bluuebunny/crossref_metadata_2025 huggingface dataset:
Long Crack R‐Curve of Aligned Porous Silicon Nitride
Authors: Deng Zhen‐Yan , Inagaki Yoshiaki , She Jihong , Tanaka Yoshihisa , Liu Yu‐Fu , Sakamoto Masao , Ohji Tatsuki
Published: February 2005

Long crack R‐curve of a porous Si3N4 with aligned fibrous grains was investigated, using a chevron‐notched beam technique. A crack was constrained to propagate normal to the grain alignment. The crack growth resistance of aligned porous Si3N4 was much larger compared with that of dense Si3N4 ceramics. Microstructure observations showed that pullouts of fibrous grains in aligned porous Si3N4 markedly increased during crack propagation relative to those of dense Si3N4, due to the existence of pores. The efficient grain pullouts in porous Si3N4 increased the bridging stress at the crack wake.
A General Process for Preparation of Core‐Shell Particles of Complete and Smooth Shells
Authors: Chen Gia‐Chi , Kuo Cheng‐Yu , Lu Shih‐Yuan
Published: February 2005

Core‐shell particles have attracted much research attention in recent years because of the great potential in protection, modification, and functionalization of the core particles with suitable shell materials to achieve specific physical, chemical, and biological performance. A general approach for preparation of core‐shell particles possessing complete and smooth shells is presented. The shell layer was constructed by introducing nanoparticles of desired shell material, prepared with the reverse micelle process, onto the core particle surface via layer‐by‐layer electrostatic adsorption, hydrophilic and hydrophobic interactions, and subsequent sintering. The proposed process is illustrated by coating silica spheres with four different shell materials, including Ag, CdS, ZnS, and TiO2. The successful formation of complete and smooth shell was examined with a novel light scattering intensity decay method and also by direct observation of scanning electron microscope and transmission electron microscope images.
Synthesis, Characterization, and Optical Properties of Pristine and Doped Yttrium Aluminum Garnet Nanopowders
Authors: Wang Yiguang , Zhang Ligong , Fan Yi , Luo Jinsong , McCready David E. , Wang Chongmin , An Linan
Published: February 2005

Pristine, Si‐doped, and Si/Nd‐codoped yttrium aluminum garnet (YAG) nanoparticles were synthesized by pyrolysis of complex compounds of aluminum and yttrium with triethanolamine. It was found that the coexistence of Si4+ and Nd3+ increased the solubility of both ions and promoted the formation of YAG phase. Single‐phase, nanocrystalline Si/Nd:YAG powders were obtained at calcination temperatures as low as 920°C. The optical behavior of the Si/Nd:YAG nanopowders was similar to that of single‐crystal Nd:YAG.
The Effect of Sr Substitution on Phase Formation and Magnetic Properties of Y‐type Hexagonal Ferrite
Authors: Bai Yang , Zhou Ji , Gui Zhilun , Li Longtu
Published: February 2005

The effect of Sr substitution in Y‐type hexagonal ferrite of composition Ba2–xSrxZn1.2Co0.4Cu0.4Fe12O22 was investigated. x varied from 0 to 2.0. The samples were prepared by the solid‐state reaction method. Phase formation was characterized by powder X‐ray diffraction. The microstructure was observed via scanning electron microscopy and magnetic properties were measured using an impedance analyzer and vibrating sample magnetometer (VSM). Results reveal that Sr2+ cannot completely substitute for Ba2+ ions in Y‐type hexagonal ferrites. The maximum solubility of Sr in the Y‐type ferrite is about 1.8. With Sr above 1.8, the lattice mismatch induced by the difference in ionic radii of Ba2+ and Sr2+ ions prevents the formation of Y‐type phase. While Sr content is equal or less than 1.0, the magnetic properties may be enhanced. As x>1.0, because of the internal stress resulting from the lattice mismatch, demagnetizing field significantly degrades initial permeability.
Polygonal Single‐Crystal Aluminum Borate Microtubes
Authors: Yang Weiyou , Xie Zhipeng , Ma Jingtao , Miao Hezhuo , Luo Jiesheng , Zhang Ligong , An Linan
Published: February 2005

In this paper we report the synthesis of polygonal single‐crystal aluminum borate microtubes by the direct calcination of alumina and boron nitride powders in air. The diameters of the obtained tubes vary between 1 and 15 μm; and the lengths are in the range of tens to hundreds of micrometers. The cross‐sections of the microtubes are in the shapes of rectangular, hexagonal, and quasi‐circle. A new solid–liquid–solid growth mechanism was proposed to account for the formation of microtubes instead of whiskers.
Kinetics of Mullite Grain Growth in Alumino Silicate Fibers
Authors: Schmücker Martin , Schneider Hartmut , Mauer Thomas , Clauß Bernd
Published: February 2005

Grain growth kinetics of mullite in laboratory‐produced and commercial (3M‐Nextel 720) alumino silicate fibers was analyzed in the temperature range between 1500° and 1700°C. The lab fibers consist of mullite plus traces of α‐alumina, while the phase content of the commercial Nextel 720 fibers is about 60 wt% mullite plus 40 wt%α‐alumina. The temperature‐induced grain coarsening of mullite follows the empirical law D1/n−D01/n=kt. Two different temperature regimes can be distinguished with respect to the grain growth exponents: above 1600°C the growth exponent is ≈1/3, while below 1600°C the growth exponent of ≈1/12 is exceptionally low. Laboratory‐produced and commercial fibers show almost identical mullite grain growth kinetics.
Effect of Isothermal Heat Treatment on Plasma‐Sprayed Yttria‐Stabilized Zirconia Studied by Impedance Spectroscopy
Authors: Anderson Piers S. , Wang Xin , Xiao Ping
Published: February 2005

