bluuebunny/crossref_metadata_2025 huggingface dataset:
Tooth Profile Generation and Analysis of Crowned Elliptical Gears
Authors: Bair Biing-Wen
Published: June 2009

This study develops a crowned elliptical gear drive that prevents edge contact when an elliptical gear drive has axial misalignment. According to the theory of gearing and a gear-generation mechanism with longitudinal crowing, the mathematical model of a crowned elliptical gear is derived using a rack cutter. Pointed teeth typically appear on the tips of the teeth of elliptical gears at major axis. Additionally, a three-dimensional geometric relation is applied to prevent pointed teeth from developing on elliptical gears. Moreover, a computer simulation program is employed to generate the tooth profiles of crowned elliptical gears without pointed teeth and locate the contact position for longitudinal middle tooth profiles. Two numerical examples demonstrate the design process and the tooth profile graphs.
Hybrid Modeling of Ball Screw Drives With Coupled Axial, Torsional, and Lateral Dynamics
Authors: Okwudire Chinedum E. , Altintas Yusuf
Published: June 2009

It has been a common practice to assume that the torsional and axial dynamics are totally decoupled from the lateral dynamics of the screw when modeling ball screw drives. However, experiments show that there is a considerable coupling between them, which could adversely affect the positioning accuracy and fatigue life of the drive. In this paper, the lateral dynamics of the screw is explicitly incorporated into the hybrid finite element model of ball screw drives. The ball screw is modeled by Timoshenko beam elements, and the balls, joints, bearings, and fasteners are modeled as pure springs. Rigid components are modeled as lumped masses. The proposed screw-nut interface model, which includes the effects of lateral vibrations, is shown to predict the coupling between axial, torsional, and lateral dynamics of ball screw drives. The effects of this dynamic coupling on the positioning accuracy of the drive are also presented with experimental proof. The proposed model provides a more realistic platform for a designer to optimize the drive parameters for high speed-high acceleration machine tool applications, where the ball screw vibrations limit the fatigue life of the mechanism, bandwidth of the servo systems, and positioning accuracy of the machine.
GEROLAB Package System: Innovative Tool to Design a Trochoidal-Gear Pump
Authors: Gamez-Montero P. J. , Castilla R. , Mujal R. , Khamashta M. , Codina E.
Published: June 2009

An innovative tool for the improvement of the design methodology of a trochoidal gear, which works as a part of a hydraulic pump, is introduced. For the past few years, new, specific, and demanding applications of this type of pumps have demanded to maximize efficiency and to reduce manufacturing costs by improving its two main performance indexes: volumetric characteristics and teeth contact stress. The existing methodologies and software fail to thoroughly address the design by taking into account both performance indexes. The software presented is an integrated package system, which is composed of three basic modules: design, volumetric characteristics, and contact stress module. As a result, by consecutively following each module, this new tool allows to find the better gear set for the concrete initial required design parameters. An application using the GEROLAB package system methodology is carried out to obtain the best candidate, which is compared and contrasted with a gear set of a specific gerotor pump unit. As a conclusion, this new software aims to lead the designer to a better design by improving performance indexes of a new born gerotor pump project.
Assessment of Large-Eddy Simulation of Internal Separated Flow
Authors: Hahn Marco , Drikakis Dimitris
Published: June 2009

This paper presents a systematic numerical investigation of different implicit large-eddy simulations (LESs) for massively separated flows. Three numerical schemes, a third-order accurate monotonic upwind scheme for scalar conservation laws (MUSCL) scheme, a fifth-order accurate MUSCL scheme, and a ninth-order accurate weighted essentially non-oscillatory (WENO) method, are tested in the context of separation from a gently curved surface. The case considered here is a simple wall-bounded flow that consists of a channel with a hill-type curvature on the lower wall. The separation and reattachment locations, velocity, and Reynolds stress profiles are presented and compared against solutions from classical LES simulations.
Tip Vortex Cavitation Inception Scaling for High Reynolds Number Applications
Authors: Shen Young T. , Gowing Scott , Jessup Stuart
Published: June 2009

Tip vortices generated by marine lifting surfaces such as propeller blades, ship rudders, hydrofoil wings, and antiroll fins can lead to cavitation. Prediction of the onset of this cavitation depends on model tests at Reynolds numbers much lower than those for the corresponding full-scale flows. The effect of Reynolds number variations on the scaling of tip vortex cavitation inception is investigated using a theoretical flow similarity approach. The ratio of the circulations in the full-scale and model-scale trailing vortices is obtained by assuming that the spanwise distributions of the section lift coefficients are the same between the model-scale and the full-scale. The vortex pressure distributions and core sizes are derived using the Rankine vortex model and McCormick’s assumption about the dependence of the vortex core size on the boundary layer thickness at the tip region. Using a logarithmic law to describe the velocity profile in the boundary layer over a large range of Reynolds number, the boundary layer thickness becomes dependent on the Reynolds number to a varying power. In deriving the scaling of the cavitation inception index as the ratio of Reynolds numbers to an exponent m, the values of m are not constant and are dependent on the values of the model- and full-scale Reynolds numbers themselves. This contrasts traditional scaling for which m is treated as a fixed value that is independent of Reynolds numbers. At very high Reynolds numbers, the present theory predicts the value of m to approach zero, consistent with the trend of these flows to become inviscid. Comparison of the present theory with available experimental data shows promising results, especially with recent results from high Reynolds number tests. Numerical examples of the values of m are given for different model- to full-scale sizes and Reynolds numbers.
Time-Dependent Reliability Estimation for Dynamic Problems Using a Niching Genetic Algorithm
Authors: Li Jing , Mourelatos Zissimos P.
Published: June 2009

