bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of concurrent N2+ and N+ ion bombardment on the plasma-assisted deposition of carbon nitride thin film
Authors: Cao Z. X. , Oechsner H.
Published: February 2004

Nitrogen ion beam extracted from low-temperature plasmas consists of both atomic and molecular ions, and often the latter is dominant. In preparing nitrides thin films with low-energy nitrogen ions striking the growing surface, the atomic and molecular ions operate at different energy scales. While the atomic ions already provoke the subplantation growth at about 120 eV, the molecular ions that first dissociate at the surface may need roughly a doubled energy. Therefore, the energy dependence of various film characters displays a rather nontrivial behavior. We illustrate this effect with the electron-cyclotron-wave resonance plasma assisted deposition of amorphous CN film, for which the variation of the film composition, microhardness, and surface roughness with the ion energy was investigated.
Electron beam fluorescence temperature measurements of N2 in a semiconductor plasma reactor
Authors: Shimada M. , Cattolica R. , Tynan G. R.
Published: February 2004

The rotational temperature of nitrogen molecules in inductively coupled plasma (ICP) discharge has been measured using the electron beam fluorescence (EBF) technique. The neutral gas temperature is an important parameter in understanding the energy balance and neutral radical uniformity in plasma processing. The EBF technique has been used to study rarefied flows, but has not been applied previously to characterize a semiconductor plasma reactor. In this work an electron beam was integrated into an inductively coupled semiconductor plasma reactor, and was used to excite neutral nitrogen molecules from the N2X 1Σg+ state into the excited molecular ion N2+B 2Σu+ state. The electron beam excitation process maps the rotational population distribution of the original ground state into the excited molecular ion state following a dipole model with ΔK=±1. The rotational temperature, which is thought to be equivalent to the neutral gas translational temperature under plasma reactor conditions, can be determined from the rotational intensity distribution of the fluorescence spectrum from the N2+B 2Σu+ state. For the range of neutral gas temperatures found in the plasma reactor a dipole model of the excitation-emission process provides good agreement with the measured spectrum. A least-square fitting procedure comparing the model and measured emission spectra is used to determine the rotation temperature. In this article the first EBF measurements of gas heating in an ICP reactor at different gas pressures (20, 50 mTorr) and plasma source input powers (250, 500, and 1000 W) are reported.
Easy method enhancing the sensitivity of a helium mass-spectrometer leak detector
Authors: Firpo G. , Pozzo A.
Published: February 2004

Commercial He mass spectrometer leak detectors usually do not provide sufficient sensitivity to perform accurate measurements of the permeation rate of He through glass. Ultrasensitive dedicated systems have adeguate sensitivity but involve high costs and complex procedures. However, both cryogenics and photomultiplier technology routinely demand this goal. Here, we propose a novel method to increase the sensitivity of commercial devices to easily measure accurate permeation rate. We modified a commercial leak detector by reducing the pumping speed at the inlet of the rotary pump, thus increasing its sensitivity by one order of magnitude. The modified detector was used to measure the leak rate of the permeation of He through the glass walls of a photomultiplier. Further improvements made to decrease the minimum detectable signal were limited by the high ultimate pressure in the spectrometer tube.
Optical emission diagnostic of a pulsed arc discharge
Authors: Restrepo E. , Devia A.
Published: February 2004

A spectroscopic analysis of plasma properties produced in a new type of pulsed-vacuum arc system, is presented herein. The system is used to produce TiN coatings and the process is carried out by using a vacuum chamber filled with nitrogen. Inside the chamber there are two opposite electrodes (the anode being the sample and the cathode acting as the target). A bank of capacitors is charged up to 300 V. Thereafter, it is discharged between the electrodes by means of a RLC circuit producing the plasma. Some species (Ti, Ti I, N, N I, N2, N2 I, and TiN) have been identified in the plasma. Electron and excitation temperatures were calculated by means of titanium atomic lines assuming local thermodynamic equilibrium (LTE), but supposing that Texc is different to Te, using the Boltzmann plot and line-continuum ratio, obtaining approximate values of 1 eV. Electron density in the order of 1013 cm−3 was calculated using stark broadening of atomic nitrogen lines at 414.6 and 492.8 nm. Moreover, the plasma behavior as a function of the pressure was studied in this article.
Fabrication of ferroelectric PbZrxTi1−xO3 thick films and their optical waveguide properties
Authors: Hu G. J. , Hu S. H. , Meng X. J. , Wang G. S. , Zhao Q. , Sun J. L. , Chu J. H. , Dai N. , Xu L. , Liu L. Y. , Li D. X.
Published: February 2004

3 μm thick ferroelectric PbZrxTi1−xO3 films with x=0.4 and 0.5 have been fabricated on LaNiO3 coated silicon substrates by a modified sol-gel process. X-ray diffraction analysis shows that the films exhibit highly (100)-preferred orientation (α100>99%) and a single perovskite phase. Atomic force microscopy study shows that specimens possess smooth surfaces. The prism-film coupling measurement indicates that the system with the configuration of PbZr0.5Ti0.5O3/PbZr0.4Ti0.6O3/air can be used for a planar optical waveguide.
Thickness dependence of the properties of highlyc-axis textured AlN thin films
Authors: Martin F. , Muralt P. , Dubois M.-A. , Pezous A.
Published: February 2004

