bluuebunny/crossref_metadata_2025 huggingface dataset:
Parallel fuzzy connected image segmentation on GPU
Authors: Zhuge Ying , Cao Yong , Udupa Jayaram K. , Miller Robert W.
Published: June 2011

Purpose:

Image segmentation techniques using fuzzy connectedness (FC) principles have shown their effectiveness in segmenting a variety of objects in several large applications. However, one challenge in these algorithms has been their excessive computational requirements when processing large image datasets. Nowadays, commodity graphics hardware provides a highly parallel computing environment. In this paper, the authors present a parallel fuzzy connected image segmentation algorithm implementation on NVIDIA's compute unified device Architecture (cuda) platform for segmenting medical image data sets.

Methods:

In the FC algorithm, there are two major computational tasks: (i) computing the fuzzy affinity relations and (ii) computing the fuzzy connectedness relations. These two tasks are implemented ascuda kernels and executed on GPU. A dramatic improvement in speed for both tasks is achieved as a result.

Results:

Our experiments based on three data sets of small, medium, and large data size demonstrate the efficiency of the parallel algorithm, which achieves a speed‐up factor of 24.4x, 18.1x, and 10.3x, correspondingly, for the three data sets on the NVIDIA Tesla C1060 over the implementation of the algorithm on CPU, and takes 0.25, 0.72, and 15.04 s, correspondingly, for the three data sets.

Conclusions:

The authors developed a parallel algorithm of the widely used fuzzy connected image segmentation method on the NVIDIA GPUs, which are far more cost‐ and speed‐effective than both cluster of workstations and multiprocessing systems. A near‐interactive speed of segmentation has been achieved, even for the large data set.
Reflection modeling in ultraviolet phototherapy
Authors: Grimes David Robert , Robbins Chris , Martin Colin J. , Phanco Graeme , O'Hare Neil John
Published: June 2011

Purpose:

Ultraviolet phototherapy is a widely used treatment which has exceptional success with a variety of skin conditions. Over‐exposure to ultraviolet radiation (UVR) can however be detrimental and cause side effects such as erythema, photokeratisis, and even skin cancer. Quantifying patient dose is therefore imperative to ensure biologically effective treatment while minimizing negative repercussions. A dose model for treatment would be valuable in achieving these ends.

Methods:

Prior work by the authors concentrated on modeling the output of the lamps used in treatment and it was found a line source model described the output from the sources to a high degree. In practice, these lamps are surrounded by reflective anodized aluminum in patient treatment cabins and this work extends the model to quantify specular reflections from these planes on patient dose.

Results:

The extension of the model to allow for reflected images in addition to tube output shows a remarkably good fit to the actual data measured.

Conclusions:

The reflection model yields impressive accuracy and is a good basis for full UVR cabin modeling.
SonoKnife: Feasibility of a line‐focused ultrasound device for thermal ablation therapy
Authors: Chen Duo , Xia Rongmin , Chen Xin , Shafirstein Gal , Corry Peter M. , Griffin Robert J. , Penagaricano Jose A. , Tulunay‐Ugur Ozlem E. , Moros Eduardo G.
Published: June 2011

Purpose:

To evaluate the feasibility of line‐focused ultrasound for thermal ablation of superficially located tumors.

Methods:

A SonoKnife is a cylindrical‐section ultrasound transducer designed to radiate from its concave surface. This geometry generates a line‐focus or acoustic edge. The motivation for this approach was the noninvasive thermal ablation of advanced head and neck tumors and positive neck nodes in reasonable treatment times. Line‐focusing may offer advantages over the common point‐focusing of spherically curved radiators such as faster coverage of a target volume by scanning of the acoustic edge. In this paper, The authors report studies using numerical models and phantom andex vivo experiments using a SonoKnife prototype.

Results

: Acoustic edges were generated by cylindrical‐section single‐element ultrasound transducers numerically, and by the prototype experimentally. Numerically, simulations were performed to characterize the acoustic edge for basic design parameters: transducer dimensions, line‐focus depth, frequency, and coupling thickness. The dimensions of the acoustic edge as a function of these parameters were determined. In addition, a step‐scanning simulation produced a large thermal lesion in a reasonable treatment time. Experimentally, pressure distributions measured in degassed water agreed well with acoustic simulations, and sonication experiments in gel phantoms andex vivo porcine liver samples produced lesions similar to those predicted with acoustic and thermal models.

Conclusions

: Results support the feasibility of noninvasive thermal ablation with a SonoKnife.
Characterization of imaging performance in differential phase contrast CT compared with the conventional CT—Noise power spectrum NPS(k)
Authors: Tang Xiangyang , Yang Yi , Tang Shaojie
Published: June 2011

Purpose:

The differential phase contrast CT is emerging as a new technology to improve the contrast sensitivity of the conventional CT. Via system analysis, modeling, and computer simulation, the authors study the noise power spectrum (NPS)—an imaging performance indicator—of the differential phase contrast CT and compare it with that of the conventional CT.

