bluuebunny/crossref_metadata_2025 huggingface dataset:
Free Vibration of Helicoidal Beams of Arbitrary Shape and Variable Cross Section
Authors: Busool Wisam , Eisenberger Moshe
Published: July 2002

In this study, the dynamic stiffness method is employed for the free vibration analysis of helical springs. This work gives the exact solutions for the natural frequencies of helical beams having arbitrary shapes, such as conical, hyperboloidal, and barrel. Both the cross-section dimensions and the shape of the beam can vary along the axis of the curved member as polynomial expressions. The problem is described by six differential equations. These are second order equations with variable coefficients, with six unknown displacements, three translations, and three rotations at every point along the member. The proposed solution is based on a new finite-element method for deriving the exact dynamic stiffness matrix for the member, including the effects of the axial and the shear deformations and the rotational inertia effects for any desired precision. The natural frequencies are found as the frequencies that cause the determinant of the dynamic stiffness matrix to become zero. Then the mode shape for every natural frequency is found. Examples are given for beams and helical springs with different shape, which can vary along the axis of the member. It is shown that the present numerical results agree well with previously published numerical and experimental results.
Spectral Finite Element Analysis of Sandwich Beams With Passive Constrained Layer Damping1
Authors: Wang Gang , Wereley Norman M.
Published: July 2002

We present a spectral finite element model (SFEM) for sandwich beams with passive constrained layer damping (PCLD) treatments. The viscoelastic core has a complex modulus that varies with frequency. The SFEM is formulated in the frequency domain using dynamic shape functions based on the exact displacement solutions from progressive wave methods, where we implicitly account for the frequency dependent complex modulus of the viscoelastic core. The SFEM results of natural frequencies and frequency response functions are compared to those calculated using conventional finite element (CFEM), where the Golla-Hughes-McTavish method is used to account for the frequency dependent complex modulus of a viscoelastic core. Also experimental data are used to validate both analyses using frequency response functions measured for two cantilevered sandwich beams with PCLD treatments having 50% and 75% coverage of the beam length. SFEM shows improved computational efficiency and accuracy, because many more elements must be incorporated into the CFEM for comparable accuracy.
On the Absorption Coefficient of Porous Corrugated Surfaces
Authors: Simo´n Francisco , Rodrı´guez Rosa M. , Pfretzschner Jaime
Published: July 2002

A classical way of improving acoustical absorption performances of porous materials is the use of corrugated surfaces; this use can obtain lower cut-off frequencies and also improve the overall absorption over a wide frequency range. An analytical approximation is presented for the calculus of the absorption on this kind of surfaces, where the thickness gradient is represented as a series of steps. Reflection coefficient of every step is obtained and will contribute to the net reflection coefficient. Theoretical results will be presented and shown to agree with experimental data.
Investigation of surface oxides on aluminum alloys by valence band photoemission
Authors: Claycomb Gregory D. , Sherwood Peter M. A.
Published: July 2002

Core level and valence band x-ray photoelectron spectroscopy are used to study the chemical composition of the surface films on aluminum alloys. Certain alloying elements may preferentially migrate to the surface of an alloy, thereby altering the composition and consequently the chemistry of the surface. The behavior of a 6061 aluminum alloy is compared with that of pure aluminum. It is shown that the type of magnesium film formed at the alloy surface can be determined by comparing the valence band spectra of the aluminum alloy surface with that of known magnesium and aluminum compounds. The experimental valence band spectra of these compounds are supported by spectra generated from band structure calculations. The effect of boiling water on the surface film is discussed, with significant differences in surface chemistry being seen for the metal and the alloy.
Effects of cross field diffusion in a low pressure high density oxygen/silane plasma
Authors: Charles C.
Published: July 2002

A low pressure high density oxygen/silane radio frequency (13.56 MHz) plasma coupled in a helicon reactor used for silicon dioxide deposition is characterized by using an energy selective mass spectrometer situated at the wall of the processing chamber: measurements of positive and negative ion energy distribution functions and mass spectra (1⩽amu⩽150) are obtained for various flow-rate ratios (R=O2/SiH4=1 to 10 but constant total flow rate of 30 sccm), and for a constant radio frequency power and magnetic field of 800 W and 70 G, respectively. Plasma potentials between 35 (R=10) and 60 V (R=1) are measured depending on the silane and oxygen flows showing charging of the silica-covered diffusion chamber wall. The magnitude of the wall charging most likely depends on the effective capacitance formed by the silica layer (which results from months of deposition) and on the inbalance between the positively and negatively charged particles which impinge onto the sidewalls at the discharge initiation until equilibrium of fluxes is reached. This inbalance is a result of the magnetic field configuration generated by the four coils surrounding the reactor and of the subsequent cross-field diffusion of the positively and negatively charged particles to the sidewalls. Maximum wall charging is observed when the silane flow is maximum (R<2), a situation where polymerization is observed in the mass spectrum of the positive ions, and where a minimum density of negative ions (O−, OH−, and H−) are detected close to the walls. Although the polymerization appeared to be a primary candidate for the increase of the plasma potential to ∼60 V, its presence does not significantly change the total positive ion density at equilibrium but is accompanied by a dramatic decrease (two orders of magnitude for the O− ions) in the negative ion density close to the walls when R is increased from 1 to 10. The change in the degree of electronegativity close to the walls affects the global plasma equilibrium and appears as an indirect factor in the magnitude of the wall charging.
Substrate bias dependence of the structure and internal stress of TiN films deposited by the filtered cathodic vacuum arc
Authors: Cheng Y. H. , Tay B. K. , Lau S. P.
Published: July 2002

