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Objective: This study aimed to qualitatively examine how perceptions of cannabis differ among college students in an effort to better understand the changing landscape of cannabis on college campus...

BackgroundMeadow brown butterflies (Maniola jurtina) on the Isles of Scilly represent an ideal model in which to dissect the links between genotype, phenotype and long-term patterns of selection in the wild - a largely unfulfilled but fundamental aim of modern biology. To meet this aim, a clear description of genotype is required.FindingsHere we present the draft genome sequence of M. jurtina to serve as an initial genetic resource for this species. Seven libraries were constructed using DNA from multiple wild caught females and sequenced using Illumina, PacBio RSII and MinION technology. A novel hybrid assembly approach was employed to generate a final assembly with an N50 of 214 kb (longest scaffold 2.9 Mb). The genome encodes a total of 36,294 genes. 90.3% and 88.7% of core BUSCO (Benchmarking Universal Single-Copy Orthologs) Arthropoda and Insecta gene sets were recovered as complete single-copies from this assembly. Comparisons with 17 other Lepidopteran species placed 86.5% of the assembled genes in orthogroups.ConclusionsOur results provide the first high-quality draft genome and annotation of the butterfly M. jurtina.

Pathogenic mitochondrial (mt)DNA molecules can exhibit heteroplasmy in single cells and cause a range of clinical phenotypes, although their contribution to immunity is poorly understood. Here, in mice carrying heteroplasmic C5024T in mt-tRNAAla– that impairs oxidative phosphorylation – we found a reduced mutation burden in peripheral T and B memory lymphocyte subsets, compared to their naïve counterparts. Furthermore, selection diluting the mutation was inducedin vitroby triggering T and B cell antigen receptors. While C5024T dysregulated naïve CD8+T cell respiration and metabolic remodeling post-activation, these phenotypes were partially ameliorated by selection. Analogous to mice, peripheral blood memory T and B lymphocyte subsets from human MELAS (Mitochondrial Encephalomyopathy with Lactic Acidosis and Stroke-like episodes) patients – carrying heteroplasmic A3243G in mt-tRNALeu– displayed a reduced mutation burden, compared to naïve cells. In both humans and mice, mtDNA selection was observed in IgG+antigen-specific B cells after SARS-CoV-2 Spike vaccination, illustrating an on-going processin vivo. Taken together, these data illustrate purifying selection of pathogenic mtDNA variants during the oxidative phosphorylation checkpoints of the naïve-memory lymphocyte transition.HighlightsIn human MELAS patients (A3243G in mt-tRNALeu) and a related mouse model (C5024T in mt-tRNAAla), T and B memory subsets displayed a reduced mtDNA mutation burden compared to their naïve counterparts.Selection was observed in antigen-specific IgG+B cells after SARS-CoV-2 Spike protein vaccination.T and B cell antigen receptor stimulation triggered purifying selectionin vitro, facilitating mechanistic studies of mtDNA selection.Heteroplasmic pathogenic mutations in mtDNA dysregulated metabolic remodeling after lymphocyte activation and reduced macrophage OXPHOS capacity.

The Protein Data Bank (PDB) is the single global archive of experimentally determined three-dimensional (3D) structure data of biological macromolecules. Since 2003, the PDB has been managed by the Worldwide Protein Data Bank (wwPDB; wwpdb.org), an international consortium that collaboratively oversees deposition, validation, biocuration, and open access dissemination of 3D macromolecular structure data. The PDB Core Archive houses 3D atomic coordinates of more than 144 000 structural models of proteins, DNA/RNA, and their complexes with metals and small molecules and related experimental data and metadata. Structure and experimental data/metadata are also stored in the PDB Core Archive using the readily extensible wwPDB PDBx/mmCIF master data format, which will continue to evolve as data/metadata from new experimental techniques and structure determination methods are incorporated by the wwPDB. Impacts of the recently developed universal wwPDB OneDep deposition/validation/biocuration system and various methods-specific wwPDB Validation Task Forces on improving the quality of structures and data housed in the PDB Core Archive are described together with current challenges and future plans.

Summary Neurodegenerative disorders are an increasingly common and irreversible burden on society, often affecting the aging population, but their etiology and disease mechanisms are poorly understood. Studying monogenic neurodegenerative diseases with known genetic cause provides an opportunity to understand cellular mechanisms also affected in more complex disorders. We recently reported that loss-of-function mutations in the autophagy adaptor protein SQSTM1/p62 lead to a slowly progressive neurodegenerative disease presenting in childhood. To further elucidate the neuronal involvement, we studied the cellular consequences of loss of p62 in a neuroepithelial stem cell (NESC) model and differentiated neurons derived from reprogrammed p62 patient cells or by CRISPR/Cas9-directed gene editing in NESCs. Transcriptomic and proteomic analyses suggest that p62 is essential for neuronal differentiation by controlling the metabolic shift from aerobic glycolysis to oxidative phosphorylation required for neuronal maturation. This shift is blocked by the failure to sufficiently downregulate lactate dehydrogenase expression due to the loss of p62, possibly through impaired Hif-1α downregulation and increased sensitivity to oxidative stress. The findings imply an important role for p62 in neuronal energy metabolism and particularly in the regulation of the shift between glycolysis and oxidative phosphorylation required for normal neurodifferentiation., Graphical Abstract, Highlights • SQSTM1/p62 is dispensable for mitophagy in a human neuronal model • SQSTM1/p62 is needed for proper neuronal differentiation • Loss of SQSTM1/p62 leads to increased oxidative stress and poor LDHA downregulation • SQSTM1/p62 is essential for the shift to OXPHOS during neuronal differentiation, SQSTM1/p62 is a known autophagy adaptor that, if lost, causes childhood-onset neurodegeneration. Data from Wredenberg et al. show that loss of p62 in a neuronal stem cell model does not affect mitophagy but instead leads to impaired differentiation. The authors suggest p62 finely tunes LDHA expression and thus controls the metabolic shift to OXPHOS required for proper differentiation.

