bluuebunny/crossref_metadata_2025 huggingface dataset:
Contributions to discrete-time methods for room acoustic simulation
Authors: Escolano-Carrasco José
Published: February 2009

The sound field distribution in a room is the consequence of the acoustic properties of radiating sources and the geometry and absorbing characteristics of the surrounding boundaries. Despite there existing a consolidated acoustic wave theory, it is very difficult, nearly impossible, to find an analytical expression of the sound variable distribution in a real room, as a function of time and position. This scenario represents as an inhomogeneous boundary value problem, where the complexity of source properties and boundary conditions make that problem extremely hard to solve. The use of discrete-time methods arises as a suitable solution for solving those partial differential equations, particularized by some specific constrains. Together with the constant growth of computer power, those methods are increasing their suitability for room acoustic simulation. However, there exists an important lack of accuracy in the definition of some of these conditions so far: Current frequency-dependent boundary conditions do not comply with any physical model, and directive sources in discrete-time methods have been hardly treated. This thesis discusses the current state-of-the-art boundary conditions and source modeling in discrete-time methods for room acoustic simulation, and it contributes some algorithms to enhance them.
Consonant-vowel and diphthong transitions in the speech of persons with dysarthria and of healthy controls.
Authors: Kuo Christina , Weismer Gary
Published: February 2009

Formant transitions have been of interest to researchers for their contribution to speech perception, sound identification, and inferences to articulatory behavior. In previous work diphthong transitions have been shown to be sensitive to speech motor control deficits found in dysarthria; comparable data regarding transitions between obstruents (i.e., consonants) and vowels are not available. Here consonant-vowel (CV) and diphthong transitions in speakers with dysarthria and in healthy controls are examined to understand whether the effects for diphthong transitions observed in speakers with dysarthria—reduced extent and slope and occasionally lengthened duration—are present for CV transitions. The first part of this study extends work by Weismer et al. [J. Acoust. Soc. Am. 121, 3135 (2007)], who reported a tendency for shallower CV transitions in speakers with dysarthria when compared to healthy controls, but only for syllables where C=dorsal. Given the limited number of dorsal CV transitions in the previous work, more utterances are included to better balance the sample sizes for different places of articulation (i.e., bilabial, alveolar, and dorsal) and also between the two speaker groups. Distributional analyses for several transition measures will be presented for 8 speakers with dysarthria and 18 healthy controls. [Work supported by NIDCD R01 DC003723.]
Adaptive Radiation: Contrasting Theory with Data
Authors: Gavrilets Sergey , Losos Jonathan B.
Published: February 2009

Biologists have long been fascinated by the exceptionally high diversity displayed by some evolutionary groups. Adaptive radiation in such clades is not only spectacular, but is also an extremely complex process influenced by a variety of ecological, genetic, and developmental factors and strongly dependent on historical contingencies. Using modeling approaches, we identify 10 general patterns concerning the temporal, spatial, and genetic/morphological properties of adaptive radiation. Some of these are strongly supported by empirical work, whereas for others, empirical support is more tentative. In almost all cases, more data are needed. Future progress in our understanding of adaptive radiation will be most successful if theoretical and empirical approaches are integrated, as has happened in other areas of evolutionary biology.
Is Genetic Evolution Predictable?
Authors: Stern David L. , Orgogozo Virginie
Published: February 2009

Ever since the integration of Mendelian genetics into evolutionary biology in the early 20th century, evolutionary geneticists have for the most part treated genes and mutations as generic entities. However, recent observations indicate that all genes are not equal in the eyes of evolution. Evolutionarily relevant mutations tend to accumulate in hotspot genes and at specific positions within genes. Genetic evolution is constrained by gene function, the structure of genetic networks, and population biology. The genetic basis of evolution may be predictable to some extent, and further understanding of this predictability requires incorporation of the specific functions and characteristics of genes into evolutionary theory.
The Bacterial Species Challenge: Making Sense of Genetic and Ecological Diversity
Authors: Fraser Christophe , Alm Eric J. , Polz Martin F. , Spratt Brian G. , Hanage William P.
Published: February 2009

The Bacteria and Archaea are the most genetically diverse superkingdoms of life, and techniques for exploring that diversity are only just becoming widespread. Taxonomists classify these organisms into species in much the same way as they classify eukaryotes, but differences in their biology—including horizontal gene transfer between distantly related taxa and variable rates of homologous recombination—mean that we still do not understand what a bacterial species is. This is not merely a semantic question; evolutionary theory should be able to explain why species exist at all levels of the tree of life, and we need to be able to define species for practical applications in industry, agriculture, and medicine. Recent studies have emphasized the need to combine genetic diversity and distinct ecology in an attempt to define species in a coherent and convincing fashion. The resulting data may help to discriminate among the many theories of prokaryotic species that have been produced to date.
Anomalously Metal-Rich Fluids Form Hydrothermal Ore Deposits
Authors: Wilkinson Jamie J. , Stoffell Barry , Wilkinson Clara C. , Jeffries Teresa E. , Appold Martin S.
Published: February 2009