TiN films were deposited by an off-plane double bend filtered cathodic vacuum arc technique. Atomic force microscopy and x-ray diffraction were used to study the surface morphology and crystal structure. Substrate bending methods were used to determine the internal stress in the films. The influence of substrate bias on the structure and internal stress were systematically studied. As substrate bias is increased to −100 V, the surface roughness decreases to the minimum, the preferred orientation changes gradually from (200) to (111), and the internal stress increases greatly to the maximum. Further increase of substrate bias results in the drastic increase of surface roughness and the gradual decrease of internal stress. The preferred orientation changes gradually from (111) to (200). Except for the atomic peening, the change in the structure and preferred orientation also contributes to the variation of internal stress in the films with substrate bias.
Characterization of cubic boron nitride growth using UV-extended real-time spectroscopic ellipsometry: Effect of plasma additions and dynamic substrate bias steps
Authors: Zapien J. A. , Collins R. W. , Messier R.
Published: July 2002

Boron nitride (BN) films with high cubic content were deposited using pulsed dc sputtering of a B4C target and rf biasing of the substrate. The film deposition was characterized using ultraviolet-extended real-time spectroscopic ellipsometry (UV-RTSE). A three-layer optical model with constrained evolution of the layer thicknesses was used to characterize the growth of the BN films on c-Si substrates. This model was chosen for consistency with previous research and includes (i) an initial hexagonal BN (hBN) layer; (ii) a hexagonal+cubic BN (hBN+cBN) mixed-phase transition layer; and (iii) a topmost cBN layer. The evolution of the layer thicknesses was obtained as a function of the substrate bias (Vavg) used to deposit the BN films under static conditions. These results showed a minimum in the thicknesses of the hBN and mixed-phase layers, thus identifying an optimum Vavg for cBN nucleation and growth from the hBN phase under the deposition conditions used here. Complementary ex situ infrared transmission spectroscopy confirmed that UV-RTSE provides high sensitivity for resolving the crystallographic phase of BN due to differences in the hBN and cBN optical functions. The model with constrained evolution of thicknesses was also applied to determine the phase evolution of BN under dynamic conditions in which the substrate bias magnitude |Vavg| was decreased in a stepwise manner during deposition. As a result, the critical substrate bias voltage VCm needed to sustain pure cBN growth was determined. It was found that for |Vavg|<|VCm| a second hBN+cBN mixed-phase layer evolves and that upon further reduction in |Vavg| an onset for the formation of a pure hBN layer appears. The effects of plasma additions of hydrogen and oxygen on the BN phase evolution and the cBN optical properties were documented in dynamic |Vavg| step-down experiments. It was found that any additions of hydrogen or oxygen gas are detrimental when used under otherwise optimum cBN growth conditions, meaning that these additions promote the growth of hBN.
Calibration of new high dose rate sources
Authors: Stump K. E. , DeWerd L. A. , Micka J. A. , Anderson D. R.
Published: July 2002

The increased popularity of high dose rate (HDR) brachytherapy in the treatment of neoplastic disease has resulted in the introduction of new or redesigned sources by various manufacturers. In order to provide accurate clinical dosimetry, accurate calibration of these sources is required. In this work, two new sources are calibrated with the standard, NIST traceable, in‐air calibration technique. These sources are the Varian VariSource (VS2000) and the newly redesigned Nucletron MicroSelectron source. An uncertainty budget was performed to determine the overall uncertainty of the measurement, and a total relative expanded uncertainty of 2.15% was calculated. The results of the source calibration using the in‐air technique and that obtained based upon the 1991 standard calibration all lie well within this uncertainty. In addition to the in‐air calibration, contact autoradiographs of the sources were made with International Specialty Products model HD810 radiochromic film to demonstrate the sources relative dose distributions.
Fast cavitation
Authors: Heyda P. G.
Published: July 2002

Underwater projectiles and vehicles may be able to travel at hundreds of miles per hour with the help of an attached cavity that is produced mainly by inertial forces and can greatly reduce fluid drag. A simple, idealized model explains the essentials of the phenomenon.
Effective range theory
Authors: Madsen Lars Bojer
Published: July 2002

The effective range formula for scattering by a short range potential is derived for arbitrary angular momentum of the scattered particle. The derivation is a generalization of a method due to Bethe, and makes no reference to the analytical properties of Jost functions. The derivation is a useful supplement to the textbook coverage of the subject.
Probing physics students’ conceptual knowledge structures through term association
Authors: Beatty Ian D. , Gerace William J.
Published: July 2002

Traditional tests are not effective tools for diagnosing the content and structure of students’ knowledge of physics. As a possible alternative, a set of term-association tasks (the ConMap tasks) was developed to probe the interconnections within students’ store of conceptual knowledge. The tasks have students respond spontaneously to a term or problem or topic area with a sequence of associated terms; the response terms and time-of-entry data are captured. The tasks were tried on introductory physics students, and preliminary investigations show that the tasks are capable of eliciting information about the stucture of their knowledge. Specifically, data gathered through the tasks is similar to that produced by a hand-drawn concept map task, has measures that correlate with in-class exam performance, and is sensitive to learning produced by topic coverage in class. Although the results are preliminary and only suggestive, the tasks warrant further study as student-knowledge assessment instruments and sources of experimental data for cognitive modeling efforts.
Ship recognition via its radiated sound: The fractal based approaches
Authors: Yang Su , Li Zhishun , Wang Xinlong
Published: July 2002

