bluuebunny/crossref_metadata_2025 huggingface dataset:
Study on Validity of Stress Intensity Factor in CT Specimen for Materials With Low Yield Stress
Authors: Takanashi Masahiro , Itabashi Yu , Hirano Takashi
Published: January 2014

This paper discusses the validity of a stress intensity factor K for compact tension (CT) specimens of stainless and low-alloy steels by computing J-integral values. In fracture mechanics testing, specimen configurations and loading conditions are strictly regulated to meet the small-scale yielding (SSY) condition and obtain valid data. For some materials, particularly those without an obvious yield point, it is hard to meet the SSY condition. The regulations specified in fracture mechanics test standards are based on experimental results. The theoretical reason for the SSY condition is not necessarily clear and the condition seems conservative.

This study focuses on a correlation with the stress intensity factors K and the J-integral values under the SSY condition. J-integral values of CT specimens were analyzed by elasto-plastic finite element analysis. Subsequently, the validity of the stress intensity factor K was checked by comparing K and J–integral values to determine whether CT specimens were under the SSY condition or not. To simplify the comparison, J-integral values were converted to K values, equivalent stress intensity factor KJ values.

When K values were low enough to meet the SYY condition of the specimen, they equaled KJ values. Meanwhile, KJ values exceed K values in a high K region, which means the specimen is no longer under a SSY condition. It is possible to determine whether the specimen is under the SSY condition or not by comparing the values of KJ and K. This paper evaluated the validity of K values using the correlative relationship and showed that they were valid to some extent even if loading conditions and specimen configurations were outside the scope of the specification by the test standards. The validity of K values with 0.2% offset strength or flow strength instead of yield stress is also confirmed by this method.
Influence of Pump-Circuit Coupling on Acoustic Waves in Pipelines and Pump Velocity Fields
Authors: Keller Jens , Parrondo Jorge , Blanco Eduardo , Barrio Raúl , Suárez Carlos
Published: January 2014

Hydraulic piping systems are sensitive to excessive acoustic emissions which can give rise to vibrations or to failure of mechanical elements due to fatigue. Common excitation sources are centrifugal pumps due to the periodic interaction of the impeller with the volute tongue. They radiate pressure fluctuations into the connected circuit at the blade-passing frequency which are reflected in the circuit. The aim of the present investigation was the experimental characterization of the perturbations induced in a piping network as a function of the acoustic impedance of the circuit using fast-response pressure transducers. Three transducers were placed along the discharge pipe to decompose the pressure signal into the radiated and reflected acoustic wave, with amplitude and phase. The speed of sound in the water pipelines was determined experimentally. Results of impedance at the pump tongue were compared with a theoretical approach, using a Transfer Matrix Analysis, where each pipe element is represented by a 2×2-matrix, relating the acoustic pressure and velocity fluctuations at the two ports. Attenuation was considered by using complex wavenumbers. The acoustic impedance was changed by using different rotation speeds and by using a cavity which works as a harmonic oscillator whose resonance frequency can be changed. The results presented for different operating points show the influence of changing impedance on the pressure perturbations due to pump-circuit acoustic coupling. The pump was built with a transparent impeller and volute to conduct Particle Image Velocimetry (PIV) measurements in a plane perpendicular to the pump rotation axis. Phase-averaged velocity fields were obtained at different blade positions in the zone around the tongue. Vorticity fields were derived from it and turbulence is shown by representing in-plane turbulent kinetic energy (TKE). Strong vorticity and turbulence occurred in the volute channel behind the blade and near the tongue tip.
Different Microstructures in the HAZ of Double Submerged Arc Welded Pipelines and How They Influence the Fatigue Crack Growth
Authors: Maier Bernd , Guster Christoph , Tichy Richard , Ecker Werner
Published: January 2014

To meet the cost and weight reduction requirements in welded structures, the application of high-strength low-alloy (HSLA) steels also increases in the field of pipeline engineering. However, in comparison to low-strength steels these materials are more susceptible to notches or initial cracks. Therefore, an extensive investigation of fatigue crack growth behaviour in the different microstructural zones of the heat-affected zone (HAZ) is necessary for an appropriate lifetime evaluation of welded pipes. For this purpose, fracture mechanics parameters like thresholds and crack growth rates in the three characteristic microstructural zones of the HAZ as well as in the base material are characterized. The different microstructures are reproduced with a Gleeble thermal simulator system using the same heating curves as measured previously during the welding process of a double submerged arc welded pipeline. The effect of varying stress ratios R on the crack growth threshold and the crack growth curves is also assessed. This knowledge about the mean stress sensitivity is needed for estimating the influence of residual stresses stemming from different manufacturing processes like welding and post-welding treatments, which is essential for a reliable, damage tolerant application of HSLA steels in pipelines.
Numerical Analyses of Interaction Behavior of Multiple Surface Cracks Using a Modified Creep-Damage Model and Fracture Mechanics Approach
Authors: Wen Jian-Feng , Xuan Fu-Zhen , Tu Shan-Tung
Published: January 2014

The propagation and coalescence processes of two initially semi-elliptical surface cracks in a plate at high temperature are predicted by using a modified creep-damage model implemented with the numerical method. The creep growth of multiple cracks during pre-coalescence and coalescence periods is investigated in detail. For the purpose of comparison, the step-by-step fracture-mechanics-based finite element technique is also employed. The results indicated that, before coalescence, the cracks grow steadily but relatively rapidly in the adjacent district; after the cracks coalesce, the concave positions of the newly combined crack front exhibit a considerably high growth rate. Compared with the fracture mechanics approach, the numerical approach based on the modified creep-damage model enables to give more detailed information on creep crack initiation and more reliable estimate of propagation. In addition, the combination rules given by existing standards are also examined.
Fracture Assessment of X65 Steel Pipe With Through-Wall Crack Under Pure Bending
Authors: Mourad A.-H. I. , Altarawneh J. , El Domiaty A.
Published: January 2014