A time-dependent reliability analysis method is presented for dynamic systems under uncertainty using a niching genetic algorithm (GA). The system response is modeled as a parametric random process. A double-loop optimization algorithm is used. The inner loop calculates the maximum response in time, using a hybrid (global-local) optimization algorithm. A global GA quickly locates the vicinity of the global maximum, and a gradient-based optimizer subsequently refines its location. A time-dependent problem is, therefore, transformed into a time-independent one. The outer loop calculates multiple most probable points (MPPs), which are commonly encountered in vibration problems. The dominant MPPs with the highest contribution to the probability of failure are identified. A niching GA is used because of its ability to simultaneously identify multiple solutions. All potential MPPs are initially identified approximately, and their location is efficiently refined using a gradient-based optimizer with local metamodels. For computational efficiency, the local metamodels are built using mostly available sample points from the niching GA. Among all MPPs, the significant and independent ones are identified using a correlation analysis. Approximate limit states are built at the identified MPPs, and the system failure probability is estimated using bimodal bounds. The vibration response of a cantilever plate under a random oscillating pressure load and a point load is used to illustrate the present method and demonstrate its robustness and efficiency. A finite-element model is used to calculate the plate response.
Design Project Modularization for Product Families
Authors: Xu Qianli , Jiao Jianxin Roger
Published: June 2009

Design project management involves sophisticated task planning that needs to streamline a number of design activities while properly allocating resources associated with these activities. In the field of product family design, there is a lack of comprehensive modeling and analysis formalisms for design project management. In this respect, this paper proposes a modular design project approach to support design project modeling and planning for product families. Design activities are identified as project modules according to the activity sequences, along with the players and resources involved in each activity. A generic variety structure is introduced to characterize the differentiation of design activities among various project variants of the family. Based on the identified modules of activities, design project planning is formulated as a project reconfiguration problem, for which genetic algorithms are employed for configuration evaluation. A case study of truck cab family design in an automobile company demonstrates that design project modularization enhances the coordination of multiple design projects of product families and contributes to the achievement of economy of scale through reusing proven elements.
Numerical Simulation of Flow Past Multiple Porous Cylinders
Authors: Al-Hajeri M. H. , Aroussi A. , Witry A.
Published: June 2009

The present study numerically investigates two-dimensional laminar flow past three circular porous cylinders arranged in an in-line array. Six approaches to face velocity (Vi/Vf) ratios are used and particle trajectories are computed for a range of velocities and particle diameters. Furthermore, the flow past a solid cylinder, which had similar geometry characteristics to the porous cylinders used in this study, is compared with the flow around multiple porous cylinders. For the same range of Reynolds number (312–520), the flow behavior around the solid cylinder differs from the flow around the porous cylinders. The flow characteristics around solid cylinders are determined by the Reynolds number, whereas the flow characteristics around the porous cylinders are detrained by the Vi/Vf ratio. Stagnation areas are found behind each porous cylinder, and the size of these areas increases as the Vi/Vf velocity ratio increases. Furthermore, for the particle ranges used in power plants (<50 μm), the particles were uniformly distributed around the surface of the porous cylinders.
Investigations of Tripping Effect on the Friction Factor in Turbulent Pipe Flows
Authors: Al-Salaymeh A. , Bayoumi O. A.
Published: June 2009

Tripping devices are usually installed at the entrance of laboratory-scale pipe test sections to obtain a fully developed turbulent flow sooner. The tripping of laminar flow to induce turbulence can be carried out in different ways, such as using cylindrical wires, sand papers, well-organized tape elements, fences, etc. Claims of tripping effects have been made since the classical experiments of Nikuradse (1932, Gesetzmässigkeit der turbulenten Strömung in glatten Rohren, Forschungsheft 356, Ausgabe B, Vol. 3, VDI-Verlag, Berlin), which covered a significant range of Reynolds numbers. Nikuradse’s data have become the metric by which theories are established and have also been the subject of intense scrutiny. Several subsequent experiments reported friction factors as much as 5% lower than those measured by Nikuradse, and the authors of those reports attributed the difference to tripping effects, e.g., work of Durst et al. (2003, “Investigation of the Mean-Flow Scaling and Tripping Effect on Fully Developed Turbulent Pipe Flow,” J. Hydrodynam., 15(1), pp. 14–22). In the present study, measurements with and without ring tripping devices of different blocking areas of 10%, 20%, 30%, and 40% have been carried out to determine the effect of entrance condition on the developing flow field in pipes. Along with pressure drop measurements to compute the skin friction, both the Pitot tube and hot-wire anemometry measurements have been used to accurately determine the mean velocity profile over the working test section at different Reynolds numbers based on the mean velocity and pipe diameter in the range of 1.0×105–4.5×105. The results we obtained suggest that the tripping technique has an insignificant effect on the wall friction factor, in agreement with Nikuradse’s original data.
A New Method for Numerical Prediction of Liquid Column Separation
Accompanying Hydraulic Transients in Pipelines
Authors: Adamkowski Adam , Lewandowski Mariusz
Published: June 2009

