotational and translational mean difference up to 1.5{degrees} and 3.4 mm). Analysis of the results using alternative joint representations highlighted the versatility of the proposed modeling approach. This work paves the way for using personalized models to compensate for STA in healthy subjects and different activities.

category: bioengineering


title: Quantitative proteomics identifies secreted diagnostic biomarkers as well as tumor-dependent prognostic targets for clear cell Renal Cell Carcinoma

abstract: Clear cell Renal Cell Carcinoma (ccRCC) is the third most common and most malignant urological cancer, with a 5-year survival rate of 10% for patients with advanced tumors. Here, we identified 10,160 unique proteins by in-depth quantitative proteomics, of which 955 proteins were significantly regulated between tumor and normal adjacent tissues. We verified 4 putatively secreted biomarker candidates, namely PLOD2, FERMT3, SPARC and SIRP, as highly expressed proteins that are not affected by intra- and inter-tumor heterogeneity. Moreover, SPARC displayed a significant increase in urine samples of ccRCC patients, making it a promising marker for clinical screening assays. Furthermore, based on molecular expression profiles, we propose a biomarker panel for the robust classification of ccRCC tumors into two main clusters, which significantly differed in patient outcome with an almost three times higher risk of death for cluster 1 tumors compared to cluster 2 tumors. Moreover, among the most significant clustering proteins, 13 were targets of repurposed inhibitory FDA-approved drugs. Our rigorous proteomics approach identified promising diagnostic and tumor-discriminative biomarker candidates which can serve as therapeutic targets for the treatment of ccRCC.

category: cancer biology


title: State-Anxiety Modulates the Effect of Emotion Cues on Visual Temporal Sensitivity in Autism Spectrum Disorder

abstract: Atypical processing of stimulus inputs across a range of sensory modalities in autism spectrum disorders (ASD) are widely reported. Sensory processing is known to be influenced by bodily internal states such as physiological arousal and anxiety. Since a sizeable proportion of ASD individuals reportedly have co-morbid anxiety disorders that are linked with dysregulated arousal, we investigated if face-emotion arousal cues, influenced visual sensory sensitivity (indexed by temporal resolution) in an ASD group (n=20) compared to a matched group of typically-developed individuals (TD, n=21). We asked further if emotion-cued changes in visual sensitivity associated with individual differences in state- and trait-anxiety. Participants reported the laterality of the second of two consecutive Gaussian-blob flashes in a visual temporal order judgment task (v-TOJ), demanding higher-level visual processing. The key manipulation was presenting a task-irrelevant face emotion cue briefly at unexpected time points preceding the task-relevant flashes. Disgust vs Neutral emotion signals enhanced the visual temporal resolution in ASD individuals. Furthermore, individual state-anxiety scores correlated with the emotion-cued change of temporal resolution (Disgust vs Neutral) in the ASD group. Both these effects were absent in the TD group. The results show that individual state-anxiety levels significantly modulate the effect of emotions on visual temporal sensitivity in ASD individuals, which was absent in our TD sample. The findings support a nuanced approach to understand the disparate sensory features in ASD individuals, by factoring in the interplay of the individual reactivity to environmental affective information and the severity of anxiety.


title: Archaeal extracellular vesicles are produced in an ESCRT-dependent manner and promote gene transfer and nutrient cycling in extreme environments

abstract: Membrane-bound extracellular vesicles (EVs), secreted by cells from all three domains of life, transport various molecules and act as agents of intercellular communication in diverse environments. Here we demonstrate that EVs produced by a hyperthermophilic and acidophilic archaeon Sulfolobus islandicus carry not only diverse proteins but also chromosomal and plasmid DNA, and can transfer this DNA to recipient cells. Furthermore, we show that EVs can support the heterotrophic growth of Sulfolobus in minimal medium, implicating EVs in carbon and nitrogen fluxes in extreme environments. Finally, our results suggest that, similar to eukaryotes, production of EVs in S. islandicus depends on the archaeal ESCRT machinery. Using synchronized S. islandicus cultures, we show that EV production is linked to cell division and appears to be triggered by increased expression of ESCRT proteins during this cell cycle phase. Using a CRISPR-based knockdown system, we show that archaeal ESCRT-III and AAA+ ATPase Vps4 are required for EV production, whereas archaea-specific component CdvA appears to be dispensable. Collectively, our results suggest that ESCRT-mediated EV biogenesis has deep evolutionary roots, likely predating the divergence of eukaryotes and archaea, and that EVs play an important role in horizontal gene transfer and nutrient cycling in extreme environments.


