Identification, expression, and DNA sequence of the GDP-mannose biosynthesis genes encoded by the O7 rfb gene cluster of strain VW187 (Escherichia coli O7:K1)

The O7-specific lipopolysaccharide (LPS) in strains of Escherichia coli consists of a repeating unit made of galactose, mannose, rhamnose, 4-acetamido-2,6-dideoxyglucose, and N-acetylglucosamine. We have recently cloned and characterized genetically the O7-specific LPS biosynthesis region (rfbEcO7) of the E. coli O7:K1 strain VW187 (C. L. Marolda, J. Welsh, L. Dafoe, and M. A. Valvano, J. Bacteriol. 172:3590-3599, 1990). In this study, we localized the gnd gene encoding gluconate-6-phosphate dehydrogenase at one end of the rfbEcO7 gene cluster and sequenced that end of the cluster. Three open reading frames (ORF) encoding polypeptides of 275, 464, and 453 amino acids were identified upstream of gndEcO7, all transcribed toward the gnd gene. ORF275 had 45% similarity at the protein level with ORF16.5, which occupies a similar position in the Salmonella enterica LT2 rfb region, and presumably encodes a nucleotide sugar transferase. The polypeptides encoded by ORFs 464 and 453 were expressed under the control of the ptac promoter and visualized in Coomassie blue-stained sodium dodecyl sulfate-polyacrylamide gels and by maxicell analysis. ORF464 expressed GDP-mannose pyrophosphorylase and ORF453 encoded a phosphomannomutase, the enzymes for the biosynthesis pathway of GDP-mannose, one of the nucleotide sugar precursors for the formation of the O7 repeating unit. They were designated rfbMEcO7 and rfbKEcO7, respectively. The RfbMEcO7 polypeptide was homologous to the corresponding protein in S. enterica LT2, XanB of Xanthomonas campestris, and AlgA of Pseudomonas aeruginosa, all GDP-mannose pyrophosphorylases. RfbKEcO7 was very similar to CpsG of S. enterica LT2, an enzyme presumably involved in the biosynthesis of the capsular polysaccharide colanic acid, but quite different from the corresponding RfbK protein of S. enterica LT2.

Automatic FPGA Synthesis of Memory Intensive C-based Kernels

Realising high performance image and signal processing
applications on modern FPGA presents a challenging implementation problem due to the large data frames streaming through these systems. Specifically, to meet the high bandwidth and data storage demands of these applications, complex hierarchical memory architectures must be manually specified
at the Register Transfer Level (RTL). Automated approaches which convert high-level operation descriptions, for instance in the form of C programs, to an FPGA architecture, are unable to automatically realise such architectures. This paper
presents a solution to this problem. It presents a compiler to automatically derive such memory architectures from a C program. By transforming the input C program to a unique dataflow modelling dialect, known as Valved Dataflow (VDF), a mapping and synthesis approach developed for this dialect can
be exploited to automatically create high performance image and video processing architectures. Memory intensive C kernels for Motion Estimation (CIF Frames at 30 fps), Matrix Multiplication (128x128 @ 500 iter/sec) and Sobel Edge Detection (720p @ 30 fps), which are unrealisable by current state-of-the-art C-based synthesis tools, are automatically derived from a C description of the algorithm.

Contingent Participation: Imaginaries of Sustainable Technoscientific Innovation in the EU

Citizen participation is often valorised in the governance of areas of high scientific uncertainty at national, international and supranational levels. This chapter considers citizen or public participation in the specific area of the EU’s agenda on sustainable development as it increasingly frames technoscientific innovation and development. Specifically, the chapter focuses on just one underexplored aspect of the conditions of possibility for participation: imaginaries. These include how the EU imagines its engagement, responsibilities and identity in relation to the specific area, including the knowledges that are constructed and used in decision-making, and by implication the role of citizen or public participation.

The discussion draws on an analysis of the social and technoscientific imaginaries found in legal, regulatory and policy discourses. These construct the frame of sustainable development and build a link between it and technoscientific innovation and development. By attention to imaginaries as one aspect of the frame, the chapter highlights the centrality of, and main interactions between, sustainable development in the inscription and potential disruption of the normative and programmatic background for the operationalisation of technoscientific innovation. These insights are used to highlight how imaginaries constitute a crucial aspect of the conditions of possibility for participation: determining who has to participate in decision-making through configurations of ‘citizen’ or ‘public’, how, why, and which outcomes are to be achieved by that participation.

