bluuebunny/crossref_metadata_2025 huggingface dataset:
Validation of new real‐time polymerase chain reaction assays for detection of hepatitis A virus RNA and parvovirus B19 DNA
Authors: Molenaar‐de Backer Marijke W.A. , de Waal Mirjam , Sjerps Margret C. , Koppelman Marco H.G.M.
Published: September 2015

BACKGROUND

To meet European guidelines for plasma for fractionation, plasma fractionators have implemented parvovirus B19 (B19V) and hepatitis A virus (HAV) nucleic acid test (NAT) screening on test pools. In this study we evaluate recently developed in‐house NAT assays for B19V DNA and HAV RNA. The B19V NAT was designed to target two different regions of the B19V genome.

STUDY DESIGN AND METHODS

The B19V DNA and HAV RNA tests were validated according to commonly used guidelines. The performance of the B19V and HAV assays was evaluated during routine screening of more than 2 × 106 donations.

RESULTS

The 95% lower limit of detection (LLD) of the HAV NAT was 1.34 IU/mL. The 95% LLD for B19V was 39.1 IU/mL for the NS1 region and 76.9 IU/mL for the VP2 region. The B19V test showed good accuracy, precision, robustness, and no cross‐contamination was observed. Both assays detected B19V Genotypes 1 to 3 and HAV Genotypes I to III. During routine screening 103 donations showed B19V DNA loads of more than 1.25 × 106 IU/mL and one donation was reactive in the HAV NAT.

CONCLUSION

The dual‐target B19V polymerase chain reaction (PCR) showed good accuracy (<0.1 log IU/mL) at the crucial concentration of 10 IU/µL for the NS1 and the VP2 region of the B19V genome and detected all known genotypes with similar sensitivity for each genotype. In addition, the dual target format reduces the chance that molecular variants of B19V are wrongly quantified. The HAV RNA assay showed high sensitivity for Genotypes I to III. Both new PCR assays have been successfully introduced for plasma screening in test pools of 480 or 96 donations.
Numerical and Experimental Investigation of Aerodynamic Performance for a Straight Turbine Cascade With a Novel Partial Shroud
Authors: Liu Yan , Zhang Tian-Long , Zhang Min , Zhang Meng-Chao
Published: September 2015

A comparative experimental and numerical analysis is carried out to assess the aerodynamic performance of a novel partial shroud in a straight turbine cascade. This partial shroud is designed as a combination of winglet and shroud. A plain tip is employed as a baseline case. A pure winglet tip is also studied for comparison. Both experiments and predictions demonstrate that this novel partial shroud configuration has aerodynamic advantages over the pure winglet arrangement. Predicted results show that, relative to the baseline blade with a plain tip, using the partial shroud can lead to a reduction of 20.89% in the mass-averaged total pressure coefficient on the upper half-span of a plane downstream of the cascade trailing edge and 16.53% in the tip leakage mass flow rate, whereas the pure winglet only decreases these two performance parameters by 11.36% and 1.32%, respectively. The flow physics is explored in detail to explain these results via topological analyses. The use of this new partial shroud significantly affects the topological structures and total pressure loss coefficients on various axial cross sections, particularly at the rear part of the blade passage. The partial shroud not only weakens the tip leakage vortex (TLV) but also reduces the strength of passage vortex near the casing (PVC) endwall. Furthermore, three partial shrouds with width-to-pitch ratios of 3%, 5%, and 7% are considered. With an increase in the width of the winglet part, improvements in aerodynamics and the tip leakage mass flow rate are limited.
Numerical and Experimental Investigation on the Effect of Swirl Brakes on the Labyrinth Seals
Authors: Sun Dan , Wang Shuang , Fei Cheng-Wei , Ai Yan-Ting , Wang Ke-Ming
Published: September 2015

Swirl brake influences the static and rotordynamic characteristics of labyrinth seal which are important in the prediction of turbomachine stability. To study the influence of the swirl brakes on improving seal stability, the effects of swirl brakes on the static and rotordynamic characteristics of labyrinth seals were investigated by the combination of numerical simulation and experiment. First, it was performed to the effects of swirl brake on the static flow characteristics of labyrinth seal with swirl ratio and pressure distribution based on computational fluid dynamics (CFD). And then a comparison between leakage predicted by the CFD model and measurement was presented to verify the accuracy of the simulation. Moreover, an experiment was implemented to analyze the rotordynamic characteristics of labyrinth seal using an improved impedance method based on an unbalanced synchronous excitation method on a rotor test rig. The influences of swirl brake density, length, inlet/outlet pressure ratio, and rotating speed were measured and discussed, respectively. The CFD numerical results show that the swirl brake effectively reduces the seal swirl ratio (∼60–75% less), circumferential pressure difference (∼25–85% less) so that the seal destabilizing forces decrease. With the increasing of the swirl vanes density and length, the seal leakage drops (∼8–20% less). The experimental rotordynamic characteristics results show that it is more obvious to reduce the cross-couple stiffness (∼50–300% less) and increase the direct damping (∼50–60% larger) with the increasing in the number and length of the swirl vanes, and thus the swirl brake improves the seal rotordynamic stability. The efforts of this paper provide a useful insight to clearly understand the effects of swirl brakes on the labyrinth seal static and rotordynamic characteristics, which is beneficial to improve the design of annular seals.
Estimation of Forcing Function for a Geometrically Mistuned Bladed Rotor Via Modified Modal Domain Analysis
Authors: Vishwakarma Vinod , Sinha Alok
Published: September 2015

