bluuebunny/crossref_metadata_2025 huggingface dataset:
Investigation of Light Interaction With Transient Expanding Thick Flames
Authors: Parsinejad Farzan , Metghalchi Hameed
Published: May 2008

Optical behavior of spherical flames has been investigated using Shadowgraph method. A mathematical model has been developed to predict the intensity profile of refracted light beams interacting with a transient expanding thick flame. Experimental facilities have been built to visualize transient expanding spherical flames. Facilities include a cylindrical chamber with two end glasses for optical observation. Shadowgraph pictures of flame propagation have been taken using a high speed Charged Coupled Device camera. Flame edge has been located using the mathematical model and it has been compared to experimental measurements. Experimental results are in very good agreement with those predicted by the theoretical model.
The Use of Blind-Source-Separation Algorithm for Mechanical Signal Separation and Machine Fault Diagnosis
Authors: Tse Peter W. , Zhang Jinyu
Published: May 2008

Vibration based machine fault diagnosis is widely adopted in machine condition monitoring. Since a machine is usually composed of many mechanical components, during the machine running, each component will generate its vibration and transmit to other components thru the shaft or linkages. Hence, the vibration signal collected from a sensor is the aggregation of all generated vibrations. To enhance the accuracy in vibration based machine fault diagnosis, the vibration generated by each component must be isolated and identified. In this paper, the performance of blind-source-separation (BSS) in separating various mixed sources is discussed. The BSS based method of second order statistics (SOS) has been applied to separate the aggregated vibration signals generated from a number of mechanical components. To verify the effectiveness of the BSS based SOS, a number of experiments were conducted using both simulated data and vibration generated form the industrial machines. The results show that the BSS possesses the ability to separate both artificially and naturally mixed signals. Such ability is definitely welcome in the fields of condition monitoring and maintenance. Moreover, the paper also discusses the advantages and disadvantages of the algorithm in the applications of machine fault diagnosis and future improvements.
Probabilistic Analyses for the Performance Characteristics of the Main Bearings in an Operating Engine Due to Variability in Bearing Properties
Authors: Wang Jin , Vlahopoulos Nickolas , Mourelatos Zissimos P. , Ebrat Omidreza , Vaidyanathan Kumar
Published: May 2008

This paper presents the development of surrogate models (metamodels) for evaluating the bearing performance in an internal combustion engine. The metamodels are employed for performing probabilistic analyses for the engine bearings. The metamodels are developed based on results from a simulation solver computed at a limited number of sample points, which sample the design space. An integrated system-level engine simulation model, consisting of a flexible crankshaft dynamics model and a flexible engine block model connected by a detailed hydrodynamic lubrication model, is employed in this paper for generating information necessary to construct the metamodels. An optimal symmetric Latin hypercube algorithm is utilized for identifying the sampling points based on the number and the range of the variables that are considered to vary in the design space. The development of the metamodels is validated by comparing results from the metamodels with results from the actual simulation models over a large number of evaluation points. Once the metamodels are established they are employed for performing probabilistic analyses. The initial clearance between the crankshaft and the bearing at each main bearing and the oil viscosity comprise the random variables in the probabilistic analyses. The maximum oil pressure and the percentage of time (the time ratio) within each cycle that a bearing operates with oil film thickness less than a user defined threshold value at each main bearing constitute the performance variables of the system. The availability of the metamodels allows comparing the performance of several probabilistic methods in terms of accuracy and computational efficiency. A useful insight is gained by the probabilistic analysis on how variability in the bearing characteristics affects its performance.
Integration of Reverse Osmosis Desalination With Cold-Heat-Power Production in the Tertiary Sector
Authors: Foronda C. , Serra L. , Uche J. , Verda V. , Valero A.
Published: May 2008

A preliminary design of a polygeneration system, conceived for a four-star hotel located on the Mediterranean coast, is presented in this paper. For this system, a single fuel is used to produce cold, heat, power and water from an internal combustion engine integrated and optimized with heaters, absorption units and a reverse osmosis desalination plant. The proposed system is the result of minimizing a multiobjective function of energy (cold, heat and power) and water consumption, in order to reach a more efficient and sustainable management of the installation. The hotel covers its internal demand of energy and water and reaches important overall energy efficiency improvements thanks to the integration of all the systems. An economic analysis has also been performed. The overall system has been modeled by means of data provided by manufacturers and also by means of efficiency curves available in the literature.
Numerical and Experimental Study of Pin-Fin Array Heat Exchangers
Authors: Hamilton Leonard J. , Gopinath Ashok
Published: May 2008

The performance characteristics of a staggered short pin-fin array heat exchanger (Figure 3) were analyzed numerically and experimentally. The numerical simulation formed the primary thrust of the study and a three-dimensional finite element model based on the commercial FEA package ANSYS was used. The simulation was validated against historical empirical data in the literature, as well as against data obtained from a modular experimental rig that was designed to allow different configurations to be easily swapped and studied. Compared to the experimental data, the numerical model provided very accurate heat transfer predictions (Figure 2), but overestimated the pressure drop values for some configurations. The discrepancy was greatest for cases where the pin-fins made up a large fraction of the total heat exchanger array volume.
Dynamic Modelling and Identification of a Waterjet Cutting System
Authors: Fabien Brian C. , Ramulu M.
Published: May 2008