Stable crack growth emanated from circumferential through-wall crack in X65 steel pipe under pure bending was evaluated. Seamless carbon steel pipes grade X65 PL5 with an inner diameter of 42.9 mm and an outer diameter of 60.3 mm were used in the present experimental work. This type of pipes is of common use in piping systems of gas and oil industry. Pipes of different circumferential crack depths were tested. J-R curve was used to characterize the nonlinear fracture behavior of the through-wall crack pipe. Variation of J with crack growth, crack surface area and deflection of the pipe were obtained. J-integral-crack growth curve was used in the calculation of critical crack size based on the Jc value measured in our previous experimental work by testing full scale pipe made from the same material. The length of crack at unstable crack growth is also determined. Both critical and instability crack sizes can be used for fracture assessment of X65 steel pipes.
The Long Journey in the Development of ISI Rules for CASS Piping
Authors: Chockie Alan , Griesbach Timothy J.
Published: January 2014

The factors that make cast austenitic stainless steel (CASS) a good candidate for use in primary piping systems have significantly hampered the ability to detect, locate, and size flaws within the material. The difficulties in effectively inspecting CASS piping has been mirrored in the challenges associated with the development of CASS inspection requirements.

Since the early 1970’s the United States Federal Regulations have made the Section XI inservice inspection (ISI) rules of American Society of Mechanical Engineers (ASME) Boiler and Pressure Vessel Code a mandatory requirement for all operating nuclear power plants. Section XI of the ASME Boiler and Pressure Vessel Code requires that the pressure boundary system of all light water reactors be inspected using a volumetric technique. However, the inhomogeneous coarse-grain structure and elastic anisotropy of the CASS material can produce beam skewing and distortion, high background noise, and uncertainty in the propagation velocity of the ultrasound beam.

The difficulties associated with reliability detecting and sizing flaws in CASS material have been well known since the late 1970’s. To date there are no qualified procedures, equipment, or personnel or performance demonstration requirements for the UT examination of CASS pressure-retaining welds. Until Section XI, Appendix VIII qualification requirements are developed licensees will continue to follow the rules of Appendix III for the inspection of CASS material.

In recent years there have been significant advances in the development of Section XI requirements for the inspection of CASS. This paper reviews the long and difficult journey, the changing strategies and approaches, the driving forces, and the anticipated new Code requirements for the inspection of CASS.
Buckling of High-Strength Steel Cylinders Under Cyclic Bending in the Inelastic Range
Authors: Varelis George E. , Karamanos Spyros A.
Published: January 2014

The present paper examines the structural behavior of high-strength elongated steel cylinders, referred to as tubes or pipes, subjected to strong cyclic bending, through a rigorous finite element simulation. The cylinder exhibits cross-sectional distortion, in the form of ovalization, combined with excessive plastic deformations. Those deformations, under repeated loading, may lead to instability in the form of local buckling (wrinkling) and, eventually, failure of the loaded member. The study focuses on relatively-thick-walled cylindrical members, which exhibit local buckling in the plastic range of the steel material, with the use of advanced nonlinear finite element models able to describe both geometrical and material nonlinearities. A cyclic plasticity model that adopts the “Bounding Surface” concept is employed, calibrated through special-purpose material testing, and implemented within ABAQUS, using a user-subroutine. The numerical model is validated by comparison with available experimental data on tubular members made of high-strength steel. Finally, a parametric numerical study is conducted, aimed at determining the effects of geometrical imperfections of the cylinder on its plastic buckling performance.
Chemical bonding and defect states of LPCVD grown silicon-rich Si3N4 for quantum dot applications
Authors: Mohammed Shakil , Nimmo Michael T. , Malko Anton V. , Hinkle Christopher L.
Published: January 2014

Si-rich Si3N4 (SRN) thin films were investigated to understand the various defect states present within the SRN that can lead to reduced performance in quantum dot based devices made of these materials. The SRN films, deposited by low pressure chemical vapor deposition followed by furnace anneals over a range of temperatures, were determined to be comprised of two distinct phase separated SRN regions with different compositions (precipitates within a host matrix). Photoluminescence (PL) spectra showed multiple peaks convoluted together within the visible and near-visible range. Depending on deposition and annealing conditions, the films displayed changes in PL peak intensities which were correlated with chemical bonding utilizing x-ray photoelectron spectroscopy, Fourier transform infrared spectroscopy, spectroscopic ellipsometry, and capacitance–voltage measurements. It is found that the PL originates from defect-state to defect-state and band edge to defect-state electronic transitions.
Ti/Al/Ti/Ni/Au ohmic contacts on AlGaN/GaN high electron mobility transistors with improved surface morphology and low contact resistance
Authors: Chiu Yu-Sheng , Lin Tai-Ming , Nguyen Hong-Quan , Weng Yu-Chen , Nguyen Chi-Lang , Lin Yueh-Chin , Yu Hung-Wei , Yi Chang Edward , Lee Ching-Ting
Published: January 2014

