bluuebunny/crossref_metadata_2025 huggingface dataset:
Nanoprotrusion model for field emission from integrated microtips
Authors: Purcell S. T. , Binh Vu Thien , Baptist R.
Published: July 2002

A study of the field emission from Spindt-type metallic integrated single microtips and arrays of microtips has been carried out. Measurements of field emission patterns, I/V curves, current stability, energy distributions against both applied voltage and angle of emission, and high frequency noise were made throughout the seasoning process. These measurements show that the emitters are not conventional metal emitters but undergo an evolution that depends on their past history such as operating time and exposure to ambient gases. In particular, the energy distributions showed three distinct behaviors; (1) a large and even dominating part of the total emission current could come from low-energy secondary electrons generated by collisions of the emitted electrons with the grid, particularly in the beginning of the seasoning process. After seasoning, the electrons were predominantly emitted from either, (2) multiple discrete bands nearer to the Fermi level that displace with applied voltage, or, (3) from the standard conduction band at the Fermi level. This invalidates the strict use of simple Fowler–Nordheim theory for analyzing the field emission from these emitters. A model based essentially on the field emission from nanoprotrusions all along the surface of the conical shank of the deposited tip is proposed which is consistent with all the experimental results. The spatial distribution of the noncontrolled nanoprotrusions evolves from the shanks to the microtip apex regions during the seasoning process.
Detection mechanism of an optical evanescent field using a noncontact mode atomic force microscope with a frequency modulation detection method
Authors: Abe Masayuki , Uchihashi Takayuki , Ohta Masahiro , Ueyama Hitoshi , Sugawara Yasuhiro , Morita Seizo
Published: July 2002

By using the noncontact atomic force microscope with a frequency modulation detection method, the force gradient induced by the optical evanescent field was detected in a high vacuum. We succeeded in measuring the exponential distance dependence of the force gradient induced by the optical evanescent field. Furthermore, we investigated the incident beam intensity and bias voltage dependence of the force gradient induced by the optical evanescent field. We confirmed that the detection mechanism is not photothermal effect but the surface photovoltage effect.
Fabrication of magnetic submicron-wire channels for the investigation of magnetization reversal
Authors: Chen Y. , Kottler V. , Carcenac F. , René J. F. , Essaidi N. , Chappert C. , Launois H.
Published: July 2002

We have demonstrated a process for fabricating magnetic structures with feature sizes ranging from 50 nm to several hundred microns using high-resolution electron beam lithography with 50 keV electrons followed by Ti evaporation and liftoff. The Ti was used as an etch mask for ion milling and then removed by SF6 reactive ion etching. A variety of magnetic fine structures were fabricated on (CoNi/Pt)6 multilayer substrates. In particular, submicron-wire channels, which connect a number of microscopic squares to a large reservoir area, were obtained. The magnetization reversal processes in submicron wire channels were studied with a conventional magneto-optical Kerr microscope. We found that the switching field for magnetization and the reversal behavior of the connected microscopic squares strongly depend on the channel wire width, thereby providing a way to study domain wall motion in a subwavelength scale with diffraction limited methods.
Monolithic nanofluid sieving structures for DNA manipulation
Authors: Turner S. W. , Perez A. M. , Lopez A. , Craighead H. G.
Published: July 2002

A new technique for fabricating two-dimensional artificial gels for DNA electrophoresis is presented. The technique differs from previous approaches in that the entire device is fabricated as a monolithic unit using exclusively planar processing techniques adapted from semiconductor electronics fabrication. The height of the fluid gap between the dielectric floor and ceiling is determined by the thickness of a sacrificial layer which is removed by a wet chemical etch. This allows precise control and excellent uniformity of the gap over an entire silicon wafer. Floor-to-ceiling height control better than 5 nm has been demonstrated over a 1.5 cm device. Electron beam lithography is used to define a square array of 100 nm obstructions in the sacrificial layer. Chemical vapor deposition silicon nitride is applied over the sacrificial layer. Reactive ion etching (RIE) is used to create access holes in the nitride layer, so that the sacrificial layer can be removed with a wet chemical etch. After the wet etch, the access holes are resealed with very low temperature oxide (VLTO) silicon dioxide. Finally, loading widows are opened with RIE at both ends of the device so that DNA in aqueous solution can be introduced and its motion under the influence of an electric field can be observed. The DNA molecules are labeled with a fluorescent dye and observed through the dielectric top layers with an optical microscope. The electrophoretic mobility is measured for two different DNA chain lengths, 43 and 7.2 kbase. The velocity for both DNA lengths is reported for an applied potential between 2 and 20 V over the 15 mm device. At some voltages the velocities differed by nearly a factor of 2.
Elucidation of field emission characteristics of phosphorous-doped diamond films
Authors: Kuriyama Kenji , Kimura Chiharu , Koizumi Satoshi , Kamo Mutsukazu , Sugino Takashi
Published: July 2002

Field emission characteristics are investigated using phosphorous (P)-doped polycrystalline and homoepitaxial diamond films. A significant variation in turn-on voltage is observed for different cathode metals on the polycrystalline and homoepitaxial diamond films. This experimental fact shows that the internal electron emission at the diamond/cathode contact dominates the field emission characteristics. We also investigate the temperature dependence of the field emission characteristics for P-doped polycrystalline diamond films in comparison with those of the undoped one. The turn-on voltage decreases with increasing temperature for the P-doped diamond film. On the other hand, no variation in the turn-on voltage occurs for the undoped diamond film. This behavior of the field emission characteristics is well explained by the thermionic field emission theory combined with the temperature dependence of the ionized donor concentration.
Patterning of planar magnetic nanostructures by ion irradiation
Authors: Devolder T. , Chappert C. , Chen Y. , Cambril E. , Launois H. , Bernas H. , Ferré J. , Jamet J. P.
Published: July 2002