Summary Here, we present a revised protocol to derive neuroepithelial stem (NES) cells from human induced pluripotent stem cells. NES cells can be further differentiated into a culture of neurons (90%) and glia (10%). We describe how to derive and maintain NES cells in culture and how to differentiate them. In addition, we show the potential use of NES cells to study the role of reactive oxygen species in neuronal differentiation and a guideline for NES cell transfection. For complete details on the use and execution of this protocol, please refer to Calvo-Garrido et al. (2019); Falk et al. (2012)., Graphical abstract, Highlights • NES cells, neurons, and glia generated through dual SMAD inhibition • Proliferate, maintain, and differentiate NES cells into neurons and glia in culture • Coating of the growth surface, cell density, and trypsinization need to be adjusted • Feasible model to study ROS effect in neuronal and glia differentiation, Here, we present a revised protocol to derive neuroepithelial stem (NES) cells from human induced pluripotent stem cells. NES cells can be further differentiated into a culture of neurons (90%) and glia (10%). We describe how to derive and maintain NES cells in culture and how to differentiate them. In addition, we show the potential use of NES cells to study the role of reactive oxygen species in neuronal differentiation and a guideline for NES cell transfection.

Attempts to control insect pests and disease vectors have a long history. Recently, new technology has opened a whole new range of possible methods to suppress or transform natural populations. But it has also become clear that a better understanding of the ecology of targeted populations is needed. One key parameter is mating behaviour. Often modified males are released which need to successfully reproduce with females while competing with wild males. Insect control techniques can be affected by target species’ mating ecology, and conversely mating ecology is likely to evolve in response to manipulation attempts. A better understanding of (female) mating behaviour will help anticipate and overcome potential challenges, and thus make desirable outcomes more likely.

A new study using artificial selection reveals that the size of the sex comb on the legs of male flies is genetically correlated with their fertility success under conditions of sperm competition.

Gene drive is a naturally occurring phenomenon in which selfish genetic elements manipulate gametogenesis and reproduction to increase their own transmission to the next generation. Currently, there is great excitement about the potential of harnessing such systems to control major pest and vector populations. If synthetic gene drive systems can be constructed and applied to key species, they may be able to rapidly spread either modifying or eliminating the targeted populations. This approach has been lauded as a revolutionary and efficient mechanism to control insect-borne diseases and crop pests. Driving endosymbionts have already been deployed to combat the transmission of dengue and Zika virus in mosquitoes. However, there are a variety of barriers to successfully implementing gene drive techniques in wild populations. There is a risk that targeted organisms will rapidly evolve an ability to suppress the synthetic drive system, rendering it ineffective. There are also potential risks of synthetic gene drivers invading non-target species or populations. This Special Feature covers the current state of affairs regarding both natural and synthetic gene drive systems with the aim to identify knowledge gaps. By understanding how natural drive systems spread through populations, we may be able to better predict the outcomes of synthetic drive release.

Summary: The Scandinavian winter-swimming culture combines brief dips in cold water with hot sauna sessions, with conceivable effects on body temperature. We study thermogenic brown adipose tissue (BAT) in experienced winter-swimming men performing this activity 2–3 times per week. Our data suggest a lower thermal comfort state in the winter swimmers compared with controls, with a lower core temperature and absence of BAT activity. In response to cold, we observe greater increases in cold-induced thermogenesis and supraclavicular skin temperature in the winter swimmers, whereas BAT glucose uptake and muscle activity increase similarly to those of the controls. All subjects demonstrate nocturnal reduction in supraclavicular skin temperature, whereas a distinct peak occurs at 4:30–5:30 a.m. in the winter swimmers. Our data leverage understanding of BAT in adult human thermoregulation, suggest both heat and cold acclimation in winter swimmers, and propose winter swimming as a potential strategy for increasing energy expenditure.

Selfish genetic elements (SGEs) are diverse and near ubiquitous in Eukaryotes and can be potent drivers of evolution. Here, we discuss SGEs that specifically act on sperm to gain a transmission advantage to the next generation. The diverse SGEs that affect sperm often impose costs on carrier males, including damaging ejaculates, skewing offspring sex ratios and in particular reducing sperm-competitive success of SGE-carrying males. How males and females tolerate and mitigate against these costs is a dynamic and expanding area of research. The intense intra-genomic conflict that these selfish elements generate could also have implications for male fertility and spermatogenesis more widely.This article is part of the theme issue ‘Fifty years of sperm competition’.

New findings What is the central question of this study? Does blockade of the IL-6 receptor by tocilizumab inhibit immune cell mobilization to the blood stream in humans during an acute bout of exercise? What is the main finding and its importance? Blockade of IL-6 receptor signalling by tocilizumab attenuates mobilization of NK cells and dendritic cells to the blood stream during exercise. This implies an inhibitory effect of tocilizumab on the innate immune response to physical stress, which could be considered in clinical settings. Abstract Immune cells are recruited from their storage organs and the endothelial walls to the blood stream in response to physiological stress. This is essential for the recognition and clearing of infected, transformed or damaged cells. One of the most potent stimuli to recruit immune cells to the circulation is exercise. Accordingly, exercise has proven beneficial in disease settings, such as cancer and diabetes. Interleukin-6 (IL-6) is released from contracting skeletal muscle in response to exercise, and rodent studies have established a link between exercise-induced IL-6 and recruitment of natural killer (NK) cells. Whether exercise-induced IL-6 is involved in regulating NK cell mobilization in humans is unclear. This study explored the effect of IL-6 receptor blockade on immune cell mobilization during an acute bout of exercise in humans. In a randomized, placebo-controlled clinical study, abdominally obese humans receiving placebo infusions or tocilizumab infusions performed an acute bout of exercise before and after the intervention. Immune cell recruitment was measured by flow cytometry. IL-6 receptor blockade attenuated the increase of NK cells by 53% (mean difference -0.49 (95% CI: -0.89 to -0.08) × 109 cells L-1 , P