The structures of four bombesins and their cloned precursor-encoding cDNAs from acid-solvated skin secretion of the European yellow-bellied toad, Bombina variegata

Four different bombesins (bombesin, His(6)-bombesin, Phe(13)-bombesin and Asp(2)-, Phe(4)-SAP-bombesin) have been identified by a systematic sequencing study of peptides in reverse phase HPLC fractions of the skin secretion of the European yellow-bellied toad, Bombina variegata, that had been solvated in 0.1% (v/v) aqueous trifluoroacetic acid (TFA) and stored frozen at -20°C for 12 years. By using a 3'- and 5'-RACE PCR strategy, the corresponding biosynthetic precursor-encoding cDNAs of all four peptides were cloned from a cDNA library made from the same long-term frozen, acid-solvated skin secretion sample following thawing and lyophilization. Canonical bombesin and His(6)-bombesin are classical bombesin sub-family members, whereas Phe(13)-bombesin and Asp(2)-, Phe(4)-SAP-bombesin, belong to the litorin/ranatensin sub-family of bombesin-like peptides (BLPs). Assignment of these peptides to respective sub-families, was based upon both their primary structural similarities and their comparative pharmacological activities. An interesting observation in this study, was that the nucleotide sequences of the open-reading frames of cloned cDNAs encoding bombesin and its His(6)-substituted analog, were identical except for a single base that was responsible for the change observed at the position 6 residue in the mature peptide from Asn to His. In contrast, the precursor cDNA nucleotide sequences encoding the Phe(13)-bombesins, exhibited 53 base differences. The pharmacological activities of synthetic replicates of each bombesin were compared using two different mammalian smooth muscle preparations and all four peptides were found to be active. However, there were significant differences in their relative potencies.

Genomic Analysis of Pseudomonas putida Phage tf with Localized Single-Strand DNA Interruptions

The complete sequence of the 46,267 bp genome of the lytic bacteriophage tf specific to Pseudomonas putida PpG1 has been determined. The phage genome has two sets of convergently transcribed genes and 186 bp long direct terminal repeats. The overall genomic architecture of the tf phage is similar to that of the previously described Pseudomonas aeruginosa phages PaP3, LUZ24 and phiMR299-2, and 39 out of the 72 products of predicted tf open reading frames have orthologs in these phages. Accordingly, tf was classified as belonging to the LUZ24-like bacteriophage group. However, taking into account very low homology levels between tf DNA and that of the other phages, tf should be considered as an evolutionary divergent member of the group. Two distinguishing features not reported for other members of the group were found in the tf genome. Firstly, a unique end structure - a blunt right end and a 4-nucleotide 3'-protruding left end - was observed. Secondly, 14 single-chain interruptions (nicks) were found in the top strand of the tf DNA. All nicks were mapped within a consensus sequence 5'-TACT/RTGMC-3'. Two nicks were analyzed in detail and were shown to be present in more than 90% of the phage population. Although localized nicks were previously found only in the DNA of T5-like and phiKMV-like phages, it seems increasingly likely that this enigmatic structural feature is common to various other bacteriophages.

Genetic characterization of tigecycline resistance in clinical isolates of Enterobacter cloacae and Enterobacter aerogenes

OBJECTIVES:
The intrinsically encoded ramA gene has been linked to tigecycline resistance through the up-regulation of efflux pump AcrAB in Enterobacter cloacae. The molecular basis for increased ramA expression in E. cloacae and Enterobacter aerogenes, as well as the role of AraC regulator rarA, has not yet been shown. To ascertain the intrinsic molecular mechanism(s) involved in tigecycline resistance in Enterobacter spp., we analysed the expression levels of ramA and rarA and corresponding efflux pump genes acrAB and oqxAB in Enterobacter spp. clinical isolates.

METHODS:
The expression levels of ramA, rarA, oqxA and acrA were tested by quantitative real-time RT-PCR. The ramR open reading frames of the ramA-overexpressing strains were sequenced; strains harbouring mutations were transformed with wild-type ramR to study altered ramA expression and tigecycline susceptibility.