The influence of film thickness on the material properties of aluminum nitride (AlN) thin films deposited on Pt(111) electrodes has been investigated experimentally by means of x-ray diffraction, dielectric response, atomic force microscopy, interferometry measurement of effective d33, and residual stress measurement. The thickness was varied between 35 nm and 2 μm. Full width at mid-height of the rocking curve decreased from 2.60 to 1.14°, rms roughness increased from 3.8 to 18.6 Å, the effective d33, namely d33,f, from 2.75 to 5.15 pm/V. The permittivity εAlN was stable at 10.2, whereas the dielectric losses decreased from 1% to 0.1%. The breakdown electric field under dc voltages varied between 4.0 and 5.5 MV/cm.
Use of gas-phase ethanol to mitigate extreme UV/water oxidation of extreme UV optics
Authors: Klebanoff L. E. , Malinowski M. E. , Clift W. M. , Steinhaus C. , Grunow P.
Published: February 2004

A technique is described that uses a gas-phase species to mitigate the oxidation of a Mo/Si multilayer optic caused by either extreme UV (EUV) or electron-induced dissociation of adsorbed water vapor. It is found that introduction of ethanol (EtOH) into a water-rich gas-phase environment inhibits oxidation of the outermost Si layer of the Mo/Si EUV reflective coating. Auger electron spectroscopy, sputter Auger depth profiling, EUV reflectivity, and photocurrent measurements are presented that reveal the EUV/water- and electron/water-derived optic oxidation can be suppressed at the water partial pressures used in the tests (∼2×10−7–2×10−5 Torr). The ethanol appears to function differently in two time regimes. At early times, ethanol decomposes on the optic surface, providing reactive carbon atoms that scavenge reactive oxygen atoms before they can oxidize the outermost Si layer. At later times, the reactive carbon atoms form a thin (∼5 Å), possibly self-limited, graphitic layer that inhibits water adsorption on the optic surface.
Maximum entropy decomposition of quadrupole mass spectra
Authors: v. Toussaint U. , Dose V. , Golan A.
Published: February 2004

We present an information-theoretic method called generalized maximum entropy (GME) for decomposing mass spectra of gas mixtures from noisy measurements. In this GME approach to the noisy, underdetermined inverse problem, the joint entropies of concentration, cracking, and noise probabilities are maximized subject to the measured data. This provides a robust estimation for the unknown cracking patterns and the concentrations of the contributing molecules. The method is applied to mass spectroscopic data of hydrocarbons, and the estimates are compared with those received from a Bayesian approach. We show that the GME method is efficient and is computationally fast.
Growth and characterization of hafnium silicate films prepared by UV/ozone oxidation
Authors: Punchaipetch Prakaipetch , Pant Gaurang , Kim M. J. , Wallace Robert M. , Gnade Bruce E.
Published: February 2004

Physical and electrical properties of hafnium silicate (HfSixOy) dielectric films prepared by room-temperature UV/ozone (O3) oxidation of hafnium silicide (HfSi2) are reported. Angle-resolved x-ray photoelectron spectroscopy was used to determine chemical bonding at the film interface and within the bulk film. These films, with 12 at. % Hf composition, have a dielectric constant (κ) of ∼8–9 and exhibit a flat-band voltage shift of 60 mV. The leakage current density at VFB+1 V is 4.7×10−5 A/cm2 for a 4.7-nm-thick film (capacitance equivalent thickness=2.6 nm) and breakdown strength was >8 MV/cm.
Mitigation of accumulated electric charge by deposited fluorocarbon film during SiO2 etching
Authors: Shimmura Tadashi , Suzuki Yuya , Soda Sinnosuke , Samukawa Seiji , Koyanagi Mitsumasa , Hane Kazuhiro
Published: February 2004

SiO 2 contact-hole etching with a high-aspect ratio is a key process in fabricating ultra-large scale integrated devices. However, charge accumulation in contact holes during plasma etching causes serious problems, such as charge-build-up damage, etching-stop, and microloading effects. Therefore, understanding the mechanism behind this electric charge accumulation and controlling the plasma etching processes would be very important to achieve the next-generation semiconductor devices. We found, through our previous research, that deposited fluorocarbon film in contact holes had high electric conductivity because of ion bombardment. In this experiment, we investigated the build up of charging potential during plasma processes by in situ on-wafer monitoring to control charge accumulation in the contact holes. We developed an on-wafer monitoring device to measure the charging potential in SiO2 contact holes (aspect ratio=5.7). The dc potential of the SiO2 contact hole top and bottom surfaces were measured during plasma exposure with/without deposited fluorocarbon film in the holes. The results revealed that the sidewall deposited fluorocarbon film has high electric conductivity that may mitigate electric charge accumulation at the bottom of contact holes during SiO2 etching processes.
Measurement of the Effect of Amazon Smoke on Inhibition of Cloud Formation
Authors: Koren Ilan , Kaufman Yoram J. , Remer Lorraine A. , Martins Jose V.
Published: February 2004

Urban air pollution and smoke from fires have been modeled to reduce cloud formation by absorbing sunlight, thereby cooling the surface and heating the atmosphere. Satellite data over the Amazon region during the biomass burning season showed that scattered cumulus cloud cover was reduced from 38%in clean conditions to 0%for heavy smoke (optical depth of 1.3). This response to the smoke radiative effect reverses the regional smoke instantaneous forcing of climate from –28 watts per square meter in cloud-free conditions to +8 watts per square meter once the reduction of cloud cover is accounted for.
Control of Pancreas and Liver Gene Expression by HNF Transcription Factors
Authors: Odom Duncan T. , Zizlsperger Nora , Gordon D. Benjamin , Bell George W. , Rinaldi Nicola J. , Murray Heather L. , Volkert Tom L. , Schreiber Jörg , Rolfe P. Alexander , Gifford David K. , Fraenkel Ernest , Bell Graeme I. , Young Richard A.
Published: February 2004