title: A catalytic-independent function of human DNA polymerase Kappa controls the pool of the Checkpoint Kinase 1

abstract:
DNA polymerase kappa (Pol {kappa}) has been well documented thus far for its specialized DNA synthesis activity during translesion replication, progression of replication forks through regions difficult to replicate and replication checkpoint at stalled forks.

Here we unveiled an unexpected role for Pol {kappa} in controlling the stability and abundance of Chk1, the major mediator of the replication checkpoint. We found that loss of Pol {kappa} decreased the Chk1 protein level in the nucleus of four human cell lines. Pol {kappa} and not the other Y-family polymerase members is required to maintain the Chk1 protein pool all along the cell cycle. We showed that Pol {kappa} depletion affected the protein stability of Chk1 and protected it from proteasome degradation and the replication recovery defects observed in Pol {kappa}-depleted cells could be overcome by the re-expression of Chk1. Importantly, this new function of Pol {kappa} does not require its catalytic activity, revealing that in addition to its known roles in the replication process, Pol {kappa} can contribute to the maintenance of genome stability independently of its DNA synthesis activity.


title: Tankyrase-mediated ADP-ribosylation is a novel regulator of TNF-induced death

abstract: Tumor necrosis factor (TNF) is an inflammatory cytokine that, upon binding to its receptor TNFR1, can drive cytokine production, cell survival, or cell death and is a major component of an organisms anti-pathogen repetoire1,2. TNF stimulation leads to the formation of two distinct signalling complexes, a well-defined membrane bound complex (complex 1), and a less well characterised cytosolic death inducing complex (complex 2). Using mass spectrometry, we identified the ADP-ribosyltransferase, tankyrase-1 (TNKS1/TNKS/ARTD5/PARP5a) as a novel native complex 2 component. Following a TNF-induced death stimulus TNKS1 is recruited to complex 2, resulting in complex 2 poly(ADP-ribosyl)ation (PARylation). Tankyrase inhibitors sensitise cells to TNF-induced death, which is correlated with increased complex 2 assembly. Tankyrase-mediated PARylation promotes recruitment of the E3 ligase RNF146 and RNF146 deficiency or proteasome inhibition results in increased levels of complex 2, suggesting that RNF146 causes proteasomal degradation of complex 2. Several viruses express ADP-ribose binding macrodomain proteins, and expression of the SARS-CoV-2 or VEEV macrodomain markedly sensitises cells to TNF-induced death. This suggests that ADP-ribosylation serves as yet another mechanism to detect pathogenic interference of TNF signalling and retaliate with an inflammatory cell death.


title: Specification of axial identity by Hoxa2 distinguishes between a phenotypic and molecular ground state in mouse cranial neural crest cells

abstract: Hox genes play a key role in head formation by specifying the axial identity of neural crest cells (NCCs) migrating into embryonic pharyngeal arches. In the absence of Hoxa2, NCC derivatives of the second pharyngeal arch (PA2) undergo a homeotic transformation and duplicate structures formed by first arch (PA1) NCCs. Current models postulate that PA1 represents a NCC  ground state and loss of Hoxa2 causes a reversion of PA2 NCCs to the PA1  ground state. We use bulk and single-cell RNAseq to investigate the molecular mechanisms driving this phenotypic transformation in the mouse. In Hoxa2-/- mutants, PA2 NCCs generally maintain expression of the PA2 transcriptional signature and fail to strongly upregulate a PA1 transcriptional signature. Our analyses identify putative HOXA2 targets and suggest that subsets of NCCs may respond to HOXA2 activity in distinct manners. This separation of phenotypic and molecular states has significant implications for understanding craniofacial development.