Pathogenic variants in FBXL4 cause a severe encephalopathic syndrome associated with mtDNA depletion and deficient oxidative phosphorylation. To gain further insight into the enigmatic pathophysiology caused by FBXL4 deficiency, we generated homozygous Fbxl4 knockout mice and found that they display a predominant perinatal lethality. Surprisingly, the few surviving animals are apparently normal until the age of 8–12 months when they gradually develop signs of mitochondrial dysfunction and weight loss. One‐year‐old Fbxl4 knockouts show a global reduction in a variety of mitochondrial proteins and mtDNA depletion, whereas lysosomal proteins are upregulated. Fibroblasts from patients with FBXL4 deficiency and human FBXL4 knockout cells also have reduced steady‐state levels of mitochondrial proteins that can be attributed to increased mitochondrial turnover. Inhibition of lysosomal function in these cells reverses the mitochondrial phenotype, whereas proteasomal inhibition has no effect. Taken together, the results we present here show that FBXL4 prevents mitochondrial removal via autophagy and that loss of FBXL4 leads to decreased mitochondrial content and mitochondrial disease., A new Fbxl4 knock‐out mouse model was generated and compared well in terms of disease pathophysiology with findings in patient fibroblasts and a CRISPR/Cas9 knock‐out cell line.

Sweden has one neonatal screening laboratory, receiving 115 to 120 thousand samples per year. Among the one million babies screened by tandem mass spectrometry from November 2010 until July 2019, a total of 665 babies were recalled and 311 verified as having one of the diseases screened for with this methodology, giving a positive predictive value (PPV) of 47% and an incidence of 1:3200. The PPV was high (41%) already in the first year after start of screening, thanks to the availability of the collaborative project Region 4 Stork database. The PPV is presently 58%. This improvement was achieved by the implementation of second-tier analyses in the screening for methylmalonic aciduria, propionic aciduria, isovaleric aciduria, and homocystinuria, and the employment of various post analytical tools of the Region 4 Stork, and its successor the collaborative laboratory integrated reports.

Identifying loci associated with a phenotype is a critical step in many evolutionary studies. Most methods require large sample sizes or breeding designs that can be prohibitively difficult. Here we apply a rarely used approach to identify SNP loci associated with a complex phenotype. We mate siblings from isofemale lines isolate genotypes from three wild populations. After phenotyping we perform whole genome sequencing of isofemale lines from the extremes of the phenotypic distribution of each population and identify SNPs that are consistently fixed for alternative alleles across line pairs. The focal phenotype is female remating rate in the fly Drosophila pseudoobscura, defined as the willingness of a female to mate with a second male after her first mating. This is an integral part of mating system evolution, sexual selection and sexual conflict, and is a quantitative polygenic trait.About 200 SNPs are consistently fixed for alternate alleles in the three pairs of isofemale lines. We use different simulation approaches to explore how many SNPs would be expected to be fixed. We find the surprising result that we uncover fewer observed fixed SNPs than are expected by either simulation approach. We also complete functional analyses of these SNPs. Many lie near genes or regulatory regions known to be involved in Drosophila courtship and mating behaviours, and some have previously been associated with re-mating rates in Genome-Wide Association Studies. Given the small sample size, these results should be treated with caution. Nevertheless, this study suggests that even from a relatively small number of isofemale lines established from wild populations, it is possible to identify candidate loci potentially associated with a complex quantitative trait. However, further work is required to understand modelling the expected distribution of differences.

Context Interleukin 6 (IL-6) contributes to bone remodeling in preclinical studies. Clinical trials investigating the role of IL-6 in bone remodeling are limited. Objective To investigate if IL-6 regulates bone remodeling in humans. Design Plasma concentrations of the bone resorption marker carboxy-terminal type I collagen crosslinks (CTX) and of the bone formation marker procollagen type 1 N-terminal propeptide (P1NP) were measured during a mixed-meal tolerance test (MMTT) in 3 placebo-controlled human studies. Participants Five healthy individuals participated in study 1; 52 obese individuals, in study 2; and 10 healthy individuals, in study 3. Interventions Study 1 was a single-blinded crossover study consisting of a 1-h infusion of saline (placebo) or the IL-6 receptor antibody tocilizumab followed by an exercise bout. Study 2 was a randomized, double-blinded 12-week exercise training intervention study. Participants received infusions of saline or tocilizumab. Study 3 was a randomized, double-blinded, crossover study consisting of 30 min infusion of saline or IL-6. Main outcomes measures Effect of IL-6 on CTX levels. Results CTX was significantly (P < 0.01) decreased during MMTTs in all 3 studies. Treatment with tocilizumab did not affect exercise or meal induced changes in plasma CTX or P1NP concentrations acutely (study 1) or after a 12-week treatment period (study 2). Exogenous IL-6 had no effect on CTX or P1NP plasma concentrations (study 3). Conclusions IL-6 may not regulate bone remodeling in humans.

To investigate human brown adipose tissue (BAT) physiology, we recruited young healthy men that were either winter swimmers (WS) or controls. We used 2-deoxy-2-[18F]fluoro-D-glucose PET/MRI-scanning, infrared thermography, iButtons and ingestible thermometer pills to assess BAT glucose uptake, and BAT and core temperature. In a crossover design with interventions performed in randomized order, subjects performed individualized cooling and thermoneutral protocols. We monitored a subset for BAT circadian rhythm. In response to cooling, WS increased more in BAT temperature, BAT glucose uptake and energy expenditure compared to control subjects. At thermoneutrality, WS had higher BAT temperature and lower core temperature, despite BAT glucose uptake in controls only. All subjects demonstrated a nocturnal reduction in BAT temperature, while a peak was observed at 4:30-5:30 am in WS, coinciding with rising plasma cortisol levels. In conclusion, WS have highly regulated BAT, and BAT is active at thermoneutrality in a circadian rhythm in humans.

