bluuebunny/crossref_metadata_2025 huggingface dataset:
Electromotile hearing: Acoustic tones mask psychophysical response to high-frequency electrical stimulation of intact guinea pig cochle
Authors: Le Prell Colleen G. , Kawamoto Kohei , Raphael Yehoash , Dolan David F.
Published: December 2006

When sinusoidal electric stimulation is applied to the intact cochlea, a frequency-specific acoustic emission can be recorded in the ear canal. Acoustic emissions are produced by basilar membrane motion, and have been used to suggest a corresponding acoustic sensation termed “electromotile hearing.” Electromotile hearing has been specifically attributed to electric stimulation of outer hair cells in the intact organ of Corti. To determine the nature of the auditory perception produced by electric stimulation of a cochlea with intact outer hair cells, guinea pigs were tested in a psychophysical task. First, subjects were trained to report detection of sinusoidal acoustic stimuli and dynamic range was assessed using response latency. Subjects were then implanted with a ball electrode placed into scala tympani. Following the surgical implant procedure, subjects were transferred to a task in which acoustic signals were replaced by sinusoidal electric stimulation, and dynamic range was assessed again. Finally, the ability of acoustic pure-tone stimuli to mask the detection of the electric signals was assessed. Based on the masking effects, it is concluded that sinusoidal electric stimulation of the intact cochlea results in perception of a tonal (rather than a broadband or noisy) sound at a frequency of 8kHz or above.
Study of the comparison of the methods of equivalent sources and boundary element methods for near-field acoustic holography
Authors: Valdivia Nicolas P. , Williams Earl G.
Published: December 2006

Boundary element methods (BEM) based near-field acoustic holography (NAH) has been used successfully in order to reconstruct the normal velocity on an arbitrarily shaped structure surface from measurements of the pressure field on a nearby conformal surface. An alternative approach for this reconstruction on a general structure utilizes the equivalent sources method (ESM). In ESM the acoustic field is represented by a set of point sources located over a surface that is close to the structure surface. This approach is attractive mainly for its simplicity of implementation and speed. In this work ESM as an approximation of BEM based NAH is studied and the necessary conditions for the successful application of this approach in NAH is discussed. A cylindrical fuselage surface excited by a point force as an example to validate the results is used.
Long-time temperature rise due to absorption of focused Gaussian beams in tissue
Authors: Myers Matthew R.
Published: December 2006

An analytical technique previously developed to study tissue displacement due to acoustic radiation force is extended to analyze temperature rise in tissue for exposure times that are comparable to, or longer than, the tissue perfusion time. A focused transducer with Gaussian amplitude shading is assumed to radiate into a perfused tissue medium with constant thermal and acoustic properties. A simple closed-form expression is derived for the steady-state temperature rise, and a transient correction term is constructed that allows for computation of the equilibrium time of the medium. Comparisons with temperature calculations for non-Gaussian transducers show that the model may be applied to more general intensity profiles.
Impact of perforation impedance on the transmission loss of reactive and dissipative silencers
Authors: Lee Iljae , Selamet Ahmet , Huff Norman T.
Published: December 2006

The effect of perforation impedance on the acoustic behavior of reactive and dissipative silencers is investigated using experimental and computational approaches. The boundary element method (BEM) is applied for the prediction of transmission loss of silencers with different perforation geometries. The variations are considered in the porosity (8.4 and 25.7%) and hole diameter (0.249 and 0.498cm) of perforations for both reactive and dissipative silencers, as well as the fiber filling density (100 and 200kg∕m3) for the latter. The acoustic impedance for a number of perforations in contact with air alone and fibrous material has been incorporated into the predictions, which are then compared with the measured transmission loss using an impedance tube setup. The results demonstrate the significance of the accuracy of the perforation impedance in the predictions for both reactive and dissipative silencers.
Whistle variability in South Atlantic spinner dolphins from the Fernando de Noronha Archipelago off Brazil
Authors: Camargo Fernanda S. , Rollo Mario M. , Giampaoli Viviana , Bellini Claudio
Published: December 2006

A series of quali- and quantitative analyses were conducted to evaluate the variability of spinner dolphin whistles from the Fernando de Noronha Archipelago off Brazil. Nine variables were extracted from each whistle contour, and the whistle contours shapes were classified into the seven categories described in Driscoll (1995). The analysis showed mean beginning and ending frequencies values of 10.78 and 12.74kHz, respectively. On average, whistle duration was relatively short, with mean values around 0.495s (N=702). Comparative analyses were also conducted to investigate the relationship between the obtained results and those presented in previous studies. When comparing averages, the results of the study of Oswald et al.(2003) in the Tropical Eastern Pacific (TEP) presented less significant differences in relation to this study; only whistle duration differed significantly between both works. The results of multivariate classification tests also pointed TEP population as the closest related to the population studied here. The similarities between such disjunct populations might be attributed to a more recent isolation event (the closing of the Panama Isthmus) than the divergence that has driven North and South Atlantic populations apart.
Anomalous postcritical refraction behavior for certain transversely isotropic media
Authors: Fa Lin , Brown Ray L. , Castagna John P.
Published: December 2006

