bluuebunny/crossref_metadata_2025 huggingface dataset:
Regulation of Neurotrophin-3 Expression by Epithelial-Mesenchymal Interactions: The Role of Wnt Factors
Authors: Patapoutian Ardem , Backus Carey , Kispert Andreas , Reichardt Louis F.
Published: July 2002

Neurotrophins regulate survival, axonal growth, and target innervation of sensory and other neurons. Neurotrophin-3 (NT-3) is expressed specifically in cells adjacent to extending axons of dorsal root ganglia neurons, and its absence results in loss of most of these neurons before their axons reach their targets. However, axons are not required for NT-3 expression in limbs; instead, local signals from ectoderm induce NT-3 expression in adjacent mesenchyme. Wnt factors expressed in limb ectoderm induce NT-3 in the underlying mesenchyme. Thus, epithelial-mesenchymal interactions mediated by Wnt factors control NT-3 expression and may regulate axonal growth and guidance.
Variability in Spike Trains During Constant and Dynamic Stimulation
Authors: Warzecha Anne-Kathrin , Egelhaaf Martin
Published: July 2002

In a recent study, it was concluded that natural time-varying stimuli are represented more reliably in the brain than constant stimuli are. The results presented here disagree with this conclusion, although they were obtained from the same identified neuron (H1) in the fly's visual system. For large parts of the neuron's activity range, the variability of the responses was very similar for constant and time-varying stimuli and was considerably smaller than that in many visual interneurons of vertebrates.
Reconstructing Phylogeny with and without Temporal Data
Authors: Fox David L. , Fisher Daniel C. , Leighton Lindsey R.
Published: July 2002

Conventional cladistic methods of inferring evolutionary relationships exclude temporal data from the initial search for optimal hypotheses, but stratocladistics includes such data. A comparison of the ability of these methods to recover known, simulated evolutionary histories given the same, evolved character data shows that stratocladistics recovers the true phylogeny in over twice as many cases as cladistics (42 versus 18 percent). The comparison involved 550 unique taxon-by-character matrices, representing 15 evolutionary models and fossil records ranging from 100 to 10 percent complete.
The Evolution of Dinosaurs
Authors: Sereno Paul C.
Published: July 2002

The ascendancy of dinosaurs on land near the close of the Triassic now appears to have been as accidental and opportunistic as their demise and replacement by therian mammals at the end of the Cretaceous. The dinosaurian radiation, launched by 1-meter-long bipeds, was slower in tempo and more restricted in adaptive scope than that of therian mammals. A notable exception was the evolution of birds from small-bodied predatory dinosaurs, which involved a dramatic decrease in body size. Recurring phylogenetic trends among dinosaurs include, to the contrary, increase in body size. There is no evidence for co-evolution between predators and prey or between herbivores and flowering plants. As the major land masses drifted apart, dinosaurian biogeography was molded more by regional extinction and intercontinental dispersal than by the breakup sequence of Pangaea.
Worlds Apart? The Reception of Genetically Modified Foods in Europe and the U.S.
Authors: Gaskell George , Bauer Martin W. , Durant John , Allum Nicholas C.
Published: July 2002

Recent controversies about genetically modified foods in the United Kingdom and several other European countries highlight the apparent differences that exist in public opinion on this subject across the Atlantic. Why are people in the United States seemingly untroubled by a technology that causes Europeans so many difficulties? The results of survey research on public perceptions of biotechnology in Europe and the United States during 1996–1997, together with an analysis of press coverage and policy formation from 1984 to 1996, can help to answer this question.
Generation of a Widespread
Drosophila
Inversion by a Transposable Element
Authors: Cáceres Mario , Ranz José Marı́a , Barbadilla Antonio , Long Manyuan , Ruiz Alfredo
Published: July 2002

Although polymorphic inversions in
Drosophila
are very common, the origin of these chromosomal rearrangements is unclear. The breakpoints of the cosmopolitan inversion
2j
of
D. buzzatii
were cloned and sequenced. Both breakpoints contain large insertions corresponding to a transposable element. It appears that the two pairs of target site duplications generated upon insertion were exchanged during the inversion event, and that the inversion arose by ectopic recombination between two copies of the transposon that were in opposite orientations. This is apparently the mechanism by which transposable elements generate natural inversions in
Drosophila
.
Inhibition of Crystallite Growth in the Sol-Gel Synthesis of Nanocrystalline Metal Oxides
Authors: Wu Nae-Lih , Wang Sze-Yen , Rusakova I. A.
Published: July 2002

Crystal growth upon firing of hydrous transition metal oxide gels can be effectively inhibited by replacing the surface hydroxyl group before firing with another functional group that does not condense and that can produce small, secondary-phase particles that restrict advancing of grain boundaries at elevated temperatures. Accordingly, fully crystallized SnO
2
, TiO
2
, and ZrO
2
materials with mean crystallite sizes of ∼20, 50, and 15 angstroms, respectively, were synthesized by replacing the hydroxyl group with methyl siloxyl before firing at 500°C. An ultrasensitive SnO
2
-based chemical sensor resulting from the microstructural miniaturization was demonstrated.
The Role of CCR7 in T
H
1 and T
H
2 Cell Localization and Delivery of B Cell Help in Vivo
Authors: Randolph David A. , Huang Guangming , Carruthers Cynthia J. L. , Bromley Lindsay E. , Chaplin David D.
Published: July 2002