This paper presents a new method for calculating pressure fluctuations in pipelines during a water hammer with liquid column separation. The method is based on the discrete-vapor-cavity model (DVCM). Such kind of models assumes that vaporous cavities are formed in each computational section of the pipeline whenever the pressure drops to the vapor pressure at a given temperature. The proposed new model (new DVCM) brings a significant improvement in the reliability of predictions compared with existing DVCMs. The calculation method based on it eliminates some disadvantages of basic methods used in practice, as shown by comparisons between calculations made for simple hydraulic system under theoretical frictionless conditions using various DVCMs. Additionally, the authors present preliminary verification of the proposed model based on experimental results. The positive results of this verification, and the advantages of the new DVCM, could lead to incorporating them into commercial codes.
Detailed CFD Analysis of the Steady Flow in a Wells Turbine Under Incipient and Deep Stall Conditions
Authors: Torresi M. , Camporeale S. M. , Pascazio G.
Published: June 2009

This paper presents the results of the numerical simulations carried out to evaluate the performance of a high solidity Wells turbine designed for an oscillating water column wave energy conversion device. The Wells turbine has several favorable features (e.g., simplicity and high rotational speed) but is characterized by a relatively narrow operating range with high efficiency. The aim of this work is to investigate the flow-field through the turbine blades in order to offer a description of the complex flow mechanism that originates separation and, consequently, low efficiency at high flow-rates. Simulations have been performed by solving the Reynolds-averaged Navier–Stokes equations together with three turbulence models, namely, the Spalart–Allmaras, k-ω, and Reynolds-stress models. The capability of the three models to provide an accurate prediction of the complex flow through the Wells turbine has been assessed in two ways: the comparison of the computed results with the available experimental data and the analysis of the flow by means of the anisotropy invariant maps. Then, a detailed description of the flow at different flow-rates is provided, focusing on the interaction of the tip-leakage flow with the main stream and enlightening its role on the turbine performance.
Annular Extrudate Swell of Newtonian Fluids Revisited: Extended Range of Compressible Simulations
Authors: Mitsoulis Evan
Published: June 2009

In a recent article (Mitsoulis, 2007, “Annular Extrudate Swell of Newtonian Fluids: Effects of Compressibility and Slip at the Wall,” ASME J. Fluids Eng., 129, pp. 1384–1393), numerical simulations were undertaken for the benchmark problem of annular extrudate swell of Newtonian fluids. The effects of weak compressibility and slip at the wall were studied through simple linear laws. While slip was studied in the full range of parameter values, compressibility was confined within a narrow range of values for weakly compressible fluids, where the results were slightly affected. This range is now markedly extended (threefold), based on a consistent finite element method formulation for the continuity equation. Such results correspond to foam extrusion, where compressibility can be substantial. The new extended numerical results are given for different inner/outer diameter ratios κ under steady-state conditions for Newtonian fluids. They provide the shape of the extrudate, and, in particular, the thickness and diameter swells, as a function of the dimensionless compressibility coefficient B. The pressures from the simulations have been used to compute the excess pressure losses in the flow field (exit correction). As before, weak compressibility slightly affects the thickness swell (about 1% in the range of 0≤B≤0.02) mainly by a swell reduction, after which a substantial and monotonic increase occurs for B>0.02. The exit correction increases with increasing compressibility levels in the lower B-range and is highest for the tube (κ=0) and lowest for the slit (κ=1). Then it passes through a maximum around B≈0.02, after which it decreases slowly. This decrease is attributed to the limited length of the flow channel (here chosen to be eight die gaps).
Postmating Sexual Selection Favors Males That Sire Offspring with Low Fitness
Authors: Bilde Trine , Foged Anne , Schilling Nadia , Arnqvist Göran
Published: June 2009

Two's a Crowd

The process by which males and females compete to maximize their individual fitness also affects the fitness of their offspring. Sexual selection largely results from polyandry (multiple mating by females), and several competing hypotheses attempt to explain the evolution of polyandry.

Bilde
et al.

(p.
1705
) staged double mating experiments in seed beetles to distinguish between the theories underlying cryptic female choice and sexual antagonism. Contrary to expectation, males of high genetic quality, as measured on the basis of the number of offspring sired when singly mated to a female, consistently produced fewer offspring when females were doubly mated to males of both high and low genetic quality. Thus, postmating sexual selection can favor male genotypes with low fitness, and females risk genetic costs when mating with multiple males.
Solution Nuclear Magnetic Resonance Structure of Membrane-Integral Diacylglycerol Kinase
Authors: Van Horn Wade D. , Kim Hak-Jun , Ellis Charles D. , Hadziselimovic Arina , Sulistijo Endah S. , Karra Murthy D. , Tian Changlin , Sönnichsen Frank D. , Sanders Charles R.
Published: June 2009

Opening the Portico

Escherichia coli
diacylglycerol kinase (DAGK) represents a family of integral membrane phosphotransferases that function in prokaryotic-specific metabolic pathways.

Van Horn
et al.