The transcriptional regulatory networks that specify and maintain human tissue diversity are largely uncharted. To gain insight into this circuitry, we used chromatin immunoprecipitation combined with promoter microarrays to identify systematically the genes occupied by the transcriptional regulators HNF1α, HNF4α, and HNF6, together with RNA polymerase II, in human liver and pancreatic islets. We identified tissue-specific regulatory circuits formed by HNF1α, HNF4α, and HNF6 with other transcription factors, revealing how these factors function as master regulators of hepatocyte and islet transcription. Our results suggest how misregulation of HNF4α can contribute to type 2 diabetes.
Single-Crystal Nanorings Formed by Epitaxial Self-Coiling of Polar Nanobelts
Authors: Kong Xiang Yang , Ding Yong , Yang Rusen , Wang Zhong Lin
Published: February 2004

Freestanding single-crystal complete nanorings of zinc oxide were formed via a spontaneous self-coiling process during the growth of polar nanobelts. The nanoring appeared to be initiated by circular folding of a nanobelt, caused by long-range electrostatic interaction. Coaxial and uniradial loop-by-loop winding of the nanobelt formed a complete ring. Short-range chemical bonding among the loops resulted in a single-crystal structure. The self-coiling is likely to be driven by minimizing the energy contributed by polar charges, surface area, and elastic deformation. Zinc oxide nanorings formed by self-coiling of nanobelts may be useful for investigating polar surface–induced growth processes, fundamental physics phenomena, and nanoscale devices.
Minimal Machinery of RNA Polymerase Holoenzyme Sufficient for Promoter Melting
Authors: Young Brian A. , Gruber Tanja M. , Gross Carol A.
Published: February 2004

We determined the minimal portion of
Escherichia coli
RNA polymerase (RNAP) holoenzyme able to accomplish promoter melting, the crucial step in transcription initiation that provides RNAP access to the template strand. Upon duplex DNA binding, the N terminus of the β′ subunit (amino acids 1 to 314) and amino acids 94 to 507 of the σ subunit, together comprising less than one-fifth of RNAP holoenzyme, were able to melt an extended –10 promoter in a reaction remarkably similar to that of authentic holoenzyme. Our results support the model that capture of nontemplate bases extruded from the DNA helix underlies the melting process.
Smoking Rain Clouds over the Amazon
Authors: Andreae M. O. , Rosenfeld D. , Artaxo P. , Costa A. A. , Frank G. P. , Longo K. M. , Silva-Dias M. A. F.
Published: February 2004

Heavy smoke from forest fires in the Amazon was observed to reduce cloud droplet size and so delay the onset of precipitation from 1.5 kilometers above cloud base in pristine clouds to more than 5 kilometers in polluted clouds and more than 7 kilometers in pyro-clouds. Suppression of low-level rainout and aerosol washout allows transport of water and smoke to upper levels, where the clouds appear “smoking” as they detrain much of the pollution. Elevating the onset of precipitation allows invigoration of the updrafts, causing intense thunderstorms, large hail, and greater likelihood for overshooting cloud tops into the stratosphere. There, detrained pollutants and water vapor would have profound radiative impacts on the climate system. The invigorated storms release the latent heat higher in the atmosphere. This should substantially affect the regional and global circulation systems. Together, these processes affect the water cycle, the pollution burden of the atmosphere, and the dynamics of atmospheric circulation.
Continuing the Debate on Words and Seeds
Authors: Mithen Steven
Published: February 2004

Examining the Farming/Language Dispersal Hypothesis
. Peter Bellwood and Colin Renfrew, Eds. McDonald Institute for Archaeological Research, Cambridge, 2002. 519 pp. $85, £50. ISBN 1-902937-20-1.

Researchers in linguistics, archaeology, molecular genetics, and human ecology explore an explanation for the present-day distribution of many of the world's languages and language families.
As Our World Warmed
Authors: Straus Lawrence Guy
Published: February 2004

After the Ice
A Global Human History, 20,000-5000 BC. by Steven Mithen. Weidenfeld and Nicolson, London, 2003. 636 pp. £25. ISBN 0-297-64318-5. Forthcoming from Harvard University Press, ISBN 0-674-01570-3. Paper, Phoenix, London, 2004. £8.99. ISBN 0-75381-392-0.

Drawing on recent findings from archaeology, genetics, and the environmental sciences, the author provides a region-by-region account of the transformations of human populations in the 15 millennia following the Last Glacial Maximum.
Legionella
Effectors That Promote Nonlytic Release from Protozoa
Authors: Chen John , de Felipe Karim Suwwan , Clarke Margaret , Lu Hao , Anderson O. Roger , Segal Gil , Shuman Howard A.
Published: February 2004

Legionella pneumophila
, the bacterial agent of legionnaires' disease, replicates intracellularly within a specialized vacuole of mammalian and protozoan host cells. Little is known about the specialized vacuole except that the Icm/Dot type IVsecretion system is essential for its formation and maintenance. The
Legionella
genome database contains two open reading frames encoding polypeptides (LepA and LepB) with predicted coiled-coil regions and weak homology to SNAREs; these are delivered to host cells by an Icm/Dot–dependent mechanism. Analysis of mutant strains suggests that the Lep proteins may enable the
Legionella
to commandeer a protozoan exocytic pathway for dissemination of the pathogen.
Laser Guide Star Adaptive Optics Imaging Polarimetry of Herbig Ae/Be Stars
Authors: Perrin Marshall D. , Graham James R. , Kalas Paul , Lloyd James P. , Max Claire E. , Gavel Donald T. , Pennington Deanna M. , Gates Elinor L.
Published: February 2004