category: developmental biology


title: A potent bispecific nanobody protects hACE2 mice against SARS-CoV-2 infection via intranasal administration

abstract:
The dramatically expanding COVID-19 needs multiple effective countermeasures. Neutralizing antibodies are a potential therapeutic strategy for treating COVID-19. A number of neutralizing nanobodies (Nbs) were reported for their in vitro activities. However, in vivo protection of these nanobodies was not reported in animal models. In the current report, we characterized several RBD-specific Nbs isolated from a screen of an Nb library derived from an alpaca immunized with SARS-CoV-2 spike glycoprotein (S); among them, three Nbs exhibited picomolar potency against SARS-CoV-2 live virus, pseudotyped viruses, and 15 circulating SARS-CoV-2 variants. To improve the efficacy, various configurations of Nbs were engineered. Nb15-NbH-Nb15, a novel trimer constituted of three Nbs, was constructed to be bispecific for human serum albumin (HSA) and RBD of SARS-CoV-2. Nb15-NbH-Nb15 exhibited sub-ng/ml neutralization potency against the wild-type and currently circulating variants of SARS-CoV-2 with a long half-life in vivo. In addition, we showed that intranasal administration of Nb15-NbH-Nb15 provided 100% protection for both prophylactic and therapeutic purposes against SARS-CoV-2 infection in transgenic hACE2 mice. Nb15-NbH-Nb15 is a potential candidate for both prevention and treatment of SARS-CoV-2 through respiratory administration.

One sentence summaryNb15-NbH-Nb15, with a novel heterotrimeric bispecific configuration, exhibited potent and broad neutralization potency against SARS-CoV-2 in vitro and provided in vivo protection against SARS-CoV-2 infection in hACE2 transgenic mice via intranasal delivery.

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=156 SRC="FIGDIR/small/429275v1_ufig1.gif" ALT="Figure 1">
View larger version (47K):
org.highwire.dtl.DTLVardef@a30bc8org.highwire.dtl.DTLVardef@5a872eorg.highwire.dtl.DTLVardef@1610f74org.highwire.dtl.DTLVardef@13d9bd8_HPS_FORMAT_FIGEXP  M_FIG Graphical abstract:

C_FIG HighlightsO_LIWe described a novel heterotrimeric configuration of Nb-NbH-Nb (Nb15-NbH-Nb15) that exhibited improved viral inhibition and stability.
C_LIO_LINb15-NbH-Nb15 provides ultrahigh neutralization potency against SARS-CoV-2 wild type and 18 mutant variants, including the current circulating variants of D614G and N501Y predominantly in the UK and South Africa.
C_LIO_LIIt is the first to demonstrate the Nbs efficacy in preventing and treating SARS-CoV-2 infection in hACE2 transgenic mice via intranasal delivery.
C_LI


title: High Throughput Nanopore Sequencing of SARS-CoV-2 Viral Genomes from Patient Samples

abstract: In late 2019, a novel coronavirus began spreading in Wuhan, China, causing a potentially lethal respiratory viral infection. By early 2020, the novel coronavirus, called SARS-CoV-2, had spread globally, causing the COVID-19 pandemic. The infection and mutation rates of SARS-CoV-2 make it amenable to tracking movement and evolution by viral genome sequencing. Efforts to develop effective public health policies, therapeutics, or vaccines to treat or prevent COVID-19 are also expected to benefit from tracking mutations of the SARS-CoV-2 virus. Here we describe a set of comprehensive working protocols, from viral RNA extraction to analysis using online visualization tools, for high throughput sequencing of SARS-CoV-2 viral genomes using a MinION instrument. This set of protocols should serve as a reliable  how-to reference for generating quality SARS-CoV-2 genome sequences with ARTIC primer sets and next-generation nanopore sequencing technology. In addition, many of the preparation, quality control, and analysis steps will be generally applicable to other sequencing platforms.