We have investigated the ultrahigh resolution of light ion irradiation induced magnetic patterning. High aspect ratio silica masks on Co/Pt multilayers were obtained by e-beam lithography and reactive ion etching with feature sizes down to 30 nm for lines and 20 nm for dots. He+ ion irradiation of the magnetic layers through these masks was used to pattern the magnetic properties. We chose irradiation irradiation conditions such that patterned areas had a coercive force three times lower than the protected areas. After mask removal, we obtained high resolution and high density, planar magnetic nanostructures. Magnetization reversal processes in the line patterns are analyzed with numerical simulations. We suggest that this irradiation induced magnetic patterning technique is promising for both present magneto-optical storage devices and future near field recording.
Extreme ultraviolet and x-ray resist: Comparison study
Authors: He D. , Solak H. , Li W. , Cerrina F.
Published: July 2002

Extreme ultraviolet lithography (EUVL) is a next generation lithography technology aimed at critical dimension of 70 nm and below. It is important to characterize the imaging properties of the resist in this wavelength range. In this article we present results of EUVL and x-ray lithography studies of the dissolution properties of modern resists (SAL605, UV6, and PMMA) in terms of development properties and surface roughness. The exposure response curves and surface roughness were measured for each resist for EUV and x-ray exposures. We find that the absorbed dose can be used as a common parameter to compare exposure response curves from EUV and x ray on the same scale. We also find that the surface roughness is a specific property of each resist system. The relative importance of shot noise effects on roughness is discussed.
Investigation of field emission characteristics for Si-base materials: Titanium silicide, poly-Si, and single crystal Si
Authors: Lim Moo-Sup , Park Cheol-Min , Han Min-Koo , Choi Yearn-Ik
Published: July 2002

We have fabricated poly-Si, Si, and Ti silicide field emitter arrays employing in situ vacuum encapsulated lateral field emitter structures and investigated the field emission characteristics such as turn-on voltage, emission current density, and the stability of the emission current. Although poly-Si and Si emitters have almost identical turn-on voltages, Si emitters have a sharper turn on than poly-Si emitters due to their uniform surface. The current densities of poly-Si and Si emitters are 0.47 and 0.43 μ A/tip, respectively, at an anode to cathode voltage of 90 V. The turn-on voltage and current density of Ti silicide emitters are about 31 V and 1.81 μ A/tip at VAK of 90 V. The data of the normalized current fluctuations indicate that the Ti silicide emitters have the most stable current. The experimental results show that Ti silicide is the most promising among these three materials due to its low work function, uniform surface, and stable characteristics.
Fabrication and electron transport in multilayer silicon-insulator-silicon nanopillars
Authors: Pooley David M. , Ahmed Haroon , Lloyd Neil S.
Published: July 2002

We report the fabrication of sub-0.1-μm-diam silicon nanopillars with a polycrystalline silicon and silicon nitride multilayer structure. The polycrystalline silicon layers, which are 20 nm thick and are separated by 2–3 nm-thick silicon nitride tunnel barriers, make potentially useful structures for the observation of single-electron charging effects. Measurements of electron transport at 4.2 K between contacts at the top and bottom of the pillars show a zero current region and current steps attributed to Coulomb blockade.
Phonon Density of States of Iron up to 153 Gigapascals
Authors: Mao H. K. , Xu J. , Struzhkin V. V. , Shu J. , Hemley R. J. , Sturhahn W. , Hu M. Y. , Alp E. E. , Vocadlo L. , Alfè D. , Price G. D. , Gillan M. J. , Schwoerer-Böhning M. , Häusermann D. , Eng P. , Shen G. , Giefers H. , Lübbers R. , Wortmann G.
Published: July 2002

We report phonon densities of states (DOS) of iron measured by nuclear resonant inelastic x-ray scattering to 153 gigapascals and calculated from ab initio theory. Qualitatively, they are in agreement, but the theory predicts density at higher energies. From the DOS, we derive elastic and thermodynamic parameters of iron, including shear modulus, compressional and shear velocities, heat capacity, entropy, kinetic energy, zero-point energy, and Debye temperature. In comparison to the compressional and shear velocities from the preliminary reference Earth model (PREM) seismic model, our results suggest that Earth's inner core has a mean atomic number equal to or higher than pure iron, which is consistent with an iron-nickel alloy.
Intracellular Anions as the Voltage Sensor of Prestin, the Outer Hair Cell Motor Protein
Authors: Oliver Dominik , He David Z. Z. , Klöcker Nikolaj , Ludwig Jost , Schulte Uwe , Waldegger Siegfried , Ruppersberg J. P. , Dallos Peter , Fakler Bernd
Published: July 2002

Outer hair cells (OHCs) of the mammalian cochlea actively change their cell length in response to changes in membrane potential. This electromotility, thought to be the basis of cochlear amplification, is mediated by a voltage-sensitive motor molecule recently identified as the membrane protein prestin. Here, we show that voltage sensitivity is conferred to prestin by the intracellular anions chloride and bicarbonate. Removal of these anions abolished fast voltage-dependent motility, as well as the characteristic nonlinear charge movement (“gating currents”) driving the underlying structural rearrangements of the protein. The results support a model in which anions act as extrinsic voltage sensors, which bind to the prestin molecule and thus trigger the conformational changes required for motility of OHCs.
Sending Sound to the Brain
Authors: Rauschecker J. P. , Shannon R. V.
Published: July 2002

The cochlear implant, a microelectrode array that directly stimulates the auditory nerve, has greatly benefited many individuals with profound deafness. Deaf patients without an intact auditory nerve may be helped by the next generation of auditory prostheses: surface or penetrating auditory brainstem implants that bypass the auditory nerve and directly stimulate auditory processing centers in the brainstem.
Ordering of Quantum Dots Using Genetically Engineered Viruses
Authors: Lee Seung-Wuk , Mao Chuanbin , Flynn Christine E. , Belcher Angela M.
Published: July 2002