Understanding the evolution and spread of insecticide resistance requires knowing the relative fitness of resistant organisms. In the absence of insecticides, resistance is predicted to be costly. The Drosophila melanogaster DDT resistance allele (DDT-R) is associated with a male mating cost. This could be because resistant males are generally smaller, but DDT-R may also alter courtship behaviours. Here we tested for body size and courtship effects of DDT-R on mating success in competitive and non-competitive mating trials respectively. We also assessed relative aggression in resistant and susceptible males because aggression can also influence mating success. While the effect of DDT-R on male size partly contributed to reduced mating success, resistant males also had lower rates of courtship and were less aggressive than susceptible males. These differences contribute to the observed DDT-R mating costs. Additionally, these pleiotropic effects of DDT-R are consistent with the history and spread of resistance alleles in nature. Electronic supplementary material The online version of this article (doi:10.1007/s10519-017-9850-6) contains supplementary material, which is available to authorized users.

co-auteur étranger; International audience; The Trematoda are a group of phylogenetically diverse metazoan parasites that exhibit complex life cycles that often pass through invertebrate and vertebrate hosts. Some trematodes influence their host's behaviour to benefit transmission. Their parasitic influence may impact host population size by inhibiting an individual's reproductive capacity. We assessed the impact of infection by Podocotyle atomon on the reproductive behaviour and fecundity of its amphipod intermediate host, Gammarus zaddachi, using laboratory and field studies. Parasite prevalence was high in the field, with males more likely to be infected (prevalence in males 64%, in females 39%). Males also suffered a higher parasite burden than females. Infected females were less active, but we found no evidence for a reduction in female reproductive success. Infected females also had comparable pairing success to uninfected females. In males, infection reduced survival and fecundity, with mortality being highest, and sperm numbers lowest, in heavily infected individuals. Trematode parasites are sometimes associated with altered host fecundity, but studies often lack the relevant experimental data to explore the evolution of the trait. We discuss this among information specific to the effect of P. atomon infection in G. zaddachi.

Fluctuating asymmetry (FA), defined as random deviations from perfect bilateral symmetry, is assumed to reflect developmental instability. FA is predicted to increase in response to environmental stress, including parasite infection. In addition, based on theory we predict a higher FA in sexually selected traits, due to their greater sensitivity to stress. We investigated the relationships between FA, parasitism and reproductive fitness in 2 species of gammarid crustacean, incorporating both sexual and non-sexual traits. We tested the hypothesis that gammarids infected by vertically transmitted Microsporidia will display higher levels of FA than those infected by horizontally transmitted trematodes, because vertically transmitted Microsporidia can be present at the earliest stages of host development. We found little evidence for a relationship between FA and fecundity in Gammarus spp.; however, egg diameter for infected female Gammarus duebeni was significantly smaller than uninfected female G. duebeni. FA was not correlated with brood size in females or with sperm number in males. In contrast to our prediction, we report a lower relative FA in response to sexual traits than non-sexual traits. However, FA in sexual traits was found to be higher in males than females, supporting the theory that sexual selection leads to increased FA. Additionally, we report a negative correlation between FA and both trematode (Podocotyle atomon) and PCR-positive microsporidian (Nosema granulosis and Dictyocoela duebenum) infections and interpret these results in the context of the parasites' transmission strategies. FA in G. duebeni and G. zaddachi appears to associate with trematode and microsporidian presence, although reproductive fitness is less altered by infection.

Infertility is common in nature despite its obvious cost to individual fitness. Rising global temperatures are predicted to decrease fertility, and male sterility is frequently used in attempts to regulate pest or disease vector populations. When males are infertile, females may mate with multiple males to ensure fertilization, and changes in female mating behavior in turn could intensify selection on male fertility. Fertility assurance is a potentially wide-spread explanation for polyandry, but whether and how it actually contributes to the evolution of polyandry is not clear. Moreover, whether a drop in male fertility would lead to a genetic increase in polyandry depends on whether females respond genetically or through behavioral plasticity to male infertility. Here, we experimentally manipulate male fertility through heat-exposure in Drosophila pseudoobscura, and test female discrimination against infertile males before and after mating. Using isogenic lines, we compare the roles of behaviorally plastic versus genetically fixed polyandry. We find that heat-exposed males are less active and attractive, and that females are more likely to remate after mating with these males. Remating rate increases with reduced reproductive output, indicating that females use current sperm storage threshold to make dynamic remating decisions. After remating with fertile males, females restore normal fecundity levels. Our results suggest that male infertility could explain the evolution of adaptively flexible polyandry, but is less likely to cause an increase in genetic polyandry.

The RNA helicase SUV3 and the polynucleotide phosphorylase PNPase are involved in the degradation of mitochondrial mRNAs but their roles in vivo are not fully understood. Additionally, upstream processes, such as transcript maturation, have been linked to some of these factors, suggesting either dual roles or tightly interconnected mechanisms of mitochondrial RNA metabolism. To get a better understanding of the turn-over of mitochondrial RNAs in vivo, we manipulated the mitochondrial mRNA degrading complex in Drosophila melanogaster models and studied the molecular consequences. Additionally, we investigated if and how these factors interact with the mitochondrial poly(A) polymerase, MTPAP, as well as with the mitochondrial mRNA stabilising factor, LRPPRC. Our results demonstrate a tight interdependency of mitochondrial mRNA stability, polyadenylation and the removal of antisense RNA. Furthermore, disruption of degradation, as well as polyadenylation, leads to the accumulation of double-stranded RNAs, and their escape out into the cytoplasm is associated with an altered immune-response in flies. Together our results suggest a highly organised and inter-dependable regulation of mitochondrial RNA metabolism with far reaching consequences on cellular physiology., Author summary Although a number of factors have been implemented in the turnover of mitochondrial (mt) DNA-derived transcripts, their exact functions and interplay with one another is not entirely clear. Several of these factors have been proposed to co-ordinately regulate both transcript maturation, as well as degradation, but the order of events during mitochondrial RNA turnover is less well understood. Using a range of different genetically modified Drosophila melanogaster models, we studied the involvement of the RNA helicase SUV3, the polynucleotide phosphorylase PNPase, the leucine-rich pentatricopeptide repeat motif-containing protein LRPPRC, and the mitochondrial RNA poly(A) polymerase MTPAP, in stabilisation, polyadenylation, and degradation of mitochondrial transcripts. Our results show a tight collaborative activity of these factors in vivo and reveal a clear hierarchical order of events leading to mitochondrial mRNA maturation. Furthermore, we demonstrate that the loss of SUV3, PNPase, or MTPAP leads to the accumulation of mitochondrial-derived antisense RNA in the cytoplasm of cells, which is associated with an altered immune-response in flies.