Hydrothermal ore deposits form when metals, often as sulfides, precipitate in abundance from aqueous solutions in Earth's crust. Much of our knowledge of the fluids involved comes from studies of fluid inclusions trapped in silicates or carbonates that are believed to represent aliquots of the same solutions that precipitated the ores. We used laser ablation inductively coupled plasma mass spectrometry to test this paradigm by analysis of fluid inclusions in sphalerite from two contrasting zinc-lead ore systems. Metal contents in these inclusions are up to two orders of magnitude greater than those in quartz-hosted inclusions and are much higher than previously thought, suggesting that ore formation is linked to influx of anomalously metal-rich fluids into systems dominated by barren fluids for much of their life.
Confined Crystallization of Polyethylene Oxide in Nanolayer Assemblies
Authors: Wang Haopeng , Keum Jong K. , Hiltner Anne , Baer Eric , Freeman Benny , Rozanski Artur , Galeski Andrzej
Published: February 2009

The design and fabrication of ultrathin polymer layers are of increasing importance because of the rapid development of nanoscience and nanotechnology. Confined, two-dimensional crystallization of polymers presents challenges and opportunities due to the long-chain, covalently bonded nature of the macromolecule. Using an innovative layer-multiplying coextrusion process to obtain assemblies with thousands of polymer nanolayers, we discovered a morphology that emerges as confined polyethylene oxide (PEO) layers are made progressively thinner. When the thickness is confined to 20 nanometers, the PEO crystallizes as single, high-aspect-ratio lamellae that resemble single crystals. Unexpectedly, the crystallization habit imparts two orders of magnitude reduction in the gas permeability.
The Sea-Level Fingerprint of West Antarctic Collapse
Authors: Mitrovica Jerry X. , Gomez Natalya , Clark Peter U.
Published: February 2009

Recent projections of sea-level rise after a future collapse of theWest Antarctic Ice Sheet (for example, the Fourth Intergovernmental Panel on Climate Change Assessment Report) assume that meltwater will spread uniformly (that is, eustatically) across the oceans once marine-based sectors of the West Antarctic are filled. A largely neglected 1977 study predicted that peak values would be 20% higher than the eustatic in the North Pacific and 5 to 10% higher along the U.S. coastline. We show, with use of a state-of-the-art theory, that the sea-level rise in excess of the eustatic value will be two to three times higher than previously predicted for U.S. coastal sites.
A Great-Appendage Arthropod with a Radial Mouth from the Lower Devonian Hunsrück Slate, Germany
Authors: Kühl Gabriele , Briggs Derek E. G. , Rust Jes
Published: February 2009

Great-appendage arthropods, characterized by a highly modified anterior limb, were previously unknown after the Middle Cambrian. One fossil from the Lower Devonian Hunsrück Slate, Germany, extends the stratigraphic range of these arthropods by ∼100 million years.
Schinderhannes bartelsi
shows an unusual combination of anomalocaridid and euarthropod characters, including a highly specialized swimming appendage. A cladistic analysis indicates that the new taxon is basal to crown-group euarthropods and that the great-appendage arthropods are paraphyletic. This new fossil shows that features of the anomalocaridids, including the multisegmented raptorial appendage and circular plated mouth, persisted long after the initial radiation of the euarthropods.
Stability Predicts Genetic Diversity in the Brazilian Atlantic Forest Hotspot
Authors: Carnaval Ana Carolina , Hickerson Michael J. , Haddad Célio F. B. , Rodrigues Miguel T. , Moritz Craig
Published: February 2009

Biodiversity hotspots, representing regions with high species endemism and conservation threat, have been mapped globally. Yet, biodiversity distribution data from within hotspots are too sparse for effective conservation in the face of rapid environmental change. Using frogs as indicators, ecological niche models under paleoclimates, and simultaneous Bayesian analyses of multispecies molecular data, we compare alternative hypotheses of assemblage-scale response to late Quaternary climate change. This reveals a hotspot within the Brazilian Atlantic forest hotspot. We show that the southern Atlantic forest was climatically unstable relative to the central region, which served as a large climatic refugium for neotropical species in the late Pleistocene. This sets new priorities for conservation in Brazil and establishes a validated approach to biodiversity prediction in other understudied, species-rich regions.
Implications of a VLBI Distance to the Double Pulsar J0737-3039A/B
Authors: Deller A. T. , Bailes M. , Tingay S. J.
Published: February 2009

The double pulsar J0737-3039A/B is a unique system with which to test gravitational theories in the strong-field regime. However, the accuracy of such tests will be limited by knowledge of the distance and relative motion of the system. Here we present very long baseline interferometry (VLBI) observations which reveal that the distance to PSR J0737-3039A/B is

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parsecs, more than double previous estimates, and confirm its low transverse velocity (∼9 kilometers per second). Combined with a decade of pulsar timing, these results will allow tests of gravitational radiation emission theories at the 0.01% uncertainty level, putting stringent constraints on theories that predict dipolar gravitational radiation. They also allow insight into the system's formation and the source of its high-energy emission.
Nitrogen-Doped Carbon Nanotube Arrays with High Electrocatalytic Activity for Oxygen Reduction
Authors: Gong Kuanping , Du Feng , Xia Zhenhai , Durstock Michael , Dai Liming
Published: February 2009