A liquid crystal system was used for the fabrication of a highly ordered composite material from genetically engineered M13 bacteriophage and zinc sulfide (ZnS) nanocrystals. The bacteriophage, which formed the basis of the self-ordering system, were selected to have a specific recognition moiety for ZnS crystal surfaces. The bacteriophage were coupled with ZnS solution precursors and spontaneously evolved a self-supporting hybrid film material that was ordered at the nanoscale and at the micrometer scale into
∼
72-micrometer domains, which were continuous over a centimeter length scale. In addition, suspensions were prepared in which the lyotropic liquid crystalline phase behavior of the hybrid material was controlled by solvent concentration and by the use of a magnetic field.
Joining the Two Domains of a Group I Ribozyme to Form the Catalytic Core
Authors: Tanner Michael A. , Cech Thomas R.
Published: July 2002

Self-splicing group I introns, like other large catalytic RNAs, contain structural domains. Although the crystal structure of one of these domains has been determined by x-ray analysis, its connection to the other major domain that contains the guanosine-binding site has not been known. Site-directed mutagenesis and kinetic analysis of RNA splicing were used to identify a base triple in the conserved core of both a cyanobacterial (
Anabaena
) and a eukaryotic (
Tetrahymena
) group I intron. This long-range interaction connects a sequence adjacent to the guanosine-binding site with the domain implicated in coordinating the 5′ splice site helix, and it thereby contributes to formation of the active site. The resulting five-strand junction, in which a short helix forms base triples with three separate strands in the
Tetrahymena
intron, reveals exceptionally dense packing of RNA.
Five-Coordinate Hydrogen: Neutron Diffraction Analysis of the Hydrido Cluster Complex [H
2
Rh
13
(CO)
24
]
3−
Authors: Bau Robert , Drabnis Mary H. , Garlaschelli Luigi , Klooster Wim T. , Xie Zuowei , Koetzle Thomas F. , Martinengo Secondo
Published: July 2002

Pentacoordinate hydrogen atoms were identified by single-crystal neutron diffraction analysis of [N(CH
3
)
4
]
3
[H
2
Rh
13
(CO)
24
]. The hydrogen atoms are located in square pyramidal cavities of the Rh
13
cluster, in positions almost coplanar with the Rh
4
faces on the surface of the cluster. They are slightly displaced inward, toward the central rhodium atom of the cluster, with average H-Rh(central) and H-Rh(surface) distances of 1.84(2) and 1.97(2) angstroms, respectively. This result shows that hydrogen, which normally forms only one bond, can be attached to five other atoms simultaneously in a large metal cluster.
Sizes and Ages of Seamounts Using Remote Sensing: Implications for Intraplate Volcanism
Authors: Wessel Paul
Published: July 2002

Satellite altimetry was used to identify and characterize Pacific intraplate seamounts. The gravimetric amplitudes of seamounts appear to be related to the age difference between the sea floor and seamounts; by inverting this relation, pseudo ages can be obtained for undated seamounts. These pseudo ages imply that excursions in seamount volcanism generally correlate with times of formation of large oceanic plateaus.
Requirement for DCP-1 Caspase During
Drosophila
Oogenesis
Authors: McCall Kimberly , Steller Hermann
Published: July 2002

Caspases, a class of cysteine proteases, are an essential component of the apoptotic cell death program. During
Drosophila
oogenesis, nurse cells transfer their cytoplasmic contents to developing oocytes and then die. Loss of function for the
dcp-1
gene, which encodes a caspase, caused female sterility by inhibiting this transfer.
dcp-1
−
nurse cells were defective in the cytoskeletal reorganization and nuclear breakdown that normally accompany this process. The
dcp-1
−
phenotype suggests that the cytoskeletal and nuclear events in the nurse cells make use of the machinery normally associated with apoptosis and that apoptosis of the nurse cells is a necessary event for oocyte development.
Age and Origin of Carlsbad Cavern and Related Caves from
40
Ar/
39
Ar of Alunite
Authors: Polyak Victor J. , McIntosh William C. , Güven Necip , Provencio Paula
Published: July 2002

40
Ar/
39
Ar dating of fine-grained alunite that formed during cave genesis provides ages of formation for the Big Room level of Carlsbad Cavern [4.0 to 3.9 million years ago (Ma)], the upper level of Lechuguilla Cave (6.0 to 5.7 Ma), and three other hypogene caves (11.3 to 6.0 Ma) in the Guadalupe Mountains of New Mexico. Alunite ages increase and are strongly correlative with cave elevations, which indicates an 1100-meter decline in the water table, apparently related to tectonic uplift and tilting, from 11.3 Ma to the present.
40
Ar/
39
Ar dating studies of the hypogene caves have the potential to help resolve late Cenozoic climatic, speleologic, and tectonic questions.
Ordering of Ruthenium Cluster Carbonyls in Mesoporous Silica
Authors: Zhou Wuzong , Thomas John Meurig , Shephard Douglas S. , Johnson Brian F. G. , Ozkaya Don , Maschmeyer Thomas , Bell Robert G. , Ge Qingfeng
Published: July 2002

