bluuebunny/crossref_metadata_2025 huggingface dataset:
Mutations in the
SMAD4/DPC4
Gene in Juvenile Polyposis
Authors: Howe James R. , Roth Stina , Ringold John C. , Summers Robert W. , Järvinen Heikki J. , Sistonen Pertti , Tomlinson Ian P. M. , Houlston Richard S. , Bevan Steve , Mitros Frank A. , Stone Edwin M. , Aaltonen Lauri A.
Published: July 2002

Familial juvenile polyposis is an autosomal dominant disease characterized by a predisposition to hamartomatous polyps and gastrointestinal cancer. Here it is shown that a subset of juvenile polyposis families carry germ line mutations in the gene
SMAD4
(also known as
DPC4
), located on chromosome 18q21.1, that encodes a critical cytoplasmic mediator in the transforming growth factor–β signaling pathway. The mutant SMAD4 proteins are predicted to be truncated at the carboxyl-terminus and lack sequences required for normal function. These results confirm an important role for
SMAD4
in the development of gastrointestinal tumors.
Elasticity of Single-Crystal MgO to 8 Gigapascals and 1600 Kelvin
Authors: Chen Ganglin , Liebermann Robert C. , Weidner Donald J.
Published: July 2002

The cross pressure (
P
) and temperature (
T
) dependence of the elastic moduli (

C
ij

) of single-crystal samples of periclase (MgO) from acoustic wave travel times was measured with ultrasonic interferometry: ∂
2
C
11
/∂
P
∂
T
= (−1.3 ± 0.4) × 10
−3
per kelvin; ∂
2
C
110
/∂
P
∂
T
= (1.7 ± 0.7) × 10
−3
per kelvin; and ∂
2
C
44
/∂
P
∂
T
= (−0.2 ± 0.3) × 10
−3
per kelvin. The elastic anisotropy of MgO decreases with increasing pressure at ambient temperature, but then increases as temperature is increased at high pressure. An assumption of zero cross pressure and temperature derivatives for the elastic moduli underestimates the elastic anisotropy and overestimates the acoustic velocities of MgO at the extrapolated high-pressure and high-temperature conditions of Earth's mantle.
Hierarchically Ordered Oxides
Authors: Yang Peidong , Deng Tao , Zhao Dongyuan , Feng Pingyun , Pine David , Chmelka Bradley F. , Whitesides George M. , Stucky Galen D.
Published: July 2002

Porous silica, niobia, and titania with three-dimensional structures patterned over multiple length scales were prepared by combining micromolding, polystyrene sphere templating, and cooperative assembly of inorganic sol-gel species with amphiphilic triblock copolymers. The resulting materials show hierarchical ordering over several discrete and tunable length scales ranging from 10 nanometers to several micrometers. The respective ordered structures can be independently modified by choosing different mold patterns, latex spheres, and block copolymers. The examples presented demonstrate the compositional and structural diversities that are possible with this simple approach.
Illuminating Single Molecules in Condensed Matter
Authors: Moerner W. E. , Orrit Michel
Published: July 2002

Efficient collection and detection of fluorescence coupled with careful minimization of background from impurities and Raman scattering now enable routine optical microscopy and study of single molecules in complex condensed matter environments. This ultimate method for unraveling ensemble averages leads to the observation of new effects and to direct measurements of stochastic fluctuations. Experiments at cryogenic temperatures open new directions in molecular spectroscopy, quantum optics, and solid-state dynamics. Room-temperature investigations apply several techniques (polarization microscopy, single-molecule imaging, emission time dependence, energy transfer, lifetime studies, and the like) to a growing array of biophysical problems where new insight may be gained from direct observations of hidden static and dynamic inhomogeneity.
Interaction of Diphtheria Toxin T Domain with Molten Globule-Like Proteins and Its Implications for Translocation
Authors: Ren Jianhua , Kachel Kelli , Kim Hyun , Malenbaum Susan E. , John R. , Collier  , London Erwin
Published: July 2002