Methods:

The differential phase contrast CT is implemented with x‐ray tube and gratings. The x‐ray propagation and data acquisition are modeled and simulated with Fourier analysis and Fresnel analysis. To avoid any interference caused by scatter and beam hardening, a monochromatic x‐ray source (30 keV) is assumed, which irradiates the object to be imaged by 360° so that no weighting scheme is needed. A 20‐fold up‐sampling is assumed to simulate x‐ray beam's propagation through the gratingsG1 and G2 with periods 8 and 4 μm, respectively, while the intergrating distance is 193.6 mm (1/16 of the Tabolt distance). The dimension of the detector cell for data acquisition ranges from 32 × 32 to 128 × 128 μm2, while the field of view in data acquisition is 40.96 × 40.96 mm2. A uniform water phantom with a diameter 37.68 mm is employed to study the NPS, with its complex refraction coefficient n = 1 − δ + iβ = 1 − 2.5604 × 10−7 + i1.2353 × 10−10. The x‐ray flux ranges from 106 to 108 photon/cm2·projection and observes the Poisson distribution, which is consistent with that of micro‐CT in preclinical applications. The image matrix of reconstructed water phantom is 1280 × 1280, and a total of 180 regions at 128 × 128 matrix are used for NPS calculation via 2D Fourier Transform in which adequate zero padding is applied to avoid aliasing.

Results:

The preliminary data show that the differential phase contrast CT manifests its NPS with a 1/|k| trait, while the distribution of the conventional CTs NPS observes |k|. This accounts for the significant difference in their noise granularity and the differential phase contrast CTs substantial advantage in noise over the conventional CT, particularly, in the situations where the detector cell size for data acquisition is smaller than 100 μm.

Conclusions:

The differential phase contrast CT detects the projection of refractive coefficient's derivative and uses the Hilbert filter for image reconstruction, which leads to the radical difference in its NPS and the advantage in noise in comparison to that of the conventional CT.
Overestimations in zero frequency DQE of x‐ray imaging converters assessed by Monte Carlo techniques based on the study of energy impartation events
Authors: Liaparinos P. F. , Kandarakis I. S.
Published: June 2011

Purpose:

The performance of various x‐ray converters, employed in medical imaging systems, has been widely examined by several methodologies (experimental, analytical, and Monte Carlo techniques). The x‐ray converters most frequently employed in energy integrating digital radiology detectors are the Gd2O2S:Tb granular phosphor, the CsI:Tl structured phosphor, and the a‐Se photoconductor. The imaging characteristics of an x‐ray converter are affected by its x‐ray detection properties. However, various definitions of x‐ray detection have been used in the literature, leading to different results for the quantum detection efficiency (QDE) for the same type of x‐ray converter. For this reason, there is a need for accurate determination of the x‐ray detection and, in particular, its relation to detector response.

Methods:

The present article reports on the performance of the three aforementioned x‐ray converters in terms of the QDE and the x‐ray statistical factor and examines the effect of the x‐ray detection, directly related to converter output signal, on the zero‐frequency DQE. For the purposes of this study, Monte Carlo simulation was used to model the x‐ray interactions within the x‐ray converter. Simulations were carried out in the energy range from 10 keV up to 80 keV and considering two layers of different coating weights (50 and 100 mg/cm2). The prediction and comparison of zero‐frequency DQE were based on two different approaches for x‐ray detection, i.e., (a) fraction of interacting photons and (b) fraction of photons leading to energy deposition. In addition, the effect of energy deposition through Compton scattering events on the DQE values was estimated.

Results:

Our results showed discrepancies between Monte Carlo techniques (based on energy deposition events) and analytical calculations (based on x‐ray attenuation) on QDE. Discrepancies were found to range up to 10% for Gd2O2S:Tb (100 mg/cm2), 7.7% for CsI:Tl (50 mg/cm2), and 8.2% for a‐Se (50 mg/cm2). Discrepancies were analyzed by examining the scattering effects (elastic and inelastic) within the converters and led to further analysis of scattering events on as well. Significant overestimations were found for both factors (QDE and ) on the zero‐frequency DQE.

Conclusions:

Considering that the highest overestimation was found in the thin layer (50 mg/cm2), Monte Carlo evaluation showed that the overestimation (%) between DQE values (based on either x‐ray interacting events or energy impartation events) was more significant at 20 keV for CsI:Tl (approximately 2.1%), at 40 keV for Gd2O2S:Tb (approximately 8.1%), and finally at 60 and 80 keV for a‐Se converter (approximately 4.8 and 8.2%, respectively). Finally, the overestimation due to the effect of Compton scattering on DQE was found more significant for the a‐Se converter and especially above 40 keV reaching approximately the value of 8.9% at 80 keV.
Quantitative analysis of beam delivery parameters and treatment process time for proton beam therapy
Authors: Suzuki Kazumichi , Gillin Michael T. , Sahoo Narayan , Zhu X. Ronald , Lee Andrew K. , Lippy Denise
Published: June 2011

Purpose:

To evaluate patient census, equipment clinical availability, maximum daily treatment capacity, use factor for major beam delivery parameters, and treatment process time for actual treatments delivered by proton therapy systems.

Methods:

The authors have been recording all beam delivery parameters, including delivered dose, energy, range, spread‐out Bragg peak widths, gantry angles, and couch angles for every treatment field in an electronic medical record system. We analyzed delivery system downtimes that had been recorded for every equipment failure and associated incidents. These data were used to evaluate the use factor of beam delivery parameters, the size of the patient census, and the equipment clinical availability of the facility. The duration of each treatment session from patient walk‐in and to patient walk‐out of the treatment room was measured for 82 patients with cancers at various sites.

Results:

The yearly average equipment clinical availability in the last 3 yrs (June 2007–August 2010) was 97%, which exceeded the target of 95%. Approximately 2200 patients had been treated as of August 2010. The major disease sites were genitourinary (49%), thoracic (25%), central nervous system (22%), and gastrointestinal (2%). Beams have been delivered in approximately 8300 treatment fields. The use factor for six beam delivery parameters was also evaluated. Analysis of the treatment process times indicated that approximately 80% of this time was spent for patient and equipment setup. The other 20% was spent waiting for beam delivery and beam on. The total treatment process time can be expressed by a quadratic polynomial of the number of fields per session. The maximum daily treatment capacity of our facility using the current treatment processes was estimated to be 133 ± 35 patients.