The large-scale practical application of fuel cells will be difficult to realize if the expensive platinum-based electrocatalysts for oxygen reduction reactions (ORRs) cannot be replaced by other efficient, low-cost, and stable electrodes. Here, we report that vertically aligned nitrogen-containing carbon nanotubes (VA-NCNTs) can act as a metal-free electrode with a much better electrocatalytic activity, long-term operation stability, and tolerance to crossover effect than platinum for oxygen reduction in alkaline fuel cells. In air-saturated 0.1 molar potassium hydroxide, we observed a steady-state output potential of –80 millivolts and a current density of 4.1 milliamps per square centimeter at –0.22 volts, compared with –85 millivolts and 1.1 milliamps per square centimeter at –0.20 volts for a platinum-carbon electrode. The incorporation of electron-accepting nitrogen atoms in the conjugated nanotube carbon plane appears to impart a relatively high positive charge density on adjacent carbon atoms. This effect, coupled with aligning the NCNTs, provides a four-electron pathway for the ORR on VA-NCNTs with a superb performance.
Structural Analysis of Biofilm Formation by Rapidly and Slowly Growing Nontuberculous Mycobacteria
Authors: Williams Margaret M. , Yakrus Mitchell A. , Arduino Matthew J. , Cooksey Robert C. , Crane Christina B. , Banerjee Shailen N. , Hilborn Elizabeth D. , Donlan Rodney M.
Published: February 2009

Mycobacterium avium
complex (MAC) and rapidly growing mycobacteria (RGM) such as
M. abscessus
,
M. mucogenicum
,
M. chelonae
, and
M. fortuitum
, implicated in health care-associated infections, are often isolated from potable water supplies as part of the microbial flora. To understand factors that influence growth in their environmental source, clinical RGM and slowly growing MAC isolates were grown as biofilm in a laboratory batch system. High and low nutrient levels were compared, as well as stainless steel and polycarbonate surfaces. Biofilm growth was measured after 72 h of incubation by enumeration of bacteria from disrupted biofilms and by direct quantitative image analysis of biofilm microcolony structure. RGM biofilm development was influenced more by nutrient level than by substrate material, though both affected biofilm growth for most of the isolates tested. Microcolony structure revealed that RGM develop several different biofilm structures under high-nutrient growth conditions, including pillars of various shapes (
M. abscessus
and
M. fortuitum
) and extensive cording (
M. abscessus
and
M. chelonae
). Although it is a slowly growing species in the laboratory, a clinical isolate of
M. avium
developed more culturable biofilm in potable water in 72 h than any of the 10 RGM examined. This indicates that
M. avium
is better adapted for growth in potable water systems than in laboratory incubation conditions and suggests some advantage that MAC has over RGM in low-nutrient environments.
Unexpected Genetic Diversity among and within Populations of the Toxic Dinoflagellate
Alexandrium catenella
as Revealed by Nuclear Microsatellite Markers
Authors: Masseret Estelle , Grzebyk Daniel , Nagai Satoshi , Genovesi Benjamin , Lasserre Bernard , Laabir Mohamed , Collos Yves , Vaquer André , Berrebi Patrick
Published: February 2009

Since 1998, blooms of
Alexandrium catenella
associated with paralytic shellfish poisoning have been repeatedly reported for Thau Lagoon (French Mediterranean coast). Based on data obtained for rRNA gene markers, it has been suggested that the strains involved could be closely related to the Japanese temperate Asian ribotype of the temperate Asian clade. In order to gain more insight into the origin of these organisms, we carried out a genetic analysis of 61 Mediterranean and 23 Japanese strains using both ribosomal and microsatellite markers. Whereas the phylogeny based on ribosomal markers tended to confirm the previous findings, the analysis of microsatellite sequences revealed an unexpected distinction between the French and Japanese populations. This analysis also highlighted great intraspecific diversity that was not detected with the classical rRNA gene markers. The Japanese strains are divided into two differentiated
A. catenella
lineages: the Sea of Japan lineage and the east coast lineage, which includes populations from the Inland Sea and the Pacific Ocean.
A. catenella
strains isolated from Thau Lagoon belong to another lineage. These findings indicate that microsatellite markers are probably better suited to investigations of the population genetics of this species that is distributed worldwide. Finally, application of the population genetics concepts available for macroorganisms could support new paradigms for speciation and migration in phytoplankton assemblages.
Prokaryotic Genomes and Diversity in Surface Ocean Waters: Interrogating the Global Ocean Sampling Metagenome
Authors: Biers Erin J. , Sun Shulei , Howard Erinn C.
Published: February 2009

The
Sorcerer II
Global Ocean Sampling (GOS) sequencing effort has vastly expanded the landscape of metagenomics, providing an opportunity to study the genetic potential of surface ocean water bacterioplankton on a global scale. Here we describe the habitat-based microbial diversity, both taxon evenness and taxon richness, for each GOS site and estimate genome characteristics of a typical free-living, surface ocean water bacterium. While
Alphaproteobacteria
and particularly SAR11 dominate the 0.1- to 0.8-μm size fraction of surface ocean water bacteria (43% and 31%, respectively), the proportions of other taxa varied with ocean habitat type. Within each habitat type, lower-bound estimates of phylum richness ranged between 18 and 59 operational taxonomic units (OTUs). However, OTU richness was relatively low in the hypersaline lagoon community at every taxonomic level, and open-ocean communities had much more microdiversity than any other habitat. Based on the abundance of single-copy eubacterial genes from the same data set, we estimate that the genome of an average free-living surface ocean water bacterium (sized between 0.1 and 0.8 μm) contains ∼1,019 genes and 1.8 copies of the 16S rRNA gene, suggesting that these bacteria have relatively streamlined genomes in comparison to those of cultured bacteria and bacteria from other habitats (e.g., soil or acid mine drainage).
Dynamic Model of Heat Inactivation Kinetics for Bacterial Adaptation
Authors: Corradini Maria G. , Peleg Micha
Published: February 2009

