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Adobe PDF - Supplementary_Data.pdf from Homozygous Deletion Mapping in Myeloma Samples Identifies Genes and an Expression Signature Relevant to Pathogenesis and Outcome

Purpose: Myeloma is a clonal malignancy of plasma cells. Poor-prognosis risk is currently identified by clinical and cytogenetic features. However, these indicators do not capture all prognostic information. Gene expression analysis can be used to identify poor-prognosis patients and this can be improved by combination with information about DNA-level changes.Experimental Design: Using single nucleotide polymorphism–based gene mapping in combination with global gene expression analysis, we have identified homozygous deletions in genes and networks that are relevant to myeloma pathogenesis and outcome.Results: We identified 170 genes with homozygous deletions and corresponding loss of expression. Deletion within the “cell death” network was overrepresented and cases with these deletions had impaired overall survival. From further analysis of these events, we have generated an expression-based signature associated with shorter survival in 258 patients and confirmed this signature in data from two independent groups totaling 800 patients. We defined a gene expression signature of 97 cell death genes that reflects prognosis and confirmed this in two independent data sets.Conclusions: We developed a simple 6-gene expression signature from the 97-gene signature that can be used to identify poor-prognosis myeloma in the clinical environment. This signature could form the basis of future trials aimed at improving the outcome of poor-prognosis myeloma. Clin Cancer Res; 16(6); 1856–64

Virtual surgical planning and three-dimensional printing have been invaluable tools in craniomaxillofacial surgery. From planning head and neck reconstruction to orthognathic surgery and secondary reconstruction of maxillofacial trauma, virtual surgical planning and three-dimensional printing allow the surgeon to rehearse the surgical plan and use patient-specific surgical guides for carrying out the plan accurately. However, the process of virtual surgical planning and three-dimensional printing requires time and coordination between the surgeon on one hand and the biomedical engineers and designers on the other hand. Outsourcing to third-party companies contributes to inefficiencies in this process. Advances in surgical planning software and three-dimensional printing technology have enabled the integration of virtual surgical planning and three-dimensional printing at the treating hospital, the point of care. This allows for expedited use of this process in semiurgent surgical cases and acute facial trauma cases by bringing the surgeon, radiologist, biomedical engineers, and designers to the point of care. In this article, the authors present the utility of EPPOCRATIS, expedited preoperative point of care reduction of fractures to normalized anatomy and three-dimensional printing to improve surgical outcomes, in the management of acute facial trauma.

Background 3D printing—also known as additive manufacturing—has a wide range of applications. Reproduction of low-cost, high-fidelity, disease- or patient-specific models presents a key developmental area in simulation and education research for complex cranial surgery. Methods Using cadaveric dissections as source materials, skull base models were created, printed, and tested for educational value in teaching complex cranial approaches. In this pilot study, assessments were made on the value of 3D printed models demonstrating the retrosigmoid and posterior petrosectomy approaches. Models were assessed and tested in a small cohort of neurosurgery resident subjects (n = 3) using a series of 10 radiographic and 2 printed case examples, with efficacy determined via agreement survey and approach selection accuracy. Results All subjects indicated agreement or strong agreement for all study endpoints that 3D printed models provided significant improvements in understanding of neuroanatomic relationships and principles of approach selection, as compared to 2D dissections or patient cross-sectional imaging alone. Models were not superior to in-person hands-on teaching. Mean approach selection accuracy was 90% (±13%) for 10 imaging-based cases, or 92% (±7%) overall. Trainees strongly agreed that approach decision-making was enhanced by adjunctive use of 3D models for both radiographic and printed cases. Conclusion 3D printed models incorporating skull base approaches and/or pathologies provide a compelling addition to the complex cranial education armamentarium. Based on our preliminary analysis, 3D printed models offer substantial potential for pedagogical value as dissection guides, adjuncts to preoperative study and case preparation, or tools for approach selection training and evaluation.

