bluuebunny/crossref_metadata_2025 huggingface dataset:
Binaural sensitivity as a function of interaural electrode position with a bilateral cochlear implant user
Authors: Long Christopher J. , Eddington Donald K. , Colburn H. Steven , Rabinowitz William M.
Published: August 2003

Experiments were conducted with a single, bilateral cochlear implant user to examine interaural level and time-delay cues that putatively underlie the design and efficacy of bilateral implant systems. The subject’s two implants were of different types but custom equipment allowed presentation of controlled bilateral stimuli, particularly those with specified interaural time difference (ITD) and interaural level difference (ILD) cues. A lateralization task was used to measure the effect of these cues on the perceived location of the sensations elicited. For trains of fixed-amplitude, biphasic current pulses at 100 pps, the subject demonstrated sensitivity to an ITD of 300 μs, providing evidence of access to binaural information. The choice of bilateral electrode pair greatly influenced ITD sensitivity, suggesting that electrode pairings are likely to be an important consideration in the effort to provide binaural advantages. The selection of bilateral electrode pairs showing sensitivity to ITD was partially aided by comparisons of the pitch elicited by individual electrodes in each ear (when stimulated alone with fixed-amplitude current pulses at 813 pps): specifically, interaural electrodes with similar pitches were more likely (but not certain) to show ITD sensitivity. Significant changes in lateral position occurred with specific electrode pairs. With five bilateral electrode pairs of 14 tested, ITDs of 300 and 600 μs moved an auditory image significantly from right to left. With these same pairs, ILD changes of ≈11% of the dynamic range (in μApp) moved an auditory image from the far left to the far right—significantly farther than the nine pairs not showing significant ITD sensitivity. However, even these nine pairs did show response changes as a function of the interaural (or confounding monaural) level cue. Overall, insofar as the access to bilateral cues demonstrated herein generalizes to other subjects, it provides hope that the normal binaural advantages for speech recognition and sound localization can be made available to bilateral implant users.
Time course of loudness recalibration: Implications for loudness enhancement
Authors: Arieh Yoav , Marks Lawrence E.
Published: August 2003

Loudness recalibration, the effect of a relatively loud 2500-Hz recalibrating tone on the loudness of a relatively soft 2500-Hz target tone, was measured as a function of the interstimulus interval (ISI) between them. The loudness of the target tone, assessed by a 500-Hz comparison tone, declined when the ISI equaled or exceeded about 200 ms and leveled off at an ISI of about 700 ms. Notably, the target tone’s loudness did not change significantly at very short ISIs (<150 ms). The latter result is incompatible with the literature reporting loudness enhancement in this time window, but is compatible with the suggestion made by Scharf, Buus, and Nieder [J. Acoust. Soc. Am. 112, 807–810 (2002)] that early measurements of enhancement were contaminated by the influence of the recalibrating tone on the comparison tone when the two shared the same frequency. In a second experiment the frequency of the comparison tone was changed to 2500 Hz and the results of a loudness enhancement paradigm was successfully predicted from the time course of recalibration obtained in experiment 1.
Oscillatory limited compressible fluid flow induced by the radial motion of a thick-walled piezoelectric tube
Authors: Shin Dong-Youn , Grassia Paul , Derby Brian
Published: August 2003

A simple oscillatory, slightly compressible, fluid flow model in a thick-walled piezoelectric tube used in a drop-on-demand inkjet print head is developed from the point of view of fluid-structure interaction to take account of pressure wave propagation and pressure loading opposing wall motion. A frequency sweep is performed computationally using the model revealing the first acoustic fluid-structure resonance frequency and the influence of fluid viscosity. The validity of the model, with given information on the speed of sound in a fluid, is evaluated by comparing the theoretically predicted resonance frequency to the experimentally measured resonance frequency. In addition, the intrinsic speed of sound can be easily computed using the measured acoustic resonance frequency and this computed speed of sound agrees closely with speeds of sound reported in the literature.
A dynamic Young’s modulus measurement system for highly compliant polymers
Authors: Guillot François M. , Trivett D. H.
Published: August 2003

A new system to determine experimentally the complex Young’s modulus of highly compliant elastomers at elevated hydrostatic pressures and as a function of temperature is presented. A sample cut in the shape of a bar is adhered to a piezoelectric ceramic shaker and mounted vertically inside a pressure vessel equipped with glass windows. Two independent measurement methods are then used: a resonant technique, to obtain low-frequency data, and a wave propagation technique, to obtain higher-frequency data. Both techniques are implemented utilizing laser Doppler vibrometers. One vibrometer detects sample resonances through a window located at the bottom of the pressure vessel, and a set of two separate vibrometers monitors the speed of longitudinal waves propagating in the sample, through windows located on the sides of the vessel. The apparatus is contained inside an environmental chamber for temperature control. Using this approach, Young’s modulus data can be obtained at frequencies typically ranging from 100 Hz to 5 kHz, under hydrostatic pressures ranging from 0 to 2.07 MPa (300 psi), and at temperatures between −2 °C and 50 °C. Experimental results obtained on two commercial materials, Rubatex® R451N and Goodrich Thorodin™ AQ21, are presented. The effects of lateral inertia, resulting in dispersive wave propagation, are discussed and their impacts on Young’s modulus measurements are examined.
Cochlear compression estimates from measurements of distortion-product otoacoustic emissions
Authors: Neely Stephen T. , Gorga Michael P. , Dorn Patricia A.
Published: August 2003