The transmembrane (T) domain of diphtheria toxin has a critical role in the low pH–induced translocation of the catalytic domain (A chain) of the toxin across membranes. Here it is shown that at low pH, addition of proteins in a partly unfolded, molten globule-like conformation converted the T domain from a shallow membrane-inserted form to its transmembrane form. Fluorescence energy transfer demonstrated that molten globule-like proteins bound to the T domain. Thus, the T domain recognizes proteins that are partly unfolded and may function in translocation of the A chain as a transmembrane chaperone.
Prevention of Graft Versus Host Disease by Inactivation of Host Antigen-Presenting Cells
Authors: Shlomchik Warren D. , Couzens Matthew S. , Tang Cheng Bi , McNiff Jennifer , Robert Marie E. , Liu Jinli , Shlomchik Mark J. , Emerson Stephen G.
Published: July 2002

Graft versus host disease, an alloimmune attack on host tissues mounted by donor T cells, is the most important toxicity of allogeneic bone marrow transplantation. The mechanism by which allogeneic T cells are initially stimulated is unknown. In a murine allogeneic bone marrow transplantation model it was found that, despite the presence of numerous donor antigen-presenting cells, only host-derived antigen-presenting cells initiated graft versus host disease. Thus, strategies for preventing graft versus host disease could be developed that are based on inactivating host antigen-presenting cells. Such strategies could expand the safety and application of allogeneic bone marrow transplantation in treatment of common genetic and neoplastic diseases.
Peptide Antagonists of the Human Estrogen Receptor
Authors: Norris John D. , Paige Lisa A. , Christensen Dale J. , Chang Ching-Yi , Huacani Maria R. , Fan Daju , Hamilton Paul T. , Fowlkes Dana M. , McDonnell Donald P.
Published: July 2002

Estrogen receptor α transcriptional activity is regulated by distinct conformational states that are the result of ligand binding. Phage display was used to identify peptides that interact specifically with either estradiol- or tamoxifen-activated estrogen receptor α. When these peptides were coexpressed with estrogen receptor α in cells, they functioned as ligand-specific antagonists, indicating that estradiol-agonist and tamoxifen–partial agonist activities do not occur by the same mechanism. The ability to regulate estrogen receptor α transcriptional activity by targeting sites outside of the ligand-binding pocket has implications for the development of estrogen receptor α antagonists for the treatment of tamoxifen-refractory breast cancers.
Measurements of Past Ice Sheet Elevations in Interior West Antarctica
Authors: Ackert Robert P. , Barclay David J. , Borns Harold W. , Calkin Parker E. , Kurz Mark D. , Fastook James L. , Steig Eric J.
Published: July 2002

A lateral moraine band on Mount Waesche, a volcanic nunatak in Marie Byrd Land, provides estimates of past ice sheet surface elevations in West Antarctica. Helium-3 and chlorine-36 surface exposure ages indicate that the proximal part of the moraine, up to 45 meters above the present ice surface, was deposited about 10,000 years ago, substantially later than the maximum ice extent in the Ross Embayment. The upper distal part of the moraine may record multiple earlier ice sheet high stands. A nonequilibrium ice sheet model predicts a delay of several thousand years in maximum ice levels at Mount Waesche relative to the maximum ice extent in the Ross Sea. The glacial geologic evidence, coupled with the ice sheet model, indicates that the contribution of the Ross Sea sector of the West Antarctic Ice Sheet to Holocene sea level rise was only about 3 meters. These results eliminate West Antarctic ice as the principle source of the large meltwater pulse during the early Holocene.
Olfactory Reception in Invertebrates
Authors: Krieger Jürgen , Breer Heinz
Published: July 2002