Snell’s law at the boundary between two transversely isotropic media with a vertical axis of symmetry (VTI media) can be solved by setting up a fourth order polynomial for the sine of the reflection/transmission angles. This approach reveals the possible presence of an anomalous postcritical angle for certain transversely isotropic media. There are thus possibly three incident angle regimes for the reflection/refraction of longitudinal or transverse waves incident upon a VTI medium: precritical, postcritical/preanomalous, and postanomalous. The anomalous angle occurs for certain strongly anisotropic media where the required root to the phase velocity equation must be switched in order to obey Snell’s law. The reflection/transmission coefficients, polarization directions, and the phase velocity are all affected by both the anisotropy and the incident angle. The incident critical angles are also effected by the anisotropy.
Modal decomposition method for acoustic impedance testing in square ducts
Authors: Schultz Todd , Cattafesta Louis N. , Sheplak Mark
Published: December 2006

Accurate duct acoustic propagation models are required to predict and reduce aircraft engine noise. These models ultimately rely on measurements of the acoustic impedance to characterize candidate engine nacelle liners. This research effort increases the frequency range of normal-incidence acoustic impedance testing in square ducts by extending the standard two-microphone method (TMM), which is limited to plane wave propagation, to include higher-order modes. The modal decomposition method (MDM) presented includes four normal modes in the model of the sound field, thus increasing the bandwidth from 6.7to13.5kHz for a 25.4mm square waveguide. The MDM characterizes the test specimen for normal- and oblique-incident acoustic impedance and mode scattering coefficients. The MDM is first formulated and then applied to the measurement of the reflection coefficient matrix for a ceramic tubular specimen. The experimental results are consistent with results from the TMM for the same specimen to within the 95% confidence intervals for the TMM. The MDM results show a series of resonances for the ceramic tubular material exhibiting a monotonic decrease in the resonant peaks of the acoustic resistance with increasing frequency, resembling a rigidly-terminated viscous tube, and also evidence of mode scattering is visible at the higher frequencies.
Molecular analysis of the effect of relative tone level on multitone pattern discrimination
Authors: Lutfi Robert A. , Jesteadt Walt
Published: December 2006

Molecular psychophysics attempts to model the observer’s response to stimuli as they vary from trial to trial. The approach has gained popularity in multitone pattern discrimination studies as a means of estimating the relative reliance or decision weight listeners give to different tones in the pattern. Various factors affecting decision weights have been examined, but one largely ignored is the relative level of tones in the pattern. In the present study listeners detected a level-increment in a sequence of 5, 100-ms, 2.0-kHz tone bursts alternating in level between 40 and 80dB SPL. The level increment was made largest on the 40-dB tones, yet despite this all four highly-practiced listeners gave near exclusive weight to the 80-dB tones. The effect was the same when the tones were replaced by bursts of broadband Gaussian noise alternating in level. It was reduced only when the level differences were made <10dB, and it was entirely reversed only when the low-level tones alternated with louder bursts of Gaussian noise. The results are discussed in terms of the effects of both sensory and perceptual factors on estimates of decision weights.
Axial radiation force of a Bessel beam on a sphere and direction reversal of the force
Authors: Marston Philip L.
Published: December 2006

An expression is derived for the radiation force on a sphere placed on the axis of an ideal acoustic Bessel beam propagating in an inviscid fluid. The expression uses the partial-wave coefficients found in the analysis of the scattering when the sphere is placed in a plane wave traveling in the same external fluid. The Bessel beam is characterized by the cone angle β of its plane wave components where β=0 gives the limiting case of an ordinary plane wave. Examples are found for fluid spheres where the radiation force reverses in direction so the force is opposite the direction of the beam propagation. Negative axial forces are found to be correlated with conditions giving reduced backscattering by the beam. This condition may also be helpful in the design of acoustic tweezers for biophysical applications. Other potential applications include the manipulation of objects in microgravity. Islands in the (ka,β) parameter plane having a negative radiation force are calculated for the case of a hexane drop in water. Here k is the wave number and a is the drop radius. Low frequency approximations to the radiation force are noted for rigid, fluid, and elastic solid spheres in an inviscid fluid.
L1,L2 maps of distortion-product otoacoustic emissions from a moth ear with only two auditory receptor neurons
Authors: Kössl Manfred , Coro Frank
Published: December 2006

The tympanal organ of the moth Empyreuma affinis emits physiologically vulnerable distortion-product otoacoustic emissions. To assess the nature of underlying mechanical nonlinearities, we measured L1,L2 maps by varying both stimulus levels. Two types of maps were found: (1) Maps containing dominant islands centered at the L1=L2 diagonal as it is typical for saturating nonlinearities that can be described by Boltzmann functions. In contrast to maps published for mammals and frogs, the shape of such islands includes sharp ridges at L1 or L2 levels close to 70dB sound pressure level. This could be produced by a strongly asymmetric operating point of the respective transfer functions, consistent with the fact that the auditory sensory cells are not hair cells but primary mechanoreceptors with a single cilium. The saturating map components could be selectively reduced by acoustic suppression. (2) Maps where separated islands were less conspicuous but in which the dominant feature consisted of contour lines which were orthogonal to the L1=2L2 diagonal and could be generated by an expansive nonlinearity. Maps showing strong islands were found for f2 frequencies between 26.7 and 45kHz, maps without strong islands for f2 between 42 and 57.5kHz. This suggests a frequency-dependent change regarding the involved mechanical nonlinearities.
Acoustic response from adherent targeted contrast agents
Authors: Zhao Shukui , Kruse Dustin E. , Ferrara Katherine W. , Dayton Paul A.
Published: December 2006