Subsets of murine CD4
+
T cells localize to different areas of the spleen after adoptive transfer. Naı̈ve and T helper 1 (T
H
1) cells, which express the chemokine receptor CCR7, are home to the periarteriolar lymphoid sheath, whereas activated T
H
2 cells, which lack CCR7, form rings at the periphery of the T cell zones near B cell follicles. Retroviral transduction of T
H
2 cells with CCR7 forces them to localize in a T
H
1-like pattern and inhibits their participation in B cell help in vivo but not in vitro. Thus, differential expression of chemokine receptors results in unique cellular migration patterns that are important for effective immune responses.
Genetic Definition and Sequence Analysis of
Arabidopsis
Centromeres
Authors: Copenhaver Gregory P. , Nickel Kathryn , Kuromori Takashi , Benito Maria-Ines , Kaul Samir , Lin Xiaoying , Bevan Michael , Murphy George , Harris Barbara , Parnell Laurence D. , McCombie W. Richard , Martienssen Robert A. , Marra Marco , Preuss Daphne
Published: July 2002

High-precision genetic mapping was used to define the regions that contain centromere functions on each natural chromosome in
Arabidopsis thaliana.
These regions exhibited dramatic recombinational repression and contained complex DNA surrounding large arrays of 180–base pair repeats. Unexpectedly, the DNA within the centromeres was not merely structural but also encoded several expressed genes. The regions flanking the centromeres were densely populated by repetitive elements yet experienced normal levels of recombination. The genetically defined centromeres were well conserved among
Arabidopsis
ecotypes but displayed limited sequence homology between different chromosomes, excluding repetitive DNA. This investigation provides a platform for dissecting the role of individual sequences in centromeres in higher eukaryotes.
Dual Signaling Regulated by Calcyon, a D1 Dopamine Receptor Interacting Protein
Authors: Lezcano Nelson , Mrzljak Ladislav , Eubanks Steven , Levenson Robert , Goldman-Rakic Patricia , Bergson Clare
Published: July 2002

The synergistic response of cells to the stimulation of multiple receptors has been ascribed to receptor cross talk; however, the specific molecules that mediate the resultant signal amplification have not been defined. Here a 24-kilodalton single transmembrane protein, designated calcyon, we functionally characterize that interacts with the D1 dopamine receptor. Calcyon localizes to dendritic spines of D1 receptor–expressing pyramidal cells in prefrontal cortex. These studies delineate a mechanism of G
q
- and G
s
-coupled heterotrimeric GTP–binding protein–coupled receptor cross talk by which D1 receptors can shift effector coupling to stimulate robust intracellular calcium (Ca
2+
i
) release as a result of interaction with calcyon. The role of calcyon in potentiating Ca
2+
-dependent signaling should provide insight into the D1 receptor–modulated cognitive functions of prefrontal cortex.
Simulation of Early 20th Century Global Warming
Authors: Delworth Thomas L. , Knutson Thomas R.
Published: July 2002

The observed global warming of the past century occurred primarily in two distinct 20-year periods, from 1925 to 1944 and from 1978 to the present. Although the latter warming is often attributed to a human-induced increase of greenhouse gases, causes of the earlier warming are less clear because this period precedes the time of strongest increases in human-induced greenhouse gas (radiative) forcing. Results from a set of six integrations of a coupled ocean-atmosphere climate model suggest that the warming of the early 20th century could have resulted from a combination of human-induced radiative forcing and an unusually large realization of internal multidecadal variability of the coupled ocean-atmosphere system. This conclusion is dependent on the model's climate sensitivity, internal variability, and the specification of the time-varying human-induced radiative forcing.
Stable RNA/DNA Hybrids in the Mammalian Genome: Inducible Intermediates in Immunoglobulin Class Switch Recombination
Authors: Tracy Robert B. , Hsieh Chih-Lin , Lieber Michael R.
Published: July 2002

Although it is well established that mammalian class switch recombination is responsible for altering the class of immunoglobulins, the mechanistic details of the process have remained unclear. Here, we show that stable RNA/DNA hybrids form at class switch sequences in the mouse genome upon cytokine-specific stimulation of class switch in primary splenic B cells. The RNA hybridized to the switch DNA is transcribed in the physiological orientation. Mice that constitutively express an
Escherichia coli
ribonuclease H transgene show a marked reduction in RNA/DNA hybrid formation, an impaired ability to generate serum immunoglobulin G antibodies, and significant inhibition of class switch recombination in their splenic B cells. These data provide evidence that stable RNA/DNA hybrids exist in the mammalian nuclear genome, can serve as intermediates for physiologic processes, and are mechanistically important for efficient class switching in vivo.
CLAVATA3, a Multimeric Ligand for the CLAVATA1 Receptor-Kinase
Authors: Trotochaud Amy E. , Jeong Sangho , Clark Steven E.
Published: July 2002

The CLAVATA1 (CLV1) and CLAVATA3 (CLV3) proteins form a potential receptor and ligand pair that regulates the balance between cell proliferation and differentiation at the shoot meristem of
Arabidopsis
. CLV1 encodes a receptor-kinase, and CLV3 encodes a predicted small, secreted polypeptide. We demonstrate that the CLV3 and CLV1 proteins coimmunoprecipitate in vivo, that yeast cells expressing CLV1 and CLV2 bind to CLV3 from plant extracts, and that binding requires CLV1 kinase activity. CLV3 only associates with the presumed active CLV1 protein complex in vivo. More than 75% of CLV3 in cauliflower extracts is bound with CLV1, consistent with hypotheses of ligand sequestration. Soluble CLV3 was found in an approximately 25-kilodalton multimeric complex.
Electronic Structure of Solids with Competing Periodic Potentials
Authors: Voit J. , Perfetti L. , Zwick F. , Berger H. , Margaritondo G. , Grüner G. , Höchst H. , Grioni M.
Published: July 2002