The Weibullian-log logistic (WeLL) inactivation model was modified to account for heat adaptation by introducing a logistic adaptation factor, which rendered its “rate parameter” a function of both temperature and heating rate. The resulting model is consistent with the observation that adaptation is primarily noticeable in slow heat processes in which the cells are exposed to sublethal temperatures for a sufficiently long time. Dynamic survival patterns generated with the proposed model were in general agreement with those of
Escherichia coli
and
Listeria monocytogenes
as reported in the literature. Although the modified model's rate equation has a cumbersome appearance, especially for thermal processes having a variable heating rate, it can be solved numerically with commercial mathematical software. The dynamic model has five survival/adaptation parameters whose determination will require a large experimental database. However, with assumed or estimated parameter values, the model can simulate survival patterns of adapting pathogens in cooked foods that can be used in risk assessment and the establishment of safe preparation conditions.
Plasticity and Evolution of Aeruginosin Biosynthesis in Cyanobacteria
Authors: Ishida Keishi , Welker Martin , Christiansen Guntram , Cadel-Six Sabrina , Bouchier Christiane , Dittmann Elke , Hertweck Christian , Tandeau de Marsac Nicole
Published: February 2009

Aeruginosins are bioactive oligopeptides that are produced in high structural diversity by strains of the bloom-forming cyanobacterial genera
Microcystis
and
Planktothrix
. A hallmark of aeruginosins is the unusual Choi moiety central to the tetrapeptides, while other positions are occupied by variable moieties in individual congeners. Here we report on three aeruginosin synthetase gene clusters (
aer
) of
Microcystis aeruginosa
(strains PCC 7806, NIES-98, and NIES-843). The analysis and comparison the
aer
gene clusters provide the first insight into the molecular basis of biosynthetic and structural plasticity in aeruginosin pathways. Major parts of the
aer
gene clusters are highly similar in all strains, particularly the genes coding for the first three nonribosomal peptide synthetase (NRPS) modules except for the region coding for the second adenylation domain. However, the gene clusters differ largely in genes coding for tailoring enzymes such as halogenases and sulfotransferases, reflecting structural peculiarities in aeruginosin congeners produced by the individual strains. Significant deviations were further observed in the C-terminal NRPS modules, suggesting two distinct release mechanisms. The architecture of the gene clusters is in agreement with the particular aeruginosin variants that are produced by individual strains, the structures of two of which (aeruginosins 686 A and 686 B) were elucidated. The
aer
gene clusters of
Microcystis
and
Planktothrix
are proposed to originate from a common ancestor and to have evolved to their present-day diversity largely through horizontal gene transfer and recombination events.
Reciprocal Expression of Integration Host Factor and HU in the Developmental Cycle and Infectivity of
Legionella pneumophila
Authors: Morash Michael G. , Brassinga Ann Karen C. , Warthan Michelle , Gourabathini Poornima , Garduño Rafael A. , Goodman Steven D. , Hoffman Paul S.
Published: February 2009

Legionella pneumophila
is an intracellular parasite of protozoa that differentiates late in infection into metabolically dormant cysts that are highly infectious. Regulation of this process is poorly understood. Here we report that the small DNA binding regulatory proteins integration host factor (IHF) and HU are reciprocally expressed over the developmental cycle, with HU expressed during exponential phase and IHF expressed postexponentially. To assess the role of these regulatory proteins in development, chromosomal deletions were constructed. Single (
ihfA
or
ihfB
) and double deletion (Δ
ihf
) IHF mutants failed to grow in
Acanthamoeba castellanii
unless complemented in
trans
when expressed temporally from the
ihfA
promoter but not under P
tac
(isopropyl-β-
d
-thiogalactopyranoside). In contrast, IHF mutants were infectious for HeLa cells, though electron microscopic examination revealed defects in late-stage cyst morphogenesis (thickened cell wall, intracytoplasmic membranes, and inclusions of poly-β-hydroxybutyrate), and were depressed for the developmental marker MagA. Green fluorescent protein promoter fusion assays indicated that IHF and the stationary-phase sigma factor RpoS were required for full postexponential expression of
magA
. Finally, defects in cyst morphogenesis noted for Δ
ihf
mutants in HeLa cells correlated with a loss of both detergent resistance and hyperinfectivity compared with results for wild-type cysts. These studies establish IHF and HU as markers of developmental stages and show that IHF function is required for both differentiation and full virulence of
L. pneumophila
in natural amoebic hosts.
Mesophilic and Psychrotrophic Bacteria from Meat and Their Spoilage Potential In Vitro and in Beef
Authors: Ercolini Danilo , Russo Federica , Nasi Antonella , Ferranti Pasquale , Villani Francesco
Published: February 2009