In ultrasonic molecular imaging, encapsulated micron-sized gas bubbles are tethered to a blood vessel wall by targeting ligands. A challenging problem is to detect the echoes from adherent microbubbles and distinguish them from echoes from nonadherent agents and tissue. Echoes from adherent contrast agents are observed to include a high amplitude at the fundamental frequency, and significantly different spectral shape compared with free agents (p<0.0003). Mechanisms for the observed acoustical difference and potential techniques to utilize these differences for molecular imaging are proposed.
The acoustic signature of bubbles fragmenting in sheared flow
Authors: Deane Grant B. , Dale Stokes M.
Published: December 2006

Measurements of the sound of bubbles fragmenting in fluid shear are presented and analyzed. The frequency, amplitude, and decay rate of the acoustic emissions from 1.8-mm-radius bubbles fragmenting between opposed fluid jets have been determined. A broad band of frequencies (1.8 to 30kHz) is observed with peak pressure amplitudes in the range of 0.03 to 2Pa. While the peak pressure amplitudes show no significant scaling with frequency, the frequency dependence of the decay rates is consistent with the sum of thermal and acoustic radiation losses.
Synthesis of audio spectra using a diffraction model
Authors: Vijayakumar V. , Eswaran C.
Published: December 2006

It is shown that the intensity variations of an audio signal in the frequency domain can be obtained by using a mathematical function containing a series of weighted complex Bessel functions. With proper choice of values for two parameters, this function can transform an input spectrum of discrete frequencies of unit intensity into the known spectra of different musical instruments. Specific examples of musical instruments are considered for evaluating the performance of this method. It is found that this function yields musical spectra with a good degree of accuracy.
The relationship between frequency selectivity and pitch discrimination: Sensorineural hearing loss
Authors: Bernstein Joshua G. W. , Oxenham Andrew J.
Published: December 2006

This study tested the relationship between frequency selectivity and the minimum spacing between harmonics necessary for accurate f0 discrimination. Fundamental frequency difference limens (f0 DLs) were measured for ten listeners with moderate sensorineural hearing loss (SNHL) and three normal-hearing listeners for sine- and random-phase harmonic complexes, bandpass filtered between 1500 and 3500Hz, with f0’s ranging from 75 to 500Hz (or higher). All listeners showed a transition between small (good) f0 DLs at high f0’s and large (poor) f0 DLs at low f0’s, although the f0 at which this transition occurred (f0,tr) varied across listeners. Three measures thought to reflect frequency selectivity were significantly correlated to both the f0,tr and the minimum f0 DL achieved at high f0’s: (1) the maximum f0 for which f0 DLs were phase dependent, (2) the maximum modulation frequency for which amplitude modulation and quasi-frequency modulation were discriminable, and (3) the equivalent rectangular bandwidth of the auditory filter, estimated using the notched-noise method. These results provide evidence of a relationship between f0 discrimination performance and frequency selectivity in listeners with SNHL, supporting “spectral” and “spectro-temporal” theories of pitch perception that rely on sharp tuning in the auditory periphery to accurately extract f0 information.
Vowel recognition via cochlear implants and noise vocoders: Effects of formant movement and duration
Authors: Iverson Paul , Smith Charlotte A. , Evans Bronwen G.
Published: December 2006

Previous work has demonstrated that normal-hearing individuals use fine-grained phonetic variation, such as formant movement and duration, when recognizing English vowels. The present study investigated whether these cues are used by adult postlingually deafened cochlear implant users, and normal-hearing individuals listening to noise-vocoder simulations of cochlear implant processing. In Experiment 1, subjects gave forced-choice identification judgments for recordings of vowels that were signal processed to remove formant movement and/or equate vowel duration. In Experiment 2, a goodness-optimization procedure was used to create perceptual vowel space maps (i.e., best exemplars within a vowel quadrilateral) that included F1, F2, formant movement, and duration. The results demonstrated that both cochlear implant users and normal-hearing individuals use formant movement and duration cues when recognizing English vowels. Moreover, both listener groups used these cues to the same extent, suggesting that postlingually deafened cochlear implant users have category representations for vowels that are similar to those of normal-hearing individuals.
Laser interferometry measurements of middle ear fluid and pressure effects on sound transmission
Authors: Gan Rong Z. , Dai Chenkai , Wood Mark W.
Published: December 2006

An otitis media with effusion model in human temporal bones with two laser vibrometers was created in this study. By measuring the displacement of the stapes from the medial side of the footplate, the transfer function of the middle ear, which is defined as the displacement transmission ratio (DTR) of the tympanic membrane to footplate, was derived under different middle ear pressure and fluid in the cavity with a correction factor for cochlear load. The results suggest that the DTR increases with increasing frequency up to 4kHz when the middle ear pressure was changing from 0 to 20 or −20cm H2O (e.g., ±196daPa) and fluid level was increasing from 0 to a full middle ear cavity. The positive and negative pressures show different effects on the DTR. The effect of fluid on DTR varies between three frequency ranges: f<1k, between 1k and 4k, and f>4kHz. These findings show how the efficiency of the middle ear system for sound transmission changes during the presence of fluid in the cavity and variations of middle ear pressure.
The kurtosis metric as an adjunct to energy in the prediction of trauma from continuous, nonGaussian noise exposures
Authors: Qiu Wei , Hamernik Roger P. , Davis Bob
Published: December 2006