(p.
1726
) determined the structure of the 40-kilodalton functional homotrimer of
E. coli
DAGK by solution nuclear magnetic resonance spectroscopy. Each monomer comprises three transmembrane helices. The third transmembrane helix from each subunit is domain-swapped to pack against the first and second transmembrane helices from an adjacent subunit. These three helices frame a portico-like membrane-submerged cavity that contains residues critical for activity in close proximity to residues critical for folding. The structure provides insight into the determinants of lipid substrate specificity and phosphotransferase activity.
Medical Innovation Versus Stem Cell Tourism
Authors: Lindvall Olle , Hyun Insoo
Published: June 2009

Stem cell tourism is criticized on grounds of consumer fraud, blatant lack of scientific justification, and patient safety. However, the issues are complex because they invoke questions concerning the limits of acceptable medical innovation and medical travel. Here we discuss these issues and articulate conditions under which “unproven” therapies may be offered to patients outside of regular clinical trials.
The Role of Stromal Stem Cells in Tissue Regeneration and Wound Repair
Authors: Stappenbeck Thaddeus S. , Miyoshi Hiroyuki
Published: June 2009

The process of wound repair in epithelium-lined organs of mammals is complex and is influenced by numerous secreted factors including cytokines, growth factors, and chemokines. However, the cellular organizers of this process are still not understood. Recent studies of tissue regeneration in organisms with simpler development have uncovered details about the activity of stem cells in the mesenchyme (the blastema) during this process. These blastemal cells are well positioned to interpret cues from the environment and to execute decisions about the direction of wound repair. In mammalian wounds, stromal stem cells appear to be positioned to perform functions similar to those of blastemal cells, including communication with both the overlying epithelium and the inflammatory cells in the mesenchyme.
Genomic Footprints of a Cryptic Plastid Endosymbiosis in Diatoms
Authors: Moustafa Ahmed , Beszteri Bánk , Maier Uwe G. , Bowler Chris , Valentin Klaus , Bhattacharya Debashish
Published: June 2009

Green for Diatoms

Diatoms account for 20% of global carbon fixation and, together with other chromalveolates (e.g., dinoflagellates and coccolithophorids), represent many thousands of eukaryote taxa in the world's oceans and on the tree of life.

Moustafa
et al.

(p.
1724
; see the Perspective by

Dagan and Martin

) have discovered that the genomes of diatoms are highly chimeric, with about 10% of their nuclear genes being of foreign algal origin. Of this set of 1272 algal genes, 253 were, as expected, from a distant red algal secondary endosymbiont, but more than 1000 of the genes were derived from green algae and predated the red algal relationship. These protist taxa are important not only for genetic and genomic investigations but also for their potential in biofuel and nanotechnology applications and in global primary productivity in relation to climate change.
Buying So Children Belong
Longing and Belonging
Parents, Children, and Consumer Culture

by Allison J. Pugh

University of California Press, Berkeley, 2009. 319 pp. $55, £37.95. ISBN 9780520258433. Paper, $21.95, £14.95. ISBN 9780520258440.
Authors: Hofferth Sandra L.
Published: June 2009

Exploring what buying means to children and their parents, Pugh discusses why parents participate in the commercialization of childhood even as they decry it.
High-Resolution NMR in Magnetic Fields with Unknown Spatiotemporal Variations
Authors: Pelupessy Philippe , Rennella Enrico , Bodenhausen Geoffrey
Published: June 2009

NMR Under Stormy Conditions

Nuclear magnetic resonance spectroscopy is immensely useful for molecular characterization. However, the resolution necessary for making fine structural distinctions relies on sample isolation in a surrounding magnetic field that exhibits minimal variation in space and time over the course of the measurement; this requirement limits the range of materials that can be easily probed.

Pelupessy
et al.

(p.
1693
) present an approach that circumvents the need for spatiotemporal field homogeneity by detecting coherence transfer among coupled spins. Whereas prior methods have compensated for heterogeneity by applying corrective magnetic fields, this technique relies on the inherent insensitivity of the measured coherences to changes in the local magnetic environment and thus can be applied without prior knowledge of the field variation profile.
Caring Cooperators
Mothers and Others
The Evolutionary Origins of Mutual Understanding

by Sarah Blaffer Hrdy

Harvard University Press, Cambridge, MA, 2009. 430 pp. $29.95, £19.95, €21. ISBN 9780674032996.
Authors: Brown Gillian R.
Published: June 2009

Hrdy argues that human capacity for understanding arose from the cooperative nature of our "complicated and contingent form of childrearing."
Back to the Drawing Board for Psychiatric Genetics
Authors: Holden Constance
Published: June 2009

In 2003, researchers reported that a gene variant seemed to play a major role in whether people get depressed in response to life's stresses or sail through. But an exhaustive new analysis published last week in
The Journal of the American Medical Association
suggests that the big fish may be a minnow at best.
Bioscientists Slowly Bridge The Taiwan Strait
Authors: Normile Dennis
Published: June 2009

Researchers at a recent conference in Taipei on bioscience say that the dearth of joint work among life scientists in Taiwan and the Chinese mainland is because they haven't had the chance to get to know each other yet, which was one of the objectives of the conference.
China Reins in Wilder Impulses in Treatment of ‘Internet Addiction’
Authors: Stone Richard
Published: June 2009

Although no one doubts that logging long hours on the Internet can erode quality of life and on occasion can lead to ruinous consequences, there is no meeting of the minds on whether Internet addiction is a genuine disorder. In China, the official view appears to be that Internet addiction is a genuine disorder, but attitudes are shifting about how aggressively it should be treated.
Antibiotics in Nature: Beyond Biological Warfare
Authors: Mlot Christine
Published: June 2009

A body of evidence emerges that the infection-quelling miracle drugs of biomedicine play more basic roles in the metabolism of microbial communities.
Are We Ready for the Next Solar Maximum? No Way, Say Scientists
Authors: Kerr Richard A.
Published: June 2009