Impedance spectroscopy (IS) was used to measure electrical responses of various plasma‐sprayed yttria‐stabilized zirconia (YSZ) materials. Coupled with X‐ray diffraction, scanning electron microscopy, and microindentation, the effects of heat treatment on microstructure, phase composition, and mechanical properties were investigated. Results showed electrical responses for YSZ grains and grain boundaries varied with size and position of silver electrodes used. YSZ heat treated at 950°C showed micro‐crack closure: at 1150°C, microstructural and phase changes, and at 1450°C, formation of a resistive monoclinic phase. The aim was to develop that IS as a non‐destructive tool for thermal barrier coatings; results suggest IS can monitor microstructural and phase changes in YSZ under certain conditions.
Quantitative Electron Microscopy of (Bi,Pb)2Sr2Ca2Cu3O10+δ/Ag Multifilament Tapes During Initial Stages of Annealing
Authors: Bals Sara , Verbeeck Jo , Van Tendeloo Gustaaf , Liu Yi‐Lin , Grivel Jean‐Claude
Published: February 2005

The microstructural and compositional evolution during initial annealing of a superconducting (Bi,Pb)2Sr2Ca2Cu3O10+δ/Ag tape is studied using quantitative transmission electron microscopy. Special attention is devoted to the occurrence of Pb‐rich liquids, which are crucial for the Bi2Sr2CaCu2O8+δ to (Bi,Pb)2Sr2Ca2Cu3O10+δ transformation. Ca and/or Pb‐rich (Bi,Pb)2Sr2CaCu2O8+δ grains dissolve into a liquid, which reacts with Ca‐rich phases to increase the liquid's Ca‐content. This leads to (Bi,Pb)2Sr2Ca2Cu3O10+δ formation. Apparently, a Ca/Sr ratio of around 1 is sufficient to keep (Bi,Pb)2Sr2Ca2Cu3O10+δ nucleation going. It is confirmed that Ag particles are transported from the Ag‐sheath into the oxide core by the liquid and not by mechanical treatment of the tape.
(Pr, Co, Nb)‐Doped SnO2 Varistor Ceramics
Authors: Wang Jin‐Feng , Su Wen‐Bin , Chen Hong‐Cun , Wang Wen‐Xin , Zang Guo‐Zhong , Li Chang‐Peng , Bodde Sara
Published: February 2005

The effect on microstructure and electrical properties of (Co, Nb)‐doped SnO2 varistors upon the addition of Pr2O3 was investigated by scanning electron microscopy and by determining I–V, ɛ–f, and R–f relations. The threshold electric field of the SnO2‐based varistors increased significantly from 850 to 2280 V/mm, and the relative dielectric constants of the SnO2‐based varistors decreased greatly from 784 to 280 as Pr2O3 concentration was increased up to 0.3 mol%. The significant decrease of the SnO2 grain size, from 4.50 to 1.76 μm with increasing Pr2O3 concentration over the range of 0–0.3 mol%, is the origin for the increase in the threshold voltage and decrease of the dielectric constants. The grain size reduction is attributed to the segregation of Pr2O3 at grain boundaries hindering the SnO2 grains from conglomerating into large particles. Varistors were found to have a superhigh threshold voltage and a comparatively large nonlinear coefficient α. For 0.15 mol% Pr2O3‐doped sample, threshold electric field and nonlinear coefficient α were measured to be 1540 V/mm and 61, and for 0.3 mol% Pr2O3‐doped sample, V and α were 2150 V/mm and 42, respectively. Superhigh threshold voltage and large nonlinear coefficient α qualify the Pr‐doped SnO2 varistor as an excellent candidate for a high voltage protection system.
Ferroelectric 90° Domain Evaluation in Tetragonal Pb(Mg1/3Nb2/3)O3–PbTiO3 Ceramics
Authors: Zhang Xiaowen , Lei Chao , Chen Kepi
Published: February 2005

The domain structure of ferroelectrics changes during poling has a direct influence on the macroscopic properties of the materials. The intensity variation of the different X‐ray diffraction (XRD) pattern profiles was used to identify the percentage of 90° domain reorientation in the tetragonal phase of Pb(Mg1/3Nb2/3)O3–PbTiO3 (PMN–PT) ceramics after poling. The results are consistent with the change of piezoelectric properties. In addition, by using XRD patterns, a spatial distribution of polarization in a well‐poled 0.62PMN–0.38PT ceramics has been determined and was found to be best described by the Cauchy function W00l (ϕ)=1/(1+0.023ϕ2).
Intravitreal triamcinolone for macular oedema: efficacy in relation to aetiology
Authors: Sørensen Torben Lykke , Haamann Per , Villumsen Jørgen , Larsen Michael
Published: February 2005

Purpose:  To evaluate the efficacy and safety of intravitreal triamcinolone in patients with macular oedema of varying aetiology.

Methods:  Two milligrams of intravitreal triamcinolone acetonide was injected into 34 eyes with persistent macular oedema (17 eyes with macula oedema secondary to posterior uveitis, 13 eyes with diabetic retinopathy, and four with pseudophakic macular oedema). Best corrected visual acuity was determined and transfoveal optical coherence tomography performed after 1 week, 1 month, 3 months and 6 months.

Results:  Treatment improved visual acuity and subjective visual quality, and reduced foveal thickness in eyes with posterior uveitis and eyes with macular oedema secondary to diabetic retinopathy. Eyes treated for pseudophakic cystoid macular oedema demonstrated no improvement. A total of 32% of patients experienced a significant post‐injection increase in intraocular pressure. Endophthalmitis, rhegmatogenous retinal detachment and cataract were absent.

