bluuebunny/crossref_metadata_2025 huggingface dataset:
Speakers in the 17th‐Century Irish Parliament*
Authors: DENNEHY COLEMAN A.
Published: January 2010

In Ireland in the 17th century before the Battle of the Boyne, there were only five parliaments held. For these parliaments there was a total of 16 different individuals who acted as Speaker or made an attempt to become Speaker in the Commons or the Lords. This article will attempt to consider the possible criteria that may have been important in assessing the suitability of the candidates and also to see how many of those 16 are found to be suitable according to these conditions. We can be assured that the vast majority of those appointed and selected were politically reliable and that other issues such as legal training and legal experience are also common among most. However, ethnicity, religion (including attitudes to others' religion), family and marriage contacts, and administrative experience show that the Speakers did not always share a common background. To a certain extent, it may be deduced that these differences may be reflective of the changing political scene in Ireland over the course of this short 17th century. The performance and attributes of those who failed to become Speaker can also be useful in a study that attempts to understand the qualifications deemed desirable in a Speaker in 17th‐century Ireland.
Strategically Managing Negotiation Linkage Dynamics
Authors: Crump Larry
Published: January 2010

Negotiation linkage (the way in which one negotiation influences the process or outcome of another) presents challenges that are complex and real. Based on field research, this qualitative study examines four linked‐bilateral trade treaty negotiations conducted by Australia, Chile, the European Union, Singapore, and the United States to establish theoretical understanding about the strategic management of negotiation linkage dynamics. Several outcomes are achieved through case analysis. This study (a) introduces “degree of linkage dynamics” (robust, moderate, or modest) as a concept and concludes that it is determined by structural and contextual factors, (b) develops a framework of linked party action, (c) establishes guidance for managing opportunistic behavior in linked negotiations, (d) builds a six‐part typology of strategic techniques that can produce tangible gains in linked negotiations, and (e) examines research opportunities to further extend negotiation linkage theory. Research methodology developed in this study serves as a model for investigating negotiation linkage dynamics.
The Effects of Achievement Motivational Goals and of Debriefing on the Transfer of Skills in Integrative Negotiations
Authors: Bereby‐Meyer Yoella , Moran Simone , Sattler Liat
Published: January 2010

In this study, we examine the effects of achievement motivational goals (Learning vs. Performance) and of debriefing on the ability of trainees to acquire integrative (i.e., value‐creating) negotiation skills and to effectively transfer these skills across situations. Participants in four between‐subject conditions: 2 motivational goal conditions (learning/performance goals) × 2 debriefing conditions (with/without debriefing) first gained experience by engaging in repeated negotiations within an unchanging market simulation, and then proceeded to negotiate a more complex integrative negotiation task. A fifth control condition in which participants performed the second task only was also included. Experience accompanied by subsequent debriefing was found to be more effective for learning than experience without debriefing; it enhanced both integrative performance in the transfer task as well as conceptual understanding. Contrary to our expectations, participants who were assigned learning goals did not show an advantage over participants who were assigned performance goals.
Modifying the composition of hydrogen-terminated silicon nanoparticles synthesized in a nonthermal rf plasma
Authors: Holm Jason , Roberts Jeffrey T.
Published: January 2010

Hydrogen-terminated silicon nanoparticles were synthesized over a broad range of conditions in a continuous flow, nonthermal rf plasma. The effects of three operating parameters—rf power, reactor pressure, and hydrogen flow rate—were examined in terms of their effects on particle crystallinity, size, and surface composition. Silicon-hydrogen composition was characterized in situ by Fourier transform infrared spectrometry, and particle structural morphology was examined with a transmission electron microscope. Amorphous or crystalline particles could be synthesized by appropriately adjusting the operating parameters. Over the majority of settings examined, the minimum power required to produce discrete crystalline particles was ∼20–30W. Depending on the parameter settings, particles also exhibited hydrogen coverage ranging from predominantly monohydride (SiH) functional groups to more complex compositions of higher hydrides. Particles with the highest proportion of surface SiH bonds were consistently synthesized in the smallest diameter (4mm i.d.) tube reactor.
Coexistence of epitaxial Ta(111) and Ta(110) oriented magnetron sputtered thin film on c-cut sapphire
Authors: Gnanarajan S. , Lam S. K. H. , Bendavid A.
Published: January 2010

Epitaxial Ta films were grown on c-cut and r-cut sapphire substrates using magnetron sputtering. X-ray diffraction measurements indicate that the Ta films grown on c-cut sapphire substrates have two different crystal orientations. Low vacuum base pressure produced films with the (111) plane parallel to the basal plane after an initial (110) plane growth phase, whereas high base pressure resulted in the growth of (110) plane parallel to the basal plane with three azimuthally oriented domains coexisting. The superconducting critical temperatures Tc of the films grown on c-cut substrates were found to be slightly higher than those for the films grown on r-cut substrates. The x-ray photoelectron spectroscopic data show oxygen content in the high vacuum base pressure deposited films and a difference in the valance band with different Ta surface planes.
Oxygen plasma damage to blanket and patterned ultralow-κ surfaces
Authors: Bao J. , Shi H. , Huang H. , Ho P. S. , McSwiney M. L. , Goodner M. D. , Moinpour M. , Kloster G. M.
Published: January 2010