We have used laser guide star adaptive optics and a near-infrared dual-channel imaging polarimeter to observe light scattered in the circumstellar environment of Herbig Ae/Be stars on scales of 100 to 300 astronomical units. We revealed a strongly polarized, biconical nebula 10 arc seconds (6000 astronomical units) in diameter around the star LkHα 198 and also observed a polarized jet-like feature associated with the deeply embedded source LkHα 198-IR. The star LkHα 233 presents a narrow, unpolarized dark lane consistent with an optically thick circumstellar disk blocking our direct view of the star. These data show that the lower-mass T Tauri and intermediate mass Herbig Ae/Be stars share a common evolutionary sequence.
Interferon-Dependent Immunity Is Essential for Resistance to Primary Dengue Virus Infection in Mice, Whereas T- and B-Cell-Dependent Immunity Are Less Critical
Authors: Shresta Sujan , Kyle Jennifer L. , Snider Heidi M. , Basavapatna Manasa , Beatty P. Robert , Harris Eva
Published: February 2004

Dengue virus (DEN) causes dengue fever and dengue hemorrhagic fever/dengue shock syndrome, which are major public health problems worldwide. The immune factors that control DEN infection or contribute to severe disease are neither well understood nor easy to examine in humans. In this study, we used wild-type and congenic mice lacking various components of the immune system to study the immune mechanisms in the response to DEN infection. Our results demonstrate that alpha/beta interferon (IFN-α/β) and IFN-γ receptors have critical, nonoverlapping functions in resolving primary DEN infection. Furthermore, we show that IFN-α/β receptor-mediated action limits initial DEN replication in extraneural sites and controls subsequent viral spread into the central nervous system (CNS). In contrast, IFN-γ receptor-mediated responses seem to act at later stages of DEN disease by restricting viral replication in the periphery and eliminating virus from the CNS. Mice deficient in B, CD4
+
T, or CD8
+
T cells had no increased susceptibility to DEN; however, RAG mice (deficient in both B and T cells) were partially susceptible to DEN infection. In summary, (i) IFN-α/β is critical for early immune responses to DEN infection, (ii) IFN-γ-mediated immune responses are crucial for both early and late clearance of DEN infection in mice, and (iii) the IFN system plays a more important role than T- and B-cell-dependent immunity in resistance to primary DEN infection in mice.
Human Rhinovirus Type 2-Antibody Complexes Enter and Infect Cells via Fc-γ Receptor IIB1
Authors: Baravalle Günther , Brabec Marianne , Snyers Luc , Blaas Dieter , Fuchs Renate
Published: February 2004

HeLa cells were stably transfected with a cDNA clone encoding the B1 isoform of the mouse FcγRII receptor (hereafter referred to as HeLa-FcRII cells). The receptor was expressed at high level at the plasma membrane in about 90% of the cells. These cells bound and internalized mouse monoclonal virus-neutralizing antibodies 8F5 and 3B10 of the subtype immunoglobulin G2a (IgG2a) and IgG1, respectively. Binding of the minor-group human rhinovirus type 2 (HRV2) to its natural receptors, members of the low-density lipoprotein receptor family, is dependent on the presence of Ca
2+
ions. Thus, chelating Ca
2+
ions with EDTA prevented HRV2 binding, entry, and infection. However, upon complex formation of
35
S-labeled HRV2 with 8F5 or 3B10, virus was bound, internalized, and degraded in HeLa-FcRII cells. Furthermore, challenge of these cells with HRV2-8F5 or HRV2-3B10 complexes resulted in de novo synthesis of viral proteins, as shown by indirect immunofluorescence microscopy. These data demonstrate that minor-group receptors can be replaced by surrogate receptors to mediate HRV2 cell entry, delivery into endosomal compartments, and productive uncoating. Consequently, the conformational change and uncoating of HRV2 appears to be solely triggered by the low-pH (pH ≤ 5.6) environment in these compartments.
Interactions Required for Binding of Simian Virus 40 T Antigen to the Viral Origin and Molecular Modeling of Initial Assembly Events
Authors: Reese Danielle K. , Sreekumar Kodangattil R. , Bullock Peter A.
Published: February 2004

The purified T-antigen origin binding domain binds site specifically to site II, the central region of the simian virus 40 core origin. However, in the context of full-length T antigen, the origin binding domain interacts poorly with DNA molecules containing just site II. Here we investigate the contributions of additional core origin regions, termed the flanking sequences, to origin recognition and the assembly of T-antigen hexamers and double hexamers. Results from these studies indicate that in addition to site-specific binding of the T-antigen origin binding domain to site II, T-antigen assembly requires non-sequence-specific interactions between a basic finger in the helicase domain and particular flanking sequences. Related studies demonstrate that the assembly of individual hexamers is coupled to the distortions in the proximal flanking sequence. In addition, the point in the double-hexamer assembly process that is regulated by phosphorylation of threonine 124, the sole posttranslational modification required for initiation of DNA replication, was further analyzed. Finally, T-antigen structural information is used to model various stages of T-antigen assembly on the core origin and the regulation of this process.
RNA Recombination Plays a Major Role in Genomic Change during Circulation of Coxsackie B Viruses
Authors: Oberste M. Steven , Peñaranda Silvia , Pallansch Mark A.
Published: February 2004