Conclusion:  Intravitreal triamcinolone appears to induce marked a improvement in macular oedema secondary to non‐infectious uveitis and diabetic retinopathy.
Effects of interleukin‐1β and dexamethasone on the expression of matrix metalloprotease mRNA by trabecular cells exposed to elevated hydrostatic pressure
Authors: Ehrich Dirk , Tripathi Brenda , Tripathi Ramesh , Duncker Gernot
Published: February 2005

Purpose:  We investigated the effects of interleukin‐1β (Il‐1β) and dexamethasone (Dex) on the expression of matrix metalloprotease‐1, ‐2, ‐3 and ‐14 (membrane type‐1 MMP−MT1‐MMP) as well as tissue inhibitors of matrix metalloproteases (TIMP‐1 and ‐2) mRNA by trabecular cells exposed not only to normal, but also to elevated levels of hydrostatic pressure.

Methods:  Confluent primary cultures of porcine trabecular cells were incubated in a serum‐free medium (SFM) as controls, or in SFM containing either 10 ng/ml Il‐1β or 10 nm Dex and exposed to pressures of 15 mmHg or 50 mmHg (corresponding to normal and high intraocular pressure, respectively) in specially designed pressure chambers. After 72 hours, total RNA was extracted from the harvested cells, reverse transcribed and amplified using primers specific to MMP‐1, ‐2, ‐3 and ‐14, and TIMP‐1 and ‐2.

Results:  The most significant changes were detected in the levels of MMP‐3 mRNA in control cells (2.4‐fold increase), of TIMP‐1 and ‐2 mRNA in cells treated with Il‐1β (2.6‐fold increase) and of MMP‐3 mRNA in cells treated with Dex (3.5‐fold increase) exposed to 50 mmHg pressure.

Conclusion:  Because MMP‐3 (stromelysin) mRNA showed the highest upregulation, our findings suggest that trabecular cells preferentially degrade and turn over the proteoglycan components of the extracellular matrix in response to short‐term exposure to increased hydrostatic pressure with and without Dex as a homeostatic mechanism.
Serous macular detachment in diabetic cystoid macular oedema
Authors: Ozdemir Hakan , Karacorlu Murat , Karacorlu Serra
Published: February 2005

Purpose:  To define serous macular detachment in patients with diabetic cystoid macular oedema (CME).

Methods:  This study involved 78 eyes of 58 patients with diabetic CME. The patients underwent complete ophthalmic examination, fluorescein angiography and optical coherence tomography (OCT). Eyes with epiretinal membrane or vitreo‐macular traction were not included in the study. Optical coherence tomography‐3 was used in all patients and fundi were scanned on the horizontal, vertical and four oblique planes through the centre of the fovea.

Results:  In all cases the increased thickness of the retina was related primarily to the hyporeflective intraretinal cavities. With OCT, 24 of 78 eyes (31%) had serous macular detachment as shown by retinal elevation over a non‐reflective cavity with minimal shadowing of the underlying tissues. Fluorescein angiography did not show serous macular detachment in any patient.

Conclusions:  Our data showed that the incidence of serous macular detachment in diabetic CME was much higher than previously reported. Optical coherence tomography‐3 allows an in vivo cross‐sectional observation of very subtle serous macular detachment that is difficult to diagnose at the slit‐lamp or by fluorescein angiography in patients with diabetic CME.
Recurrent secondary frosted branch angiitis after toxoplasmosis vasculitis
Authors: Oh Jaeryung , Huh Kuhl , Kim Seong Woo
Published: February 2005

Purpose:  To describe a case of recurrent frosted branch angiitis after treatment of ocular toxoplasmosis.

Methods:  In a 6‐year‐old boy, we found perivascular, creamy, patchy, retinal sheathing in both eyes without any focal necrotizing retinochoroiditis or scarring. IgM antibodies for toxoplasma gondii were also found. The patient was treated with antitoxoplasmosis medication and a systemic steroid.

Results:  Several years after treatment of the toxoplasmosis, frosted branch angiitis occurred twice without any retinal scarring or serological evidence of toxoplasmosis. After systemic steroid therapy, the angiitis improved without further complications.

Conclusions:  Toxoplasmic retinal vasculitis should be considered as a cause of frosted branch angiitis.
Intravitreal triamcinolone acetonide in patients with macular oedema due to central retinal vein occlusion
Authors: Krepler Katharina , Ergun Erdem , Sacu Stefan , Richter‐Müksch Sibylla , Wagner Julia , Stur Michael , Wedrich Andreas
Published: February 2005

Purpose:  To evaluate treatment of macular oedema due to central retinal vein occlusion (CRVO) with intravitreal triamcinolone acetonide.

Methods:  In a prospective case series, 13 patients with macular oedema due to non‐ischaemic CRVO received an intravitreal injection of 4 mg triamcinolone acetonide. Examination included assessment of best corrected visual acuity (BCVA) for distance and reading, measurement of intraocular pressure (IOP), fluorescein angiography and high resolution imaging by optical coherence tomography, preoperatively and 1 week, 1 month, 3, 6 and 9 months postoperatively.

Results:  Preoperative mean BCVA was 0.9 ± 0.4 for distance vision and 1.0 ± 0.3 for reading acuity, respectively. A significant improvement in distance VA (0.5 ± 0.3, p < 0.001) and reading acuity (0.7 ± 0.3, p = 0.03) was observed until 3 months and 6 months, respectively. Mean macular thickness was significantly reduced until 9 months postoperatively.