When electrons are subject to a potential with two incommensurate periods, translational invariance is lost, and no periodic band structure is expected. However, model calculations based on nearly free one-dimensional electrons and experimental results from high-resolution photoemission spectroscopy on a quasi–one-dimensional material do show dispersing band states with signatures of both periodicities. Apparent band structures are generated by the nonuniform distribution of electronic spectral weight over the complex eigenvalue spectrum.
Direct Coupling Between Meiotic DNA Replication and Recombination Initiation
Authors: Borde Valérie , Goldman Alastair S. H. , Lichten Michael
Published: July 2002

During meiosis in
Saccharomyces cerevisiae
, DNA replication occurs 1.5 to 2 hours before recombination initiates by DNA double-strand break formation. We show that replication and recombination initiation are directly linked. Blocking meiotic replication prevented double-strand break formation in a replication-checkpoint–independent manner, and delaying replication of a chromosome segment specifically delayed break formation in that segment. Consequently, the time between replication and break formation was held constant in all regions. We suggest that double-strand break formation occurs as part of a process initiated by DNA replication, which thus determines when meiotic recombination initiates on a regional rather than a cell-wide basis.
Neurons in Monkey Prefrontal Cortex That Track Past or Predict Future Performance
Authors: Hasegawa Ryohei P. , Blitz Ari M. , Geller Nancy L. , Goldberg Michael E.
Published: July 2002

Although frontal cortex is thought to be important in controlling behavior across long periods of time, most studies of this area concentrate on neuronal responses instantaneously relevant to the current task. In order to investigate the relationship of frontal activity to behavior over longer time periods, we trained rhesus monkeys on a difficult oculomotor task. Their performance fluctuated during the day, and the activity of prefrontal neurons, even measured while the monkeys waited for the targets to appear at the beginning of each set of trials, correlated with performance in a probabilistic rather than a determinist manner: neurons reflected past or predicted future performance, much more than they reflected current performance. We suggest that this activity is related to processes such as arousal or motivation that set the tone for behavior rather than controlling it on a millisecond basis, and could result from ascending pathways that utilize slow, second-messenger synaptic processes.
Active Disruption of an RNA-Protein Interaction by a DExH/D RNA Helicase
Authors: Jankowsky Eckhard , Gross Christian H. , Shuman Stewart , Pyle Anna Marie
Published: July 2002

All aspects of cellular RNA metabolism and the replication of many viruses require DExH/D proteins that manipulate RNA in a manner that requires nucleoside triphosphates. Although DExH/D proteins have been shown to unwind purified RNA duplexes, most RNA molecules in the cellular environment are complexed with proteins. It has therefore been speculated that DExH/D proteins may also affect RNA-protein interactions. We demonstrate that the DExH protein NPH-II from vaccinia virus can displace the protein U1A from RNA in an active adenosine triphosphate–dependent fashion. NPH-II increases the rate of U1A dissociation by more than three orders of magnitude while retaining helicase processivity. This indicates that DExH/D proteins can effectively catalyze protein displacement from RNA and thereby participate in the structural reorganization of ribonucleoprotein assemblies.
Prevention of Chemotherapy-Induced Alopecia in Rats by CDK Inhibitors
Authors: Davis Stephen T. , Benson Bill G. , Bramson H. Neal , Chapman Dennis E. , Dickerson Scott H. , Dold Karen M. , Eberwein Derek J. , Edelstein Mark , Frye Stephen V. , Gampe Robert T. , Griffin Robert J. , Harris Philip A. , Hassell Anne M. , Holmes William D. , Hunter Robert N. , Knick Victoria B. , Lackey Karen , Lovejoy Brett , Luzzio Michael J. , Murray Doris , Parker Patricia , Rocque Warren J. , Shewchuk Lisa , Veal James M. , Walker Duncan H. , Kuyper Lee F.
Published: July 2002

Most traditional cytotoxic anticancer agents ablate the rapidly dividing epithelium of the hair follicle and induce alopecia (hair loss). Inhibition of cyclin-dependent kinase 2 (CDK2), a positive regulator of eukaryotic cell cycle progression, may represent a therapeutic strategy for prevention of chemotherapy-induced alopecia (CIA) by arresting the cell cycle and reducing the sensitivity of the epithelium to many cell cycle–active antitumor agents. Potent small-molecule inhibitors of CDK2 were developed using structure-based methods. Topical application of these compounds in a neonatal rat model of CIA reduced hair loss at the site of application in 33 to 50% of the animals. Thus, inhibition of CDK2 represents a potentially useful approach for the prevention of CIA in cancer patients.
Structure of MsbA from
E. coli
: A Homolog of the Multidrug Resistance ATP Binding Cassette (ABC) Transporters
Authors: Chang Geoffrey , Roth Christopher B.
Published: July 2002

Multidrug resistance (MDR) is a serious medical problem and presents a major challenge to the treatment of disease and the development of novel therapeutics. ABC transporters that are associated with multidrug resistance (MDR-ABC transporters) translocate hydrophobic drugs and lipids from the inner to the outer leaflet of the cell membrane. To better elucidate the structural basis for the “flip-flop” mechanism of substrate movement across the lipid bilayer, we have determined the structure of the lipid flippase MsbA from
Escherichia coli
by x-ray crystallography to a resolution of 4.5 angstroms. MsbA is organized as a homodimer with each subunit containing six transmembrane α-helices and a nucleotide-binding domain. The asymmetric distribution of charged residues lining a central chamber suggests a general mechanism for the translocation of substrate by MsbA and other MDR-ABC transporters. The structure of MsbA can serve as a model for the MDR-ABC transporters that confer multidrug resistance to cancer cells and infectious microorganisms.
CD4+-T-Cell-Mediated Resistance to Systemic Murine Candidiasis Induced by a Membrane Fraction ofCandida albicans
Authors: Mizutani Shigetoshi , Endo Masahiro , Ino-ue Toshiaki , Kurasawa Masahiro , Uno Yoko , Saito Hideharu , Onogi Kaoru , Kato Ikunoshin , Takesako Kazutoh
Published: July 2002