Conclusions:

This analysis shows that the facility has operated at a high performance level and has treated a large number of patients with a variety of diseases. The use factor of beam delivery parameters varies by disease site. Further improvements in efficiency may be realized in the equipment‐ and patient‐related processes of treatment.
Oxytocin Selectively Gates Fear Responses Through Distinct Outputs from the Central Amygdala
Authors: Viviani Daniele , Charlet Alexandre , van den Burg Erwin , Robinet Camille , Hurni Nicolas , Abatis Marios , Magara Fulvio , Stoop Ron
Published: June 2011

Specific neuronal populations in the amygdala control different behavioral and physiological components of the fear response.
Comment on “The Mechanism for Activation of GTP Hydrolysis on the Ribosome”
Authors: Liljas Anders , Ehrenberg Måns , Åqvist Johan
Published: June 2011

Voorhees
et al
. (Reports, 5 November 2010, p. 835) determined the structure of elongation factor Tu (EF-Tu) and aminoacyl–transfer RNA bound to the ribosome with a guanosine triphosphate (GTP) analog. However, their identification of histidine-84 of EF-Tu as deprotonating the catalytic water molecule is problematic in relation to their atomic structure; the terminal phosphate of GTP is more likely to be the proper proton acceptor.
Response to Comment on “The Mechanism for Activation of GTP Hydrolysis on the Ribosome”
Authors: Voorhees Rebecca M. , Schmeing T. Martin , Kelley Ann C. , Ramakrishnan V.
Published: June 2011

Our report of the crystal structure of elongation factor Tu (EF-Tu) and aminoacyl–transfer RNA bound to the ribosome with a guanosine triphosphate (GTP) analog included a proposed mechanism of GTP hydrolysis by EF-Tu involving histidine-84. Liljas
et al
. summarize experimental evidence against this mechanism and propose a substrate-assisted catalytic model. However, these experiments and the model are also problematic. Further study is required to definitively determine the mechanism of GTP hydrolysis by EF-Tu.
Formation and Spread of Aircraft-Induced Holes in Clouds
Authors: Heymsfield Andrew J. , Thompson Gregory , Morrison Hugh , Bansemer Aaron , Rasmussen Roy M. , Minnis Patrick , Wang Zhien , Zhang Damao
Published: June 2011

Aircraft passing through clouds that contain supercooled liquid water can produce holes and canals in those clouds.
Following the Crowd: Brain Substrates of Long-Term Memory Conformity
Authors: Edelson Micah , Sharot Tali , Dolan Raymond J. , Dudai Yadin
Published: June 2011

Firmly held memory representations in the brain can be altered by social influence.
Large Sulfur Isotope Fractionation Does Not Require Disproportionation
Authors: Sim Min Sub , Bosak Tanja , Ono Shuhei
Published: June 2011

In the absence of oxygenation, microbial activity can explain the magnitude of sulfur-isotope traces in sediments.
A Key Enzyme in the Biogenesis of Lysosomes Is a Protease That Regulates Cholesterol Metabolism
Authors: Marschner Katrin , Kollmann Katrin , Schweizer Michaela , Braulke Thomas , Pohl Sandra
Published: June 2011

Defects in a sterol regulatory pathway also cause defects in lysosome assembly leading to mucolipidosis and disease.
Turning the Tide Against HIV
Authors: Shattock Robin J. , Warren Mitchell , McCormack Sheena , Hankins Catherine A.
Published: June 2011

Potentially beneficial combinations of prevention options need innovative trials to assess multiple tools against a common control.
Electrons in Cement
Authors: Edwards Peter P.
Published: June 2011

A system of stabilized solvated electrons has been realized in a new type of inorganic electride.
Knot Your Simple Defect Lines?
Authors: Kamien Randall D.
Published: June 2011

The ordering of molecules in a liquid crystal is disrupted by colloidal particles and creates lines of defects, which can be manipulated to form loops and knots.
Remember When?
Authors: Roediger Henry L. , McDermott Kathleen B.
Published: June 2011

Group interactions can have both negative and positive impacts on memory.
Fishing in the Nuclear Pore
Authors: Kriwacki Richard W. , Yoon Mi-Kyung
Published: June 2011

The flexibility of an intrinsically disordered domain of a membrane protein allows it to pass through the nuclear pore.
Underground Lab Would Cost U.S. Billions
Authors: Cho Adrian
Published: June 2011

The Department of Energy would need to spend between $1.2 billion and $2.2 billion to build and equip an underground particle physics laboratory, according to a study presented last week to a federal advisory panel.
As
E. coli
Outbreak Recedes, New Questions Come to the Fore
Authors: Kupferschmidt Kai
Published: June 2011

Now that the deadliest outbreak of enterohemorrhagic
Escherichia coli
ever recorded appears to be winding down, some scientists say the outbreak may have given them study material for years.
NASA's Busload of Science
Authors: Charles Dan
Published: June 2011

For 30 years, the shuttle has been the main ticket into space for NASA astronauts. But it has also delivered—and fixed—massive observatories and served as an orbiting laboratory. As the program ends, what is its scientific legacy?
Development of an Automated, High-Throughput Bactericidal Assay That Measures Cellular Respiration as a Survival Readout for Neisseria meningitidis
Authors: Mak Puiying A. , Santos George F. , Masterman Kelly-Anne , Janes Jeff , Wacknov Bill , Vienken Kay , Giuliani Marzia , Herman Ann E. , Cooke Michael , Mbow M. Lamine , Donnelly John
Published: June 2011