Mesophilic and psychrotrophic populations from refrigerated meat were identified in this study, and the spoilage potential of microbial isolates in packaged beef was evaluated by analyzing the release of volatile organic compounds (VOC) by gas chromatography-mass spectrometry (GC/MS). Fifty mesophilic and twenty-nine psychrotrophic isolates were analyzed by random amplified polymorphic DNA-PCR, and representative strains were identified by 16S rRNA gene sequencing.
Carnobacterium maltaromaticum
and
C. divergens
were the species most frequently found in both mesophilic and psychrotrophic populations.
Acinetobacter baumannii
,
Buttiauxella
spp. and
Serratia
spp. were identified among the mesophilic isolates, while
Pseudomonas
spp. were commonly identified among the psychrotrophs. The isolates were further characterized for their growth at different temperatures and their proteolytic activity in vitro on meat proteins extracts at 7°C. Selected proteolytic strains of
Serratia proteamaculans
,
Pseudomonas fragi
, and
C. maltaromaticum
were used to examine their spoilage potential in situ. Single strains of these species and mixtures of these strains were used to contaminate beef chops that were packed and stored at 7°C. At time intervals up to 1 month, viable counts were determined, and VOC were identified by GC/MS. Generally, the VOC concentrations went to increase during the storage of the contaminated meats, and the profiles of the analyzed meat changed dramatically depending on the contaminating microbial species. About 100 volatiles were identified in the different contaminated samples. Among the detected volatiles, some specific molecules were identified only when the meat was contaminated by a specific microbial species. Compounds such as 2-ethyl-1-hexanol, 2-buten-1-ol, 2-hexyl-1-octanol, 2-nonanone, and 2-ethylhexanal were detectable only for
C. maltaromaticum
, which also produced the highest number of aldehydes, lactones, and sulfur compounds. The highest number of alcohols and ketons were detected in the headspace of meat samples contaminated by
P. fragi
, whereas the highest concentrations of some alcohols, such as 1-octen-3-ol, and some esters, such as isoamyl acetate, were produced by
S. proteamaculans
. In conclusion, different microbial species can contribute to meat spoilage with release of different volatile compounds that concur to the overall quality decrease of spoiling meat.
Role of
Acinetobacter baylyi
Crc in Catabolite Repression of Enzymes for Aromatic Compound Catabolism
Authors: Zimmermann Tina , Sorg Tobias , Siehler Simone Yasmin , Gerischer Ulrike
Published: February 2009

Here, we describe for the first time the Crc (
c
atabolite
r
epression
c
ontrol) protein from the soil bacterium
Acinetobacter baylyi
. Expression of
A. baylyi crc
varied according to the growth conditions. A strain with a disrupted
crc
gene showed the same growth as the wild type on a number of carbon sources. Carbon catabolite repression by acetate and succinate of protocatechuate 3,4-dioxygenase, the key enzyme of protocatechuate breakdown, was strongly reduced in the
crc
strain, whereas in the wild-type strain it underwent strong catabolite repression. This strong effect was not based on transcriptional regulation because the transcription pattern of the
pca-qui
operon (encoding protocatechuate 3,4-dioxygenase) did not reflect the derepression in the absence of Crc.
pca-qui
transcript abundance was slightly increased in the
crc
strain. Lack of Crc dramatically increased the mRNA stability of the
pca-qui
transcript (up to 14-fold), whereas two other transcripts (
pobA
and
catA
) remained unaffected.
p
-Hydroxybenzoate hydroxylase activity, encoded by
pobA
, was not significantly different in the absence of Crc, as protocatechuate 3,4-dioxygenase was. It is proposed that
A. baylyi
Crc is involved in the determination of the transcript stability of the
pca-qui
operon and thereby effects catabolite repression.
Role of RsbU in Controlling SigB Activity inStaphylococcus aureusfollowing Alkaline Stress
Authors: Pané-Farré Jan , Jonas Beate , Hardwick Steven W. , Gronau Katrin , Lewis Richard J. , Hecker Michael , Engelmann Susanne
Published: February 2009

SigB is an alternative sigma factor that controls a large regulon inStaphylococcus aureus. Activation of SigB requires RsbU, a protein phosphatase 2C (PP2C)-type phosphatase. In a closely related organism,Bacillus subtilis, RsbU activity is stimulated upon interaction with RsbT, a kinase, which following an activating stimulus switches from a 25S high-molecular-weight complex, the stressosome, to the N-terminal domain of RsbU. Active RsbU dephosporylates RsbV and thereby triggers the release of SigB from its inhibitory complex with RsbW. While RsbU, RsbV, RsbW, and SigB are conserved inS. aureus, proteins similar to RsbT and the components of the stressosome are not, raising the question of how RsbU activity and hence SigB activity are controlled inS. aureus. We found that in contrast to the case inB. subtilis, the induced expression of RsbU was sufficient to stimulate SigB-dependent transcription inS. aureus. However, activation of SigB-dependent transcription following alkaline stress did not lead to a clear accumulation of SigB and its regulators RsbV and RsbW or to a change in the RsbV/RsbV-P ratio inS. aureus. When expressed inB. subtilis, theS. aureusRsbU displayed a high activity even in the absence of an inducing stimulus. This high activity could be transferred to the PP2C domain of theB. subtilisRsbU protein by a fusion to the N-terminal domain of theS. aureusRsbU. Collectively, the data suggest that the activity of theS. aureusRsbU and hence SigB may be subjected to different regulation in comparison to that inB. subtilis.
Activation of the
Campylobacter jejuni
FlgSR Two-Component System Is Linked to the Flagellar Export Apparatus
Authors: Joslin Stephanie N. , Hendrixson David R.
Published: February 2009