Conclusion:  Intravitreal injection of triamcinolone acetonide led to a significant improvement in mean VA in patients with macular oedema due to CRVO. However, the significant effect was not permanent and persisted for a maximum of 6 months.
The Groningen Longitudinal Glaucoma Study. I. Baseline sensitivity and specificity of the frequency doubling perimeter and the GDx nerve fibre analyser
Authors: Heeg Govert P. , Blanksma Lieuwe J. , Hardus Peter L. L. J. , Jansonius Nomdo M.
Published: February 2005

Purpose:  To describe the baseline data of a large cohort of patients included for follow‐up with perimetry using the frequency doubling technique (FDT) and with quantification of the retinal nerve fibre layer as assessed by GDx, and to calculate the sensitivity and specificity of both devices from these baseline data.

Methods:  Regular visitors to our glaucoma service were included. All subjects were followed for at least 4 years with FDT in full‐threshold mode, GDx and conventional perimetry. Patients were classified as having either glaucoma or suspect glaucoma, according to baseline perimetry results. In addition, a group of healthy subjects was recruited outside the hospital.

Results:  A total of 452 glaucoma patients, 423 glaucoma suspects and 237 healthy subjects were incorporated into the analyses. Sensitivities for both FDT and GDx were fixed at 0.90. For the group as a whole, the specificity was 0.81 for FDT, using number of depressed test‐points p < 0.01 in the total deviation probability plot with a cut‐off point > 1, and 0.78 for GDx, using the Number, with a cut‐off point > 29. The area under the receiver operating characteristic (ROC) curve was 0.92 for FDT and 0.94 for GDx. Of the subjects with suspect glaucoma, 75% showed normal FDT test results and 52% showed normal GDx results. Unlike FDT, GDx failed to detect some moderate/severe glaucoma cases.

Conclusions:  The performances of FDT and GDx are approximately equivalent in terms of sensitivity, specificity and area under the ROC curve. In glaucoma suspects, GDx in particular yielded a rather high percentage of positive test results. The majority of these positive test results are presumably false‐positive results rather than results indicating preperimetric glaucoma.
Epinephrine‐supplemented local anesthetics for ear and nose surgery: Clinical use without complications in more than 10,000 surgical procedures
Authors: Häfner Hans‐Martin , Röcken Manfred , Breuninger Helmut
Published: February 2005

Introduction: Local anesthetics supplemented with epinephrine are generally regarded as contraindicated for surgical procedures involving the fingers, toes, penis, outer ear and the tip of the nose [1], but epinephrine is essential if automated tumescence local anesthesia (Auto‐TLA) is used.

Materials and methods: Infiltration anesthesia supplemented with 1 : 200,000 epinephrine was used from 1985 – 1997 in our department, while Auto‐TLA supplemented with 1 : 1.000,000 epinephrine was introduced in 1997 for all surgical procedures involving the ear or nose. During this period, 10,201 patients underwent surgery at these locations.

In addition, dermal blood flow was analyzed by acral photoplethysmography (APPG) and laser Doppler flowmetry (LDF) in the right ear lobe of five normal volunteers and during epinephrine supplemented Auto‐TLA.

Results: Epinephrine‐induced complications were not observed in a single patient. Cosmetic skin flap surgery was performed in 4,953 of these patients. Even in patients with extended surgical procedures that took up to one to two hours and that included extensive skin flaps or skin grafts, we observed no increase in complications when compared to procedures performed either under general anesthesia or local anesthesia without epinephrine supplementation.

Measuring blood perfusion of the earlobe showed a 69 % reduction of LDF and a 42 % reduction of arterial inflow (APPG) immediately following anesthesia.

Conclusion: Epinephrine supplementation of local anesthetics does not block blood perfusion in the ear and did not induce organ, tissue or flap necrosis. Local anesthesia with epinephrine supplementation is therefore safe for acral areas such as the ear or nose. Despite the relatively small influence on blood perfusion, epinephrine supplementation results in a relatively bloodless operating field and longer effectiveness of local anesthesia. The relative absence of blood in the operating field of the ear and nose significantly reduces the duration of surgery and increases the healing rate, as less electrocautery is needed.
Multistable Equilibrium System Design Methodology and Demonstration
Authors: King Carey , Beaman Joseph J. , Sreenivasan S. V. , Campbell Matthew
Published: February 2005

Multistable equilibrium (MSE) systems are those which have multiple statically stable equilibrium positions. This paper gives an overview of what MSE systems are, how they can be used, and how they can be designed. We describe the numerical optimization and synthesis of an example system composed of energy storage elements from various energy domains including mechanical rotation, mechanical translation, and magnetism.
Increased Surface Fatigue Lives of Spur Gears by Application of a Coating
Authors: Krantz Timothy , Cooper Clark , Townsend Dennis , Hansen Bruce
Published: February 2005

Hard coatings have potential for increasing gear surface fatigue lives. Experiments were conducted using gears both with and without a metal-containing, carbon-based coating. The gears were case-carburized AISI 9310 steel spur gears. Some gears were provided with the coating by magnetron sputtering. Lives were evaluated by accelerated life tests. For uncoated gears, all of 15 tests resulted in fatigue failure before completing 275 million revolutions. For coated gears, 11 of the 14 tests were suspended with no fatigue failure after 275 million revolutions. The improved life owing to the coating, approximately a sixfold increase, was a statistically significant result.
Direct Singular Positions of 3RPS Parallel Manipulators
Authors: Liu C. H. , Cheng Shengchia
Published: February 2005

In this study a procedure to obtain direct singular positions of a 3RPS parallel manipulator is presented. If the heights of three spherical joints, denoted by d1n,d2n, and d3n respectively, are used as coordinate axes, then the workspace of the moving platform may be represented as an inclined solid cylinder in this coordinate system. The location of a point on the solid circular cylinder determines a configuration of the manipulator’s moving platform. The procedure to locate direct singular positions consists of two steps, the orientation of the moving platform is assumed first, from which the horizontal position of the moving platform may be obtained. Then in the second step the heights that make determinant of the Jacobian matrix vanish may always be determined. Results show that unless the moving platform is normal to the base, in which case there exist only one or two singular configurations, otherwise there are always three singular configurations corresponding to a moving platform’s orientation.
Dynamic Topology Optimization of Compliant Mechanisms and Piezoceramic Actuators
Authors: Maddisetty Hima , Frecker Mary
Published: February 2005