Oxygen plasma damage to blanket and patterned ultralow-κ (ULK) dielectric surfaces was investigated by examining the effect of plasma species and dielectric materials. Blanket ULK films and patterned structures were treated by O2 plasma in a remote plasma chamber where the ions and radicals from the plasma source can be separately controlled to study their respective roles in the damage process. The plasma damage was characterized by angle resolved x-ray photoelectron spectroscopy, x-ray reflectivity, and Fourier transform infrared spectroscopy. Studies of the angle dependence of oxygen plasma damage to blanket ULK films indicated that damage by ions was anisotropic while that by radicals was isotropic. Ions were found to play an important role in assisting carbon depletion by oxygen radicals on the blanket film surface. More plasma damage was observed with increasing porosity in ultralow-κ films. Probable reaction paths were proposed by analyzing the reaction by-products. Plasma damage to the sidewall of low-κ trenches was examined by electron energy loss (EELS) analysis. The depletion depth of carbon was found to be related to the penetration of radical species into the porous dielectric and the distribution at the sidewall and trench bottom was affected by the trench pattern geometry, i.e., the aspect ratio, which can be correlated with the electron potential distribution and subsequent trajectory of ions. Vapor silylation was applied for dielectric recovery of trench structure and the result was examined by EELS. The trimethylchlorosilane was found to be effective for recovery of the sidewall carbon loss. The recovery was better for loss induced by radical O2 than by hybrid O2 and the difference was attributed to the surface densification by ions limiting the mass transport of vapor chemicals.
Rapid thermal annealing of ZnO thin films grown at room temperature
Authors: Jang Young Rae , Yoo Keon-Ho , Park Seung Min
Published: January 2010

The authors successfully obtained high quality ZnO thin films by growing them at room temperature (RT) and postannealing by rapid thermal annealing (RTA). The thin films were grown by pulsed laser deposition on Si (100) substrates at RT, and RTA was performed under various temperatures and ambient conditions. Based on the UV emission to visible emission ratio in RT photoluminescence (PL) spectra, the optimum film was obtained at annealing temperature ∼700 °C in an ambient of Ar, N2, or O2 at 0.1 Torr, while the optimum annealing temperature was above 1100 °C in the air ambient at atmospheric pressure. The morphology and structure of the films in different RTA conditions were investigated by using field emission scanning electron microscopy and grazing incidence x-ray diffraction, and were discussed in conjunction with the PL data.
Comparison of compressive and tensile relaxed composition-graded GaAsP and (Al)InGaP substrates
Authors: Mori M. J. , Boles S. T. , Fitzgerald E. A.
Published: January 2010

The authors present a comparison of metal organic chemical vapor deposition grown compositionally graded metamorphic buffers, which enable virtual substrates with very high quality crystal lattices with lattice constants from 5.45 to 5.65 Å (threading dislocation density, ρt, around 104 cm−2). The structures, grown on GaP or GaAs, consist of graded In-fraction InGaP and AlInGaP or graded P-fraction GaAsP. They show that surface roughness and locally strained regions of phase separation (branch defects) limit misfit dislocation glide velocity and escalate threading dislocation density. High surface roughness and branch defects in (Al)InGaP lead to the lowest quality virtual substrates we observed, with ρt of around 3×106 cm−2. In contrast, graded mixed-anion films of GaAsP avoid branch defects and minimize surface roughness, giving superior defect densities, as low as 104 cm−2 at useful lattice constants halfway between that of Si and Ge. Tensile graded GaAs1−zPz layers yield the smoothest films (0.78 nm rms in a 5 μm scan) with the lowest defect densities but are subject to cracking when graded beyond z=0.5 with a graded layer thickness of <10 μm. Compressive graded GaAsP yields excellent ρt values (3.3×105 cm−2) with very thin buffers (1.3 μm). The accelerated grade rate of the compressive buffers increases crosshatch roughness, which along with the higher defect density of GaP substrates, accounts for the higher defect density compared to tensile GaAsP on GaAs substrates.
G Protein Subunit Gα
13
Binds to Integrin α
IIb
β
3
and Mediates Integrin “Outside-In” Signaling
Authors: Gong Haixia , Shen Bo , Flevaris Panagiotis , Chow Christina , Lam Stephen C.-T. , Voyno-Yasenetskaya Tatyana A. , Kozasa Tohru , Du Xiaoping
Published: January 2010

Integrin G Protein

Adhesion molecules, known as integrins, are found on the surface of cells. When integrins adhere to components of the extracellular matrix, they act as receptors and initiate signaling events within the cell.

Gong
et al.

(p.
340
) show that they do so in part by partnering with a signal-transducing protein called Gα
13
. Such α subunits of heterotrimeric guanine nucleotide-binding proteins are well known for transducing signals from the large class of G protein–coupled receptors, but were not known to work with integrins. Gα
13
appears to interact directly with the integrin α
IIb
β
3
and to transmit signals that regulate cell spreading.
Large-Scale Controls of Methanogenesis Inferred from Methane and Gravity Spaceborne Data
Authors: Bloom A. Anthony , Palmer Paul I. , Fraser Annemarie , Reay David S. , Frankenberg Christian
Published: January 2010

Measuring Methanogenesis

After carbon dioxide, methane is the second most important greenhouse gas, and an important species in terms of its role in atmospheric chemistry. The sources and sinks of methane, particularly the natural ones, are too poorly quantified, however, even to explain why the decades-long, steady increase of its concentration in the atmosphere was interrupted between 1999 and 2006.

Bloom
et al.