Subspecies of the protozoan parasite Trypanosoma brucei are the causative agents of Human African Trypanosomiasis (HAT), a debilitating neglected tropical disease prevalent across sub-Saharan Africa. HAT case numbers have steadily decreased since the start of the century, and sustainable elimination of one form of the disease is in sight. However, key to this is the development of novel drugs to combat the disease. Acoziborole is a recently developed benzoxaborole, currently in advanced clinical trials, for treatment of stage 1 and stage 2 HAT. Importantly, acoziborole is orally bioavailable, and curative with one dose. Recent studies have made significant progress in determining the molecular mode of action of acoziborole. However, less is known about the potential mechanisms leading to acoziborole resistance in trypanosomes. In this study, an in vitro-derived acoziborole-resistant cell line was generated and characterised. The AcoR line exhibited significant cross-resistance with the methyltransferase inhibitor sinefungin as well as hypersensitisation to known trypanocides. Interestingly, transcriptomics analysis of AcoR cells indicated the parasites had obtained a procyclic- or stumpy-like transcriptome profile, with upregulation of procyclin surface proteins as well as differential regulation of key metabolic genes known to be expressed in a life cycle-specific manner, even in the absence of major morphological changes. However, no changes were observed in transcripts encoding CPSF3, the recently identified protein target of acoziborole. The results suggest that generation of resistance to this novel compound in vitro can be accompanied by transcriptomic switches resembling a procyclic- or stumpy-type phenotype., Author summary Human African Trypanosomiasis (HAT), also known as Sleeping Sickness, is a parasitic disease prevalent across sub-Saharan Africa, where it is transmitted by blood-sucking Tsetse flies. The disease is caused by a single-celled protozoan parasite called Trypanosoma brucei. HAT case numbers have decreased over the past two decades, and elimination is in sight. However, to meet the WHO objective of sustainable HAT elimination, newer and safer drugs are needed. Acoziborole is a new drug that has proven safe and effective against HAT and, unlike previous drugs, it can be orally administered. Whilst the mode of action of acoziborole was recently identified, less is known regarding the mechanisms by which T. brucei could become resistant to acoziborole. To address this, we generated a drug resistant cell line under laboratory conditions, in order to analyse the differences between resistant and sensitive cells. By sequencing the transcriptome, we observed that many genes associated with mammalian-infective parasites are down-regulated, whilst genes associated with the insect stage are up-regulated. As a result of these differences, the metabolic effects of acoziborole on parasites are nullified. These data suggest that parasite differentiation, albeit on a gene expression level, is potentially a mechanism of drug resistance in T. brucei, although it is doubtful whether this mechanism could occur in the field where mammalian immune effectors would destroy parasites differentiating this way.

Background To determine whether virtual surgical planning and three-dimensional printed cutting guides (3D/VSP) improved radiographic bone union compared to conventional methods (CM) in fibula free flap (FFF) reconstruction of the mandibles. Methods Retrospective study from the years 2000-2018 at a tertiary hospital. Osseous union was evaluated by a radiologist blinded to each patient's treatment. Results Two hundred sixty patients who underwent FFF tissue transfer, 28 with VSP and 3D cutting guides. Bony union was not achieved in 46 (20%) patients who underwent CM compared to 1 (4%) of patients with VSP and guides (p = 0.036). FFF complication was significantly higher in CM with 87 patients (38%) compared to three patients (11%) in 3D/VSP (p = 0.005). Median time to bony union for patients who underwent CM was 1.4 years compared to 0.8 years in 3D/VSP. Conclusions 3D/VSP reduced the rate of radiographic nonunion and flap-related complications in FFF reconstruction for mandibular defects.

Background In recent years, three-dimensional (3D) printing has been increasingly applied to the intracranial vasculature for patient-specific surgical planning, training, education, and research. Unfortunately, though, much of the prior literature regarding 3D printing has focused on the end-product and not the process. In addition, for 3D printing/manufacturing to occur on a large scale, challenges and bottlenecks specific to each modeled anatomy must be overcome. Main body In this review article, limitations and considerations of each 3D printing processing step, as they relate to printing individual intracranial vasculature models and providing an active clinical service for a quaternary care center, are discussed. Relevant advantages and disadvantages of the available acquisition techniques (computed tomography, magnetic resonance, and digital subtraction angiography) are reviewed. Specific steps in segmentation, processing, and creation of a printable file may impede the workflow or degrade the fidelity of the printed model and are, therefore, given added attention. The various available printing techniques are compared with respect to printing the intracranial vasculature. Finally, applications are discussed, and a variety of example models are shown. Conclusion In this review we provide insight into the manufacturing of 3D models of the intracranial vasculature that may facilitate incorporation into or improve utility of 3D vascular models in clinical practice.

The Indian River Lagoon, located on the east coast of Florida, USA, is an Estuary of National Significance and an important economic and ecological resource. The Indian River Lagoon faces several environmental pressures, including freshwater discharges through the St. Lucie Estuary; accumulation of a anoxic, fine-grained, organic-rich sediment; and metal contamination from agriculture and marinas. Although the Indian River Lagoon has been well-studied, little is known about its microbial communities; thus, a two-year 16S amplicon sequencing study was conducted to assess the spatiotemporal changes of the sediment microbiome. In general, the Indian River Lagoon exhibited a microbiome that was consistent with other estuarine studies. Statistically different microbiomes were found between the Indian River Lagoon and St. Lucie Estuary due to changes in porewater salinity causing microbes that require salts for growth to be higher in the Indian River Lagoon. The St. Lucie Estuary exhibited more obvious microbial seasonality, such as higher Betaproteobacteriales, a freshwater associated organism, in wet season and higher Flavobacteriales in dry season samples. Distance-based linear models revealed these microbiomes were more affected by changes in total organic matter and copper than changes in temperature. Anaerobic organisms, such as Campylobacterales, were more associated with high total organic matter and copper samples while aerobic organisms, such as Nitrosopumilales, were more associated with low total organic matter and copper samples. This initial study fills the knowledge gap on the Indian River Lagoon microbiome and serves as an important baseline for possible future changes due to human impacts or environmental changes.