Due to the complexity of its radiated sound, ship recognition is difficult. Fractal approaches are proposed in this study, including fractal Brownian motion based analysis, fractal dimension analysis, and wavelet analysis, to augment existing feature extraction methods that are based on spectrum analysis. Experimental results show that fractal approaches are effective. When used to augment two traditional features, line and average spectra, fractal approaches led to better classification results. This implies that fractal approaches can capture some information not detected by traditional approaches alone.
Making Sense of Changing Animal Embryos
Authors: Swalla Billie J.
Published: July 2002

The Evolution of Developmental Pathways
. Adam S. Wilkins. Sinauer Associates, Sunderland, MA, 2001. 621 pp. $54.95. ISBN 0-87893-916-4.

Wilkins explores the nature of the genetic, molecular, and selectional events that affect genetic pathways and determine how shape and form in complex organisms change during evolution.
Identification of Signal Peptide Peptidase, a Presenilin-Type Aspartic Protease
Authors: Weihofen Andreas , Binns Kathleen , Lemberg Marius K. , Ashman Keith , Martoglio Bruno
Published: July 2002

Signal peptide peptidase (SPP) catalyzes intramembrane proteolysis of some signal peptides after they have been cleaved from a preprotein. In humans, SPP activity is required to generate signal sequence–derived human lymphocyte antigen–E epitopes that are recognized by the immune system, and to process hepatitis C virus core protein. We have identified human SPP as a polytopic membrane protein with sequence motifs characteristic of the presenilin-type aspartic proteases. SPP and potential eukaryotic homologs may represent another family of aspartic proteases that promote intramembrane proteolysis to release biologically important peptides.
Unique Chromatin Remodeling and Transcriptional Regulation in Spermatogenesis
Authors: Sassone-Corsi Paolo
Published: July 2002

Most of our knowledge of transcriptional regulation comes from studies in somatic cells. However, increasing evidence reveals that gene regulation mechanisms are different in haploid germ cells. A number of highly specialized strategies operate during spermatogenesis. These include a unique chromatin reorganization program and the use of distinct promoter elements and specific transcription factors. Deciphering the rules governing transcriptional control during spermatogenesis will provide valuable insights of biomedical importance.
Out of the Chattering Ice
Authors: Proctor Robert N.
Published: July 2002

A Brain for All Seasons
Human Evolution and Abrupt Climate Change. William H. Calvin. University of Chicago Press, Chicago, 2002. 349 pp. $25, £16. ISBN 0-226-09201-1.

Through the course of a travelogue, Calvin argues that repeated episodes of abrupt cooling played a critical role in human evolution and that human-caused global warming may paradoxically trigger another crash in global temperatures.
Mammals on the European Stage
Authors: Flannery Tim
Published: July 2002

Mammoths, Sabertooths, and Hominids
65 Million Years of Mammalian Evolution in Europe. Jordi Agustí and Mauricio Antón. Columbia University Press, New York, 2002. 325 pp. $39.50, £27.50. ISBN 0-231-11640-3.

Agustí and Antón provide a broad overview of the Tertiary history of mammals in Europe: the evolutionary changes within the European fauna as well as the fates of immigrant taxa that arrived from other continents.
In Vitro Activities of Iboga Alkaloid Congeners Coronaridine and 18-Methoxycoronaridine againstLeishmania amazonensis
Authors: Delorenzi Jan Carlo , Freire-de-Lima Leonardo , Gattass Cerli R. , de Andrade Costa Deise , He Liwen , Kuehne Martin E. , Saraiva Elvira M. B.
Published: July 2002

In previous studies, we demonstrated the leishmanicide effect of coronaridine, a natural indole alkaloid isolated from stem bark ofPeschiera australis(Delorenzi et al., Antimicrob. Agents Chemother.45:1349-1354, 2001). In this study we show the leishmanicidal effect of the synthetic coronaridine and its racemic 18-methoxylated analog, 18-methoxycoronaridine. Both alkaloids revealed a potent leishmanicide effect againstLeishmania amazonensis, a causative agent of cutaneous and diffuse cutaneous leishmaniasis in the New World. Despite their potent leishmanicide effect, both alkaloids were neither toxic to murine macrophages nor did they modulate their oxidative or cytokine production responses.
Mn(III) Pyrophosphate as an Efficient Tool for Studying the Mode of Action of Isoniazid on the InhA Protein of
Mycobacterium tuberculosis
Authors: Nguyen Michel , Quémard Annaïk , Broussy Sylvain , Bernadou Jean , Meunier Bernard
Published: July 2002

