Whole genome sequencing of Plasmodium falciparum from dried blood spots using selective whole genome amplification

BACKGROUND: Translating genomic technologies into healthcare applications for the malaria parasite Plasmodium falciparum has been limited by the technical and logistical difficulties of obtaining high quality clinical samples from the field. Sampling by dried blood spot (DBS) finger-pricks can be performed safely and efficiently with minimal resource and storage requirements compared with venous blood (VB). Here, the use of selective whole genome amplification (sWGA) to sequence the P. falciparum genome from clinical DBS samples was evaluated, and the results compared with current methods that use leucodepleted VB. METHODS: Parasite DNA with high (>95%) human DNA contamination was selectively amplified by Phi29 polymerase using short oligonucleotide probes of 8-12 mers as primers. These primers were selected on the basis of their differential frequency of binding the desired (P. falciparum DNA) and contaminating (human) genomes. RESULTS: Using sWGA method, clinical samples from 156 malaria patients, including 120 paired samples for head-to-head comparison of DBS and leucodepleted VB were sequenced. Greater than 18-fold enrichment of P. falciparum DNA was achieved from DBS extracts. The parasitaemia threshold to achieve >5× coverage for 50% of the genome was 0.03% (40 parasites per 200 white blood cells). Over 99% SNP concordance between VB and DBS samples was achieved after excluding missing calls. CONCLUSION: The sWGA methods described here provide a reliable and scalable way of generating P. falciparum genome sequence data from DBS samples. The current data indicate that it will be possible to get good quality sequence on most if not all drug resistance loci from the majority of symptomatic malaria patients. This technique overcomes a major limiting factor in P. falciparum genome sequencing from field samples, and paves the way for large-scale epidemiological applications

Spatial Motion Doodles: Sketching Animation in VR Using Hand Gestures and Laban Motion Analysis

International audienceWe present a method for easily drafting expressive character animation by playing with instrumented rigid objects. We parse the input 6D trajectories (position and orientation over time)-called spatial motion doodles-into sequences of actions and convert them into detailed character animations using a dataset of parameterized motion clips which are automatically fitted to the doodles in terms of global trajectory and timing. Moreover, we capture the expres-siveness of user-manipulation by analyzing Laban effort qualities in the input spatial motion doodles and transferring them to the synthetic motions we generate. We validate the ease of use of our system and the expressiveness of the resulting animations through a series of user studies, showing the interest of our approach for interactive digital storytelling applications dedicated to children and non-expert users, as well as for providing fast drafting tools for animators

Activation of conventional protein kinase C (PKC) is critical in the generation of human neutrophil extracellular traps

BACKGROUND: Activation of NADPH oxidase is required for neutrophil extracellular trap (NET) formation. Protein kinase C (PKC) is an upstream mediator of NADPH oxidase activation and thus likely to have a role in NET formation. METHODS: Pharmacological inhibitors were used to block PKC activity in neutrophils harvested from healthy donor blood. RESULTS: Pan PKC inhibition with Ro-31-8220 (p<0.001), conventional PKC inhibition with Go 6976 (p<0.001) and specific PKCβ inhibition with LY333531 (p<0.01) blocked NET formation in response to PMA. Inhibition of novel and atypical PKC had no effect. LY333531 blocked NET induction by the diacylglycerol analogue OAG (conventional PKC activator) (p<0.001). CONCLUSIONS: Conventional PKCs have a prominent role in NET formation. Furthermore PKCβ is the major isoform implicated in NET formation

A plant natriuretic peptide-like molecule of the pathogen Xanthomonas axonopodis pv. citri causes rapid changes in the proteome of its citrus host

<p>Abstract</p> <p>Background</p> <p>Plant natriuretic peptides (PNPs) belong to a novel class of peptidic signaling molecules that share some structural similarity to the N-terminal domain of expansins and affect physiological processes such as water and ion homeostasis at nano-molar concentrations. The citrus pathogen Xanthomonas axonopodis pv. citri possesses a PNP-like peptide (XacPNP) uniquely present in this bacteria. Previously we observed that the expression of <it>XacPNP </it>is induced upon infection and that lesions produced in leaves infected with a XacPNP deletion mutant were more necrotic and lead to earlier bacterial cell death, suggesting that the plant-like bacterial PNP enables the plant pathogen to modify host responses in order to create conditions favorable to its own survival.</p> <p>Results</p> <p>Here we measured chlorophyll fluorescence parameters and water potential of citrus leaves infiltrated with recombinant purified XacPNP and demonstrate that the peptide improves the physiological conditions of the tissue. Importantly, the proteomic analysis revealed that these responses are mirrored by rapid changes in the host proteome that include the up-regulation of Rubisco activase, ATP synthase CF1 α subunit, maturase K, and α- and β-tubulin.</p> <p>Conclusions</p> <p>We demonstrate that XacPNP induces changes in host photosynthesis at the level of protein expression and in photosynthetic efficiency in particular. Our findings suggest that the biotrophic pathogen can use the plant-like hormone to modulate the host cellular environment and in particular host metabolism and that such modulations weaken host defence.</p