(p.
322
) use a combination of satellite data, which indicate water table depth and surface temperature, and atmospheric methane concentrations to determine the location and strength of methane emissions from wetlands, the largest natural global source. The constraints placed on these sources should help to improve predictions of how climate change will affect wet-land emissions of methane.
Hungry Codons Promote Frameshifting in Human Mitochondrial Ribosomes
Authors: Temperley Richard , Richter Ricarda , Dennerlein Sven , Lightowlers Robert N. , Chrzanowska-Lightowlers Zofia M.
Published: January 2010

During translation of mitochondrial genes, shifting the ribosome reading frame avoids unconventional arginine codons.
Reforming Off-Label Promotion to Enhance Orphan Disease Treatment
Authors: Liang Bryan A. , Mackey Tim
Published: January 2010

Allowing off-label promotion by drug companies may improve access to key treatments for orphan disease patients.
Lower Predation Risk for Migratory Birds at High Latitudes
Authors: McKinnon L. , Smith P. A. , Nol E. , Martin J. L. , Doyle F. I. , Abraham K. F. , Gilchrist H. G. , Morrison R. I. G. , Bêty J.
Published: January 2010

Predator Avoidance Strategy

Selective pressures influencing bird migration can include availability of food, pressure from parasites and pathogens, and predation risk. The importance of the last of these is revealed byMcKinnonet al.(p.326; see the Perspective byGilg and Yoccoz), who present an experimental analysis of the benefits of long-distance migration for reproduction in arctic-nesting birds. Measurements of a controlled effect of predation risk along a 3350-kilometer north-south gradient across arctic Canada provides evidence that the risk of nest predation decreases with latitude. Thus, birds migrating further north may acquire reproductive benefits in the form of reduced predation risk.
Nanoporous Gold Catalysts for Selective Gas-Phase Oxidative Coupling of Methanol at Low Temperature
Authors: Wittstock A. , Zielasek V. , Biener J. , Friend C. M. , Bäumer M.
Published: January 2010

Methanol Coupling Catalyzed with Gold

Gold surfaces can be effective catalysts for partial oxidation reactions, in part because lower interaction strengths of molecules absorbed on gold allow products to desorb before further unwanted oxidations occur. One challenge in these reactions is the low rate of formation of reactive atomic surface oxygen.

Wittstock
et al.

(p.
319
; see the Perspective by

Christensen and Nørskov

) created high–surface area gold catalysts by leaching silver from gold-silver alloys. This material proved to be an effective catalyst for partial oxidative coupling of methanol, yielding methyl formate. Residual silver appears to play a key role in activating the dissociation of molecular oxygen.
How the Noninflammasome NLRs Function in the Innate Immune System
Authors: Ting Jenny P. Y. , Duncan Joseph A. , Lei Yu
Published: January 2010

NLR (nucleotide-binding domain, leucine-rich repeat–containing) proteins have rapidly emerged as central regulators of immunity and inflammation with demonstrated relevance to human diseases. Much attention has focused on the ability of several NLRs to activate the inflammasome complex and drive proteolytic processing of inflammatory cytokines; however, NLRs also regulate important inflammasome-independent functions in the immune system. We discuss several of these functions, including the regulation of canonical and noncanonical NF-κB activation, mitogen-activated protein kinase activation, cytokine and chemokine production, antimicrobial reactive oxygen species production, type I interferon production, and ribonuclease L activity. We also explore the mechanistic basis of these functions and describe current challenges in the field.
Explaining Bird Migration
Authors: Gilg Olivier , Yoccoz Nigel G.
Published: January 2010

Predation pressure falls with increasing latitude, helping to explain why many birds migrate as far north as the high Arctic.
RIGorous Detection: Exposing Virus Through RNA Sensing
Authors: Rehwinkel Jan , Reis e Sousa Caetano
Published: January 2010

Virus infection in mammals elicits a variety of defense responses that are initiated by signals from virus-sensing receptors expressed by the host. These receptors include the ubiquitously expressed RIG-I–like receptor (RLR) family of RNA helicases. RLRs are cytoplasmic proteins that act in cell-intrinsic antiviral defense by recognizing RNAs indicative of virus presence. Here, we highlight recent progress in understanding how RLRs discriminate between the RNA content of healthy versus virus-infected cells, functioning as accurate sensors of virus invasion.
Icons of Early Conservation Biology
Nature's Ghosts
Confronting Extinction from the Age of Jefferson to the Age of Ecology

by Mark V. Barrow Jr.

University of Chicago Press, Chicago, 2009. 509 pp. $35, £24. ISBN 9780226038148.
Authors: Farmer Jared
Published: January 2010

Barrow's history of extinction in the United States highlights the naturalists who created the concept of endangered species.
CO
2
mmon Sense
Authors: Frommer Wolf B.
Published: January 2010

Animals and plants use the same enzyme to detect carbon dioxide.
Paul A. Samuelson (1915–2009)
Authors: Solow Robert M.
Published: January 2010

A major figure in international economics laid the foundation for modern economics and transformed the atmosphere of economic study.
An Indefatigable Debate Over Chronic Fatigue Syndrome
Authors: Kean Sam
Published: January 2010

The search for the cause of chronic fatigue syndrome, which just months ago seemed to be gaining traction, now seems likely to descend into the same confusion and acrimony that characterized it for years, as a supposed viral link to CFS published just last autumn might be unraveling.
Neandertal Jewelry Shows Their Symbolic Smarts
Authors: Balter Michael
Published: January 2010

A handful of marine mollusk shells, possibly used as necklaces and paint cups, shows that Neandertals expressed themselves symbolically, say the authors of a paper published online this week in the
Proceedings of the National Academy of Sciences
.
NIST Grants Help Schools Build for Tomorrow's Research
Authors: Mervis Jeffrey
Published: January 2010

Last week, the National Institute of Standards and Technology (NIST) announced the 12 winners in the second and final round of a $180 million competitive construction grants program.
Catalyst Offers New Hope for Capturing CO
2
on the Cheap
Authors: Service Robert F.
Published: January 2010

On page
313
of this week's issue of
Science
, researchers in the Netherlands report a new copper-based catalyst that can capture CO
2
, convert it to a different form, and then release it with a small fraction of the energy other techniques require.
Oldest Galaxies Show Stars Came Together in a Hurry
Authors: Bhattacharjee Yudhijit
Published: January 2010