The antituberculosis drug isoniazid (INH) is quickly oxidized by stoichiometric amounts of manganese(III) pyrophosphate. In the presence of nicotinamide coenzymes (NAD
+
, NADH, nicotinamide mononucleotide [NMN
+
]) and nicotinic acid adenine dinucleotide (DNAD
+
), INH oxidation produced the formation of INH-coenzyme adducts in addition to known biologically inactive products (isonicotinic acid, isonicotinamide, and isonicotinaldehyde). A pool of INH-NAD(H) adducts preformed in solution allowed the rapid and strong inhibition of in vitro activity of the enoyl-acyl carrier protein reductase InhA, an INH target in the biosynthetic pathway of mycolic acids: the inhibition was 90 or 60% when the adducts were formed in the presence of NAD
+
or NADH, respectively. Under similar conditions, no inhibitory activity of INH-NMN(H) and INH-DNAD(H) adducts was detected. When an isolated pool of 100 nM INH-NAD(H) adducts was first incubated with InhA, the enzyme activity was inhibited by 80%; when present in excess, both NADH and decenoyl-coenzyme A are able to prevent this phenomenon. InhA inhibition by several types of INH-coenzyme adducts coexisting in solution is discussed in relation with the structure of the coenzyme, the stereochemistry of the adducts, and their existence as both open and cyclic forms. Thus, manganese(III) pyrophosphate appears to be an efficient and convenient alternative oxidant to mimic the activity of the
Mycobacterium tuberculosis
KatG catalase-peroxidase and will be useful for further mechanistic studies of INH activation and for structural investigations of reactive INH species in order to promote the design of new inhibitors of InhA as potential antituberculous drugs.
Combination of Quinupristin-Dalfopristin and Gentamicin against Methicillin-Resistant
Staphylococcus aureus
: Experimental Rabbit Endocarditis Study
Authors: Batard Eric , Jacqueline Cedric , Boutoille David , Hamel Antoine , Drugeon Henri B. , Asseray Nathalie , Leclercq Roland , Caillon Jocelyne , Potel Gilles , Bugnon Denis
Published: July 2002

The combination of quinupristin-dalfopristin (Q-D) and gentamicin was tested against two strains of gentamicin- and dalfopristin-susceptible methicillin-resistant
Staphylococcus aureus
(MRSA). One strain was susceptible to macrolides, lincosamides, and streptogramin B type antibiotics (MLS
B
), and the other was constitutively resistant to these antibiotics by virtue of the
ermA
gene. The checkerboard method and time-kill curves showed that the combination of Q-D and gentamicin was indifferent. A rabbit endocarditis model simulated the pharmacokinetics achieved in humans receiving intravenous injections of Q-D (7.5 mg/kg of body weight three times a day) and gentamicin (3 mg/kg once daily). For the MLS
B
-susceptible strain, a 4-day regimen reduced mean bacterial titers (MBT) in vegetations from 8.5 ± 0.8 log CFU/g (control group) to 4.1 ± 2.6 (gentamicin), 3.0 ± 0.9 (Q-D), and 2.6 ± 0.5 log CFU/g (Q-D plus gentamicin). For the strain constitutively resistant to MLS
B
, a 4-day regimen reduced MBT in vegetations from 8.7 ± 0.9 log CFU/g (control group) to 5.0 ± 2.2 (gentamicin), 5.2 ± 2.2 (Q-D), and 5.1 ± 2.4 log CFU/g (Q-D plus gentamicin). The differences between control and treatment groups were significant for both strains (
P
< 0.0001), although there was no significant difference between treatment groups. No resistant variant was isolated from vegetations, and no significant difference in MBT in vegetations of treatment groups after 1-day regimens was observed. This experimental study found no additive benefit in combining Q-D and gentamicin against dalfopristin- and gentamicin-susceptible MRSA.
Efficacy of Liposomal Amphotericin B in Treatment of Systemic Murine Fusariosis
Authors: Ortoneda Montserrat , Capilla Javier , Pastor Francisco J. , Pujol Isabel , Guarro Josep
Published: July 2002

We have compared the activities of liposomal amphotericin B (LAMB) at 3, 5, 10, and 20 mg/kg/day and amphotericin B deoxycholate (AMB) at 1.5 and 2.5 mg/kg/day in a murine systemic infection by
Fusarium verticillioides
. Survival was improved by all treatments except AMB at 1.5 mg/kg/day. The tissue burden in liver was reduced by LAMB at all dosages and by AMB at 2.5 mg/kg/day. The two highest dosages of LAMB showed significant reductions in the spleen.
Comparative In Vitro Sensitivities of Human Immune Cell Lines, Vaginal and Cervical Epithelial Cell Lines, and Primary Cells to Candidate Microbicides Nonoxynol 9, C31G, and Sodium Dodecyl Sulfate
Authors: Krebs Fred C. , Miller Shendra R. , Catalone Bradley J. , Fichorova Raina , Anderson Deborah , Malamud Daniel , Howett Mary K. , Wigdahl Brian
Published: July 2002

In experiments to assess the in vitro impact of the candidate microbicides nonoxynol 9 (N-9), C31G, and sodium dodecyl sulfate (SDS) on human immune and epithelial cell viability, cell lines and primary cell populations of lymphocytic and monocytic origin were generally shown to be equally sensitive to exposures ranging from 10 min to 48 h. However, U-937 cells were more sensitive to N-9 and C31G after 48 h than were primary monocyte-derived macrophages. Cytokine activation of monocytes and lymphocytes had no effect on cell viability following exposure to these microbicidal compounds. Primary and passaged vaginal epithelial cultures and cell lines differed in sensitivity to N-9 and C31G but not SDS. These studies provide a foundation for in vitro experiments in which cell lines of human immune and epithelial origin can be used as suitable surrogates for primary cells to further investigate the effects of microbicides on cell metabolism, membrane composition, and integrity and the effects of cell type, proliferation, and differentiation on microbicide sensitivity.
Short-Chain Aliphatic Polysulfonates Inhibit the Entry of
Plasmodium
into Red Blood Cells
Authors: Kisilevsky Robert , Crandall Ian , Szarek Walter A. , Bhat Shridhar , Tan Christopher , Boudreau Lee , Kain Kevin C.
Published: July 2002