This paper describes a new dynamic model for a waterjet cutting system that includes a double-acting reciprocating intensifier pump. Since the system operates at high pressures the fluid flow is assumed to be compressible. The dynamic model includes the characteristics of the intensifier pump, the check valves, the accumulator, the system piping and compressible jet flow through the nozzle. The system model is presented as a set of differential-algebraic equations. Experimental results for an actual system are used to identify the discharge coefficient of the nozzle, certain unknown parameters associated with the check valve, and to determine the velocity profile of the piston in the intensifier pump. The identification is accomplished by formulating and solving a parameter optimization problem. The paper also includes numerical simulation results that validate the dynamic model.
Numerical Determination of the Moment Lyapunov Exponents
Authors: Xie Wei-Chau , So Ronald M. C.
Published: May 2008

Two numerical methods for the determination of the pth moment Lyapunov exponents of a two-dimensional system under bounded noise or real noise parametric excitation are presented. The first method is an analytical-numerical approach, in which the partial differential eigenvalue problems governing the moment Lyapunov exponents are established using the theory of stochastic dynamical systems. The eigenfunctions are expanded in double series to transform the partial differential eigenvalue problems to linear algebraic eigenvalue problems, which are then solved numerically. The second method is a Monte Carlo simulation approach. The numerical values obtained are compared with approximate analytical results with weak noise amplitudes.
The Value of a Space Precursor Analysis Program: A Saturn Example
Authors: Fragola Joseph R. , Collins Erin P.
Published: May 2008

There is general agreement that “near miss” or “close call” data is valuable to the space program and whenever NASA becomes convinced that such events have occurred they have acted responsibly and quickly to address them. The problem lies in defining what constitutes a near miss in a system that is inherently very complex, such that ‘abnormalities’ are actually normal occurrences, and yet one that is relatively reliable because of the inherent strengths incorporated into the design, such as robustness, redundancy, and functional diversity. The question becomes: what would the consideration of failure precursors add to the insights to be drawn from history as it relates to forecasting future performance? This paper will use the example of the Saturn program to address the problems involved in forecasting the risk in complex, yet reasonably reliable programs and to indicate preliminary approaches for use in establishing a space industry precursor program.
The Tensile Properties of Three Dimensional Air Textured Aramid Reinforced Composites
Authors: Langston Thomas Brice , Qiu Yiping
Published: May 2008

Air-textured aramid yarns (ATAY) and regular aramid yarns (RAY) were used in this study to fabricate 3D orthogonal woven composites. The composites were tested in warp, weft, and 45° directions to determine the engineering constants of the composites. The ATAY composite had a much lower fiber volume fraction than the RAY composite due to the bulkiness of the textured yarns. With the same fiber volume fraction, the ATAY composite had a slightly lower tensile strength and modulus, but a 120% higher in-plane shear modulus, than the RAY composite. Unlike the RAY composite that demonstrated a brittle failure, the ATAY composite failed in a ductile manner with multiple diverting cracks propagating during failure. The ATAY composite had a much higher yield point in the 45° direction tensile test and a much higher softening point in the warp direction tensile test than the RAY composite. The loop entanglements of ATAY are responsible for all the improvements observed in this study.
Diagonal-Flow, Gap-Pickup Generator Rotor Heat Transfer Model
Authors: Costley Justin M. , McLeskey James T.
Published: May 2008

A computational model of the temperature distribution in a hydrogen-cooled, diagonal-flow, gap-pickup generator rotor has been developed. The model will aid in the analysis of the uprate capabilities of turbo-generators during steam turbine upgrades. Under certain operating conditions, knowledge of hot-spot temperatures and the temperature profile in the rotor windings is indispensable in determining the proper rating levels and loading limits. Following a general description of gap-pickup generator rotors and the advantages associated with their use, a description of the methods used in analyzing the hydrogen flow and heat transfer characteristics are presented. The development of the computer model is described with an overview of the design options available. Finally, a sample calculation is presented with an interpretation of the significance of the results.
The Homicidal Chauffeur Problem With Time Delayed Feedback
Authors: Olgac Nejat , Sipahi Rifat , Ergenc Ali Fuat
Published: May 2008

The classical pursuit-evasion scenario (target tracking) is studied with an interesting perspective. A homicidal chauffeur (pursuer) tries to capture an evader on a 2-dimensional platform (a parking lot). It utilizes a certain feedback control law, which enables the capture. If, however, a time delay is introduced for the sensory perception of the pursuer the performance of the pursuer can show dramatic changes. It may or may not capture its victim depending on this very critical parameter, the sensory time delay. The unique perspective is introduced at this feature.
Vibration Localization in Plates With Misplaced Intermediate Supports in Two Orthogonal Directions
Authors: Xie Wei-Chau , Chen Zhihua
Published: May 2008

This paper investigates the effect of small misplacements on both the free vibration modes and forced vibration responses of a four-panel simply supported plate with two intermediate simple supports in two orthogonal directions. Kantorovich’s method is employed to obtain the natural frequencies and the corresponding vibration modes. Galerkin’s method is applied to determine the forced vibration response of the four-panel plate subjected to a harmonic concentrated load at the center of one of its four panels. Comparisons between numerical results obtained using the current approach and those using the exact solution and the finite element method are made to demonstrate the accuracy of the current approach. It is found that the small misplacements of the intermediate supports have dramatic effects on the vibration response of the plate. The larger the misplacement of the intermediate supports, the larger the degree of vibration localization. By judiciously introducing misplacements in the intermediate simple supports, the vibration of some of the four panels may be controlled.
Forced Oscillations of Microelectromechanical Resonators
Authors: Preidikman S. , Li H. , Balachandran B.
Published: May 2008