Evidence of the compressive growth of basilar-membrane displacement can be seen in distortion-product otoacoustic emission (DPOAE) levels measured as a function of stimulus level. When the levels of the two stimulus tones (f1 and f2) are related by the formula L1=39 dB+0.4⋅L2 [Kummer et al., J. Acoust. Soc. Am. 103, 3431–3444 (1998)] the shape of the function relating DPOAE level to L2 is similar (up to an L2 of 70 dB SPL) to the classic Fletcher and Munson [J. Acoust. Soc. Am. 9, 1–10 (1933)] loudness function when plotted on a logarithmic scale. Explicit estimates of compression have been derived based on recent DPOAE measurements from the laboratory. If DPOAE growth rate is defined as the slope of the DPOAE I/O function (in dB/dB), then a cogent definition of compression is the reciprocal of the growth rate. In humans with normal hearing, compression varies from about 1 at threshold to about 4 at 70 dB SPL. With hearing loss, compression is still about 1 at threshold, but grows more slowly above threshold. Median DPOAE I/O data from ears with normal hearing, mild loss, and moderate loss are each well fit by log functions. When the I/O function is logarithmic, then the corresponding compression is a linear function of stimulus level. Evidence of cochlear compression also exists in DPOAE suppression tuning curves, which indicate the level of a third stimulus tone (f3) that reduces DPOAE level by 3 dB. All three stimulus tones generate compressive growth within the cochlea; however, only the relative compression (RC) of the primary and suppressor responses is observable in DPOAE suppression data. An RC value of 1 indicates that the cochlear responses to the primary and suppressor components grow at the same rate. In normal ears, RC rises to 4, when f3 is an octave below f2. The similarities between DPOAE and loudness compression estimates suggest the possibility of predicting loudness growth from DPOAEs; however, intersubject variability makes such predictions difficult at this time.
A time-selective technique for free-field reciprocity calibration of condenser microphones
Authors: Barrera-Figueroa Salvador , Rasmussen Knud , Jacobsen Finn
Published: August 2003

In normal practice, microphones are calibrated in a closed coupler where the sound pressure is uniformly distributed over the diaphragm. Alternatively, microphones can be placed in a free field, although in that case the distribution of sound pressure over the diaphragm will change as a result of the diffraction of the body of the microphone, and thus, its sensitivity will change. In the two cases, a technique based on the reciprocity theorem can be applied for obtaining the absolute sensitivity either under uniform pressure or free-field conditions. In this paper, signal-processing techniques are considered that improve the accuracy of the free-field calibration method. In particular, a fast Fourier transform (FFT)-based time-selective technique for removing undesired reflections from the walls of the measurement chamber has been developed and applied to the electric transfer impedance function between two microphones. The acoustic centers of the microphones have been determined from the “cleaned” transfer impedance values. Then, the complex free-field sensitivities of the microphones have been calculated. The resulting complex sensitivities and acoustic centers have proved to be in good agreement with previously published data, and this confirms the reliability of the time-selective technique, even in nonanechoic environments. Furthermore, the obtained results give a new reference for further comparisons, because they cover a frequency range with an accuracy that has not been obtained by previously published data.
Broadband performance of a moving time reversing array
Authors: Sabra Karim G. , Dowling David R.
Published: August 2003

Acoustic time reversal exploits reciprocity between sources and receivers to generate backward propagating waves that automatically focus at their point of origin. In underwater acoustics, an array of transducers that can both transmit and receive, referred to as a time reversing array (TRA) or time reversal mirror (TRM), generates the back-propagating waves. Such arrays have been shown to spatially and temporally focus sound in unknown complicated multipath environments, and are therefore of interest for active sonar and underwater communication applications. Although stationary vertical linear TRAs have been favored in prior studies, practical applications of acoustic time reversal in underwater environments are likely to involve towed, tilted, horizontal, or bottom-mounted arrays. In particular, array motion introduces Doppler effects and eliminates source–receiver reciprocity, two factors that potentially impact the automatic focusing capability of TRAs. This paper presents the results from a theoretical and computational investigation into how array motion and orientation influence TRA retrofocusing in the shallow ocean. Here, the TRA tow speed is assumed constant, and the array is assumed to be straight and linear (vertical, horizontal, or tilted). And, for simplicity, the TRA is assumed to respond to a stationary point source emitting a broadband pulse. When a TRA moves, the retrofocus is predicted to shift in the direction of array motion due to the translation of the array between its reception and broadcast times. In addition, the performance of a towed horizontal TRA is predicted to degrade more rapidly with towing speed than that of an equivalent (but clearly idealized) towed vertical array because of range-dependent Doppler phase differences that do not influence the vertical array. However, short tilted arrays may approach vertical array performance and appear to be a potentially versatile compromise for implementing TRA concepts in active sonar or underwater communication systems.
A fuzzy rule based framework for noise annoyance modeling
Authors: Botteldooren Dick , Verkeyn Andy , Lercher Peter
Published: August 2003