We induced resistance to systemicCandida albicansinfection through CD4+-cell-mediated immunity in mice by immunization with subcutaneous injections of liveC. albicanscells emulsified in incomplete Freund adjuvant. Using the resistant mice, we tested subcellular fractions ofC. albicanscells for antigenicity. The fractions were derived from digested surface cell walls, insoluble membranes, or soluble and insoluble cytoplasmic materials, which were prepared by treatment with cell wall-digesting enzymes followed by lysis of the consequent protoplasts. Interestingly, the live-cell-immunized mice showed strong cell-mediated immune responses to the membrane fraction (C. albicansmembrane antigen [CMA]). In addition, immunization with CMA induced resistance to systemic candidiasis, which disappeared upon administration of anti-CD4 monoclonal antibody. Infusion of splenocytes from the CMA-immunized mice conferred resistance on SCID mice, whereas infusion of CD4+-T-cell-depleted splenocytes was unable to induce resistance, indicating the importance of CD4+lymphocytes for resistance. These results suggest a potential for the membrane fraction to act as an antigen conferring resistance to systemic candidiasis in place of live cells and also as a source for the isolation of a new antigen.
Heterogeneity of AmpC Cephalosporinases of
Hafnia alvei
Clinical Isolates Expressing Inducible or Constitutive Ceftazidime Resistance Phenotypes
Authors: Girlich Delphine , Naas Thierry , Bellais Samuel , Poirel Laurent , Karim Amal , Nordmann Patrice
Published: July 2002

Ten unrelated
Hafnia alvei
clinical isolates were grouped according to either their low-level and inducible cephalosporinase production or their high-level and constitutive cephalosporinase production phenotype. Their AmpC sequences shared 85 to 100% amino acid identity. The immediate genetic environment of
ampC
genes was conserved in
H. alvei
isolates but was different from that found in other
ampC
-possessing enterobacterial species.
Antibiotic Resistance in
Campylobacter
Strains Isolated from Animals, Foods, and Humans in Spain in 1997–1998
Authors: Sáenz Yolanda , Zarazaga Myriam , Lantero Marta , Gastañares M. José , Baquero Fernando , Torres Carmen
Published: July 2002

Colonization by
Campylobacter
strains was investigated in human, broiler, and pig fecal samples from 1997- 1998, as well as in foods of animal origin, and antibiotic susceptibility testing was carried out for these strains.
Campylobacter
strains were isolated in the foods of animal origin (55 of 101 samples; 54.4%), intestinal samples from broilers (85 of 105; 81%), and pigs (40 of 45; 88.9%). A total of 641
Campylobacter
strains were isolated from 8,636 human fecal samples of clinical origin (7.4%).
Campylobacter jejuni
was the most frequently isolated species from broilers (81%) and humans (84%), and
Campylobacter coli
was most frequently isolated from pigs (100%). An extremely high frequency of ciprofloxacin resistance was detected among
Campylobacter
strains, particularly those isolated from broilers and pigs (99%), with a slightly lower result for humans (72%); cross-resistance with nalidixic acid was almost always observed. A higher frequency of resistance to erythromycin (81.1%), ampicillin (65.7%), gentamicin (22.2%), and amikacin (21.6%) was detected in
C. coli
strains isolated from pigs compared to those isolated from humans (34.5, 29.3, 8.6, and 0%, respectively). A low frequency of erythromycin resistance was found in
C. jejuni
or
C. coli
isolated from broilers. A greater resistance to ampicillin and gentamicin (47.4 and 11.9%, respectively) was detected in
C. jejuni
isolated from broilers than in human strains (38 and 0.4%, respectively). β-Lactamase production was found in 81% of the
Campylobacter
strains tested, although 44% of them were characterized as ampicillin susceptible. The increasing rates of
Campylobacter
resistance make advisable a more conservative policy for the use of antibiotics in farm animals.
A Novel Human Immunodeficiency Virus Type 1 Reverse Transcriptase Mutational Pattern Confers Phenotypic Lamivudine Resistance in the Absence of Mutation 184V
Authors: Hertogs Kurt , Bloor Stuart , De Vroey Veronique , van den Eynde Christel , Dehertogh Pascale , van Cauwenberge Anja , Stürmer Martin , Alcorn Timothy , Wegner Scott , van Houtte Margriet , Miller Veronica , Larder Brendan A.
Published: July 2002

We describe a new human immunodeficiency virus type 1 (HIV-1) mutational pattern associated with phenotypic resistance to lamivudine (3TC) in the absence of the characteristic replacement of methionine by valine at position 184 (M184V) of reverse transcriptase. Combined genotypic and phenotypic analyses of clinical isolates revealed the presence of moderate levels of phenotypic resistance (between 4- and 50-fold) to 3TC in a subset of isolates that did not harbor the M184V mutation. Mutational cluster analysis and comparison with the phenotypic data revealed a significant correlation between moderate phenotypic 3TC resistance and an increased incidence of replacement of glutamic acid by aspartic acid or alanine and of valine by isoleucine at residues 44 and 118 of reverse transcriptase, respectively. This occurred predominantly in those isolates harboring zidovudine resistance-associated mutations (41L, 215Y). The requirement of the combination of mutations 41L and 215Y with mutations 44D and 44A and/or 118I for phenotypic 3TC resistance was confirmed by site-directed mutagenesis experiments. These data support the assumption that HIV-1 may have access to several different genetic pathways to escape drug pressure or that the increase in the frequency of particular mutations may affect susceptibility to drugs that have never been part of a particular regimen.
A Novel Phenotypic Drug Susceptibility Assay for Human Immunodeficiency Virus Type 1
Authors: Petropoulos Christos J. , Parkin Neil T. , Limoli Kay L. , Lie Yolanda S. , Wrin Terri , Huang Wei , Tian Huan , Smith Douglas , Winslow Genine A. , Capon Daniel J. , Whitcomb Jeannette M.
Published: July 2002

