bluuebunny/crossref_metadata_2025 huggingface dataset:
Evidence for Cell-Surface Association Between Fusin and the CD4-gp120 Complex in Human Cell Lines
Authors: Lapham Cheryl K. , Ouyang Jun , Chandrasekhar Bhaskar , Nguyen Nga Y. , Dimitrov Dimiter S. , Golding Hana
Published: July 2002

Accessory cell-surface molecules involved in the entry of human immunodeficiency virus-type 1 into cells have recently been identified and shown to belong to the family of chemokine receptors. Treatment of human cell lines with soluble monomeric gp120 at 37°C induced an association between the surface CD4-gp120 complex and a 45-kilodalton protein, which can be down-modulated by the phorbol ester phorbol 12-myristate 13-acetate. The three proteins were coprecipitated from the cell membranes with antibodies to CD4 or to gp120. The 45-kilodalton protein comigrated with fusin on sodium dodecyl sulfate gels and reacted with rabbit antisera to fusin in protein immunoblots. No 45-kilodalton protein could be coprecipitated from similarly treated nonhuman cells. However, infection of 3T3.CD4.401 cells with vaccinia-fusin recombinant virus (vCBYF1), followed by gp120 treatment, resulted in coprecipitation of fusin and CD4.401 molecules from their membranes. Together these data provide evidence for physical association between fusin and the CD4-gp120 complex on cell membranes.
On the Quantum Nature of the Shared Proton in Hydrogen Bonds
Authors: Tuckerman Mark E. , Marx Dominik , Klein Michael L. , Parrinello Michele
Published: July 2002

The relative influence of thermal and quantum fluctuations on the proton transfer properties of the charged water complexes H
5
O
2
+
and H
3
O
2
−
was investigated with the use of ab initio techniques. These small systems can be considered as prototypical representatives of strong and intermediate-strength hydrogen bonds. The shared proton in the strongly hydrogen bonded H
5
O
2
+
behaved in an essentially classical manner, whereas in the H
3
O
2
−
low-barrier hydrogen bond, quantum zero-point motion played a crucial role even at room temperature. This behavior can be traced back to a small difference in the oxygen-oxygen separation and hence to the strength of the hydrogen bond.
A Mammalian Telomerase-Associated Protein
Authors: Harrington Lea , McPhail Timothy , Mar Vernon , Zhou Wen , Oulton Rena , Program Amgen EST , Bass Mike B. , Arruda Isabel , Robinson Murray O.
Published: July 2002

The telomerase ribonucleoprotein catalyzes the addition of new telomeres onto chromosome ends. A gene encoding a mammalian telomerase homolog called TP1 (telomerase-associated protein 1) was identified and cloned. TP1 exhibited extensive amino acid similarity to the
Tetrahymena
telomerase protein p80 and was shown to interact specifically with mammalian telomerase RNA. Antiserum to TP1 immunoprecipitated telomerase activity from cell extracts, suggesting that TP1 is associated with telomerase in vivo. The identification of TP1 suggests that telomerase-associated proteins are conserved from ciliates to humans.
Metathesis of Alkanes Catalyzed by Silica-Supported Transition Metal Hydrides
Authors: Vidal Véronique , Théolier Albert , Thivolle-Cazat Jean , Basset Jean-Marie
Published: July 2002

The silica-supported transition metal hydrides (≡Si-O-Si≡)(≡Si-O-)
2
Ta-H and (≡Si-O-)
x
M-H (M, chromium or tungsten) catalyze the metathesis reaction of linear or branched alkanes into the next higher and lower alkanes at moderate temperature (25° to 200°C). With (≡Si-O-Si≡)(≡Si-O-)
2
Ta-H, ethane was transformed at room temperature into an equimolar mixture of propane and methane. Higher and lower homologs were obtained from propane, butane, and pentane as well as from branched alkanes such as isobutane and isopentane. The mechanism of the step leading to carbon-carbon bond cleavage and formation likely involves a four-centered transition state between a tantalum-alkyl intermediate and a carbon-carbon -bond of a second molecule of alkane.
INTELLECTUAL PROPERTY: Who Should Own Scientific Papers?
Authors: Liedes Jukka
Published: July 2002

Electronic communication has provided a new level of challenges and concerns in the area of protecting the rights of authors, producers, and publishers. Recently, the World Intellectual Property Organization (WIPO) began the process of updating the international copyright system to cope with today’s digital environment, but postponed discussion of a treaty that would have affected databases. Jukka Liedes, who drafted the treaty proposals, discusses the issues relevant to the scientific community in the treaties that were concluded and the issues that will need to be faced in considering protection of databases.
Genetic Feminization of Pheromones and Its Behavioral Consequences in
Drosophila
Males
Authors: Ferveur Jean-François , Savarit Fabrice , O'Kane Cahir J. , Sureau Gilles , Greenspan Ralph J. , Jallon Jean-Marc
Published: July 2002

Pheromones are intraspecific chemical signals important for mate attraction and discrimination. In the fruit fly
Drosophila melanogaster
, hydrocarbons on the cuticular surface of the animal are sexually dimorphic in both their occurrence and their effects: Female-specific molecules stimulate male sexual excitation, whereas the predominant male-specific molecule tends to inhibit male excitation. Complete feminization of the pheromone mixture produced by males was induced by targeted expression of the
transformer
gene in adult oenocytes (subcuticular abdominal cells) or by ubiquitous expression during early imaginal life. The resulting flies generally exhibited male heterosexual orientation but elicited homosexual courtship from other males.
Three-Color, Tunable, Organic Light-Emitting Devices
Authors: Shen Zilan , Burrows Paul E. , Bulović Vladimir , Forrest Stephen R. , Thompson Mark E.
Published: July 2002