Optimizing surface morphology of ohmic contacts on GaN high electron mobility transistors continues to be a challenge in the GaN electronics industry. In this study, a variety of metal schemes were tested under various annealing conditions to obtain contacts with optimal qualities. A Ti/Al/Ti/Ni/Au (20/120/40/60/50 nm) metal scheme demonstrated the lowest contact resistance (Rc) and a smooth surface morphology, and the mechanisms were investigated by materials analysis. A Ti/Al/Ti/Ni/Au metal scheme with optimized Ti and Ni thicknesses can result in formation of a larger proportion of Al-Ni intermetallics and a continuous TiN interlayer, which results in smooth surface and low Rc.
Comparison of 2D and 3D gamma analyses
Authors: Pulliam Kiley B. , Huang Jessie Y. , Howell Rebecca M. , Followill David , Bosca Ryan , O’Daniel Jennifer , Kry Stephen F.
Published: January 2014

Purpose:

As clinics begin to use 3D metrics for intensity‐modulated radiation therapy (IMRT) quality assurance, it must be noted that these metrics will often produce results different from those produced by their 2D counterparts. 3D and 2D gamma analyses would be expected to produce different values, in part because of the different search space available. In the present investigation, the authors compared the results of 2D and 3D gamma analysis (where both datasets were generated in the same manner) for clinical treatment plans.

Methods:

Fifty IMRT plans were selected from the authors’ clinical database, and recalculated using Monte Carlo. Treatment planning system‐calculated (“evaluated dose distributions”) and Monte Carlo‐recalculated (“reference dose distributions”) dose distributions were compared using 2D and 3D gamma analysis. This analysis was performed using a variety of dose‐difference (5%, 3%, 2%, and 1%) and distance‐to‐agreement (5, 3, 2, and 1 mm) acceptance criteria, low‐dose thresholds (5%, 10%, and 15% of the prescription dose), and data grid sizes (1.0, 1.5, and 3.0 mm). Each comparison was evaluated to determine the average 2D and 3D gamma, lower 95th percentile gamma value, and percentage of pixels passing gamma.

Results:

The average gamma, lower 95th percentile gamma value, and percentage of passing pixels for each acceptance criterion demonstrated better agreement for 3D than for 2D analysis for every plan comparison. The average difference in the percentage of passing pixels between the 2D and 3D analyses with no low‐dose threshold ranged from 0.9% to 2.1%. Similarly, using a low‐dose threshold resulted in a difference between the mean 2D and 3D results, ranging from 0.8% to 1.5%. The authors observed no appreciable differences in gamma with changes in the data density (constant difference: 0.8% for 2D vs 3D).

Conclusions:

The authors found that 3D gamma analysis resulted in up to 2.9% more pixels passing than 2D analysis. It must be noted that clinical 2D versus 3D datasets may have additional differences—for example, if 2D measurements are made with a different dosimeter than 3D measurements. Factors such as inherent dosimeter differences may be an important additional consideration to the extra dimension of available data that was evaluated in this study.
Field of view extension and truncation correction for MR‐based human attenuation correction in simultaneous MR/PET imaging
Authors: Blumhagen Jan O. , Braun Harald , Ladebeck Ralf , Fenchel Matthias , Faul David , Scheffler Klaus , Quick Harald H.
Published: January 2014

Purpose:

In quantitative PET imaging, it is critical to accurately measure and compensate for the attenuation of the photons absorbed in the tissue. While in PET/CT the linear attenuation coefficients can be easily determined from a low‐dose CT‐based transmission scan, in whole‐body MR/PET the computation of the linear attenuation coefficients is based on the MR data. However, a constraint of the MR‐based attenuation correction (AC) is the MR‐inherent field‐of‐view (FoV) limitation due to static magnetic field (B0) inhomogeneities and gradient nonlinearities. Therefore, the MR‐based human AC map may be truncated or geometrically distorted toward the edges of the FoV and, consequently, the PET reconstruction with MR‐based AC may be biased. This is especially of impact laterally where the patient arms rest beside the body and are not fully considered.

Methods:

A method is proposed to extend the MR FoV by determining an optimal readout gradient field which locally compensates B0 inhomogeneities and gradient nonlinearities. This technique was used to reduce truncation in AC maps of 12 patients, and the impact on the PET quantification was analyzed and compared to truncated data without applying the FoV extension and additionally to an established approach of PET‐based FoV extension.

Results:

The truncation artifacts in the MR‐based AC maps were successfully reduced in all patients, and the mean body volume was thereby increased by 5.4%. In some cases large patient‐dependent changes in SUV of up to 30% were observed in individual lesions when compared to the standard truncated attenuation map.

Conclusions:

The proposed technique successfully extends the MR FoV in MR‐based attenuation correction and shows an improvement of PET quantification in whole‐body MR/PET hybrid imaging. In comparison to the PET‐based completion of the truncated body contour, the proposed method is also applicable to specialized PET tracers with little uptake in the arms and might reduce the computation time by obviating the need for iterative calculations of the PET emission data beyond those required for reconstructing images.
Hydration and Urinary Pseudoephedrine Levels After a Simulated Team Game
Authors: Jolley Daniel , Dawson Brian , Maloney Shane K. , White James , Goodman Carmel , Peeling Peter
Published: January 2014