A topology optimization method is developed to design a piezoelectric ceramic actuator together with a compliant mechanism coupling structure for dynamic applications. The objective is to maximize the mechanical efficiency with a constraint on the capacitance of the piezoceramic actuator. Examples are presented to demonstrate the effect of considering dynamic behavior compared to static behavior and the effect of sizing the piezoceramic actuator on the optimal topology and the capacitance of the actuator element. Comparison studies are also presented to illustrate the effect of damping, external spring stiffness, and driving frequency. The optimal topology of the compliant mechanism is shown to be dependent on the driving frequency, the external spring stiffness, and whether the piezoelectric actuator element is considered design or nondesign. At high driving frequencies, it was found that the dynamically optimized structure is very near resonance.
A Novel Mechanism for Desulfation of Mucin: Identification and Cloning of a Mucin-Desulfating Glycosidase (Sulfoglycosidase) from
Prevotella
Strain RS2
Authors: Rho Jung-hyun , Wright Damian P. , Christie David L. , Clinch Keith , Furneaux Richard H. , Roberton Anthony M.
Published: February 2005

A novel enzyme which may be important in mucin degradation has been discovered in the mucin-utilizing anaerobe
Prevotella
strain RS2. This enzyme cleaves terminal 2-acetamido-2-deoxy-β-
d
-glucopyranoside 6-sulfate (6-SO
3
-GlcNAc) residues from sulfomucin and from the model substrate 4-nitrophenyl 2-acetamido-2-deoxy-β-
d
-glucopyranoside 6-sodium sulfate. The existence of this mucin-desulfating glycosidase (sulfoglycosidase) suggests an alternative mechanism by which this bacterium may desulfate sulfomucins, by glycosidic removal of a sulfated sugar from mucin oligosaccharide chains. Previously, mucin desulfation was thought to take place by the action of a specific desulfating enzyme, which then allowed glycosidases to remove desulfated sugar. Sulfate removal from sulfomucins is thought to be a rate-limiting step in mucin degradation by bacteria in the regions of the digestive tract with a significant bacterial flora. The sulfoglycosidase was induced by growth of the
Prevotella
strain on mucin and was purified 284-fold from periplasmic extracts. Tryptic digestion and sequencing of peptides from the 100-kDa protein enabled the sulfoglycosidase gene to be cloned and sequenced. Active recombinant enzyme was made in an
Escherichia coli
expression system. The sulfoglycosidase shows sequence similarity to hexosaminidases. The only other enzyme that has been shown to remove 6-SO
3
-GlcNAc from glycoside substrates is the human lysosomal enzyme β-
N
-acetylhexosaminidase A, point mutations in which cause the inheritable, lysosomal storage disorder Tay-Sachs disease. The human enzyme removes GlcNAc from glycoside substrates also, in contrast to the
Prevotella
enzyme, which acts on a nonsulfated substrate at a rate that is only 1% of the rate observed with a sulfated substrate.
Identification of Essential Amino Acid Residues of the NorM Na
+
/Multidrug Antiporter in
Vibrio parahaemolyticus
Authors: Otsuka Masato , Yasuda Makoto , Morita Yuji , Otsuka Chie , Tsuchiya Tomofusa , Omote Hiroshi , Moriyama Yoshinori
Published: February 2005

NorM is a member of the multidrug and toxic compound extrusion (MATE) family and functions as a Na
+
/multidrug antiporter in
Vibrio parahaemolyticus
, although the underlying mechanism of the Na
+
/multidrug antiport is unknown. Acidic amino acid residues Asp32, Glu251, and Asp367 in the transmembrane region of NorM are conserved in one of the clusters of the MATE family. In this study, we investigated the role(s) of acidic amino acid residues Asp32, Glu251, and Asp367 in the transmembrane region of NorM by site-directed mutagenesis. Wild-type NorM and mutant proteins with amino acid replacements D32E (D32 to E), D32N, D32K, E251D, E251Q, D367A, D367E, D367N, and D367K were expressed and localized in the inner membrane of
Escherichia coli
KAM32 cells, while the mutant proteins with D32A, E251A, and E251K were not. Compared to cells with wild-type NorM, cells with the mutant NorM protein exhibited reduced resistance to kanamycin, norfloxacin, and ethidium bromide, but the NorM D367E mutant was more resistant to ethidium bromide. The NorM mutant D32E, D32N, D32K, D367A, and D367K cells lost the ability to extrude ethidium ions, which was Na
+
dependent, and the ability to move Na
+
, which was evoked by ethidium bromide. Both E251D and D367N mutants decreased Na
+
-dependent extrusion of ethidium ions, but ethidium bromide-evoked movement of Na
+
was retained. In contrast, D367E caused increased transport of ethidium ions and Na
+
. These results suggest that Asp32, Glu251, and Asp367 are involved in the Na
+
-dependent drug transport process.
Type III Secretion of the
Salmonella
Effector Protein SopE Is Mediated via an N-Terminal Amino Acid Signal and Not an mRNA Sequence
Authors: Karavolos M. H. , Wilson M. , Henderson J. , Lee J. J. , Khan C. M. A.
Published: February 2005