The Indian River Lagoon, located on the east coast of Florida, USA, is an Estuary of National Significance and an important economic and ecological resource. The Indian River Lagoon faces several environmental pressures, including freshwater discharges through the St. Lucie Estuary; accumulation of anoxic, fine-grained, organic-rich sediment; and metal contamination from agriculture and marinas. Although the Indian River Lagoon has been well-studied, little is known about its microbial communities; thus, a two-year 16S amplicon sequencing study was conducted to assess the spatiotemporal changes of the sediment bacterial and archaeal groups. In general, the Indian River Lagoon exhibited a prokaryotic community that was consistent with other estuarine studies. Statistically different communities were found between the Indian River Lagoon and St. Lucie Estuary due to changes in porewater salinity causing microbes that require salts for growth to be higher in the Indian River Lagoon. The St. Lucie Estuary exhibited more obvious prokaryotic seasonality, such as a higher relative abundance of Betaproteobacteriales in wet season and a higher relative abundance of Flavobacteriales in dry season samples. Distance-based linear models revealed these communities were more affected by changes in total organic matter and copper than changes in temperature. Anaerobic prokaryotes, such as Campylobacterales, were more associated with high total organic matter and copper samples while aerobic prokaryotes, such as Nitrosopumilales, were more associated with low total organic matter and copper samples. This initial study fills the knowledge gap on the Indian River Lagoon bacterial and archaeal communities and serves as important data for future studies to compare to determine possible future changes due to human impacts or environmental changes.

OBJECTIVE: To explore the healthcare experiences of parents whose baby died either before, during or shortly after birth between 20+0 and 23+6 weeks of gestation in order to identify practical ways to improve healthcare provision. DESIGN: Qualitative interview study. SETTING: England through two parent support organisations and four NHS Trusts. SAMPLE: A purposive sample of parents. METHODS: Thematic analysis of semi-structured in-depth narrative interviews. MAIN OUTCOME MEASURES: Parents' healthcare experiences. RESULTS: The key overarching theme to emerge from interviews with 38 parents was the importance of the terminology used to refer to the death of their baby. Parents who were told they were 'losing a baby' rather than 'having a miscarriage' were more prepared for the realities of labour, the birth experience and for making decisions around seeing and holding their baby. Appropriate terminology validated their loss, and impacted on parents' health and wellbeing immediately following bereavement and in the longer term. CONCLUSION: For parents experiencing the death of their baby at the margins between miscarriage, stillbirth and neonatal death, ensuring the use of appropriate terminology that reflects parents' preferences is vital. This helps to validate their loss and prepare them for the experiences of labour and birth. Reflecting parents' language preferences combined with compassionate bereavement care is likely to have a positive impact on parents' experiences and improve longer-term outcomes. TWEETABLE ABSTRACT: Describing baby loss shortly before 24 weeks of gestation as a 'miscarriage' does not prepare parents for labour and birth, seeing their baby and making memories.