This study investigated the influence of dehydration on urinary levels of pseudoephedrine (PSE) after prolonged repeated effort activity. Fourteen athletes performed a simulated team game circuit (STGC) outdoors over 120 min under three different hydration protocols: hydrated (HYD), dehydrated (DHY) and dehydrated + postexercise fluid bolus (BOL). In all trials, a 60 mg dose of PSE was administered 30 min before trial and at half time of the STGC. Urinary PSE levels were measured before drug administration and at 90 min postexercise. In addition, body mass (BM) changes and urinary specific gravity (USG), osmolality (OSM), creatinine (Cr), and pH values were recorded. No differences in PSE levels were found 90 min postexercise between conditions (HYD: 208.5 ± 116.5; DHY: 238.9 ± 93.5; BOL: 195.6 ± 107.3 μg·ml−1), although large variations were seen within and between participants across conditions (range: 33–475 μg·ml−1: ICC r = .03–0.16, p > .05). There were no differences between conditions in USG, OSM, pH or PSE/Cr ratio. In conclusion, hydration status did not influence urinary PSE levels after prolonged repeated effort activity, with ~70% of samples greater than the WADA limit (>150 μg.ml−1), and ~30% under. Due to the unpredictability of urinary PSE values, athletes should avoid taking any medications containing PSE during competition.
Discovery of a Three-Dimensional Topological Dirac Semimetal, Na
3
Bi
Authors: Liu Z. K. , Zhou B. , Zhang Y. , Wang Z. J. , Weng H. M. , Prabhakaran D. , Mo S.-K. , Shen Z. X. , Fang Z. , Dai X. , Hussain Z. , Chen Y. L.
Published: January 2014

A 3D Graphene?

Discoveries of materials with exciting electronic properties have propelled condensed matter physics over the past decade. Two of the best-known examples, graphene and topological insulators, have something in common: a linear energy-momentum relationship—the Dirac dispersion—in their two-dimensional (2D) electronic states. Topological insulators also have a more mundane aspect of their electronic structure, characterized by a band gap. Another class of materials, topological Dirac semimetals, has been proposed that has a linear dispersion along all three momentum directions—a bulk Dirac cone; these materials are predicted to have intriguing electronic properties and to be related to other exotic states through quantum phase transitions.

Liu
et al.

(p.
864
, published online 16 January) detected such a state in the compound Na3Bi by using photoemission spectroscopy.
Single-Crystal Linear Polymers Through Visible Light–Triggered Topochemical Quantitative Polymerization
Authors: Dou Letian , Zheng Yonghao , Shen Xiaoqin , Wu Guang , Fields Kirk , Hsu Wan-Ching , Zhou Huanping , Yang Yang , Wudl Fred
Published: January 2014

Topochemical Polymerization

In a topochemical reaction, chemical changes start at active sites in the solid and then proceed autocatalytically to neighboring regions. If one starts with a monomer that can form ordered structures similar to the final polymer, it is possible to polymerize chains in a fully ordered state and thus make very long single chains.

Dou
et al.

(p.
272
; see the Perspective by

Goroff

) describe an unexpected visible-light–induced polymerization of derivatives of a dye. Two of the derivatives underwent photoinduced single-crystal-to-single-crystal topochemical polymerization.
Low Core-Mantle Boundary Temperature Inferred from the Solidus of Pyrolite
Authors: Nomura Ryuichi , Hirose Kei , Uesugi Kentaro , Ohishi Yasuo , Tsuchiyama Akira , Miyake Akira , Ueno Yuichiro
Published: January 2014

Melting Moments

The boundary between Earth's core and mantle defines where the iron-rich liquid outer core meets the more chemically heterogeneous solid lower mantle and is marked by a sharp thermal gradient of nearly 1500 kelvin. The precise relationship between temperature and melting of the lowermost mantle constrains the structure and heat flow across the core-mantle boundary. In order to identify trace amounts of liquid as melting initiates,

Nomura
et al
.

(p.
522
, published online 16 January) performed x-ray microtomographic imaging of rocks of a primitive mantle composition that had been subjected to high pressures and temperatures in a diamond anvil cell. The experimentally determined maximum melting point of 3570 kelvin suggests that some phases typically thought to lose stability in the lowermost mantle, such as MgSiO
3
-rich post-perovskite, may be more widely distributed than expected.
Direct in Vivo RNAi Screen Unveils Myosin IIa as a Tumor Suppressor of Squamous Cell Carcinomas
Authors: Schramek Daniel , Sendoel Ataman , Segal Jeremy P. , Beronja Slobodan , Heller Evan , Oristian Daniel , Reva Boris , Fuchs Elaine
Published: January 2014

Identifying Drivers and Passengers

Modern genomics is unearthing hundreds of genetic and epigenetic alterations associated with human cancers. It is important to delineate which of these alterations participate actively in tumor progression and/or metastases (driver mutations) and which are inconsequential (passenger mutations). To this end,

Schramek
et al.

(p.
309
) conducted an in vivo RNA interference screen in mice to test simultaneously the functionality of putative cancer genes and down-regulated messenger RNAs associated with tumor-initiating cells of squamous cell carcinomas (SCCs). Several candidates, including nonmuscle myosin-IIa, not previously viewed as tumor suppressors were uncovered.
Ammonia Pollution From Farming May Exact Hefty Health Costs
Authors: Stokstad Erik
Published: January 2014

As U.S. farm exports soar, so does concern about ammonia pollution.
Gut Instinct
Authors: de Vrieze Jop
Published: January 2014

Do bacteria in the guts of African hunter-gatherers hold the key to a healthier life? An American anthropologist plans to find out.
Complete Genome Sequence of Border Disease Virus Genotype 3 Strain Gifhorn
Authors: Fahnøe Ulrik , Höper Dirk , Schirrmeier Horst , Beer Martin , Rasmussen Thomas Bruun
Published: January 2014