title: Plasmodium falciparum hydroxymethylbilane synthase does not house any cosynthase activity within the haem biosynthetic pathway

abstract: The production of uroporphyrinogen III, the universal progenitor of macrocyclic, modified tetrapyrroles, is produced from aminolaevulinic acid (ALA) by a conserved pathway involving three enzymes: porphobilinogen synthase (PBGS), hydroxymethylbilane synthase (HmbS) and uroporphyrinogen III synthase (UroS). The gene encoding uroporphyrinogen III synthase has not yet been identified in Plasmodium falciparum but it has been suggested that this activity is housed inside a bifunctional hybroxymethylbilane synthase (HmbS). In this present study it is demonstrated that P. falciparum HmbS does not have uroporphyrinogen III synthase activity. This was demonstrated by the failure of a codon optimised P. falciparum hemC gene, encoding HmbS, to compliment a defined E. coli hemD- mutant (SASZ31) deficient in uroporphyrinogen III synthase activity. Furthermore, HPLC analysis of the oxidsed reaction product from recombinant, purified HmbS showed that only uroporphyrin I could be detected (corresponding to hydroxymethylbilane production). No uroporphyrin III was detected, thus showing that P. falciparum HmbS does not have UroS activity and can only catalyse the formation of hydroxymethylbilane from porphobilinogen.

category: biochemistry


title: Constructing smaller genome graphs via string compression

abstract:
The size of a genome graph -- the space required to store the nodes, their labels and edges -- affects the efficiency of operations performed on it. For example, the time complexity to align a sequence to a graph without a graph index depends on the total number of characters in the node labels and the number of edges in the graph. The size of the graph also affects the size of the graph index that is used to speed up the alignment. This raises the need for approaches to construct space-efficient genome graphs.

We point out similarities in the string encoding approaches of genome graphs and the external pointer macro (EPM) compression model. Supported by these similarities, we present a pair of linear-time algorithms that transform between genome graphs and EPM-compressed forms. We show that the algorithms result in an upper bound on the size of the genome graph constructed based on an optimal EPM compression. In addition to the transformation, we show that equivalent choices made by EPM compression algorithms may result in different sizes of genome graphs. To further optimize the size of the genome graph, we purpose the source assignment problem that optimizes over the equivalent choices during compression and introduce an ILP formulation that solves that problem optimally. As a proof-of-concept, we introduce RLZ-Graph, a genome graph constructed based on the relative Lempel-Ziv EPM compression algorithm. We show that using RLZ-Graph, across all human chromosomes, we are able to reduce the disk space to store a genome graph on average by 40.7% compared to colored de Bruijn graphs constructed by Bifrost under the default settings.

The RLZ-Graph software is available at https://github.com/Kingsford-Group/rlzgraph


title: Transformations, Comparisons, and Analysis of Down to Up Protomer States of Variants of the SARS-CoV-2 Prefusion Spike Protein Including the UK Variant B.1.1.7

abstract: Monitoring and strategic response to variants in SARS-CoV-2 represents a considerable challenge in the current pandemic, as well as potentially future viral outbreaks of similar magnitude. In particular mutations and deletions involving the virions prefusion Spike protein have significant potential impact on vaccines and therapeutics that utilize this key structural viral protein in their mitigation strategies. In this study, we have demonstrated how dominant energetic landscape mappings ("glue points") coupled with sequence alignment information can potentially identify or flag key residue mutations and deletions associated with variants. Surprisingly, we also found excellent homology of stabilizing residue glue points across the lineage of {beta} coronavirus Spike proteins, and we have termed this as "sequence homologous glue points". In general, these flagged residue mutations and/or deletions are then computationally studied in detail using all-atom biocomputational molecular dynamics over approximately one microsecond in order to ascertain structural and energetic changes in the Spike protein associated variants. Specifically, we examined both a theoretically-based triple mutant and the so-called UK or B.1.1.7 variant. For the theoretical triple mutant, we demonstrated through Alanine mutations, which help "unglue" key residue-residue interactions, that these three key stabilizing residues could cause the transition of Down to Up protomer states, where the Up protomer state allows binding of the prefusion Spike protein to hACE2 host cell receptors, whereas the Down state is believed inaccessible. Thus, we are able to demonstrate the importance of glue point residue identification in the overall stability of the prefusion Spike protein. For the B.1.1.7 variant, we demonstrated the critical importance of D614G and N5017 on the structure and binding, respectively, of the Spike protein. Notably, we had previously identified D614 as a key glue point in the inter-protomer stabilization of the Spike protein prior to the emergence of its mutation. The mutant D614G is a structure breaking Glycine mutation demonstrating a relatively more distal Down state RBD and a more stable conformation in general. In addition, we demonstrate that the mutation N501Y may significantly increase the Spike protein binding to hACE2 cell receptors through its interaction with Y41 of hACE2 forming a potentially strong hydrophobic residue binding pair. We note that these two key mutations, D614G and N501Y, are also found in the so-called South African (SA; B.1.351) variant of SARS-CoV-2. Future studies along these lines are, therefore, aimed at mapping glue points to residue mutations and deletions of associated prefusion Spike protein variants in order to help identify and analyze possible "variants of interest" and optimize efforts aimed at the mitigation of this current and future virions.