Forced oscillations of micromechanical resonators fabricated as clamped-clamped composite structures are studied in this effort. Piezoelectric actuation is used to excite these structures on the input side and piezoelectric sensing is carried out on the output side. Each resonator structure is modeled as a beam with stepwise properties and a distributed actuation. For weak damping and weak forcing, a nonlinear analysis is conducted to obtain an approximate solution of the system. In this analysis, the method of multiple scales is used on the governing partial differential system and a solution is sought to describe oscillations about a buckled position. The different modeling assumptions made are presented and discussed.
Surface Tension Measurements of Non-Spherical Droplets Using Acoustic Levitation
Authors: Fuentes Arturo A. , Bayazitoglu Yildiz
Published: May 2008

This paper deals with containerless thermophysical property measurement of deformed droplets. This work is partially motivated in the need for alternatives to study dynamic features of aggressive solvents that can damage certain instruments and/or yield incorrect results. Measurements of small-amplitude shape oscillations of droplets are carried out to obtain surface tension. The theoretical analysis, which is presented in detailed in a previous publication, studies the effect of static deformation due to an arbitrary external levitating force on the oscillations of a droplet, especially regarding the splitting of the frequency spectrum. Detailed explanation of the experimental apparatus is given. A novel experimental procedure is presented which includes the estimation of the droplet’s deformation transfer function magnitude. Experimental data and observations on the frequency splitting and surface tension of acoustically levitated samples are presented; in the initial experiments the surface tension measured came within 1% of the published values.
Moving Boundary Model of Once Through Boiler Evaporators
Authors: Ahnert F. , Colonna P. , Kikstra J. F.
Published: May 2008

Dynamic modeling and simulation of steam power plants is often adopted as a tool for control design, personnel training, efficiency improvement and on-line diagnostic. The boiler is possibly the most complex component of the thermal power plant. A usual boiler configuration is the so-called Once-Through arrangement. A common problem in 2-phase systems modeling is the correct calculation of the phase boundary. This is technically interesting in such boilers: the location of the phase transition changes rapidly depending on load conditions and temperature distribution along the walls. A lumped parameters, one-dimensional evaporator model implementing a moving boundary approach is presented and first validation results are discussed. The model takes into account the influence of radiation and convection on the gas side. The flow inside the pipes is divided into 3 regions (sub-cooled, 2-phase, superheated) and the model calculates the locations of the 2-phase transitions and the average steam quality along the pipes. The system is discretized using a staggered grid for higher numerical stability and is implemented in the computer program Aspen Custom Modeler (ACM). Results include the calculation of the system response to input signals simulating a load variation and a validation by comparison with a model implemented in a commercial software for power plant simulations (MMS). Input data, parameters and geometry are taken from an existing plant operating in Uppsala, Sweden.
Numerical Effects on the Prediction of Flow Instabilities in Channels With Sudden-Expansions
Authors: Patel S. , Drikakis D.
Published: May 2008

We have considered the problem of flow through a suddenly-expanded channel and performed a computational investigation to examine numerical effects on the prediction of flow instability and bifurcation phenomena. The results revealed that the solution of the flow depends on the numerical method employed. We have employed Godunov-type methods in conjunction with first-, second- and third-order accurate interpolation schemes. It is shown that the order of accuracy of the interpolation used in the discretisation of the wave-speed dependent term and averaged part of the intercell flux affects the prediction of the instability. Computations using first-order discretisation for the calculation of the flux components results in symmetric stable flow, whereas second- and third-order discretisations lead to a symmetry-breaking bifurcation.
An Electrohydrodynamic (EHD) Meso Pump for Cryogenic Spot Cooling of Low Volume Complex Electronics
Authors: Rada M. , Ohadi M. , Shooshtari A.
Published: May 2008

Low temperature cooling provides excellent operating conditions for some conventional and superconducting electronics applications. The cooling requirements for these applications include moderate to low power levels (down to fraction of Watts) and very strict spatial and temporal temperature variation (in the order of 0.1K or less). In this paper, experimental and modeling results of a liquid nitrogen cooling system, using an EHD meso pump are presented. The pumping mechanism uses the EHD ion-drag technique. The experimental set-up, procedures and results are presented, followed by a pumping curve obtained by numerical modeling.
Site Redressing Techniques and Rework of Lead-Free Chip Scale Packages
Authors: Gowda Arun , Primavera Anthony , Srihari K.
Published: May 2008

The implementation of lead-free solder into an electronics assembly process necessitates the reassessment of the individual factors involved in component attachment and rework. A component assembly undergoes multiple thermal cycles during rework. With the use of lead-free solder, the assemblies are subjected to higher assembly and rework temperatures than those required for eutectic tin-lead assemblies. The rework of lead-free area array components involves the removal of defective component, preparation of the printed circuit board attachment pad (site redressing), solder paste replenishment or flux deposition, and component placement and reflow. This paper primarily focuses on the site redressing aspect of lead-free rework, followed by the development of rework processes for lead-free chip scale packages utilizing the knowledge gained in the site redressing studies.
Dynamics of Half-Car Models Under Stochastic Road Excitations
Authors: Wedig Walter V.
Published: May 2008