The complete genome sequence of the genotype 3 border disease virus strain Gifhorn has been determined; this strain was originally isolated from pigs. This represents the consensus sequence for the virus used to produce the bacterial artificial chromosome (BAC) cDNA clone pBeloGif3, which yields a virus that is severely attenuated in cell culture.
Draft Genome Sequence of the Model Naphthalene-Utilizing Organism Pseudomonas putida OUS82
Authors: Tay Martin , Roizman Dan , Cohen Yehuda , Tolker-Nielsen Tim , Givskov Michael , Yang Liang
Published: January 2014

Pseudomonas putida
OUS82 was isolated from petrol- and oil-contaminated soil in 1992, and ever since, it has been used as a model organism to study the microbial assimilation of naphthalene and phenanthrene. Here, we report the 6.7-Mb draft genome sequence of
P. putida
OUS82 and analyze its featured pathways for biodegradation.
Draft Genome Sequence of the
d
-Xylose-Fermenting Yeast
Spathaspora arborariae
UFMG-HM19.1A
T
Authors: Lobo Francisco P. , Gonçalves Davi L. , Alves Sergio L. , Gerber Alexandra L. , de Vasconcelos Ana Tereza R. , Basso Luiz C. , Franco Glória R. , Soares Marco A. , Cadete Raquel M. , Rosa Carlos A. , Stambuk Boris U.
Published: January 2014

The draft genome sequence of the yeast
Spathaspora arborariae
UFMG-HM19.1A
T
(CBS 11463 = NRRL Y-48658) is presented here. The sequenced genome size is 12.7 Mb, consisting of 41 scaffolds containing a total of 5,625 predicted open reading frames, including many genes encoding enzymes and transporters involved in
d
-xylose fermentation.
Evaluation of Two Line Probe Assays for Rapid Detection of Mycobacterium tuberculosis, Tuberculosis (TB) Drug Resistance, and Non-TB Mycobacteria in HIV-Infected Individuals with Suspected TB
Authors: Luetkemeyer Anne F. , Kendall Michelle A. , Wu Xingye , Lourenço Maria Cristina , Jentsch Ute , Swindells Susan , Qasba Sarojini S. , Sanchez Jorge , Havlir Diane V. , Grinsztejn Beatriz , Sanne Ian M. , Firnhaber Cynthia
Published: January 2014

Limited performance data from line probe assays (LPAs), nucleic acid tests used for the rapid diagnosis of tuberculosis (TB), nontuberculosis mycobacteria (NTM), and
Mycobacterium tuberculosis
drug resistance are available for HIV-infected individuals, in whom paucibacillary TB is common. In this study, the strategy of testing sputum with GenoType MTBDRplus (MTBDR-Plus) and GenoType Direct LPA (Direct LPA) was compared to a gold standard of one mycobacterial growth indicator tube (MGIT) liquid culture. HIV-positive (HIV
+
) individuals with suspected TB from southern Africa and South America with <7 days of TB treatment had 1 sputum specimen tested with Direct LPA, MTBDR-Plus LPA, smear microscopy, MGIT, biochemical identification of mycobacterial species, and culture-based drug-susceptibility testing (DST). Of 639 participants, 59.3% were MGIT
M. tuberculosis
culture positive, of which 276 (72.8%) were acid-fast bacillus (AFB) smear positive. MTBDR-Plus had a sensitivity of 81.0% and a specificity of 100%, with sensitivities of 44.1% in AFB smear-negative versus 94.6% in AFB smear-positive specimens. For specimens that were positive for
M. tuberculosis
by MTBDR-Plus, the sensitivity and specificity for rifampin resistance were 91.7% and 96.6%, respectively, and for isoniazid (INH) they were 70.6% and 99.1%. The Direct LPA had a sensitivity of 88.4% and a specificity of 94.6% for
M. tuberculosis
detection, with a sensitivity of 72.5% in smear-negative specimens. Ten of 639 MGIT cultures grew
Mycobacterium avium
complex or
Mycobacterium kansasii
, half of which were detected by Direct LPA. Both LPA assays performed well in specimens from HIV-infected individuals, including in AFB smear-negative specimens, with 72.5% sensitivity for
M. tuberculosis
identification with the Direct LPA and 44.1% sensitivity with MTBDR-Plus. LPAs have a continued role for use in settings where rapid identification of INH resistance and clinically relevant NTM are priorities.
Comparison of Real-Time Reverse Transcriptase PCR Assays for Detection of Swine Hepatitis E Virus in Fecal Samples
Authors: Gerber Priscilla F. , Xiao Chao-Ting , Cao Dianjun , Meng Xiang-Jin , Opriessnig Tanja
Published: January 2014

Hepatitis E virus (HEV) is a major cause of acute viral hepatitis in people in many developing countries and is also endemic in many industrialized countries. Mammalian HEV (mHEV) isolates can be divided into at least four recognized major genotypes. Several nucleic acid amplification techniques have been developed for mHEV detection, with great differences in sensitivity. The aim of this study was to compare the performances of two singleplex real-time reverse transcriptase (RT) PCR assays for broad detection of all four mHEV genotypes (assays A and B) and two duplex real-time RT-PCR assays for detection and differentiation of mHEV genotypes 3 and 4 (assays C and D). RNAs extracted from 28 fecal samples from pigs experimentally inoculated with HEV genotype 3 and 186 fecal samples from commercial pigs with unknown HEV exposure were tested by all four assays. In experimental samples, HEV RNA was detected in 96.4% (assay A), 39.2% (assay B), 14.2% (assay C), and 0% (assay D) of the samples. In field samples with unknown HEV exposure, HEV RNA was detected in 67.2% (assay A), 36.4% (assay B), 1.1% (assay C), and 0.5% (assay D) of the samples. The assays showed overall poor agreement (κ = 0.19 to 0.03), with differences in detection rates between assays (
P
< 0.01). Assays A and B, which broadly detect HEV genotypes 1 to 4, had significantly higher detection rates for HEV RNA than the duplex assays C and D, which were both designed to detect and differentiate between HEV genotypes 3 and 4.
Performance of Sa
Select
, a Chromogenic Medium for Detection of Staphylococci in Clinical Specimens
Authors: Hirvonen Jari J. , Kerttula Anne-Marie , Kaukoranta Suvi-Sirkku
Published: January 2014