title: Transformations, Lineage Comparisons, and Analysis of Down to Up Protomer States of Variants of the SARS-CoV-2 Prefusion Spike Protein Including the UK Variant B.1.1.7


title: Energetic and structural features of SARS-CoV-2 N-protein co-assemblies with nucleic acids

abstract: Nucleocapsid (N) protein of the SARS-CoV-2 virus packages the viral genome into well-defined ribonucleoprotein particles, but the molecular pathway is still unclear. N-protein is dimeric and consists of two folded domains with nucleic acid (NA) binding sites, surrounded by intrinsically disordered regions that promote liquid-liquid phase separation. Here we use biophysical tools to study N-protein interactions with oligonucleotides of different length, examining the size, composition, secondary structure, and energetics of the resulting states. We observe formation of supramolecular clusters or nuclei preceding growth into phase-separated droplets. Short hexanucleotide NA forms compact 2:2 N-protein/NA complexes with reduced disorder. Longer oligonucleotides expose additional N-protein interactions and multi-valent protein-NA interactions, which generate higher-order mixed oligomers and simultaneously promote growth of droplets. Phase separation is accompanied by a significant increase in protein secondary structure, different from that caused by initial NA binding, which may contribute to the assembly of ribonucleoprotein particles within molecular condensates.


title: Single-cell transitional dynamics unravel stimulus- and cell type-dependent signaling outputs of distinct p53 regulatory feedback motifs

abstract: Despite myriad of systemic analysis, our quantitative understanding of signaling networks and their differential outputs is still limited, especially at the dynamic level. Here we employed a network motif approach to dissect key regulatory units of p53 pathway and determined how their collective activities generated context-specific dynamic responses. By combining single-cell imaging and mathematical modeling of dose-dependent p53 dynamics induced by three chemotherapeutics of distinct mechanism-of-action, including Etoposide, Nutlin-3a and 5-Fluouracil, and in five cancer cell types, we identified novel p53 dynamic modes and their mechanistic origins with new insight on previously unknown drug mediators and phenotypic heterogeneity of cancer cells. Our results established the functional roles of unique feedback motifs of p53 pathway in generating the stimulus- and cell type-specific signaling responses and demonstrated that transitional dynamics activated at intermediate stimulus levels can be exploited as novel quantitative readouts to uncover and elucidate the key building blocks of bio-networks.

category: systems biology


title: GABA administration limits viral replication and pneumonitis in a mouse model of COVID-19

abstract: Despite the availability of vaccines for COVID-19, serious illness and death induced by coronavirus infection will remain a global health burden because of vaccination hesitancy, possible virus mutations, and the appearance of novel coronaviruses. Accordingly, there is a need for new approaches to limit severe illness stemming from coronavirus infections. Cells of the immune system and lung epithelia express receptors for GABA (GABA-Rs), a widely used neurotransmitter within the CNS. GABA-R agonists have anti-inflammatory effects and can limit acute lung injury. We previously showed that GABA treatment effectively reduced disease severity and death rates in mice following infection with a coronavirus (MHV-1) which provides a potentially lethal model of COVID-19. Here, we report that GABA treatment also reduced viral load in the lungs, suggesting that GABA-Rs may provide a new druggable target to limit pulmonary coronavirus replication. Histopathological analysis revealed that GABA treatment reduced lung inflammatory infiltrates and damages. Since GABA is safe for human consumption, inexpensive, and available worldwide, it is a promising candidate to help treat COVID-19.