Plasmodium species are apicomplexan parasites whose zoites are polarized cells with a marked apical organisation where the organelles associated with host cell invasion and colonization reside. Plasmodium gametes mate in the mosquito midgut to form the spherical and presumed apolar zygote that morphs during the following 24 hours into a polarized, elongated and motile zoite form, the ookinete. Endocytosis-mediated protein transport is generally necessary for the establishment and maintenance of polarity in epithelial cells and neurons, and the small GTPase RAB11A is an important regulator of protein transport via recycling endosomes. PbRAB11A is essential in blood stage asexual of Plasmodium. Therefore, a promoter swap strategy was employed to down-regulate PbRAB11A expression in gametocytes and zygotes of the rodent malaria parasite, Plasmodium berghei which demonstrated the essential role of RAB11A in ookinete development. The approach revealed that lack of PbRAB11A had no effect on gamete production and fertility rates however, the zygote to ookinete transition was almost totally inhibited and transmission through the mosquito was prevented. Lack of PbRAB11A did not prevent meiosis and mitosis, nor the establishment of polarity as indicated by the correct formation and positioning of the Inner Membrane Complex (IMC) and apical complex. However, morphological maturation was prevented and parasites remained spherical and immotile and furthermore, they were impaired in the secretion and distribution of microneme cargo. The data are consistent with the previously proposed model of RAB11A endosome mediated delivery of plasma membrane in Toxoplasma gondii if not its role in IMC formation and implicate it in microneme function., Author summary According to the WHO there was estimated to be over 200 million cases of malaria in 2017 and nearly half a million deaths. The disease is caused by specific species of Plasmodium which are passed between human hosts by a mosquito vector. In order to transmit through the mosquito the single-celled parasite undergoes many developmental changes as it morphs from non-motile blood forms to become a polarised and motile ookinete in the mosquito midgut. Transport of proteins within the cell during these critical morphological transitions relies on specific endosome vesicles to correctly target proteins within the parasite. We investigated the role of the RAB11A protein which is known to be involved in endosomal vesicle targeting to generate cellular polarity in other organisms. Because RAB11A is also essential for parasite growth in the mammalian host we used a promoter swap system to specifically switch off RAB11A in the sexual transmission stages. In the absence of RAB11A parasites were unable to form elongated, motile ookinetes and were unable to pass through the mosquito. Interestingly the parasites were able to form some of the (polarising) structures specific to ookinetes however full morphological transformation did not occur and the parasites were not motile. We show that although proteins are still delivered to the parasite surface, secretion is impaired and that the mutant parasites are smaller despite obvious microtubule formation implying that there is a deficit in delivery of membrane to the surface.

Background: Miltefosine has been used successfully to treat visceral leishmaniasis (VL) in India, but it was unsuccessful for VL in a clinical trial in Brazil. Methods: To identify molecular markers that predict VL treatment failure whole genome sequencing of 26 L. infantum isolates, from cured and relapsed patients allowed a GWAS analysis of SNPs, gene and chromosome copy number variations. Findings: A strong association was identified (p = 0·0005) between the presence of a genetically stable L. infantum Miltefosine Sensitivity Locus (MSL), and a positive response to miltefosine treatment. The risk of treatment failure increased 9·4-fold (95% CI 2·11–53·54) when an isolate did not have the MSL. The complete absence of the MSL predicted miltefosine failure with 0·92 (95% CI 0·65–0·996) sensitivity and 0·78 (95% CI 0·52–0·92) specificity. A genotyping survey of L. infantum (n = 157) showed that the frequency of MSL varies in a cline from 95% in North East Brazil to

International audience; The transition to motherhood is characterized by some of the most pronounced endocrine changes a women will experience in her lifetime. Unfortunately matrescence is also a time in a woman's life when she is most susceptible to mental illness such as perinatal depression. A growing body of research has aimed to determine how key endocrine systems, such as the hypothalamic-pituitary-adrenal (HPA) axis, are involved in the dysregulation of perinatal mental health. However, very little research has consistently linked perinatal changes in the HPA axis with maternal mental illness. Therefore the aims of this mini review are to 1) clearly summarize the normative changes in the HPA axis that occur during pregnancy and the postpartum period; 2) summarize what we know about the HPA axis in perinatal depression, and 3) propose key areas for future research. Understanding physiological biomarkers that can predict which women are at risk for perinatal mood disorders will lead to better tools for treating and ultimately preventing these debilitating disorders; improving the health of mother, child and family.

Composite textiles have found widespread use and advantages in various industries and applications. The constant demand for high-quality products and services requires companies to minimize their manufacturing costs and delivery time in order to compete with general and niche marketplaces. Creation of molding and tooling options for advanced composites encompasses a large portion of fabrication time, making it a costly process and a restraining factor. This research discusses a preliminary investigation into the use and control of soluble polymer compounds and additive manufacturing to fabricate sacrificial molds. These molds suffer from dimensional errors due to several factors, which have also been characterized. The basic soluble mold of a composite is 3D printed to meet the desired dimensions and geometry of holistic structures or spliced components. The time taken to dissolve the mold depends on the rate of agitation of the solvent. This process is steered towards enabling the implantation of optoelectronic devices within the composite to provide a sensing capability for structural health monitoring. The shape deviation of the 3D printed mold is also studied and compared to its original dimensions to optimize the dimensional quality to produce dimensionally accurate parts of up to 0.02% error.