In a preliminary study, known staphylococcus (
n
= 86) and other microbial (
n
= 12) isolates were plated on three chromogenic media, Sa
Select
(Bio-Rad, Hercules, CA, USA), CHROMagar
Staph. aureus
(CHROMagar Microbiology, Paris, France), and
S. aureus
ID (bioMérieux, Marcy l'Etoile, France). The sensitivities of all the media to detect
Staphylococcus aureus
after 24 h of incubation were high (100.0%). However, their specificities varied at 93.3% (95% confidence interval [CI], 86.0% to 100.0%) (CHROMagar
Staph. aureus
), 97.8% (95% CI, 93.5% to 100.0%) (
S. aureus
ID), and 100.0% (Sa
Select
). Sa
Select
also showed the highest sensitivity for recovery and differentiation of other staphylococci. As the best performing chromogenic medium, Sa
Select
was then prospectively compared to conventional culture and identification tests for the detection of staphylococci from 2,780 clinical specimens. A total of 1,589 staphylococcal isolates were recovered. Of these, 912 were
S. aureus
and 677 were other staphylococci. The sensitivity and specificity of Sa
Select
to detect
S. aureus
in clinical specimens after 24 h of incubation were 99.6% and 99.9% (95% CI, 99.2% to 100.0% and 99.8% to 100.0%), respectively, whereas the sensitivity and specificity using conventional plates combined with laboratory identification methods were 96.8% and 99.5% (95% CI, 95.7 to 97.9% and 99.2% to 99.8%). For the recovery and preliminary identification of other staphylococci, the sensitivity and specificity of Sa
Select
were 94.4% and 99.9%. Sa
Select
is a well-performing chromogenic medium that significantly improved the detection of staphylococci, especially
S. aureus
, compared to conventional culture (
P
< 0.0001).
Dynamic Imaging of the Hepatitis C Virus NS5A Protein during a Productive Infection
Authors: Eyre Nicholas S. , Fiches Guillaume N. , Aloia Amanda L. , Helbig Karla J. , McCartney Erin M. , McErlean Christopher S. P. , Li Kui , Aggarwal Anupriya , Turville Stuart G. , Beard Michael R.
Published: January 2014

Hepatitis C virus (HCV) NS5A is essential for viral genome replication within cytoplasmic replication complexes and virus assembly at the lipid droplet (LD) surface, although its definitive functions are poorly understood. We developed approaches to investigate NS5A dynamics during a productive infection. We report here that NS5A motility and efficient HCV RNA replication require the microtubule network and the cytoplasmic motor dynein and demonstrate that both motile and relatively static NS5A-positive foci are enriched with host factors VAP-A and Rab5A. Pulse-chase imaging revealed that newly synthesized NS5A foci are small and distinct from aged foci, while further studies using a unique dual fluorescently tagged infectious HCV chimera showed a relatively stable association of NS5A foci with core-capped LDs. These results reveal new details about the dynamics and maturation of NS5A and the nature of potential sites of convergence of HCV replication and assembly pathways.

IMPORTANCE
Hepatitis C virus (HCV) is a major cause of serious liver disease worldwide. An improved understanding of the HCV replication cycle will enable development of novel and improved antiviral strategies. Here we have developed complementary fluorescent labeling and imaging approaches to investigate the localization, traffic and interactions of the HCV NS5A protein in living, virus-producing cells. These studies reveal new details as to the traffic, composition and biogenesis of NS5A foci and the nature of their association with putative sites of virus assembly.
TRAF2 Facilitates Vaccinia Virus Replication by Promoting Rapid Virus Entry
Authors: Haga Ismar R. , Pechenick Jowers Tali , Griffiths Samantha J. , Haas Juergen , Beard Philippa M.
Published: January 2014

Tumor necrosis factor receptor (TNFR)-associated factor 2 (TRAF2) is a pivotal intracellular mediator of signaling pathways downstream of TNFR1 and -2 with known pro- and antiviral effects. We investigated its role in the replication of the prototype poxvirus vaccinia virus (VACV). Loss of TRAF2 expression, either through small interfering RNA treatment of HeLa cells or through genetic knockout in murine embryonic fibroblasts (MEFs), led to significant reductions in VACV growth following low-multiplicity infection. In single-cycle infections, there was delayed production of both early and late VACV proteins as well as accelerated virus-induced alterations to cell morphology, indicating that TRAF2 influences early stages of virus replication. Consistent with an early role, uncoating assays showed normal virus attachment but delayed virus entry in the absence of TRAF2. Although alterations to c-Jun N-terminal kinase (JNK) signaling were apparent in VACV-infected TRAF2
−/−
MEFs, treatment of wild-type cells with a JNK inhibitor did not affect virus entry. Instead, treatment with an inhibitor of endosomal acidification greatly reduced virus entry into TRAF2
−/−
MEFs, suggesting that VACV is reliant on the endosomal route of entry in the absence of TRAF2. Thus, TRAF2 is a proviral factor for VACV that plays a role in promoting efficient viral entry, most likely via the plasma membrane.