category: immunology


title: SARS-CoV-2 RBD-Tetanus toxoid conjugate vaccine induces a strong neutralizing immunity in preclinical studies

abstract: Controlling the global COVID-19 pandemic depends, among other measures, on developing preventive vaccines at an unprecedented pace. Vaccines approved for use and those in development intend to use neutralizing antibodies to block viral sites binding to the hosts cellular receptors. Virus infection is mediated by the spike glycoprotein trimer on the virion surface via its receptor binding domain (RBD). Antibody response to this domain is an important outcome of the immunization and correlates well with viral neutralization. Here we show that macromolecular constructs with recombinant RBD conjugated to tetanus toxoid induce a potent immune response in laboratory animals. Some advantages of the immunization with the viral antigen coupled to tetanus toxoid have become evident such as predominant IgG immune response due to affinity maturation and long-term specific B-memory cells. This paper demonstrates that subunit conjugate vaccines can be an alternative for COVID-19, paving the way for other viral conjugate vaccines based on the use of small viral proteins involved in the infection process.


title: A novel antibody against the furin cleavage site of SARS-CoV-2 spike protein: effects on proteolytic cleavage and ACE2 binding

abstract: SARS-CoV-2 harbors a unique S1/S2 furin cleavage site within its spike protein, which can be cleaved by furin and other proprotein convertases. Proteolytic activation of SARS-CoV-2 spike protein at the S1/S2 boundary facilitates interaction with host ACE2 receptor for cell entry. To address this, high titer antibody was generated against the SARS-CoV-2-specific furin motif. Using a series of innovative ELISA-based assays, this furin site blocking antibody displayed high sensitivity and specificity for the S1/S2 furin cleavage site, and demonstrated effective blockage of both enzyme-mediated cleavage and spike-ACE2 interaction. The results suggest that immunological blocking of the furin cleavage site may afford a suitable approach to stem proteolytic activation of SARS-CoV-2 spike protein and curtail viral infectivity.


title: Soil, senescence and exudate utilisation: Characterisation of the Paragon var. spring bread wheat root microbiome

abstract: Conventional methods of agricultural pest control and crop fertilisation are contributing to a crisis of biodiversity loss, biogeochemical cycle dysregulation, and ecosystem collapse. Thus, we must find ecologically responsible means to control disease and promote crop yields. The root-associated microbiome may contribute to this goal as microbes can aid plants with disease suppression, abiotic stress relief, and nutrient bioavailability. We applied 16S rRNA gene & fungal 18S rRNA gene (ITS2 region) amplicon sequencing to profile the diversity of the bacterial, archaeal & fungal communities associated with the roots of UK elite spring bread wheat variety Triticum aestivum var. Paragon in different soils and developmental stages. This revealed that community composition shifted significantly for all three groups across compartments. This shift was most pronounced for bacteria and fungi, while we observed weaker selection on the ammonia oxidising archaea-dominated archaeal community. Across multiple soil types we found that soil inoculum was a significant driver of endosphere community composition, however several bacterial families were identified as core enriched taxa in all soil conditions. The most abundant of these were Streptomycetaceae and Burkholderiaceae. Moreover, as the plants senesce, both families were reduced in abundance, indicating that input from the living plant was required to maintain their abundance in the endosphere. To understand which microbes are using wheat root exudates in the rhizosphere, root exudates were labelled in a 13CO2 DNA stable isotope probing experiment. This shows that bacterial taxa within the Burkholderiaceae family among other core enriched taxa, such as Pseudomonadaceae, were able to use root exudates but Streptomycetaceae were not. Overall, this work provides a better understanding of the wheat microbiome, including the endosphere community. Understanding crop microbiome formation will contribute to ecologically responsible methods for yield improvement and biocontrol in the future.


