Fast Gaussian Pairwise Constrained Spectral Clustering

International audienceWe consider the problem of spectral clustering with partial supervision in the form of must-link and cannot-link constraints. Such pairwise constraints are common in problems like coreference resolution in natural language processing. The approach developed in this paper is to learn a new representation space for the data together with a dis-tance in this new space. The representation space is obtained through a constraint-driven linear transformation of a spectral embedding of the data. Constraints are expressed with a Gaussian function that locally reweights the similarities in the projected space. A global, non-convex optimization objective is then derived and the model is learned via gradi-ent descent techniques. Our algorithm is evaluated on standard datasets and compared with state of the art algorithms, like [14,18,31]. Results on these datasets, as well on the CoNLL-2012 coreference resolution shared task dataset, show that our algorithm significantly outperforms related approaches and is also much more scalable

Process efficiency in polymer extrusion: Correlation between the energy demand and melt thermal stability

YesThermal stability is of major importance in polymer extrusion, where product quality is dependent upon the level of melt homogeneity achieved by the extruder screw. Extrusion is an energy intensive process and optimisation of process energy usage while maintaining melt stability is necessary in order to produce good quality product at low unit cost. Optimisation of process energy usage is timely as world energy prices have increased rapidly over the last few years. In the first part of this study, a general discussion was made on the efficiency of an extruder. Then, an attempt was made to explore correlations between melt thermal stability and energy demand in polymer extrusion under different process settings and screw geometries. A commodity grade of polystyrene was extruded using a highly instrumented single screw extruder, equipped with energy consumption and melt temperature field measurement. Moreover, the melt viscosity of the experimental material was observed by using an off-line rheometer. Results showed that specific energy demand of the extruder (i.e. energy for processing of unit mass of polymer) decreased with increasing throughput whilst fluctuation in energy demand also reduced. However, the relationship between melt temperature and extruder throughput was found to be complex, with temperature varying with radial position across the melt flow. Moreover, the melt thermal stability deteriorated as throughput was increased, meaning that a greater efficiency was achieved at the detriment of melt consistency. Extruder screw design also had a significant effect on the relationship between energy consumption and melt consistency. Overall, the relationship between the process energy demand and thermal stability seemed to be negatively correlated and also it was shown to be highly complex in nature. Moreover, the level of process understanding achieved here can help to inform selection of equipment and setting of operating conditions to optimise both energy and thermal efficiencies in parallel.This work was funded through an inter-disciplinary research programme (Grant No. EP/G059330/1) by the EPSRC-UK. The technical assistance provided by Ken Howell, Roy Dixon and John Wyborn is greatly appreciated

Engineered Single-Domain Antibodies with High Protease Resistance and Thermal Stability

The extreme pH and protease-rich environment of the upper gastrointestinal tract is a major obstacle facing orally-administered protein therapeutics, including antibodies. Through protein engineering, several Clostridium difficile toxin A-specific heavy chain antibody variable domains (VHHs) were expressed with an additional disulfide bond by introducing Ala/Gly54Cys and Ile78Cys mutations. Mutant antibodies were compared to their wild-type counterparts with respect to expression yield, non-aggregation status, affinity for toxin A, circular dichroism (CD) structural signatures, thermal stability, protease resistance, and toxin A-neutralizing capacity. The mutant VHHs were found to be well expressed, although with lower yields compared to wild-type counterparts, were non-aggregating monomers, retained low nM affinity for toxin A, albeit the majority showed somewhat reduced affinity compared to wild-type counterparts, and were capable of in vitro toxin A neutralization in cell-based assays. Far-UV and near-UV CD spectroscopy consistently showed shifts in peak intensity and selective peak minima for wild-type and mutant VHH pairs; however, the overall CD profile remained very similar. A significant increase in the thermal unfolding midpoint temperature was observed for all mutants at both neutral and acidic pH. Digestion of the VHHs with the major gastrointestinal proteases, at biologically relevant concentrations, revealed a significant increase in pepsin resistance for all mutants and an increase in chymotrypsin resistance for the majority of mutants. Mutant VHH trypsin resistance was similar to that of wild-type VHHs, although the trypsin resistance of one VHH mutant was significantly reduced. Therefore, the introduction of a second disulfide bond in the hydrophobic core not only increases VHH thermal stability at neutral pH, as previously shown, but also represents a generic strategy to increase VHH stability at low pH and impart protease resistance, with only minor perturbations in target binding affinities. These are all desirable characteristics for the design of protein-based oral therapeutics

The development of a 16S rRNA gene based PCR for the identification of Streptococcus pneumoniae and comparison with four other species specific PCR assays