Data from an earlier study [Hamernik et al. (2003). J. Acoust. Soc. Am. 114, 386–395] were consistent in showing that, for equivalent energy [Leq=100dB(A)] and spectra, exposure to a continuous, nonGaussian (nonG) noise could produce substantially greater hearing and sensory cell loss in the chinchilla model than a Gaussian (G) noise exposure and that the statistical metric, kurtosis, computed on the amplitude distribution of the noise could order the extent of the trauma. This paper extends these results to Leq=90 and 110dB(A), and to nonG noises that are generated using broadband noise bursts, and band limited impacts within a continuous G background noise. Data from nine new experimental groups with 11 or 12 chinchillas/group is presented. Evoked response audiometry established hearing thresholds and surface preparation histology quantified sensory cell loss. At the lowest level [Leq=90dB(A)] there were no differences in the trauma produced by G and nonG exposures. For Leq>90dB(A) nonG exposures produced increased trauma relative to equivalent G exposures. Removing energy from the impacts by limiting their bandwidth reduced trauma. The use of noise bursts to produce the nonG noise instead of impacts also reduced the amount of trauma.
Objectively measured and subjectively perceived distortion in nonlinear systems
Authors: Olofsson Åke , Hansen Martin
Published: December 2006

A method for measuring nonlinear distortion, which is applicable to almost any transmission system and which can use any input signal as a test signal, is proposed. The method exploits the fact that a pair of signals, generated to form the real and imaginary parts of the analytical signal corresponding to the input signal to a system, lose their property of being a Hilbert pair after being passed through a nonlinear system. The method was tested by measuring 12 different hearing aid compression systems. These objective measurements were compared with the subjectively perceived amount of distortion, assessed by a group of 10 otologically normal subjects. A reasonably monotonic relation between the subjective and objective measures of distortion was observed. The objective Hilbert pair based measure can be related to both traditional total harmonic distortion and traditional signal to noise ratio.
High-frequency hearing in phocid and otariid pinnipeds: An interpretation based on inertial and cochlear constraints
Authors: Hemilä Simo , Nummela Sirpa , Berta Annalisa , Reuter Tom
Published: December 2006

Audiograms in air and underwater, determined by previous workers for four pinniped species, two eared seals (Otariidae) and two phocids (Phocidae), are supplemented here by measurements on their middle ear ossicular mass, enabling mechanistic interpretations of high-frequency hearing and audiogram differences. Otariid hearing is not largely affected by the medium (air/water). This indicates that cochlear constraints limit high-frequency hearing in otariids. Phocids, however, have massive middle ear ossicles, and underwater hearing has radically shifted towards higher frequencies. This suggests that the high-frequency hearing of phocids in air is constrained by ossicle inertia.
Detection and Differentiation of
Bordetella
spp. by Real-Time PCR
Authors: Koidl Christoph , Bozic Michael , Burmeister Anja , Hess Markus , Marth Egon , Kessler Harald H.
Published: December 2006

Molecular detection of
Bordetella pertussis
DNA is a sensitive and specific method for the rapid diagnosis of pertussis. In this study, a new molecular assay for the detection and differentiation of
Bordetella
spp. based on automated DNA extraction and real-time PCR was evaluated. The analytical sensitivity of the new assay was determined by Probit analysis of serial dilutions of both cloned PCR products IS
481
and IS
1001
and cell suspensions of
B. pertussis
,
B. parapertussis
, and
B. bronchiseptica
. The specificity was analyzed by testing a number of pathogens producing respiratory infections. Moreover, a total of 92 clinical samples were investigated. The results were compared to those obtained by an in-house assay based on manual DNA extraction, followed by real-time PCR and detection of IS
481
. The analytical sensitivity of the new assay for the detection of IS
481
and IS
1001
was determined to be 2.2 and 1.2 genome equivalents/μl, respectively. The analytical sensitivity for the detection of
B. pertussis
,
B. parapertussis
, and
B. bronchiseptica
was determined to be 1.6, 1.0, and 2.7 genome equivalents/μl, respectively. When clinical specimens were tested with the new assay, 46 of 92 were found to be positive for
Bordetella
DNA. With the in-house assay, 45 samples tested positive. The new molecular assay proved to be suitable for the rapid diagnosis of pertussis in the routine diagnostic laboratory.
Isolation of
Trichophyton violaceum
and
Trichophyton soudanense
in Baltimore, Maryland
Authors: Magill Shelley S. , Manfredi Liliana , Swiderski Andrew , Cohen Bernard , Merz William G.
Published: December 2006