RESULTS:
Tigecycline resistance was mediated primarily by increased ramA expression in E. cloacae and E. aerogenes. Only the ramA-overexpressing E. cloacae isolates showed increased rarA and oqxA expression. Upon complementation with wild-type ramR, all Enterobacter spp. containing ramR mutations exhibited decreased ramA and acrA expression and increased tigecycline susceptibility. Exceptions were one E. cloacae strain and one E. aerogenes strain, where a decrease in ramA levels was not accompanied by lower acrA expression.

CONCLUSIONS:
Increased ramA expression due to ramR deregulation is the primary mediator of tigecycline resistance in clinical isolates of E. cloacae and E. aerogenes. However, some ramA-overexpressing isolates do not show changes in ramR, suggesting alternate pathways of ramA regulation; the rarA regulator and the oqxAB efflux pump may also play a role in tigecycline resistance in E. cloacae.

Local framings

Framings provide a way to construct Quillen functors from simplicial sets to any given model category. A more structured set- up studies stable frames giving Quillen functors from spectra to stable model categories. We will investigate how this is compatible with Bousfield localisation to gain insight into the deeper structure of the stable homotopy category. We further show how these techniques relate to rigidity questions and how they can be used to study algebraic model categories.

Real-time GPU color-based segmentation of football players

In this paper, we propose a multi-camera application capable of processing high resolution images and extracting features based on colors patterns over graphic processing units (GPU). The goal is to work in real time under the uncontrolled environment of a sport event like a football match. Since football players are composed for diverse and complex color patterns, a Gaussian Mixture Models (GMM) is applied as segmentation paradigm, in order to analyze sport live images and video. Optimization techniques have also been applied over the C++ implementation using profiling tools focused on high performance. Time consuming tasks were implemented over NVIDIA's CUDA platform, and later restructured and enhanced, speeding up the whole process significantly. Our resulting code is around 4-11 times faster on a low cost GPU than a highly optimized C++ version on a central processing unit (CPU) over the same data. Real time has been obtained processing until 64 frames per second. An important conclusion derived from our study is the scalability of the application to the number of cores on the GPU. © 2011 Springer-Verlag.

Simplifying the representation of complex free-energy landscapes using sketch-map

A new scheme, sketch-map, for obtaining a low-dimensional representation of the region of phase space explored during an enhanced dynamics simulation is proposed. We show evidence, from an examination of the distribution of pairwise distances between frames, that some features of the free-energy surface are inherently high-dimensional. This makes dimensionality reduction problematic because the data does not satisfy the assumptions made in conventional manifold learning algorithms We therefore propose that when dimensionality reduction is performed on trajectory data one should think of the resultant embedding as a quickly sketched set of directions rather than a road map. In other words, the embedding tells one about the connectivity between states but does not provide the vectors that correspond to the slow degrees of freedom. This realization informs the development of sketch-map, which endeavors to reproduce the proximity information from the high-dimensionality description in a space of lower dimensionality even when a faithful embedding is not possible.

Ethical dilemmas for clinical psychologists in conducting qualitative research

Clinical psychologists often use qualitative methods to explore sensitive topics with vulnerable individuals, yet there has been little discussion of the specific ethical issues involved. For clinicians conducting qualitative research, there are likely to be ethical dilemmas associated with being both a researcher and a practitioner. We argue that this overarching issue frames all other ethical issues raised. This article provides an overview of the range of ethical issues that have been discussed in general in relation to qualitative research and considers the specific nature of these in relation to the discipline of clinical psychology. Such issues will be exemplified by reference to some of our own research and practice and the extant literature. We conclude with some suggestions for good practice, although our aim is to trigger debate rather than to establish prescriptive guidelines.

Impact of Metric Selection on Wireless DeAuthentication DoS Attack Performance

DeAuthentication Denial of Service attacks in Public Access WiFi operate by exploiting the lack of authentication of management frames in the 802.11 protocol. Detection of these attacks rely almost exclusively on the selection of appropriate thresholds. In this work the authors demonstrate that there are additional, previously unconsidered, metrics which also influence DoS detection performance. A method of systematically tuning these metrics to optimal values is proposed which ensures that parameter choices are repeatable and verifiable.