Predicting the effect of noise on individual people and small groups is an extremely difficult task due to the influence of a multitude of factors that vary from person to person and from context to context. Moreover, noise annoyance is inherently a vague concept. That is why, in this paper, it is argued that noise annoyance models should identify a fuzzy set of possible effects rather than seek a very accurate crisp prediction. Fuzzy rule based models seem ideal candidates for this task. This paper provides the theoretical background for building these models. Existing empirical knowledge is used to extract a few typical rules that allow making the model more specific for small groups of individuals. The resulting model is tested on two large-scale social surveys augmented with exposure simulations. The testing demonstrates how this new way of thinking about noise effect modeling can be used in practice both in management support as a “noise annoyance adviser” and in social science for testing hypotheses such as the effect of noise sensitivity or the degree of urbanization.
Chasing 100
Authors: Chen Ingfei
Published: September 2003

When geriatrician Thomas Perls treated his first centenarians in 1992, he expected to find them debilitated by illness and dementia. But he instead discovered that some extremely old individuals can escape Alzheimer's disease and other age-related disorders. In 1994 at Harvard Medical School, he started the New England Centenarian Study (NECS), a survey that has grown into an international investigation with data on 1000 centenarians from the United States, Canada, Germany, Japan, and Australia. Now based at the Boston University School of Medicine, the study has toppled several misconceptions about extremely old age and suggests a genetic basis to exceptional longevity. The work led to the founding of a biotech company called Centagenetix, which subsequently merged with Elixir Pharmaceuticals.
Autophagy and Aging--When "All You Can Eat" Is Yourself
Authors: Cuervo Ana Maria
Published: September 2003

A recent paper provides evidence that macroautophagy is an essential downstream pathway for one of the mutations known to extend life span. Autophagy, or the degradation of intracellular components by the lysosomal system, was thought for a long time to be a catabolic process responsible for cellular cleanup. However, in recent years, we have learned that autophagy comes in different sizes and shapes, macroautophagy being one of them, and that this cellular maid plays many more roles than previously anticipated. Activation of autophagy is essential in physiological processes as diverse as morphogenesis, cellular differentiation, tissue remodeling, and cellular defense, among others. Furthermore, its participation in different pathological conditions, including cancer and neurodegeneration, is presently a subject of intense investigation. A role in aging has now been added to this growing list of autophagy functions. The activity of different forms of autophagy decreases with age, and this reduced function has been blamed for the accumulation of damaged proteins in old organisms. Research such as that covered in this Perspective shows that there is much more than trash to worry about when autophagy is not functioning properly.
Barking up the Tree of Long Life
Authors: Beckman Mary
Published: September 2003

The dog days of an old theory might be upon us. New research shows that the more energy a dog uses in its lifetime, the longer it lives. The results contradict a 1928 hypothesis that the faster an organism's metabolic rate, the more quickly it ages. The data also challenge the idea that leanness enhances longevity.
How Does the Huntington's Disease Mutation Damage Cells?
Authors: Rubinsztein David C.
Published: September 2003

Huntington's Disease (HD) is an autosomal dominant neurodegenerative condition with devastating consequences. HD is caused by the expansion of a CAG trinucleotide repeat stretch in the coding sequence of the HD gene that gives rise to a long polyglutamine tract in the huntingtin protein. How this mutated protein gives rise to the disease state is controversial. In this Perspective, I discuss the results of a new study on the effects of the mutated huntingtin protein in light of previous findings and suggest that the HD mutation damages cells by perturbing multiple parallel pathways by gain-of-function and possibly also dominant negative mechanisms.
p53- and Drug-Induced Apoptotic Responses Mediated by BH3-Only Proteins Puma and Noxa
Authors: Villunger Andreas , Michalak Ewa M. , Coultas Leigh , Müllauer Franziska , Böck Günther , Ausserlechner Michael J. , Adams Jerry M. , Strasser Andreas
Published: September 2003

Apoptosis provoked by DNA damage requires the p53 tumor suppressor, but which of the many p53-regulated genes are required has remained unknown. Two genes induced by this transcription factor,
noxa
and
puma
(
bbc3
), stand out, because they encode BH3-only proteins, proapoptotic members of the Bcl-2 family required to initiate apoptosis. In mice with either
noxa
or
puma
disrupted, we observed decreased DNA damage–induced apoptosis in fibroblasts, although only loss of Puma protected lymphocytes from cell death. Puma deficiency also protected cells against diverse p53-independent cytotoxic insults, including cytokine deprivation and exposure to glucocorticoids, the kinase inhibitor staurosporine, or phorbol ester. Hence, Puma and Noxa are critical mediators of the apoptotic responses induced by p53 and other agents.
T Cell-Dendritic Cell Interaction in Vivo: Random Encounters Favor Development of Long-Lasting Ties
Authors: Acuto Oreste
Published: September 2003

Understanding the complexity of the functional communication between cells composing the immune system is central to improving our capacity to manipulate it and conceive better strategies to combat microbial pathogens. So far, these studies have been based on immunohistochemistry of fixed tissues and in vitro attempts to reproduce functional connections between cells. The application of two-photon laser microscopy to the observation of viable immune cells in their natural environment where foreign antigens are carried to trigger an immune response opens a new era for these studies. They reveal exceptional properties of the locomotion of T cells that facilitate encounters with dendritic cells and the receipt of information that promotes T cell survival, death, or initiation of immune responses. These studies also complement in vitro observations addressing the importance of time of stimulation in determining T cell fates.
The Wnt/β-catenin Pathway
Authors: Moon Randall T.
Published: September 2003

Wnts are a family of secreted glycoproteins that act as ligands to activate receptor-mediated signaling pathways. The first described Wnt pathway, and the best understood, is one that works through a cytoplasmic protein, β-catenin. Specific members of the Wnt family interact with a Frizzled receptor complex, promoting the activity of Dishevelled, which in turn enhances the stability and activity of β-catenin. β-catenin probably binds members of the T cell factor and lymphoid enhancer factor (TCF/LEF) family of transcription factors, or to various other proteins, to regulate expression of target genes in a cellular context-dependent manner. Wnt signaling through this pathway regulates cell proliferation, survival, and cell fate. Inappropriate activation of this pathway may be an initiating event in some cancers, and inhibiting this pathway in transformed cells shows promise for anticancer therapies. Conversely, activation of this pathway shows promise as a target for developing osteoporosis, neurodegenerative disease, and stem cell therapies. The Connections Map for this Pathway provides an overview of the many signals that converge on β-catenin and influence Wnt signaling.