Inhibition of cyclin-dependent kinase 9 downregulates cytokine production without detrimentally affecting human monocyte-derived macrophage viability

Cyclin-dependent kinase (CDK) inhibitor drugs (CDKi), such as R-roscovitine and AT7519, induce neutrophil apoptosis in vitro and enhance the resolution of inflammation in a number of in vivo models. This class of compounds are potential novel therapeutic agents that could promote the resolution of acute and chronic inflammatory conditions where neutrophil activation contributes to tissue damage and aberrant tissue repair. In this study we investigated CDKi effects on macrophage pro-inflammatory mediator production and viability. Treatment of human monocyte-derived macrophages (MDMs) with the CDKi AT7519 and R-roscovitine at concentrations that induce neutrophil apoptosis had no significant effect on control or LPS-activated MDM apoptosis and viability, and did not detrimentally affect MDM efferocytosis of apoptotic cells. In addition, enhanced efferocytosis, induced by the glucocorticoid dexamethasone, was also unaffected after a short time treatment with R-roscovitine. Macrophage cytokine responses to inflammatory stimuli are also of importance during inflammation and resolution. As a key target of CDKi, CDK9, is involved in protein transcription via the RNA polymerase II complex, we investigated the effect of CDKi drugs on cytokine production. Our data show that treatment with AT7519 significantly downregulated expression and release of key MDM cytokines IL-6, TNF, IL-10 and IL-1β, as well as markers of pro-inflammatory macrophage polarisation. R-Roscovitine was also able to downregulate inflammatory cytokine protein secretion from MDMs. Using siRNA transfection, we demonstrate that genetic knock-down of CDK9 replicates these findings, reducing expression and release of pro-inflammatory cytokines. Furthermore, overexpression of CDK9 in THP-1 cells can promote a pro-inflammatory phenotype in these cells, suggesting that CDK9 plays an important role in the inflammatory phenotype of macrophages. Overall, this study demonstrates that pharmacological and genetic targeting of CDK9 inhibits an inflammatory phenotype in human MDMs. As such these data indicate that CDK9 may be key to therapeutically targeting pro-inflammatory macrophage functions during chronic inflammation

Geographic mosaics and changing rates of cereal domestication

Domestication is the process by which plants or animals evolved to fit a human-managed environment, and it is marked by innovations in plant morphology and anatomy that are in turn correlated with new human behaviours and technologies for harvesting, storage and field preparation. Archaeobotanical evidence has revealed that domestication was a protracted process taking thousands of plant generations. Within this protracted process there were changes in the selection pressures for domestication traits as well as variation across a geographic mosaic of wild and cultivated populations. Quantitative data allow us to estimate the changing selection coefficients for the evolution of non-shattering (domestic-type seed dispersal) in Asian rice (Oryza sativa L.), barley (Hordeum vulgare L.), emmer wheat (Triticum dicoccon (Shrank) Schübl.) and einkorn wheat (Triticum monococcum L.). These data indicate that selection coefficients tended to be low, but also that there were inflection points at which selection increased considerably. For rice, selection coefficients of the order of 0.001 prior to 5500 BC shifted to greater than 0.003 between 5000 and 4500 BC, before falling again as the domestication process ended 4000–3500 BC. In barley and the two wheats selection was strongest between 8500 and 7500 BC. The slow start of domestication may indicate that initial selection began in the Pleistocene glacial era

Developmental and computational perspectives on infant social cognition

Adults effortlessly and automatically infer complex pat- terns of goals, beliefs, and other mental states as the causes of others’ actions. Yet before the last decade little was known about the developmental origins of these abilities in early infancy. Our understanding of infant social cognition has now improved dramatically: even preverbal infants appear to perceive goals, preferences (Kushnir, Xu, & Wellman, in press), and even beliefs from sparse observations of inten- tional agents’ behavior. Furthermore, they use these infer- ences to predict others’ behavior in novel contexts and to make social evaluations (Hamlin, Wynn, & Bloom, 2007). Keywords: Social cognition; Cognitive Development; Computational Modeling; Theory of Min

In Situ neutron diffraction study of effect of hydrogen on deformation mechanisms in austenitic and duplex steels

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