The vertical dynamics of half car models riding on irregular road surfaces is characterized by base excitations with time delays determined by the ratio of the car speed and the axle distance between front and rear wheels. Root mean square values of the stationary car vibrations are analytically investigated to derive closed-form results by means of symbolic programming software packages. In particular, the paper discusses modelling aspects when noise filtering and time-delay shifts are interchanged in order to obtain more simple calculations in the frequency domain and in the time domain, as well.
Immunogenicity of
Bacillus anthracis
Protective Antigen Domains and Efficacy of Elicited Antibody Responses Depend on Host Genetic Background
Authors: Abboud Nareen , Casadevall Arturo
Published: May 2008

Neutralizing antibodies to
Bacillus anthracis
protective antigen (PA), a component of anthrax toxin, mediate protection against anthrax. PA is antigenically complex and can elicit protective and nonprotective antibodies. Furthermore, vaccinated individuals demonstrate considerable variability in their antibody responses to PA. To explore the relationship between PA structure and antigenicity, we produced
Escherichia coli
strains expressing full-length PA (PA1-4), domains 2 to 4 (PA2-4), domain 1, (PA1), and domain 4 (PA4) and evaluated the immunogenicities and protective efficacies of the protein fractions in four mouse strains (strains A/J, BALB/c, C57BL/6, and Swiss Webster). Immunization with PA1-4 resulted in significantly higher lethal toxin-neutralizing antibody titers than immunization with any recombinant protein (rPA) fraction of PA. The magnitude and neutralizing capacity of the antibody response to full-length PA and its fragments varied depending on the mouse strain. We found no correlation between the antibody titer and the neutralizing antibody titer for A/J and Swiss Webster mice. In C57BL/6 mice, antibody titers and neutralization capacity correlated for two of four rPA domain proteins tested, while BALB/c mice displayed a similar correlation with only one rPA. By correlating the reactivity of immune sera with solvent-exposed linear peptide segments of PA, we tentatively assign the presence of four new linear B-cell epitopes in PA amino acids 121 to 150, 143 to 158, 339 to 359, and 421 to 440. We conclude that the genetic background of the host determines the relative efficacy of the antitoxin response. The results suggest that the variability observed in vaccination studies with PA-derived vaccines is a result of host heterogeneity and implies a need to develop other antigens as vaccine candidates.
PCR-Based Assay Using Occult Blood Detection Cards for Detection of Diarrheagenic
Escherichia coli
in Specimens from U.S. Travelers to Mexico with Acute Diarrhea
Authors: Grimes Kevin A. , Mohamed Jamal A. , DuPont Herbert L. , Padda Ranjit S. , Jiang Zhi-Dong , Flores Jose , Belkind-Gerson Jaime , Martinez-Sandoval Francisco G. , Okhuysen Pablo C.
Published: May 2008

Large field studies of travelers' diarrhea for multiple destinations are limited by the need to perform stool cultures on site in a timely manner. A method for the collection, transport, and storage of fecal specimens that does not require immediate processing and refrigeration and that is stable for months would be advantageous. This study was designed to determine if enterotoxigenic
Escherichia coli
(ETEC) and enteroaggregative
E. coli
(EAEC) DNA could be identified from cards that were processed for the evaluation of fecal occult blood. U.S. students traveling to Mexico during 2005 to 2007 were monitored for the occurrence of diarrheal illness. When ill, students provided a stool specimen for culture and occult blood by the standard methods. Cards then were stored at room temperature prior to DNA extraction. Fecal PCR was performed to identify ETEC and EAEC in DNA extracted from stools and from occult blood cards. Significantly more EAEC cases were identified by PCR that was performed on DNA that was extracted from cards (49%) or from frozen feces (40%) than from culture methods that used HEp-2 adherence assays (13%) (
P
< 0.001). Similarly, more ETEC cases were detected from card DNA (38%) than from fecal DNA (30%) or by culture that was followed by hybridization (10%) (
P
< 0.001). The sensitivity and specificity of the card test were 75 and 62%, respectively, compared to those for EAEC by culture and were 50 and 63%, respectively, compared to those for ETEC. DNA extracted from fecal cards that was used for the detection of occult blood is of use in identifying diarrheagenic
E. coli
.
Prevention of Drug Carryover Effects in Studies Assessing Antimycobacterial Efficacy of TMC207
Authors: Lounis Nacer , Gevers Tom , Van Den Berg Joke , Verhaeghe Tom , van Heeswijk Rolf , Andries Koen
Published: May 2008