Modified modal domain analysis (MMDA) is a method to generate an accurate reduced-order model (ROM) of a bladed disk with geometric mistuning. An algorithm based on the MMDA ROM and a state observer is developed to estimate forcing functions for synchronous (including integer multiples) conditions from the dynamic responses obtained at few nodal locations of blades. The method is tested on a simple spring-mass model, finite element model (FEM) of a geometrically mistuned academic rotor, and FEM of a bladed rotor of an industrial-scale transonic research compressor. The accuracy of the forcing function estimation algorithm is examined by varying the order of ROM and the number of vibration output signals.
Bayesian Calibration for Power Splitting in Single-Shaft Combined Cycle Plant Diagnostics
Authors: Jiang Xiaomo , Lin TsungPo , Mendoza Eduardo
Published: September 2015

Condition monitoring and diagnostics of a combined cycle gas turbine (CCGT) power plant has become an important tool to improve its availability, reliability, and performance. However, there are two major challenges in the diagnostics of performance degradation and anomaly in a single-shaft combined cycle (CC) power plant. First, since the gas turbine (GT) and steam turbine (ST) in such a plant share a common generator, each turbine's contribution to the total plant power output is not directly measured, but must be accurately estimated to identify the possible causes of plant level degradation. Second, multivariate operational data instrumented from a power plant need to be used in the plant model calibration, power splitting, and degradation diagnostics. Sensor data always contain some degree of uncertainty. This adds to the difficulty of both estimation of GT to ST power split (PS) and degradation diagnostics. This paper presents an integrated probabilistic methodology for accurate power splitting and the degradation diagnostics of a single-shaft CC plant, accounting for uncertainties in the measured data. The method integrates the Bayesian inference approach, thermodynamic physics modeling, and sensed operational data seamlessly. The physics-based thermodynamic heat balance model is first established to model the power plant components and their thermodynamic relationships. The model is calibrated to model the plant performance at the design conditions of its main components. The calibrated model is then employed to simulate the plant performance at various operating conditions. A Bayesian inference method is next developed to determine the PS between the GT and the ST by comparing the measured and expected power outputs at different operation conditions, considering uncertainties in multiple measured variables. The calibrated model and calculated PS are further applied to pinpoint the possible causes at individual components resulting in the plant level degradation. The proposed methodology is demonstrated using operational data from a real-world single-shaft CC power plant with a known degradation issue. This study provides an effective probabilistic methodology to accurately split the power for degradation diagnostics of a single-shaft CC plant, addressing the uncertainties in multiple measured variables.
Nonviscous Modes of Nonproportionally Damped Viscoelastic Systems
Authors: Lázaro Mario
Published: September 2015

Nonviscously damped vibrating systems are characterized by dissipative mechanisms depending on the time-history of the response velocity, introduced in the physical models using convolution integrals involving hereditary kernel functions. One of the most used damping viscoelastic models is the Biot's model, whose hereditary functions are assumed to be exponential kernels. The free-motion equations of these types of nonviscous systems lead to a nonlinear eigenvalue problem enclosing certain number of the so-called nonviscous modes with nonoscillatory nature. Traditionally, the nonviscous modes (eigenvalues and eigenvectors) for nonproportional systems have been computed using the state-space approach, computationally expensive. In this paper, we address this problem developing a new method, computationally more efficient than that based on the state-space approach. It will be shown that real eigenvalues and eigenvectors of viscoelastically damped system can be obtained from a linear eigenvalue problem with the same size as the physical system. The numerical approach can even be enhanced to solve highly damped problems. The theoretical results are validated using a numerical example.
Crack Propagation Versus Fiber Alignment in Collagen Gels: Experiments and Multiscale Simulation
Authors: Vanderheiden Sarah M. , Hadi Mohammad F. , Barocas V. H.
Published: September 2015

It is well known that the organization of the fibers constituting a collagenous tissue can affect its failure behavior. Less clear is how that effect can be described computationally so as to predict the failure of a native or engineered tissue under the complex loading conditions that can occur in vivo. Toward the goal of a general predictive strategy, we applied our multiscale model of collagen gel mechanics to the failure of a double-notched gel under tension, comparing the results for aligned and isotropic samples. In both computational and laboratory experiments, we found that the aligned gels were more likely to fail by connecting the two notches than the isotropic gels. For example, when the initial notches were 30% of the sample width (normalized tip-to-edge distance = 0.7), the normalized tip-to-tip distance at which the transition occurred from between-notch failure to across-sample failure shifted from 0.6 to 1.0. When the model predictions for the type of failure event (between the two notches versus across the sample width) were compared to the experimental results, the two were found to be strongly covariant by Fisher’s exact test (p < 0.05) for both the aligned and isotropic gels with no fitting parameters. Although the double-notch system is idealized, and the collagen gel system is simpler than a true tissue, it presents a simple model system for studying failure of anisotropic tissues in a controlled setting. The success of the computational model suggests that the multiscale approach, in which the structural complexity is incorporated via changes in the model networks rather than via changes to a constitutive equation, has the potential to predict tissue failure under a wide range of conditions.
Allometry of the Tendon Enthesis: Mechanisms of Load Transfer Between Tendon and Bone
Authors: Deymier-Black Alix C. , Pasteris Jill D. , Genin Guy M. , Thomopoulos Stavros
Published: September 2015