An independently controlled, three-color, organic light-emitting device was constructed with a vertically stacked pixel architecture that allows for independent tuning of color, gray scale, and intensity. The 12-layer device structure consists of sequentially stacked layers of metal oxide, amorphous organic, crystalline organic, and metal thin films deposited by a combination of thermal evaporation and radio-frequency sputtering. Each of the three addressable colors is sufficiently bright for flat panel video display applications. A novel inverted structure is used for the middle device in the stack to lower the maximum drive voltage of the compound pixel.
Seismic Evidence of Partial Melt Within a Possibly Ubiquitous Low-Velocity Layer at the Base of the Mantle
Authors: Revenaugh J. , Meyer R.
Published: July 2002

Three source regions show evidence of a low-velocity layer that is less than 15 kilometers thick on top of the core-mantle boundary and require about a 3:1 ratio of shear-to-compressional velocity reduction, which is consistent with partial melt. Layer thickness is correlated with travel time residuals of the seismic phases that are most sensitive to the lowermost mantle velocity. These observations suggest that the layer is thinned beneath downwellings but is present everywhere along the core-mantle boundary. Low viscosity accompanying partial melt can localize the upwelling of warmed mantle, making the low-velocity layer a plausible source of mantle plumes.
IκB Kinase-β: NF-κB Activation and Complex Formation with IκB Kinase-α and NIK
Authors: Woronicz John D. , Gao Xiong , Cao Zhaodan , Rothe Mike , Goeddel David V.
Published: July 2002

Activation of the transcription factor nuclear factor kappa B (NF-κB) by inflammatory cytokines requires the successive action of NF-κB–inducing kinase (NIK) and IκB kinase–α (IKK-α). A widely expressed protein kinase was identified that is 52 percent identical to IKK-α. IκB kinase–β (IKK-β) activated NF-κB when overexpressed and phosphorylated serine residues 32 and 36 of IκB-α and serines 19 and 23 of IκB-β. The activity of IKK-β was stimulated by tumor necrosis factor and interleukin-1 treatment. IKK-α and IKK-β formed heterodimers that interacted with NIK. Overexpression of a catalytically inactive form of IKK-β blocked cytokine-induced NF-κB activation. Thus, an active IκB kinase complex may require three distinct protein kinases.
Coupled Quantum Dots Fabricated by Cleaved Edge Overgrowth: From Artificial Atoms to Molecules
Authors: Schedelbeck Gert , Wegscheider Werner , Bichler Max , Abstreiter Gerhard
Published: July 2002

Atomically precise quantum dots of mesoscopic size have been fabricated in the gallium arsenide–aluminum gallium arsenide material system by cleaved edge overgrowth, with a high degree of control over shape, composition, and position. The formation of bonding and antibonding states between two such “artificial atoms” was studied as a function of quantum dot separation by microscopic photoluminescence (PL) spectroscopy. The coupling strength within these “artificial molecules” is characterized by a systematic dependence of the separation of the bonding and antibonding levels, and of the PL linewidth, on the “interatomic” distance. This model system opens new insights into the physics of coupled quantum objects.
Extraocular Circadian Phototransduction in Humans
Authors: Campbell Scott S. , Murphy Patricia J.
Published: July 2002

Physiological and behavioral rhythms are governed by an endogenous circadian clock. The response of the human circadian clock to extraocular light exposure was monitored by measurement of body temperature and melatonin concentrations throughout the circadian cycle before and after light pulses presented to the popliteal region (behind the knee). A systematic relation was found between the timing of the light pulse and the magnitude and direction of phase shifts, resulting in the generation of a phase response curve. These findings challenge the belief that mammals are incapable of extraretinal circadian phototransduction and have implications for the development of more effective treatments for sleep and circadian rhythm disorders.
Single-Molecule Vibrational Spectroscopy and Microscopy
Authors: Stipe B. C. , Rezaei M. A. , Ho W.
Published: July 2002

Vibrational spectra for a single molecule adsorbed on a solid surface have been obtained with a scanning tunneling microscope (STM). Inelastic electron tunneling spectra for an isolated acetylene (C
2
H
2
) molecule adsorbed on the copper (100) surface showed an increase in the tunneling conductance at 358 millivolts, resulting from excitation of the C-H stretch mode. An isotopic shift to 266 millivolts was observed for deuterated acetylene (C
2
D
2
). Vibrational microscopy from spatial imaging of the inelastic tunneling channels yielded additional data to further distinguish and characterize the two isotopes. Single-molecule vibrational analysis should lead to better understanding and control of surface chemistry at the atomic level.
Crystal Structure of Hemolin: A Horseshoe Shape with Implications for Homophilic Adhesion
Authors: Su Xiao-Dong , Gastinel Louis N. , Vaughn Daniel E. , Faye Ingrid , Poon Pak , Bjorkman Pamela J.
Published: July 2002

Hemolin, an insect immunoglobulin superfamily member, is a lipopolysaccharide-binding immune protein induced during bacterial infection. The 3.1 angstrom crystal structure reveals a bound phosphate and patches of positive charge, which may represent the lipopolysaccharide binding site, and a new and unexpected arrangement of four immunoglobulin-like domains forming a horseshoe. Sequence analysis and analytical ultracentrifugation suggest that the domain arrangement is a feature of the L1 family of neural cell adhesion molecules related to hemolin. These results are relevant to interpretation of human L1 mutations in neurological diseases and suggest a domain swapping model for how L1 family proteins mediate homophilic adhesion.
A Molecular Pathway Revealing a Genetic Basis for Human Cardiac and Craniofacial Defects
Authors: Yamagishi Hiroyuki , Garg Vidu , Matsuoka Rumiko , Thomas Tiffani , Srivastava Deepak
Published: July 2002