International audience; As the world faces the health crisis of a global pandemic-with healthcare protocols in overhaul, and patients and care teams experiencing unprecedented levels of stress and unpredictability-we predict that current knowledge gaps in maternal health will inevitably have a lasting impact on the health of women giving birth now and in the near future. Since we are decades away from closing the knowledge gaps we need filled today, we recommend shifting thinking toward a comprehensive conceptual model that merges knowledge of stress physiology, neurobiology, and pregnancy physiology. The model we present here, the Maternal Reactive Scope Model, is an expansion of the Reactive Scope Model built upon the concept of Homeostasis and Allostasis. The model provides a framework to consider pathways and interactions across physiological systems to attribute a physiological basis for considering stress exposure and bridge research gaps on mechanisms to measure or target for treatment. Our intention is to provide an adaptable, heuristic framework for discussion of research considerations and new healthcare models that aim to provide the best care for new mothers during and after the COVID-19 pandemic.

Plasmodium species are apicomplexan parasites whose zoites are polarized cells with a marked apical organisation where the organelles associated with host cell invasion and colonization reside. Plasmodium gametes mate in the mosquito midgut to form the spherical and presumed apolar zygote that morphs during the following 24 hours into a polarized, elongated and motile zoite form, the ookinete. Endocytosis-mediated protein transport is generally necessary for the establishment and maintenance of polarity in epithelial cells and neurons, and the small GTPase RAB11A is an important regulator of protein transport via recycling endosomes. PbRAB11A is essential in blood stage asexual of Plasmodium. Therefore, a promoter swap strategy was employed to down-regulate PbRAB11A expression in gametocytes and zygotes of the rodent malaria parasite, Plasmodium berghei which demonstrated the essential role of RAB11A in ookinete development. The approach revealed that lack of PbRAB11A had no effect on gamete production and fertility rates however, the zygote to ookinete transition was almost totally inhibited and transmission through the mosquito was prevented. Lack of PbRAB11A did not prevent meiosis and mitosis, nor the establishment of polarity as indicated by the correct formation and positioning of the Inner Membrane Complex (IMC) and apical complex. However, morphological maturation was prevented and parasites remained spherical and immotile and furthermore, they were impaired in the secretion and distribution of microneme cargo. The data are consistent with the previously proposed model of RAB11A endosome mediated delivery of plasma membrane in Toxoplasma gondii if not its role in IMC formation and implicate it in microneme function.Author SummaryAccording to the WHO there was estimated to be over 200 million cases of malaria in 2017 and nearly half a million deaths. The disease is caused by specific species of Plasmodium which are passed between human hosts by a mosquito vector. In order to transmit through the mosquito the single-celled parasite undergoes many developmental changes as it morphs from non-motile blood forms to become a polarised and motile ookinete in the mosquito midgut. Transport of proteins within the cell during these critical morphological transitions relies on specific endosome vesicles to correctly target proteins within the parasite. We investigated the role of the RAB11A protein which is known to be involved in endosomal vesicle targeting to generate cellular polarity in other organisms. Because RAB11A is also essential for parasite growth in the mammalian host we used a promoter swap system to specifically switch off RAB11A in the sexual transmission stages. In the absence of RAB11A parasites were unable to form elongated, motile ookinetes and were unable to pass through the mosquito. Interestingly the parasites were able to form some of the (polarising) structures specific to ookinetes however full morphological transformation did not occur and the parasites were not motile. We show that although proteins are still delivered to the parasite surface, secretion is impaired and that the mutant parasites are smaller despite obvious microtubule formation implying that there is a deficit in delivery of membrane to the surface.

The application of machine learning to aerospace problems faces a particular challenge. For successful learning a large amount of good quality training data is required, typically tens of thousands of cases. However, due to the time and cost of experimental aerospace testing, this data is scarce. This paper shows that successful learning is possible with two novel techniques: The first technique is rapid testing. Over the last five years the Whittle Laboratory has developed a capability where rebuild and test times of a compressor stage now take 15 minutes instead of weeks. The second technique is to base machine learning on physical parameters, derived from engineering wisdom developed in industry over many decades. The method is applied to the important industry problem of predicting the effect of blade damage on compressor operability. The current approach has high uncertainty, it is based on human judgement and correlation of a handful of experimental test cases. It is shown using 100 training cases and 25 test cases that the new method is able to predict the operability of damaged compressor stages with an accuracy of 2% in a 95% confidence interval; far better than is possible by even the most experienced compressor designers. Use of the method is also shown to generate new physical understanding, previously unknown by any of the experts involved in this work. Using this method in the future offers an exciting opportunity to generate understanding of previously intractable problems in aerospace.