Science Viewpoint

R. T. Moon, B. Bowerman, M. Boutros, and N. Perrimon, The promise and perils of Wnt signaling through β-catenin.
Science
296
, 1644-1646 (2002). [
Abstract
] [
Full Text
]
Dependence Receptors: What Is the Mechanism?
Authors: Chao Moses V.
Published: September 2003

Receptors of diverse primary structure and with diverse ligands have been reported to be capable of stimulating apoptosis in the absence of ligand binding. These receptors are called dependence receptors, and the newest member of this family appears to be the Sonic hedgehog receptor Patched, which has been reported to stimulate apoptosis when expressed in the absence of its ligand. The signaling mechanisms that account for this type of receptor activity are unknown. Several theories behind how dependence receptors may trigger cell death are described.
Mutations in the DNA Mismatch Repair Proteins MutS and MutL of Oxazolidinone-Resistant or -Susceptible
Enterococcus faecium
Authors: Willems Rob J. , Top Janetta , Smith Derek J. , Roper David I. , North Sarah E. , Woodford Neil
Published: September 2003

Mutations in
mutS
and
mutL
, which encode DNA mismatch repair (MMR) proteins, can confer hypermutator phenotypes and may facilitate the emergence of mutational antibiotic resistance in bacteria. Linezolid-resistant enterococci (LRE) rarely emerge during therapy and contain mutations in 23S rRNA genes. As enterococci with defective MMR could be prone to the development of oxazolidinone resistance mutations, we investigated 13 clinical isolates of
Enterococcus faecium
, including 2 LRE, for mutations in
mutSL
. A 4,944-bp fragment spanning
mutSL
was sequenced from two pairs of linezolid-resistant (MICs, 64 μg/ml) and linezolid-susceptible (MICs, 2 μg/ml)
E. faecium
isolates (one pair from Austria and one pair from the United Kingdom) identical by pulsed-field gel electrophoresis. The pairs represented distinct strains in which linezolid resistance had emerged during therapy. The MutSL peptides of all four isolates had amino acid substitutions compared with the sequence of
E. faecium
strain DO (used for genome sequencing). These were Val352Ile (one pair of isolates only) and Met628Leu in MutS and Leu387Pro, Tyr406Phe, Thr415Ser, Phe427Leu, and Phe565Ile in MutL. The significance of these changes remains unknown; these isolates did not show a demonstrable hypermutator phenotype. The same substitutions were found in two of nine geographically diverse linezolid-susceptible enterococcal isolates; the other seven isolates had MutSL sequences identical to that of strain DO. Multilocus sequence typing revealed that all isolates with alternate MutSL peptides belonged to a distinct lineage of a prevalent
E. faecium
clonal complex, designated CC17. Further studies are needed to investigate the prevalence of these MutSL mutations and their possible roles in the emergence of
E. faecium
strains resistant to oxazolidinones and other antibiotic classes.
In Vitro Susceptibilities of Gram-Negative Bacteria Isolated from Hospitalized Patients in Four European Countries, Canada, and the United States in 2000-2001 to Expanded-Spectrum Cephalosporins and Comparator Antimicrobials: Implications for Therapy
Authors: Wenzel Richard P. , Sahm Daniel F. , Thornsberry Clyde , Draghi Deborah C. , Jones Mark E. , Karlowsky James A.
Published: September 2003

Access to current antimicrobial agent surveillance data is an important prerequisite for the optimal management of patients with hospital-acquired infections. The present study used data collected in 2000 to 2001 from 670 laboratories in Europe (France, Germany, Italy, and Spain), Canada, and the United States to report on the in vitro activities of ceftriaxone, cefotaxime, and comparative agents against >125,000 isolates of gram-negative bacteria from hospitalized patients. All but two isolates of
Enterobacteriaceae
(one isolate of
Proteus mirabilis
from France and one isolate of
Morganella morganii
from Canada) were susceptible to imipenem. The susceptibility of
Escherichia coli
to ceftriaxone or cefotaxime was ≥97% in each country, and for
P. mirabilis
, susceptibility was 99% in each country except Italy. In contrast, susceptibility of
E. coli
to ciprofloxacin varied from 80.5% (Spain) to 94.0% (France); levofloxacin susceptibility ranged from 75.2% (Spain) to 91.6% (United States). Among
Klebsiella pneumoniae
and
Klebsiella oxytoca
isolates, ceftriaxone and cefotaxime susceptibilities ranged from 86.6 to 98.7% and 83.5 to 99.7%, respectively, depending upon the country. Considerable geographic variation in the susceptibilities (generally 85 to 95% susceptible) of
Serratia marcescens
and
M. morganii
to ceftriaxone and cefotaxime were observed. For
S. marcescens
, susceptibility to piperacillin-tazobactam varied from 81.5% (France) to 94.1% (Italy) and susceptibility to ciprofloxacin ranged from 66.2% (Germany) to 90.7% (Spain).
Enterobacter cloacae
and
Enterobacter aerogenes
were less susceptible to ceftriaxone and cefotaxime than were the other species of
Enterobacteriaceae
studied. The present study demonstrated that established parenteral expanded-spectrum cephalosporin antimicrobial agents retain significant in vitro activity against many clinically important gram-negative pathogens.
Rapid Direct Method for Monitoring Antibiotics in a Mouse Model of Bacterial Biofilm Infection
Authors: Kadurugamuwa Jagath L. , Sin Lin V. , Yu Jun , Francis Kevin P. , Kimura Richard , Purchio Tony , Contag Pamela R.
Published: September 2003