Microdeletions of chromosome 22q11 are the most common genetic defects associated with cardiac and craniofacial anomalies in humans. A screen for mouse genes dependent on dHAND, a transcription factor implicated in neural crest development, identified
Ufd1
, which maps to human 22q11 and encodes a protein involved in degradation of ubiquitinated proteins. Mouse
Ufd1
was specifically expressed in most tissues affected in patients with 22q11 deletion syndrome. The human
UFD1L
gene was deleted in all 182 patients studied with 22q11 deletion, and a smaller deletion of approximately 20 kilobases that removed exons 1 to 3 of
UFD1L
was found in one individual with features typical of 22q11 deletion syndrome. These data suggest that
UFD1L
haploinsufficiency contributes to the congenital heart and craniofacial defects seen in 22q11 deletion.
Two Distinct Cytokines Released from a Human Aminoacyl-tRNA Synthetase
Authors: Wakasugi Keisuke , Schimmel Paul
Published: July 2002

Aminoacyl-tRNA synthetases catalyze aminoacylation of transfer RNAs (tRNAs). It is shown that human tyrosyl-tRNA synthetase can be split into two fragments with distinct cytokine activities. The endothelial monocyte–activating polypeptide II–like carboxy-terminal domain has potent leukocyte and monocyte chemotaxis activity and stimulates production of myeloperoxidase, tumor necrosis factor-α, and tissue factor. The catalytic amino-terminal domain binds to the interleukin-8 type A receptor and functions as an interleukin-8–like cytokine. Under apoptotic conditions in cell culture, the full-length enzyme is secreted, and the two cytokine activities can be generated by leukocyte elastase, an extracellular protease. Secretion of this tRNA synthetase may contribute to apoptosis both by arresting translation and producing needed cytokines.
Initiation of Mammalian Liver Development from Endoderm by Fibroblast Growth Factors
Authors: Jung Joonil , Zheng Minghua , Goldfarb Mitchell , Zaret Kenneth S.
Published: July 2002

The signaling molecules that elicit embryonic induction of the liver from the mammalian gut endoderm or induction of other gut-derived organs are unknown. Close proximity of cardiac mesoderm, which expresses fibroblast growth factors (FGFs) 1, 2, and 8, causes the foregut endoderm to develop into the liver. Treatment of isolated foregut endoderm from mouse embryos with FGF1 or FGF2, but not FGF8, was sufficient to replace cardiac mesoderm as an inducer of the liver gene expression program, the latter being the first step of hepatogenesis. The hepatogenic response was restricted to endoderm tissue, which selectively coexpresses FGF receptors 1 and 4. Further studies with FGFs and their specific inhibitors showed that FGF8 contributes to the morphogenetic outgrowth of the hepatic endoderm. Thus, different FGF signals appear to initiate distinct phases of liver development during mammalian organogenesis.
Genetic Variation in Susceptibility to Endocrine Disruption by Estrogen in Mice
Authors: Spearow Jimmy L. , Doemeny Paul , Sera Robyn , Leffler Rachael , Barkley Marylynn
Published: July 2002

Large (more than 16-fold) differences in susceptibility to disruption of juvenile male reproductive development by 17β-estradiol (E
2
) were detected between strains of mice. Effects of strain, E
2
dose, and the interaction of strain and E
2
dose on testes weight and spermatogenesis were all highly significant (
P
< 0.0001). Spermatid maturation was eliminated by low doses of E
2
in strains such as C57BL/6J and C17/Jls. In contrast, mice of the widely used CD-1 line, which has been selected for large litter size, showed little or no inhibition of spermatid maturation even in response to 16 times as much E
2
. Product safety bioassays conducted with animals selected for fecundity may greatly underestimate disruption of male reproductive development by estradiol and environmental estrogenic compounds.
Generating Solitons by Phase Engineering of a Bose-Einstein Condensate
Authors: Denschlag J. , Simsarian J. E. , Feder D. L. , Clark Charles W. , Collins L. A. , Cubizolles J. , Deng L. , Hagley E. W. , Helmerson K. , Reinhardt W. P. , Rolston S. L. , Schneider B. I. , Phillips W. D.
Published: July 2002

Quantum phase engineering is demonstrated with two techniques that allow the spatial phase distribution of a Bose-Einstein condensate (BEC) to be written and read out. A quantum state was designed and produced by optically imprinting a phase pattern onto a BEC of sodium atoms, and matter-wave interferometry with spatially resolved imaging was used to analyze the resultant phase distribution. An appropriate phase imprint created solitons, the first experimental realization of this nonlinear phenomenon in a BEC. The subsequent evolution of these excitations was investigated both experimentally and theoretically.
Control of Energy Transfer in Oriented Conjugated Polymer-Mesoporous Silica Composites
Authors: Nguyen Thuc-Quyen , Wu Junjun , Doan Vinh , Schwartz Benjamin J. , Tolbert Sarah H.
Published: July 2002

Nanoscale architecture was used to control energy transfer in semiconducting polymers embedded in the channels of oriented, hexagonal nanoporous silica. Polarized femtosecond spectroscopies show that excitations migrate unidirectionally from aggregated, randomly oriented polymer segments outside the pores to isolated, aligned polymer chains within the pores. Energy migration along the conjugated polymer backbone occurred more slowly than Förster energy transfer between polymer chains. The different intrachain and interchain energy transfer time scales explain the behavior of conjugated polymers in a range of solution environments. The results provide insights for optimizing nanostructured materials for use in optoelectronic devices.
Designing Small-Molecule Switches for Protein-Protein Interactions
Authors: Guo Zhihong , Zhou Demin , Schultz Peter G.
Published: July 2002