Recent progress in understanding the principles and mechanisms in olfaction is the result of multidisciplinary research efforts that explored chemosensation by using a variety of model organisms. Studies on invertebrates, notably nematodes, insects, and crustaceans, to which diverse experimental approaches can be applied, have greatly helped elucidate various aspects of olfactory signaling. From the converging results of genetic, molecular, and physiological studies, a common set of chemosensory mechanisms emerges. Recognition and discrimination of odorants as well as chemo-electrical transduction and processing of olfactory signals appear to be mediated by fundamentally similar mechanisms in phylogenetically diverse animals. The common challenge of organisms to decipher the world of odors was apparently met by a phylogenetically conserved strategy. Thus, comparative studies should continue to provide important contributions toward an understanding of the sense of smell.
Northern Hemisphere Ice-Sheet Influences on Global Climate Change
Authors: Clark Peter U. , Alley Richard B. , Pollard David
Published: July 2002

Large ice sheets actively interact with the rest of the climate system by amplifying, pacing, and potentially driving global climate change over several time scales. Direct and indirect influences of ice sheets on climate cause changes in ocean surface temperatures, ocean circulation, continental water balance, vegetation, and land-surface albedo, which in turn cause additional feedbacks in the climate system and help to synchronize global climate change. The effect of the underlying geological substrate on ice-sheet dynamics may be the missing link in understanding the ice sheet–climate interactions that are integral to the middle Pleistocene transition; the 100,000-year climate cycle; high-amplitude, millennial-scale climate variability; and low–aspect ratio ice sheets of the Last Glacial Maximum.
Neutral, Single-Component Nickel (II) Polyolefin Catalysts That Tolerate Heteroatoms
Authors: Younkin Todd R. , Connor Eric F. , Henderson Jason I. , Friedrich Stefan K. , Grubbs Robert H. , Bansleben Donald A.
Published: July 2002

More than half of the 170 million metric tons of polymers produced each year are polyolefins. Current technology uses highly active cationic catalysts, which suffer from an inability to tolerate heteroatoms such as oxygen, nitrogen, and sulfur. These systems require scrupulously clean starting materials and activating cocatalysts. A family of catalysts has been developed whose members are tolerant of both heteroatoms and less pure starting materials. These heteroatom-tolerant neutral late transition metal complexes are in fact highly active systems that produce high-molecular-weight polyethylene, polymerize functionalized olefins, and require no cocatalyst.
Regulation of Absorption and ABC1-Mediated Efflux of Cholesterol by RXR Heterodimers
Authors: Repa J. J. , Turley S. D. , Lobaccaro J.-M. A. , Medina J. , Li L. , Lustig K. , Shan B. , Heyman R. A. , Dietschy J. M. , Mangelsdorf D. J.
Published: July 2002

Several nuclear hormone receptors involved in lipid metabolism form obligate heterodimers with retinoid X receptors (RXRs) and are activated by RXR agonists such as rexinoids. Animals treated with rexinoids exhibited marked changes in cholesterol balance, including inhibition of cholesterol absorption and repressed bile acid synthesis. Studies with receptor-selective agonists revealed that oxysterol receptors (LXRs) and the bile acid receptor (FXR) are the RXR heterodimeric partners that mediate these effects by regulating expression of the reverse cholesterol transporter, ABC1, and the rate-limiting enzyme of bile acid synthesis, CYP7A1, respectively. Thus, these RXR heterodimers serve as key regulators of cholesterol homeostasis by governing reverse cholesterol transport from peripheral tissues, bile acid synthesis in liver, and cholesterol absorption in intestine.
T-Independent Immune Response: New Aspects of B Cell Biology
Authors: Fagarasan Sidonia , Honjo Tasuku
Published: July 2002

Recent results emphasize the roles of T-independent antibody response in humoral defenses, for which B1 cells and marginal zone B cells are mostly responsible. We discuss how these cells are activated, migrate, and differentiate into antibody-producing cells in various lymphoid tissues. Based on recent findings in each of these areas of B cell biology, we propose a possible mechanism for peripheral tolerance of autoreactive B cells at target organs.
Proximity-Induced Superconductivity in DNA
Authors: Kasumov A. Yu. , Kociak M. , Guéron S. , Reulet B. , Volkov V. T. , Klinov D. V. , Bouchiat H.
Published: July 2002