Complement-mediated bactericidal activity has long been regarded as the serological correlate of protective immunity againstNeisseria meningitidis. This was affirmed in 2005 at a WHO-sponsored meningococcal serology standardization workshop. The assay currently employed by most laboratories involves determining surviving bacterial colony counts on agar as a readout which is labor-intensive, time-consuming, and not amendable to rapid data analysis for clinical trials. Consequently, there is an acute need to develop a sensitive, high-throughput bactericidal assay to enable a rapid and robust assessment of the effectiveness of vaccine candidates. To this end, we have developed an automated, kinetic assay based on the fluorescent respiration product of resazurin which reduces assay volume, shortens assay time, and facilitates automation of data analysis. We demonstrate proof of concept for applicability of this high-throughput system with multiple meningococcal strains and utilizing different lots of human complement. The assay is robust and highly reproducible. Titers obtained by the fluorescence readout method are strongly correlated with the data obtained using the conventional, agar plate-based assay. These results demonstrate that the detection of bacteria that have survived the bactericidal reaction by measuring metabolic activity using a fluorescent dye as an alternative readout is a promising approach for the development of a high-throughput bactericidal assay.
Allele-Specific Real-Time PCR System for Detection of Subpopulations of Genotype 1a and 1b Hepatitis C NS5B Y448H Mutant Viruses in Clinical Samples
Authors: Bae Andrew S. , Ku Karin S. , Miller Michael D. , Mo Hongmei , Svarovskaia Evguenia S.
Published: June 2011

The Y448H mutation in NS5B has been selected by GS-9190 as well as several benzothiadiazine hepatitis C virus (HCV) polymerase inhibitors
in vitro
and
in vivo
. However, the level and the evolution kinetics of this resistance mutation prior to and during treatment are poorly understood. In this study, we developed an allele-specific real-time PCR (AS-PCR) assay capable of detecting Y448H when it was present at a level down to 0.5% within an HCV population of genotype 1a or 1b. No Y448H mutation was detected above the assay cutoff of 0.5% in genotype 1b-infected Con-1 replicons prior to
in vitro
treatment. However, the proportion of replicons with the Y448H mutation rapidly increased in a dose-dependent manner upon treatment with GS-9190. After 3 days of treatment, 1.2%, 6.8%, and >50% of the replicon population expressed Y448H with the use of GS-9190 at 1, 10, and 20 times its 50% effective concentration, respectively. In addition, plasma from 65 treatment-naïve HCV-infected patients (42 and 23 with genotype 1a and 1b, respectively) was tested for the presence of Y448H by AS-PCR and population sequencing. As expected, all patient samples were wild type at NS5B Y448 by population sequencing. AS-PCR results were obtained for 62/65 samples tested, with low levels of Y448H ranging from 0.5% to 3.0% detected in 5/62 (8%) treatment-naïve patient samples. These findings suggest the need for combination therapy with HCV-specific inhibitors to avoid viral rebound of preexisting mutant HCV.
Legionella micdadei, a New Cause of Prosthetic Joint Infection
Authors: Fernández-Cruz Ana , Marín Mercedes , Castelo Laura , Usubillaga Rafael , Martín-Rabadán Pablo , Bouza Emilio ,   the GAIO (Group for the Assessment of Osteoarticular Infections) Study Group
Published: June 2011

Extrapulmonary infections caused by
Legionella
spp. other than
Legionella pneumophila
are rare. We report what is, to our knowledge, the first description of a prosthetic joint infection due to
Legionella
spp. Systematic testing of samples with suspected prosthetic infection by molecular biology techniques was essential.
Legionella micdadei
should be added to the list of microorganisms causing prosthetic joint infection.
Molecular Characterization of a Catalase-Negative Staphylococcus aureus subsp.
aureus
Strain Collected from a Patient with Mitral Valve Endocarditis and Pericarditis Revealed a Novel Nonsense Mutation in the
katA
Gene
Authors: To Kelvin K. W. , Cheng Vincent C. C. , Chan Jasper F. W. , Wong Amy C. Y. , Chau Sandy , Tsang Flora H. F. , Curreem Shirly O. T. , Lau Susanna K. P. , Yuen Kwok-Yung , Woo Patrick C. Y.
Published: June 2011

We report a case of endocarditis and pericarditis caused by catalase-negative
Staphylococcus aureus
. Molecular characterization revealed a novel nonsense mutation in the
katA
gene, leading to a loss of 238 amino acids (47% of the wild-type catalase protein), including the heme-binding site, NADPH-binding region, and Tyr-337, essential for catalysis.
Analysis of Intrahost Variation in Venezuelan Equine Encephalitis Virus Reveals Repeated Deletions in the 6-Kilodalton Protein Gene
Authors: Forrester N. L. , Guerbois M. , Adams A. P. , Liang X. , Weaver S. C.
Published: June 2011