Mutations introduced into human growth hormone (hGH) (Thr
175
→ Gly–hGH) and the extracellular domain of the hGH receptor (Trp
104
→ Gly–hGHbp) created a cavity at the protein-protein interface that resulted in binding affinity being reduced by a factor of 10
6
. A small library of indole analogs was screened for small molecules that bind the cavity created by the mutations and restore binding affinity. The ligand 5-chloro-2-trichloromethylimidazole was found to increase the affinity of the mutant hormone for its receptor more than 1000-fold. Cell proliferation and JAK2 phosphorylation assays showed that the mutant hGH activates growth hormone signaling in the presence of added ligand. This approach may allow other protein-protein and protein–nucleic acid interactions to be switched on or off by the addition or depletion of exogenous small molecules.
Three-Dimensional Structure of the Tn
5
Synaptic Complex Transposition Intermediate
Authors: Davies Douglas R. , Goryshin Igor Y. , Reznikoff William S. , Rayment Ivan
Published: July 2002

Genomic evolution has been profoundly influenced by DNA transposition, a process whereby defined DNA segments move freely about the genome. Transposition is mediated by transposases, and similar events are catalyzed by retroviral integrases such as human immunodeficiency virus–1 (HIV-1) integrase. Understanding how these proteins interact with DNA is central to understanding the molecular basis of transposition. We report the three-dimensional structure of prokaryotic Tn
5
transposase complexed with Tn
5
transposon end DNA determined to 2.3 angstrom resolution. The molecular assembly is dimeric, where each double-stranded DNA molecule is bound by both protein subunits, orienting the transposon ends into the active sites. This structure provides a molecular framework for understanding many aspects of transposition, including the binding of transposon end DNA by one subunit and cleavage by a second, cleavage of two strands of DNA by a single active site via a hairpin intermediate, and strand transfer into target DNA.
Dependence of Stem Cell Fate in
Arabidopsis
on a Feedback Loop Regulated by
CLV3
Activity
Authors: Brand Ulrike , Fletcher Jennifer C. , Hobe Martin , Meyerowitz Elliot M. , Simon Rüdiger
Published: July 2002

The fate of stem cells in plant meristems is governed by directional signaling systems that are regulated by negative feedback. In
Arabidopsis thaliana
, the
CLAVATA
(
CLV
) genes encode the essential components of a negative, stem cell–restricting pathway. We used transgenic plants overexpressing
CLV3
to show that meristem cell accumulation and fate depends directly on the level of CLV3 activity and that CLV3 signaling occurs exclusively through a CLV1/CLV2 receptor kinase complex. We also demonstrate that the
CLV
pathway acts by repressing the activity of the transcription factor
WUSCHEL
, an element of the positive, stem cell–promoting pathway.
The Last Glacial-Holocene Transition in Southern Chile
Authors: Bennett K. D. , Haberle S. G. , Lumley S. H.
Published: July 2002

Warming at the last glacial termination in the North Atlantic region was interrupted by a period of renewed glacial activity during the Younger Dryas chronozone (YDC). The underlying mechanism of this cooling remains elusive, but hypotheses turn on whether it was a global or a North Atlantic phenomenon. Chronological, sedimentological, and palaeoecological records from sediments of small lakes in oceanic southern Chile demonstrate that there was no YDC cooling in southern Chile. It is therefore likely that there was little or no cooling in southern Pacific surface waters and hence that YDC cooling in the North Atlantic was a regional, rather than global, phenomenon.
Planar Hexacoordinate Carbon: A Viable Possibility
Authors: Exner Kai , Schleyer Paul von Ragué
Published: July 2002

The viability of molecules with planar hexacoordinate carbon atoms is demonstrated by density-functional theory (DFT) calculations for CB
6
2–
, a CB
6
H
2
isomer, and three C
3
B
4
minima. All of these species have six π electrons and are aromatic. Although other C
3
B
4
isomers are lower in energy, the activation barriers for the rearrangements of the three planar carbon C
3
B
4
minima into more stable isomers are appreciable, and experimental observation should be possible. High-level ab initio calculations confirm the DFT results. The planar hexacoordination in these species does not violate the octet rule because six partial bonds to the central carbons are involved.
A Primitive Enantiornithine Bird and the Origin of Feathers
Authors: Zhang Fucheng , Zhou Zhonghe
Published: July 2002

A fossil enantiornithine bird,
Protopteryx fengningensis
gen. et sp. nov., was collected from the Early Cretaceous Yixian Formation of Northern China. It provides fossil evidence of a triosseal canal in early birds. The manus and the alular digit are long, as in
Archaeopteryx
and
Confuciusornis
, but are relatively short in other enantiornithines. The alula or bastard wing is attached to an unreduced alular digit. The two central tail feathers are scalelike without branching. This type of feather may suggest that modern feathers evolved through the following stages: (i) elongated scale, (ii) central shaft, (iii) barbs, and finally (iv) barbules and barbicel.
Effect of a Cellulose Acetate Phthalate Topical Cream on Vaginal Transmission of Simian Immunodeficiency Virus in Rhesus Monkeys
Authors: Manson Kelledy H. , Wyand Michael S. , Miller Christopher , Neurath A. Robert
Published: July 2002

Human immunodeficiency virus type 1 (HIV-1) infection continues to spread in developing countries, mostly through heterosexual transmission. The development of a safe and cost-effective topical microbicide, effective against a range of STDs including HIV-1, would greatly impact the ongoing epidemic. When formulated in a vehicle, a micronized form of cellulose acetate phthalate (CAP), which is an inactive pharmaceutical excipient, has been shown to inactivate HIV-1, herpes simplex virus types 1 and 2, cytomegalovirus,
Neisseria gonorrhoeae
,
Trichomonas vaginalis
,
Haemophilus ducreyi
, and
Chlamydia trachomatis
in vitro. Formulated CAP was also shown to be effective against herpes simplex virus type 2 in vivo. Here we show that a formulation of CAP protected four of six rhesus monkeys from vaginal infection with simian immunodeficiency virus. Thus, CAP may be a candidate for use as a topical microbicide for preventing HIV-1 infection in humans.
Isolation and Characterization of Tetracycline-Resistant Clinical Isolates of
Helicobacter pylori
Authors: Kwon Dong H. , Kim J. J. , Lee M. , Yamaoka Y. , Kato M. , Osato M. S. , El-Zaatari F. A. K. , Graham David Y.
Published: July 2002