The anionic ruthenium cluster carbonylates [Ru
6
C(CO)
16
]
2–
or [H
2
Ru
10
(CO)
25
]
2–
interspersed with bis(triphenylphosphino)iminium counterions (PPN
+
) are incorporated from solution into the pores of MCM-41 mesoporous silica (3 nanometers in diameter), where they form tightly packed arrays. These arrays were shown by high-resolution transmission electron microscopy, Fourier transform optical diffraction, and computer simulations to be well ordered both along and perpendicular to the axis of the cylindrical pores. In their denuded state produced by gentle thermolysis, the cluster carbonylates yield nanoparticles of ruthenium that are less well ordered than their assimilated precursors but show good activity as hydrogenation catalysts for hexene and cyclooctene. In both their as-prepared and denuded states, these encapsulated clusters are likely to exhibit interesting electronic and other properties.
An Elusive Blind-Thrust Fault Beneath Metropolitan Los Angeles
Authors: Shaw John H. , Shearer and Peter M.
Published: July 2002

Seismic reflection profiles, petroleum wells, and relocated earthquakes reveal the presence of an active blind-thrust fault beneath metropolitan Los Angeles. A segment of this fault likely caused the 1987 Whittier Narrows (magnitude 6.0) earthquake. Mapped sizes of other fault segments suggest that the system is capable of much larger (magnitude 6.5 to 7) and more destructive earthquakes.
Single-Molecule Biomechanics with Optical Methods
Authors: Mehta Amit D. , Rief Matthias , Spudich James A. , Smith David A. , Simmons Robert M.
Published: July 2002

Single-molecule observation and manipulation have come of age. With the advent of optical tweezers and other methods for probing and imaging single molecules, investigators have circumvented the model-dependent extrapolation from ensemble assays that has been the hallmark of classical biochemistry and biophysics. In recent years, there have been important advances in the understanding of how motor proteins work. The range of these technologies has also started to expand into areas such as DNA transcription and protein folding. Here, recent experiments with rotary motors, linear motors, RNA polymerase, and titin are described.
Mirrorless Lasing from Mesostructured Waveguides Patterned by Soft Lithography
Authors: Yang Peidong , Wirnsberger † Gernot , Huang Howard C. , Cordero Steven R. , McGehee Michael D. , Scott Brian , Deng Tao , Whitesides George M. , Chmelka Bradley F. , Buratto Steven K. , Stucky Galen D.
Published: July 2002

Mesostructured silica waveguide arrays were fabricated with a combination of acidic sol-gel block copolymer templating chemistry and soft lithography. Waveguiding was enabled by the use of a low–refractive index (1.15) mesoporous silica thin film support. When the mesostructure was doped with the laser dye rhodamine 6G, amplified spontaneous emission was observed with a low pumping threshold of 10 kilowatts per square centimeter, attributed to the mesostructure's ability to prevent aggregation of the dye molecules even at relatively high loadings within the organized high–surface area mesochannels of the waveguides. These highly processible, self-assembling mesostructured host media and claddings may have potential for the fabrication of integrated optical circuits.
Efficacies of Vesicular and Free Sodium Stibogluconate Formulations against Clinical Isolates of
Leishmania donovani
Authors: Carter K. C. , Mullen A. B. , Sundar S. , Kenney R. T.
Published: July 2002

In this study, the in vitro and in vivo efficacies of free sodium stibogluconate (SSG) and a nonionic surfactant vesicular formulation of SSG (SSG-NIV) against a laboratory strain of
Leishmania donovani
(MHOM/ET/67:LV82) and different clinical isolates of
L. donovani
were determined. Treatment with SSG-NIV was more effective against intramacrophage amastigotes than treatment with SSG. In vivo murine studies showed that there was interstrain variability in the infectivity of the different
L. donovani
strains, with two of the strains (20001 and 20003) giving low parasite burdens. In addition, interstrain variability in the antileishmanial efficacy of SSG in a single dose containing 300 mg of Sb(V)/kg of body weight was observed. This dose of free drug either caused a >97% reduction in liver parasite burdens or had no significant effect on parasite burdens compared with the result with the respective control. In some instances, treatment with this free SSG dose also caused a significant reduction in spleen (strain 20006) or bone marrow (strains 20001 and 20009) parasite burdens. Treatment with SSG-NIV was more effective than that with SSG against all of the strains tested. In SSG-responsive strains, the reduction in liver parasite burdens by SSG-NIV treatment was similar to that caused by free SSG. In SSG-nonresponsive strains, SSG-NIV treatment caused at least a 95% reduction in liver parasite burdens. Overall, these results indicate that the use of a vesicular formulation of SSG is likely to increase its clinical efficacy against visceral leishmaniasis.
Multidrug Resistant
Mycobacterium leprae
from Patients with Leprosy
Authors: Maeda Shinji , Matsuoka Masanori , Nakata Noboru , Kai Masanori , Maeda Yumi , Hashimoto Ken , Kimura Hiroaki , Kobayashi Kazuo , Kashiwabara Yoshiko
Published: July 2002

Sequences of the
folP1
,
rpoB
, and
gyrA
genes were analyzed for 88 isolates of
Mycobacterium leprae
from leprosy patients in Japan, Haiti, Indonesia, Pakistan, and the Philippines. Thirteen isolates (14.8%) showed representative mutations in more than two genes, suggesting the emergence of multidrug-resistant
M. leprae
.
Rapid Detection of Mutations in the Human-Derived
Pneumocystis carinii
Dihydropteroate Synthase Gene Associated with Sulfa Resistance
Authors: Ma Liang , Kovacs Joseph A.
Published: July 2002