International audience

To evade mammalian immunity, Trypanosoma brucei switches the variant surface glycoprotein (VSG) expressed on its surface. Key to this reaction are controls exerted to ensure only one of many subtelomeric multigene VSG expression sites are transcribed at a time. DNA repair activities have to date been implicated only in catalysis of VSG switching by recombination, not transcriptional control. However, how VSG switching is signalled to guide the appropriate reaction, or to integrate switching into parasite growth, is unknown. Here we show that loss of ATR, a DNA damage signalling protein kinase, is lethal and causes increased nuclear genome lesions. ATR depletion also causes expression of mixed VSGs on the cell surface, increased transcription of genes from silent expression sites, and altered localisation of RNA Polymerase I and VEX1, factors involved in VSG transcription. The work therefore reveals that VSG expression control is mediated by a nuclear DNA damage signalling factor.

© 2017 ASME. The paper investigates the effect of non-equilibrium behaviour of boundary layers on the profile loss of a compressor. The investigation is undertaken using both direct numerical simulation (DNS) of a mid-height section of a compressor blade and a reduced order model, MISES. The solutions are validated using experimental measurements made in the embedded stage of a multistage low speed compressor. The paper shows that up to 35% of the suction surface boundary layer of the compressor blade exhibits non-equilibrium behaviour. The size of this region reduces as the Reynolds number is increased. The non-equilibrium behaviour was found to reduce profile loss in most cases, however, in a range of cases where transition occurs through a small separation the presence of non-equilibrium behaviour was found to increase profile loss.

Amphotericin B is an increasingly important tool in efforts to reduce the global disease burden posed by Leishmania parasites. With few other chemotherapeutic options available for the treatment of leishmaniasis, the potential for emergent resistance to this drug is a considerable threat. Here we characterised four novel amphotericin B-resistant Leishmania mexicana lines. All lines exhibited altered sterol biosynthesis, and hypersensitivity to pentamidine. Whole genome sequencing demonstrated resistance-associated mutation of the sterol biosynthesis gene sterol C5-desaturase in one line. However, in three out of four lines, RNA-seq revealed loss of expression of sterol C24-methyltransferase (SMT) responsible for drug resistance and altered sterol biosynthesis. Additional loss of the miltefosine transporter was associated with one of those lines. SMT is encoded by two tandem gene copies, which we found to have very different expression levels. In all cases, reduced overall expression was associated with loss of the 3’ untranslated region of the dominant gene copy, resulting from structural variations at this locus. Local regions of sequence homology, between the gene copies themselves, and also due to the presence of SIDER1 retrotransposon elements that promote multi-gene amplification, correlate to these structural variations. Moreover, in at least one case loss of SMT expression was not associated with loss of virulence in primary macrophages or in vivo. Whilst such repeat sequence-mediated instability is known in Leishmania genomes, its presence associated with resistance to a major antileishmanial drug, with no evidence of associated fitness costs, is a significant concern., Author summary Leishmania parasites are a significant threat, causing some 0.9–1.6 million new infections and 20,000 deaths annually. Amphotericin B is gaining importance in control of leishmaniasis, as other treatment options are increasingly compromised by emergent drug resistance. We sought to identify mechanisms by which resistance to amphotericin B could emerge through experimental selection and characterisation of four novel resistant L. mexicana lines. Whilst in all cases, sterol metabolism was dramatically affected, in three lines specific sterol changes resulted from decreased expression of the sterol biosynthesis enzyme sterol C24-methyltransferase (SMT). We found that this resulted from structural variation events at the genome level, resulting in specific loss of the more highly expressed SMT gene copy. These events appear to arise due to repetitive sequence elements in the genomic region, suggesting that SMT could be particularly prone to loss of expression in this manner. Moreover, in at least one line, we found that disruption of SMT function was not associated with any loss of fitness. These findings are significant in demonstrating that an apparently common mechanism of resistance-associated mutation does not compromise parasite virulence, thus representing a possible route to emergent resistance in the clinical setting.

Endocannabinoid (eCB) signaling plays an important role in the stress response pathways of the mammalian brain, yet its role in the avian stress response has not been described. Understanding eCB signaling in avian species (such as the European starling, Sturnus vulgaris) allows a model system that exhibits natural attenuation of hypothalamic-pituitary-adrenal (HPA) responsiveness to stressors. Specifically, seasonally breeding birds exhibit the highest HPA activity during the breeding season and subsequently exhibit a robust HPA down-regulation during molt. Because eCB signaling in mammals has an overall inhibitory effect on HPA activity, we expected shifts in eCB signaling to regulate the seasonal HPA down-regulation during molt. However, our data did not support a role for eCB signaling in the molt-related suppression of HPA activity. For example, injection of the cannabinoid receptor (CB1) antagonist, AM251, did not potentiate molt-suppressed HPA activity. Instead, our data suggest eCB regulation of HPA plasticity as birds transition from breeding to molt. In support of this hypothesis, birds in the late breeding season demonstrated a more dynamic response at the level of avian amygdala eCB content in response to acute stress. The response and directionality of this effect match that seen in mammals. Overall, our data suggest that eCB signaling may allow for a dynamic range in HPA responsiveness (eg, breeding), but the signaling pathway's role may be limited when the HPA response is restrained (eg, molt). This first characterization of eCB signaling in the avian stress response also emphasizes that although the system functions similarly to other species, its exact role may be species specific.