Astronomers announced at the American Astronomical Society meeting that they have spotted five galaxies that date back to a mere 600 million years after the big bang—the earliest galaxies found so far by 200 million years.
Inventory Asks: Where Is All the
Non
-Dark Matter Hiding?
Authors: Bhattacharjee Yudhijit
Published: January 2010

Only 17% of the matter in the universe is detectable "baryonic matter," the atoms and ions that make up stars, planets, dust, and gas. Astronomers announced at the American Astronomical Society meeting that this proportion varies by scale: The largest galaxies have the highest percentage of baryonic matter, although not quite 17%; whereas the smallest galaxies have less than 1%.
The Secret Lives of Ocean Fish
Authors: Kean Sam
Published: January 2010

Many scientists argue that ocean fish such as cod segregate themselves into distinct environments, as salmon do—and thrive or struggle for the same reasons.
Questions Abound in Q-Fever Explosion in the Netherlands
Authors: Enserink Martin
Published: January 2010

A burgeoning goat-farm sector appears to be behind the worst outbreak ever recorded in humans of a rare zoonosis that until now has been seen primarily as a rare occupational disease for farmers, veterinarians, and slaughterhouse workers and as a potential—if not very deadly—bioterror agent.
Humans, Animals—It's One Health. Or Is It?
Authors: Enserink Martin
Published: January 2010

A "holistic approach" and "synergism" working for the health of all species are the buzzwords of the One Health movement, which aims to bring veterinary and human health closer together. But the Dutch Q-fever outbreak provides a vivid example of how those two worlds often don't get along, especially when the stakes are different for each.
Integrin G Protein
Authors: Ray L. Bryan
Published: January 2010

Cell adhesion mediated by integrins is coupled to intracellular signaling by direct binding to G protein.
The Smell of Aggression
Authors: Foley John F.
Published: January 2010

The pheromone cVA promotes aggression in
Drosophila
through olfactory sensory neurons that express the receptor Or67d.
Behavioral Fingerprinting
Authors: VanHook Annalisa M.
Published: January 2010

A large-scale screen of neuroactive compounds identifies pathways that may play a role in regulating rest-wake activity.
Generator of Stomata
Authors: Wong Wei
Published: January 2010

A secreted factor promotes the development of stomata in leaf epidermis.
Use of Protein AG in an Enzyme-Linked Immunosorbent Assay for Serodiagnosis ofToxoplasma gondiiInfection in Four Species of Animals
Authors: Zhang Donglin , Wang Zhengsong , Fang Rui , Nie Hao , Feng Huihui , Zhou Yanqin , Zhao Junlong
Published: January 2010

An enzyme-linked immunosorbent assay (ELISA) employing protein AG (AG-ELISA) as a conjugate was developed to detect anti-Toxoplasma gondiiantibodies (Ab) in experimentally infected pigs and naturally infected pigs, goats, dogs, and cats. The results indicate that AG-ELISA can be a useful method for serological diagnosis ofT. gondiiinfection in these four species of animals.
Characterization of Host Responses against a Recombinant Fowlpox Virus-Vectored Vaccine Expressing the Hemagglutinin Antigen of an Avian Influenza Virus
Authors: Hghihghi Hamid R. , Read Leah R. , Mohammadi Hakimeh , Pei Yanlong , Ursprung Claudia , Nagy Éva , Behboudi Shahriar , Haeryfar S. M. Mansour , Sharif Shayan
Published: January 2010

There currently are commercial fowlpox virus (FPV)-vectored vaccines for use in chickens, including TROVAC-AIV H5, which expresses the hemagglutinin (HA) antigen of an avian influenza virus and can confer immunity against avian influenza in chickens. Despite the use of recombinant FPV (rFPV) for vaccine delivery, very little is known about the immune responses generated by these viruses in chickens. The present study was designed to investigate host responses to rFPVin vivoandin vitro. In cultured cells infected with TROVAC-AIV H5, there was an early increase in the expression of type I interferons (IFN), Toll-like receptors 3 and 7 (TLR3 and TLR7, respectively), TRIF, and MyD88, which was followed by a decrease in the expression of these genes at later time points. There also was an increase in the expression of interleukin-1β (IL-1β), IL-8, and beta-defensin genes at early time points postinfection. In chickens immunized with TROVAC-AIV H5, there was higher expression of IFN-γ and IL-10 at day 5 postvaccination in spleen of vaccinated birds than in that of control birds. We further investigated the ability of the vaccine to induce immune responses against the HA antigen and discovered that there was a cell-mediated response elicited in vaccinated chickens against this antigen. The findings of this study demonstrate that FPV-vectored vaccines can elicit a repertoire of responses marked by the early expression of TLRs, type I interferons, and proinflammatory cytokines, as well as cytokines associated with adaptive immune responses. This study provides a platform for designing future generations of rFPV-vectored vaccines.
Antibodies against Immunodominant Antigens of
Mycobacterium tuberculosis
in Subjects with Suspected Tuberculosis in the United States Compared by HIV Status
Authors: Achkar Jacqueline M. , Jenny-Avital Elisabeth , Yu Xian , Burger Susanne , Leibert Eric , Bilder Patrick W. , Almo Steven C. , Casadevall Arturo , Laal Suman
Published: January 2010