Recent studies have shown that point mutations in the dihydropteroate synthase (DHPS) gene of human-derived
Pneumocystis carinii
are related to exposure to sulfa drugs and possibly represent the emergence of sulfa resistance. We developed a simple single-strand conformation polymorphism (SSCP) method to permit rapid detection of these mutations. With plasmid constructs, SSCP was able to detect as little as 10% of a minority population. The SSCP assay was compared to direct sequencing for typing the DHPS gene by examining 37 clinical isolates with known DHPS sequences and 41 clinical isolates with unknown DHPS sequences. The typing results were consistent between these two methods for all isolates except 11 in which mutations were detected by SSCP but not by direct sequencing. Sequencing of individual clones after subcloning confirmed the presence of mutations in a minority population as determined by SSCP. SSCP is a very simple and sensitive method for rapid identification of
P. camii
DHPS mutations.
4′-Ethynyl Nucleoside Analogs: Potent Inhibitors of Multidrug-Resistant Human Immunodeficiency Virus Variants In Vitro
Authors: Kodama Ei-Ichi , Kohgo Satoru , Kitano Kenji , Machida Haruhiko , Gatanaga Hiroyuki , Shigeta Shiro , Matsuoka Masao , Ohrui Hiroshi , Mitsuya Hiroaki
Published: July 2002

A series of 4′-ethynyl (4′-
E
) nucleoside analogs were designed, synthesized, and identified as being active against a wide spectrum of human immunodeficiency viruses (HIV), including a variety of laboratory strains of HIV-1, HIV-2, and primary clinical HIV-1 isolates. Among such analogs examined, 4′-
E
-2′-deoxycytidine (4′-
E
-dC), 4′-
E
-2′-deoxyadenosine (4′-
E
-dA), 4′-
E
-2′-deoxyribofuranosyl-2,6-diaminopurine, and 4′-
E
-2′-deoxyguanosine were the most potent and blocked HIV-1 replication with 50% effective concentrations ranging from 0.0003 to 0.01 μM in vitro with favorable cellular toxicity profiles (selectivity indices ranging 458 to 2,600). These 4′-
E
analogs also suppressed replication of various drug-resistant HIV-1 clones, including HIV-1
M41L/T215Y
, HIV-1
K65R
, HIV-1
L74V
, HIV-1
M41L/T69S-S-G/T215Y
, and HIV-1
A62V/V75I/F77L/F116Y/Q151M
. Moreover, these analogs inhibited the replication of multidrug-resistant clinical HIV-1 strains carrying a variety of drug resistance-related amino acid substitutions isolated from HIV-1-infected individuals for whom 10 or 11 different anti-HIV-1 agents had failed. The 4′-
E
analogs also blocked the replication of a non-nucleoside reverse transcriptase inhibitor-resistant clone, HIV-1
Y181C
, and showed an HIV-1 inhibition profile similar to that of zidovudine in time-of-drug-addition assays. The antiviral activity of 4′-
E
-thymidine and 4′-
E
-dC was blocked by the addition of thymidine and 2′-deoxycytidine, respectively, while that of 4′-
E
-dA was not affected by 2′-deoxyadenosine, similar to the antiviral activity reversion feature of 2′,3′-dideoxynucleosides, strongly suggesting that 4′-
E
analogs belong to the family of nucleoside reverse transcriptase inhibitors. Further development of 4′-
E
analogs as potential therapeutics for infection with multidrug-resistant HIV-1 is warranted.
Genetic Organization of the Chromosome Region Surrounding
mecA
in Clinical Staphylococcal Strains: Role of IS
431
-Mediated
mecI
Deletion in Expression of Resistance in
mecA
-Carrying, Low-Level Methicillin- Resistant
Staphylococcus haemolyticus
Authors: Katayama Yuki , Ito Teruyo , Hiramatsu Keiichi
Published: July 2002

We report on the structural diversity of
mecA
gene complexes carried by 38 methicillin-resistant
Staphylococcus aureus
and 91 methicillin-resistant coagulase-negative
Staphylococcus
strains of seven different species with a special reference to its correlation with phenotypic expression of methicillin resistance. The most prevalent and widely disseminated
mec
complex had the structure
mecI-mecR1-mecA
-IS
431R
(or IS
431mec
), designated the class A
mecA
gene complex. In contrast, in
S. haemolyticus, mecA
was bracketed by two copies of IS
431
, forming the structure IS
431L-mecA
-IS
431R
. Of the 38
S. haemolyticus
strains, 5 had low-level methicillin resistance (MIC, 1 to 4 mg/liter) and characteristic heterogeneous methicillin resistance as judged by population analysis. In these five strains, IS
431L
was located to the left of an intact
mecI
gene, forming the structure IS
431L
-class A
mecA
-gene complex. In other
S. haemolyticus
strains, IS
431L
was associated with the deletion of
mecI
and
mecR1
, forming the structure IS
431L
-Δ
mecR1-mecA
-IS
431mec
, designated the class C
mecA
gene complex. Mutants with the class C
mecA
gene complex were obtained in vitro by selecting strain SH621, containing the IS
431L
-class A
mecA
gene complex with low concentrations of methicillin (1 and 3 mg/liter). The mutants had intermediate level of methicillin resistance (MIC, 16 to 64 mg/liter). The
mecA
gene transcription was shown to be derepressed in a representative mutant strain, SH621-37. Our study indicated that the
mecI
-encoded repressor function is responsible for the low-level methicillin resistance of some
S. haemolyticus
clinical strains and that the IS
431
-mediated
mecI
gene deletion causes the expression of methicillin resistance through the derepression of
mecA
gene transcription.
Convulsant and Subconvulsant Doses of Norfloxacin in the Presence and Absence of Biphenylacetic Acid Alter Extracellular Hippocampal Glutamate but Not Gamma-Aminobutyric Acid Levels in Conscious Rats
Authors: Smolders I. , Gousseau C. , Marchand S. , Couet W. , Ebinger G. , Michotte Y.
Published: July 2002