Tetracycline is an important component of combination therapies for
Helicobacter pylori
eradication. Twenty-nine tetracycline-resistant isolates requiring MICs ranging from 4 to 16 μg/ml were isolated from Korean (22 of 460) and Japanese (7 of 105) patients. Interestingly, all of the 29 tetracycline-resistant isolates exhibited cross-resistance to metronidazole, and the cross-resistance was transferred to tetracycline-sensitive
H. pylori
strains.
The Ribavirin Analog ICN 17261 Demonstrates Reduced Toxicity and Antiviral Effects with Retention of both Immunomodulatory Activity and Reduction of Hepatitis-Induced Serum Alanine Aminotransferase Levels
Authors: Tam Robert C. , Ramasamy Kanda , Bard Josie , Pai Bharati , Lim Charmaine , Averett Devron R.
Published: July 2002

The demonstrated utility of the nucleoside analog ribavirin in the treatment of certain viral diseases can be ascribed to its multiple distinct properties. These properties may vary in relative importance in differing viral disease conditions and include the direct inhibition of viral replication, the promotion of T-cell-mediated immune responses via an enhanced type 1 cytokine response, and a reduction of circulating alanine aminotransferase (ALT) levels associated with hepatic injury. Ribavirin also has certain known toxicities, including the induction of anemia upon chronic administration. To determine if all these properties are linked, we compared thed-nucleoside ribavirin to itsl-enantiomer (ICN 17261) with regard to these properties. Strong similarities were seen for these two compounds with respect to induction of type 1 cytokine bias in vitro, enhancement of type 1 cytokine responses in vivo, and the reduction of serum ALT levels in a murine hepatitis model. In contrast, ICN 17261 had no in vitro antiviral activity against a panel of RNA and DNA viruses, while ribavirin exhibited its characteristic activity profile. Importantly, the preliminary in vivo toxicology profile of ICN 17261 is significantly more favorable than that of ribavirin. Administration of 180 mg of ICN 17261 per kg of body weight to rats by oral gavage for 4 weeks generated substantial serum levels of drug but no observable clinical pathology, whereas equivalent doses of ribavirin induced a significant anemia and leukopenia. Thus, structural modification of ribavirin can dissociate its immunomodulatory properties from its antiviral and toxicologic properties, resulting in a compound (ICN 17261) with interesting therapeutic potential.
Difference in Incidence of Spontaneous Mutations between Herpes Simplex Virus Types 1 and 2
Authors: Sarisky Robert T. , Nguyen Tammy T. , Duffy Karen E. , Wittrock Robert J. , Leary Jeffry J.
Published: July 2002

Spontaneous mutations within the herpes simplex virus (HSV) genome are introduced by errors during DNA replication. Indicative of the inherent mutation rate of HSV DNA replication, heterogeneous HSV populations containing both acyclovir (ACV)-resistant and ACV-sensitive viruses occur naturally in both clinical isolates and laboratory stocks. Wild-type, laboratory-adapted HSV type 1 (HSV-1) strains KOS and Cl101 reportedly accumulate spontaneous ACV-resistant mutations at a frequency of approximately six to eight mutants per 10
4
plaque-forming viruses (U. B. Dasgupta and W. C. Summers, Proc. Natl. Acad. Sci. USA 75:2378–2381, 1978; J. D. Hall, D. M. Coen, B. L. Fisher, M. Weisslitz, S. Randall, R. E. Almy, P. T. Gelep, and P. A. Schaffer, Virology 132:26–37, 1984). Typically, these resistance mutations map to the thymidine kinase (TK) gene and render the virus TK deficient. To examine this process more closely, a plating efficiency assay was used to determine whether the frequencies of naturally occurring mutations in populations of the laboratory strains HSV-1 SC16, HSV-2 SB5, and HSV-2 333 grown in MRC-5 cells were similar when scored for resistance to penciclovir (PCV) and ACV. Our results indicate that (i) HSV mutants resistant to PCV and those resistant to ACV accumulate at approximately equal frequencies during replication in cell culture, (ii) the spontaneous mutation frequency for the HSV-1 strain SC16 is similar to that previously reported for HSV-1 laboratory strains KOS and Cl101, and (iii) spontaneous mutations in the laboratory HSV-2 strains examined were 9- to 16-fold more frequent than those in the HSV-1 strain SC16. These observations were confirmed and extended for a group of eight clinical isolates in which the HSV-2 mutation frequency was approximately 30 times higher than that for HSV-1 isolates. In conclusion, our results indicate that the frequencies of naturally occurring, or spontaneous, HSV mutants resistant to PCV and those resistant to ACV are similar. However, HSV-2 strains may have a greater propensity to generate drug-resistant mutants than do HSV-1 strains.
In Vivo Increase in Resistance to Ciprofloxacin in
Escherichia coli
Associated with Deletion of the C-Terminal Part of MarR
Authors: Linde Hans-Jörg , Notka Frank , Metz Michaela , Kochanowski Bernd , Heisig Peter , Lehn Norbert
Published: July 2002