The immunodominance of
Mycobacterium tuberculosis
proteins malate synthase (MS) and MPT51 has been demonstrated in case-control studies with patients from countries in which tuberculosis (TB) is endemic. The value of these antigens for the serodiagnosis of TB now is evaluated in a cross-sectional study of pulmonary TB suspects in the United States diagnosed to have TB, HIV-associated TB, or other respiratory diseases (ORD). Serum antibody reactivity to recombinant purified MS and MPT51 was determined by enzyme-linked immunosorbent assays (ELISAs) of samples from TB suspects and well-characterized control groups. TB suspects were diagnosed with TB (
n
= 87; 49% sputum microscopy negative, 20% HIV
+
) or ORD (
n
= 63; 58% HIV
+
). Antibody reactivity to MS and MPT51 was significantly higher in U.S. HIV
+
/TB samples than in HIV
−
/TB samples (
P
< 0.001), and it was significantly higher in both TB groups than in control groups with latent TB infection (
P
< 0.001). Antibody reactivity to both antigens was higher in U.S. HIV
+
/TB samples than in HIV
+
/ORD samples (
P
= 0.052 for MS,
P
= 0.001 for MPT51) but not significantly different between HIV
−
/TB and HIV
−
/ORD. Among U.S. HIV
+
TB suspects, a positive anti-MPT51 antibody response was strongly and significantly associated with TB (odds ratio, 11.0; 95% confidence interval, 2.3 to 51.2;
P
= 0.002). These findings have implications for the adjunctive use of TB serodiagnosis with these antigens in HIV
+
subjects.
Phenotypic and Molecular Characterization of
Solobacterium moorei
Isolates from Patients with Wound Infection
Authors: Zheng Guili , Summanen Paula H. , Talan David , Bennion Robert , Rowlinson Marie-Claire , Finegold Sydney M.
Published: January 2010

Though seldom reported,
Solobacterium moorei
, which was first described in 2000, has been identified in specimens from patients with root canals, periradicular lesions, periodontal disease, dentoalveolar abscesses, bacteremia, septic thrombophlebitis, and halitosis. In the present study, we describe 9 cases of mixed wound infection, from a pool of 400 surgical wound infections that we have studied, in which
S. moorei
was isolated or found in a clone library. All isolates of
S. moorei
were identified by 16S rRNA gene sequence analysis, and then six were examined for their physiological and biochemical characteristics and for antimicrobial susceptibility. The results of the present study indicate that
Solobacterium moorei
may be a significant component in some mixed surgical wound infections and that surgical management and antimicrobial therapy may be indicated when these bacteria are identified in significant situations.
Development of a Reverse Transcription-Loop-Mediated Isothermal Amplification Assay for Detection of Pandemic (H1N1) 2009 Virus as a Novel Molecular Method for Diagnosis of Pandemic Influenza in Resource-Limited Settings
Authors: Kubo Toru , Agoh Masanobu , Mai Le Q. , Fukushima Kiyoyasu , Nishimura Hidekazu , Yamaguchi Akinori , Hirano Manabu , Yoshikawa Akira , Hasebe Futoshi , Kohno Shigeru , Morita Kouichi
Published: January 2010

This paper reports on the development of a one-step, real-time reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay targeting the hemagglutinin (HA) gene for the rapid molecular-based detection of pandemic (H1N1) 2009 virus. The detection limit of the pandemic (H1N1) 2009 virus HA-specific RT-LAMP assay was same as that of the currently used real-time reverse transcription-PCR method. The assay detected the pandemic (H1N1) 2009 virus HA gene in 136 RNA samples extracted from nasopharyngeal swab specimens from Japanese and Vietnamese patients. No cross-reactive amplification with the RNA of other seasonal influenza viruses was observed, and the detection of specific viral genome targets in clinical specimens was achieved in less than 40 min. The sensitivity and specificity of the pandemic (H1N1) 2009 virus HA-specific RT-LAMP assay obtained in this study were 97.8% and 100%, respectively. Use of the (H1N1) 2009 virus HA-specific RT-LAMP assay will enable the faster and easier diagnosis of pandemic (H1N1) 2009 virus infection, especially in resource-limited situations in developing countries.
Fatal Infection Caused by
Cupriavidus gilardii
in a Child with Aplastic Anemia
Authors: Karafin Matthew , Romagnoli Mark , Fink Doran L. , Howard Tracy , Rau Rachel , Milstone Aaron M. , Carroll Karen C.
Published: January 2010

Cupriavidus gilardii
is a Gram-negative bacterium that has rarely been associated with human infections. We report a fatal case of sepsis caused by
C. gilardii
in a previously healthy 12-year-old female.
Coexistence of Multiple Multilocus Variable-Number Tandem-Repeat Analysis Subtypes of
Clostridium difficile
PCR Ribotype 027 Strains within Fecal Specimens
Authors: Tanner Hannah E. , Hardy Katherine J. , Hawkey Peter M.
Published: January 2010

We investigated whether multilocus variable-number tandem-repeat analysis (MLVA) typing could identify different subtypes of
Clostridium difficile
ribotype 027 within the same feces specimen. Five of 39 specimens yielded at least one isolate with an MLVA profile different (more than five summed tandem repeat differences) from that of other isolates in the same specimen, thereby potentially obscuring epidemiological links between
C. difficile
infection cases.
16S rRNA Gene Sequencing in Routine Identification of Anaerobic Bacteria Isolated from Blood Cultures
Authors: Justesen Ulrik Stenz , Skov Marianne Nielsine , Knudsen Elisa , Holt Hanne Marie , Søgaard Per , Justesen Tage
Published: January 2010

A comparison between conventional identification and 16S rRNA gene sequencing of anaerobic bacteria isolated from blood cultures in a routine setting was performed (n= 127). With sequencing, 89% were identified to the species level, versus 52% with conventional identification. The times for identification were 1.5 days and 2.8 days, respectively.
Identification of Novel Diagnostic Serum Biomarkers for Chagas' Disease in Asymptomatic Subjects by Mass Spectrometric Profiling
Authors: Ndao Momar , Spithill Terry W. , Caffrey Rebecca , Li Hongshan , Podust Vladimir N. , Perichon Regis , Santamaria Cynthia , Ache Alberto , Duncan Mark , Powell Malcolm R. , Ward Brian J.
Published: January 2010