RNA viruses exist as a spectrum of mutants that is generated and maintained during replication within the host. Consensus sequencing overlooks minority genotypes present in the viral sample that may impact the population's phenotype. In-depth sequencing of an original field isolate of subtype IE Venezuelan equine encephalitis virus (VEEV) demonstrated the presence of multiple deletions within the 6,000-molecular-weight (6K) protein gene. Using
in vitro
and
in vivo
experiments, similar deletions were generated in an additional VEEV strain originating from an infectious cDNA clone. Time course experiments demonstrated that the deletions are produced during acute infection although not until 24 h postinfection. Molecular clones containing some of these deletions were generated, and although the larger deletions appear to be noninfectious, viruses with the smaller deletions were viable and formed small plaques. Serial passages provided no evidence that these deletion mutants function as defective interfering particles. Furthermore, since wild-type infections generally occur at a low multiplicity of infection, it is unlikely that these deletions are propagated in natural transmission cycles. However, they could affect pathogenesis at later stages of infection. Because they are ubiquitously generated both
in vivo
and
in vitro
, further investigation is warranted to understand the generation of these deletions and their significance for disease.
M94 Is Essential for the Secondary Envelopment of Murine Cytomegalovirus
Authors: Maninger Silke , Bosse Jens Bernhard , Lemnitzer Frederic , Pogoda Madlen , Mohr Christian A. , von Einem Jens , Walther Paul , Koszinowski Ulrich H. , Ruzsics Zsolt
Published: June 2011

The gene M94 of murine cytomegalovirus (MCMV) as well as its homologues UL16 in alphaherpesviruses is involved in viral morphogenesis. For a better understanding of its role in the viral life cycle, a library of random M94 mutants was generated by modified transposon-based linker scanning mutagenesis. A comprehensive set of M94 mutants was reinserted into the MCMV genome and tested for their capacity to complement the M94 null mutant. Thereby, 34 loss-of-function mutants of M94 were identified, which were tested in a second screen for their capacity to inhibit virus replication. This analysis identified two N-terminal insertion mutants of M94 with a dominant negative effect. We compared phenotypes induced by the conditional expression of these dominant negative M94 alleles with the null phenotype of the M94 deletion. The viral gene expression cascade and the nuclear morphogenesis steps were not affected in either setting. In both cases, however, secondary envelopment did not proceed in the absence of functional M94, and capsids subsequently accumulated in the center of the cytoplasmic assembly complex. In addition, deletion of M94 resulted in a block of cell-to-cell spread. Moreover, the dominant negative mutant of M94 demonstrated a defect in interacting with M99, the UL11 homologue of MCMV.
Comparative Evolution of GII.3 and GII.4 Norovirus over a 31-Year Period
Authors: Boon Denali , Mahar Jackie E. , Abente Eugenio J. , Kirkwood Carl D. , Purcell Robert H. , Kapikian Albert Z. , Green Kim Y. , Bok Karin
Published: June 2011

Noroviruses are the most common cause of epidemic gastroenteritis. Genotype II.3 is one of the most frequently detected noroviruses associated with sporadic infections. We studied the evolution of the major capsid gene from seven archival GII.3 noroviruses collected during a cross-sectional study at the Children's Hospital in Washington, DC, from 1975 through 1991, together with capsid sequence from 56 strains available in GenBank. Evolutionary analysis concluded that GII.3 viruses evolved at a rate of 4.16 × 10
−3
nucleotide substitutions/site/year (strict clock), which is similar to that described for the more prevalent GII.4 noroviruses. The analysis of the amino acid changes over the 31-year period found that GII.3 viruses evolve at a relatively steady state, maintaining 4% distance, and have a tendency to revert back to previously used residues while preserving the same carbohydrate binding profile. In contrast, GII.4 viruses demonstrate increasing rates of distance over time because of the continued integration of new amino acids and changing HBGA binding patterns. In GII.3 strains, seven sites acting under positive selection were predicted to be surface-exposed residues in the P2 domain, in contrast to GII.4 positively selected sites located primarily in the shell domain. Our study suggests that GII.3 noroviruses caused disease as early as 1975 and that they evolve via a specific pattern, responding to selective pressures induced by the host rather than presenting a nucleotide evolution rate lower than that of GII.4 noroviruses, as previously proposed. Understanding the evolutionary dynamics of prevalent noroviruses is relevant to the development of effective prevention and control strategies.
Comprehensive Proteomic Analysis of Influenza Virus Polymerase Complex Reveals a Novel Association with Mitochondrial Proteins and RNA Polymerase Accessory Factors
Authors: Bradel-Tretheway Birgit G. , Mattiacio Jonelle L. , Krasnoselsky Alexei , Stevenson Catherine , Purdy David , Dewhurst Stephen , Katze Michael G.
Published: June 2011

The trimeric RNA polymerase complex (3P, for PA-PB1-PB2) of influenza A virus (IAV) is an important viral determinant of pathogenicity and host range restriction. Specific interactions of the polymerase complex with host proteins may be determining factors in both of these characteristics and play important roles in the viral life cycle. To investigate this question, we performed a comprehensive proteomic analysis of human host proteins associated with the polymerase of the well-characterized H5N1 Vietnam/1203/04 isolate. We identified over 400 proteins by liquid chromatography-tandem mass spectrometry (LC-MS/MS), of which over 300 were found to bind to the PA subunit alone. The most intriguing and novel finding was the large number of mitochondrial proteins (∼20%) that associated with the PA subunit. These proteins mediate molecular transport across the mitochondrial membrane or regulate membrane potential and may in concert with the identified mitochondrion-associated apoptosis inducing factor (AIFM1) have roles in the induction of apoptosis upon association with PA. Additionally, we identified host factors that associated with the PA-PB1 (68 proteins) and/or the 3P complex (34 proteins) including proteins that have roles in innate antiviral signaling (e.g., ZAPS or HaxI) or are cellular RNA polymerase accessory factors (e.g., polymerase I transcript release factor [PTRF] or Supt5H). IAV strain-specific host factor binding to the polymerase was not observed in our analysis. Overall, this study has shed light into the complex contributions of the IAV polymerase to host cell pathogenicity and allows for direct investigations into the biological significance of these newly described interactions.
Adenovirus E1A Directly Targets the E2F/DP-1 Complex
Authors: Pelka Peter , Miller Matthew S. , Cecchini Matthew , Yousef Ahmed F. , Bowdish Dawn M. , Dick Fred , Whyte Peter , Mymryk Joe S.
Published: June 2011