Forecasters testing their skills against the sun's mounting ferocity find themselves still in the early days of space weather prediction.
Selection and Characterization of Murine Monoclonal Antibodies toStaphylococcus aureusIron-Regulated Surface Determinant B with Functional Activity In Vitro and In Vivo
Authors: Brown Martha , Kowalski Rose , Zorman Julie , Wang Xin-min , Towne Victoria , Zhao Qinjian , Secore Susan , Finnefrock Adam C. , Ebert Tim , Pancari Greg , Isett Kevin , Zhang Yuhua , Anderson Annaliesa S. , Montgomery Donna , Cope Leslie , McNeely Tessie
Published: June 2009

In an effort to characterize important epitopes ofStaphylococcus aureusiron-regulated surface determinant B (IsdB), murine IsdB-specific monoclonal antibodies (MAbs) were isolated and characterized. A panel of 12 MAbs was isolated. All 12 MAbs recognized IsdB in enzyme-linked immunosorbent assays and Western blots; 10 recognized native IsdB expressed byS. aureus. The antigen epitope binding of eight of the MAbs was examined further. Three methods were used to assess binding diversity: MAb binding to IsdB muteins, pairwise binding to recombinant IsdB, and pairwise binding to IsdB-expressing bacteria. Data from these analyses indicated that MAbs could be grouped based on distinct or nonoverlapping epitope recognition. Also, MAb binding to recombinant IsdB required a significant portion of intact antigen, implying conformational epitope recognition. Four MAbs with nonoverlapping epitopes were evaluated for in vitro opsonophagocytic killing (OPK) activity and efficacy in murine challenge models. These were isotype switched from immunoglobulin G1 (IgG1) to IgG2b to potentially enhance activity; however, this isotype switch did not appear to enhance functional activity. MAb 2H2 exhibited OPK activity (≥50% killing in the in vitro OPK assay) and was protective in two lethal challenge models and a sublethal indwelling catheter model. MAb 13C7 did not exhibit OPK (<50% killing in the in vitro assay) and was protective in one lethal challenge model. Neither MAb 13G11 nor MAb 1G3 exhibited OPK activity in vitro or was active in a lethal challenge model. The data suggest that several nonoverlapping epitopes are recognized by the IsdB-specific MAbs, but not all of these epitopes induce protective antibodies.
Lipoprotein Lipase and Hydrofluoric Acid Deactivate Both Bacterial Lipoproteins and Lipoteichoic Acids, but Platelet-Activating Factor-Acetylhydrolase Degrades Only Lipoteichoic Acids
Authors: Seo Ho Seong , Nahm Moon H.
Published: June 2009

To identify the Toll-like receptor 2 ligand critically involved in infections with gram-positive bacteria, lipoprotein lipase (LPL) or hydrogen peroxide (H2O2) is often used to selectively inactivate lipoproteins, and hydrofluoric acid (HF) or platelet-activating factor-acetylhydrolase (PAF-AH) is used to selectively inactivate lipoteichoic acid (LTA). However, the specificities of these chemical reactions are unknown. We investigated the reaction specificities by using two synthetic lipoproteins (Pam3CSK4and FSL-1) and LTAs from pneumococci and staphylococci. Changes in the structures of the two synthetic proteins and the LTAs were monitored by mass spectrometry, and biological activity changes were evaluated by measuring tumor necrosis factor alpha production by mouse macrophage cells (RAW 264.7) following stimulation. PAF-AH inactivated LTA without reducing the biological activities of Pam3CSK4and FSL-1. Mass spectroscopy confirmed that PAF-AH monodeacylated pneumococcal LTA but did not alter the structure of either Pam3CSK4or FSL-1. As expected, HF treatment reduced the biological activity of LTA by more than 80% and degraded LTA. HF treatment not only deacylated Pam3CSK4and FSL-1 but also reduced the activities of the lipoproteins by more than 60%. Treatment with LPL decreased the biological activities by more than 80%. LPL also removed an acyl chain from the LTA and reduced its activity. Our results indicate that treatment with 1% H2O2for 6 h at 37°C inactivates Pam3CSK4, FSL-1, and LTA by more than 80%. Although HF, LPL, and H2O2treatments degrade and inactivate both lipopeptides and LTA, PAF-AH selectively inactivated LTA with no effect on the biological and structural properties of the two lipopeptides. Also, the ability of PAF-AH to reduce the inflammatory activities of cell wall extracts from gram-positive bacteria suggests LTA to be essential in inflammatory responses to gram-positive bacteria.
Intracellular Infections Enhance Interleukin-6 and Plasminogen Activator Inhibitor 1 Production by Cocultivated Human Adipocytes and THP-1 Monocytes
Authors: Bouwman J. J. M. , Diepersloot R. J. A. , Visseren F. L. J.
Published: June 2009