RNA recombination has been shown to occur during circulation of enteroviruses, but most studies have focused on poliovirus. To examine the role of recombination in the evolution of the coxsackie B viruses (CVB), we determined the partial sequences of four genomic intervals for multiple clinical isolates of each of the six CVB serotypes isolated from 1970 to 1996. The regions sequenced were the 5′-nontranslated region (5′-NTR) (350 nucleotides [nt]), capsid (VP4-VP2, 416 nt, and VP1, ∼320 nt), and polymerase (3D, 491 nt). Phylogenetic trees were constructed for each genome region, using the clinical isolate sequences and those of the prototype strains of all 65 enterovirus serotypes. The partial VP1 sequences of each CVB serotype were monophyletic with respect to serotype, as were the VP4-VP2 sequences, in agreement with previously published studies. In some cases, however, incongruent tree topologies suggested that intraserotypic recombination had occurred between the sequenced portions of VP2 and VP1. Outside the capsid region, however, isolates of the same serotype were not monophyletic, indicating that recombination had occurred between the 5′-NTR and capsid, the capsid and 3D, or both. Almost all clinical isolates were recombinant relative to the prototype strain of the same serotype. All of the recombination partners appear to be members of human enterovirus species B. These results suggest that recombination is a frequent event during enterovirus evolution but that there are genetic restrictions that may influence recombinational compatibility.
The Truncated Form of Glycoprotein gp2 of Equine Herpesvirus 1 (EHV-1) Vaccine Strain KyA Is Not Functionally Equivalent to Full-Length gp2 Encoded by EHV-1 Wild-Type Strain RacL11
Authors: von Einem Jens , Wellington Janet , Whalley J. Millar , Osterrieder Kerstin , O'Callaghan Dennis J. , Osterrieder Nikolaus
Published: February 2004

Most equine herpesvirus 1 (EHV-1) strains, including the naturally occurring virulent RacL11 isolate, encode a large glycoprotein, gp2 (250 kDa), which is expressed from gene 71. Besides other alterations in the viral genome, the avirulent strain KyA harbors an in-frame deletion of 1,242 nucleotides in gene 71. To examine the contributions of gp2 variation to virus growth and virulence, mutant RacL11 and KyA viruses expressing full-length or truncated gp2 were generated. Western blot analyses demonstrated expression of a 250-kDa gp2 in cells infected with RacL11 virus or a mutant KyA virus harboring full-length gene 71, whereas a 75- to 80-kDa gp2 was detected in cells infected with KyA or mutant RacL11 virus expressing KyA gp2. The RacL11 gp2 precursor of 250 kDa in size and its truncated KyA counterpart of 80 kDa, as well as the 42-kDa carboxy-terminal gp2 subunit, were incorporated into virus particles. Absence of gp2 in RacL11 resulted in a 6-fold reduction of extracellular virus titers and a 13% reduction of plaque diameters, whereas gp2-negative KyA exhibited a 55% reduction in plaque diameter and a 51-fold decrease in extracellular virus titers. The massive growth defects of gp2-negative KyA could be restored by reinsertion of the truncated but not the full-length gp2 gene. The virulence of the generated gp2 mutant viruses was compared to the virulence of KyA and RacL11 in a murine infection model. RacL11 lacking gp2 was apathogenic for BALB/c mice, and insertion of the truncated KyA gp2 gene into RacL11 was unable to restore virulence. Similarly, replacement in the KyA genome of the truncated with the full-length RacL11 gene 71 did not result in the generation of virulent virus. From the results we conclude that full-length and truncated EHV-1 gp2 are not functionally equivalent and cannot compensate for the action of their homologues in allogeneic virus backgrounds.
Norovirus Capture with Histo-Blood Group Antigens Reveals Novel Virus-Ligand Interactions
Authors: Harrington Patrick R. , Vinjé Jan , Moe Christine L. , Baric Ralph S.
Published: February 2004

Noroviruses are genetically diverse, uncultivable, positive-sense RNA viruses and are the most common cause of epidemic acute gastroenteritis in humans in the United States. Recent studies of norovirus attachment in vitro by using recombinant virus-like particles (VLPs) suggest that various norovirus strains exhibit different patterns of attachment to ABH histo-blood group antigens, which are carbohydrate epitopes present in high concentrations on mucosal cell surfaces of the gut. However, attachment of live norovirus strains to histo-blood group antigens has not been investigated to date. Utilizing a newly designed magnetic bead-virus capture method, we characterized histo-blood group antigen attachment properties of various norovirus strains obtained from clinical stool specimens to compare the attachment properties of wild-type virus and VLPs and to further map norovirus attachment. Consistent with previous reports using VLPs, various strains of noroviruses exhibited different patterns of attachment to histo- blood group antigens. Norwalk virus bound specifically to H type 1, H type 3, and Le
b
. Two genogroup II noroviruses, one representing the Toronto genotype and the other from a novel genotype, bound specifically to Le
b
. A Desert Shield-like strain did not attach to H types 1, 2, or 3, H type 1 and 3 precursors, Le
a
, or Le
b
. Surprisingly, wild-type Snow Mountain virus (SMV) attached specifically to H type 3, which contradicted previous findings with SMV VLPs. On further investigation, we found that stool components promote this attachment, providing the first known observation that one or more components of human feces could promote and enhance norovirus attachment to histo-blood group antigens.
Infectious Salmon Anemia Virus Specifically Binds to and Hydrolyzes 4-O-Acetylated Sialic Acids
Authors: Hellebø Audny , Vilas Ulrike , Falk Knut , Vlasak Reinhard
Published: February 2004