Background Since different types of genetic variants, from single nucleotide variants (SNVs) to large chromosomal rearrangements, underlie intellectual disability, we evaluated the use of whole-genome sequencing (WGS) rather than chromosomal microarray analysis (CMA) as a first-line genetic diagnostic test. Methods We analyzed three cohorts with short-read WGS: (i) a retrospective cohort with validated copy number variants (CNVs) (cohort 1, n = 68), (ii) individuals referred for monogenic multi-gene panels (cohort 2, n = 156), and (iii) 100 prospective, consecutive cases referred to our center for CMA (cohort 3). Bioinformatic tools developed include FindSV, SVDB, Rhocall, Rhoviz, and vcf2cytosure. Results First, we validated our structural variant (SV)-calling pipeline on cohort 1, consisting of three trisomies and 79 deletions and duplications with a median size of 850 kb (min 500 bp, max 155 Mb). All variants were detected. Second, we utilized the same pipeline in cohort 2 and analyzed with monogenic WGS panels, increasing the diagnostic yield to 8%. Next, cohort 3 was analyzed by both CMA and WGS. The WGS data was processed for large (> 10 kb) SVs genome-wide and for exonic SVs and SNVs in a panel of 887 genes linked to intellectual disability as well as genes matched to patient-specific Human Phenotype Ontology (HPO) phenotypes. This yielded a total of 25 pathogenic variants (SNVs or SVs), of which 12 were detected by CMA as well. We also applied short tandem repeat (STR) expansion detection and discovered one pathologic expansion in ATXN7. Finally, a case of Prader-Willi syndrome with uniparental disomy (UPD) was validated in the WGS data. Important positional information was obtained in all cohorts. Remarkably, 7% of the analyzed cases harbored complex structural variants, as exemplified by a ring chromosome and two duplications found to be an insertional translocation and part of a cryptic unbalanced translocation, respectively. Conclusion The overall diagnostic rate of 27% was more than doubled compared to clinical microarray (12%). Using WGS, we detected a wide range of SVs with high accuracy. Since the WGS data also allowed for analysis of SNVs, UPD, and STRs, it represents a powerful comprehensive genetic test in a clinical diagnostic laboratory setting.

The rapid evolutionary divergence of male genital structures under sexual selection is well documented. However, variation in female genital traits and the potential for sexual conflict to drive the coevolution between male and female traits has only recently received attention. In many lepidopterans, females possess genital teeth (collectively, signa). Comparative studies suggest these teeth, involved in the deflation of spermatophores, may have coevolved with male spermatophore thickness via sexually antagonistic coevolution in a contest over the rate of deflation of spermatophores within the reproductive tract. We tested the hypothesis that sexual conflict should generate coevolution between genital teeth and spermatophore morphology by examining these traits under experimental manipulation of sexual conflict intensity. Using micro-CT scanning, we examined spermatophore and teeth morphology in populations of the Indian moth, Plodia interpunctella, which had been evolving for 110 generations under different adult sex-ratio biases. We found divergence in female signa morphology in response to sexual conflict: females from female-biased populations (reduced sexual conflict) developed wider signa. However, we found no evidence of coevolution between signa traits and spermatophore thickness as reported from comparative studies.

What drives mating system variation is a major question in evolutionary biology. Female multiple mating (polyandry) has diverse evolutionary consequences, and there are many potential benefits and costs of polyandry. However, our understanding of its evolution is biased towards studies enforcing monandry in polyandrous species. What drives and maintains variation in polyandry between individuals, genotypes, populations and species remains poorly understood. Genetic variation in polyandry may be actively maintained by selection, or arise by chance if polyandry is selectively neutral. In Drosophila pseudoobscura, there is genetic variation in polyandry between and within populations. We used isofemale lines to found replicate populations with high or low initial levels of polyandry and tracked polyandry under experimental evolution over seven generations. Polyandry remained relatively stable, reflecting the starting frequencies of the experimental populations. There were no clear fitness differences between high versus low polyandry genotypes, and there was no signature of balancing selection. We confirmed these patterns in direct comparisons between evolved and ancestral females and found no consequences of polyandry for female fecundity. The absence of differential selection even when initiating populations with major differences in polyandry casts some doubt on the importance of polyandry for female fitness.

The aim was to determine disease-causing variants in the GALT gene which codes for the enzyme galactose-1-phosphate uridylyltransferase. Loss of activity of this enzyme causes classical galactosemia-a life threatening, treatable disorder, included in the Swedish newborn screening program since 1967. A total of 66 patients with the disease are known in Sweden and 56 index patients were investigated. An additional two patients with Duarte galactosemia were included. The disease-causing variants were identified in all patients. As reported from other countries only a few variants frequently recur in severe disease. The two variants p.(Gln188Arg) (c.563A>G) and p.(Met142Lys) (c.425T>A) are present in several index patients whereas the remaining are found in one to three patients each. The most common variant, p.(Gln188Arg), has an allele frequency of 51% in the cohort. A total of 16 novel variants were found among the 33 different variants in the cohort. Two of these are synonymous variants affecting splicing, demonstrating the importance of the evaluation of synonymous variants at the cDNA level. Concise sentence: Galactosemia is a rare disease in Sweden and the disease-causing variants are heterogenous including two synonymous variants.

Adenosine kinase (ADK) deficiency (OMIM [online mendelian inheritance in man]: 614300) is an autosomal recessive disorder of adenosine and methionine metabolism, with a unique clinical phenotype, mainly involving the central nervous system and dysmorphic features. Patients usually present early in life with sepsis-like symptoms, respiratory difficulties, and neonatal jaundice. Subsequently, patients demonstrate hypotonia and global developmental delay. Biochemically, methionine is elevated with normal homocysteine levels and the diagnosis is confirmed through molecular analysis of the ADK gene. There is no curative treatment; however, a methionine-restricted diet has been tried with variable outcomes. Herein, we report a 4-year-old Saudi female with global developmental delay, hypotonia, and dysmorphic features. Interestingly, she has a tall stature, developmental dysplasia of the hip, optic nerve gliosis, and tigroid fundus. We found a mutation not reported previously and we compared the current case with previously reported cases. We alert clinicians to consider ADK deficiency in any neonate presenting with global developmental delay, hypotonia, dysmorphic features, and high methionine levels.