We have developed a rapid, continuous method for monitoring the effectiveness of several antibacterial agents in real time, noninvasively, by using a recently described mouse model of chronic biofilm infection (J. L. Kadurugamuwa et al., Infect. Immun. 71:882-890, 2003), which relies on biophotonic imaging of bioluminescent bacteria. To facilitate real-time monitoring of infection, we used a
Staphylococcus aureus
isolate that was made bioluminescent by inserting a modified
lux
operon into the bacterial chromosome. This bioluminescent reporter bacterium was used to study the antimicrobial effects of several antibiotics belonging to different molecular families. Treatment with rifampin, tobramycin, and ciprofloxacin was started 7 days after subcutaneous implantation of catheters precolonized with 10
4
CFU of
S. aureus
. Three different doses of antibiotics were administered twice a day for 4 consecutive days. The number of metabolically active bacteria in untreated mice and the tobramycin- and ciprofloxacin-treated groups remained relatively unchanged over the 4-week observation period, indicating poor efficacies for tobramycin and ciprofloxacin. A rapid dose-dependent decline in metabolic activity in rifampin-treated groups was observed, with almost a 90% reduction after two doses and nearly undetectable levels after three doses. The disappearance of light emission correlated with colony counts. After the final treatment, cell numbers rebounded as a function of concentration in a time-dependent manner. The staphylococci isolated from the catheters of mice treated with rifampin were uniformly resistant to rifampin but retained their in vitro susceptibilities to tobramycin and ciprofloxacin. Since the metabolic activities of viable cells and a postantibiotic effect could be detected directly on the support matrix nondestructively and noninvasively, the methodology is specifically appealing for investigating the effects of antibiotics on biofilms in vivo. Moreover, our study points to the possible use of biophotonic imaging for the detection of the development of resistance to therapeutic agents during treatment of chronic infections in vivo.
Antimicrobial Resistance Genes in Enterotoxigenic
Escherichia coli
O149:K91 Isolates Obtained over a 23-Year Period from Pigs
Authors: Maynard Christine , Fairbrother John M. , Bekal Sadjia , Sanschagrin François , Levesque Roger C. , Brousseau Roland , Masson Luke , Larivière Serge , Harel Josée
Published: September 2003

A total of 112
Escherichia coli
O149:K91 strains isolated from pigs with diarrhea in Quebec, Canada, between 1978 and 2000 were characterized for their genotypic antimicrobial resistance profiles. Tests for resistance to 10 antimicrobial agents were conducted. Resistance to tetracycline and sulfonamides was found to be the most frequent, but resistance to cefotaxime and ceftiofur was absent. An increase in the number of isolates resistant to at least three antimicrobials was observed over time. The distribution of 28 resistance genes covering six antimicrobial families (beta-lactams, aminoglycosides, phenicols, tetracycline, trimethoprim, and sulfonamides) was assessed by colony hybridization. Significant differences in the distributions of tetracycline [
tet
(A),
tet
(B),
tet
(C)], trimethoprim (
dhfrI
,
dhfrV
,
dhfrXIII
), and sulfonamide (
sulI
,
sulII
) resistance genes were observed during the study period (1978 to 2000). Sixty percent of the isolates possessed a class 1 integron, illustrating the importance of integrons in the epidemiology of antibiotic resistance in
E. coli
strains from pigs. Amplification of the integron's variable region resulted in four distinct fragments of 1, 1.3, 1.6, and 1.8 kb, with the 1.6- and 1.8-kb fragments appearing only during the last half of the study period. Examination of linkages among the different resistance genes showed a variety of positive and negative associations. Association analysis of isolates divided into two groups, those isolated between 1978 and 1989 and those isolated between 1990 and 2000, revealed the appearance of new positive resistance gene associations. Our genotypic resistance analyses of ETEC isolates from pigs indicate that many of the antibiotic resistance genes behind phenotypic resistance are not static but, rather, are in a state of flux driven by various selection forces such as the use of specific antimicrobials.
Effects of Tylosin on Bacterial Mucolysis,Clostridium perfringensColonization, and Intestinal Barrier Function in a Chick Model of Necrotic Enteritis
Authors: Collier C. T. , van der Klis J. D. , Deplancke B. , Anderson D. B. , Gaskins H. R.
Published: September 2003