We recovered two isolates (EP1 and EP2) of
Escherichia coli
from the same patient that had identical pulsed-field gel electrophoresis patterns but required different MICs of ciprofloxacin (CIP): 16 and 256 mg/liter for EP1 and EP2, respectively. Both isolates had mutations in the quinolone resistance-determining regions of GyrA (Ser83Leu and Asp87Tyr) and ParC (Ser80Ile), but not in those regions of GyrB or ParE. Isolate EP2 was also more resistant to chloramphenicol, tetracyclines, cefuroxime, and organic solvents. A deletion of adenine (A) 1821 was found in
marR
of isolate EP2, which resulted in an 18-amino-acid C-terminal deletion in the MarR protein. The causative relationship between ΔA1821 and the Mar phenotype was demonstrated both by the replacement of the wild-type
marR
by
marR
ΔA1821 in isolate EP1 and by complementation with the wild-type
marR
in
trans
in isolate EP2. In isolate EP2 complemented with wild-type
marR
, susceptibility to chloramphenicol was restored completely, whereas susceptibility to CIP was restored only incompletely. Northern blotting demonstrated increased expression of
marA
and
acrAB
but not of
soxS
in isolate EP2 compared to EP1. In conclusion, the deletion of A1821 in
marR
in the clinical isolate EP2 caused an increase in the MICs of CIP and unrelated antibiotics. Presumably, the C-terminal part of MarR is necessary for proper repressor function.
Pharmacokinetic and Pharmacodynamic Study of the Human Immunodeficiency Virus Protease Inhibitor Amprenavir after Multiple Oral Dosing
Authors: Sadler Brian M. , Gillotin Catherine , Lou Yu , Stein Daniel S.
Published: July 2002

In a dose-ranging study of amprenavir (formerly 141W94), an inhibitor of the protease enzyme of human immunodeficiency virus (HIV) type 1, single-dose and steady-state pharmacokinetic parameters were estimated from plasma samples collected on day 1 and during week 3, respectively. Amprenavir was administered on either a twice-daily (b.i.d.) or three-times-daily dosage schedule to 62 HIV-infected adults, 59 of whom had pharmacokinetic data. Log-log regression analysis (the power model) revealed that the steady-state area under the curve (AUC
ss
) and the maximum, minimum, and average concentrations at steady state (
C
max,ss
,
C
min,ss
, and
C
avg,ss
, respectively) increased in a dose-proportional manner over the 300- to 1,200-mg dose range. Steady-state clearance was dose independent. AUC
ss
/AUC
0→∞
decreased linearly with dose and correlated significantly with treatment-associated decreases in α
1
-acid glycoprotein. After 3 weeks, the dose of 1,200 mg b.i.d. provided a median amprenavir C
min,ss
(0.280 μg/ml) that was higher than the median in vitro 50% inhibitory concentration for clinical HIV isolates (0.023 μg/ml), even after adjustment for protein binding. The median amprenavir
C
min,ss
was also greater than the estimated in vivo trough concentration calculated to yield 90% of the maximum antiviral effect (0.228 μg/ml) over 4 weeks. A pharmacodynamic analysis of the relationship between steady-state pharmacokinetic parameters and safety revealed headache and oral numbness to be the only side effects significantly associated with
C
max
. The pharmacodynamic relationship defined in this study supports the use of 1,200 mg b.i.d. as the approved dose of amprenavir.
Prevalence and Mechanisms of Macrolide Resistance in Invasive and Noninvasive Group B Streptococcus Isolates from Ontario, Canada
Authors: de Azavedo Joyce C. S. , McGavin Mary , Duncan Carla , Low Donald E. , McGeer Allison
Published: July 2002

Macrolide resistance has been demonstrated in group B streptococcus (GBS), but there is limited information regarding mechanisms of resistance and their prevalence. We determined these in GBS obtained from neonatal blood cultures and vaginal swabs from pregnant women. Of 178 isolates from cases of neonatal GBS sepsis collected from 1995 to 1998, 8 and 4.5% were resistant to erythromycin and clindamycin, respectively, and one isolate showed intermediate penicillin resistance (MIC, 0.25 μg/ml). Of 101 consecutive vaginal or rectal/vaginal isolates collected in 1999, 18 and 8% were resistant to erythromycin and clindamycin, respectively. Tetracycline resistance was high (>80%) among both groups of isolates. Of 32 erythromycin-resistant isolates, 28 possessed the
erm
methylase gene (7
ermB
and 21
ermTR/ermA
) and 4 harbored the
mefA
gene; one isolate harbored both genes. One isolate which was susceptible to erythromycin but resistant to clindamycin (MIC, 4 μg/ml) was found to have the
linB
gene, previously identified only in
Enterococcus faecium
. The
mreA
gene was found in all the erythromycin-resistant strains as well as in 10 erythromycin-susceptible strains. The rate of erythromycin resistance increased from 5% in 1995–96 to 13% in 1998–99, which coincided with an increase in macrolide usage during that time.
Expression of Green Fluorescent Protein as a Marker for Effects of Antileishmanial Compounds In Vitro
Authors: Kamau Sarah W. , Grimm Felix , Hehl Adrian B.
Published: July 2002

TransgenicLeishmania infantumpromastigotes, which constitutively express green fluorescent protein (GFP) in their cytoplasm, were used to monitor the effects of antileishmanial compounds in real time. The GFP-based assay provided a reliable measure of drug-induced inhibitory effects on protein expression, resulting in a dynamic picture of the responses of leishmanial promastigotes to the compounds tested.
Penicillin-Binding Proteins in
Leptospira interrogans
Authors: Brenot Audrey , Trott Daren , Girons Isabelle Saint , Zuerner Richard
Published: July 2002