Several features of the tendon-to-bone attachment were examined allometrically to determine load transfer mechanisms. The humeral head diameter increased geometrically with animal mass. Area of the attachment site exhibited a near isometric increase with muscle physiological cross section. In contrast, the interfacial roughness as well as the mineral gradient width demonstrated a hypoallometric relationship with physiologic cross-sectional area (PCSA). The isometric increase in attachment area indicates that as muscle forces increase, the attachment area increases accordingly, thus maintaining a constant interfacial stress. Due to the presence of constant stresses at the attachment, the micrometer-scale features may not need to vary with increasing load.
Low DC-bias silicon nitride anisotropic etching
Authors: Wang Yifei , Liu He , Li Yuanrui , Wu Wei
Published: September 2015

This paper presents an effective approach for silicon nitride plasma dry etching, with low DC-bias (self-bias) and vertical sidewalls. Four factors were studied: capacitively coupled radio frequency power, inductively coupled plasma power, pressure in the etching chamber, and the combination of etching gases. By meticulously controlling these factors, the authors successfully achieved low DC-bias silicon nitride anisotropic etching without argon bombardment, with DC-bias of only 34 V and etching rate of 40 nm/min. Our new etching recipe can avoid short-circuits effectively when patterning sub-50 nm thick silicon nitride layer as a dielectric layer between two metal layers. Moreover, “beaklike” structures, which were formed during the etching process due to increasing etching DC-bias at the beginning of the etching process, were discovered, understood, and prevented.
A model for the temporal evolution of the spatial coherence in decaying reverberant sound fields
Authors: Nees Sam , Schwarz Andreas , Kellermann Walter
Published: September 2015

Reverberant sound fields are often modeled as isotropic. However, it has been observed that spatial properties change during the decay of the sound field energy, due to non-isotropic attenuation in non-ideal rooms. In this letter, a model for the spatial coherence between two sensors in a decaying reverberant sound field is developed for rectangular rooms. The modeled coherence function depends on room dimensions, surface reflectivity, and orientation of the sensor pair, but is independent of the position of source and sensors in the room. The model includes the spherically isotropic (diffuse) and cylindrically isotropic sound field models as special cases.
Utility of Whole-Genome Sequencing of Escherichia coli O157 for Outbreak Detection and Epidemiological Surveillance
Authors: Holmes Anne , Allison Lesley , Ward Melissa , Dallman Timothy J. , Clark Richard , Fawkes Angie , Murphy Lee , Hanson Mary
Published: September 2015

Detailed laboratory characterization of
Escherichia coli
O157 is essential to inform epidemiological investigations. This study assessed the utility of whole-genome sequencing (WGS) for outbreak detection and epidemiological surveillance of
E. coli
O157, and the data were used to identify discernible associations between genotypes and clinical outcomes. One hundred five
E. coli
O157 strains isolated over a 5-year period from human fecal samples in Lothian, Scotland, were sequenced with the Ion Torrent Personal Genome Machine. A total of 8,721 variable sites in the core genome were identified among the 105 isolates; 47% of the single nucleotide polymorphisms (SNPs) were attributable to six “atypical”
E. coli
O157 strains and included recombinant regions. Phylogenetic analyses showed that WGS correlated well with the epidemiological data. Epidemiological links existed between cases whose isolates differed by three or fewer SNPs. WGS also correlated well with multilocus variable-number tandem repeat analysis (MLVA) typing data, with only three discordant results observed, all among isolates from cases not known to be epidemiologically related. WGS produced a better-supported, higher-resolution phylogeny than MLVA, confirming that the method is more suitable for epidemiological surveillance of
E. coli
O157. A combination of
in
silico
analyses (VirulenceFinder, ResFinder, and local BLAST searches) were used to determine
stx
subtypes, multilocus sequence types (15 loci), and the presence of virulence and acquired antimicrobial resistance genes. There was a high level of correlation between the WGS data and our routine typing methods, although some discordant results were observed, mostly related to the limitation of short sequence read assembly. The data were used to identify sublineages and clades of
E. coli
O157, and when they were correlated with the clinical outcome data, they showed that one clade, Ic3, was significantly associated with severe disease. Together, the results show that WGS data can provide higher resolution of the relationships between
E. coli
O157 isolates than that provided by MLVA. The method has the potential to streamline the laboratory workflow and provide detailed information for the clinical management of patients and public health interventions.
Haemophilus influenzae with Non-Beta-Lactamase-Mediated Beta-Lactam Resistance: Easy To Find but Hard To Categorize
Authors: Skaare Dagfinn , Lia Astrid , Hannisdal Anja , Tveten Yngvar , Matuschek Erika , Kahlmeter Gunnar , Kristiansen Bjørn-Erik
Published: September 2015