Activation of σ
54
-dependent gene expression essential for formation of flagella in
Campylobacter jejuni
requires the components of the inner membrane-localized flagellar export apparatus and the FlgSR two-component regulatory system. In this study, we characterized the FlgS sensor kinase and how activation of the protein is linked to the flagellar export apparatus. We found that FlgS is localized to the
C. jejuni
cytoplasm and that His141 of FlgS is essential for autophosphorylation, phosphorelay to the cognate FlgR response regulator, motility, and expression of σ
54
-dependent flagellar genes. Mutants with incomplete flagellar export apparatuses produced wild-type levels of FlgS and FlgR, but they were defective for signaling through the FlgSR system. By using genetic approaches, we found that FlgSR activity is linked to and downstream of the flagellar export apparatus in a regulatory cascade that terminates in expression of σ
54
-dependent flagellar genes. By analyzing defined
flhB
and
fliI
mutants of
C. jejuni
that form flagellar export apparatuses that are secretion incompetent, we determined that formation of the apparatus is required to contribute to the signal sensed by FlgS to terminate in activation of expression of σ
54
-dependent flagellar genes. Considering that the flagellar export apparatuses of
Escherichia coli
and
Salmonella
species influence σ
28
-dependent flagellar gene expression, our work expands the signaling activity of the apparatuses to include σ
54
-dependent pathways of
C. jejuni
and possibly other motile bacteria. This study indicates that these apparatuses have broader functions beyond flagellar protein secretion, including activation of essential two-component regulatory systems required for expression of σ
54
-dependent flagellar genes.
Sporulation and Enterotoxin (CPE) Synthesis Are Controlled by the Sporulation-Specific Sigma Factors SigE and SigK inClostridium perfringens
Authors: Harry Kathryn H. , Zhou Ruanbao , Kroos Lee , Melville Stephen B.
Published: February 2009

Clostridium perfringensis the third most frequent cause of bacterial food poisoning annually in the United States. IngestedC. perfringensvegetative cells sporulate in the intestinal tract and produce an enterotoxin (CPE) that is responsible for the symptoms of acute food poisoning. Studies ofBacillus subtilishave shown that gene expression during sporulation is compartmentalized, with different genes expressed in the mother cell and the forespore. The cell-specific RNA polymerase sigma factors σF, σE, σG, and σKcoordinate much of the developmental process. TheC. perfringens cpegene, encoding CPE, is transcribed from three promoters, where P1 was proposed to be σKdependent, while P2 and P3 were proposed to be σEdependent based on consensus promoter recognition sequences. In this study, mutations were introduced into thesigEandsigKgenes ofC. perfringens. With thesigEandsigKmutants,gusAfusion assays indicated that there was no expression ofcpein either mutant. Results fromgusAfusion assays and immunoblotting experiments indicate that σE-associated RNA polymerase and σK-associated RNA polymerase coregulate each other's expression. Transcription and translation of thespoIIIDgene inC. perfringenswere not affected by mutations insigEandsigK, which differs fromB. subtilis, in whichspoIIIDtranscription requires σE-associated RNA polymerase. The results presented here show that the regulation of developmental events in the mother cell compartment ofC. perfringensis not the same as that inB. subtilisandClostridium acetobutylicum.
A sports setting matrix for understanding the implementation context for community sport
Authors: Finch C F , Donaldson A
Published: February 2009

There has been increasing recognition of the need for effectiveness research within the real-world intervention context of community sport. This is important because, even if interventions have been shown to be efficacious in controlled trials, if they are not also widely adopted and sustained, then it is unlikely that they will have a public health impact. There is very little information about how to best conduct such studies, but application of health promotion frameworks, such as the RE-AIM framework, to evaluate the public health impact of interventions could potentially help to understand the implementation context. Care needs to be taken when directly applying the RE-AIM framework, however, because the definitions for each of its dimensions will depend on the level/s the intervention is targeted at. This paper provides a novel extension to the RE-AIM framework (the RE-AIM Sports Setting Matrix (RE-AIM SSM)), which accounts for the fact that many sports injury interventions need to be targeted at multiple levels of sports delivery. Accordingly, the RE-AIM components also need to be measured across all tiers of possible influence on the rate of uptake and effectiveness. Specific examples are given for coachdelivered exercise training interventions. The RE-AIM SSM is specific to the community sports setting implementation context and could be used to guide the delivery of future sports safety, and other health promotion, interventions in this area.
The unbearable lightness of diagnostic testing: time to contain inappropriate test ordering
Authors: Schattner A
Published: February 2009

The numbers of tests, imaging and diagnostic procedures ordered by doctors in primary care and hospital settings alike are rising. According to established guidelines, many of these tests may be unnecessary. Various non-clinical factors often underlie test ordering, and multiple prevalent problems associated with tests and their interpretation are often under-appreciated. These include the significant potential for cascades of further testing and patient harm. Multifaceted strategies may improve test-ordering behaviour. Brief reconsideration of several points by the clinician before ordering the test is advisable.
Lipid-Dependent Membrane Protein Topogenesis
Authors: Dowhan William , Bogdanov Mikhail
Published: February 2009

The topology of polytopic membrane proteins is determined by topogenic sequences in the protein, protein-translocon interactions, and interactions during folding within the protein and between the protein and the lipid environment. Orientation of transmembrane domains is dependent on membrane phospholipid composition during initial assembly as well as on changes in lipid composition postassembly. The membrane translocation potential of negative amino acids working in opposition to the positive-inside rule is largely dampened by the normal presence of phosphatidylethanolamine, thus explaining the dominance of positive residues as retention signals. Phosphatidylethanolamine provides the appropriate charge density that permits the membrane surface to maintain a charge balance between membrane translocation and retention signals and also allows the presence of negative residues in the cytoplasmic face of proteins for other purposes.
cAMP-GEF cytoprotection by Src tyrosine kinase activation of phosphoinositide-3-kinase p110 β/α in rat hepatocytes
Authors: Gates Anna , Hohenester Simon , Anwer M. Sawkat , Webster Cynthia R. L.
Published: February 2009