The paper investigates the effect of non-equilibrium behaviour of boundary layers on the profile loss of a compressor. The investigation is undertaken using both direct numerical simulation (DNS) of a mid-height section of a compressor blade and a reduced order model, MISES. The solutions are validated using experimental measurements made in the embedded stage of a multistage low speed compressor. The paper shows that up to 35% of the suction surface boundary layer of the compressor blade exhibits non-equilibrium behaviour. The size of this region reduces as the Reynolds number is increased. The non-equilibrium behaviour was found to reduce profile loss in most cases, however, in a range of cases where transition occurs through a small separation the presence of non-equilibrium behaviour was found to increase profile loss.

International audience; Zoonotic cutaneous leishmaniasis caused by Leishmania (L.) major parasites affects urban and suburban areas in the center and south of Tunisia where the disease is endemo-epidemic. Several cases were reported in human patients for which infection due to L major induced lesions with a broad range of severity. However, very little is known about the mechanisms underlying this diversity. Our hypothesis is that parasite genomic variability could, in addition to the host immunological background, contribute to the intra-species clinical variability observed in patients and explain the lesion size differences observed in the experimental model. Based on several epidemiological, in vivo and in vitro experiments, we focused on two clinical isolates showing contrasted severity in patients and BALB/c experimental mice model. We used DNA-seq as a high-throughput technology to facilitate the identification of genetic variants with discriminating potential between both isolates. Our results demonstrate that various levels of heterogeneity could be found between both L major isolates in terms of chromosome or gene copy number variation (CNV), and that the intra-species divergence could surprisingly be related to single nucleotide polymorphisms (SNPs) and Insertion/Deletion (InDels) events. Interestingly, we particularly focused here on genes affected by both types of variants and correlated them with the observed gene CNV. Whether these differences are sufficient to explain the severity in patients is obviously still open to debate, but we do believe that additional layers of -omic information is needed to complement the genomic screen in order to draw a more complete map of severity determinants.

Background: The tibial tubercle-trochlear groove (TT-TG) distance is a useful tool in guiding surgical management for patients with recurrent lateral patellar instability. Current recommendations for tibial tubercle transfer are based on TT-TG distance thresholds derived from adult populations. Recurrent patellar instability, however, frequently affects children, but normal and pathological TT-TG values have not been established for pediatric patients. The objectives of this study were to (1) confirm that magnetic resonance imaging (MRI) measurements for TT-TG distance in a pediatric population are reliable and reproducible, (2) determine whether the TT-TG distance changes with age, (3) define normal TT-TG distances in a pediatric population, and (4) confirm that a subgroup of pediatric patients with patellar instability have higher TT-TG distances. Methods: Six hundred and eighteen MRIs were retrospectively collected for patients who were nine months to sixteen years old. Each MRI was measured twice in a blinded, randomized manner by each reviewer. Patient age, sex, knee laterality, magnet strength, underlying diagnosis, and pertinent previous surgical treatments were all recorded separately from the measurements. MRIs that were unreadable and those of patients who had previous extensor mechanism surgery, preexisting deformity, or destructive neoplasms were excluded. Results: There was excellent intraobserver and interobserver reliability of TT-TG distance measurements. TT-TG distance was associated with the natural logarithm of age (p < 0.001). A percentile-based growth chart was created to demonstrate this relationship. The median TT-TG distance for patients without patellar instability in this pediatric population was 8.5 mm (mean and 95% confidence interval, 8.6 ± 0.3 mm). Patients with patellar instability had higher TT-TG distances (median, 12.1 mm; p < 0.001). TT-TG distance measured nearly 2 mm less on MRIs performed with a 3-T magnet than on those acquired with a 1.5-T magnet (p < 0.001). Conclusions: TT-TG distance changes with chronologic age in the pediatric population. As such, we developed a percentile-based growth chart in order to better depict normal TT-TG distances in the pediatric population. Like many issues in pediatric orthopaedics, an age-based approach for directing surgical treatment may be more appropriate for skeletally immature individuals with recurrent lateral patellar instability. Level of Evidence: Diagnostic Level III. See Instructions for Authors for a complete description of levels of evidence.