Drosophila melanogaster has been a working horse of genetics and cell biology for more than a century. However, proteomic-based methods have been limited due to technical obstacles, especially the lack of reliable labelling methods. Here, we advanced a chemically defined food source into stable-isotope labelling of amino acids in flies (SILAF). It allows for the rapid generation of a large number of flies with full incorporation of lysine-6. SILAF followed by fractionation and enrichment gave proteomic insights at a depth of 5,966 proteins and 7,496 phosphorylation sites, which substantiated metabolic regulation on enzymatic level. Furthermore, the label can be traced and predicts protein turnover rates during different developmental stages. The ease and versatility of the method actuates the fruit fly as an appealing model in proteomic and post-translational modification studies and it enlarges potential metabolic applications based on heavy amino acid diets.

Understanding selection in the wild remains a major aim of evolutionary ecology and work by Ford and colleagues on the meadow brown butterfly Maniola jurtina did much to ignite this agenda. A great deal of their work was conducted during the 1950s on the Isles of Scilly. They documented island-specific wing-spot patterns that remained consistent over about a decade, but patterns on some islands changed after environmental perturbation. It was suggested that these wing-spot patterns reflected island-specific selection and that there was little migration between islands. However, genetic studies to test the underlying assumption of restricted migration are lacking and it is also unknown whether the originally described wing-spot patterns have persisted over time. We therefore collected female butterflies from five of Ford's original study locations, including three large islands (St Mary's, St Martin's and Tresco) and two small islands (Tean and St Helen's). Wing-spot patterns had not changed appreciably over time on three of the islands (two large and one small), but were significantly different on the other two. Furthermore, analysis of 176 amplified fragment length polymorphisms revealed significant genome-wide differentiation among the five islands. Our findings are consistent with Ford's conclusions that despite the close proximity of these islands, there is restricted gene flow among them.

Inherited methylation disorders are a group of rarely reported, probably largely underdiagnosed disorders affecting transmethylation processes in the metabolic pathway between methionine and homocysteine. These are methionine adenosyltransferase I/III, glycine N-methyltransferase, S-adenosylhomocysteine hydrolase and adenosine kinase deficiencies. This paper provides the first consensus recommendations for the diagnosis and management of methylation disorders. Following search of the literature and evaluation according to the SIGN-methodology of all reported patients with methylation defects, graded recommendations are provided in a structured way comprising diagnosis (clinical presentation, biochemical abnormalities, differential diagnosis, newborn screening, prenatal diagnosis), therapy and follow-up. Methylation disorders predominantly affect the liver, central nervous system and muscles, but clinical presentation can vary considerably between and within disorders. Although isolated hypermethioninemia is the biochemical hallmark of this group of disorders, it is not always present, especially in early infancy. Plasma S-adenosylmethionine and S-adenosylhomocysteine are key metabolites for the biochemical clarification of isolated hypermethioninemia. Mild hyperhomocysteinemia can be present in all methylation disorders. Methylation disorders do not qualify as primary targets of newborn screening. A low-methionine diet can be beneficial in patients with methionine adenosyltransferase I/III deficiency if plasma methionine concentrations exceed 800 μmol/L. There is some evidence that this diet may also be beneficial in patients with S-adenosylhomocysteine hydrolase and adenosine kinase deficiencies. S-adenosylmethionine supplementation may be useful in patients with methionine adenosyltransferase I/III deficiency. Recommendations given in this article are based on general principles and in practice should be adjusted individually according to patient’s age, severity of the disease, clinical and laboratory findings. Electronic supplementary material The online version of this article (doi:10.1007/s10545-016-9972-7) contains supplementary material, which is available to authorized users.

The maintenance of multiple morphs in warning signals is enigmatic because directional selection through predator avoidance should lead to the rapid loss of such variation. Opposing natural and sexual selection is a good candidate driving the maintenance of multiple male morphs but it also includes another enigma: when warning signal efficiency differs between male morphs, why would females choose a phenotype with lower survival? We tested the hypothesis that indirect responses to selection on correlated characters through sexual selection may substantially shape the evolution of male coloration. If male phenotypes differ in their fertilization ability, female choice against the best surviving phenotype can evolve. The wood tiger moth, Parasemia plantaginis , has two coexisting male morphs in Europe. Previous studies have shown that yellow males are better defended against predators, but that white males have a higher mating success. We examined differences in fertility between white and yellow males in terms of sperm production, number of sperm transferred and rate of sperm replenishment, and association between these fertility traits, female mate choice and reproductive output. If white morphs have greater fertility than yellow males, then this could explain why females prefer to mate with white males. However, we did not find any difference between male colour morphs either in mating probability, fertility (i.e. sperm availability and sperm transferred) or reproductive success (i.e. number of eggs laid and hatching success). We discuss our results in relation to context-dependent mating success and maintenance of colour polymorphism within populations.

The seemingly transparent wings of many insects have recently been found to display dramatic structural coloration. These structural colours (wing interference patterns: WIPs) may be involved in species recognition and mate choice, yet little is known about the evolutionary processes that shape them. Additionally, existing research has been restricted by analysing WIPs without due consideration of how they are actually perceived by the viewers’ colour vision. Here, we use multispectral digital imaging and a model ofDrosophilavision to compare WIPs of male and femaleDrosophila simulansfrom replicate populations forced to evolve with or without sexual selection for 68 generations. We show for the first time that WIPs modelled inDrosophilavision evolve in response to sexual selection, and confirm that WIPs correlate with male sexual attractiveness. These findings add a new element to the otherwise well describedDrosophilacourtship display and confirm that wing colours evolve through sexual selection.

Congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency is caused by mutations in the CYP21A2 gene and is often fatal in its classic forms if not treated with glucocorticoids. In contrast, non-classic CAH (NCCAH), with a prevalence from 0.1 % up to a few percentages in certain ethnic groups, only results in mild partial cortisol insufficiency and patients survive without treatment. Most NCCAH cases are never identified, but unnecessary suffering due to hyperandrogenism, especially in females, can be avoided by a correct diagnosis. A 17-hydroprogesterone (17OHP) level above 300 nmol/L indicates classic CAH while 30-300 nmol/L in adult males or females (follicular phase or if anovulatoric) indicates NCCAH. The gold standard for diagnosing NCCAH is the ACTH stimulation test. Deletion, large gene conversions, and nine microconversion-derived mutations are the most common CYP21A2 mutations. However, almost 200 rare mutations have been described. Since there is a good genotype-phenotype relationship, genotyping provides valuable diagnostic, as well as prognostic information. Neonatal screening for CAH is now performed in an increasing number of countries with the main goal of reducing mortality and morbidity due to salt-losing adrenal crises in the newborn period. In addition, screening may shorten the time to diagnosis in virilized girls. Neonatal screening misses some patients with milder classic CAH and most NCCAH cases. In conclusion, diagnosing classic CAH is life-saving, but diagnosing NCCAH is also important to prevent unnecessary suffering.

Massively parallel DNA sequencing has revolutionized analyses of human genetic variation. From having been out of reach for individual research groups and even more so for clinical diagnostic laboratories until recently, it is now possible to analyse complete human genomes within reasonable time frames and at a reasonable cost using technologies that are becoming increasingly available. This represents a huge advance in our ability to provide correct diagnoses for patients with rare inherited disorders and their families. This paradigm shift is especially dramatic within the area of monogenic disorders. Not only can rapid and safe diagnostics of virtually all known single-gene defects now be established, but novel causes of disease in previously unsolved cases can also be identified, illuminating novel pathways important for normal physiology. This greatly increases the capability not only to improve management of rare disorders, but also to improve understanding of pathogenetic mechanisms relevant for common, complex diseases.

Mitochondrial gene expression is largely regulated by post-transcriptional mechanisms that control the amount and translation of each mitochondrial mRNA. Despite its importance for mitochondrial function, the mechanisms and proteins involved in mRNA turnover are still not fully characterized. Studies in yeast and human cell lines have indicated that the mitochondrial helicase SUV3, together with the polynucleotide phosphorylase, PNPase, composes the mitochondrial degradosome. To further investigate the in vivo function of SUV3 we disrupted the homolog of SUV3 in Drosophila melanogaster (Dm). Loss of dmsuv3 led to the accumulation of mitochondrial mRNAs, without increasing rRNA levels, de novo transcription or decay intermediates. Furthermore, we observed a severe decrease in mitochondrial tRNAs accompanied by an accumulation of unprocessed precursor transcripts. These processing defects lead to reduced mitochondrial translation and a severe respiratory chain complex deficiency, resulting in a pupal lethal phenotype. In summary, our results propose that SUV3 is predominantly required for the processing of mitochondrial polycistronic transcripts in metazoan and that this function is independent of PNPase.

BACKGROUND: Coenzyme Q is an essential mitochondrial electron carrier, redox cofactor and a potent antioxidant in the majority of cellular membranes. Coenzyme Q deficiency has been associated with a range of metabolic diseases, as well as with some drug treatments and ageing. METHODS: We used whole exome sequencing (WES) to investigate patients with inherited metabolic diseases and applied a novel ultra-pressure liquid chromatography-mass spectrometry approach to measure coenzyme Q in patient samples. RESULTS: We identified a homozygous missense mutation in the COQ7 gene in a patient with complex mitochondrial deficiency, resulting in severely reduced coenzyme Q levels We demonstrate that the coenzyme Q analogue 2,4-dihydroxybensoic acid (2,4DHB) was able to specifically bypass the COQ7 deficiency, increase cellular coenzyme Q levels and rescue the biochemical defect in patient fibroblasts. CONCLUSION: We report the first patient with primary coenzyme Q deficiency due to a homozygous COQ7 mutation and a potentially beneficial treatment using 2,4DHB.

The hepatotoxin phomopsin A (PHO-A), a secondary metabolite mainly produced by the fungus Diaporthe toxica, occurs predominantly on sweet lupins. Along with the growing interest in sweet lupins for food and feed commodities, concerns have been raised about fungal infestations, and consequently, about the determination of PHO-A. High performance liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) represents the most suitable analytical technique for sensitive and selective detection of mycotoxins including PHO-A. However, isotopic labeled substances are needed as internal standards for a reliable and convenient quantification. As no isotope standard for PHO-A is currently available, a biosynthesis of fully (15)N6-labeled PHO-A was established by cultivation of D. toxica on defined media containing Na(15)NO3 and (15)N-labeled yeast extract as the only nitrogen sources. The identity of (15)N6-PHO-A was confirmed by high resolution mass spectrometry. The new (15)N6-labeled standard will facilitate the method development for PHO-A including a more accurate quantification by LC-MS/MS.

A rare lethal autosomal recessive syndrome with skeletal dysplasia, polycystic kidneys and multiple malformations was first described by Gillessen-Kaesbach et al and subsequently by Nishimura et al. The skeletal features uniformly comprise a round pelvis, mesomelic shortening of the upper limbs and defective ossification of the cervical spine. We studied two unrelated families including three affected fetuses with Gillessen-Kaesbach–Nishimura syndrome using whole-exome and Sanger sequencing, comparative genome hybridization and homozygosity mapping. All affected patients were shown to have a novel homozygous splice variant NM_024740.2: c.1173+2T>A in the ALG9 gene, encoding alpha-1,2-mannosyltransferase, involved in the formation of the lipid-linked oligosaccharide precursor of N-glycosylation. RNA analysis demonstrated skipping of exon 10, leading to shorter RNA. Mass spectrometric analysis showed an increase in monoglycosylated transferrin as compared with control tissues, confirming that this is a congenital disorder of glycosylation (CDG). Only three liveborn children with ALG9-CDG have been previously reported, all with missense variants. All three suffered from intellectual disability, muscular hypotonia, microcephaly and renal cysts, but none had skeletal dysplasia. Our study shows that some pathogenic variants in ALG9 can present as a lethal skeletal dysplasia with visceral malformations as the most severe phenotype. The skeletal features overlap with that previously reported for ALG3- and ALG12-CDG, suggesting that this subset of glycosylation disorders constitutes a new diagnostic group of skeletal dysplasias.