Cyclic AMP protects against hepatocyte apoptosis by a protein kinase A-independent cAMP-GEF/phosphoinositide-3-kinase (PI3K)/Akt signaling pathway. However, the signaling pathway coupling cAMP-GEF with PI3K is unknown. The aim of this study was to investigate the role of Src tyrosine kinases (Src-TYK) and PI3K-p110 isoforms in this pathway. Studies were done in rat hepatocytes using the hydrophobic bile acid glycochenodeoxycholic acid (GCDC) to induce apoptosis. cAMP-binding guanine nucleotide exchange factors (cAMP-GEFs) were selectively activated by using 4-(4-chloro-phenylthio)-2′- O-methyladenosine-3′-5′-cyclic monophosphate (CPT-2-Me-cAMP), which sequentially phosphorylated Src-TYK (within 1 min) followed by Akt (within 5 min). The Src inhibitors PP2 and SU6656 inhibited basal and CPT-2-Me-cAMP-mediated Src and Akt phosphorylation. These inhibitors had no effect on CPT-2-Me-cAMP-mediated activation of Rap GTPases. CPT-2-Me-cAMP induced transient Src dependent autophosphorylation of the epidermal growth factor receptor (EGFR). Inhibition of the EGFR with AG 1478 partially inhibited the ability of CPT-2-Me to phosphorylate Akt. Whereas PP2 completely abolished the protective effect of CPT-2-Me-cAMP in GCDC induced apoptosis, AG 1478 partially inhibited the cytoprotective effect. CPT-2-Me-cAMP treatment resulted in Src-dependent activation of the p110 β and α subunits of PI3K, but only the latter was sensitive to inhibition with AG 1478. In conclusion, activation of cAMP-GEFs results in phosphorylation of Src-TYK and Akt and activation of the p110 β/α subunits of PI3K. Maximal cAMP-GEF-mediated Akt phosphorylation as well as protection from bile acid-induced apoptosis requires activation of Src-TYK and the EGFR. These studies support the existence of two pathways: cAMP-GEF/Rap/Src/PI3Kβ/Akt and cAMP-GEF/Rap/Src/EGFR/PI3Kα/Akt, both of which are necessary for maximal cytoprotective effect of cAMP-GEFs in hepatocytes.
Inhibition of p38 MAPK reverses hypoxia-induced pulmonary artery endothelial dysfunction
Authors: Weerackody Roshan P. , Welsh David J. , Wadsworth Roger M. , Peacock Andrew J.
Published: February 2009

Hypoxia-induced endothelial dysfunction plays a crucial role in the pathogenesis of hypoxic pulmonary hypertension. p38 MAPK expression is increased in the pulmonary artery following hypoxic exposure. Recent evidence suggests that increased p38 MAPK activity is associated with endothelial dysfunction. However, the role of p38 MAPK activation in pulmonary artery endothelial dysfunction is not known. Sprague-Dawley rats were exposed to 2 wk hypobaric hypoxia, which resulted in the development of pulmonary hypertension and vascular remodeling. Endothelium-dependent relaxation of intrapulmonary vessels from hypoxic animals was impaired due to a reduced nitric oxide (NO) generation. This was despite increased endothelial NO synthase immunostaining and protein expression. Hypoxia exposure increased superoxide generation and p38 MAPK expression. The inhibition of p38 MAPK restored endothelium-dependent relaxation, increased bioavailable NO, and reduced superoxide production. In conclusion, the pharmacological inhibition of p38 MAPK was effective in increasing NO generation, reducing superoxide burden, and restoring hypoxia-induced endothelial dysfunction in rats with hypoxia-induced pulmonary hypertension. p38 MAPK may be a novel target for the treatment of pulmonary hypertension.
Slowing of cardiomyocyte Ca2+ release and contraction during heart failure progression in postinfarction mice
Authors: Mørk Halvor K. , Sjaastad Ivar , Sejersted Ole M. , Louch William E.
Published: February 2009

Deterioration of cardiac contractility during congestive heart failure (CHF) is believed to involve decreased function of individual cardiomyocytes and may include reductions in contraction magnitude and/or kinetics. We examined the progression of in vivo and in vitro alterations in contractile function in CHF mice and investigated underlying alterations in Ca2+ homeostasis. Following induction of myocardial infarction (MI), mice with CHF were examined at early (1 wk post-MI) and chronic (10 wk post-MI) stages of disease development. Sham-operated mice served as controls. Global and local left ventricle function were assessed by echocardiography in sedated animals (∼2% isoflurane). Excitation-contraction coupling was examined in cardiomyocytes isolated from the viable septum. CHF progression between 1 and 10 wk post-MI resulted in increased mortality, development of hypertrophy, and deterioration of global left ventricular function. Local function in the noninfarcted myocardium also declined, as posterior wall shortening velocity was reduced in chronic CHF (1.2 ± 0.1 vs. 1.9 ± 0.2 cm/s in sham). Parallel alterations occurred in isolated cardiomyocytes since contraction and Ca2+ transient time to peak values were prolonged in chronic CHF (115 ± 6 and 158 ± 11% sham values, respectively). Surprisingly, contraction and Ca2+ transient magnitudes in CHF were larger than sham values at both time points, resulting from increased sarcoplasmic reticulum Ca2+ content and greater Ca2+ influx via L-type channels. We conclude that, in mice with CHF following myocardial infarction, declining myocardial function involves slowing of cardiomyocyte contraction without reduction in contraction magnitude. Corresponding alterations in Ca2+ transients suggest that slowing of Ca2+ release is a critical mediator of CHF progression.
Suppression of prostaglandin E2-induced MUC5AC overproduction by RGS4 in the airway
Authors: Song Kyoung Seob , Choi Yeon Ho , Kim Jong-Mu , Lee Hyunjae , Lee Tae-Jin , Yoon Joo-Heon
Published: February 2009