Fluoroquinolones are antibiotics with central excitatory side effects. These adverse effects presumably result from inhibition of γ-aminobutyric acid (GABA) binding to GABA
A
receptors. This GABA antagonistic effect is greatly potentiated by the active metabolite of fenbufen, biphenylacetic acid (BPAA). Nevertheless, it remains questionable whether GABA receptor antagonism alone can explain the convulsant activity potentials of these antimicrobial agents. The present study was undertaken to investigate the possible effects of norfloxacin, both in the absence and in the presence of BPAA, on the extracellular hippocampal levels of GABA and glutamate, the main central inhibitory and excitatory amino acid neurotransmitters, respectively. This in vivo microdialysis approach with conscious rats allows monitoring of behavioral alterations and concomitant transmitter modulation in the hippocampus. Peroral administration of 100 mg of BPAA per kg of body weight had no effect on behavior and did not significantly alter extracellular GABA or glutamate concentrations. Intravenous perfusion of 300 mg of norfloxacin per kg did not change the rat's behavior or the concomitant neurotransmitter levels in about half of the experiments, while the remaining animals exhibited severe seizures. These norfloxacin-induced convulsions did not affect extracellular hippocampal GABA levels but were accompanied by enhanced glutamate concentrations. Half of the rats receiving both 100 mg of BPAA per kg and 50 mg of norfloxacin per kg displayed lethal seizures, while the remaining animals showed no seizure-related behavior. In the latter subgroup, again no significant alterations in extracellular GABA levels were observed, but glutamate overflow remained significantly elevated for at least 3 h. In conclusion, norfloxacin exerts convulsant activity in rats, accompanied by elevations of extracellular hippocampal glutamate levels but not GABA levels, even in the presence of BPAA.
Presence of a Group II Intron in a Multiresistant
Serratia marcescens
Strain That Harbors Three Integrons and a Novel Gene Fusion
Authors: Centrón Daniela , Roy Paul H.
Published: July 2002

We analyzed the role of integrons in the dissemination of antibiotic resistance in a recent multiresistant clinical isolate,
Serratia marcescens
SCH88050909 (SCH909). This isolate harbors three integrons, all on a 60-kb conjugative plasmid. By PCR, hybridization, and sequencing analyses, we found that integron 1 has the
dfrA1
and
ant
(
3
")
-Ia
cassettes. The first cassette in integron 2 contains the
ant
(
2
")
-Ia
gene, separated from its
attC
site (59-base element) by a 1,971-bp insert containing a group II intron; this intron codes for a putative maturase-reverse transcriptase on the complementary strand and is the first such intron to be found associated with an integron. The
attC
site is followed by a novel aminoglycoside resistance gene,
ant
(
3
")
-Ii-aac
(
6
′)
-IId
, which has been characterized for its bifunctional ANT(3")-I and AAC(6′)-II activities. DNA sequence analysis of this fused cassette suggests that insertion and excision due to the integrase activity could have an important role in the evolution of aminoglycoside resistance genes. This gene is followed by an unknown open reading frame with a typical
attC
site and a partial cassette composed of the beginning of the
bla
OXA-10
cassette interrupted by IS
1
. The sequence downstream of IS
1
revealed that the
bla
OXA-10
cassette is incomplete and that the 3′ conserved segment of this integron is absent. Integron 3 is in a Tn
1696
-like transposon with the
aac
(
3
)
-Ia
cassette followed by three unknown cassettes and
ant
(
3
")
-Ia
. The presence of the group II intron and the relationship of group II introns in eubacteria with mobile elements suggest a possible role of this element in events such as cassette formation and/or plasmid evolution.
Imaging the Enzymatic Digestion of Bacterial Cellulose Ribbons Reveals the Endo Character of the Cellobiohydrolase Cel6A fromHumicola insolensand Its Mode of Synergy with Cellobiohydrolase Cel7A
Authors: Boisset Claire , Fraschini Carole , Schülein Martin , Henrissat Bernard , Chanzy Henri
Published: July 2002

Dispersed cellulose ribbons from bacterial cellulose were subjected to digestion with cloned Cel7A (cellobiohydrolase [CBH] I) and Cel6A (CBH II) fromHumicola insolenseither alone or in a mixture and in the presence of an excess of β-glucosidase. Both Cel7A and Cel6A were effective in partially converting the ribbons into soluble sugars, Cel7A being more active than Cel6A. In combination, these enzymes showed substantial synergy culminating with a molar ratio of approximately two-thirds Cel6A and one-third Cel7A. Ultrastructural transmission electron microscopy (TEM) observations indicated that Cel7A induced a thinning of the cellulose ribbons, whereas Cel6A cut the ribbons into shorter elements, indicating an endo type of action. These observations, together with the examination of the digestion kinetics, indicate that Cel6A can be classified as an endo-processive enzyme, whereas Cel7A is essentially a processive enzyme. Thus, the synergy resulting from the mixing of Cel6A and Cel7A can be explained by the partial endo character of Cel6A. A preparation of bacterial cellulose ribbons appears to be an appropriate substrate, superior toValoniaor bacterial cellulose microcrystals, to visualize directly by TEM the endo-processivity of an enzyme such as Cel6A.
Sediment Microbial Community Structure and Mercury Methylation in Mercury-Polluted Clear Lake, California
Authors: Macalady J. L. , Mack E. E. , Nelson D. C. , Scow K. M.
Published: July 2002

Spatial and temporal variations in sediment microbial community structure in a eutrophic lake polluted with inorganic mercury were identified using polar lipid fatty acid (PLFA) analysis. Microbial community structure was strongly related to mercury methylation potential, sediment organic carbon content, and lake location. Pore water sulfate, total mercury concentrations, and organic matter C/N ratios showed no relationships with microbial community structure. Seasonal changes and changes potentially attributable to temperature regulation of bacterial membranes were detectable but were less important influences on sediment PLFA composition than were differences due to lake sampling location. Analysis of biomarker PLFAs characteristic of
Desulfobacter
and
Desulfovibrio
groups of sulfate-reducing bacteria suggests that
Desulfobacter
-like organisms are important mercury methylators in the sediments, especially in the Lower Arm of Clear Lake.
Biofilm Community Structure in Polluted Rivers: Abundance of Dominant Phylogenetic Groups over a Complete Annual Cycle
Authors: Brümmer I. H. M. , Fehr W. , Wagner-Döbler I.
Published: July 2002