Infectious salmon anemia virus (ISAV) is the causative agent of infections in farmed Atlantic salmon. ISAV presumably represents a new genus within the
Orthomyxoviridae
. ISAV has been shown earlier to exhibit a receptor-destroying activity, which was defined as an acetylesterase with unknown specificity. We have analyzed the substrate specificity of the ISAV esterase in detail. Purified ISAV hydrolyzed free 5-
N
-acetyl-4-
O
-acetyl neuraminic acid. In addition, the purified 9-O-acetylated sialic acid derivative was also hydrolyzed, but at lower rates. When we used a glycosidically bound substrate, ISAV was unable to hydrolyze 9-O-acetylated sialic acid, which represents the major substrate for the influenza C virus esterase. ISAV completely de-O-acetylated glycoprotein-bound 5-
N
-acetyl-4-
O
-acetyl neuraminic acid. Thus, the enzymatic activity of the hemagglutinin-esterase of ISAV is comparable to that of the sialate-4-O-esterases of murine coronaviruses and related group 2 coronaviruses. In addition, we found that ISAV specifically binds to glycoproteins containing 4-O-acetylated sialic acids. Both the ISAV esterase and recombinant rat coronavirus esterase specific for 4-O-acetylated sialic acids hydrolyzed ISAV receptors on horse and rabbit erythrocytes, indicating that this sialic acid represents a receptor determinant for ISAV.
Cholesterol Is Required for Endocytosis and Endosomal Escape of Adenovirus Type 2
Authors: Imelli Nicola , Meier Oliver , Boucke Karin , Hemmi Silvio , Greber Urs F.
Published: February 2004

The species C adenovirus type 2 (Ad2) and Ad5 bind the coxsackievirus B Ad receptor and αv integrin coreceptors and enter epithelial cells by clathrin-mediated endocytosis. This pathway is rapid and efficient. It leads to cell activation and the cholesterol-dependent formation of macropinosomes. Macropinosomes are triggered to release their contents when incoming Ad2 escapes from endosomes. Here, we show that cholesterol extraction of epithelial cells by methyl-β-cyclodextrin (mβCD) treatment reduced Ad5-mediated luciferase expression ∼4-fold. The addition of cholesterol to normal cells increased gene expression in a dose-dependent manner up to threefold, but it did not restore gene expression in mβCD-treated cells. mβCD had no effect in the presence of excess cholesterol, indicating that the inhibition of gene expression was due specifically to cholesterol depletion. Cholesterol depletion inhibited rapid Ad2 endocytosis, endosomal escape, and nuclear targeting, consistent with the notion that clathrin-dependent endocytosis of Ad2 is cholesterol dependent. In cholesterol-reduced cells, Ad2 internalized at a low rate, suggestive of an alternative, clathrin-independent, low-capacity entry pathway. While exogenous cholesterol completely restored rapid Ad2 endocytosis, macropinocytosis, and macropinosome disruption, it did not, surprisingly, restore viral escape from endosomes. Our results indicate that macropinosome disruption and endosomal escape of Ad2 are independent events in cells depleted of and then refilled with cholesterol, suggesting that viral escape from endosomes requires lipid-controlled membrane homeostasis, trafficking, or signaling.
Rapid Uncoating of Vector Genomes Is the Key toEfficient Liver Transduction with Pseudotyped Adeno-Associated VirusVectors
Authors: Thomas Clare E. , Storm Theresa A. , Huang Zan , Kay Mark A.
Published: February 2004

Transduction
of the liver with single-stranded adeno-associated virus
serotype 2 (AAV2) vectors is inefficient; less than 10% of
hepatocytes are permissive for stable transduction, and transgene
expression is characterized by a lag phase of up to 6 weeks. AAV2-based
vector genomes packaged inside AAV6 or AAV8 capsids can transduce the
liver with higher efficiency, but the molecular mechanisms underlying
this phenomenon have not been determined. We now show that the primary
barrier to transduction of the liver with vectors based on AAV2 capsids
is uncoating of vector genomes in the nucleus. The majority of AAV2
genomes persist as encapsidated single-stranded molecules within the
nucleus for as long as 6 weeks after vector administration.
Double-stranded vector genomes packaged inside AAV2 capsids are at
least 50-fold more active than single-stranded counterparts, but these
vectors also exhibit a lag phase before maximal gene expression. Vector
genomes packaged inside AAV6 or AAV8 capsids do not persist as
encapsidated molecules and are more biologically active than vector
genomes packaged inside AAV2 capsids. Our data suggest that the rate of
uncoating of vector genomes determines the ability of complementary
plus and minus single-stranded genomes to anneal together and convert
to stable, biologically active double-stranded molecular
forms.
The Amino-Terminal Region of Major Capsid Protein P3 Is Essential for Self-Assembly of Single-Shelled Core-Like Particles of
Rice Dwarf Virus
Authors: Hagiwara Kyoji , Higashi Takahiko , Miyazaki Naoyuki , Naitow Hisashi , Cheng R. Holland , Nakagawa Atsushi , Mizuno Hiroshi , Tsukihara Tomitake , Omura Toshihiro
Published: February 2004