Obesity is associated with a chronic inflammatory state, and adipocyte dysfunction is thought to play a crucial role in this. Infection of adipose tissue may trigger the production of inflammatory cytokines, leading to increased recruitment of macrophages into adipose tissue, which in turn may exacerbate the inflammatory state in obesity. Low-grade inflammation was mimicked in an in vitro coculture model with human adipocytes and THP-1 monocytes. Adipocytes and monocytes were infected with adenovirus, cytomegalovirus (CMV), or influenza A virus. After 48 h, transinfection was evaluated and interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-α), adiponectin, and plasminogen activator inhibitor 1 (PAI-1) were measured. IL-6 production was upregulated in cocultures of uninfected adipocytes and THP-1 macrophages in a THP-1 cell number-dependent fashion. IL-6 production by CMV-infected adipocytes was increased relative to that of uninfected adipocytes (
P
< 0.01). IL-6 production by CMV-infected cocultures was 16- to 37-fold higher than that of uninfected adipocytes (
P
< 0.001). IL-6 production in influenza A virus-infected cocultures was increased 12- to 20-fold (
P
< 0.05). Only CMV infection increased levels of PAI-1 in cocultures (fourfold;
P
< 0.05). Soluble factors produced by THP-1 macrophages rather than by adipocytes were responsible for the increased production of IL-6 in cocultures. Infection of cocultivated human adipocytes and THP-1 monocytes with CMV or influenza A virus led to increased production of IL-6 and PAI-1. Thus, infection of adipose tissue evokes an inflammatory response, leading to adipose tissue dysfunction and subsequent overproduction of IL-6 and PAI-1. This may further compound the atherogenic effects of obesity.
Development and Technical and Clinical Validation of a Quantitative Enzyme-Linked Immunosorbent Assay for the Detection of Human Antibodies to Hepatitis B Surface Antigen in Recipients of Recombinant Hepatitis B Virus Vaccine
Authors: Cambron Pierre , Jacquet Jeanne-Marie , Hoet Bernard , Lievens Marc
Published: June 2009

Pending removal from the market of a commercial assay (the AUSAB [Abbott Laboratories] enzyme immunoassay [EIA]) for the determination of antibodies to hepatitis B surface antigen (HBsAg), a new in-house quantitative enzyme-linked immunosorbent assay (ELISA) to measure antibodies against HBsAg (anti-HBs) was developed (anti-HBs in-house). Specific anti-HBs antibodies were sandwiched between the precoated HBsAg ad and ay subtypes purified from plasma from hepatitis B virus (HBV) human carriers and the recombinant HBsAg adw2 subtype tagged with horseradish peroxidase. The assay was calibrated against the 1st International Reference Preparation for anti-hepatitis B immunoglobulin (lot 1977-W1042). Analytical sensitivity and the limit of quantitation were estimated at 0.43 mIU/ml and 2.0 mIU/ml, respectively. Overall reproducibility was 11.86%, and accuracy was estimated to be 94.89%. More than 4,000 samples from seven clinical trials were tested with the anti-HBs in-house assay and compared to results generated with AUSAB EIA and AUSAB radioimmunoassay (RIA). During the technical validation, the anti-HBs in-house assay was compared to the AUSAB RIA as a reference (
n
= 919). Overall assessment of concordance and Deming's regression analysis were performed. The coefficient of correlation between the AUSAB RIA and anti-HBs in-house assay was 0.9815 with a slope of 0.9187. The overall agreement between anti-HBs in-house and AUSAB RIA was 97.61%, considering the clinical cutoffs at 3.3 mIU/ml and 1.0 mIU/ml for the respective assays. From a clinical perspective, seroprotection rates and anti-HBs geometric mean antibody concentrations for individual studies calculated with either the in-house assay or the reference assays were similar. Conclusions of individual studies were confirmed. The performance characteristics of the in-house assay are acceptable. There is no evidence that use of the new assay would lead to different clinical conclusions from the reference method.
Quantitative Detection of the M204V Hepatitis B Virus Minor Variants by Amplification Refractory Mutation System Real-Time PCR Combined with Molecular Beacon Technology
Authors: Ntziora F. , Paraskevis D. , Haida C. , Magiorkinis E. , Manesis E. , Papatheodoridis G. , Manolakopoulos S. , Beloukas A. , Chryssoy S. , Magiorkinis G. , Sypsa V. , Hatzakis A.
Published: June 2009

Mutations in the highly conserved tyrosine-methionine-aspartate-aspartate (YMDD) motif are frequently associated with resistance to antivirals and represent a major concern in the treatment of hepatitis B virus (HBV) infection. Conventional methods fail to detect minority populations of drug-resistant viral quasispecies if they represent less than 25% of the total sample virus population. The amplification refractory mutation system real-time PCR (ARMS RT-PCR) was combined with molecular beacon technology using the LightCycler system. The samples from HBV patients selected for assay evaluation included (i) 57 samples from treatment-naïve patients for biological discriminatory ability (cutoff) estimation, (ii) 12 samples from patients with treatment failure that were M204V positive by sequencing, and (iii) 13 samples from patients with treatment failure that were negative for mutation at codon 204 by sequencing. The discriminatory ability of the assay was 0.25% when tested with laboratory-synthesized DNA target sequences. The median mutant-to-wild-type ratio for samples from naive patients tested positive for the wild type and for mutant variants was 0.01% (5th and 95th percentiles = 0.0001 and 0.04%, respectively). A value of 0.04% was selected as the biological cutoff of the assay of clinical samples. In all samples M204V positive by sequencing (12/12), the mutant variant was detected as the predominant population (range, 82.76 to 99.43%). Interestingly, in 5 (38%) of 13 samples negative by sequencing, the M204V variant was detected at a ratio above the biological cutoff (0.05 to 28%). The assay represents an efficient technique for the early detection and quantification of M204V variants before mutant strains emerge to dominate the population.
Emerging Genotypes of Human Respiratory Syncytial Virus Subgroup A among Patients in Japan
Authors: Shobugawa Yugo , Saito Reiko , Sano Yasuko , Zaraket Hassan , Suzuki Yasushi , Kumaki Akihiko , Dapat Isolde , Oguma Taeko , Yamaguchi Masahiro , Suzuki Hiroshi
Published: June 2009