The levels of TMC207 (R207910) that can be reached in mouse organs and the sputa of treated patients easily exceed the MIC of the compound and can therefore interfere with in vitro bacterial titrations. We studied the usefulness of protein-enriched media for the prevention of such drug carryover effects. The average MIC of
Mycobacterium tuberculosis
was determined on three different media: unsupplemented 7H11 agar (MIC = 0.03 μg/ml), 7H11 agar supplemented with 5% bovine serum albumin (BSA; MIC = 1 μg/ml), and Lowenstein-Jensen medium (MIC = 14.33 μg/ml). In a second stage of the study, the maximal noninhibitory concentrations (MNICs) of TMC207 were determined by adding TMC207 to the bacterial inoculum rather than to the culture medium. These MNICs were 0.97 μg/ml for 7H11 agar, 32.33 μg/ml for 7H11 agar with 5% BSA, and 96.33 μg/ml for Lowenstein-Jensen medium. Both protein-enriched media were able to prevent drug carryover effects, but the use of 7H11 medium supplemented with 5% BSA is preferred for practical reasons.
Severe Thrombocytopenia Associated with Acute Hepatitis E Virus Infection
Authors: Colson P. , Payraudeau E. , Leonnet C. , De Montigny S. , Villeneuve L. , Motte A. , Tamalet C.
Published: May 2008

We describe here what is, to the best of our knowledge, the third reported case of severe thrombocytopenia associated with acute hepatitis E virus infection. The patient was a 72-year-old French woman. It seems likely that the cause of the thrombocytopenia was acute hepatitis E virus infection, possibly occurring via an immune mechanism. No complications were noted, in contrast to the two previous reports.
Monitoring Early Fusion Dynamics of Human Immunodeficiency Virus Type 1 at Single-Molecule Resolution
Authors: Dobrowsky Terrence M. , Zhou Yan , Sun Sean X. , Siliciano Robert F. , Wirtz Denis
Published: May 2008

The fusion of human immunodeficiency virus type 1 (HIV-1) to host cells is a dynamic process governed by the interaction between glycoproteins on the viral envelope and the major receptor, CD4, and coreceptor on the surface of the cell. How these receptors organize at the virion-cell interface to promote a fusion-competent site is not well understood. Using single-molecule force spectroscopy, we map the tensile strengths, lifetimes, and energy barriers of individual intermolecular bonds between CCR5-tropic HIV-1 gp120 and its receptors CD4 and CCR5 or CXCR4 as a function of the interaction time with the cell. According to the Bell model, at short times of contact between cell and virion, the gp120-CD4 bond is able to withstand forces up to 35 pN and has an initial lifetime of 0.27 s and an intermolecular length of interaction of 0.34 nm. The initial bond also has an energy barrier of 6.7

k
B

T
(where

k
B

is Boltzmann's constant and
T
is absolute temperature). However, within 0.3 s, individual gp120-CD4 bonds undergo rapid destabilization accompanied by a shortened lifetime and a lowered tensile strength. This destabilization is significantly enhanced by the coreceptor CCR5, not by CXCR4 or fusion inhibitors, which suggests that it is directly related to a conformational change in the gp120-CD4 bond. These measurements highlight the instability and low tensile strength of gp120-receptor bonds, uncover a synergistic role for CCR5 in the progression of the gp120-CD4 bond, and suggest that the cell-virus adhesion complex is functionally arranged about a long-lived gp120-coreceptor bond.
Efficienttrans-Encapsidation of Hepatitis C Virus RNAs into Infectious Virus-Like Particles
Authors: Steinmann Eike , Brohm Christiane , Kallis Stephanie , Bartenschlager Ralf , Pietschmann Thomas
Published: May 2008

Recently, complete replication of hepatitis C virus (HCV) in tissue culture was established using the JFH1 isolate. To analyze determinants of HCV genome packaging and virion assembly, we developed a system that supports particle production based ontrans-packaging of subgenomic viral RNAs. Using JFH1 helper viruses, we show that subgenomic JFH1 replicons lacking the entire core to NS2 coding region are efficiently encapsidated into infectious virus-like particles. Similarly, chimeric helper viruses with heterologous structural proteinstrans-package subgenomic JFH1 replicons. Like authentic cell culture-produced HCV (HCVcc) particles, thesetrans-complemented HCV particles (HCVTCP) penetrate target cells in a CD81 receptor-dependent fashion. Since HCVTCPproduction was limited by competition between the helper and subgenomic RNA and to avoid contamination of HCVTCPstocks with helper viruses, we created HCV packaging cells. These cells encapsidate various HCV replicons with high efficiency, reaching infectivity titers up to 106tissue culture infectious doses 50 per milliliter. The produced particles display a buoyant density comparable to HCVcc particles and can be propagated in the packaging cell line but support only a single-round infection in naïve cells. Together, this work demonstrates that subgenomic HCV replicons are assembly competent, thus excludingcis-acting RNA elements in the core-to-NS2 genomic region essential for RNA packaging. The experimental system described here should be helpful to decipher the mechanisms of HCV assembly and to identify RNA elements and viral proteins involved in particle formation. Similar to other vector systems of plus-strand RNA viruses, HCVTCPmay prove valuable for gene delivery or vaccination approaches.
Differential Gene Regulation by Epstein-Barr Virus Type 1 and Type 2 EBNA2
Authors: Lucchesi Walter , Brady Gareth , Dittrich-Breiholz Oliver , Kracht Michael , Russ Rainer , Farrell Paul J.
Published: May 2008