Haemophilus influenzae
is a major pathogen, and beta-lactams are first-line drugs. Resistance due to altered penicillin-binding protein 3 (rPBP3) is frequent, and susceptibility testing of such strains is challenging. A collection of 154 beta-lactamase-negative isolates with a large proportion of rPBP3 (67.5%) was used to evaluate and compare Etest (
Haemophilus
test medium [HTM]) and disk diffusion (EUCAST method) for categorization of susceptibility to aminopenicillins and cefuroxime, using MICs generated with broth (HTM) microdilution and clinical breakpoints from CLSI and EUCAST as the gold standards. In addition, the proficiency of nine disks in screening for the rPBP3 genotype (N526K positive) was evaluated. By Etest, both essential and categorical agreement were generally poor (<70%), with high very major errors (VME) (CLSI, 13.0%; EUCAST, 34.3%) and falsely susceptible rates (FSR) (CLSI, 87.0%; EUCAST, 88.3%) for ampicillin. Ampicillin (2 μg) with adjusted (+2 mm) zone breakpoints was superior to Etest for categorization of susceptibility to ampicillin (agreement, 74.0%; VME, 11.0%; FSR, 28.3%). Conversely, Etest was superior to 30 μg cefuroxime for categorization of susceptibility to cefuroxime (agreement, 57.1% versus 60.4%; VME, 2.6% versus 9.7%; FSR, 7.1% versus 26.8%). Benzylpenicillin (1 unit) (EUCAST screening disk) and cefuroxime (5 μg) identified rPBP3 isolates with highest accuracies (95.5% and 92.2%, respectively). In conclusion, disk screening reliably detects rPBP3
H. influenzae
, but false ampicillin susceptibility is frequent with routine methods. We suggest adding a comment recommending high-dose aminopenicillin therapy or the use of other agents for severe infections with screening-positive isolates that are susceptible to aminopenicillins by gradient or disk diffusion.
Significance of a Positive Toxoplasma Immunoglobulin M Test Result in the United States
Authors: Dhakal Reshika , Gajurel Kiran , Pomares Christelle , Talucod Jeanne , Press Cynthia J. , Montoya Jose G.
Published: September 2015

A positive
Toxoplasma
immunoglobulin M (IgM) result is often interpreted as a marker of an acute infection. However, IgM can persist for several years, and
Toxoplasma
commercial IgM diagnostic test kits can yield a number of false-positive results. For these reasons, a chronic
Toxoplasma
infection can be erroneously classified as an acute infection, resulting in serious adverse consequences, especially in pregnant women, leading to emotional distress and unnecessary interventions, including termination of pregnancy. Interpretation of
Toxoplasma
serology at a reference laboratory can help differentiate a recently acquired infection from a chronic infection. Serological test results for 451 patients with positive
Toxoplasma
IgM and IgG test results obtained at nonreference laboratories (NRLs) that were referred to Palo Alto Medical Foundation Toxoplasma Serology Laboratory (PAMF-TSL) to determine whether the patient was acutely or chronically infected were retrospectively reviewed. PAMF-TSL results established that of the 451 patients, 335 (74%) had a chronic infection, 100 (22%) had an acute infection, and 7 (2%) were not infected, and for 9 (2%), results were indeterminate. Positive
Toxoplasma
IgM and IgG test results obtained at NRLs cannot accurately distinguish between acute and chronic infections. To do so, testing at reference laboratories is required, as mandated in 1997 in a letter from the Food and Drug Administration (FDA) to clinicians and laboratories in the United States.
Optimal Cutoff Titers for Indirect Immunofluorescence Assay for Diagnosis of Scrub Typhus
Authors: Lim Cherry , Blacksell Stuart D. , Laongnualpanich Achara , Kantipong Pacharee , Day Nicholas P. J. , Paris Daniel H. , Limmathurotsakul Direk
Published: September 2015

We determined the optimal cutoff titers in admission and convalescent-phase samples for scrub typhus indirect immunofluorescence assay using Bayesian latent class models. Cutoff titers of ≥1:3,200 in an admission sample or of a ≥4-fold rise to ≥1:3,200 in a convalescent-phase sample provided the highest accuracy (sensitivity, 81.6%; specificity, 100%).
Comparing Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry and Phenotypic and Molecular Methods for Identification of Species within the Streptococcus anginosus Group
Authors: Arinto-Garcia Raquel , Pinho Marcos Daniel , Carriço João André , Melo-Cristino José , Ramirez Mário
Published: September 2015

The heterogeneity of members of the
Streptococcus anginosus
group (SAG) has traditionally hampered their correct identification. Recently, the group was subdivided into 6 taxa whose prevalence among human infections is poorly described. We evaluated the accuracy of the Rapid ID32 Strep test, matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS), and a PCR multiplex method to identify 212 SAG isolates recovered from human infections to the species and subspecies level by using multilocus sequence analysis (MLSA) as the gold standard. We also determined the antimicrobial susceptibilities of the isolates. Representatives of all SAG taxa were found among our collection. MALDI-TOF MS and the Rapid ID32 Strep test correctly identified 92% and 68% of the isolates to the species level, respectively, but showed poor performance at the subspecies level, and the latter was responsible for major identification errors. The multiplex PCR method results were in complete agreement with the MLSA identifications but failed to distinguish the subspecies
Streptococcus constellatus
subsp.
pharyngis
and
S. constellatus
subsp.
viborgensis
. A total of 145 MLSA sequence types were present in our collection, indicating that within each taxon a number of different lineages are capable of causing infection. Significant antibiotic resistance was observed only to tetracycline, erythromycin, and clindamycin and was present in most taxa. MALDI-TOF MS is a reliable method for routine SAG species identification, while the need for identification to the subspecies level is not clearly established.
Cooperation between the Hepatitis C Virus p7 and NS5B Proteins Enhances Virion Infectivity
Authors: Aligeti Mounavya , Roder Allison , Horner Stacy M.
Published: September 2015