The mechanism by which E-prostanoid (EP) receptor is critically involved in PGE2-induced mucin 5AC ( MUC5AC) gene expression in the airway has been unclear. Furthermore, there have been little reports regarding the negative regulatory mechanism and/or proteins that affect PGE2-induced MUC5AC overproduction. In the present study, we found that PGE2 induced MUC5AC gene expression in a dose-dependent manner (EC50: 73.31 ± 3.13 nM) and that the EP2/4-specific agonist, misoprostol, increased MUC5AC mRNA level, whereas the EP1/3-specific agonist, sulprostone, had no effect. Interestingly, the cAMP concentration (685.1 ± 14.9 pM) of the EC50 value of EP4-mediated cAMP production was much higher than that of EP2 (462.33 ± 23.79 pM), suggesting that EP4 has higher sensitivity to PGE2 compared with EP2. Moreover, PGE2-induced Muc5ac overproduction was much increased in regulator of G protein signaling ( Rgs) 4 knockout (KO) mice compared with wild-type mice at both transcriptional and translational levels, and it was dramatically suppressed in Rgs4 KO mice that had been infected with lentivirus expressing RGS4 (lenti::RGS4) compared with lentivirus expressing enhanced green fluorescent protein (lenti::eGFP). Finally, we demonstrate that PGE2 can induce MUC5AC overproduction via the EP4 receptor and that RGS4 may have suppressive effects in controlling MUC5AC overexpression in the airway. These findings may provide a molecular paradigm for the development of novel drugs for respiratory diseases.
Modulation of epidermal growth factor receptor binding to human airway smooth muscle cells by glucocorticoids and β2-adrenergic receptor agonists
Authors: Kassel Karen M. , Schulte Nancy A. , Toews Myron L.
Published: February 2009

EGF receptors (EGFRs) are increased in airway smooth muscle in asthma, which may contribute to both their hyperproliferation and hypercontractility. Lysophosphatidic acid (LPA) is a candidate pathological agent in asthma and other airway diseases, and LPA upregulates EGFRs in human airway smooth muscle (HASM) cells. We tested whether therapeutic glucocorticoids and/or β2-adrenergic receptor (β2AR) agonists also alter EGFR binding in HASM cells. Exposure to glucocorticoids for 24 h induced a twofold increase in EGFR binding similar to that with LPA; fluticasone was markedly more potent than dexamethasone. The increase in EGFR binding by glucocorticoids required 24-h exposure, consistent with transcription-mediated effects. Although the increase in EGFR binding was blocked by the protein synthesis inhibitor cycloheximide for LPA, fluticasone, and dexamethasone, only LPA induced a significant increase in EGFR protein expression detected by immunoblotting. In contrast to the increased binding induced by the glucocorticoids, the β2AR agonists isoproterenol, albuterol, and salmeterol all induced a decrease in EGFR binding. β2AR agonist effects were multiphasic, with an initial decline at 2–4 h that reversed by 6 h and a second, somewhat greater decrease by 18–24 h. In cells pretreated with glucocorticoids, the decreases in EGFR binding by subsequent β2AR treatment were not statistically significant; glucocorticoid upregulation of EGFRs also prevented further increases by LPA. Similar increases by glucocorticoids and decreases by β2AR agonists were found in HFL-1 human lung fibroblasts. These complex and opposing effects of clinically relevant glucocorticoids and β2AR agonists on airway mesenchymal cell EGFRs likely contribute to their overall therapeutic profile in the diseased airway.
Cubilin and Megalin Co-Localize in the Neonatal Inner Ear
Authors: Tauris Jacob , Christensen Erik Ilsø , Nykjær Anders , Jacobsen Christian , Petersen Claus Munck , Ovesen Therese
Published: February 2009

Cubilin and megalin are multifunctional endocytotic receptors expressed in many absorptive epithelia. The receptors have separate functions but may act in concert in several tissues including the small intestine, the visceral yolk sac and the renal proximal tubule to perform significant physiological functions essential to homeostasis in the extracellular fluid compartments. Because of the importance of fluid homeostasis in the inner ear, we investigated the expression of cubilin and megalin in this structure. Furthermore, we examined the binding properties of 6 different aminoglycosides (AGs) to cubilin and compared the results to data obtained for megalin, which is a well-known receptor for AGs and other polybasic substances, including several ototoxic drugs. In the cochlea, immunohistochemical labelling for cubilin showed expression corresponding to the apical surface of the strial marginal cells, to epithelial cells at the spiral prominence and to epithelial cells of Reissner’s membrane facing the cochlear duct. In the vestibular apparatus, positive labelling was found in dark cells of the utricle and those flanking the crista ampullaris of the semicircular canals. Exactly the same tissue distribution was found for megalin. These findings support the prevailing view that cubilin and megalin constitute a dual-receptor complex facilitating the function of each other. The physiological role of this receptor complex in the inner ear remains unknown, although several established ligands for both cubilin and megalin are present in the inner ear fluids. Receptor-ligand interaction analysis showed that all 6 AGs bind to both receptors and with approximately the same affinity. The results demonstrate a novel role for cubilin as a drug receptor for AGs and possibly other ototoxic substances.