A transfection assay with a lymphoblastoid cell line infected with Epstein-Barr virus was used to compare the abilities of type 1 and type 2 EBNA2 to sustain cell proliferation. The reduced proliferation in cells expressing type 2 EBNA2 correlated with loss of expression of some cell genes that are known to be targets of type 1 EBNA2. Microarray analysis of EBNA2 target genes identified a small number of genes that are more strongly induced by type 1 than by type 2 EBNA2, and one of these genes (CXCR7) was shown to be required for proliferation of lymphoblastoid cell lines. The Epstein-Barr virus LMP1 gene was also more strongly induced by type 1 EBNA2 than by type 2, but this effect was transient. Type 1 and type 2 EBNA2 were equally effective at arresting cell proliferation of Burkitt's lymphoma cell lines lacking Epstein-Barr virus and were also shown to cause apoptosis in these cells. The results indicate that differential gene regulation by Epstein-Barr virus type 1 and type 2 EBNA2 may be the basis for the much weaker B-cell transformation activity of type 2 Epstein-Barr virus strains compared to type 1 strains.
Rhesus Macaque
TRIM5
Alleles Have Divergent Antiretroviral Specificities
Authors: Wilson Sam J. , Webb Benjamin L. J. , Maplanka Charlotte , Newman Ruchi M. , Verschoor Ernst J. , Heeney Jonathan L. , Towers Greg J.
Published: May 2008

TRIM5α is a potent barrier to cross-species retroviral transmission, and TRIM5αs from different species have divergent antiretroviral specificities. Multiple
TRIM5
alleles circulate within rhesus macaque populations. Here we show that they too have different antiretroviral specificities, highlighting how
TRIM5
genotypes contribute to protection in an individual or a population.
A Bovine Herpesvirus Type 1 Mutant Virus Specifying a Carboxyl-Terminal Truncation of Glycoprotein E Is Defective in Anterograde Neuronal Transport in Rabbits and Calves
Authors: Liu Z. F. , Brum M. C. S. , Doster A. , Jones C. , Chowdhury S. I.
Published: May 2008

Bovine herpesvirus type 1 (BHV-1) is an important component of the bovine respiratory disease complex (BRDC) in cattle. The ability of BHV-1 to transport anterogradely from neuronal cell bodies in trigeminal ganglia (TG) to nerve ending in the noses and corneas of infected cattle following reactivation from latency plays a significant role in the pathogenesis of BRDC and maintenance of BHV-1 in the cattle population. We have constructed a BHV-1 bacterial artificial chromosome (BAC) clone by inserting an excisable BAC plasmid sequence in the long intergenic region between the glycoprotein B (gB) and UL26 genes. A BAC-excised, reconstituted BHV-1 containing only the 34-bp loxP sequence within the gB-UL26 intergenic region was highly infectious in calves, retained wild-type virulence properties, and reactivated from latency following treatment with dexamethasone. Using a two-step Red-mediated mutagenesis system inEscherichia coli, we constructed a gE cytoplasmic tail-truncated BHV-1 and a gE-rescued BHV-1. Following primary infection, the gE cytoplasmic tail-truncated virus was efficiently transported retrogradely from the nerve endings in the nose and eye to cell bodies in the TG of calves and rabbits. However, following dexamethasone-induced reactivation from latency, the gE mutant virus was not isolated from nasal and ocular sheddings. Reverse transcriptase PCR assays detected VP5 transcription in the TG of rabbits infected with gE-rescued and gE cytoplasmic tail-truncated viruses during primary infection and after dexamethasone treatment but not during latency. Therefore, the BHV-1gE cytoplasmic tail-truncated virus reactivated in the TG; however, it had defective anterograde transport from TG to nose and eye in calves and rabbits.
The Cytoplasmic Terminus of Kaposi's Sarcoma-Associated Herpesvirus Glycoprotein B Is Not Essential for Virion Egress and Infectivity
Authors: Subramanian R. , D'Auvergne O. , Kong Haixia , Kousoulas K. G.
Published: May 2008

Kaposi's sarcoma-associated herpesvirus (KSHV)-encoded glycoprotein B (gB) is an important determinant of viral infectivity and virion egress. A small interfering RNA (siRNA)-based strategy was devised to inhibit KSHV gB gene expression. Transient cotransfection of plasmids constitutively expressing gB and anti-gB siRNAs in 293 cells substantially inhibited gB mRNA levels and protein production. Similarly, transient expression of siRNAs into the primary effusion lymphoma cell line BCBL-1 caused a substantial reduction of gB transcripts and protein synthesis. TaqMan real-time PCR assays against the lytic KSHV gene ORF59 and infectivity assays on 293 cells were employed to assess the effect of inhibiting gB synthesis on virion egress from BCBL-1 cells and infectivity on 293 cells, respectively. These experiments showed that gB was essential for virion egress and infectivity. Transfection of a codon-optimized gB gene with the first 540 nucleotides altered, and therefore not recognized by anti-gB siRNAs that target the native but not the codon-optimized sequence, efficiently rescued virion egress and infectivity in BCBL-1 cells in the presence of siRNAs inhibiting wild-type gB expression. To assess the role of the cytoplasmic domain of gB in virion egress, mutant gB genes were generated specifying carboxyl terminal truncations of 25 and 58 amino acids disrupting two prominent predicted α-helical domains associated with virus-induced cell fusion. A third truncation removed the entire predicted cytoplasmic terminus of 84 amino acids, while a fourth truncation removed 110 amino acids, including the terminal most hydrophobic, intramembrane anchoring sequence. Virion egress experiments revealed that all truncated gBs facilitated virion egress from BCBL-1 cells, with the exception of the largest 110-amino-acid truncation, which removed the gB anchoring sequence. Importantly, the gB truncation that removed the entire predicted cytoplasmic domain increased virion egress, suggesting the presence of a egress regulation domain located proximal to the intramembrane sequence within the cytoplasmic domain of gB. All supernatant virions were infectious on 293 cells, indicating that the carboxyl terminus of gB is not essential for either virion egress or virus infectivity.
The Immune Response to Herpes Simplex Virus Type 1 Infection in Susceptible Mice Is a Major Cause of Central Nervous System Pathology Resulting in Fatal Encephalitis
Authors: Lundberg Patric , Ramakrishna Chandran , Brown Jeffrey , Tyszka J. Michael , Hamamura Mark , Hinton David R. , Kovats Susan , Nalcioglu Orhan , Weinberg Kenneth , Openshaw Harry , Cantin Edouard M.
Published: May 2008