Deregulation of the cell cycle is of paramount importance during adenovirus infection. Adenovirus normally infects quiescent cells and must initiate the cell cycle in order to propagate itself. The pRb family of proteins controls entry into the cell cycle by interacting with and repressing transcriptional activation by the E2F transcription factors. The viral E1A proteins indirectly activate E2F-dependent transcription and cell cycle entry, in part, by interacting with pRb and family members to free the E2Fs. We report here that an E1A 13S isoform can unexpectedly activate E2F-responsive gene expression independently of binding to the pRb family of proteins. We demonstrate that E1A binds to E2F/DP-1 complexes through a direct interaction with DP-1. E1A appears to utilize this binding to recruit itself to E2F-regulated promoters, and this allows the E1A 13S protein, but not the E1A 12S protein, to activate transcription independently of interaction with pRb. Importantly, expression of E1A 13S, but not E1A 12S, led to significant enhancement of E2F4 occupancy of E2F sites of two E2F-regulated promoters. These observations identify a novel mechanism by which adenovirus deregulates the cell cycle and suggest that E1A 13S may selectively activate a subset of E2F-regulated cellular genes during infection.
Targeting OX40 Promotes Lung-Resident Memory CD8 T Cell Populations That Protect against Respiratory Poxvirus Infection
Authors: Salek-Ardakani Shahram , Moutaftsi Magdalini , Sette Alessandro , Croft Michael
Published: June 2011

One goal of vaccination is to promote development of mucosal effector cells that can immediately respond to peripheral infection. This is especially important for protection against viruses that enter the host through the respiratory tract. We show that targeting the OX40 costimulatory receptor (CD134) strongly promotes mucosal memory in the CD8 T cell compartment. Systemic injection of an agonist antibody to OX40 strongly enhanced development of polyfunctional effector CD8 T cells that were induced after intraperitoneal infection with a highly virulent strain of vaccinia virus. These cells were located in lymphoid organs and also the lung, and importantly, long-term memory CD8 T cells were maintained in the lung over 1 year. Anti-OX40 also boosted memory development when mice were vaccinated subcutaneously with viral peptide. These CD8 T cells were sufficient to provide protection from lethal respiratory infection with live vaccinia virus independent of CD4 T cells and antibody. Again, the CD8 T cell populations that were induced after secondary infection displayed polyfunctionality and were maintained in the lung for over a year. These data suggest that agonists to the OX40 costimulatory receptor represent potential candidates for incorporation into vaccines for respiratory viruses.
Reconstructing the History of Maize Streak Virus Strain A Dispersal To Reveal Diversification Hot Spots and Its Origin in Southern Africa
Authors: Monjane Adérito L. , Harkins Gordon W. , Martin Darren P. , Lemey Philippe , Lefeuvre Pierre , Shepherd Dionne N. , Oluwafemi Sunday , Simuyandi Michelo , Zinga Innocent , Komba Ephrem K. , Lakoutene Didier P. , Mandakombo Noella , Mboukoulida Joseph , Semballa Silla , Tagne Appolinaire , Tiendrébéogo Fidèle , Erdmann Julia B. , van Antwerpen Tania , Owor Betty E. , Flett Bradley , Ramusi Moses , Windram Oliver P. , Syed Rizwan , Lett Jean-Michel , Briddon Rob W. , Markham Peter G. , Rybicki Edward P. , Varsani Arvind
Published: June 2011

Maize streak virus strain A (MSV-A), the causal agent of maize streak disease, is today one of the most serious biotic threats to African food security. Determining where MSV-A originated and how it spread transcontinentally could yield valuable insights into its historical emergence as a crop pathogen. Similarly, determining where the major extant MSV-A lineages arose could identify geographical hot spots of MSV evolution. Here, we use model-based phylogeographic analyses of 353 fully sequenced MSV-A isolates to reconstruct a plausible history of MSV-A movements over the past 150 years. We show that since the probable emergence of MSV-A in southern Africa around 1863, the virus spread transcontinentally at an average rate of 32.5 km/year (95% highest probability density interval, 15.6 to 51.6 km/year). Using distinctive patterns of nucleotide variation caused by 20 unique intra-MSV-A recombination events, we tentatively classified the MSV-A isolates into 24 easily discernible lineages. Despite many of these lineages displaying distinct geographical distributions, it is apparent that almost all have emerged within the past 4 decades from either southern or east-central Africa. Collectively, our results suggest that regular analysis of MSV-A genomes within these diversification hot spots could be used to monitor the emergence of future MSV-A lineages that could affect maize cultivation in Africa.
p63 Is Necessary for the Activation of Human Papillomavirus Late Viral Functions upon Epithelial Differentiation
Authors: Mighty Kristen K. , Laimins Laimonis A.
Published: June 2011