The molecular mechanisms that govern hepatitis C virus (HCV) assembly, release, and infectivity are still not yet fully understood. In the present study, we sequenced a genotype 2A strain of HCV (JFH-1) that had been cell culture adapted in Huh-7.5 cells to produce nearly 100-fold-higher viral titers than the parental strain. Sequence analysis identified nine mutations in the genome, present within both the structural and nonstructural genes. The infectious clone of this virus containing all nine culture-adapted mutations had 10-fold-higher levels of RNA replication and RNA release into the supernatant but had nearly 1,000-fold-higher viral titers, resulting in an increased specific infectivity compared to wild-type JFH-1. Two mutations, identified in the p7 polypeptide and NS5B RNA-dependent RNA polymerase, were sufficient to increase the specific infectivity of JFH-1. We found that the culture-adapted mutation in p7 promoted an increase in the size of cellular lipid droplets following transfection of viral RNA. In addition, we found that the culture-adaptive mutations in p7 and NS5B acted synergistically to enhance the specific viral infectivity of JFH-1 by decreasing the level of sphingomyelin in the virion. Overall, these results reveal a genetic interaction between p7 and NS5B that contributes to virion specific infectivity. Furthermore, our results demonstrate a novel role for the RNA-dependent RNA polymerase NS5B in HCV assembly.

IMPORTANCE
Hepatitis C virus assembly and release depend on viral interactions with host lipid metabolic pathways. Here, we demonstrate that the viral p7 and NS5B proteins cooperate to promote virion infectivity by decreasing sphingomyelin content in the virion. Our data uncover a new role for the viral RNA-dependent RNA polymerase NS5B and p7 proteins in contributing to virion morphogenesis. Overall, these findings are significant because they reveal a genetic interaction between p7 and NS5B, as well as an interaction with sphingomyelin that regulates virion infectivity. Our data provide new strategies for targeting host lipid-virus interactions as potential targets for therapies against HCV infection.
Exclusive Decoration of Simian Immunodeficiency Virus Env with High-Mannose Type N-Glycans Is Not Compatible with Mucosal Transmission in Rhesus Macaques
Authors: Karsten Christina B. , Buettner Falk F. R. , Cajic Samanta , Nehlmeier Inga , Neumann Berit , Klippert Antonina , Sauermann Ulrike , Reichl Udo , Gerardy-Schahn Rita , Rapp Erdmann , Stahl-Hennig Christiane , Pöhlmann Stefan
Published: September 2015

Human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) envelope (Env) proteins are extensively decorated with N-glycans, predominantly of the high-mannose type. However, it is unclear how high-mannose N-glycans on Env impact viral spread. We show that exclusive modification of SIV Env with these N-glycans reduces viral infectivity and abrogates mucosal transmission, despite increasing viral capture by immune cell lectins. Thus, high-mannose N-glycans have opposed effects on SIV infectivity and lectin reactivity, and a balance might be required for efficient mucosal transmission.
Kaposi's Sarcoma-Associated Herpesvirus Viral Interferon Regulatory Factor 1 Interacts with a Member of the Interferon-Stimulated Gene 15 Pathway
Authors: Jacobs Sarah R. , Stopford Charles M. , West John A. , Bennett Christopher L. , Giffin Louise , Damania Blossom
Published: September 2015

Kaposi's sarcoma-associated herpesvirus (KSHV) is a gammaherpesvirus known to establish lifelong latency in the human host. We and others have previously shown that three KSHV homologs of cellular interferon regulatory factors (IRFs), known as viral IRFs (vIRFs), participate in evasion of the host interferon (IFN) response. We report that vIRF1 interacts with the cellular interferon-stimulated gene 15 (ISG15) E3 ligase, HERC5, in the context of Toll-like receptor 3 (TLR3) activation and IFN induction. The ISG15 protein is covalently conjugated to target proteins upon activation of the interferon response. Interaction between vIRF1 and HERC5 was confirmed by immunoprecipitation, and the region between amino acids 224 and 349 of vIRF1 was required for interaction with HERC5. We further report that expression of vIRF1 in the context of TLR3 activation results in decreased ISG15 conjugation of proteins. Specifically, TLR3-induced ISG15 conjugation and protein levels of cellular IRF3, a known ISG15 target, were decreased in the presence of vIRF1 compared to the control. vIRF1 itself was also identified as a target of ISG15 conjugation. KSHV-infected cells exhibited increased ISG15 conjugation upon reactivation from latency in coordination with increased IFN. Furthermore, knockdown of ISG15 in latently infected cells resulted in a higher level of KSHV reactivation and an increase in infectious virus. These data suggest that the KSHV vIRF1 protein affects ISG15 conjugation and interferon responses and may contribute to effective KSHV replication.