Necrotic enteritis (NE) is a worldwide poultry disease caused by the alpha toxin-producing bacteriumClostridium perfringens. Disease risk factors include concurrent coccidial infection and the dietary use of cereal grains high in nonstarch polysaccharides (NSP), such as wheat, barley, rye, and oats. Outbreaks of NE can be prevented or treated by the use of in-feed antibiotics. However, the current debate regarding the prophylactic use of antibiotics in animal diets necessitates a better understanding of factors that influence intestinal colonization byC. perfringensas well as the pathophysiological consequences of its growth. We report a study with a chick model of NE, which used molecular (16S rRNA gene [16S rDNA]) and culture-based microbiological techniques to investigate the impact of the macrolide antibiotic tylosin phosphate (100 ppm) and a dietary NSP (pectin) on the community structure of the small intestinal microbiota relative to colonization byC. perfringens. The effects of tylosin and pectin on mucolytic activity of the microbiota andC. perfringenscolonization and their relationship to pathological indices of NE were of particular interest. The data demonstrate that tylosin reduced the percentage of mucolytic bacteria in general and the concentration ofC. perfringensin particular, and these responses correlated in a temporal fashion with a reduction in the occurrence of NE lesions and an improvement in barrier function. The presence of pectin did not significantly affect the variables measured. Thus, it appears that tylosin can control NE through its modulation ofC. perfringenscolonization and the mucolytic activity of the intestinal microbiota.
Effects of Lipid-Based Oral Formulations on Plasma and Tissue Amphotericin B Concentrations and Renal Toxicity in Male Rats
Authors: Risovic Verica , Boyd Michael , Choo Eugene , Wasan Kishor M.
Published: September 2003

The purpose of this study was to determine the effects of various lipid and mixed-micelle formulations on the oral absorption and renal toxicity of amphotericin B (AMB) in rats. The maximum concentration of AMB in plasma and the area under the concentration-time curve for 0 to 24 h for AMB were elevated in rats administered triglyceride (TG)-rich AMB formulations in comparison to those in rats given (i) AMB preformulated as a micelle containing sodium deoxycholate with sodium phosphate as a buffer (DOC-AMB), (ii) an AMB-lipid complex suspension, or (iii) AMB solubilized in methanol. Furthermore, our findings suggest that AMB incorporated into TG-based oral formulations has less renal toxicity than DOC-AMB.
Activity of Human β-Defensin 3 Alone or Combined with Other Antimicrobial Agents against Oral Bacteria
Authors: Maisetta Giuseppantonio , Batoni Giovanna , Esin Semih , Luperini Filippo , Pardini Manuela , Bottai Daria , Florio Walter , Giuca Maria Rita , Gabriele Mario , Campa Mario
Published: September 2003

The in vitro activities of human β-defensin 3 (hBD-3) alone or combined with lysozyme, metronidazole, amoxicillin, and chlorhexidine were investigated with the oral bacteria
Streptococcus mutans
,
Streptococcus sanguinis
,
Streptococcus sobrinus
,
Lactobacillus acidophilus
,
Actinobacillus actinomycetemcomitans
, and
Porphyromonas gingivalis
. hBD-3 showed bactericidal activity against all of the bacterial species tested. The bactericidal effect was enhanced when the peptide was used in combination with the antimicrobial agents mentioned above.
Chimeric Analysis of AcrA Function Reveals the Importance of Its C-Terminal Domain in Its Interaction with the AcrB Multidrug Efflux Pump
Authors: Elkins Christopher A. , Nikaido Hiroshi
Published: August 2003

AcrAB-TolC is the major, constitutively expressed efflux protein complex that provides resistance to a variety of antimicrobial agents in
Escherichia coli
. Previous studies showed that AcrA, a periplasmic protein of the membrane fusion protein family, could function with at least two other resistance-nodulation-division family pumps, AcrD and AcrF, in addition to its cognate partner, AcrB. We found that, among other
E
.
coli
resistance-nodulation-division pumps, YhiV, but not MdtB or MdtC, could also function with AcrA. When AcrB was assessed for the capacity to function with AcrA homologs, only AcrE, but not YhiU or MdtA, could complement an AcrA deficiency. Since AcrA could, but YhiU could not, function with AcrB, we engineered a series of chimeric mutants of these proteins in order to determine the domain(s) of AcrA that is required for its support of AcrB function. The 290-residue N-terminal segment of the 398-residue protein AcrA could be replaced with a sequence coding for the corresponding region of YhiU, but replacement of the region between residues 290 and 357 produced a protein incapable of functioning with AcrB. In contrast, the replacement of residues 357 through 397 of AcrA still produced a functional protein. We conclude that a small region of AcrA close to, but not at, its C terminus is involved in the interaction with its cognate pump protein, AcrB.
Mycobacterium tuberculosis
FurA Autoregulates Its Own Expression
Authors: Sala Claudia , Forti Francesca , Di Florio Elisabetta , Canneva Fabio , Milano Anna , Riccardi Giovanna , Ghisotti Daniela
Published: August 2003