Although combination antiretroviral therapy has resulted in a considerable improvement in the treatment of human immunodeficiency virus (HIV) type 1 (HIV-1) infection, the emergence of resistant virus is a significant obstacle to the effective management of HIV infection and AIDS. We have developed a novel phenotypic drug susceptibility assay that may be useful in guiding therapy and improving long-term suppression of HIV replication. Susceptibility to protease (PR) and reverse transcriptase (RT) inhibitors is measured by using resistance test vectors (RTVs) that contain a luciferase indicator gene and PR and RT sequences derived from HIV-1 in patient plasma. Cells are transfected with RTV DNA, resulting in the production of virus particles that are used to infect target cells. Since RTVs are replication defective, luciferase activity is measured following a single round of replication. The assay has been automated to increase throughput and is completed in 8 to 10 days. Test results may be useful in facilitating the selection of optimal treatment regimens for patients who have failed prior therapy or drug-naive patients infected with drug-resistant virus. In addition, the assay can be used to evaluate candidate drugs and assist in the development of new drugs that are active against resistant strains of HIV-1.
A Population Pharmacokinetic Analysis of Nelfinavir Mesylate in Human Immunodeficiency Virus-Infected Patients Enrolled in a Phase III Clinical Trial
Authors: Jackson Kimberley A. , Rosenbaum Sara E. , Kerr Bradley M. , Pithavala Yazdi K. , Yuen Geoffrey , Dudley Michael N.
Published: July 2002

A population pharmacokinetic analysis was conducted on nelfinavir in patients infected with human immunodeficiency virus (HIV) who were enrolled in a phase III clinical trial. The data consisted of 509 plasma concentrations from 174 patients who received nelfinavir at a dose of 500 or 750 mg three times a day. The analysis was performed using nonlinear mixed-effect modeling as implemented in NONMEM (version 4.0; double precision). A one-compartment model with first-order absorption best described the data. The timing and small number of early postdose blood levels did not allow accurate estimation of volume of distribution (
V/F
) and the absorption rate constant (

k
a

). As a result, two models were used to analyze the data: model 1, in which oral clearance (CL/
F
),
V/F
, and

k
a

were estimated, and model 2, in which
V/F
and

k
a

were fixed to known values and only CL/
F
was estimated. Estimates of CL/
F
ranged from 41.9 to 45.1 liters/h, values in close agreement with previous studies. Neither body weight, age, sex, race, dose level, baseline viral load, metabolite-to-parent drug plasma concentration ratio, history of liver disease, nor elevated results of liver function tests appeared to be significant covariates for clearance. The only significant covariate-parameter relationship was concomitant use of fluconazole on CL/
F
, which was associated with a modest reduction in interindividual variability of CL/
F
. Patients who received concomitant therapy with fluconazole had a statistically significant reduction in nelfinavir CL/
F
of 26 to 30%. Since serious dose-limiting toxicity and concentration-related toxicities are not apparent for nelfinavir, this effect of fluconazole is unlikely to be of clinical significance.
Comparison of a Rabbit Model of Bacterial Endocarditis and an In Vitro Infection Model with Simulated Endocardial Vegetations
Authors: Hershberger Ellie , Coyle Elizabeth A. , Kaatz Glenn W. , Zervos Marcus J. , Rybak Michael J.
Published: July 2002

Animal models are commonly used to determine the efficacy of various antimicrobial agents for treatment of bacterial endocarditis. Previously we have utilized an in vitro infection model, which incorporates simulated endocardial vegetations (SEVs) to evaluate the pharmacodynamics of various antibiotics. In the present study, we compared four experimental rabbit endocarditis protocols to an in vitro infection model in an effort to determine if these models are comparable. We have evaluated the activity of clinafloxacin, trovafloxacin, sparfloxacin, and ciprofloxacin in rabbit models against
Staphylococcus aureus
and
Enterococcus
spp. In vitro models were performed simulating the antibiotic pharmacokinetics obtained in the in vivo studies. Models were dosed the same as rabbit models, and SEVs were evaluated at the same time the rabbit vegetations were examined. Clinafloxacin and trovafloxacin were evaluated against methicillin-susceptible (MSSA1199) and -resistant (MRSA494) strains of
S. aureus
. Ciprofloxacin was studied against MSSA1199 and MSSA487. Sparfloxacin and clinafloxacin were evaluated against
Enterococcus faecium
SF2149 and
Enterococcus faecalis
WH245, respectively. We found that reductions in SEV bacterial density obtained in the in vitro model were similar to those obtained in rabbit vegetations, indicating that the SEV model may be a valuable tool for assessing antibiotic potential in the treatment of bacterial endocarditis.
Selection of Naturally Occurring Extended-Spectrum TEM β-Lactamase Variants by Fluctuating β-Lactam Pressure
Authors: Blazquez Jesús , Morosini María-Isabel , Negri María-Cristina , Baquero Fernando
Published: July 2002