title: Avian influenza A viruses reassort and diversify differently in mallards and mammals

abstract: Reassortment among co-infecting influenza A viruses (IAVs) is an important source of viral diversity and can facilitate expansion into novel host species. Indeed, reassortment played a key role in the evolution of the last three pandemic IAVs. Observed patterns of reassortment within a coinfected host are likely to be shaped by several factors, including viral load, the extent of viral mixing within the host and the stringency of selection. These factors in turn are expected to vary among the diverse host species that IAV infects. To investigate host differences in IAV reassortment, here we examined reassortment of two distinct avian IAV within their natural host (mallards) and a mammalian model system (guinea pigs). Animals were co-inoculated with A/wildbird/California/187718-36/2008 (H3N8) and A/mallard/Colorado/P66F1-5/2008 (H4N6) viruses. Longitudinal samples were collected from the cloaca of mallards or the nasal tract of guinea pigs and viral genetic exchange was monitored by genotyping clonal isolates from these samples. Relative to those in guinea pigs, viral populations in mallards showed higher frequencies of reassortant genotypes and were characterized by higher genotype richness and diversity. In line with these observations, analysis of pairwise segment combinations revealed lower linkage disequilibrium in mallards as compared to guinea pigs. No clear longitudinal patterns in richness, diversity or linkage disequilibrium were present in either host. Our results reveal mallards to be a highly permissive host for IAV reassortment and suggest that reduced viral mixing limits avian IAV reassortment in a mammalian host.


title: Differential impact of plant secondary metabolites on the soil microbiota

abstract: Plant metabolites can shape the microbial community composition in the soil. Two indole metabolites, benzoxazolinone (BOA) and gramine, produced by different Gramineae species, and quercetin, a flavonoid synthesized by many dicot species, were studied for their impacts on the community structure of soil bacteria. The three plant metabolites were directly added to agricultural soil over a period of 28 days. Alterations in bacterial composition were monitored by next generation sequencing of 16S rRNA gene PCR products and phospholipid fatty acid analysis. Treatment of the soil with the plant metabolites altered the composition of bacterial taxa on the phylum and genus levels. Alpha diversity was significantly altered by BOA or quercetin, but not by gramine. BOA treatment caused an increase in the relative abundances of only four genera, three of them belonging to the Actinobacteriota. Gramine or quercetin treatment resulted in the increase in relative abundance of 13 or 14 genera, respectively, most of them belonging to the Proteobacteria. The relative abundance of 22 genera was decreased after BOA treatment, 16 of which were also decreased by gramine or quercetin. Isolation and characterization of cultivable bacterial indicated an enrichment in specific Arthrobacter or Pseudomonas strains. Therefore, the effects of the treatments on soil bacteria were characteristic for each metabolite, with BOA exerting a broad-spectrum inhibitory effect, with only few genera able to proliferate, while gramine and quercetin caused the proliferation of many potentially beneficial strains. As a consequence, benzoxazolinone or gramine biosynthesis which have evolved in different barley species, is accompanied with the association with distinct bacterial communities in the soil, presumably after mutual adaptation during evolution.


title: Structure and efflux mechanism of the yeast pleiotropic drug resistance transporter Pdr5

abstract: Pdr5, a member of the extensive ABC transporter superfamily, is representative of a clinically relevant subgroup involved in pleiotropic drug resistance. Through the coupling of nucleotide hydrolysis with drug efflux, Pdr5 homologues enable pathogenic species to survive in the presence of chemically diverse antifungal agents. Our structural and functional results reveal details of an ATP-driven conformational cycle, which mechanically drives drug translocation through an amphipathic channel, and a clamping switch within a conserved linker loop that acts as a nucleotide sensor. One half of the transporter remains nearly invariant throughout the cycle, while its partner undergoes changes that are transmitted across interdomain interfaces to support a peristaltic motion of the pumped molecule. The efflux model proposed here rationalises the pleiotropic impact of Pdr5 and opens avenues for the development of effective antifungal compounds.