IMPORTANCE
Tumor necrosis factor receptor-associated factors (TRAFs) are key facilitators of intracellular signaling with roles in innate and adaptive immunity and stress responses. We have discovered that TRAF2 is a proviral factor in vaccinia virus replication in both HeLa cells and mouse embryonic fibroblasts and that its influence is exercised through promotion of efficient virus entry.
Hepatitis C Virus-Induced Changes in MicroRNA 107 (miRNA-107) and miRNA-449a Modulate CCL2 by Targeting the Interleukin-6 Receptor Complex in Hepatitis
Authors: Sarma Nayan J. , Tiriveedhi Venkataswarup , Crippin Jeffrey S. , Chapman William C. , Mohanakumar Thalachallour
Published: January 2014

Hepatitis C virus (HCV)-mediated liver diseases are one of the major health issues in the United States and worldwide. HCV infection has been reported to modulate microRNAs (miRNAs) that control various cell surface receptors and gene-regulatory complexes involved in hepatic inflammation and liver diseases. We report here that specific downregulation of miRNA-107 and miRNA-449a following HCV infection in patients with HCV-mediated liver diseases modulates expression of CCL2, an inflammatory chemokine upregulated in patients with chronic liver diseases, by targeting components of the interleukin-6 receptor (IL-6R) complex. Computational analysis for DNA-bound transcription factors in theCCL2promoter identified adjacent binding sites for CCAAT/CEBPα, spleen focus-forming virus, proviral integration oncogene (SPI1/PU.1), and STAT3. We demonstrate that CEBPα, PU.1, and STAT3 interacted with each other physically to cooperatively bind to the promoter and activateCCL2expression. Analysis ofIL-6RandJAK1expression in HCV patients by quantitative PCR showed significant upregulation when there was impaired miRNA-107 and miRNA-449a expression, along with upregulation of PU.1 and STAT3, but not CEBPα. miRNA-449a and miRNA-107 target expression ofIL-6RandJAK1, respectively,in vitroand also inhibit IL-6 signaling and impair STAT3 activation in human hepatocytes. Taken together, our results demonstrate a novel gene-regulatory mechanism in which HCV-induced changes in miRNAs (miRNA-449a and miRNA-107) regulate CCL2 expression by activation of the IL-6-mediated signaling cascade, which we propose will result in HCV-mediated induction of inflammatory responses and fibrosis.

IMPORTANCEHepatitis C virus (HCV)-induced hepatitis is a major health concern worldwide. HCV infection results in modulation of noncoding microRNAs affecting major cellular pathways, including inflammatory responses. In this study, we have identified a microRNA-regulated pathway for the chemokine CCL2 in HCV-induced hepatitis. Understanding microRNA-mediated transcriptional-regulatory pathways will result in development of noninvasive biomarkers for better disease prediction and development of effective therapeutics.
Epstein-Barr Virus Latent Membrane Protein 1 Genetic Variability in Peripheral Blood B Cells and Oropharyngeal Fluids
Authors: Renzette Nicholas , Somasundaran Mohan , Brewster Frank , Coderre James , Weiss Eric R. , McManus Margaret , Greenough Thomas , Tabak Barbara , Garber Manuel , Kowalik Timothy F. , Luzuriaga Katherine
Published: January 2014

We report the diversity of latent membrane protein 1 (LMP1) gene founder sequences and the level of Epstein-Barr virus (EBV) genome variability over time and across anatomic compartments by using virus genomes amplified directly from oropharyngeal wash specimens and peripheral blood B cells during acute infection and convalescence. The intrahost nucleotide variability of the founder virus was 0.02% across the region sequences, and diversity increased significantly over time in the oropharyngeal compartment (P= 0.004). The LMP1 region showing the greatest level of variability in both compartments, and over time, was concentrated within the functional carboxyl-terminal activating regions 2 and 3 (CTAR2 and CTAR3). Interestingly, a deletion in a proline-rich repeat region (amino acids 274 to 289) of EBV commonly reported in EBV sequenced from cancer specimens was not observed in acute infectious mononucleosis (AIM) patients. Taken together, these data highlight the diversity in circulating EBV genomes and its potential importance in disease pathogenesis and vaccine design.

IMPORTANCEThis study is among the first to leverage an improved high-throughput deep-sequencing methodology to investigate directly from patient samples the degree of diversity in Epstein-Barr virus (EBV) populations and the extent to which viral genome diversity develops over time in the infected host. Significant variability of circulating EBV latent membrane protein 1 (LMP1) gene sequences was observed between cellular and oral wash samples, and this variability increased over time in oral wash samples. The significance of EBV genetic diversity in transmission and disease pathogenesis are discussed.
Antigenic Mapping of the Hemagglutinin of an H9N2 Avian Influenza Virus Reveals Novel Critical Amino Acid Positions in Antigenic Sites
Authors: Wan Zhimin , Ye Jianqiang , Xu Liangliang , Shao Hongxia , Jin Wenjie , Qian Kun , Wan Hongquan , Qin Aijian
Published: January 2014

H9N2 influenza virus is undergoing extensive genetic and antigenic evolution, warranting detailed antigenic mapping of its hemagglutinin (HA). Through examining antibody escape mutants of an Asian avian H9N2 virus, we identified 9 critical amino acid positions in H9 antigenic sites. Five of these positions, 164, 167, 168, 196, and 207, have not been reported previously and, thus, represent novel molecular markers for monitoring the antigenic change of H9N2 virus.
Molecular Epidemiology of Contemporary G2P[4] Human Rotaviruses Cocirculating in a Single U.S. Community: Footprints of a Globally Transitioning Genotype
Authors: Dennis Allison F. , McDonald Sarah M. , Payne Daniel C. , Mijatovic-Rustempasic Slavica , Esona Mathew D. , Edwards Kathryn M. , Chappell James D. , Patton John T.
Published: January 2014