Human respiratory syncytial virus (HRSV) is a common etiological agent of acute lower respiratory tract disease in infants. We report the molecular epidemiology of HRSV in Niigata, Japan, over six successive seasons (from 2001 to 2007) and the emerging genotypes of HRSV subgroup A (HRSV-A) strains. A total of 488 HRSV samples were obtained from 1,103 screened cases in a pediatric clinic in Niigata. According to the phylogenetic analysis, among the PCR-positive samples, 338 HRSV-A strains clustered into the previously reported genotypes GA5 and GA7 and two novel genotypes, NA1 and NA2, which were genetically close to GA2 strains. One hundred fifty HRSV-B strains clustered into three genotypes, namely, GB3, SAB3, and BA, which has a 60-nucleotide insertion in the second hypervariable region of the G protein. The NA1 strains emerged first, in the 2004-2005 season, and subsequently, the NA2 strain emerged in the 2005-2006 season. Both strains caused large epidemics in the 2005-2006 and 2006-2007 seasons. The average age of children who were infected with NA2 strains was significantly higher than that of those infected with GA5 and the frequency of reinfection by NA2 was the highest among all genotypes, suggesting that this genotype possessed new antigenicity for evading past host immunity. This is the first paper to show a possible correlation between an emerging genotype, NA2, and large outbreaks of HRSV in Japan. Continuing studies to follow up the genetic changes and to clarify the mechanism of reinfection in HRSV are important steps to understand HRSV infections.
Detection and Genotyping of Human Papillomavirus in Self-Obtained Cervicovaginal Samples by Using the FTA Cartridge: New Possibilities for Cervical Cancer Screening
Authors: Lenselink Charlotte H. , de Bie Roosmarie P. , van Hamont Dennis , Bakkers Judith M. J. E. , Quint Wim G. V. , Massuger Leon F. A. G. , Bekkers Ruud L. M. , Melchers Willem J. G.
Published: June 2009

This study assesses human papillomavirus (HPV) detection and genotyping in self-sampled genital smears applied to an indicating FTA elute cartridge (FTA cartridge). The study group consisted of 96 women, divided into two sample sets. All samples were analyzed by the HPV SPF
10
-Line Blot 25. Set 1 consisted of 45 women attending the gynecologist; all obtained a self-sampled cervicovaginal smear, which was applied to an FTA cartridge. HPV results were compared to a cervical smear (liquid based) taken by a trained physician. Set 2 consisted of 51 women who obtained a self-sampled cervicovaginal smear at home, which was applied to an FTA cartridge and to a liquid-based medium. DNA was obtained from the FTA cartridges by simple elution as well as extraction. Of all self-obtained samples of set 1, 62.2% tested HPV positive. The overall agreement between self- and physician-obtained samples was 93.3%, in favor of the self-obtained samples. In sample set 2, 25.5% tested HPV positive. The overall agreement for high-risk HPV presence between the FTA cartridge and liquid-based medium and between DNA elution and extraction was 100%. This study shows that HPV detection and genotyping in self-obtained cervicovaginal samples applied to an FTA cartridge is highly reliable. It shows a high level of overall agreement with HPV detection and genotyping in physician-obtained cervical smears and liquid-based self-samples. DNA can be obtained by simple elution and is therefore easy, cheap, and fast. Furthermore, the FTA cartridge is a convenient medium for collection and safe transport at ambient temperatures. Therefore, this method may contribute to a new way of cervical cancer screening.
Evaluation of GenoType Mycobacteria Direct Assay in Comparison with Gen-Probe
Mycobacterium tuberculosis
Amplified Direct Test and GenoType MTBDR
plus
for Direct Detection of
Mycobacterium tuberculosis
Complex in Clinical Samples
Authors: Neonakis Ioannis K. , Gitti Zoe , Baritaki Stavroula , Petinaki Efi , Baritaki Maria , Spandidos Demetrios A.
Published: June 2009

Three molecular assays were evaluated for the direct detection of
Mycobacterium t
uberculosis
complex bacteria in 125 respiratory and 22 nonrespiratory samples. The overall sensitivities obtained were as follows: GenoType MTBDR
plus
, 97.9%; GenoType Mycobacteria Direct, 93.7%; Gen-Probe
Mycobacterium tuberculosis
Amplified Direct Test, 89.6%. The specificity of the assays used was 100%.
Evaluation of MeningoFinder, a Novel Multiplex Ligation-Dependent Probe Amplification Assay for Simultaneous Detection of Six Virus Species Causing Central Nervous System Infections
Authors: Wolffs P. F. G. , Vink C. , Keijdener J. , Habek B. , Reijans M. , Simons G. , Bruggeman C. A. , van den Brule A. J. C.
Published: June 2009

A multiplex ligation-dependent probe amplification assay for simultaneous detection of six virus species was developed and tested on clinical cerebrospinal fluid (CSF) samples. The assay, termed MeningoFinder, showed an accordance of 97%, concordance of 96%, interlaboratory sensitivity of 90%, and interlaboratory specificity of 94% compared to PCRs.
Therapeutic Memory T Cells Require Costimulation for Effective Clearance of a Persistent Viral Infection
Authors: Garidou Lucile , Heydari Sara , Truong Phi , Brooks David G. , McGavern Dorian B.
Published: June 2009