More than 10 million people are thought to be infected withTrypanosoma cruzi, primarily in the Americas. The clinical manifestations of Chagas' disease (CD) are variable, but most subjects remain asymptomatic for decades. Only 15 to 30% eventually develop terminal complications. All current diagnostic tests have limitations. New approaches are needed for blood bank screening as well as for improved diagnosis and prognosis. Sera from subjects with asymptomatic CD (n= 131) were compared to those from uninfected controls (n= 164) and subjects with other parasitic diseases (n= 140), using protein array mass spectrometry. To identify biomarkers associated with CD, sera were fractionated by anion-exchange chromatography and bound to two commercial ProteinChip array chemistries: WCX2 and IMAC3. Multiple candidate biomarkers were found in CD sera (3 to 75.4 kDa). Algorithms employing 3 to 5 of these biomarkers achieved up to 100% sensitivity and 98% specificity for CD. The biomarkers most useful for diagnosis were identified and validated. These included MIP1 alpha, C3a anaphylatoxin, and unusually truncated forms of fibronectin, apolipoprotein A1 (ApoA1), and C3. An antipeptide antiserum against the 28.9-kDa C terminus of the fibronectin fragment achieved good specificity (90%) for CD in a Western blot format. We identified full-length ApoA1 (28.1 kDa), the major structural and functional protein component of high-density lipoprotein (HDL), as an important negative biomarker for CD, and relatively little full-length ApoA1 was detected in CD sera. This work provides proof of principle that both platform-dependent (i.e., mass spectrometry-based) and platform-independent (i.e., Western blot) tests can be generated using high-throughput mass profiling.
Development of an Immunochromatographic Assay Specifically Detecting Pandemic H1N1 (2009) Influenza Virus
Authors: Miyoshi-Akiyama Tohru , Narahara Kenji , Mori Sunao , Kitajima Hirotake , Kase Tetsuo , Morikawa Saeko , Kirikae Teruo
Published: January 2010

The pandemic caused by a new type of influenza virus, pandemic H1N1 (2009) influenza virus A (AH1pdm), has had a major worldwide impact. Since hemagglutinin (HA) genes are among the most specific genes in the influenza virus genome, AH1pdm can be definitively diagnosed by viral gene analysis targeting the HA genes. This type of analysis, however, cannot be easily performed in clinical settings. While commercially available rapid diagnosis kits (RDKs) based on immunochromatography can be used to detect nucleoproteins (NPs) of influenza A and B viruses in clinical samples, there are no such kits that are specific for AH1pdm. We show here that an RDK using a combination of monoclonal antibodies against NP can be used to specifically detect AH1pdm. The RDK recognized AH1pdm virus isolates but did not recognize seasonal H1N1 and H3N2 and influenza B viruses, indicating that the specificity of the RDK is 100%. A parallel comparison of RDK with a commercial influenza A/B virus kit revealed that both types of kits had equal sensitivities in detecting their respective viruses. Preliminary evaluation of clinical samples from 5 individuals with PCR-confirmed human AH1pdm infection showed that the RDK was positive for all samples, with the same detection intensity as that of a commercial influenza A/B virus kit. This RDK, together with a new vaccine and the stockpiling of anti-influenza drugs, will make aggressive measures to contain AH1pdm infections possible.
Preexisting Infection with Human T-Cell Lymphotropic Virus Type 2 neither Exacerbates nor Attenuates Simian Immunodeficiency Virus SIV
mac251
Infection in Macaques
Authors: Gordon Shari N. , Weissman Anna R. , Cecchinato Valentina , Fenizia Claudio , Ma Zhong-Min , Lee Tzong-Hae , Zaffiri Lorenzo , Andresen Vibeke , Parks Robyn Washington , Jones Kathryn S. , Heraud Jean Michel , Ferrari Maria Grazia , Chung Hye Kyung , Venzon David , Mahieux Renaud , Murphy Edward L. , Jacobson Steven , Miller Christopher J. , Ruscetti Francis W. , Franchini Genoveffa
Published: January 2010

Coinfection with human T-cell lymphotropic virus type 2 (HTLV-2) and human immunodeficiency virus type 1 (HIV-1) has been reported to have either a slowed disease course or to have no effect on progression to AIDS. In this study, we generated a coinfection animal model and investigated whether HTLV-2 could persistently infect macaques, induce a T-cell response, and impact simian immunodeficiency virus SIV
mac251
-induced disease. We found that inoculation of irradiated HTLV-2-infected T cells into Indian rhesus macaques elicited humoral and T-cell responses to HTLV-2 antigens at both systemic and mucosal sites. Low levels of HTLV-2 provirus DNA were detected in the blood, lymphoid tissues, and gastrointestinal tracts of infected animals. Exposure of HTLV-2-infected or naïve macaques to SIV
mac251
demonstrated comparable levels of SIV
mac251
viral replication, similar rates of mucosal and peripheral CD4
+
T-cell loss, and increased T-cell proliferation. Additionally, neither the magnitude nor the functional capacity of the SIV-specific T-cell-mediated immune response was different in HTLV-2/SIV
mac251
coinfected animals versus SIV
mac251
singly infected controls. Thus, HTLV-2 targets mucosal sites, persists, and importantly does not exacerbate SIV
mac251
infection. These data provide the impetus for the development of an attenuated HTLV-2-based vectored vaccine for HIV-1; this approach could elicit persistent mucosal immunity that may prevent HIV-1/SIV
mac251
infection.
Human RNA Polymerase I-Driven Reverse Genetics for Influenza A Virus in Canine Cells
Authors: Suphaphiphat Pirada , Keiner Bjoern , Trusheim Heidi , Crotta Stefania , Tuccino Annunziata Barbara , Zhang Pu , Dormitzer Philip R. , Mason Peter W. , Franti Michael
Published: January 2010