IMPORTANCE
The KSHV vIRF1 protein can inhibit interferon activation in response to viral infection. We identified a cellular protein named HERC5, which is the major ligase for ISG15, as a vIRF1 binding partner. vIRF1 association with HERC5 altered ISG15 modification of cellular proteins, and knockdown of ISG15 augmented reactivation of KSHV from latency.
Ecuador Paraiso Escondido Virus, a New Flavivirus Isolated from New World Sand Flies in Ecuador, Is the First Representative of a Novel Clade in the Genus Flavivirus
Authors: Alkan Cigdem , Zapata Sonia , Bichaud Laurence , Moureau Grégory , Lemey Philippe , Firth Andrew E. , Gritsun Tamara S. , Gould Ernest A. , de Lamballerie Xavier , Depaquit Jérôme , Charrel Rémi N.
Published: September 2015

A new flavivirus, Ecuador Paraiso Escondido virus (EPEV), named after the village where it was discovered, was isolated from sand flies (
Psathyromyia abonnenci
, formerly
Lutzomyia abonnenci
) that are unique to the New World. This represents the first sand fly-borne flavivirus identified in the New World. EPEV exhibited a typical flavivirus genome organization. Nevertheless, the maximum pairwise amino acid sequence identity with currently recognized flaviviruses was 52.8%. Phylogenetic analysis of the complete coding sequence showed that EPEV represents a distinct clade which diverged from a lineage that was ancestral to the nonvectored flaviviruses Entebbe bat virus, Yokose virus, and Sokoluk virus and also the
Aedes
-associated mosquito-borne flaviviruses, which include yellow fever virus, Sepik virus, Saboya virus, and others. EPEV replicated in C6/36 mosquito cells, yielding high infectious titers, but failed to reproduce either in vertebrate cell lines (Vero, BHK, SW13, and XTC cells) or in suckling mouse brains. This surprising result, which appears to eliminate an association with vertebrate hosts in the life cycle of EPEV, is discussed in the context of the evolutionary origins of EPEV in the New World.

IMPORTANCE
The flaviviruses are rarely (if ever) vectored by sand fly species, at least in the Old World. We have identified the first representative of a sand fly-associated flavivirus, Ecuador Paraiso Escondido virus (EPEV), in the New World. EPEV constitutes a novel clade according to current knowledge of the flaviviruses. Phylogenetic analysis of the virus genome showed that EPEV roots the
Aedes
-associated mosquito-borne flaviviruses, including yellow fever virus. In light of this new discovery, the New World origin of EPEV is discussed together with that of the other flaviviruses.
The Nucleoprotein Is Required for Lymphocytic Choriomeningitis Virus-Based Vaccine Vector Immunogenicity
Authors: Darbre Stephanie , Johnson Susan , Kallert Sandra , Lambert Paul-Henri , Siegrist Claire-Anne , Pinschewer Daniel D.
Published: September 2015

Recombinant glycoprotein-deficient lymphocytic choriomeningitis virus-based vaccine vectors (rLCMV/ΔGP) are potent CD8
+
T cell inducers. To investigate the underlying molecular requirements, we generated a nucleoprotein-deficient vector counterpart (rLCMV/ΔNP). NP but not GP is a minimal
trans
-acting factor for viral transcription and genome replication. We found that, unlike rLCMV/ΔGP, rLCMV/ΔNP failed to elicit detectable CD8
+
T cell responses unless NP was
trans
complemented in a transgenic host. Hence, NP-dependent intracellular gene expression is essential for LCMV vector immunogenicity.
Consequences of HLA-B*13-Associated Escape Mutations on HIV-1 Replication and Nef Function
Authors: Shahid Aniqa , Olvera Alex , Anmole Gursev , Kuang Xiaomei T. , Cotton Laura A. , Plana Montserrat , Brander Christian , Brockman Mark A. , Brumme Zabrina L.
Published: September 2015

HLA-B*13 is associated with superiorin vivoHIV-1 viremia control. Protection is thought to be mediated by sustained targeting of key cytotoxic T lymphocyte (CTL) epitopes and viral fitness costs of CTL escape in Gag although additional factors may contribute. We assessed the impact of 10 published B*13-associated polymorphisms in Gag, Pol, and Nef, in 23 biologically relevant combinations, on HIV-1 replication capacity and Nef-mediated reduction of cell surface CD4 and HLA class I expression. Mutations were engineered into HIV-1NL4.3, and replication capacity was measured using a green fluorescent protein (GFP) reporter T cell line. Nef-mediated CD4 and HLA-A*02 downregulation was assessed by flow cytometry, and T cell recognition of infected target cells was measured via coculture with an HIV-specific luciferase reporter cell line. When tested individually, only Gag-I147L and Gag-I437L incurred replicative costs (5% and 17%, respectively), consistent with prior reports. The Gag-I437L-mediated replication defect was rescued to wild-type levels by the adjacent K436R mutation. A novel B*13 epitope, comprising 8 residues and terminating at Gag147, was identified in p24Gag(GQMVHQAIGag140–147). No other single or combination Gag, Pol, or Nef mutant impaired viral replication. Single Nef mutations did not affect CD4 or HLA downregulation; however, the Nef double mutant E24Q-Q107R showed 40% impairment in HLA downregulation with no evidence of Nef stability defects. Moreover, target cells infected with HIV-1-NefE24Q-Q107Rwere recognized better by HIV-specific T cells than those infected with HIV-1NL4.3or single Nef mutants. Our results indicate that CTL escape in Gag and Nef can be functionally costly and suggest that these effects may contribute to long-term HIV-1 control by HLA-B*13.