Oestrogens activate nucleus- and membrane-initiated signalling. Nucleus-initiated events control a wide array of physiological and behavioural responses. These effects generally take place within relatively long periods of time (several hours to days). By contrast, membrane-initiated signalling affects a multitude of cellular functions in a much shorter timeframe (seconds to minutes). However, much less is known about their functional significance. Furthermore, the origin of the oestrogens able to trigger these acute effects is rarely examined. Finally, these two distinct types of oestrogenic actions have often been studied independently such that we do not exactly know how they cooperate to control the same response. The present review presents a synthesis of recent work carried out in our laboratory that aimed to address these issues in the context of the study of male sexual behaviour in Japanese quail, which is a considered as a suitable species for tackling these issues. The first section presents data indicating that 17β-oestradiol, or its membrane impermeable analogues, acutely enhances measures of male sexual motivation but does not affect copulatory behaviour. These effects depend on the activation of membrane-initiated events and local oestrogen production. The second part of this review discusses the regulation of brain oestrogen synthesis through post-translational modifications of the enzyme aromatase. Initially discovered in vitro, these rapid and reversible enzymatic modulations occur in vivo following variations in the social and environment context and therefore provide a mechanism of acute regulation of local oestrogen provision with a spatial and time resolution compatible with the rapid effects observed on male sexual behaviour. Finally, we discuss how these distinct modes of oestrogenic action (membrane- versus nucleus-initiated) acting in different time frames (short- versus long-term) interact to control different components (motivation versus performance) of the same behavioural response and improve reproductive fitness.

Given the connection between chronic stress and health, there has been a growing emphasis on identifying chronically stressed wild animals, especially in relation to anthropogenic disturbances. There is considerable confusion, however, in how to identify chronically stressed wild animals, but the most common assumption is that measures of glucocorticoid (GC) function will increase. In an attempt to determine an “endocrine profile” of a chronically stressed wild animal, this review collected papers from the literature that measured baseline GC, stress-induced GC, measures of integrated GC, negative feedback, hypothalamic–pituitary–adrenal axis sensitivity, and/or body weight in chronically stressed animals. The collected studies encompassed laboratory and field studies, numerous diverse species, and multiple techniques for inducing chronic stress. Each paper was ranked according to its relevance to wild animals and scored as to whether the measured response increased, decreased, or stayed the same after exposure to chronic stress. The analyses uncovered so much variation between studies that the literature does not support a generalized endocrine profile in how wild animals respond to chronic stress. The common predictions appear to be based almost entirely on theoretical models rather than empirical data. The three most important variables affecting GC responses were the stressors used to induce chronic stress, the potential for those stressors to induce habituation, and the taxon of the focal species. The best approach for identifying a chronically stressed population appears to be documentation of changes at multiple levels of GC regulation, but the direction of the change (increase or decrease) may be relatively unimportant compared to the fact that the response changes at all. The conclusion is that a consistent, predictable, endocrine response to chronic stress, regardless of the protocol used to induce chronic stress and the species under study, does not exist.

This Letter reports a measurement of the CP violation observables SJ/ψK0S and CJ/ψK0S in the decay channel B0→J/ψK0S performed with 1.0 fb−1 of pp collisions at s√=7 TeV collected by the LHCb experiment. The fit to the data yields SJ/ψK0S=0.73±0.07(stat)±0.04(syst) and CJ/ψK0S=0.03±0.09(stat)±0.01(syst). Both values are consistent with the current world averages and within expectations from the Standard Model.

In the male brain, the medial preoptic nucleus (POM) is known to be a critical relay for the activation of sexual behaviour, with the aromatisation of testosterone into 17β-oestradiol (E2 ) playing a key role. Acute stress has been shown to differentially modulate the aromatase enzyme in this and other brain nuclei in a sex-specific manner. In POM specifically, stress induces increases in aromatase activity (AA) that are both rapid and reversible. How the physiological processes initiated during an acute stress response mediate sex- and nuclei- specific changes in AA and which stress response hormones are involved remains to be determined. By examining the relative effects of corticosterone (CORT), arginine vasotocin (AVT, the avian homologue to arginine vasopressin) and corticotrophin-releasing factor (CRF), the present study aimed to define the hormone profile regulating stress-induced increases in AA in the POM. We found that CORT, AVT and CRF all appear to play some role in these changes in the male brain. In addition, these effects occur in a targeted manner, such that modulation of the enzyme by these hormones only occurs in the POM rather than in all aromatase-expressing nuclei. Similarly, in the female brain, the experimental effects were restricted to the POM but only CRF was capable of inducing the stress-like increases in AA. These data further demonstrate the high degree of specificity (nuclei-, sex- and hormone-specific effects) in this system, highlighting the complexity of the stress-aromatase link and suggesting modes through which the nongenomic modulation of this enzyme can result in targeted, rapid changes in local oestrogen concentrations.