Group A rotaviruses (RVs) remain a leading cause of childhood gastroenteritis worldwide. Although the G/P types of locally circulating RVs can vary from year to year and differ depending upon geographical location, those with G1P[8], G2P[4], G3P[8], G4P[8], G9P[8], and G12P[8] specificities typically dominate. Little is known about the evolution and diversity of G2P[4] RVs and the possible role that widespread vaccine use has had on their increased frequency of detection. To address these issues, we analyzed the 12 G2P[4] RV isolates associated with a rise in RV gastroenteritis cases at Vanderbilt University Medical Center (VUMC) during the 2010-2011 winter season. Full-genome sequencing revealed that the isolates had genotype 2 constellations typical of DS-1-like viruses (G2P[4]-I2-R2-C2-M2-A2-N2-T2-E2-H2). Phylogenetic analyses showed that the genome segments of the isolates were comprised of two or three different subgenotype alleles; this enabled recognition of three distinct clades of G2P[4] viruses that caused disease at VUMC in the 2010-2011 season. Although the three clades cocirculated in the same community, there was no evidence of interclade reassortment. Bayesian analysis of 328 VP7 genes of G2 viruses isolated in the last 39 years indicate that existing G2 VP7 gene lineages continue to evolve and that novel lineages, as represented by the VUMC isolates, are constantly being formed. Moreover, G2 lineages are characteristically shaped by lineage turnover events that introduce new globally dominant strains every 7 years, on average. The ongoing evolution of G2 VP7 lineages may give rise to antigenic changes that undermine vaccine effectiveness in the long term.

IMPORTANCE
Little is known about the diversity of cocirculating G2 rotaviruses and how their evolution may undermine the effectiveness of rotavirus vaccines. To expand our understanding of the potential genetic range exhibited by rotaviruses circulating in postvaccine communities, we analyzed part of a collection of rotaviruses recovered from pediatric patients in the United States from 2010 to 2011. Examining the genetic makeup of these viruses revealed they represented three segregated groups that did not exchange genetic material. The distinction between these three groups may be explained by three separate introductions. By comparing a specific gene, namely, VP7, of the recent rotavirus isolates to those from a collection recovered from U.S. children between 1974 and 1991 and other globally circulating rotaviruses, we were able to reconstruct the timing of events that shaped their ancestry. This analysis indicates that G2 rotaviruses are continuously evolving, accumulating changes in their genetic material as they infect new patients.
Evolution of a Cell Culture-Derived Genotype 1a Hepatitis C Virus (H77S.2) during Persistent Infection with Chronic Hepatitis in a Chimpanzee
Authors: Yi MinKyung , Hu Fengyu , Joyce Michael , Saxena Vikas , Welsch Christoph , Chavez Deborah , Guerra Bernadette , Yamane Daisuke , Veselenak Ronald , Pyles Rick , Walker Christopher M. , Tyrrell Lorne , Bourne Nigel , Lanford Robert E. , Lemon Stanley M.
Published: January 2014

Persistent infection is a key feature of hepatitis C virus (HCV). However, chimpanzee infections with cell culture-derived viruses (JFH1 or related chimeric viruses that replicate efficiently in cell culture) have been limited to acute-transient infections with no pathogenicity. Here, we report persistent infection with chronic hepatitis in a chimpanzee challenged with cell culture-derived genotype 1a virus (H77S.2) containing 6 cell culture-adaptive mutations. Following acute-transient infection with a chimeric H77/JFH1 virus (HJ3-5), intravenous (i.v.) challenge with 10
6
FFU H77S.2 virus resulted in immediate seroconversion and, following an unusual 4- to 6-week delay, persistent viremia accompanied by alanine aminotransferase (ALT) elevation, intrahepatic innate immune responses, and diffuse hepatopathy. This first persistent infection with cell culture-produced HCV provided a unique opportunity to assess evolution of cell culture-adapted virus
in vivo
. Synonymous and nonsynonymous nucleotide substitution rates were greatest during the first 8 weeks of infection. Of 6 cell culture-adaptive mutations in H77S.2, Q1067R (NS3) had reverted to Q1067 and S2204I (NS5A) was replaced by T2204 within 8 weeks of infection. By 62 weeks, 4 of 6 mutations had reverted to the wild-type sequence, and all reverted to the wild-type sequence by 194 weeks. The data suggest H77S.2 virus has greater potential for persistence and pathogenicity than JFH1 and demonstrate both the capacity of a nonfit virus to persist for weeks in the liver in the absence of detectable viremia as well as strong selective pressure against cell culture-adaptive mutations
in vivo
.

IMPORTANCE
This study shows that mutations promoting the production of infectious genotype 1a HCV in cell culture have the opposite effect and attenuate replication in the liver of the only fully permissive animal species other than humans. It provides the only example to date of persistent infection in a chimpanzee challenged with cell culture-produced virus and provides novel insight into the forces shaping molecular evolution of that virus during 5 years of persistent infection. It demonstrates that a poorly fit virus can replicate for weeks within the liver in the absence of detectable viremia, an observation that expands current concepts of HCV pathogenesis and that is relevant to relapses observed with direct-acting antiviral therapies.