IMPORTANCEProtective effects of HLA-B*13 on HIV-1 disease progression are mediated in part by fitness costs of CTL escape mutations in conserved Gag epitopes, but other mechanisms remain incompletely known. We extend our knowledge of the impact of B*13-driven escape on HIV-1 replication by identifying Gag-K436R as a compensatory mutation for the fitness-costly Gag-I437L. We also identify Gag-I147L, the most rapidly and commonly selected B*13-driven substitution in HIV-1, as a putative C-terminal anchor residue mutation in a novel B*13 epitope. Most notably, we identify a novel escape-driven fitness defect: B*13-driven substitutions E24Q and Q107R in Nef, when present together, substantially impair this protein's ability to downregulate HLA class I. This, in turn, increases the visibility of infected cells to HIV-specific T cells. Our results suggest that B*13-associated escape mutations impair HIV-1 replication by two distinct mechanisms, that is, by reducing Gag fitness and dampening Nef immune evasion function.
Successive Respiratory Syncytial Virus Epidemics in Local Populations Arise from Multiple Variant Introductions, Providing Insights into Virus Persistence
Authors: Agoti Charles N. , Otieno James R. , Ngama Mwanajuma , Mwihuri Alexander G. , Medley Graham F. , Cane Patricia A. , Nokes D. James
Published: September 2015

Respiratory syncytial virus (RSV) is a global respiratory pathogen of humans, with infection occurring characteristically as recurrent seasonal epidemics. Unlike influenza viruses, little attention has been paid to the mechanism underlying worldwide spread and persistence of RSV and how this may be discerned through an improved understanding of the introduction and persistence of RSV in local communities. We analyzed 651 attachment (G) glycoprotein nucleotide sequences of RSV B collected over 11 epidemics (2002 to 2012) in Kilifi, Kenya, and contemporaneous data collected elsewhere in Kenya and 18 other countries worldwide (2002 to 2012). Based on phylogeny, genetic distance and clustering patterns, we set out pragmatic criteria to classify local viruses into distinct genotypes and variants, identifying those newly introduced and those locally persisting. Three genotypes were identified in the Kilifi data set: BA (
n
= 500), SAB1 (
n
= 148), and SAB4 (
n
= 3). Recurrent RSV epidemics in the local population were composed of numerous genetic variants, most of which have been newly introduced rather than persisting in the location from season to season. Global comparison revealed that (i) most Kilifi variants do not cluster closely with strains from outside Kenya, (ii) some Kilifi variants were closely related to those observed outside Kenya (mostly Western Europe), and (iii) many variants were circulating elsewhere but were never detected in Kilifi. These results are consistent with the hypothesis that year-to-year presence of RSV at the local level (i.e., Kilifi) is achieved primarily, but not exclusively, through introductions from a pool of variants that are geographically restricted (i.e., to Kenya or to the region) rather than global.

IMPORTANCE
The mechanism by which RSV persists and reinvades local populations is poorly understood. We investigated this by studying the temporal patterns of RSV variants in a rural setting in tropical Africa and comparing these variants with contemporaneous variants circulating in other countries. We found that periodic seasonal RSV transmission at the local level appears to require regular new introductions of variants. However, importantly, the evidence suggests that the source of new variants is mostly geographically restricted, and we hypothesize that year-to-year RSV persistence is at the country level rather than the global level. This has implications for control.
Monitoring the athlete training response: subjective self-reported measures trump commonly used objective measures: a systematic review
Authors: Saw Anna E , Main Luana C , Gastin Paul B
Published: September 2015

Background

Monitoring athlete well-being is essential to guide training and to detect any progression towards negative health outcomes and associated poor performance. Objective (performance, physiological, biochemical) and subjective measures are all options for athlete monitoring.

Objective

We systematically reviewed objective and subjective measures of athlete well-being. Objective measures, including those taken at rest (eg, blood markers, heart rate) and during exercise (eg, oxygen consumption, heart rate response), were compared against subjective measures (eg, mood, perceived stress). All measures were also evaluated for their response to acute and chronic training load.

Methods

The databases Academic search complete, MEDLINE, PsycINFO, SPORTDiscus and PubMed were searched in May 2014. Fifty-six original studies reported concurrent subjective and objective measures of athlete well-being. The quality and strength of findings of each study were evaluated to determine overall levels of evidence.

Results

Subjective and objective measures of athlete well-being generally did not correlate. Subjective measures reflected acute and chronic training loads with superior sensitivity and consistency than objective measures. Subjective well-being was typically impaired with an acute increase in training load, and also with chronic training, while an acute decrease in training load improved subjective well-being.

This review provides further support for practitioners to use subjective measures to monitor changes in athlete well-being in response to training. Subjective measures may stand alone, or be incorporated into a mixed methods approach to athlete monitoring, as is current practice in many sport settings.
Comparative efficacy and tolerability of first-generation and newer-generation antidepressant medications for depressive disorders in children and adolescents: study protocol for a systematic review and network meta-analysis
Authors: Zhou Xinyu , Qin Bin , Whittington Craig , Cohen David , Liu Yiyun , Del Giovane Cinzia , Michael Kurt D , Zhang Yuqing , Xie Peng
Published: September 2015

Introduction

Depressive disorders are among the most common psychiatric disorders in children and adolescents, and have adverse effects on their psychosocial functioning. Questions concerning the efficacy and safety of antidepressant medications in the treatment of depression in children and adolescents, led us to integrate the direct and indirect evidence using network meta-analysis to create hierarchies of these drugs.