<p>Abstract</p> <p>Background</p> <p><it>Streptococcus pneumoniae </it>is one of the most frequently encountered pathogens in humans but its differentiation from closely related but less pathogenic streptococci remains a challenge.</p> <p>Methods</p> <p>This report describes a newly-developed PCR assay (Spne-PCR), amplifying a 217 bp product of the 16S rRNA gene of <it>S. pneumoniae</it>, and its performance compared to other genotypic and phenotypic tests.</p> <p>Results</p> <p>The new PCR assay designed in this study, proved to be specific at 57°C for <it>S. pneumoniae</it>, not amplifying <it>S. pseudopneumoniae </it>or any other streptococcal strain or any strains from other upper airway pathogenic species. PCR assays (psaA, LytA, ply, spn9802-PCR) were previously described for the specific amplification of <it>S. pneumoniae</it>, but <it>psaA</it>-PCR was the only one found not to cross-react with <it>S. pseudopneumoniae</it>.</p> <p>Conclusion</p> <p>Spne-PCR, developed for this study, and psaA-PCR were the only two assays which did not mis-identify <it>S. pseudopneumoniae </it>as <it>S. pneumoniae</it>. Four other PCR assays and the AccuProbe assay were unable to distinguish between these species.</p

Yeast : the soul of beer’s aroma—a review of flavour-active esters and higher alcohols produced by the brewing yeast

Among the most important factors influencing beer quality is the presence of well-adjusted amounts of higher alcohols and esters. Thus, a heavy body of literature focuses on these substances and on the parameters influencing their production by the brewing yeast. Additionally, the complex metabolic pathways involved in their synthesis require special attention. More than a century of data, mainly in genetic and proteomic fields, has built up enough information to describe in detail each step in the pathway for the synthesis of higher alcohols and their esters, but there is still place for more. Higher alcohols are formed either by anabolism or catabolism (Ehrlich pathway) of amino acids. Esters are formed by enzymatic condensation of organic acids and alcohols. The current paper reviews the up-to-date knowledge in the pathways involving the synthesis of higher alcohols and esters by brewing yeasts. Fermentation parameters affecting yeast response during biosynthesis of these aromatic substances are also fully reviewed.Eduardo Pires gratefully acknowledges the Fundacao para a Ciencia e a Tecnologia (FCT, Portugal) for the PhD fellowship support (SFRH/BD/61777/2009). The financial contributions of the EU FP7 project Ecoefficient Biodegradable Composite Advanced Packaging (EcoBioCAP, grant agreement no. 265669) as well as of the Grant Agency of the Czech Republic (project GACR P503/12/1424) are also gratefully acknowledged. The authors thank the Ministry of Education, Youth and Sports of the Czech Republic (MSM 6046137305) for their financial support

The context of chemical communication driving a mutualism

Recent work suggests that Drosophila and Saccharomyces yeasts may establish a mutualistic association, and that this is driven by chemical communication. While individual volatiles have been implicated in the attraction of D. melanogaster, the semiochemicals affecting the behavior of the sibling species D. simulans are less well characterised. Here, we comprehensively scrutinize a broad range of volatiles produced by attractive and repulsive yeasts to experimentally evaluate the chemical nature of communication between these species. When grown in liquid or on agar-solidified grape juice, attraction to S. cerevisiae was primarily driven by 3-methylbutyl acetate (isoamyl acetate) and repulsion by acetic acid, a known attractant to D. melanogaster (also known as vinegar fly). Using T-maze choice tests and synthetic compounds we show that these responses were strongly influenced by compound concentration. Moreover, the behavioral response is further impacted by the chemical context of the environment. Thus, chemical communication between yeasts and flies is complex, and is not simply driven by the presence of single volatiles, but modulated by compound interactions. The ecological context of chemical communication needs to be taken into consideration when testing for ecologically realistic responses

Predicting the response to CTLA-4 blockade by longitudinal noninvasive monitoring of CD8 T cells

Immunotherapy using checkpoint-blocking antibodies against targets such as CTLA-4 and PD-1 can cure melanoma and non-small cell lung cancer in a subset of patients. The presence of CD8 T cells in the tumor correlates with improved survival. We show that immuno-positron emission tomography (immuno-PET) can visualize tumors by detecting infiltrating lymphocytes and, through longitudinal observation of individual animals, distinguish responding tumors from those that do not respond to therapy. We used 89 Zr-labeled PEGylated single-domain antibody fragments (VHHs) specific for CD8 to track the presence of intratumoral CD8 + T cells in the immunotherapy-susceptible B16 melanoma model in response to checkpoint blockade. A 89 Zr-labeled PEGylated anti-CD8 VHH detected thymus and secondary lymphoid structures as well as intratumoral CD8 T cells. Animals that responded to CTLA-4 therapy showed a homogeneous distribution of the anti-CD8 PET signal throughout the tumor, whereas more heterogeneous infiltration of CD8 T cells correlated with faster tumor growth and worse responses. To support the validity of these observations, we used two different transplantable breast cancer models, yielding results that conformed with predictions based on the antimelanoma response. It may thus be possible to use immuno-PET and monitor antitumor immune responses as a prognostic tool to predict patient responses to checkpoint therapies.National Institutes of Health (U.S.) (Grant R01-AI087879-06)National Institutes of Health (U.S.) (Grant DP1-GM106409-03)National Institutes of Health (U.S.) (Grant R01-GM100518-04)National Institutes of Health (U.S.) (Grant P01 CA080111