Tinea capitis is of public health importance because of its transmissibility.
Trichophyton violaceum
and
Trichophyton soudanense
, which are common causes of tinea capitis in parts of Africa and West Asia, have only rarely been reported to cause dermatophytoses in the United States. We identified 24 patients with 25 positive cultures for
T. violaceum
or
T. soudanense
that were processed in a single hospital laboratory in Baltimore, Maryland, between 1 January 2000 and 30 June 2006. Most patients for whom clinical information was available had tinea capitis. There was a marked increase in the isolation of these organisms between the period from 2000 to 2002 and the period from 2003 to 2006, possibly associated with changes in immigration to the Baltimore metropolitan area. The changing epidemiology of this transmissible fungal infection not only is of public health interest as an example of the introduction of a “new” pathogen to an area where it traditionally was not endemic but also is of clinical and microbiological importance given reports suggesting an increasing incidence of tinea capitis in some areas and increasing clinical failure rates of current therapies.
Detection of
Treponema pallidum
subsp.
pallidum
from Skin Lesions, Serum, and Cerebrospinal Fluid in an Infant with Congenital Syphilis after Clindamycin Treatment of the Mother during Pregnancy
Authors: Woznicová Vladana , Šmajs David , Wechsler Dan , Matějková Petra , Flasarová Magdalena
Published: December 2006

We report here a case of congenital syphilis in a newborn after clindamycin treatment in pregnancy. Using PCR detection of
tmpC
(TP0319) and DNA sequencing of the genes TP0136 and TP0548, DNA sequences identical to
Treponema pallidum
subsp.
pallidum
strain SS14 were detected in the infant's skin lesions, serum, and cerebrospinal fluid.
Absence or Overexpression of the Varicella-Zoster Virus (VZV) ORF29 Latency-Associated Protein Impairs Late Gene Expression and Reduces VZV Latency in a Rodent Model
Authors: Cohen Jeffrey I. , Krogmann Tammy , Pesnicak Lesley , Ali Mir A.
Published: December 2006

Varicella-zoster virus (VZV) ORF29 encodes the viral single-stranded DNA binding protein and is expressed during latency in human ganglia. We constructed an ORF29 deletion mutant virus and showed that the virus could replicate only in cells expressing ORF29. An ORF29-repaired virus, in which ORF29 was driven by a cytomegalovirus promoter, grew to peak titers similar to those seen with the parental virus. The level of ORF29 protein in cells infected with the repaired virus was greater than that seen with parental virus. Infection of cells with either the ORF29 deletion or repaired virus resulted in similar levels of VZV immediate-early proteins but reduced levels of glycoprotein E compared to those observed with parental virus. Cotton rats infected with the ORF29 deletion mutant had a markedly reduced frequency of latent infection in dorsal root ganglia compared with those infected with parental virus (
P
< 0.00001). In contrast, infection of animals with the ORF29 deletion mutant resulted in a frequency of ganglionic infection at 3 days similar to that seen with the parental virus. Animals infected with the ORF29-repaired virus, which overexpresses ORF29, also had a reduced frequency of latent infection compared with those infected with parental virus (
P
= 0.0044). These studies indicate that regulation of ORF29 at appropriate levels is critical for VZV latency in a rodent model.
Novel Pathway for Induction of Latent Virus from Resting CD4+T Cells in the Simian Immunodeficiency Virus/Macaque Model of Human Immunodeficiency Virus Type 1 Latency
Authors: Shen Anding , Yang Hung-Chih , Zhou Yan , Chase Amanda J. , Boyer Jean D. , Zhang Hao , Margolick Joseph B. , Zink M. Christine , Clements Janice E. , Siliciano Robert F.
Published: December 2006

Although combination therapy allows the suppression of human immunodeficiency virus type 1 (HIV-1) viremia to undetectable levels, eradication has not been achieved because the virus persists in cellular reservoirs, particularly the latent reservoir in resting CD4+T lymphocytes. We previously established a simian immunodeficiency virus (SIV)/macaque model to study latency. We describe here a novel mechanism for the induction of SIV from latently infected resting CD4+T cells. Several human cell lines including CEMx174 and Epstein-Barr virus-transformed human B-lymphoblastoid cell lines mediated contact-dependent activation of resting macaque T cells and induction of latent SIV. Antibody-blocking assays showed that interactions between the costimulatory molecule CD2 and its ligand CD58 were involved, whereas soluble factors and interactions between T-cell receptors and major histocompatibility complex class II were not. Combinations of specific antibodies to CD2 also induced T-cell activation and virus induction in human resting CD4+T cells carrying latent HIV-1. This is the first demonstration that costimulatory signals can induce latent virus without the coengagement of the T-cell receptor, and this study might provide insights into potential pathways to target latent HIV-1.
Nonstructural Proteins of Respiratory Syncytial Virus Suppress Premature Apoptosis by an NF-κB-Dependent, Interferon-Independent Mechanism and Facilitate Virus Growth
Authors: Bitko Vira , Shulyayeva Olena , Mazumder Barsanjit , Musiyenko Alla , Ramaswamy Murali , Look Dwight C. , Barik Sailen
Published: December 2006