Type III secretion systems (TTSS) are virulence-associated components of many gram-negative bacteria that translocate bacterial proteins directly from the bacterial cytoplasm into the host cell. The
Salmonella
translocated effector protein SopE has no consensus cleavable amino-terminal secretion sequence, and the mechanism leading to its secretion through the
Salmonella
pathogenicity island 1 (SPI-1) TTSS is still not fully understood. There is evidence from other bacteria which suggests that the TTSS signal may reside within the 5′ untranslated region (UTR) of the mRNA of secreted effectors. We investigated the role of the 5′ UTR in the SPI-1 TTSS-mediated secretion of SopE using promoter fusions and obtained data indicating that the mRNA sequence is not involved in the secretion process. To clarify the proteinaceous versus RNA nature of the signal, we constructed frameshift mutations in the amino-terminal region of SopE of
Salmonella enterica
serovar Typhimurium SL1344. Only constructs with the native amino acid sequence were secreted, highlighting the importance of the amino acid sequence versus the mRNA sequence for secretion. Additionally, we obtained frameshift mutation data suggesting that the first 15 amino acids are important for secretion of SopE independent of the presence of the chaperone binding site. These data shed light on the nature of the signal for SopE secretion and highlight the importance of the amino-terminal amino acids for correct targeting and secretion of SopE via the SPI-1-encoded TTSS during host cell invasion.
Experimental Identification and Quantification of Glucose Metabolism in Seven Bacterial Species
Authors: Fuhrer Tobias , Fischer Eliane , Sauer Uwe
Published: February 2005

The structurally conserved and ubiquitous pathways of central carbon metabolism provide building blocks and cofactors for the biosynthesis of cellular macromolecules. The relative uses of pathways and reactions, however, vary widely among species and depend upon conditions, and some are not used at all. Here we identify the network topology of glucose metabolism and its in vivo operation by quantification of intracellular carbon fluxes from
13
C tracer experiments. Specifically, we investigated
Agrobacterium tumefaciens
, two pseudomonads,
Sinorhizobium meliloti
,
Rhodobacter sphaeroides
,
Zymomonas mobilis
, and
Paracoccus versutus
, which grow on glucose as the sole carbon source, represent fundamentally different metabolic lifestyles (aerobic, anaerobic, photoheterotrophic, and chemoheterotrophic), and are phylogenetically distinct (firmicutes, γ-proteobacteria, and α-proteobacteria). Compared to those of the model bacteria
Escherichia coli
and
Bacillus subtilis
, metabolisms of the investigated species differed significantly in several respects: (i) the Entner-Doudoroff pathway was the almost exclusive catabolic route; (ii) the pentose phosphate pathway exhibited exclusively biosynthetic functions, in many cases also requiring flux through the nonoxidative branch; (iii) all aerobes exhibited fully respiratory metabolism without significant overflow metabolism; and (iv) all aerobes used the pyruvate bypass of the malate dehydrogenase reaction to a significant extent. Exclusively,
Pseudomonas fluorescens
converted most glucose extracellularly to gluconate and 2-ketogluconate. Overall, the results suggest that metabolic data from model species with extensive industrial and laboratory history are not representative of microbial metabolism, at least not quantitatively.
Genome-Wide Analysis of the General Stress Response Network inEscherichia coli: σS-Dependent Genes, Promoters, and Sigma Factor Selectivity
Authors: Weber Harald , Polen Tino , Heuveling Johanna , Wendisch Volker F. , Hengge Regine
Published: February 2005

The σS(or RpoS) subunit of RNA polymerase is the master regulator of the general stress response inEscherichia coli. While nearly absent in rapidly growing cells, σSis strongly induced during entry into stationary phase and/or many other stress conditions and is essential for the expression of multiple stress resistances. Genome-wide expression profiling data presented here indicate that up to 10% of theE. coligenes are under direct or indirect control of σSand that σSshould be considered a second vegetative sigma factor with a major impact not only on stress tolerance but on the entire cell physiology under nonoptimal growth conditions. This large data set allowed us to unequivocally identify a σSconsensus promoter in silico. Moreover, our results suggest that σS-dependent genes represent a regulatory network with complex internal control (as exemplified by the acid resistance genes). This network also exhibits extensive regulatory overlaps with other global regulons (e.g., the cyclic AMP receptor protein regulon). In addition, the global regulatory protein Lrp was found to affect σSand/or σ70selectivity of many promoters. These observations indicate that certain modules of the σS-dependent general stress response can be temporarily recruited by stress-specific regulons, which are controlled by other stress-responsive regulators that act together with σ70RNA polymerase. Thus, not only the expression of genes within a regulatory network but also the architecture of the network itself can be subject to regulation.
Cupin-Type Phosphoglucose Isomerases (Cupin-PGIs) Constitute a Novel Metal-Dependent PGI Family Representing a Convergent Line of PGI Evolution
Authors: Hansen Thomas , Schlichting Bettina , Felgendreher Martina , Schönheit Peter
Published: February 2005

Cupin-type phosphoglucose isomerases (cPGIs) were identified in some archaeal and bacterial genomes and the respective coding function of
cpgi
's from the euryarchaeota
Archaeoglobus fulgidus
and
Methanosarcina mazei
, as well as the bacteria
Salmonella enterica
serovar Typhimurium and
Ensifer meliloti
, was proven by functional overexpression. These cPGIs and the cPGIs from
Pyrococcus
and
Thermococcus
spp. represent the cPGI family and were compared with respect to kinetic, inhibitory, thermophilic, and metal-binding properties. cPGIs showed a high specificity for the substrates fructose-6-phosphate and glucose-6-phosphate and were inhibited by millimolar concentrations of sorbitol-6-phosphate, erythrose-4-phosphate, and 6-phosphogluconate. Treatment of cPGIs with EDTA resulted in a complete loss of catalytic activity, which could be regained by the addition of some divalent cations, most effectively by Fe
2+
and Ni
2+
, indicating a metal dependence of cPGI activity. The motifs TX
3
PX
3
GXEX
3
TXGHXHX
6-11
EXY and PPX
3
HX
3
N were deduced as the two signature patterns of the novel cPGI family. Phylogenetic analysis suggests lateral gene transfer for the bacterial cPGIs from euryarchaeota.
GenotypingMycobacterium ulceransandMycobacterium marinumby Using Mycobacterial Interspersed Repetitive Units
Authors: Stragier Pieter , Ablordey Anthony , Meyers Wayne M. , Portaels Françoise
Published: February 2005