Segregation distorters located on sex chromosomes are predicted to sweep to fixation and cause extinction via a shortage of one sex, but in nature they are often found at low, stable frequencies. One potential resolution to this longstanding puzzle involves female multiple mating (polyandry). Because many meiotic drivers severely reduce the sperm competitive ability of their male carriers, females are predicted to evolve more frequent polyandry and thereby promote sperm competition when a meiotic driver invades. Consequently, the driving chromosome's relative fitness should decline, halting or reversing its spread. We used formal modeling to show that this initially appealing hypothesis cannot resolve the puzzle alone: other selective pressures (e.g., low fitness of drive homozygotes) are required to establish a stable meiotic drive polymorphism. However, polyandry and meiotic drive can strongly affect one another's frequency, and polyandrous populations may be resistant to the invasion of rare drive mutants.

The time is ripe for analyzing where the misgivings come from that researchers have after attending “personality” sessions at behavior conferences or reading papers in behavior and ecology journals that seem to attract excitement, because of the label “personality.” Beekman and Jordan (2017) point out that “animal personality” studies is nothing more than a rebranding of existing fields of research, fields that are far more solidly grounded and hypothesis driven than the often vague and superficial focus on animal personalities. They go on to state that there has been a rapid increase of mainly descriptive papers pointing to correlations and measuring behavioral repeatability with little attempt to link observed behaviors to evolutionary theory. “This latter approach was met with bemusement among many in the field of behavioral ecology” including the present authors. The field may have profited from the “Bystander’s Dilemma” (Darley and Latané 1968), meaning that if someone needs to do something, here criticize, the greater the number of bystanders, the less likely it is that any one of them will criticize. However, somewhat surprisingly, as we took a similar initiative (Jungwirth et al., unpublished data), there are now 2 independent critical reviews of animal personality research written by outsiders to the field. Importantly, Beekman and Jordan criticize points very similar to the ones we raise, despite all of us having different scientific backgrounds. We may diverge in the emphases we place on various points but overall we agree that it is difficult to see how and where the animal personality approach has advanced our understanding of behavior, ecology, and evolution.

Background Mitochondrial diseases due to defective respiratory chain complex III (CIII) are relatively uncommon. The assembly of the eleven-subunit CIII is completed by the insertion of the Rieske iron-sulfur protein, a process for which BCS1L protein is indispensable. Mutations in the BCS1L gene constitute the most common diagnosed cause of CIII deficiency, and the phenotypic spectrum arising from mutations in this gene is wide. Results A case of CIII deficiency was investigated in depth to assess respiratory chain function and assembly, and brain, skeletal muscle and liver histology. Exome sequencing was performed to search for the causative mutation(s). The patient’s platelets and muscle mitochondria showed respiration defects and defective assembly of CIII was detected in fibroblast mitochondria. The patient was compound heterozygous for two novel mutations in BCS1L, c.306A > T and c.399delA. In the cerebral cortex a specific pattern of astrogliosis and widespread loss of microglia was observed. Further analysis showed loss of Kupffer cells in the liver. These changes were not found in infants suffering from GRACILE syndrome, the most severe BCS1L-related disorder causing early postnatal mortality, but were partially corroborated in a knock-in mouse model of BCS1L deficiency. Conclusions We describe two novel compound heterozygous mutations in BCS1L causing CIII deficiency. The pathogenicity of one of the mutations was unexpected and points to the importance of combining next generation sequencing with a biochemical approach when investigating these patients. We further show novel manifestations in brain, skeletal muscle and liver, including abnormality in specialized resident macrophages (microglia and Kupffer cells). These novel phenotypes forward our understanding of CIII deficiencies caused by BCS1L mutations. Electronic supplementary material The online version of this article (doi:10.1186/s13023-017-0624-2) contains supplementary material, which is available to authorized users.

Parental care is common in vertebrates, but less common in invertebrates, tending to occur in species that have relatively few young that can be protected from the physical or biotic environment. Individuals will be selected to trade off current and future reproductive success, leading to adaptive changes in brood care behaviour in response to predation risk. Investment in brood care may also be reduced as a result of costs of parasitic infection. Here we investigated the impact of predator cues and of parasite infection on brood care behaviour in amphipod crustaceans. We found no evidence for any reduction in brooding behaviour or reproductive success in response to infection by vertically transmitted microsporidian parasites. This may reflect the mechanism of parasite transmission, as vertically transmitted parasites rely on host reproduction to pass to the next generation. However, we found evidence for changes in brood care behaviour in response to increased risk of predation. Larger Crangonyx pseudogracilis females, with higher oxygen demands for their broods, showed a reduction in active brood care when exposed to predator cues. Additionally, Gammarus duebeni were found to reduce brooding duration in response to predator cues; we suggest that this earlier release of juveniles is likely to enhance survival of the brood, by spreading the risk of predation on juveniles.

Teaching English to Young learners (TEYL) initiatives can represent an example of complex change (Fullan, 1992), whose classroom implementation implies those affected by the change learning new, or adjusting existing, practices and beliefs. Research (Levin and Fullan, 2008; Wedell, 2013) suggests that if contextually appropriate versions of complex change outcomes are to become visible in the majority of classrooms, both the parts of the change system (e.g. materials and assessment), and the way in which the people affected (change ‘partners’) carry out their roles, need to be as consistent as possible with change aims. Saudi Arabia introduced the teaching of English in the last year of primary level in 2004, and expanded it to the fourth year of schooling in 2011. This study uses the reported experiences of representatives from one key group of change ‘partners’: Saudi Primary English Supervisors, to explore aspects of the first six years of TEYL implementation. Their reports suggest widespread inconsistency during the first phase of implementation and little attempt to address the issues before launching the second phase. We suggest that acknowledging the value of these implementers' experiences and giving greater consideration to their suggestions, could help enable the second phase of implementation to become more consistent than the first.