Several steps in the pathogenesis of a
Plasmodium falciparum
infection depend on interactions of parasite surface proteins with negatively charged sugars on the surface of host cells such as sialate residues or glycosaminoglycans. For these reasons, our previous studies examining agents that interfere with heparan sulfate-protein binding during amyloidogenesis suggested that short-chain aliphatic polysulfonates may prove useful as antimalarial agents. A series of related polysulfonates were synthesized and assessed both in tissue culture with the asexual stages of
P. falciparum
in human red blood cells and in vivo by use of
Plasmodium berghei
infections in mice. Poly(vinylsulfonate sodium salt) (molecular weight range, 1,500 to 3,000) proved effective in interfering with
P. falciparum
merozoite entry into human red blood cells and significantly delaying the increase in the level of
P. berghei
parasitemia in mice. The concept that anionic molecules that mimic large polysaccharide structures may have antimalarial properties has been suggested and examined previously. Our results suggest that related anionic agents [poly(vinylsulfonate sodium salt)-like molecules] orders of magnitude smaller than those previously considered may prove useful in abrogating merozoite entry into erythrocytes and may potentially block sporozoite entry into liver cells. Structure-activity studies conducted to enhance these properties may provide compounds with scope for significant further analysis and development.
Secondary (γ-
Proteobacteria
) Endosymbionts Infect the Primary (β-
Proteobacteria
) Endosymbionts of Mealybugs Multiple Times and Coevolve with Their Hosts
Authors: Thao MyLo Ly , Gullan Penny J. , Baumann Paul
Published: July 2002

Mealybugs (Hemiptera, Coccoidea, Pseudococcidae) are plant sap-sucking insects that have within their body cavities specialized cells containing prokaryotic primary endosymbionts (P-endosymbionts). The P-endosymbionts have the unusual property of containing within their cytoplasm prokaryotic secondary endosymbionts (S-endosymbionts) [C. D. von Dohlen, S. Kohler, S. T. Alsop, and W. R. McManus, Nature (London) 412:433-436, 2001]. Four-kilobase fragments containing 16S-23S ribosomal DNA (rDNA) were obtained from the P-endosymbionts of 22 mealybug species and the S-endosymbionts of 12 representative species. Phylogenetic analyses of the P-endosymbionts indicated that they have a monophyletic origin and are members of the β-subdivision of the
Proteobacteria
; these organisms were subdivided into five different clusters. The S-endosymbionts were members of the γ-subdivision of the
Proteobacteria
and were grouped into clusters similar to those observed with the P-endosymbionts. The S-endosymbiont clusters were distinct from each other and from other insect-associated bacteria. The similarity of the clusters formed by the P- and S-endosymbionts suggests that the P-endosymbionts of mealybugs were infected multiple times with different precursors of the S-endosymbionts and once the association was established, the P- and S-endosymbionts were transmitted together. The lineage consisting of the P-endosymbionts of mealybugs was given the designation “
Candidatus
Tremblaya” gen. nov., with a single species, “
Candidatus
Tremblaya princeps” sp. nov. The results of phylogenetic analyses of mitochondrial DNA fragments encoding cytochrome oxidase subunits I and II from four representative mealybug species were in agreement with the results of 16S-23S rDNA analyses, suggesting that relationships among strains of “
Candidatus
T. princeps” are useful in inferring the phylogeny of their mealybug hosts.
Phage-Displayed Peptides as Developmental Agonists for
Phytophthora capsici
Zoospores
Authors: Bishop-Hurley Sharon L. , Mounter Sarah A. , Laskey James , Morris Roy O. , Elder Jim , Roop Philip , Rouse Chris , Schmidt Francis J. , English James T.
Published: July 2002

As part of its pathogenic life cycle,
Phytophthora capsici
disperses to plants through a motile zoospore stage. Molecules on the zoospore surface are involved in reception of environmental signals that direct preinfection behavior. We developed a phage display protocol to identify peptides that bind to the surface molecules of
P. capsici
zoospores in vitro
.
The selected phage-displayed peptides contained an abundance of polar amino acids and proline but were otherwise not conserved. About half of the selected phage that were tested concomitantly induced zoospore encystment in the absence of other signaling agents. A display phage was shown to bind to the zoospore but not to the cyst form of
P. capsici.
Two free peptides corresponding to active phage were similarly able to induce encystment of zoospores, indicating that their ability to serve as signaling ligands did not depend on their exact molecular context. Isolation and subsequent expression of peptides that act on pathogens could allow the identification of receptor molecules on the zoospore surface, in addition to forming the basis for a novel plant disease resistance strategy.
Genotypic and Phenotypic Diversity within Species of Purple Nonsulfur Bacteria Isolated from Aquatic Sediments
Authors: Oda Yasuhiro , Wanders Wouter , Huisman Louis A. , Meijer Wim G. , Gottschal Jan C. , Forney Larry J.
Published: July 2002