Despite the large number of in vitro mutations that increase resistance to extended-spectrum cephalosporins in TEM-type β-lactamases, only a small number occur in naturally occurring enzymes. In nature, and particularly in the hospital, bacteria that contain β-lactamases encounter simultaneous or consecutive selective pressure with different β-lactam molecules. All variants obtained by submitting anEscherichia colistrain that contains ablaTEM-1gene to fluctuating challenge with both ceftazidime and amoxicillin contained only mutations previously detected in naturally occurring β-lactamases. Nevertheless, some variants obtained by ceftazidime challenge alone contained mutations never detected in naturally occurring TEM β-lactamases, suggesting that extended-spectrum TEM variants in hospital isolates result from fluctuating selective pressure with several β-lactams rather than selection with a single antibiotic.
Clearance of Infection withMycobacterium bovisBCG in Mice Is Enhanced by Treatment with S28463 (R-848), and Its Efficiency Depends on Expression of Wild-TypeNramp1(Resistance Allele)
Authors: Moisan Jacques , Wojciechowski Wojciech , Guilbault Claudine , Lachance Claude , Di Marco Sergio , Skamene Emil , Matlashewski Greg , Radzioch Danuta
Published: July 2002

The mousebcghost resistance gene is known to control the activation of host macrophages for killing of intracellular parasites likeLeishmania donovanias well as intracellular bacteria, includingMycobacterium bovisBCG andSalmonella entericaserovar Typhimurium. TheNramp1gene has been mapped to this locus and affects the efficiency of macrophage activation. It has been shown that imidazoquinoline compounds, including S28463, are able to improve the clearance of a number of intracellular pathogens such as herpes simplex virus 2, human papillomavirus, andLeishmania. The goal of this study was to determine whether S28463 is efficient against infection with another intracellular pathogen,M. bovisBCG, and to determine the molecular basis underlying this effect. To achieve this, B10A.Nramp1rand B10A.Nramp1−/−mice were infected withM. bovisBCG and treated with S28463. The bacterial content in the spleen from these mice was assayed by a colony-forming assay. In addition, in vitro experiments were performed using bone marrow-derived macrophage cell lines from these mice. These cells were treated with S28463 and/or gamma interferon (IFN-γ), and nitric oxide (NO) production was measured. Our study was able to show that S28463 acts in synergy with IFN-γ to increase the production of NO in vitro. We were also able to demonstrate that mice that carried the resistant allele of theNramp1gene and were infected withM. bovisBCG responded to treatment with S28463, resulting in a decreased bacterial load after 2 weeks of treatment. Mice that do not express theNramp1gene responded only to a very large dose of S28463, and the response was not as efficient as that observed in mice carrying a wild-typeNramp1allele. Our data provide evidence for the potential of S28463 as an immunomodulator that may be helpful in designing efficient strategies to improve host defense against mycobacterial infection.
Characterization of OXA-29 fromLegionella(Fluoribacter)gormanii: Molecular Class D β-Lactamase with Unusual Properties
Authors: Franceschini Nicola , Boschi Letizia , Pollini Simona , Herman Raphaël , Perilli Mariagrazia , Galleni Moreno , Frère Jean-Marie , Amicosante Gianfranco , Rossolini Gian Maria
Published: July 2002

A class D β-lactamase determinant was isolated from the genome ofLegionella(Fluoribacter)gormaniiATCC 33297T. The enzyme, named OXA-29, is quite divergent from other class D β-lactamases, being more similar (33 to 43% amino acid identity) to those of groups III (OXA-1) and IV (OXA-9, OXA-12, OXA-18, and OXA-22) than to other class D enzymes (21 to 24% sequence identity). Phylogenetic analysis confirmed the closer ancestry of OXA-29 with members of the former groups. The OXA-29 enzyme was purified from anEscherichia colistrain overexpressing the gene via a T7-based expression system by a single ion-exchange chromatography step onS-Sepharose. The mature enzyme consists of a 28.5-kDa polypeptide and exhibits an isoelectric pH of >9. Analysis of the kinetic parameters of OXA-29 revealed efficient activity (kcat/Kmratios of >105M−1· s−1) for several penam compounds (oxacillin, methicillin, penicillin G, ampicillin, carbenicillin, and piperacillin) and also for cefazolin and nitrocefin. Oxyimino cephalosporins and aztreonam were also hydrolyzed, although less efficiently (kcat/Kmratios of around 103M−1· s−1). Carbapenems were neither hydrolyzed nor inhibitory. OXA-29 was inhibited by BRL 42715 (50% inhibitory concentration [IC50], 0.44 μM) and by tazobactam (IC50, 3.2 μM), but not by clavulanate. It was also unusually resistant to chloride ions (IC50, >100 mM). Unlike OXA-10, OXA-29 was apparently found as a dimer both in diluted solutions and in the presence of EDTA. Its activity was either unaffected or inhibited by divalent cations. OXA-29 is a new class D β-lactamase that exhibits some unusual properties likely reflecting original structural and mechanistic features.
SJ-3366, a Unique and Highly Potent Nonnucleoside Reverse Transcriptase Inhibitor of Human Immunodeficiency Virus Type 1 (HIV-1) That Also Inhibits HIV-2
Authors: Buckheit Robert W. , Watson Karen , Fliakas-Boltz Valerie , Russell Julie , Loftus Tracy L. , Osterling Mark C. , Turpin Jim A. , Pallansch Luke A. , White E. Lucile , Lee J.-W. , Lee S.-H. , Oh J.-W. , Kwon H.-S. , Chung S.-G. , Cho E.-H.
Published: July 2002