The two nonstructural (NS) proteins NS1 and NS2 of respiratory syncytial virus (RSV) are abundantly expressed in the infected cell but are not packaged in mature progeny virions. We found that both proteins were expressed early in infection, whereas the infected cells underwent apoptosis much later. Coincident with NS protein expression, a number of cellular antiapoptotic factors were expressed or activated at early stages, which included NF-κB and phosphorylated forms of protein kinases AKT, phosphoinositide-dependent protein kinase, and glycogen synthase kinase. Using specific short interfering RNAs (siRNAs), we achieved significant knockdown of one or both NS proteins in the infected cell, which resulted in abrogation of the antiapoptotic functions and led to early apoptosis. NS-dependent suppression of apoptosis was observed in Vero cells that are naturally devoid of type I interferons (IFN). The siRNA-based results were confirmed by the use of NS-deleted RSV mutants. Early activation of epidermal growth factor receptor (EGFR) in the RSV-infected cell did not require NS proteins. Premature apoptosis triggered by the loss of NS or by apoptosis-promoting drugs caused a severe reduction of RSV growth. Finally, recombinantly expressed NS1 and NS2, individually and together, reduced apoptosis by tumor necrosis factor alpha, suggesting an intrinsic antiapoptotic property of both. We conclude that the early-expressed nonstructural proteins of RSV boost viral replication by delaying the apoptosis of the infected cell via a novel IFN- and EGFR-independent pathway.
Fitness Landscape of Human Immunodeficiency Virus Type 1 Protease Quasispecies
Authors: Fernàndez Guerau , Clotet Bonaventura , Martínez Miguel Angel
Published: December 2006

Here we show, at a high resolution (1%), the human immunodeficiency virus
type 1 (HIV-1) protease gene quasispecies landscape from
three infected naïve individuals. A huge range of genetic
configurations was found (67%, 71%, and 80% of the nucleotide clones
from the three individuals, respectively, were different), and these
configurations created a dense net that linked different parts of the
viral population. Similarly, a vast diversity of different protease
activities was also found. Importantly, 65% of the analyzed enzymes had
detectable protease activity, and 11% of the minority individual
variants showed similar or better fitness than the master (most
abundant) enzyme, suggesting that the viral complexity in this genomic
region does not exclusively depend on the enzyme's catalytic
efficiency. Several high-fitness minority variants had only one
substitution compared to the master sequence, supporting the
possibility that the rugged HIV-1 protease quasispecies fitness
landscape may be formed by a continuous network that can be traversed
by single mutational steps without passing through defective or
less-adapted
proteins.
The Minimal Replicator Element of the Kaposi's Sarcoma-Associated Herpesvirus Terminal Repeat Supports Replication in a Semiconservative and Cell-Cycle-Dependent Manner
Authors: Verma Subhash C. , Choudhuri Tathagata , Robertson Erle S.
Published: December 2006

Kaposi's
sarcoma-associated herpesvirus (KSHV) persists as episomes in infected
cells by circularizing at the terminal repeats (TRs). The KSHV episome
carries multiple reiterated copies of the terminal repeat, and each
copy is capable of supporting replication. Expression of the
latency-associated nuclear antigen (LANA) is critical for the
replication of TR-containing plasmids. A 32-bp sequence upstream of
LANA binding site 1 (LBS1), referred to as RE (replication element),
along with LANA binding sites 1 and 2 (RE-LBS1/2), is sufficient to
support replication (J. Hu and R. Renne, J. Virol.
79:2637-2642, 2005). In this report we demonstrate that the
minimal replicator element (RE-LBS1/2) replicates in synchrony with the
host cellular DNA, and only once, in a cell-cycle-dependent
manner. Overexpression of the mammalian replication inhibitor geminin
blocked replication of the plasmid containing the minimal replicator
element, confirming the involvement of the host cellular replication
control mechanism, and prevented rereplication of the plasmid in the
same cell cycle. Overexpression of Cdt1 also rescued the replicative
ability of the RE-LBS1/2-containing plasmids. A chromatin
immunoprecipitation assay performed using anti-origin recognition
complex 2 (α-ORC2) and α-LANA antibodies from cells
transfected with RE-LBS1/2, RE-LBS1, LBS1, or RE showed the association
of ORC2 with the RE region. Expression of LANA increased the number
of copies of chromatin-bound DNA of replication elements,
suggesting that LANA is important for the recruitment of ORCs and may
contribute to the stabilization of the replication protein complexes at
the RE
site.
Human Cytomegalovirus Disrupts both Ataxia Telangiectasia Mutated Protein (ATM)- and ATM-Rad3-Related Kinase-Mediated DNA Damage Responses during Lytic Infection
Authors: Luo Min Hua , Rosenke Kyle , Czornak Kamila , Fortunato Elizabeth A.
Published: December 2006