To assess the extent of genotypic and phenotypic diversity within species of purple nonsulfur bacteria found in aquatic sediments, a total of 128 strains were directly isolated from agar plates that had been inoculated with sediment samples from Haren and De Biesbosch in The Netherlands. All isolates were initially characterized by BOX-PCR genomic DNA fingerprinting, and 60 distinct genotypes were identified. Analyses of 16S rRNA gene sequences of representatives of each genotype showed that five and eight different phylotypes of purple nonsulfur bacteria were obtained from the Haren and De Biesbosch sites, respectively. At the Haren site, 80.5% of the clones were
Rhodopseudomonas palustris
, whereas
Rhodoferax fermentans
and
Rhodopseudomonas palustris
were numerically dominant at the De Biesbosch site and constituted 45.9 and 34.4% of the isolates obtained, respectively. BOX-PCR genomic fingerprints showed that there was a high level of genotypic diversity within each of these species. The genomic fingerprints of
Rhodopseudomonas palustris
isolates were significantly different for isolates from the two sampling sites, suggesting that certain strains may be endemic to each sampling site. Not all
Rhodopseudomonas palustris
isolates could degrade benzoate, a feature that has previously been thought to be characteristic of the species. There were differences in the BOX-PCR genomic fingerprints and restriction fragment length polymorphisms of benzoate-coenzyme A ligase genes and form I and form II ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) genes between benzoate-degrading and non-benzoate-degrading genotypes. The ability to distinguish these two
Rhodopseudomonas palustris
groups based on multiple genetic differences may reflect an incipient speciation event resulting from adaptive evolution to local environmental conditions.
Influence of the Transposition of the Thermostabilizing Domain of
Clostridium thermocellum
Xylanase (XynX) on Xylan Binding and Thermostabilization
Authors: Shin Eun-Sun , Yang Mi-Jeong , Jung Kyung Hwa , Kwon Eun-Ju , Jung Jae Sung , Park Seur Kee , Kim Jungho , Yun Han Dae , Kim Hoon
Published: July 2002

A xylanase gene,
xynX
, of
Clostridium thermocellum
had one thermostabilizing domain (TSD) between the signal peptide sequence and the catalytic domain (CD). The TSD of a truncated xylanase gene,
xynX
′
TSD-CD
, was transpositioned from the N terminus to the C terminus of the CD by overlapping PCRs, and a modified product,
xynX
′
CD-TSD
, was constructed. XynX′
TSD-CD
had a higher optimum temperature (70°C versus 65°C) and was more thermostable (residual activity of 68% versus 46% after a 20-min preincubation at 70°C) than the one without the TSD, XynX′
CD
. However, the domain-transpositioned enzyme, XynX′
CD-TSD
, showed a lower optimum temperature (30°C) and thermostability (20%) than XynX′
CD
. Both XynX′
TSD-CD
and XynX′
CD-TSD
showed significantly higher binding capacity toward xylan than XynX′
CD
, and the domain transposition did not cause any change in the binding ability. XynX′
TSD-CD
and XynX′
CD-TSD
also showed considerable binding to lichenan but not to carboxymethyl cellulose and laminarin. XynX′
TSD-CD
and XynX′
CD-TSD
had higher activities for insoluble xylan than XynX′
CD
, while XynX′
CD
was more active against soluble xylan than XynX′
TSD-CD
and XynX′
CD-TSD
. These results indicate that the TSD of XynX has dual functions, xylan binding and thermostabilization, and the domain should also be classified as a xylan-binding domain (XBD). The binding capacity of the XBD was not affected by domain transpositioning within the gene.
Biodegradation of 1,2,3-Trichloropropane through Directed Evolution and Heterologous Expression of a Haloalkane Dehalogenase Gene
Authors: Bosma Tjibbe , Damborský Jirí , Stucki Gerhard , Janssen Dick B.
Published: July 2002

Using a combined strategy of random mutagenesis of haloalkane dehalogenase and genetic engineering of a chloropropanol-utilizing bacterium, we constructed an organism that is capable of growth on 1,2,3-trichloropropane (TCP). This highly toxic and recalcitrant compound is a waste product generated from the manufacture of the industrial chemical epichlorohydrin. Attempts to select and enrich bacterial cultures that can degrade TCP from environmental samples have repeatedly been unsuccessful, prohibiting the development of a biological process for groundwater treatment. The critical step in the aerobic degradation of TCP is the initial dehalogenation to 2,3-dichloro-1-propanol. We used random mutagenesis and screening on eosin-methylene blue agar plates to improve the activity on TCP of the haloalkane dehalogenase fromRhodococcussp. m15-3 (DhaA). A second-generation mutant containing two amino acid substitutions, Cys176Tyr and Tyr273Phe, was nearly eight times more efficient in dehalogenating TCP than wild-type dehalogenase. Molecular modeling of the mutant dehalogenase indicated that the Cys176Tyr mutation has a global effect on the active-site structure, allowing a more productive binding of TCP within the active site, which was further fine tuned by Tyr273Phe. The evolved haloalkane dehalogenase was expressed under control of a constitutive promoter in the 2,3-dichloro-1-propanol-utilizing bacteriumAgrobacterium radiobacterAD1, and the resulting strain was able to utilize TCP as the sole carbon and energy source. These results demonstrated that directed evolution of a key catabolic enzyme and its subsequent recruitment by a suitable host organism can be used for the construction of bacteria for the degradation of a toxic and environmentally recalcitrant chemical.
Differentiation of
Paenibacillus larvae
subsp.
larvae
, the Cause of American Foulbrood of Honeybees, by Using PCR and Restriction Fragment Analysis of Genes Encoding 16S rRNA
Authors: Alippi Adriana M. , López Ana Claudia , Aguilar O. Mario
Published: July 2002