We have identified and characterized a potent new nonnucleoside reverse transcriptase (RT) inhibitor (NNRTI) of human immunodeficiency virus type 1 (HIV-1) that also is active against HIV-2 and which interferes with virus replication by two distinct mechanisms. 1-(3-Cyclopenten-1-yl)methyl-6-(3,5-dimethylbenzoyl)-5-ethyl-2,4-pyrimidinedione (SJ-3366) inhibits HIV-1 replication at concentrations of approximately 1 nM, with a therapeutic index of greater than 4 × 10
6
. The efficacy and toxicity of SJ-3366 are consistent when evaluated with established or fresh human cells, and the compound is equipotent against all strains of HIV-1 evaluated, including syncytium-inducing, non-syncytium-inducing, monocyte/macrophage-tropic, and subtype virus strains. Distinct from other members of the pharmacologic class of NNRTIs, SJ-3366 inhibited laboratory and clinical strains of HIV-2 at a concentration of approximately 150 nM, yielding a therapeutic index of approximately 20,000. Like most NNRTIs, the compound was less active when challenged with HIV-1 strains possessing the Y181C, K103N, and Y188C amino acid changes in the RT and selected for a virus with a Y181C amino acid change in the RT after five tissue culture passages in the presence of the compound. In combination anti-HIV assays with nucleoside and nonnucleoside RT and protease inhibitors, additive interactions occurred with all compounds tested with the exception of dideoxyinosine, with which a synergistic interaction was found. Biochemically, SJ-3366 exhibited a

K
i

value of 3.2 nM, with a mixed mechanism of inhibition against HIV-1 RT, but it did not inhibit HIV-2 RT. SJ-3366 also inhibited the entry of both HIV-1 and HIV-2 into target cells. On the basis of its therapeutic index and multiple mechanisms of anti-HIV action, SJ-3366 represents an exciting new compound for use in HIV-infected individuals.
Human Antibodies to Bacterial Superantigens and Their Ability To Inhibit T-Cell Activation and Lethality
Authors: LeClaire Ross D. , Bavari Sina
Published: July 2002

Bacterial superantigens (BSAgs) cause massive stimulation of the immune system and are associated with various pathologies and diseases. To address the role of antibodies in protection against BSAgs, we screened the sera of 29 human volunteers for antibodies to the SAgs staphylococcal enterotoxin A (SEA), SEB, SEC1, and toxic shock syndrome toxin 1 (TSST-1). Although all volunteers had detectable levels of antibodies against SEB and SEC1, many (9 out of 29 volunteers) lacked detectable antibody to SEA or had minimal titers. Antibody titers to TSST-1 were well below those to SEB and SEC1, and three volunteers lacked detectable antibody to this BSAg. In addition, pooled immunoglobulin preparations obtained from different companies had antibody titers against SEs and TSST-1. There was a good correlation between antibody titers and inhibition of superantigenic effects of these toxins. Transfer of SEB-specific antibodies, obtained from pooled sera, suppressed in vitro T-cell proliferation and totally protected mice against SEB. These data suggest that the inhibitory activity of human sera was specific to antibodies directed against the toxins. Thus, it may be possible to counteract with specific antibodies BSAg-associated pathologies caused by stimulation of the immune system.
Antiviral Guanosine Analogs as Substrates for Deoxyguanosine Kinase: Implications for Chemotherapy
Authors: Sjöberg Anita Herrström , Wang Liya , Eriksson Staffan
Published: July 2002

A highly active form of human recombinant deoxyguanosine kinase (dGK) phosphorylated purine nucleoside analogs active against cytomegalovirus, hepatitis B virus, and human immunodeficiency virus, such as penciclovir, 2′,3′-dideoxyguanosine and 3′-fluoro-2′,3′-dideoxyguanosine. The antiherpesvirus drug ganciclovir, which is also used in gene therapy, was a substrate for dGK, but with low efficiency. ATP and UTP were both good phosphate donors, with apparent

K
m

values of 6 and 4 μM and
V
max
values of 34 and 90 nmol of dGMP/mg of dGK/min, respectively. With a mixture of 5 mM ATP and 0.05 mM UTP, which represent physiologically relevant concentrations, the activities of dGK with ganciclovir and penciclovir was 1% and approximately 10%, respectively, of that with dGuo. The levels of dGK in different tissues were determined with a selective enzyme assay and the total activities per gram of tissues were similar in liver, brain, heart, and thymus extracts. The fact that the cellular dGK enzyme can phosphorylate antiviral guanosine analogs may help to explain the efficacies and side effects of several forms of chemotherapy.
Antibiotic Susceptibilities of
Salmonella enterica
Serovar Typhi and
S. enterica
Serovar Paratyphi A Isolated from Patients in Japan
Authors: Hirose Kenji , Tamura Kazumichi , Sagara Hiroko , Watanabe Haruo
Published: July 2002

The antibiotic susceptibilities of 62 strains of
Salmonella enterica
serovar Typhi and 37 strains of
S. enterica
serovar Paratyphi A were investigated with 18 antibiotics. Eighteen
S. enterica
serovar Typhi isolates and five
S. enterica
serovar Paratyphi A isolates were resistant to one or more antimicrobial agents, among which 10
S. enterica
serovar Typhi isolates were nalidixic acid resistant and also showed decreased ciprofloxacin susceptibility.
Pharmacokinetics of Itraconazole Oral Solution in Neutropenic Children during Long-Term Prophylaxis
Authors: Schmitt Claudine , Perel Yves , Harousseau Jean-luc , Lemerle Sophie , Chwetzoff Elisabeth , le Moing Jean-Pierre , Levron Jean-Claude
Published: July 2002