Optimization of image quality and dose for Varian aS500 electronic portal imaging devices (EPIDs)

This study was carried out to investigate whether the electronic portal imaging (EPI) acquisition process could be optimized, and as a result tolerance and action levels be set for the PIPSPro QC-3V phantom image quality assessment. The aim of the optimization process was to reduce the dose delivered to the patient while maintaining a clinically acceptable image quality. This is of interest when images are acquired in addition to the planned patient treatment, rather than images being acquired using the treatment field during a patient's treatment. A series of phantoms were used to assess image quality for different acquisition settings relative to the baseline values obtained following acceptance testing. Eight Varian aS500 EPID systems on four matched Varian 600C/D linacs and four matched Varian 2100C/D linacs were compared for consistency of performance and images were acquired at the four main orthogonal gantry angles. Images were acquired using a 6 MV beam operating at 100 MU min(-1) and the low-dose acquisition mode. Doses used in the comparison were measured using a Farmer ionization chamber placed at d(max) in solid water. The results demonstrated that the number of reset frames did not have any influence on the image contrast, but the number of frame averages did. The expected increase in noise with corresponding decrease in contrast was also observed when reducing the number of frame averages. The optimal settings for the low-dose acquisition mode with respect to image quality and dose were found to be one reset frame and three frame averages. All patients at the Northern Ireland Cancer Centre are now imaged using one reset frame and three frame averages in the 6 MV 100 MU min(-1) low-dose acquisition mode. Routine EPID QC contrast tolerance (+/-10) and action (+/-20) levels using the PIPSPro phantom based around expected values of 190 (Varian 600C/D) and 225 (Varian 2100C/D) have been introduced. The dose at dmax from electronic portal imaging has been reduced by approximately 28%, and while the image quality has been reduced, the images produced are still clinically acceptable.

THE IMPROVED SIGN BIT ENCRYPTION OF MOTION VECTORS FOR H.264/AVC

In this paper, an improved video encryption method for encrypting the sign bit of motion vectors is proposed based on H.264/AVC, which belongs to selective encryption. This method improves upon previous work involving the sign bit encryption of motion vectors by ensuring the four candidates for the encrypted motion vectors are always located in two orthogonal lines. The improved method can provide a much more effective scrambling effect while keeping the encrypted stream format-compliant and the compression ratio unchanged. The combination of the proposed method with encryption of intra prediction modes can further enhance the scrambling effect, especially for the first few frames which are left clear when only the motion vectors are encrypted.

Cinematic Peripheries of Dublin

The term periphery is, most of the time, used with or in relation to the centre. The ‘zoning’ taken by granted by the architect is unlike the one portrayed by the director who frames the differences of places in a non-linear manner. Film offers a constructed urban experience, suggesting the city to be a local network composed of nodes and links, rather than a centre and the margin. It is possible to talk about the construction of a new kind of network in film through a temporal representation of space, of the distant as the close. In this way, film may be a tool to shift the gaze from the bird’s-eye view to the eye level to create ‘a unified perceptual image of the city’, in Christine Boyer’s words. The experienced surface of the city is two-dimensional neither in fiction nor in reality. In this chapter, the nodes of Dublin are examined through two Irish films, Goldfish Memory (Elizabeth Gill, 2003) and Adam and Paul (Leonard Abrahamson, 2004). Specific elements of the city of Dublin, including walls, houses, pubs, streets, bridges, and parks, are analysed to understand the nature of the network of the city composed of nodes and their connections.

Weathering Rinds: Archives of Paleoenvironments on Mount Kenya, East Africa

Weathering-rind thicknesses on pebble-and cobble-size sediment have been used for the past half-century, at least, as an age indicator of postdepositional time following a geologic event. In mountainous terrain, rind thickness is taken as a measurement of weathering over time frames of 0.5 m.yr.; variable thicknesses are used to discriminate relative ages of glacial deposits. The effects of chemical and physical weathering that together produce rinds are only rarely considered, and most research objectives have centered on lichen alteration of clast surfaces. Recent microscopic analyses of weathering rinds on volcanic clasts of similar to 70.0-ka to similar to 2.0-m.yr. age produced new data on weathering products as well as unexpected incorporated biotic materials undergoing diagenesis. The question as to how much physical/mineral/chemical/ biotic paleoenvironmental data might be archived in rinds is discussed. The character and classification of organic materials undergoing diagenesis are also discussed.