A rapid procedure for the identification of
Paenibacillus larvae
subsp.
larvae
, the causal agent of American foulbrood (AFB) disease of honeybees (
Apis mellifera
L.), based on PCR and restriction fragment analysis of the 16S rRNA genes (rDNA) is described. Eighty-six bacterial strains belonging to 39 species of the genera
Paenibacillus
,
Bacillus
,
Brevibacillus
, and
Virgibacillus
were characterized. Amplified rDNA was digested with seven restriction endonucleases. The combined data from restriction analysis enabled us to distinguish 35 profiles. Cluster analysis revealed that
P. larvae
subsp.
larvae
and
Paenibacillus larvae
subsp.
pulvifaciens
formed a group with about 90% similarity; however, the
P. larvae
subsp.
larvae
restriction fragment length polymorphism pattern produced by endonuclease
Hae
III was found to be unique and distinguishable among other closely related bacteria. This pattern was associated with DNA extracted directly from honeybee brood samples showing positive AFB clinical signs that yielded the restriction profile characteristic of
P. larvae
subsp.
larvae
, while no amplification product was obtained from healthy larvae. The method described here is particularly useful because of the short time required to carry it out and because it allows the differentiation of
P. larvae
subsp.
larvae
-infected larvae from all other species found in apiarian sources.
Value of Microimmunofluorescence for Diagnosis and Follow-up of
Bartonella
Endocarditis
Authors: Fournier Pierre-Edouard , Mainardi Jean-Luc , Raoult Didier
Published: July 2002

Bartonella
endocarditis is a disease of emerging importance that causes serious complications and high rates of mortality. Due to the fastidious nature of
Bartonella
species and their high degrees of antibiotic susceptibility, cultures of clinical samples most often remain sterile and valvular biopsy specimens, the best specimens for PCR amplification, are seldom available. Therefore, serology appears to be the easiest diagnostic tool. In order to determine the best cutoff value for serology and its predictive values for the detection of
Bartonella
endocarditis, we studied 48 patients with culture- and/or PCR-confirmed
Bartonella
endocarditis. We also applied these serological criteria to 156 patients with blood culture-negative endocarditis. Furthermore, we compared the kinetics of the antibody responses to
Bartonella
spp. in order to estimate the value of serology for prediction of the occurrence of relapses. A titer of ≥1:800 for immunoglobulin G antibodies to either
Bartonella henselae
or
B. quintana
has a positive predictive value of 0.810 for the detection of chronic
Bartonella
infections in the general population and a value of 0.955 for the detection of
Bartonella
infections among patients with endocarditis. When this cutoff was applied to 156 patients with blood culture-negative endocarditis, we were able to diagnose
Bartonella
infections in an additional 45 patients with definite endocarditis for whom a positive
Bartonella
serology was the only evidence of infection. On follow-up, the kinetics of the decrease in antibody titers were significantly delayed in two patients with relapses. In conclusion, we recommend the determination of antibodies to both
B. quintana
and
B. henselae
and the use of a cutoff value of 1:800 for the diagnosis of
Bartonella
endocarditis. We propose that this criterion, which may also help with the detection of late relapses, be included as a major criterion in the Duke criteria for the diagnosis of infective endocarditis.
MaGiK Method of T-Cell Receptor Repertoire Analysis
Authors: Killian M. Scott , Matud Jose , Detels Roger , Giorgi Janis V. , Jamieson Beth D.
Published: July 2002

T-cell receptor diversity enables the cellular immune response to recognize a broad range of viral and other pathogenic agents. An increasingly common method of characterizing T-cell receptor diversity and usage in response to antigenic challenges involves the identification of clonal expansions by PCR amplification of the CDR3 region of distinct TCRVβ families. Though clonal expansions often appear evident upon visual inspection of the results, a systematic method is needed for the valid enumeration of these expansions. Here, we describe a novel analysis method, termed the MaGiK method, for systematically identifying and enumerating clonal T-cell expansions and for applying the results to investigations of the T-cell receptor repertoire.
Induction of Cytokines by Glucosyltransferases ofStreptococcus mutans
Authors: Chia Jean-San , Lien Huei-Ting , Hsueh Po-Ren , Chen Pei-Min , Sun Andy , Chen Jen-Yang
Published: July 2002

Production of proinflammatory cytokines is implicated in the pathogenesis of viridans streptococcus-induced α-streptococcal shock syndrome and infective endocarditis.Streptococcus mutans, one of the opportunistic pathogens causing infective endocarditis, was reported previously to stimulate monocytes and epithelial and endothelial cells in vitro to produce various cytokines. We found that glucosyltransferases (GTFs) GtfC and GtfD ofS. mutansstimulated predominantly the production of interleukin-6 (IL-6) from T cells cultured in vitro. The level of IL-6 but not of tumor necrosis factor alpha in blood was significantly elevated when rats were injected intravenously withS. mutansGS-5, whereas IL-6 was detected at a much lower level when rats were challenged with NHS1DD, an isogenic mutant defective in the expression of GTFs. The serum IL-6 level was elevated in patients with endocarditis caused by different species of viridans streptococci which express GTF homologues. Affinity column-purified GTFs reduced the levels of detectable IL-2 of T cells stimulated by another bacterial antigen, tetanus toxoid. These results suggested that GTFs might modulate the production of Th1-type cytokines and that GTFs ofS. mutansplay a significant role in stimulating the production of the proinflammatory cytokine IL-6 in vivo.
Complement-Mediated Neutralization of Canine Distemper Virus In Vitro: Cross-Reaction between Vaccine Onderstepoort and Field KDK-1 Strains with Different Hemagglutinin Gene Characteristics
Authors: Mochizuki Masami , Motoyoshi Megumi , Maeda Ken , Kai Kazunari
Published: July 2002