The core protein P3 of
Rice dwarf virus
constructs asymmetric dimers, one of which is inserted by the amino-terminal region of another P3 protein. The P3 proteins with serial amino-terminal deletions, expressed in a baculovirus system, formed particles with gradually decreasing stability. The capacity for self-assembly disappeared when 52 of the amino-terminal amino acids had been deleted. These results demonstrated that insertion of the amino-terminal arm of one P3 protein into another appears to play an important role in stabilizing the core particles.
The Long Third Complementarity-Determining Region of the Heavy Chain Is Important in the Activity of the Broadly Neutralizing Anti-Human Immunodeficiency Virus Type 1 Antibody 2F5
Authors: Zwick Michael B. , Komori H. Kiyomi , Stanfield Robyn L. , Church Sarah , Wang Meng , Parren Paul W. H. I. , Kunert Renate , Katinger Hermann , Wilson Ian A. , Burton Dennis R.
Published: February 2004

The human monoclonal antibody 2F5 neutralizes primary human immunodeficiency virus type 1 (HIV-1) with rare breadth and potency. A crystal structure of a complex of 2F5 and a peptide corresponding to its core epitope on gp41, ELDKWAS, revealed that the peptide interacts with residues at the base of the unusually long (22-residue) third complementarity-determining region of the heavy chain (CDR H3) but not the apex. Here, we perform alanine-scanning mutagenesis across CDR H3 and make additional substitutions of selected residues to map the paratope of Fab 2F5. Substitution of residues from the base of the H3 loop or from CDRs H1, H2, and L3, which are proximal to the peptide, significantly diminished the affinity of Fab 2F5 for gp41 and a short peptide containing the 2F5 core motif. However, nonconservative substitutions to a phenylalanine residue at the apex of the H3 loop also markedly decreased 2F5 binding to both gp41 and the peptide, suggesting that recognition of the core epitope is crucially dependent on features at the apex of the H3 loop. Furthermore, substitution at the apex of the H3 loop had an even more pronounced effect on the neutralizing activity of 2F5 against three sensitive HIV-1. These observations present a challenge to vaccine strategies based on peptide mimics of the linear epitope.
Assessment of the Role of the Central DNA Flap in Human Immunodeficiency Virus Type 1 Replication by Using a Single-Cycle Replication System
Authors: Ao Zhujun , Yao Xiaojian , Cohen Éric A.
Published: February 2004

In this study, reverse transcriptase (RT)- and integrase (IN)-defective human immunodeficiency virus type 1 (HIV-1) was transcomplemented with Vpr-RT-IN fusion proteins to delineate
pol
sequences important for HIV-1 replication. Our results reveal that a 194-bp sequence encompassing the 3′end of the IN gene and containing the central DNA flap is necessary and sufficient for efficient HIV-1 single-cycle replication in dividing and nondividing cells. Furthermore, we show that the central DNA flap enhances HIV-1 single-round replication by five- to sevenfold, primarily by facilitating nuclear import of proviral DNA. In agreement with previous reports, our data support a functional role of the central DNA flap during the early stages of HIV-1 infection.
Characterization of a Membrane-Associated Trimeric Low-pH-Induced Form of the Class II Viral Fusion Protein E from Tick-Borne Encephalitis Virus and Its Crystallization
Authors: Stiasny Karin , Bressanelli Stéphane , Lepault Jean , Rey Felix A. , Heinz Franz X.
Published: February 2004

The interaction of a dimeric membrane anchor-free form of the envelope protein E (sE dimer) from tick-borne encephalitis virus with liposomes at acidic pH levels leads to its conversion into membrane-inserted sE trimers. Electron microscopy shows that these trimers have their long dimensions along the threefold molecular axis, which is oriented perpendicularly to the plane of the membrane, where the protein inserts via the internal fusion peptide. Liposomes containing sE at their surface display paracrystalline arrays of protein in a closely packing arrangement in which each trimer is surrounded by six others, suggesting cooperativity in the insertion process. sE trimers, solubilized with nonionic detergents, yielded three-dimensional crystals suitable for X-ray diffraction analysis.
Complete Protection from Papillomavirus Challenge after a Single Vaccination with a Vesicular Stomatitis Virus Vector Expressing High Levels of L1 Protein
Authors: Roberts Anjeanette , Reuter Jon D. , Wilson Jean H. , Baldwin Stuart , Rose John K.
Published: February 2004

We generated an attenuated, recombinant vesicular stomatitis virus (VSV) expressing high levels of the cottontail rabbit papillomavirus (CRPV) L1 protein from an upstream site in the VSV genome. Rabbits vaccinated once with this VSV-L1 recombinant produced high levels of anti-L1 antibody and were completely protected against papilloma formation after challenge with CRPV. In contrast, animals vaccinated only once with a VSV vector expressing lower levels of L1 from a downstream site in the VSV genome generated lower levels of L1 antibody and demonstrated only incomplete protection from papilloma formation after challenge. We conclude that the level of L1 protein expression is critical in generating complete immunity with a single-dose vaccine.
Flying the smoky skies: secondhand smoke exposure of flight attendants
Authors: Repace J
Published: February 2004

Objective:To assess the contribution of secondhand smoke (SHS) to aircraft cabin air pollution and flight attendants’ SHS exposure relative to the general population.

Methods:Published air quality measurements, modelling studies, and dosimetry studies were reviewed, analysed, and generalised.