The
furA-katG
region of
Mycobacterium tuberculosis
, encoding a Fur-like protein and the catalase-peroxidase, is highly conserved among mycobacteria. Both genes are induced upon oxidative stress. In this work we analyzed the
M. tuberculosis furA
promoter region. DNA fragments were cloned upstream of the luciferase reporter gene, and promoter activity in
Mycobacterium smegmatis
was measured in both the presence and absence of oxidative stress. The shortest fragment containing an inducible promoter extends 45 bp upstream of
furA
. In this region, −35 and −10 promoter consensus sequences can be identified, as well as a 23-bp AT-rich sequence that is conserved in the nonpathogenic but closely related
M. smegmatis
.
M. tuberculosis
FurA was purified and found to bind upstream of
furA
by gel shift analysis. A ca. 30-bp DNA sequence, centered on the AT-rich region, was essential for FurA binding and protected by FurA in footprinting analysis. Peroxide treatment of FurA abolished DNA binding. Three different AT-rich sequences mutagenized by site-directed mutagenesis were constructed. In each mutant, both
M. tuberculosis
FurA binding in vitro and
pfurA
regulation upon oxidative-stress in
M. smegmatis
were abolished. Thus,
pfurA
is an oxidative stress-responsive promoter controlled by the FurA protein.
Arsenic Sensing and Resistance System in the Cyanobacterium
Synechocystis
sp. Strain PCC 6803
Authors: López-Maury Luis , Florencio Francisco J. , Reyes José C.
Published: August 2003

Arsenic is one of the most important global environmental pollutants. Here we show that the cyanobacterium
Synechocystis
sp. strain PCC 6803 contains an arsenic and antimony resistance operon consisting of three genes:
arsB
, encoding a putative arsenite and antimonite carrier,
arsH
, encoding a protein of unknown function, and
arsC
, encoding a putative arsenate reductase. While
arsB
mutant strains were sensitive to arsenite, arsenate, and antimonite,
arsC
mutants were sensitive only to arsenate. The
arsH
mutant strain showed no obvious phenotype under the conditions tested. In vivo the
arsBHC
operon was derepressed by oxyanions of arsenic and antimony (oxidation state, +3) and, to a lesser extent, by bismuth (oxidation state, +3) and arsenate (oxidation state, +5). In the absence of these effectors, the operon was repressed by a transcription repressor of the ArsR/SmtB family, encoded by an unlinked gene termed
arsR
. Thus,
arsR
null mutants showed constitutive derepression of the
arsBHC
operon. Expression of the
arsR
gene was not altered by the presence of arsenic or antimony compounds. Purified recombinant ArsR protein binds to the
arsBHC
promoter-operator region in the absence of metals and dissociates from the DNA in the presence of Sb(III) or As(III) but not in the presence of As(V), suggesting that trivalent metalloids are the true inducers of the system. DNase I footprinting experiments indicate that ArsR binds to two 17-bp direct repeats, with each one consisting of two inverted repeats, in the region from nucleotides −34 to + 17 of the
arsBHC
promoter-operator.
Effect of Translational Signals on mRNA Decay in
Bacillus subtilis
Authors: Sharp Josh S. , Bechhofer David H.
Published: August 2003

A 254-nucleotide model mRNA, designated Δ
ermC
mRNA, was used to study the effects of translational signals and ribosome transit on mRNA decay in
Bacillus subtilis
. Δ
ermC
mRNA features a strong ribosome-binding site (RBS) and a 62-amino-acid-encoding open reading frame, followed by a transcription terminator structure. Inactivation of the RBS or the start codon resulted in a fourfold decrease in the mRNA half-life, demonstrating the importance of ternary complex formation for mRNA stability. Data for the decay of Δ
ermC
mRNAs with stop codons at positions increasingly proximal to the translational start site showed that actual translation—even the formation of the first peptide bond—was not important for stability. The half-life of an untranslated 3.2-kb Δ
ermC
-
lacZ
fusion RNA was similar to that of a translated Δ
ermC
-
lacZ
mRNA, indicating that the translation of even a longer RNA was not required for wild-type stability. The data are consistent with a model in which ribosome binding and the formation of the ternary complex interfere with a 5′-end-dependent activity, possibly a 5′-binding endonuclease, which is required for the initiation of mRNA decay. This model is supported by the finding that increasing the distance from the 5′ end to the start codon resulted in a 2.5-fold decrease in the mRNA half-life. These results underscore the importance of the 5′ end to mRNA stability in
B
.
subtilis
.
YqfS from
Bacillus subtilis
Is a Spore Protein and a New Functional Member of the Type IV Apurinic/Apyrimidinic-Endonuclease Family
Authors: Salas-Pacheco José M. , Urtiz-Estrada Norma , Martínez-Cadena Guadalupe , Yasbin Ronald E. , Pedraza-Reyes Mario
Published: August 2003

The enzymatic properties and the physiological function of the type IV apurinic/apyrimidinic (AP)-endonuclease homolog of
Bacillus subtilis
, encoded by
yqfS
, a gene specifically expressed in spores, were studied here. To this end, a recombinant YqfS protein containing an N-terminal His
6
tag was synthesized in
Escherichia coli
and purified to homogeneity. An anti-His
6
-YqfS polyclonal antibody exclusively localized YqfS in cell extracts prepared from
B
.
subtilis
spores. The His
6
-YqfS protein demonstrated enzymatic properties characteristic of the type IV family of DNA repair enzymes, such as AP-endonucleases and 3′-phosphatases. However, the purified protein lacked both 5′-phosphatase and exonuclease III activities. YqfS showed not only a high level of amino acid identity with
E. coli
Nfo but also a high resistance to inactivation by EDTA, in the presence of DNA containing AP sites (AP-DNA). These results suggest that YqfS possesses a trinuclear Zn center in which the three metal atoms are intimately coordinated by nine conserved basic residues and two water molecules. Electrophoretic mobility shift assays demonstrated that YqfS possesses structural properties that permit it to bind and scan undamaged DNA as well as to strongly interact with AP-DNA. The ability of
yqfS
to genetically complement the DNA repair deficiency of an
E. coli
mutant lacking the major AP-endonucleases Nfo and exonuclease III strongly suggests that its product confers protection to cells against the deleterious effects of oxidative promoters and alkylating agents. Thus, we conclude that YqfS of
B. subtilis
is a spore-specific protein that has structural and enzymatic properties required to participate in the repair of AP sites and 3′ blocking groups of DNA generated during both spore dormancy and germination.
Identification and Characterization of a New Enoyl Coenzyme A Hydratase Involved in Biosynthesis of Medium-Chain-Length Polyhydroxyalkanoates in Recombinant
Escherichia coli
Authors: Park Si Jae , Lee Sang Yup
Published: August 2003