The properties of neutralization of antigens of canine distemper virus Onderstepoort and a recent field isolate, KDK-1, were investigated with strain-specific dog sera. A conventional neutralization assay indicated antigenic dissimilarity between the strains; however, when guinea pig complement was included in the reaction mixture, the strains were neutralized with not only the homologous but also the heterologous antibodies.
Manipulations in Maternal Environment Reverse Periodontitis in Genetically Predisposed Rats
Authors: Sluyter Frans , Breivik Torbjørn , Cools Alexander
Published: July 2002

The predisposition to develop periodontitis is partly genetically determined in humans as well as in animals. Here we demonstrate, however, that early manipulations in the maternal environment of an animal (rat) model of periodontitis can fully reverse the genetic predisposition to develop periodontitis at adult age.
Leukotriene B4Induces Nitric Oxide Synthesis inTrypanosoma cruzi-Infected Murine Macrophages and Mediates Resistance to Infection
Authors: Talvani A. , Machado F. S. , Santana G. C. , Klein A. , Barcelos L. , Silva J. S. , Teixeira M. M.
Published: July 2002

The production of nitric oxide (NO) by gamma interferon (IFN-γ)-activated macrophages is a major effector mechanism during experimentalTrypanosoma cruziinfection. In addition to IFN-γ, chemoattractant molecules, such as platelet-activating factor (PAF) and CC chemokines, may also activate macrophages to induce NO and mediate the killing ofT. cruziin an NO-dependent manner. Here we investigated the ability of leukotriene B4(LTB4) to induce the production of NO by macrophages infected withT. cruziin vitro and whether NO mediated LTB4-induced parasite killing. The activation ofT. cruzi-infected but not naive murine peritoneal macrophages with LTB4induced the time- and concentration-dependent production of NO. In addition, low concentrations of LTB4acted in synergy with IFN-γ to induce NO production. The NO produced mediated LTB4-induced microbicidal activity in macrophages, as demonstrated by the inhibitory effects of an inducible NO synthase inhibitor. LTB4-induced NO production and parasite killing were LTB4receptor dependent and were partially blocked by a PAF receptor antagonist. LTB4also induced significant tumor necrosis factor alpha (TNF-α) production, and blockade of TNF-α suppressed LTB4-induced NO release and parasite killing. A blockade of LTB4or PAF receptors partially inhibited IFN-γ-induced NO and TNF-α production but not parasite killing. Finally, daily treatment of infected mice with CP-105,696 was accompanied by a significantly higher level of blood parasitemia, but not lethality, than that seen in vehicle-treated animals. In conclusion, our results suggest a role for LTB4during experimentalT. cruziinfection. Chemoattractant molecules such as LTB4not only may play a major role in leukocyte migration into sites of inflammation in vivo but also, in the event of an infection, may play a relevant role in the activation of recruited leukocytes to kill the invading microorganism in an NO-dependent manner.
FimbriatedSalmonella entericaSerovar Typhimurium Abates Initial Inflammatory Responses by Macrophages
Authors: Pascual David W. , Trunkle Theresa , Sura Jamie
Published: July 2002

Oral immunization of mice with aSalmonellavaccine expressing colonization factor antigen I (CFA/I) from enterotoxigenicEscherichia coliresults in the rapid onset of interleukin-4 (IL-4) and IL-5 production, which explains the observed elevations in mucosal immunoglobulin A (IgA) and serum IgG1 antibodies. In contrast, oral immunization with theSalmonellavector does not result in the production of Th2-type cytokines. To begin to assess why such differences exist between the two strains, it should be noted that in vitro infection of RAW 264.7 macrophages resulted in the absence of nitric oxide (NO) production in cells infected with theSalmonella-CFA/I vaccine. This observation suggests differential proinflammatory cytokine production by these isogenicSalmonellastrains. Upon measurement of proinflammatory cytokines, minimal to no tumor necrosis factor alpha (TNF-α), IL-1α, IL-1β, or IL-6 was produced bySalmonella-CFA/I-infected RAW 264.7 or peritoneal macrophages, but production was greatly induced inSalmonellavector-infected macrophages. Only minute levels of IL-12 p70 were induced bySalmonellavector-infected macrophages, and none was induced bySalmonella-CFA/I-infected macrophages. The absence of IL-12 was not due to overt increases in production of either IL-12 p40 or IL-10. CFU measurements taken at 8 h postinfection showed no differences in colonization in RAW 264.7 cells infected with eitherSalmonellaconstruct, but there were differences in peritoneal macrophages. However, after 24 h, theSalmonellavector strain colonized to a greater extent in RAW 264.7 cells than in peritoneal macrophages. Infection of RAW 264.7 cells or peritoneal macrophages with eitherSalmonellaconstruct showed no difference in macrophage viabilities. This evidence shows that the expression of CFA/I fimbriae alters how macrophages recognize or process salmonellae and prevents the rapid onset of proinflammatory cytokines which is typical duringSalmonellainfections.