The
Leptospira interrogans ponA
and
pbpB
genes were isolated and characterized.
ponA
and
pbpB
encode the penicillin-binding proteins (PBPs) 1 and 3, respectively. There is little sequence variation between the PBP genes from two
L. interrogans
strains (serovar icterohaemorrhagiae strain Verdun and serovar pomona strain RZ11). The deduced
L. interrogans
PBP 1 and PBP 3 protein sequences from the two strains shared over 50% similarity to homologous proteins from
Escherichia coli
. It was demonstrated for strain Verdun that
ponA
and
pbpB
are transcribed individually from their own promoter. The
ponA
and
pbpB
genes from both strains are separated by 8 to 10 kb and oriented such that their transcription is convergent. The
L. interrogans
PBP 1 and PBP 3 proteins were synthesized in
E. coli
and were modified with ampicillin using a digoxigenin-ampicillin conjugate. These data show that both genes encode functional PBPs.
Gentamicin Increases the Efficacy of Vancomycin against Penicillin-Resistant Pneumococci in the Rabbit Meningitis Model
Authors: Cottagnoud Philippe , Gerber Cynthia M. , Cottagnoud Marianne , Täuber Martin G.
Published: July 2002

In experimental meningitis a single dose of gentamicin (10 mg/kg of body weight) led to gentamicin levels in around cerebrospinal fluid (CSF) of 4 mg/liter for 4 h, decreasing slowly to 2 mg/liter 4 h later. The CSF penetration of gentamicin ranged around 27%, calculated by comparison of areas under the curve (AUC in serum/AUC in CSF). Gentamicin monotherapy (−1.24 log
10
CFU/ml) was inferior to vancomycin monotherapy (−2.54 log
10
CFU/ml) over 8 h against penicillin-resistant pneumococci. However, the combination of vancomycin with gentamicin was significantly superior (−4.48 log
10
CFU/ml) compared to either monotherapy alone. The synergistic activity of vancomycin combined with gentamicin was also demonstrated in vitro in time-kill assays.
In Situ Analysis of Sulfate-Reducing Bacteria Related to
Desulfocapsa thiozymogenes
in the Chemocline of Meromictic Lake Cadagno (Switzerland)
Authors: Tonolla Mauro , Demarta Antonella , Peduzzi Sandro , Hahn Dittmar , Peduzzi Raffaele
Published: July 2002

Comparative sequence analysis of a 16S rRNA gene clone library from the chemocline of the meromictic Lake Cadagno (Switzerland) retrieved two clusters of sequences resembling sulfate-reducing bacteria within the family
Desulfovibrionaceae
. In situ hybridization showed that, similar to sulfate-reducing bacteria of the family
Desulfobacteriaceae
, bacteria of one cluster with similarity values to the closest cultured relatives of between 92.6 and 93.1% resembled free cells or cells loosely attached to other cells or debris. Bacteria of the second cluster closely related to
Desulfocapsa thiozymogenes
DSM7269 with similarity values between 97.9 and 98.4% were generally associated with aggregates of different small-celled phototrophic sulfur bacteria, suggesting a potential interaction between the two groups of bacteria.
Optimization of Annealing Temperature To Reduce Bias Caused by a Primer Mismatch in Multitemplate PCR
Authors: Ishii Kousuke , Fukui Manabu
Published: July 2002

To reduce PCR bias derived from a primer mismatch, the effect of the annealing temperature on the product ratio was investigated by denaturing gradient gel electrophoresis analysis of PCR products from a mixture of perfect-match and one-mismatch templates. These templates were generated by PCR from
Pediococcus acidilactici
for one mismatch and
Micrococcus luteus
for the perfect match. PCRs showed that the bias was reduced at lower temperatures. An environmental sample was also examined.
New Spatially Explicit Method for Detecting Extracellular Protease Activity in Biofilms
Authors: Francoeur Steven N. , Wetzel Robert G. , Neely Robert K.
Published: July 2002

A novel method of detecting extracellular protease activity at biofilm-substratum interfaces was developed. This method utilizes fluorescent molecules bound to cellulose substrata with a lectin. Extracellular proteases degrade the lectin and release the fluorochrome into solution. This new technique and a standard dissolved-substrate assay detected similar responses of biofilm extracellular protease activity to experimental manipulation of N supply. Combination of this technique with confocal scanning laser microscopy allowed direct visualization of microspatial patterns of bacterial distribution and extracellular protease activity at the biofilm-substratum interface.
Essential Role for the
Legionella pneumophila
Rep Helicase Homologue in Intracellular Infection of Mammalian Cells
Authors: Harb Omar S. , Abu Kwaik Yousef
Published: July 2002

We have previously isolated 32 mutants of
Legionella pneumophila
that are defective in the infection of mammalian cells but not protozoa. The mutated loci have been designated macrophage-specific infectivity (
mil
) loci. In this study we characterized the
mil
mutant GK11. This mutant was incapable of growth within U937 macrophage-like cells and WI-26 alveolar epithelial cells. This defect in intracellular replication correlated with a defect in cytopathogenicity to these cells. Sequence analysis of the GK11 locus revealed it to be highly similar to
rep
helicase genes of other bacteria. Since helicase mutants of
Escherichia coli
are hypersensitive to thymine starvation, we examined the sensitivity of GK11 to thymineless death (TLD). In the absence of thymine and thymidine, mutant GK11 did not undergo TLD but was defective for in vitro growth, and the defect was partially restored when these compounds were added to the growth medium. In addition, supplementation with thymidine or thymine partially restored the ability of GK11 to grow within and kill U937 macrophage-like cells. The data suggested that the low levels of thymine or thymidine in the
L. pneumophila
phagosome contributed to the defect of GK11 within macrophages. Using confocal laser scanning microscopy, we determined the effect of the mutation in the Rep helicase homologue on the intracellular trafficking of GK11 within macrophages. In contrast to the wild-type strain, phagosomes harboring GK11 colocalized with several late endosomal/lysosomal markers, including LAMP-1, LAMP-2, and cathepsin D. In addition, only 50% of the GK11 phagosomes colocalized with the endoplasmic reticulum marker BiP 4 h postinfection. Colocalization of BiP with GK11 phagosomes was absent 6 h postinfection, while 90% of the wild-type phagosomes colocalized with this marker at both time points. We propose that the low level of thymine within the
L. pneumophila
phagosome in combination with simultaneous exposure to multiple stress stimuli results in deleterious mutations that cannot be repaired in the
rep
helicase homologue mutant, rendering it defective in intracellular replication.
Molecular Characterization of a New Variant of Toxin-Coregulated Pilus Protein (TcpA) in a Toxigenic Non-O1/Non-O139 Strain of
Vibrio cholerae
Authors: Nandi Bisweswar , Nandy Ranjan K. , Vicente Ana C. P. , Ghose Asoke C.
Published: July 2002