The seasonal dynamics of river biofilm communities in two German rivers, the Elbe and one of its tributaries, the Spittelwasser, were investigated for the first time by using fluorescence in situ hybridization and a standardized biofilm sampling procedure. We show the importance of members of the β subclass of the class
Proteobacteria
, which formed the largest single group in the massively polluted Spittelwasser at all times. Clear seasonal peaks of abundance were observed for the planctomycetes and the
Cytophaga-Flavobacterium
cluster.
Extensive Polymorphism in
Cryptosporidium parvum
Identified by Multilocus Microsatellite Analysis
Authors: Feng Xiaochuan , Rich Stephen M. , Akiyoshi Donna , Tumwine James K. , Kekitiinwa Addy , Nabukeera Nicolette , Tzipori Saul , Widmer Giovanni
Published: July 2002

Restriction fragment length polymorphism and DNA sequence analysis discern two main types of
Cryptosporidium parvum
. We present a survey of length polymorphism at several microsatellite loci for type 1 and type 2 isolates. A total of 14 microsatellite loci were identified from
C. parvum
DNA sequences deposited in public databases. All repeats were mono-, di-, and trinucleotide repeats of A, AT, and AAT, reflecting the high AT content of the
C. parvum
genome. Several of these loci showed significant length polymorphism, with as many as seven alleles identified for a single locus. Differences between alleles ranged from 1 to 27 bp. Karyotype analysis using probes flanking three microsatellites localized each marker to an individual chromosomal band, suggesting that these markers are single copy. In a sample of 19 isolates for which at least three microsatellites were typed, a majority of isolates displayed a unique multilocus fingerprint. Microsatellite analysis of isolates passaged between different host species identified genotypic changes consistent with changes in parasite populations.
Determination of the Sensitivity of a Rapid
Escherichia coli
O157:H7 Assay for Testing 375-Gram Composite Samples
Authors: Tsai Wan-Ling , Miller Cynthia E. , Richter Edward R.
Published: July 2002

Both 25-g single-size ground beef samples and 375-g composite ground beef samples were tested by a method combining an immunomagnetic separation (IMS) technique with a sandwich enzyme-linked immunosorbent assay (ELISA) system (IMS-ELISA). The results demonstrated that IMS-ELISA could detect the target,
Escherichia coli
O157:H7, at the level of 10
−1
CFU/g of sample in either the 25- or 375-g sample size.
Plant Genome Complexity May Be a Factor Limiting In Situ the Transfer of Transgenic Plant Genes to the Phytopathogen
Ralstonia solanacearum
Authors: Bertolla Franck , Pepin Regis , Passelegue-Robe Eugenie , Paget Eric , Simkin Andrew , Nesme Xavier , Simonet Pascal
Published: July 2002

The development of natural competence by bacteria in situ is considered one of the main factors limiting transformation-mediated gene exchanges in the environment.
Ralstonia solanacearum
is a plant pathogen that is also a naturally transformable bacterium that can develop the competence state during infection of its host. We have attempted to determine whether this bacterium could become the recipient of plant genes. We initially demonstrated that plant DNA was released close to the infecting bacteria. We constructed and tested various combinations of transgenic plants and recipient bacteria to show that the effectiveness of such transfers was directly related to the ratio of the complexity of the plant genome to the number of copies of the transgene.
Multiple Antibiotic Resistance (
mar
) Locus in
Salmonella enterica
Serovar Typhimurium DT104
Authors: Randall Luke P. , Woodward Martin J.
Published: July 2002

In order to understand the role of the
mar
locus in
Salmonella
with regard to multiple antibiotic resistance, cyclohexane resistance, and outer membrane protein F (OmpF) regulation, a
marA
::
gfp
reporter mutant was constructed in an antibiotic-sensitive
Salmonella enterica
serovar Typhimurium DT104 background. Salicylate induced
marA
, whereas a number of antibiotics, disinfectants, and various growth conditions did not. Increased antibiotic resistance was observed upon salicylate induction, although this was shown to be by both
mar
-dependent and
mar
-independent pathways. Cyclohexane resistance, however, was induced by salicylate by a
mar
-dependent pathway. Complementation studies with a plasmid that constitutively expressed
marA
confirmed the involvement of
mar
in
Salmonella
with low-level antibiotic resistance and cyclohexane resistance, although the involvement of
mar
in down regulation of OmpF was unclear. However,
marA
overexpression did increase the expression of a ca. 50-kDa protein, but its identity remains to be elucidated. Passage of the
marA
::
gfp
reporter mutant with increasing levels of tetracycline, a method reported to select for
mar
mutants in
Escherichia coli
, led to both multiple-antibiotic and cyclohexane resistance. Collectively, these data indicate that low-level antibiotic resistance, cyclohexane resistance, and modulation of OMPs in
Salmonella
, as in
E. coli
, can occur in both a
mar
-dependent and
mar
-independent manner.
Use of Pulsed-Field Gel Electrophoresis and Flagellin Gene Typing in Identifying Clonal Groups of
Campylobacter jejuni
and
Campylobacter coli
in Farm and Clinical Environments
Authors: Fitzgerald Collette , Stanley Karen , Andrew Sarah , Jones Keith
Published: July 2002