Many viruses (herpes simplex virus type 1, polyomavirus, and human immunodeficiency virus type 1) require the activation of ataxia telangiectasia mutated protein (ATM) and/or Mre11 for a fully permissive infection. However, the longer life cycle of human cytomegalovirus (HCMV) may require more specific interactions with the DNA repair machinery to maximize viral replication. A prototypical damage response to the double-stranded ends of the incoming linear viral DNA was not observed in fibroblasts at early times postinfection (p.i.). Apparently, a constant low level of phosphorylated ATM was enough to phosphorylate its downstream targets, p53 and Nbs1. p53 was the only cellular protein observed to relocate at early times, forming foci in infected cell nuclei between 3.5 and 5.5 h p.i. Approximately half of these foci localized with input viral DNA, and all localized with viral UL112/113 prereplication site foci. No other DNA repair proteins localized with the virus or prereplication foci in the first 24 h p.i. When viral replication began in earnest, between 24 and 48 h p.i., there were large increases in steady-state levels and phosphorylation of many proteins involved in the damage response, presumably triggered by ATM-Rad3-related kinase activation. However, a sieving process occurred in which only certain proteins were specifically sequestered into viral replication centers and others were particularly excluded. In contrast to other viruses, activation of a damage response is neither necessary nor detrimental to infection, as neither ATM nor Mre11 was required for full virus replication and production. Thus, by preventing simultaneous relocalization of all the necessary repair components to the replication centers, HCMV subverts full activation and completion of both double-stranded break and S-phase checkpoints that should arrest all replication within the cell and likely lead to apoptosis.
Protective Role of Beta Interferon in Host Defense against Influenza A Virus
Authors: Koerner Iris , Kochs Georg , Kalinke Ulrich , Weiss Siegfried , Staeheli Peter
Published: December 2006

Type I interferon (IFN), which includes the IFN-α and -β subtypes, plays an essential role in host defense against influenza A virus. However, the relative contribution of IFN-β remains unresolved. In mice, type I IFN is effective against influenza viruses only if the IFN-induced resistance factor Mx1 is present, though most inbred mouse strains, including the recently developed IFN-β-deficient mice, bear only defective
Mx1
alleles. We therefore generated IFN-β-deficient mice carrying functional
Mx1
alleles (designated Mx-BKO) and compared them to either wild-type mice bearing functional copies of both
IFN-β
and
Mx1
(designated Mx-wt) or mice carrying functional
Mx1
alleles but lacking functional type I IFN receptors (designated Mx-IFNAR). Influenza A virus strain SC35M (H7N7) grew to high titers and readily formed plaques in monolayers of Mx-BKO and Mx-IFNAR embryo fibroblasts which showed no spontaneous expression of
Mx1
. In contrast, Mx-wt embryo fibroblasts were found to constitutively express
Mx1
, most likely explaining why SC35M did not grow to high titers and formed no visible plaques in such cells. In vivo challenge experiments in which SC35M was applied via the intranasal route showed that the 50% lethal dose was about 20-fold lower in Mx-BKO mice than in Mx-wt mice and that virus titers in the lungs were increased in Mx-BKO mice. The resistance of Mx-BKO mice to influenza A virus strain PR/8/34 (H1N1) was also substantially reduced, demonstrating that IFN-β plays an important role in the defense against influenza A virus that cannot be compensated for by IFN-α.
Persistent Activation of STAT3 by Latent Kaposi's Sarcoma-Associated Herpesvirus Infection of Endothelial Cells
Authors: Punjabi Almira S. , Carroll Patrick A. , Chen Lei , Lagunoff Michael
Published: December 2006

Kaposi's sarcoma-associated herpesvirus (KSHV) is the infectious cause of Kaposi's sarcoma, primary effusion lymphoma, and plasmablastic multicentric Castleman's disease. STAT3 has been shown to be important for the maintenance of primary effusion lymphoma cells in culture and is chronically activated in many tumor cell lines. However, little is known about the role of KSHV in the activation of STAT3 or the role of STAT3 in KS tumors. We demonstrate that STAT3 is activated by KSHV infection of endothelial cells, the KS tumor cell type, in a biphasic fashion. Viral binding and entry activate STAT3 in the first 2 h after infection, but this activation dissipates by 4 h postinfection. By 12 h after KSHV infection, concomitant with the expression of latent genes, STAT3 is once again activated, and this activation persists for as long as latent infection is maintained. Activated STAT3 translocates to the nucleus, where it can bind to STAT3-specific DNA elements and can activate STAT3-dependent promoter activity. Conditioned medium from KSHV-infected endothelial cells is able to transiently activate STAT3, indicating the involvement of a secreted factor and that a latency-associated factor in KSHV-infected cells is necessary for sustained activation. KSHV upregulates gp130 receptor expression, and both gp130 and JAK2 are required for the activation of STAT3. However, neither human nor viral interleukin-6 is required for STAT3 activation. Persistent activation of the oncogenic signal transducer, STAT3, by KSHV may play a critical role in the viral pathogenesis of Kaposi's sarcoma, as well as in primary effusion lymphomas.
Notch1 Augments Intracellular Trafficking of Adeno-Associated Virus Type 2
Authors: Ren Changchun , White April F. , Ponnazhagan Selvarangan
Published: December 2006

We report here the significance of the Notch1 receptor in intracellular trafficking of recombinant adeno-associated virus type 2 (rAAV2). RNA profiling of human prostate cancer cell lines with various degrees of AAV transduction indicated a correlation of the amount of Notch1 with rAAV transgene expression. A definitive role of Notch1 in enhancing AAV transduction was confirmed by developing clonal derivatives of DU145 cells overexpressing either full-length or intracellular Notch1. To discern stages of AAV2 transduction influenced by Notch1, competitive binding with soluble heparin and Notch1 antibody, intracellular trafficking using Cy3-labeled rAAV2, and blocking assays for proteasome and dynamin pathways were performed. Results indicated that in the absence or low-level expression of Notch1, only binding of virus was found on the cell surface and internalization was impaired. However, increased Notch1 expression in these cells allowed efficient perinuclear accumulation of labeled capsids. Nuclear transport of the vector was evident by transgene expression and real-time PCR analyses. Dynamin levels were not found to be different among these cell lines, but blocking dynamin function abrogated AAV2 transduction in DU145 clones overexpressing full-length Notch1 but not in clones overexpressing intracellular Notch1. These studies provide evidence for the role of activated Notch1 in intracellular trafficking of AAV2, which may have implications in the optimal use of AAV2 in human gene therapy.
Recovery of Infectious Bluetongue Virus from RNA
Authors: Boyce Mark , Roy Polly
Published: December 2006