title: Novel gut pathobionts confound results in a widely used mouse model of human inflammatory disease

abstract: The mammalian gut microbiota consists of hundreds of anaerobic bacterial species that shape intestinal homeostasis and influence host immune responses. Although the causal roles of specific human gut bacterial species in health and disease are emerging, the role of indigenous gut bacteria in driving immunophenotypic variability in mouse models of human disease remains poorly understood. We performed a large-scale experiment using 579 laboratory mice designed to identify and validate the causes of disease variability in the widely used dextran sulphate sodium (DSS) mouse model of inflammatory bowel disease. Using microbiome analysis, coupled with machine learning and targeted anaerobic culturing, we identified and isolated the novel gut pathobiont species Duncaniella muricolitica and Alistipes okayasuensis and fulfilled Kochs postulates in mice to show that each pathobiont exerts dominant effects in the DSS model leading to variable treatment responses. We show these pathobiont species are common, but not ubiquitous, in mouse facilities around the world, raising experimental design opportunities for improved mouse models of human intestinal diseases.


title: Human monoclonal antibodies against Staphylococcus aureus surface antigens recognize in vitro biofilm and in vivo implant infections

abstract: Implant-associated Staphylococcus aureus infections are difficult to treat because of biofilm formation. Bacteria in a biofilm are often insensitive to antibiotics and host immunity. Monoclonal antibodies (mAbs) could provide an alternative approach to improve the diagnosis and/or treatment of biofilm-related infections. Here we show that mAbs targeting common surface components of S. aureus can recognize clinically relevant biofilm types. We identify two groups of antibodies: one group that uniquely binds S. aureus in biofilm state and one that recognizes S. aureus in both biofilm and planktonic state. In a mouse model, we show that mAb 4497 (recognizing wall teichoic acid (WTA)) specifically localizes to biofilm-infected implants. In conclusion, we demonstrate the capacity of several human mAbs to detect S. aureus biofilms in vitro and in vivo. This is an important first step to develop mAbs for imaging or treating S. aureus biofilms.


title: An adaptive compromise - Conflicting evolutionary pressures on arthropod borne Zika virus dinucleotide composition in mammalian hosts and mosquito vectors

abstract:
Most vertebrate RNA viruses show pervasive suppression of CpG and UpA dinucleotides, closely resembling the dinucleotide composition of host cell transcriptomes. In contrast, CpG suppression is absent in both invertebrate mRNA and RNA viruses that exclusively infect arthropods. Arthropod-borne (arbo) viruses are transmitted between vertebrate hosts by invertebrate vectors and thus encounter potentially conflicting evolutionary pressures in the different cytoplasmic environments. Using a newly developed Zika virus (ZIKV) model, we have investigated how demands for CpG suppression in vertebrate cells can be reconciled with potentially quite different compositional requirements in invertebrates, and how this affects ZIKV replication and transmission.

Mutant viruses with synonymously elevated CpG or UpA dinucleotide frequencies showed attenuated replication in vertebrate cell lines, which was rescued by knockout of the zinc-finger antiviral protein (ZAP). Conversely, in mosquito cells, ZIKV mutants with elevated CpG dinucleotide frequencies showed substantially enhanced replication compared to wildtype. Host-driven effects on virus replication attenuation and enhancement were even more apparent in mouse and mosquito models. Infections with CpG-or UpA-high ZIKV mutants in mice did not cause typical ZIKV-induced tissue damage and completely protected mice during subsequent challenge with wildtype virus, which demonstrates their potential as live-attenuated vaccines. In contrast, the CpG-high mutants displayed enhanced replication in Aedes aegypti mosquitoes and a larger proportion of mosquitoes carried infectious virus in their saliva.

These findings show that mosquito cells are also capable of discriminating RNA based on dinucleotide composition. However, the evolutionary pressure on the CpG dinucleotides of viral genomes in arthropod vectors directly opposes the pressure present in vertebrate host cells, which provides evidence that an adaptive compromise is required for arbovirus transmission. This suggests that the genome composition of arthropod-borne flaviviruses is crucial to maintain the balance between high-level replication in the vertebrate host and persistent replication in the mosquito vector.