Although campylobacters have been isolated from a wide range of animal hosts, the association between campylobacters isolated from humans and animals in the farm environment is unclear. We used flagellin gene typing and pulsed-field gel electrophoresis (PFGE) to investigate the genetic diversity among isolates from animals (cattle, sheep, and turkey) in farm environments and sporadic cases of campylobacteriosis in the same geographical area. Forty-eight combined
fla
types were seen among the 315
Campylobacter
isolates studied. Six were found in isolates from all four hosts and represented 50% of the total number of isolates. Seventy-one different
Sma
I PFGE macrorestriction profiles (mrps) were observed, with 86% of isolates assigned to one of 29 different mrps. Fifty-seven isolates from diverse hosts, times, and sources had an identical
Sma
I mrp and combined
fla
type. Conversely, a number of genotypes were unique to a particular host. We provide molecular evidence which suggests a link between campylobacters in the farm environment with those causing disease in the community.
Map of the IncP1β Plasmid pTSA Encoding the Widespread Genes (
tsa
) for
p-
Toluenesulfonate Degradation in
Comamonas testosteroni
T-2
Authors: Tralau Tewes , Cook Alasdair M. , Ruff Juergen
Published: July 2002

The catabolic IncP1β plasmid pTSA from
Comamonas testosteroni
T-2 was mapped by subtractive analysis of restriction digests, by sequencing outwards from the
tsa
operon (toluenesulfonate degradation), and by generating overlapping, long-distance-PCR amplification products. The plasmid was estimated to comprise 72 ± 4 kb. The
tsa
region was found to be a composite transposon flanked by two IS
1071
elements. A cryptic
tsa
operon was also present in the
tsa
transposon. Those backbone genes and regions which we sequenced were in the same order as the corresponding genes in resistance plasmid R751, and identities of about 99% were observed. Enrichment cultures with samples from four continents were done to obtain organisms able to utilize
p
-toluenesulfonate as the sole source of carbon and energy for aerobic growth. Most (15) of the 16 cultures (13 of them isolates) were obtained from contaminated sites and were attributed to three metabolic groups, depending on their metabolism of
p
-toluenesulfonate. The largest group contained the
tsa
transposon, usually (six of seven isolates) with negligible differences in sequence from strain T-2.
Intra- and Extracellular β-Galactosidases from
Bifidobacterium bifidum
and
B. infantis
: Molecular Cloning, Heterologous Expression, and Comparative Characterization
Authors: Møller Peter L. , Jørgensen Flemming , Hansen Ole C. , Madsen Søren M. , Stougaard Peter
Published: July 2002

Three β-galactosidase genes from
Bifidobacterium bifidum
DSM20215 and one β-galactosidase gene from
Bifidobacterium infantis
DSM20088 were isolated and characterized. The three
B. bifidum
β-galactosidases exhibited a low degree of amino acid sequence similarity to each other and to previously published β-galactosidases classified as family 2 glycosyl hydrolases. Likewise, the
B. infantis
β-galactosidase was distantly related to enzymes classified as family 42 glycosyl hydrolases. One of the enzymes from
B. bifidum
, termed BIF3, is most probably an extracellular enzyme, since it contained a signal sequence which was cleaved off during heterologous expression of the enzyme in
Escherichia coli
. Other exceptional features of the BIF3 β-galactosidase were (i) the monomeric structure of the active enzyme, comprising 1,752 amino acid residues (188 kDa) and (ii) the molecular organization into an N-terminal β-galactosidase domain and a C-terminal galactose binding domain. The other two
B. bifidum
β-galactosidases and the enzyme from
B. infantis
were multimeric, intracellular enzymes with molecular masses similar to typical family 2 and family 42 glycosyl hydrolases, respectively. Despite the differences in size, molecular composition, and amino acid sequence, all four β-galactosidases were highly specific for hydrolysis of β-
d
-galactosidic linkages, and all four enzymes were able to transgalactosylate with lactose as a substrate.
Genotypic Analyses of
Escherichia coli
O157:H7 and O157 Nonmotile Isolates Recovered from Beef Cattle and Carcasses at Processing Plants in the Midwestern States of the United States
Authors: Barkocy-Gallagher Genevieve A. , Arthur Terrance M. , Siragusa Gregory R. , Keen James E. , Elder Robert O. , Laegreid William W. , Koohmaraie Mohammad
Published: July 2002

Escherichia coli
O157:H7 and O157 nonmotile isolates (
E. coli
O157) previously were recovered from feces, hides, and carcasses at four large Midwestern beef processing plants (R. O. Elder, J. E. Keen, G. R. Siragusa, G. A. Barkocy-Gallagher, M. Koohmaraie, and W. W. Laegreid, Proc. Natl. Acad. Sci. USA 97:2999–3003, 2000). The study implied relationships between cattle infection and carcass contamination within single-source lots as well as between preevisceration and postprocessing carcass contamination, based on prevalence. These relationships now have been verified based on identification of isolates by genomic fingerprinting.
E. coli
O157 isolates from all positive samples were analyzed by pulsed-field gel electrophoresis of genomic DNA after digestion with
Xba
I. Seventy-seven individual subtypes (fingerprint patterns) grouping into 47 types were discerned among 343 isolates. Comparison of the fingerprint patterns revealed three clusters of isolates, two of which were closely related to each other. Remarkably, isolates carrying both Shiga toxin genes and nonmotile isolates largely fell into specific clusters. Within lots analyzed, 68.2% of the postharvest (carcass) isolates matched preharvest (animal) isolates. For individual carcasses, 65.3 and 66.7% of the isolates recovered postevisceration and in the cooler, respectively, matched those recovered preevisceration. Multiple isolates were analyzed from some carcass samples and were found to include strains with different genotypes. This study suggests that most
E. coli
O157 carcass contamination originates from animals within the same lot and not from cross-contamination between lots. In addition, the data demonstrate that most carcass contamination occurs very early during processing.