A novel category of variable tandem repeats (VNTR) called mycobacterial interspersed repetitive units (MIRUs) has been identified forMycobacterium ulcerans(n= 39),M. marinum(n= 27), and one related organism. Fifteen MIRU loci were identified in the genome ofM. marinumand were used to genotypeM. ulcerans,M. marinum, and anM. marinum-like organism that is considered a possible missing link betweenM. marinumandM. ulcerans. Seven MIRU loci were polymorphic, and locus-specific PCRs for four of these loci differentiated sevenM. ulceransgenotypes, fourM. marinumgenotypes, and a unique genotype for the missing link organism. The sevenM. ulceransgenotypes were related to six different geographic origins of isolates. All isolates from West and Central Africa, including old and recent isolates, belonged to the same genotype, emphasizing the great spatiotemporal homogeneity among African isolates. Unlike theM. ulceransgenotypes, the fourM. marinumgenotypes could not be clearly related to the geographic origins of the isolates. According to MIRU-VNTR typing, allM. ulceransandM. marinumisolates of American origin were closely related, suggesting a common American ancestor for these two pathogenic species on the American continents. MIRU typing has significant potential value for discriminating between reoccurrence and reinfection forM. ulceransdisease.
Additional Determinants within
Escherichia coli
FNR Activating Region 1 and RNA Polymerase α Subunit Required for Transcription Activation
Authors: Weber K. Derek , Vincent Owen D. , Kiley Patricia J.
Published: February 2005

The global anaerobic regulator FNR is a DNA binding protein that activates transcription of genes required for anaerobic metabolism in
Escherichia coli
through interactions with RNA polymerase (RNAP). Alanine-scanning mutagenesis of FNR amino acid residues 181 to 193 of FNR was utilized to determine which amino acid side chains are required for transcription of both class II and class I promoters. In vivo assays of FNR function demonstrated that a core of residues (F181, R184, S187, and R189) was required for efficient activation of class II promoters, while at a class I promoter, FF(−61.5), only S187 and R189 were critical for FNR activation. Site-directed mutagenesis of positions 184, 187, and 189 revealed that the positive charge contributes to the function of the side chain at positions 184 and 189 while the serine hydroxyl is critical for the function of position 187. Subsequent analysis of the carboxy-terminal domain of the α subunit (αCTD) of RNAP, using an alanine library in single copy, revealed that in addition to previously characterized side chains (D305, R317, and L318), E286 and E288 contributed to FNR activation of both class II and class I promoters, suggesting that αCTD region 285 to 288 also participates in activation by FNR. In conclusion, this study demonstrates that multiple side chains within region 181 to 192 are required for FNR activation and the surface of αCTD required for FNR activation is more extensive than previously observed.
Comparative Study of Class 1 Integron andVibrio choleraeSuperintegron Integrase Activities
Authors: Biskri Latefa , Bouvier Marie , Guérout Anne-Marie , Boisnard Stéphanie , Mazel Didier
Published: February 2005

Superintegrons (SIs) and multiresistant integrons (MRIs) have two main structural differences: (i) the SI platform is sedentary, while the MRI platform is commonly associated with mobile DNA elements and (ii) the recombination sites (attC) of SI gene cassette clusters are highly homogeneous, while those of MRI cassette arrays are highly variable in length and sequence. In order to determine if the latter difference was correlated with a dissimilarity in the recombination activities, we conducted a comparative study of the integron integrases of the class 1 MRI (IntI1) and theVibrio choleraeSI (VchIntIA). We developed two assays that allowed us to independently measure the frequencies of cassette deletion and integration at the cognateattIsites. We demonstrated that the range ofattCsites efficiently recombined by VchIntIA is narrower than the range ofattCsites efficiently recombined by IntI1. Introduction of mutations into theV. choleraerepeats (VCRs), theattCsites of theV. choleraeSI cassettes, allowed us to map positions that affected the VchIntIA and IntI1 activities to different extents. Using a cointegration assay, we established that inE. coli,attI1-×-VCR recombination catalyzed by IntI1 was 2,600-fold more efficient thanattIVch-×-VCR recombination catalyzed by VchIntIA. We performed the same experiments inV. choleraeand established that theattIVch-×-VCR recombination catalyzed by VchIntIA was 2,000-fold greater than the recombination measured inE. coli. Taken together, our results indicate that in theV. choleraeSI, the substrate recognition and recombination reactions mediated by VchIntIA might differ from the class 1 MRI paradigm.
Evolution of Genomic Content in the Stepwise Emergence ofEscherichia coliO157:H7
Authors: Wick Lukas M. , Qi Weihong , Lacher David W. , Whittam Thomas S.
Published: February 2005