The late phase of the human papillomavirus (HPV) life cycle is linked to epithelial differentiation, and we investigated the factors that regulate this process. One potential regulator is p63, a member of the p53 family of proteins, which modulates epithelial development, as well as proliferation capability, in stem cells. In this study, we examined the role of p63 in the HPV life cycle using a lentiviral knockdown system for p63. In epithelial cells, the ΔN truncated isoforms of p63 predominate, while the full-length TA isoforms are present at very low levels. Upon the differentiation of normal keratinocytes, p63 levels rapidly decreased while higher levels were retained in HPV-positive cells. Our studies indicate that reducing p63 levels in differentiated HPV-positive cells resulted in the loss of viral genome amplification and late gene expression. p63 regulates the expression of cell cycle regulators, and we determined that cyclin A, cyclin B1, cdk1, and cdc25c were reduced in p63-deficient, HPV-positive keratinocytes, which suggests a possible mechanism of action. In addition, activation of the DNA repair pathway is necessary for genome amplification, and the expression of two members, BRCA2 and RAD51, was altered in the absence of p63 in HPV-positive cells. Our studies indicate that p63 is necessary for the activation of differentiation-dependent HPV late viral functions and provide insights into relevant cellular targets.
Mechanism of Neutralization by the Broadly Neutralizing HIV-1 Monoclonal Antibody VRC01
Authors: Li Yuxing , O'Dell Sijy , Walker Laura M. , Wu Xueling , Guenaga Javier , Feng Yu , Schmidt Stephen D. , McKee Krisha , Louder Mark K. , Ledgerwood Julie E. , Graham Barney S. , Haynes Barton F. , Burton Dennis R. , Wyatt Richard T. , Mascola John R.
Published: June 2011

The structure of VRC01 in complex with the HIV-1 gp120 core reveals that this broadly neutralizing CD4 binding site (CD4bs) antibody partially mimics the interaction of the primary virus receptor, CD4, with gp120. Here, we extended the investigation of the VRC01-gp120 core interaction to the biologically relevant viral spike to better understand the mechanism of VRC01-mediated neutralization and to define viral elements associated with neutralization resistance. In contrast to the interaction of CD4 or the CD4bs monoclonal antibody (MAb) b12 with the HIV-1 envelope glycoprotein (Env), occlusion of the VRC01 epitope by quaternary constraints was not a major factor limiting neutralization. Mutagenesis studies indicated that VRC01 contacts within the gp120 loop D, the CD4 binding loop, and the V5 region were necessary for optimal VRC01 neutralization, as suggested by the crystal structure. In contrast to interactions with the soluble gp120 monomer, VRC01 interaction with the native viral spike did not occur in a CD4-like manner; VRC01 did not induce gp120 shedding from the Env spike or enhance gp41 membrane proximal external region (MPER)-directed antibody binding to the Env spike. Finally, VRC01 did not display significant reactivity with human antigens, boding well for potential
in vivo
applications. The data indicate that VRC01 interacts with gp120 in the context of the functional spike in a manner distinct from that of CD4. It achieves potent neutralization by precisely targeting the CD4bs without requiring alterations of Env spike configuration and by avoiding steric constraints imposed by the quaternary structure of the functional Env spike.
Autocatalytic Activity of the Ubiquitin-Specific Protease Domain of Herpes Simplex Virus 1 VP1-2
Authors: Bolstad M. , Abaitua F. , Crump C. M. , O'Hare P.
Published: June 2011

The herpes simplex virus (HSV) tegument protein VP1-2 is essential for virus entry and assembly. VP1-2 also contains a highly conserved ubiquitin-specific protease (USP) domain within its N-terminal region. Despite conservation of the USP and the demonstration that it can act on artificial substrates such as polyubiquitin chains, identification of the relevance of the USP
in vivo
to levels or function of any substrate remains limited. Here we show that HSV VP1-2 USP can act on itself and is important for stability. VP1-2 N-terminal variants encompassing the core USP domain itself were not affected by mutation of the catalytic cysteine residue (C65). However, extending the N-terminal region resulted in protein species requiring USP activity for accumulation. In this context, C65A mutation resulted in a drastic reduction in protein levels which could be stabilized by proteosomal inhibition or by the presence of normal C65. The functional USP domain could increase abundance of unstable variants, indicating action at least in part, in
trans
. Interestingly, full-length variants containing the inactive USP, although unstable when expressed in isolation, were stabilized by virus infection. The catalytically inactive VP1-2 retained complementation activity of a VP1-2-negative virus. Furthermore, a recombinant virus expressing a C65A mutant VP1-2 exhibited little difference in single-step growth curves and the kinetics and abundance of VP1-2 or a number of test proteins. Despite the absence of a phenotype for these replication parameters, the USP activity of VP1-2 may be required for function, including its own stability, under certain circumstances.
Mammalian Orthoreovirus Escape from Host Translational Shutoff Correlates with Stress Granule Disruption and Is Independent of eIF2α Phosphorylation and PKR
Authors: Qin Qingsong , Carroll Kate , Hastings Craig , Miller Cathy L.
Published: June 2011