The biosynthetic pathway of medium-chain-length (MCL) polyhydroxyalkanoates (PHAs) from fatty acids has been established in
fadB
mutant
Escherichia coli
strain by expressing the MCL-PHA synthase gene. However, the enzymes that are responsible for the generation of (
R
)-3-hydroxyacyl coenzyme A (R3HA-CoAs), the substrates for PHA synthase, have not been thoroughly elucidated.
Escherichia coli
MaoC, which is homologous to
Pseudomonas aeruginosa
(
R
)-specific enoyl-CoA hydratase (PhaJ1), was identified and found to be important for PHA biosynthesis in a
fadB
mutant
E. coli
strain. When the MCL-PHA synthase gene was introduced, the
fadB maoC
double-mutant
E. coli
WB108, which is a derivative of
E. coli
W3110, accumulated 43% less amount of MCL-PHA from fatty acid compared with the
fadB
mutant
E. coli
WB101. The PHA biosynthetic capacity could be restored by plasmid-based expression of the

maoC
Ec

gene in
E. coli
WB108. Also,
E. coli
W3110 possessing fully functional β-oxidation pathway could produce MCL-PHA from fatty acid by the coexpression of the

maoC
Ec

gene and the MCL-PHA synthase gene. For the enzymatic analysis, MaoC fused with His
6
-Tag at its C-terminal was expressed in
E. coli
and purified. Enzymatic analysis of tagged MaoC showed that MaoC has enoyl-CoA hydratase activity toward crotonyl-CoA. These results suggest that MaoC is a new enoyl-CoA hydratase involved in supplying (
R
)-3-hydroxyacyl-CoA from the β-oxidation pathway to PHA biosynthetic pathway in the
fadB
mutant
E. coli
strain.
Extracellular Polysaccharides Associated with Thin Aggregative Fimbriae ofSalmonella entericaSerovar Enteritidis
Authors: White A. P. , Gibson D. L. , Collinson S. K. , Banser P. A. , Kay W. W.
Published: August 2003

Lipopolysaccharide (LPS) O polysaccharide was identified as the principle factor impeding intercellular formation of intact thin aggregative fimbriae (Tafi) inSalmonella entericaserovar Enteritidis. The extracellular nucleation-precipitation assembly pathway for these organelles was investigated by quantifying fimbrial formation between ΔagfA(AgfA recipient) and ΔagfB(AgfA donor) cells harboring mutations in LPS (galE::Tn10) and/or cellulose (ΔbcsA) synthesis. Intercellular complementation could be detected between ΔagfAand ΔagfBstrains only when both possessed thegalEmutation. LPS O polysaccharide appears to be an impenetrable barrier to AgfA assembly between cells but not within individual cells. The presence of cellulose did not restrict Tafi formation between cells. Transmission electron microscopy ofw+S. entericaserovar Enteritidis 3b cells revealed diffuse Tafi networks without discernible fine structure. In the absence of cellulose, however, individual Tafi fibers were clearly visible, appeared to be occasionally branched, and showed the generally distinctive appearance described forEscherichia coliK-12 curli. A third extracellular matrix component closely associated with cellulose and Tafi was detected on Western blots by using immune serum raised to whole, purified Tafi aggregates. Cellulose was required to tightly link this material to cells. Antigenically similar material was also detected inS. entericaserovar Typhimurium and one diarrheagenicE. coliisolate. Preliminary analysis indicated that this material represented an anionic, extracellular polysaccharide that was distinct from colanic acid. Therefore, Tafi in their native state appear to exist as a complex with cellulose and at least one other component.
Natural Transformation of
Campylobacter jejuni
Requires Components of a Type II Secretion System
Authors: Wiesner Rebecca S. , Hendrixson David R. , DiRita Victor J.
Published: August 2003

The human pathogen
Campylobacter jejuni
is one of more than 40 naturally competent bacterial species able to import macromolecular DNA from the environment and incorporate it into their genomes. However, in
C. jejuni
little is known about the genes involved in this process. We used random transposon mutagenesis to identify genes that are required for the transformation of this organism. We isolated mutants with insertions in 11 different genes; most of the mutants are affected in the DNA uptake stage of transformation, whereas two mutants are affected in steps subsequent to DNA uptake, such as recombination into the chromosome or in DNA transport across the inner membrane. Several of these genes encode proteins homologous to those involved in type II secretion systems, biogenesis of type IV pili, and competence for natural transformation in gram-positive and gram-negative species. Other genes identified in our screen encode proteins unique to
C. jejuni
or are homologous to proteins that have not been shown to play a role in the transformation in other bacteria.