Genome comparisons have demonstrated that dramatic genetic change often underlies the emergence of new bacterial pathogens. Evolutionary analysis ofEscherichia coliO157:H7, a pathogen that has emerged as a worldwide public health threat in the past two decades, has posited that this toxin-producing pathogen evolved in a series of steps from O55:H7, a recent ancestor of a nontoxigenic pathogenic clone associated with infantile diarrhea. We used comparative genomic hybridization with 50-mer oligonucleotide microarrays containing probes from both pathogenic and nonpathogenic genomes to infer when genes were acquired and lost. Many ancillary virulence genes identified in the O157 genome were already present in an O55:H7-like progenitor, with 27 of 33 genomic islands of >5 kb and specific for O157:H7 (O islands) that were acquired intact before the split from this immediate ancestor. Most (85%) of variably absent or present genes are part of prophages or phage-like elements. Divergence in gene content among these closely related strains was ∼140 times greater than divergence at the nucleotide sequence level. A >100-kb region around the O-antigen gene cluster contained highly divergent sequences and also appears to be duplicated in its entirety in one lineage, suggesting that the whole region was cotransferred in the antigenic shift from O55 to O157. The β-glucuronidase-positive O157 variants, although phylogenetically closest to the Sakai strain, were divergent for multiple adherence factors. These observations suggest that, in addition to gains and losses of phage elements, O157:H7 genomes are rapidly diverging and radiating into new niches as the pathogen disseminates.
Crystal Structures of the BlaI Repressor from
Staphylococcus aureus
and Its Complex with DNA: Insights into Transcriptional Regulation of the
bla
and
mec
Operons
Authors: Safo Martin K. , Zhao Qixun , Ko Tzu-Ping , Musayev Faik N. , Robinson Howard , Scarsdale Neel , Wang Andrew H.-J. , Archer Gordon L.
Published: February 2005

The 14-kDa BlaI protein represses the transcription of
blaZ
, the gene encoding β-lactamase. It is homologous to MecI, which regulates the expression of
mecA
, the gene encoding the penicillin binding protein PBP2a. These genes mediate resistance to β-lactam antibiotics in staphylococci. Both repressors can bind either
bla
or
mec
DNA promoter-operator sequences. Regulated resistance genes are activated via receptor-mediated cleavage of the repressors. Cleavage is induced when β-lactam antibiotics bind the extramembrane sensor of the sensor-transducer signaling molecules, BlaR1 or MecR1. The crystal structures of BlaI from
Staphylococcus aureus
, both in free form and in complex with 32 bp of DNA of the
mec
operator, have been determined to 2.0- and 2.7-Å resolutions, respectively. The structure of MecI, also in free form and in complex with the
bla
operator, has been previously reported. Both repressors form homodimers, with each monomer composed of an N-terminal DNA binding domain of winged helix-turn-helix topology and a C-terminal dimerization domain. The structure of BlaI in complex with the
mec
operator shows a protein-DNA interface that is conserved between both
mec
and
bla
targets. The recognition helix α3 interacts specifically with the conserved TACA/TGTA DNA binding motif. BlaI and, probably, MecI dimers bind to opposite faces of the
mec
DNA double helix in an up-and-down arrangement, whereas MecI and, probably, BlaI dimers bind to the same DNA face of
bla
promoter-operator DNA. This is due to the different spacing of
mec
and
bla
DNA binding sites. Furthermore, the flexibility of the dimeric proteins may make the C-terminal proteolytic cleavage site more accessible when the repressors are bound to DNA than when they are in solution, suggesting that the induction cascade involves bound rather than free repressor.
Response by trout populations in alpine lakes to an experimental halt to stocking
Authors: Armstrong Trip W , Knapp Roland A
Published: February 2005

Trout are often stocked into alpine lakes based on the generally untested assumption that resident populations would go extinct without stocking. The objectives of our study were to estimate the proportion of currently or formerly stocked alpine lakes in the Sierra Nevada, California, containing naturally reproducing trout populations (Oncorhynchus mykiss, Oncorhynchus mykiss aguabonita, Oncorhynchus clarki henshawi), identify characteristics of lakes associated with successful reproduction, and quantify the effects of stocking termination on trout density and individual growth rates in reproducing populations. We surveyed trout populations in 95 lakes in the John Muir Wilderness before and after a 4- to 8-year stocking halt and in 84 lakes in SequoiaKings Canyon National Park after a ≥20-year stocking hiatus. Based on recruitment during the no-stocking period, 70% and 68% of study lakes in the John Muir Wilderness and Sequoia-Kings Canyon National Park, respectively, contained reproducing populations. Results indicated that lakes with >2.1 m2 of spawning habitat and at elevations <3520 m nearly always showed evidence of reproduction. For reproducing populations, stocking termination did not result in significant changes in population density, but may have increased individual growth rates. We conclude that most trout stocking in Sierra Nevada alpine lakes could be permanently halted with only minimal impact on the recreational fishery.
Changes in the crustacean communities of Lakes Michigan, Huron, and Erie following the invasion of the predatory cladoceranBythotrepheslongimanus
Authors: Barbiero Richard P , Tuchman Marc L
Published: February 2005

The crustacean zooplankton communities in Lakes Michigan and Huron and the central and eastern basins of Lake Erie have shown substantial, persistent changes since the invasion of the predatory cladoceran Bythotrephes in the mid-1980s. A number of cladoceran species have declined dramatically since the invasion, including Eubosmina coregoni, Holopedium gibberum, Daphnia retrocurva, Daphnia pulicaria, and Leptodora kindti, and overall species richness has decreased as a result. Copepods have been relatively unaffected, with the notable exception of Meso cyclops edax, which has virtually disappeared from the lakes. These species shifts have for the most part been consistent and equally pronounced across all three lakes. Responses of crustacean species to the Bythotrephes invasion do not appear to be solely a consequence of size, and it is likely that other factors, e.g., morphology, vertical distribution, or escape responses, are important determinants of vulnerability to predation. Our results indicate that invertebrate predators in general, and invasive ones in particular, can have pronounced, lasting effects on zooplankton community structure.