In response to mammalian orthoreovirus (MRV) infection, cells initiate a stress response that includes eIF2α phosphorylation and protein synthesis inhibition. We have previously shown that early in infection, MRV activation of eIF2α phosphorylation results in the formation of cellular stress granules (SGs). In this work, we show that as infection proceeds, MRV disrupts SGs despite sustained levels of phosphorylated eIF2α and, further, interferes with the induction of SGs by other stress inducers. MRV interference with SG formation occurs downstream of eIF2α phosphorylation, suggesting the virus uncouples the cellular stress signaling machinery from SG formation. We additionally examined mRNA translation in the presence of SGs induced by eIF2α phosphorylation-dependent and -independent mechanisms. We found that irrespective of eIF2α phosphorylation status, the presence of SGs in cells correlated with inhibition of viral and cellular translation. In contrast, MRV disruption of SGs correlated with the release of viral mRNAs from translational inhibition, even in the presence of phosphorylated eIF2α. Viral mRNAs were also translated in the presence of phosphorylated eIF2α in PKR
−/−
cells. These results suggest that MRV escape from host cell translational shutoff correlates with virus-induced SG disruption and occurs in the presence of phosphorylated eIF2α in a PKR-independent manner.
Replacement of Previously Circulating Respiratory Syncytial Virus Subtype B Strains with the BA Genotype in South Africa
Authors: van Niekerk Stephanie , Venter Marietjie
Published: June 2011

Respiratory syncytial virus (RSV) is a major cause of bronchiolitis and pneumonia in infants, the immunocompromised, and the elderly in both developed and developing countries. Reinfections are common, and G protein variability is one mechanism to overcome herd immunity. This is illustrated by the appearance of the BA genotype with a 60-nucleotide duplication dominating the subtype B genotypes in epidemics worldwide. To investigate the evolution of subtype B in South Africa since 2002, the genetic variability of the G protein was analyzed in all recent strains isolated over 4 years (2006 to 2009) in South African hospitals. Bayesian analysis revealed a replacement of all subtype B genotypes previously identified in South Africa with the BA genotype since 2006, while subtype A genotypes identified in previous years are still circulating. Compared to BA strains from other countries, the evolutionary rate of the South African BA genotype was shown to be 2.305 × 10
−3
nucleotide substitutions/site/year and drift was evident. The most recent common ancestor (MRCA) of the South African BA viruses was determined to date back to 1996. All South African BA isolates clustered with the BA-IV subgenotype, and the appearance of new subgenotypes within this branch may occur if drift continues. Sequencing of the complete G protein of selected South African strains revealed an additional 6-nucleotide deletion. Acquisition of the 60-nucleotide duplication appeared to have improved the fitness of this virus, and more recent subtype B strains may need to be included in experimental vaccines to evaluate their efficacy in the current setting of evolved circulating strains.
Host Factors in Enterovirus 71 Replication
Authors: Shih Shin-Ru , Stollar Victor , Li Mei-Ling
Published: June 2011

Enterovirus 71 (EV71) infections continue to remain an important public health problem around the world, especially in the Asia-Pacific region. There is a significant mortality rate following such infections, and there is neither any proven therapy nor a vaccine for EV71. This has spurred much fundamental research into the replication of the virus. In this review, we discuss recent work identifying host cell factors which regulate the synthesis of EV71 RNA and proteins. Three of these proteins, heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1), far-upstream element-binding protein 2 (FBP2), and FBP1 are nuclear proteins which in EV71-infected cells are relocalized to the cytoplasm, and they influence EV71 internal ribosome entry site (IRES) activity. hnRNP A1 stimulates IRES activity but can be replaced by hnRNP A2. FBP2 is a negative regulatory factor with respect to EV71 IRES activity, whereas FBP1 has the opposite effect. Two other proteins, hnRNP K and reticulon 3, are required for the efficient synthesis of viral RNA. The cleavage stimulation factor 64K subunit (CstF-64) is a host protein that is involved in the 3′ polyadenylation of cellular pre-mRNAs, and recent work suggests that in EV71-infected cells, it may be cleaved by the EV71 3C protease. Such a cleavage would impair the processing of pre-mRNA to mature mRNAs. Host cell proteins play an important role in the replication of EV71, but much work remains to be done in order to understand how they act.
Prion Disease Detection, PMCA Kinetics, and IgG in Urine from Sheep Naturally/Experimentally Infected with Scrapie and Deer with Preclinical/Clinical Chronic Wasting Disease
Authors: Rubenstein Richard , Chang Binggong , Gray Perry , Piltch Martin , Bulgin Marie S. , Sorensen-Melson Sharon , Miller Michael W.
Published: June 2011

Prion diseases, also known as transmissible spongiform encephalopathies, are fatal neurodegenerative disorders. Low levels of infectious agent and limited, infrequent success of disease transmissibility and PrP
Sc
detection have been reported with urine from experimentally infected clinical cervids and rodents. We report the detection of prion disease-associated seeding activity (PASA) in urine from naturally and orally infected sheep with clinical scrapie agent and orally infected preclinical and infected white-tailed deer with clinical chronic wasting disease (CWD). This is the first report on PASA detection of PrP
Sc
from the urine of naturally or preclinical prion-diseased ovine or cervids. Detection was achieved by using the surround optical fiber immunoassay (SOFIA) to measure the products of limited serial protein misfolding cyclic amplification (sPMCA). Conversion of PrP
C
to PrP
Sc
was not influenced by the presence of poly(A) during sPMCA or by the homogeneity of the PrP genotypes between the PrP
C
source and urine donor animals. Analysis of the sPMCA-SOFIA data resembled a linear, rather than an exponential, course. Compared to uninfected animals, there was a 2- to 4-log increase of proteinase K-sensitive, light chain immunoglobulin G (IgG) fragments in scrapie-infected sheep but not in infected CWD-infected deer. The higher-than-normal range of IgG levels found in the naturally and experimentally infected clinical scrapie-infected sheep were independent of their genotypes. Although analysis of urine samples throughout the course of infection would be necessary to determine the usefulness of altered IgG levels as a disease biomarker, detection of PrP
Sc
from PASA in urine points to its potential value for antemortem diagnosis of prion diseases.