We have established a human RNA polymerase I (pol I)-driven influenza virus reverse genetics (RG) system in the Madin-Darby canine kidney 33016-PF cell line, which is approved for influenza vaccine manufacture. RNA pol I polymerases are generally active only in cells of species closely related to the species of origin of the polymerases. Nevertheless, we show that a nonendogenous RNA pol I promoter drives efficient rescue of influenza A viruses in a canine cell line. Application of this system allows efficient generation of virus strains and presents an alternative approach for influenza vaccine production.
Predominant Clonal Accumulation of CD8+T Cells with Moderate Avidity in the Central Nervous Systems of Theiler's Virus-Infected C57BL/6 Mice
Authors: Kang Hyun Seok , Kim Byung S.
Published: January 2010

Induction of antigen-specific CD8+T cells bearing a high-avidity T-cell receptor (TCR) is thought to be an important factor in antiviral and antitumor immune responses. However, the relationship between TCR diversity and functional avidity of epitope-specific CD8+T cells accumulating in the central nervous system (CNS) during viral infection is unknown. Hence, analysis of T-cell diversity at the clonal level is important to understand the fate and function of virus-specific CD8+T cells. In this study, we examined the Vβ diversity and avidity of CD8+T cells specific to the predominant epitope (VP2121-130) of Theiler's murine encephalomyelitis virus. We found that Vβ6+CD8+T cells, associated with epitope specificity, predominantly expanded in the CNS during viral infection. Further investigations of antigen-specific Vβ6+CD8+T cells by CDR3 spectratyping and sequencing indicated that distinct T-cell clonotypes are preferentially increased in the CNS compared to the periphery. Among the epitope-specific Vβ6+CD8+T cells, MGX-Jβ1.1 motif-bearing cells, which could be found at a high precursor frequency in naïve mice, were expanded in the CNS and tightly associated with gamma interferon production. These T cells displayed moderate avidity for the cognate epitope rather than the high avidity normally observed in memory/effector T cells. Therefore, our findings provide new insights into the CD8+T-cell repertoire during immune responses to viral infection in the CNS.
Meta-Analysis To Test the Association of HIV-1
nef
Amino Acid Differences and Deletions with Disease Progression
Authors: Pushker Ravindra , Jacqué Jean-Marc , Shields Denis C.
Published: January 2010

Previous relatively small studies have associated particular amino acid replacements and deletions in the HIV-1
nef
gene with differences in the rate of HIV disease progression. We tested more rigorously whether particular
nef
amino acid differences and deletions are associated with HIV disease progression. Amino acid replacements and deletions in patients' consensus sequences were investigated for 153 progressor (P), 615 long-term nonprogressor (LTNP), and 2,311 unknown progressor sequences from 582 subtype B HIV-infected patients. LTNPs had more defective
nef
s (interrupted by frameshifts or stop codons), but on a per-patient basis there was no excess of LTNP patients with one or more defective
nef
sequences compared to the Ps (
P
= 0.47). The high frequency of amino acid replacement at residues S
8
, V
10
, I
11
, A
15
, V
85
, V
133
, N
157
, S
163
, V
168
, D
174
, R
178
, E
182
, and R
188
in LTNPs was also seen in permuted datasets, implying that these are simply rapidly evolving residues. Permutation testing revealed that residues showing the greatest excess over expectation (A
15
, V
85
, N
157
, S
163
, V
168
, D
174
, R
178
, and R
188
) were not significant (
P
= 0.77). Exploratory analysis suggested a hypothetical excess of frameshifting in the regions
9
SVIG and
118
QGYF among LTNPs. The regions V
10
and
152
KVEEA of
nef
were commonly deleted in LTNPs. However, permutation testing indicated that none of the regions displayed significantly excessive deletion in LTNPs. In conclusion, meta-analysis of HIV-1
nef
sequences provides no clear evidence of whether defective
nef
sequences or particular regions of the protein play a significant role in disease progression.
1a/1b Subtype Profiling of Nonnucleoside Polymerase Inhibitors of Hepatitis C Virus
Authors: Nyanguile Origène , Devogelaere Benoit , Vijgen Leen , Van den Broeck Walter , Pauwels Frederik , Cummings Maxwell D. , De Bondt Hendrik L. , Vos Ann M. , Berke Jan M. , Lenz Oliver , Vandercruyssen Geneviève , Vermeiren Katrien , Mostmans Wendy , Dehertogh Pascale , Delouvroy Frédéric , Vendeville Sandrine , VanDyck Koen , Dockx Koen , Cleiren Erna , Raboisson Pierre , Simmen Kenneth A. , Fanning Gregory C.
Published: January 2010