This study was undertaken to investigate possible immune mechanisms in fatal herpes simplex virus type 1 (HSV-1) encephalitis (HSE) after HSV-1 corneal inoculation. Susceptible 129S6 (129) but not resistant C57BL/6 (B6) mice developed intense focal inflammatory brain stem lesions of primarily F4/80+macrophages and Gr-1+neutrophils detectable by magnetic resonance imaging as early as day 6 postinfection (p.i.). Depletion of macrophages and neutrophils significantly enhanced the survival of infected 129 mice. Immunodeficient B6 (IL-7R−/−Kitw41/w41) mice lacking adaptive cells (B6-E mice) and transplanted with 129 bone marrow showed significantly accelerated fatal HSE compared to B6-E mice transplanted with B6 marrow or control nontransplanted B6-E mice. In contrast, there was no difference in ocular viral shedding in B6-E mice transplanted with 129 or B6 bone marrow. Acyclovir treatment of 129 mice beginning on day 4 p.i. (24 h after HSV-1 first reaches the brain stem) reduced nervous system viral titers to undetectable levels but did not alter brain stem inflammation or mortality. We conclude that fatal HSE in 129 mice results from widespread damage in the brain stem caused by destructive inflammatory responses initiated early in infection by massive infiltration of innate cells.
Identification of Closely Spaced but Distinct Transcription Initiation Sites for the Murine Gammaherpesvirus 68 Latency-Associated M2 Gene
Authors: DeZalia Mark , Speck Samuel H.
Published: May 2008

Murine gammaherpesvirus 68 (MHV68) infection of mice provides a tractable small-animal system for assessing viral requirements for establishment of and reactivation from latency. The M2 gene product has no homology to any known proteins but has been shown to play a role in both the establishment of MHV68 latency and reactivation from latency. Furthermore, we have recently shown that M2 expression in primary murine B cells leads to enhanced proliferation, survival, and differentiation toward a preplasma memory B-cell phenotype (A. M. Siegel, J. H. Herskowitz, and S. H. Speck, PLoS Pathog. 4:e1000039, 2008). Previous studies have characterized the structure of the M2 transcript, but to date there has been no characterization of the M2 promoter, additional open reading frames (ORFs) in the M2 region, or identified splice acceptor and splice donor sites present in the previously characterized M2 gene transcript. Here we report (i) the identification and disruption of a novel transcript that encodes a short, previously unreported ORF (M2b) located in the intron between exon 1 and exon 2 of the M2 transcript; (ii) the identification of clustered but distinct M2 gene transcription initiation sites suggesting the presence of multiple promoters involved in regulating M2 gene transcription; (iii) the characterization in vivo of recombinant MHV68 harboring deletions within the identified M2 promoter region; and (iv) the in vivo analysis of recombinant MHV68 harboring mutations that ablate either the identified M2 splice acceptor or splice donor site. Finally, our 5′ rapid amplification of cDNA ends in conjunction with splice acceptor mutation analyses confirmed that all detected M2 gene transcripts expressed during MHV68 infection in mice splice into the M2 ORF downstream of the first AUG codon, providing strong evidence that initiation of the M2 gene product arises from the second AUG codon located at residue 8 in the M2 ORF. This initial detailed analysis of M2 gene transcription in vivo will aid future studies on regulation of M2 gene expression.
Identification of Mutated Cyclization Sequences That Permit Efficient Replication of West Nile Virus Genomes: Use in Safer Propagation of a Novel Vaccine Candidate
Authors: Suzuki Ryosuke , Fayzulin Rafik , Frolov Ilya , Mason Peter W.
Published: May 2008