Methods and analysis

Seven databases with PubMed, EMBASE, the Cochrane Library, Web of Science, CINAHL, LiLACS and PsycINFO will be searched from 1966 to December 2013 (updated to May, 2015). There are no restrictions on language or type of publication. Randomised clinical trials assessing first-generation and newer-generation antidepressant medications against active comparator or placebo as acute treatment for depressive disorders in children and adolescents (under 18 years of age) will be included. The primary outcome for efficacy will be mean improvement in depressive symptoms, as measured by the mean change score of a depression rating scale from baseline to post-treatment. The tolerability of treatment will be defined as side effect discontinuation, as defined by the proportion of patients who discontinued treatment due to adverse events during the trial. We will also assess the secondary outcome for efficacy (response rate), acceptability (all-cause discontinuation) and suicide-related outcomes. We will perform the Bayesian network meta-analyses for all relative outcome measures. Subgroup analyses and sensitivity analyses will be conducted to assess the robustness of the findings.

Dissemination

The network meta-analysis will provide useful information on antidepressant treatment for child and adolescent depression. The results will be disseminated through peer-reviewed publication or conference presentations.

Trial registration number

PROSPERO CRD42015016023.
Carpal tunnel syndrome and computer exposure at work in two large complementary cohorts
Authors: Mediouni Z , Bodin J , Dale A M , Herquelot E , Carton M , Leclerc A , Fouquet N , Dumontier C , Roquelaure Y , Evanoff B A , Descatha A
Published: September 2015

Objectives

The boom in computer use and concurrent high rates in musculoskeletal complaints and carpal tunnel syndrome (CTS) among users have led to a controversy about a possible link. Most studies have used cross-sectional designs and shown no association. The present study used longitudinal data from two large complementary cohorts to evaluate a possible relationship between CTS and the performance of computer work.

Settings and participants

The Cosali cohort is a representative sample of a French working population that evaluated CTS using standardised clinical examinations and assessed self-reported computer use. The PrediCTS cohort study enrolled newly hired clerical, service and construction workers in several industries in the USA, evaluated CTS using symptoms and nerve conduction studies (NCS), and estimated exposures to computer work using a job exposure matrix.

Primary and secondary outcome measures

During a follow-up of 3–5 years, the association between new cases of CTS and computer work was calculated using logistic regression models adjusting for sex, age, obesity and relevant associated disorders.

Results

In the Cosali study, 1551 workers (41.8%) completed follow-up physical examinations; 36 (2.3%) participants were diagnosed with CTS. In the PrediCTS study, 711 workers (64.2%) completed follow-up evaluations, whereas 31 (4.3%) had new cases of CTS. The adjusted OR for the group with the highest exposure to computer use was 0.39 (0.17; 0.89) in the Cosali cohort and 0.16 (0.05; 0.59) in the PrediCTS cohort.

Conclusions

Data from two large cohorts in two different countries showed no association between computer work and new cases of CTS among workers in diverse jobs with varying job exposures. CTS is far more common among workers in non-computer related jobs; prevention efforts and work-related compensation programmes should focus on workers performing forceful hand exertion.
Impact of multiple liver resections prior to salvage liver transplantation on survival in patients with recurrent HCC
Authors: Hu Zhenhua , Zhang Qijun , Zhou Jie , Li Zhiwei , Xiang Jie , Zhou Lin , Wu Jian , Zhang Min , Zheng Shusen
Published: September 2015

Objectives

Salvage liver transplantation (SLT) is a controversial technique that has been reported to be acceptable for the management of patients with recurrent hepatocellular carcinoma (HCC) after primary hepatic resection (HR). However, whether the number of times liver resection is performed has an impact on survival after SLT has not yet been reported.

Design

Retrospective study.

Setting

The level of care is primary and the study was carried out at only 1 centre.

Participants

The study included 59 patients who underwent SLT for HCC from September 2001 to December 2012. 51 patients underwent HR only once before SLT, while the remaining 8 patients underwent HR more than once before SLT (HR=2 [7], HR=3, [1]).

Primary and secondary outcome measures

In this study, the 1-year, 3-year and 5-year overall and tumour-free survival outcomes between the 2 groups were compared.

Results

There were no significant differences between patients who underwent HR once and those who underwent HR more than once with respect to overall or tumour-free survival after receiving SLT. The 1-year, 3-year and 5-year overall survival rates for patients who underwent HR once were 72.9%, 35.3% and 35.5% vs 50%, 50% and 50%, respectively (p=0.986), while the 1-year, 3-year and 5-year tumour-free survival rates for those who underwent HR more than once were 66.3%, 55.3% and 44.4% vs 40%, 40% and 40%, respectively (p=0.790).

Conclusions

There was no significant difference in the survival rate of patients who underwent HR once before SLT and those who underwent HR more than once. This suggests that SLT is a reasonable choice for patients who suffer from recurrent HCC after HR.

Trial registration number

This is a retrospective study and no registry or number is required.