A toxigenic non-O1/non-O139 strain of
Vibrio cholerae
(10259) was found to contain a new variant of the toxin-coregulated pilus (TCP) protein gene (
tcpA
) as determined by PCR and Southern hybridization experiments. Nucleotide sequence analysis data of the new
tcpA
gene in strain 10259 (O53) showed it to be about 74 and 72% identical to those of O1 classical and El Tor biotype strains, respectively. The predicted amino acid sequence of the 10259 TcpA protein shared about 81 and 78% identity with the corresponding sequences of classical and El Tor TcpA strains, respectively. An antiserum raised against the TCP of a classical strain, O395, although it recognized the TcpA protein of strain 10259 in an immunoblotting experiment, exhibited considerably less protection against 10259 challenge compared to that observed against the parent strain. Incidentally, the
tcpA
sequences of two other toxigenic non-O1/non-O139 strains (V2 and S7, both belonging to the serogroup O37) were determined to be almost identical to that of classical
tcpA
. Further,
tcpA
of another toxigenic non-O1/non-O139 strain V315-1 (O nontypeable) was closely related to that of El Tor
tcpA
. Analysis of these results with those already available in the literature suggests that there are at least four major variants of the
tcpA
gene in
V. cholerae
which probably evolved in parallel from a common ancestral gene. Existence of highly conserved as well as hypervariable regions within the sequence of the TcpA protein would also predict that such evolution is under the control of considerable selection pressure.
Evidence of Involvement of the Mannose Receptor in Adhesion ofBorrelia burgdorferito Monocyte/Macrophages
Authors: Cinco Marina , Cini Barbara , Murgia Rossella , Presani Gianni , Prodan Mario , Perticarari Sandra
Published: July 2002

The mannose receptor (MR) plays an important role in the recognition of some pathogens in nonopsonic phagocytosis and in antigen presentation to T cells. We found thatBorrelia burgdorferi, the agent of Lyme borreliosis, adheres to monocyte-derived macrophages and to rat MR-transfected cells but not to untransfected cells. Antibodies to MR and sugars such as mannose, mannan, fucose, and some lectins significantly lowered the adhesion, confirming participation of the MR in the binding.
Role of Lung Epithelial Cells in Defense against
Klebsiella pneumoniae
Pneumonia
Authors: Cortés Guadalupe , Álvarez Dolores , Saus Carles , Albertí Sebastián
Published: July 2002

The airway epithelium represents a primary site for the entry of pathogenic bacteria into the lungs. It has been suggested for many respiratory pathogens, including
Klebsiella pneumoniae
, that adhesion and invasion of the lung epithelial cells is an early stage of the pneumonia process. We observed that poorly encapsulated
K. pneumoniae
clinical isolates and an isogenic unencapsulated mutant invaded lung epithelial cells more efficiently than highly encapsulated strains independent of the K type. By contrast, the unencapsulated mutant was completely avirulent in a mouse model of pneumonia, unlike the wild-type strain, which produced pneumonia and systemic infection. Furthermore, the unencapsulated mutant bound more epithelially produced complement component C3 than the wild-type strain. Our results show that lung epithelial cells play a key role as a host defense mechanism against
K. pneumoniae
pneumonia, using two different strategies: (i) ingestion and control of the microorganisms and (ii) opsonization of the microorganisms. Capsular polysaccharide avoids both mechanisms and enhances the virulence of
K. pneumoniae
.
Differences in Genotypes ofHelicobacter pylorifrom Different Human Populations
Authors: Kersulyte Dangeruta , Mukhopadhyay Asish K. , Velapatiño Billie , Su WanWen , Pan ZhiJun , Garcia Claudia , Hernandez Virginia , Valdez Yanet , Mistry Rajesh S. , Gilman Robert H. , Yuan Yuan , Gao Hua , Alarcón Teresa , López-Brea Manuel , Balakrish Nair G. , Chowdhury Abhijit , Datta Simanti , Shirai Mutsunori , Nakazawa Teruko , Ally Reidwaan , Segal Isidore , Wong Benjamin C. Y. , Lam S. K. , Olfat Farzad O. , Borén Thomas , Engstrand Lars , Torres Olga , Schneider Roberto , Thomas Julian E. , Czinn Steven , Berg Douglas E.
Published: July 2002

DNA motifs at several informative loci in more than 500 strains ofHelicobacter pylorifrom five continents were studied by PCR and sequencing to gain insights into the evolution of this gastric pathogen. Five types of deletion, insertion, and substitution motifs were found at the right end of theH. pylori cagpathogenicity island. Of the three most common motifs, type I predominated in Spaniards, native Peruvians, and Guatemalan Ladinos (mixed Amerindian-European ancestry) and also in native Africans and U.S. residents; type II predominated among Japanese and Chinese; and type III predominated in Indians from Calcutta. Sequences in thecagAgene and invacAm1type alleles of the vacuolating cytotoxin gene (vacA) of strains from native Peruvians were also more like those from Spaniards than those from Asians. These indications of relatedness of Latin American and Spanish strains, despite the closer genetic relatedness of Amerindian and Asian people themselves, lead us to suggest thatH. pylorimay have been brought to the New World by European conquerors and colonists about 500 years ago. This thinking, in turn, suggests thatH. pyloriinfection might have become widespread in people quite recently in human evolution.