The RNA-dependent RNA polymerase (NS5B) of hepatitis C virus (HCV) is an unusually attractive target for drug discovery since it contains five distinct drugable sites. The success of novel antiviral therapies will require nonnucleoside inhibitors to be active in at least patients infected with HCV of subtypes 1a and 1b. Therefore, the genotypic assessment of these agents against clinical isolates derived from genotype 1-infected patients is an important prerequisite for the selection of suitable candidates for clinical development. Here we report the 1a/1b subtype profiling of polymerase inhibitors that bind at each of the four known nonnucleoside binding sites. We show that inhibition of all of the clinical isolates tested is maintained, except for inhibitors that bind at the palm-1 binding site. Subtype coverage varies across chemotypes within this class of inhibitors, and inhibition of genotype 1a improves when hydrophobic contact with the polymerase is increased. We investigated if the polymorphism of the palm-1 binding site is the sole cause of the reduced susceptibility of subtype 1a to inhibition by 1,5-benzodiazepines by using reverse genetics, X-ray crystallography, and surface plasmon resonance studies. We showed Y415F to be a key determinant in conferring resistance on subtype 1a, with this effect being mediated through an inhibitor- and enzyme-bound water molecule. Binding studies revealed that the mechanism of subtype 1a resistance is faster dissociation of the inhibitor from the enzyme.
Epigenetic Regulation of Kaposi's Sarcoma-Associated Herpesvirus Latency by Virus-Encoded MicroRNAs That Target Rta and the Cellular Rbl2-DNMT Pathway
Authors: Lu Fang , Stedman William , Yousef Malik , Renne Rolf , Lieberman Paul M.
Published: January 2010

Kaposi's sarcoma-associated herpesvirus (KSHV) encodes a cluster of 12 microRNAs (miRNAs) that are processed from a transcript that is embedded within the major latency control region. We have generated a deletion mutation that eliminates 10 of the 12 viral miRNAs from the KSHV bacmid by using recombineering methods. The KSHV miRNA deletion mutant (BAC36 ΔmiR) behaved similarly to wild-type (wt) BAC36 in viral production, latency gene transcription, and viral DNA copy number in 293 and dermal microvascular endothelial cells (DMVECs). However, BAC36 ΔmiR consistently expressed elevated levels of viral lytic genes, including the immediate-early transcriptional activator Rta (ORF50). At least one KSHV microRNA (miRK12-5) was capable of suppressing ORF50 mRNA, but poor seed sequence alignments suggest that these targets may be indirect. Comparison of epigenetic marks in ΔmiR KSHV genomes revealed decreases in histone H3 K9 methylation, increases in histone H3 acetylation, and a striking loss of DNA methylation throughout the viral and cellular genome. One viral miRNA, K12-4-5p, was found to have a sequence targeting retinoblastoma (Rb)-like protein 2 (Rbl2), which is a known repressor of DNA methyl transferase 3a and 3b mRNA transcription. We show that ectopic expression of miR-K12-4-5p reduces Rbl2 protein expression and increases DNMT1, -3a, and -3b mRNA levels relative to the levels for control cells. We conclude that KSHV miRNA targets multiple pathways to maintain the latent state of the KSHV genome, including repression of the viral immediate-early protein Rta and a cellular factor, Rbl2, that regulates global epigenetic reprogramming.
Murine Gammaherpesvirus 68 Has Evolved Gamma Interferon and Stat1-Repressible Promoters for the Lytic Switch Gene 50
Authors: Goodwin Megan M. , Canny Susan , Steed Ashley , Virgin Herbert W.
Published: January 2010

Cytokines regulate viral gene expression with important consequences for viral replication and pathogenesis. Gamma interferon (IFN-γ) is a key regulator of chronic murine gammaherpesvirus 68 (γHV68) infection and a potent inhibitor of γHV68 reactivation from latency. Macrophages are the cell type that is responsive to the IFN-γ-mediated control of γHV68 reactivation; however, the molecular mechanism of this IFN-γ action is undefined. Here we report that IFN-γ inhibits lytic replication of γHV68 in primary bone marrow-derived macrophages and decreases transcript levels for the essential lytic switch gene 50. Interestingly, IFN-γ suppresses the activity of the two known gene 50 promoters, demonstrating that an inflammatory cytokine can directly regulate the promoters for the γHV68 lytic switch gene. Stat1, but not IFN-α/β signaling, is required for IFN-γ action. Moreover, Stat1 deficiency increases basal γHV68 replication, gene 50 expression, and promoter activity. Together, these data identify IFN-γ and Stat1 as being negative regulators of the γHV68 lytic cycle and raise the possibility that γHV68 maintains IFN-γ/Stat1-responsive gene 50 promoters to facilitate cell-extrinsic control over the interchange between the lytic and latent cycles.
NTPase and 5′ to 3′ RNA Duplex-Unwinding Activities of the Hepatitis E Virus Helicase Domain
Authors: Karpe Yogesh A. , Lole Kavita S.
Published: January 2010

Hepatitis E virus (HEV) is a causative agent of acute hepatitis, and it is the sole member of the genus
Hepevirus
in the family
Hepeviridae
. The open reading frame 1 (ORF1) protein of HEV encodes nonstructural polyprotein with putative domains for methyltransferase, cysteine protease, helicase and RNA-dependent RNA polymerase. It is not yet known whether ORF1 functions as a single protein with multiple domains or is processed to form separate functional units. On the basis of amino acid conserved motifs, HEV helicase has been grouped into helicase superfamily 1 (SF-1). In order to examine the RNA helicase activity of the NTPase/helicase domain of HEV, the region (amino acids 960 to 1204) was cloned and expressed as histidine-tagged protein in
Escherichia coli
(HEV Hel) and purified. HEV Hel exhibited NTPase and RNA unwinding activities. Enzyme hydrolyzed all rNTPs efficiently, dATP and dCTP with moderate efficiency, while it showed less hydrolysis of dGTP and dTTP. Enzyme showed unwinding of only RNA duplexes with 5′ overhangs showing 5′-to-3′ polarity. We also expressed and purified two HEV Hel mutants. Helicase mutant I, with substitution in the nucleotide-binding motif I (GKS to GAS), showed 30% ATPase activity. Helicase mutant II, with substitutions in the Mg
2+
binding motif II (DEAP to AAAP), showed 50% ATPase activity. Both mutants completely lost ability to unwind RNA duplexes with 5′ overhangs. These findings represent the first report demonstrating NTPase/RNA helicase activity of the helicase domain of HEV ORF1.