We investigated the pharmacokinetics and safety of an oral solution of itraconazole in two groups of neutropenic children stratified by age. Effective concentrations of itraconazole in plasma were reached quickly and maintained throughout treatment. The results indicate a trend toward higher concentrations of itraconazole in plasma in older children.
Treatment of Tuberculosis Using a Combination of Sustained-Release Rifampin-Loaded Microspheres and Oral Dosing with Isoniazid
Authors: Quenelle Debra C. , Winchester Gary A. , Staas Jay K. , Barrow Esther L. W. , Barrow William W.
Published: July 2002

Previously, we reported on the use of rifampin-loaded microspheres to effectively treat
Mycobacterium tuberculosis
-infected macrophages and mice. Using similar biocompatible polymeric excipients of lactide and glycolide copolymers, we have increased the rifampin loading of small microsphere formulations (1 to 10 μm) by fourfold. Improved formulations were evaluated individually and in combination with oral regimens of isoniazid for the treatment of
Mycobacterium tuberculosis
H37Rv-infected mice. Groups (10 mice per group) consisted of mice that received (i) oral dosages of isoniazid (25 to 0.19 mg/kg of body weight/day), (ii) two intraperitoneal injections of rifampin-loaded microspheres on days 0 and 7, (iii) a combination of small rifampin-loaded microspheres on days 0 and 7 and isoniazid orally for 25 days (12.5 to 0.39 mg/kg/day), (iv) placebo injections, and (v) no treatment. Treatment with rifampin-loaded microspheres alone resulted in significant reductions in the numbers of CFU in the lungs and spleens by day 26. A bioassay revealed that plasma rifampin levels from the microspheres exceeded the MICs by more than twofold throughout the 26-day experimental period. Susceptibility testing demonstrated continued sensitivity to rifampin during the treatment period. Whereas isoniazid alone significantly reduced the numbers of CFU for dosages ranging from 12.5 to 1.56 mg/kg, combination therapy with rifampin-loaded microspheres increased the effective range to 0.39 mg/kg. In many cases, complete elimination of CFU was obtained with the combination therapy, something not achieved with most of the single therapies. These results demonstrate the ability to use small microsphere formulations alone to achieve significant results in a murine tuberculosis model and also the ability to use them safely in combination with another antimycobacterial agent.
In Vitro Activities of Daptomycin against 2,789 Clinical Isolates from 11 North American Medical Centers
Authors: Barry Arthur L. , Fuchs Peter C. , Brown Steven D.
Published: July 2002

The in vitro activity of daptomycin is affected by the concentration of calcium cations in the test medium. Mueller-Hinton broth is currently adjusted to contain 10 to 12.5 mg of magnesium per liter and 20 to 25 mg of calcium per liter, but for testing of daptomycin, greater concentrations of calcium (50 mg/liter) are recommended to better resemble the normal concentration of ionized calcium in human serum. Two levels of calcium were used for broth microdilution tests of 2,789 recent clinical isolates of gram-positive bacterial pathogens. MICs of daptomycin were two- to fourfold lower when the broth contained additional calcium. For most species, however, the percentages of strains that were inhibited by 2.0 μg of daptomycin per ml were essentially identical with the two broth media. Enterococci were the important exception; i.e., 92% were inhibited when tested in calcium-supplemented broth but only 35% were inhibited by 2.0 μg/ml without the additional calcium. This type of information should be considered when selecting criteria for defining in vitro susceptibility to daptomycin.
Comparison of Hydrogen Peroxide Contact Lens Disinfection Systems and Solutions against
Acanthamoeba polyphaga
Authors: Hughes Reanne , Kilvington Simon
Published: July 2002

Acanthamoeba
is a free-living amoeba causing a potentially blinding infection of the cornea. Contact lens wearers are most at risk and account for some 95% of cases. Hydrogen peroxide is used for contact lens disinfection due to its broad antimicrobial activity. Lenses must be neutralized before use to avoid pronounced stinging and possible corneal damage. Neutralization is achieved by adding a catalyst during the disinfection process (one-step) or afterwards (two-step). Here, the activities of commercial peroxide systems and individual solutions against trophozoites and cysts of
Acanthamoeba polyphaga
were compared. All disinfection systems were active against trophozoites, giving a ≥3-log (99.9%) kill within 1 h. Of the four one-step systems, only one showed some cysticidal activity, giving a 1.28 ± 0.41-log reduction. Both two-step systems were cysticidal, giving a ≥3-log kill at 4 h. All system peroxide solutions were cysticidal, giving a ≥3-log kill by 4 to 6 h. Variation in the cysticidal rate was observed with two solutions that gave a 1.8- to 2.1-log kill at 4 h compared with 3.0 to 4.0 for the rest (
P
< 0.05). No cysticidal activity was found with the peroxigen sodium perborate or the contact lens protein remover subtilisin A. Two-step systems are cysticidal providing contact times of at least 4 h are employed. Variation in cyst killing occurs between peroxide solutions, possibly due to formulation differences. One-step systems are less effective against
Acanthamoeba
cysts due to rapid peroxide neutralization. The cysticidal activity of one-step systems could be improved if neutralization rates were retarded.
Clarithromycin Resistance and Eradication of
Helicobacter pylori
in Children
Authors: Kalach N. , Benhamou P. H. , Campeotto F. , Bergeret M. , Dupont C. , Raymond J.
Published: July 2002

Outcome of
Helicobacter pylori
infection was analyzed in 61 children treated with a triple therapy including clarithromycin. Bacterial eradication was obtained in all children with clarithromycin-susceptible strains but not in children with clarithromycin-resistant ones (
P
= 0.0001).
H. pylori
antimicrobial susceptibility is mandatory before choosing a treatment, and clarithromycin should be avoided in case of resistance.