Existing live-attenuated flavivirus vaccines (LAV) could be improved by reducing their potential to recombine with naturally circulating viruses in the field. Since the highly conserved cyclization sequences (CS) found in the termini of flavivirus genomes must be complementary to each other to support genome replication, we set out to identify paired mutant CS that could support the efficient replication of LAV but would be unable to support replication in recombinant viruses harboring one wild-type (WT) CS. By systematic evaluation of paired mutated CS encoded in West Nile virus (WNV) replicons, we identified variants having single and double mutations in the 5′- and 3′-CS components that could support genome replication at WT levels. Replicons containing only the double-mutated CS in the 5′ or the 3′ ends of the genome were incapable of replication, indicating that mutated CS could be useful for constructing safer LAV. Despite the identity of the central portion of the CS in all mosquito-borne flaviviruses, viruses carrying complementary the double mutations in both the 5′- and the 3′-CS were indistinguishable from WT WNV in their replication in insect and mammalian cell lines. In addition to the utility of our novel CS pair in constructing safer LAV, we demonstrated that introduction of these mutated CS into one component of a recently described two-component genome system (A. V. Shustov, P. W. Mason, and I. Frolov, J. Virol. 81:11737-11748, 2007) enabled us to engineer a safer single-cycle WNV vaccine candidate with reduced potential for recombination during its propagation.
Differences in Hepatitis C Virus (HCV)-Specific CD8 T-Cell Phenotype during Pegylated Alpha Interferon and Ribavirin Treatment Are Related to Response to Antiviral Therapy in Patients Chronically Infected with HCV
Authors: Caetano Joana , Martinho António , Paiva Artur , Pais Beatriz , Valente Cristina , Luxo Cristina
Published: May 2008

CD8 T cells play a major role in antiviral immune responses. Their importance for progression to chronic hepatitis C and response to treatment are still unclear. To address these issues, hepatitis C virus (HCV)-specific CD8 T-cell responses were monitored, at the single-cell level, using HLA class I pentamers specific for HCV core and HCV NS3 epitopes, in 23 chronically infected patients during treatment with pegylated alpha interferon and ribavirin. Patients who presented a sustained-response to therapy had stronger HCV-specific CD8 T-cell responses at all time points studied. Moreover, there were clear differences in the phenotypes of these cells during therapy: in responder patients, terminally differentiated effector cells increased more rapidly, and their frequency was always higher than in nonresponder patients. Sustained-responder patients also showed a higher frequency of HCV-specific CD8 T cells producing cytotoxic factors. Overall, a late and inefficient differentiation process of HCV-specific CD8 T cells might be associated with lack of response to treatment. A better knowledge of the mechanisms underlying this impairment may be important for the development of new therapeutic strategies to maintain, restore, or increase CD8 T-cell effectiveness in chronic HCV infection.
Genetic Analysis of the Capsid Gene of Genotype GII.2 Noroviruses
Authors: Iritani Nobuhiro , Vennema Harry , Siebenga J. Joukje , Siezen Roland J. , Renckens Bernadet , Seto Yoshiyuki , Kaida Atsushi , Koopmans Marion
Published: May 2008

Noroviruses (NoVs) are considered to be a major cause of acute nonbacterial gastroenteritis in humans. The NoV genus is genetically diverse, and genotype GII.4 has been most commonly identified worldwide in recent years. In this study we analyzed the complete capsid gene of NoV strains belonging to the less prevalent genotype GII.2. We compared a total of 36 complete capsid sequences of GII.2 sequences obtained from the GenBank (
n
= 5) and from outbreaks or sporadic cases that occurred in The Netherlands (
n
= 10) and in Osaka City, Japan (
n
= 21), between 1976 and 2005. Alignment of all capsid sequences did not show fixation of amino acid substitutions over time as an indication for genetic drift. In contrast, when strains previously recognized as recombinants were excluded from the alignment, genetic drift was observed. Substitutions were found at five informative sites (two in the P1 subdomain and three in the P2 subdomain), segregating strains into five genetic groups (1994 to 1997, 1999 to 2000, 2001 to 2003, 2004, and 2005). Only one amino acid position changed consistently between each group (position 345). Homology modeling of the GII.2 capsid protein showed that the five amino acids were located on the surface of the capsid and close to each other at the interface of two monomers. The data suggest that these changes were induced by selective pressure, driving virus evolution. Remarkably, this was observed only for nonrecombinant genomes, suggesting differences in behavior with recombinant strains.
Mechanistic Studies of a T20-Dependent Human Immunodeficiency Virus Type 1 Variant
Authors: Baldwin Chris , Berkhout Ben
Published: May 2008

We previously described a T20-dependent human immunodeficiency virus type 1 variant from a patient on T20 therapy (3). This virus carries two mutations in the gp41 domain of the envelope protein (Env) that was proposed to undergo a premature conformational switch to the 6-helix bundle structure. The T20 peptide can rescue this hyperfusogenic Env protein by preventing the premature switch and preserving an earlier prefusion conformation, thus restoring virus infectivity and replication. In this study, we set out to critically test this mechanistic explanation with alternative effectors that may control the Env switch, including other fusion inhibitors and antibodies that target gp41.
Flowering and stem growth in the celery-leaved buttercup, Ranunculus sceleratus
Authors: Samarakoon Ananda B. , Horton Roger F.
Published: May 2008

The semiaquatic celery-leaved buttercup, Ranunculus sceleratus L. flowers under 18-h photoperiods (long days (LD)) but remains vegetative under 8-h photoperiods (short days (SD)). A minimum of 20-30 LD are required for floral bud initiation which occurs prior to the elongation of the flowering stalk. The effects of LD on flowering and the subsequent growth of the flowering stem can be mimicked with gibberellin (GA3) under SD. When plants which have been induced to flower by 28 LD are submerged, the flower stalk elongates rapidly over the next 48 h. Submergence effects can be mimicked by C2H4, but not by GA3 or indole-3-acetic acid (IAA) applications.