Results:Flight attendants reported suffering greatly from SHS pollution on aircraft. Both government and airline sponsored studies concluded that SHS created an air pollution problem in aircraft cabins, while tobacco industry sponsored studies yielding similar data concluded that ventilation controlled SHS, and that SHS pollution levels were low. Between the time that non-smoking sections were established on US carriers in 1973, and the two hour US smoking ban in 1988, commercial aircraft ventilation rates had declined three times as fast as smoking prevalence. The aircraft cabin provided the least volume and lowest ventilation rate per smoker of any social venue, including stand up bars and smoking lounges, and afforded an abnormal respiratory environment. Personal monitors showed little difference in SHS exposures between flight attendants assigned to smoking sections and those assigned to non-smoking sections of aircraft cabins.

Conclusions:In-flight air quality measurements in ~250 aircraft, generalised by models, indicate that when smoking was permitted aloft, 95% of the harmful respirable suspended particle (RSP) air pollution in the smoking sections and 85% of that in the non-smoking sections of aircraft cabins was caused by SHS. Typical levels of SHS-RSP on aircraft violated current (PM2.5) federal air quality standards ~threefold for flight attendants, and exceeded SHS irritation thresholds by 10 to 100 times. From cotinine dosimetry, SHS exposure of typical flight attendants in aircraft cabins is estimated to have been >6-fold that of the average US worker and ~14-fold that of the average person. Thus, ventilation systems massively failed to control SHS air pollution in aircraft cabins. These results have implications for studies of the past and future health of flight attendants.
Households contaminated by environmental tobacco smoke: sources of infant exposures
Authors: Matt G E , Quintana P J E , Hovell M F , Bernert J T , Song S , Novianti N , Juarez T , Floro J , Gehrman C , Garcia M , Larson S
Published: February 2004

Objectives:To examine (1) whether dust and surfaces in households of smokers are contaminated with environmental tobacco smoke (ETS); (2) whether smoking parents can protect their infants by smoking outside and away from the infant; and (3) whether contaminated dust, surfaces, and air contribute to ETS exposure in infants.

Design:Quasi-experiment comparing three types of households with infants: (1) non-smokers who believe they have protected their children from ETS; (2) smokers who believe they have protected their children from ETS; (3) smokers who expose their children to ETS.

Setting:Homes of smokers and non-smokers.

Participants:Smoking and non-smoking mothers and their infants ⩽ 1 year.

Main outcome measures:ETS contamination as measured by nicotine in household dust, indoor air, and household surfaces. ETS exposure as measured by cotinine levels in infant urine.

Results:ETS contamination and ETS exposure were 5–7 times higher in households of smokers trying to protect their infants by smoking outdoors than in households of non-smokers. ETS contamination and exposure were 3–8 times higher in households of smokers who exposed their infants to ETS by smoking indoors than in households of smokers trying to protect their children by smoking outdoors.

Conclusions:Dust and surfaces in homes of smokers are contaminated with ETS. Infants of smokers are at risk of ETS exposure in their homes through dust, surfaces, and air. Smoking outside the home and away from the infant reduces but does not completely protect a smoker’s home from ETS contamination and a smoker’s infant from ETS exposure.
Tobacco industry efforts to present ventilation as an alternative to smoke-free environments in North America
Authors: Drope J , Bialous S A , Glantz S A
Published: February 2004

Objective:To describe how the tobacco industry developed a network of consultants to promote ventilation as a “solution” to secondhand smoke (SHS) in the USA.

Methods:Analysis of previously secret tobacco industry documents.

Results:As with its other strategies to undermine the passage of clean indoor legislation and regulations, the tobacco industry used consultants who represented themselves as independent but who were promoting the industry’s ventilation “solution” strategies under close, but generally undisclosed, industry supervision. The nature of the industry’s use of ventilation consultants evolved over time. In the 1980s, the industry used them in an effort to steer the concerns about indoor air quality away from secondhand smoke, saying SHS was an insignificant component of a much larger problem of indoor air quality and inadequate ventilation. By the 1990s, the industry and its consultants were maintaining that adequate ventilation could easily accommodate “moderate smoking”. The consultants carried the ventilation message to businesses, particularly the hospitality business, and to local and national and international regulatory and legislative bodies.

Conclusion:While the tobacco industry and its consultants have gone to considerable lengths to promote the tobacco industry’s ventilation “solution”, this strategy has had limited success in the USA, probably because, in the end, it is simpler, cheaper, and healthier to end smoking. Tobacco control advocates need to continue to educate policymakers about this fact, particularly in regions where this strategy has been more effective.
“Creative solutions”: selling cigarettes in a smoke-free world
Authors: Smith E A , Malone R E
Published: February 2004

Objective:To analyse the development and execution of the “Creative Solutions” Benson & Hedges advertising campaign to understand its social, political, and commercial implications.

Methods:Searches of the Philip Morris documents and Legacy Tobacco Documents websites for relevant materials; Lexis/Nexis searches of major news and business publications; and denotative and connotative analyses of the advertising imagery.

Results:Philip Morris developed the Creative Solutions campaign in an effort to directly confront the successes of the tobacco control movement in establishing new laws and norms that promoted clean indoor air. The campaign’s imagery attempted to help smokers and potential smokers overcome the physical and social downsides of smoking cigarettes by managing risk and resolving internal conflict. The slogans suggested a variety of ways for smokers to respond to restrictions on their habit. The campaign also featured information about the Accommodation Program, Philip Morris’s attempt to organise opposition to clean indoor air laws.

Conclusion:The campaign was a commercial failure, with little impact on sales of the brand. Philip Morris got some exposure for the Accommodation Program and its anti-regulatory position. The lack of commercial response to the ads suggests that they were unable to successfully resolve the contradictions that smokers were increasingly experiencing and confirms the power of changing social norms to counter tobacco industry tactics.