Bluetongue virus (BTV) is an insect-vectored emerging pathogen of ruminants with the potential for devastating economic impact on European agriculture. BTV and many other members of the
Reoviridae
have remained stubbornly refractory to the development of methods for the rescue of infectious virus from cloned nucleic acid (reverse genetics). Partially disassembled virus particles are transcriptionally active, synthesizing viral transcripts in the cytoplasm of infected cells, in essence delivering viral nucleic acids in situ. With the goal of generating a reverse-genetics system for BTV, we examined the possibility of recovering infectious BTV by the transfection of BSR cells with BTV transcripts (single-stranded RNA [ssRNA]) synthesized in vitro using BTV core particles. Following transfection, viral-protein synthesis was detected by immunoblotting, and confocal examination of the cells showed a punctate cytoplasmic distribution of inclusion bodies similar to that seen in infected cells. Viral double-stranded RNA (dsRNA) was isolated from ssRNA-transfected cells, demonstrating that replication of the ssRNA had occurred. Additionally, infectious virus was present in the medium of transfected cells, as demonstrated by the passage of infectivity in BSR cells. Infectivity was sensitive to single-strand-specific RNase A, and cotransfection of genomic BTV dsRNA with transcribed ssRNA demonstrated that the ssRNA species, rather than dsRNA, were the active components. We conclude that it is possible to recover infectious BTV wholly from ssRNA, which suggests a means for establishing helper virus-independent reverse-genetics systems for members of the
Reoviridae
.
EBNA2 Interferes with the Germinal Center Phenotype by Downregulating BCL6 and TCL1 in Non-Hodgkin's Lymphoma Cells
Authors: Boccellato Francesco , Anastasiadou Eleni , Rosato Paola , Kempkes Bettina , Frati Luigi , Faggioni Alberto , Trivedi Pankaj
Published: December 2006

Epstein-Barr virus (EBV)-negative diffuse large B-cell lymphoma (DLBCL) and Burkitt lymphoma-derived cell lines infected in vitro with a recombinant EBV expressed type II/III latency. High expression of EBNA2 inversely correlated with expression of germinal center (GC)-associated genes, BCL6 and TCL1. The decreased expression of BCL6 appeared to be dose dependent, with almost complete abrogation in highly EBNA2-expressing clones. The role of EBNA2 in negative regulation of these genes was confirmed by transfection and in a hormone-inducible EBNA2 cell system. LMP1 transfection reduced expression of TCL1, but not of BCL6, in DLBCLs. The GC-associated gene repression was at the transcriptional level and CBF1 independent. A decrease in HLA-DR, surface immunoglobulin M, and class II transactivator expression and an increase in CCL3, a BCL6 repression target, was observed in EBNA2-expressing clones. Since BCL6 is indispensable for GC formation and somatic hypermutations (SHM), we suggest that the previously reported lack of SHM seen in EBNA2-expressing GC cells from infectious mononucleosis tonsils could be due to negative regulation of BCL6 by EBNA2. These findings suggest that EBNA2 interferes with the GC phenotype.
Deterministic, Compensatory Mutational Events in the Capsid of Foot-and-Mouth Disease Virus in Response to the Introduction of Mutations Found in Viruses from Persistent Infections
Authors: Mateo Roberto , Mateu Mauricio G.
Published: December 2006

The evolution of foot-and-mouth disease virus (FMDV) (biological clone C-S8c1) in persistently infected cells led to the emergence of a variant (R100) that displayed increased virulence, reduced stability, and other modified phenotypic traits. Some mutations fixed in the R100 genome involved a cluster of highly conserved residues around the capsid pores that participate in interactions with each other and/or between capsid protomers. We have investigated phenotypic and genotypic changes that occurred when these replacements were introduced into the C-S8c1 capsid. The C3007V and M3014L mutations exerted no effect on plaque size or viral yield during lytic infections, or on virion stability, but led to a reduction in biological fitness; the D3009A mutation caused drastic reductions in plaque size and viability. Remarkably, competition of the C3007V mutant with the nonmutated virus invariably resulted in the fixation of the D3009A mutation in the C3007V capsid. In turn, the presence of the D3009A mutation invariably led to the fixation of the M3014L mutation. In both cases, two individually disadvantageous mutations led, together, to an increase in fitness, as the double mutants outcompeted the nonmutated genotype. The higher fitness of C3007V/D3009A was related to a faster multiplication rate. These observations provide evidence for a chain of linked, compensatory mutational events in a defined region of the FMDV capsid. Furthermore, they indicate that the clustering of unique amino acid replacements in viruses from persistent infections may also occur in cytolytic infections in response to changes caused by previous mutations without an involvement of the new mutations in the adaptation to a different environment.