Persistent viral infections are a major health concern worldwide. During persistent infection, overwhelming viral replication and the rapid loss of antiviral T-cell function can prevent immune-mediated clearance of the infection, and therapies to reanimate the immune response and purge persistent viruses have been largely unsuccessful. Adoptive immunotherapy using memory T cells is a highly successful therapeutic approach to eradicate a persistent viral infection. Understanding precisely how therapeutically administered memory T cells achieve clearance should improve our ability to terminate states of viral persistence in humans. Mice persistently infected from birth with lymphocytic choriomeningitis virus are tolerant to the pathogen at the T-cell level and thus provide an excellent model to evaluate immunotherapeutic regimens. Previously, we demonstrated that adoptively transferred memory T cells require recipient dendritic cells to effectively purge an established persistent viral infection. However, the mechanisms that reactivate and sustain memory T-cell responses during clearance of such an infection remain unclear. Here we establish that therapeutic memory T cells require CD80 and CD86 costimulatory signals to efficiently clear an established persistent viral infection in vivo. Early blockade of costimulatory pathways with CTLA-4-Fc decreased the secondary expansion of virus-specific CD8+and CD4+memory T cells as well as their ability to produce antiviral cytokines and purge the persistent infection. Late costimulation blockade also reduced virus-specific T-cell numbers, illustrating that sustained interactions with costimulatory molecules is required for efficient T-cell expansion. These findings indicate that antiviral memory T cells require costimulation to efficiently clear a persistent viral infection and that costimulatory pathways can be targeted to modulate the magnitude of an adoptive immunotherapeutic regimen.
Kaposi's Sarcoma-Associated Herpesvirus Protein LANA2 Disrupts PML Oncogenic Domains and Inhibits PML-Mediated Transcriptional Repression of the Survivin Gene
Authors: Marcos-Villar Laura , Lopitz-Otsoa Fernando , Gallego Pedro , Muñoz-Fontela Cesar , González-Santamaría José , Campagna Michela , Shou-Jiang Gao , Rodriguez Manuel S. , Rivas Carmen
Published: June 2009

Infection by herpesviruses causes a dramatic disturbance of PML oncogenic domains (PODs) that has been suggested to be essential for viral lytic replication. Several proteins from Kaposi's sarcoma-associated herpesvirus (KSHV) have been tested as putative POD-disrupting factors with negative results. Here, we show that LANA2, a viral protein that is absolutely required for the viability and proliferation of KSHV-infected primary effusion lymphoma (PEL) cells, increases the levels of SUMO2-ubiquitin-modified PML and induces the disruption of PODs by a proteasome-mediated mechanism. In addition, we demonstrate that this disruption is largely dependent on both the integrity of a SUMO interaction motif in LANA2 and the lysine 160 from PML. Moreover, silencing of LANA2 expression in PEL cells by RNA interference led to an increase in the PML levels. Finally, we demonstrate that LANA2 relieves PML-mediated transcriptional repression of survivin, a protein that directly contributes to malignant progression of PEL. This represents the first example of inactivation of these important antiviral structures by KSHV.
Differential Rates of Protein Folding and Cellular Trafficking for the Hendra Virus F and G Proteins: Implications for F-G Complex Formation
Authors: Whitman Shannon D. , Smith Everett Clinton , Dutch Rebecca Ellis
Published: June 2009

Hendra virus F protein-promoted membrane fusion requires the presence of the viral attachment protein, G. However, events leading to the association of these glycoproteins remain unclear. Results presented here demonstrate that Hendra virus G undergoes slower secretory pathway trafficking than is observed for Hendra virus F. This slowed trafficking is not dependent on the G protein cytoplasmic tail, the presence of the G receptor ephrin B2, or interaction with other viral proteins. Instead, Hendra virus G was found to undergo intrinsically slow oligomerization within the endoplasmic reticulum. These results suggest that the critical F-G interactions occur only after the initial steps of synthesis and cellular transport.
Proteolytic Activation of the Spike Protein at a Novel RRRR/S Motif Is Implicated in Furin-Dependent Entry, Syncytium Formation, and Infectivity of Coronavirus Infectious Bronchitis Virus in Cultured Cells
Authors: Yamada Yoshiyuki , Liu Ding Xiang
Published: June 2009

The spike (S) protein of the coronavirus (CoV) infectious bronchitis virus (IBV) is cleaved into S1 and S2 subunits at the furin consensus motif RRFRR
537
/S in virus-infected cells. In this study, we observe that the S2 subunit of the IBV Beaudette strain is additionally cleaved at the second furin site (RRRR
690
/S) in cells expressing S constructs and in virus-infected cells. Detailed time course experiments showed that a peptide furin inhibitor, decanoyl-Arg-Val-Lys-Arg-chloromethylketone, blocked both viral entry and syncytium formation. Site-directed mutagenesis studies revealed that the S1/S2 cleavage by furin was not necessary for, but could promote, syncytium formation by and infectivity of IBV in Vero cells. In contrast, the second site is involved in the furin dependence of viral entry and syncytium formation. Mutations of the second site from furin-cleavable RRRR/S to non-furin-cleavable PRRRS and AAARS, respectively, abrogated the furin dependence of IBV entry. Instead, a yet-to-be-identified serine protease(s) was involved, as revealed by protease inhibitor studies. Furthermore, sequence analysis of CoV S proteins by multiple alignments showed conservation of an XXXR/S motif, cleavable by either furin or other trypsin-like proteases, at a position equivalent to the second IBV furin site. Taken together, these results suggest that proteolysis at a novel XXXR/S motif in the S2 subunit might be a common mechanism for the entry of CoV into cells.