Conductivity measurements on double-stranded DNA molecules deposited by a combing process across a submicron slit between rhenium/carbon metallic contacts reveal conduction to be ohmic between room temperature and 1 kelvin. The resistance per molecule is less than 100 kilohm and varies weakly with temperature. Below the superconducting transition temperature (1 kelvin) of the contacts, proximity-induced superconductivity is observed. These results imply that DNA molecules can be conducting down to millikelvin temperature and that phase coherence is maintained over several hundred nanometers.
Requirement of DNase II for Definitive Erythropoiesis in the Mouse Fetal Liver
Authors: Kawane Kohki , Fukuyama Hidehiro , Kondoh Gen , Takeda Junji , Ohsawa Yoshiyuki , Uchiyama Yasuo , Nagata Shigekazu
Published: July 2002

Mature erythrocytes in mammals have no nuclei, although they differentiate from nucleated precursor cells. The mechanism by which enucleation occurs is not well understood. Here we show that deoxyribonuclease II (DNase II) is indispensable for definitive erythropoiesis in mouse fetal liver. No live DNase II–null mice were born, owing to severe anemia. When mutant fetal liver cells were transferred into lethally irradiated wild-type mice, mature red blood cells were generated from the mutant cells, suggesting that DNase II functions in a non–cell-autonomous manner. Histochemical analyses indicated that the critical cellular sources of DNase II are macrophages present at the site of definitive erythropoiesis in the fetal liver. Thus, DNase II in macrophages appears to be responsible for destroying the nuclear DNA expelled from erythroid precursor cells.
Pharmacokinetics and Pharmacodynamics of Lumefantrine (Benflumetol) in Acute Falciparum Malaria
Authors: Ezzet F. , van Vugt M. , Nosten F. , Looareesuwan S. , White N. J.
Published: July 2002

The objective of this study was to conduct a prospective population pharmacokinetic and pharmacodynamic evaluation of lumefantrine during blinded comparisons of artemether-lumefantrine treatment regimens in uncomplicated multidrug-resistant falciparum malaria. Three combination regimens containing an average adult lumefantrine dose of 1,920 mg over 3 days (four doses) (regimen A) or 2,780 mg over 3 or 5 days (six doses) (regimen B or C, respectively) were given to 266 Thai patients. Detailed observations were obtained for 51 hospitalized adults, and sparse data were collected for 215 patients of all ages in a community setting. The population absorption half-life of lumefantrine was 4.5 h. The model-based median (5th and 95th percentiles) peak plasma lumefantrine concentrations were 6.2 (0.25 and 14.8) μg/ml after regimen A, 9.0 (1.1 and 19.8) μg/ml after regimen B, and 8 (1.4 and 17.4) μg/ml after regimen C. During acute malaria, there was marked variability in the fraction of drug absorbed by patients (coefficient of variation, 150%). The fraction increased considerably and variability fell with clinical recovery, largely because food intake was resumed; taking a normal meal close to drug administration increased oral bioavailability by 108% (90% confidence interval, 64 to 164) (
P
, 0.0001). The higher-dose regimens (B and C) gave 60 and 100% higher areas under the concentration-time curves (AUC), respectively, and thus longer durations for which plasma lumefantrine concentrations exceeded the putative in vivo MIC of 280 μg/ml (median for regimen B, 252 h; that for regimen C, 298 h; that for regimen A, 204 h [
P
, 0.0001]) and higher cure rates. Lumefantrine oral bioavailability is very dependent on food and is consequently poor in acute malaria but improves markedly with recovery. The high cure rates with the two six-dose regimens resulted from increased AUC and increased time at which lumefantrine concentrations were above the in vivo MIC.
Iron-Chelating Activity of Tetracyclines and Its Impact on the Susceptibility of
Actinobacillus actinomycetemcomitans
to These Antibiotics
Authors: Grenier Daniel , Huot Marie-Pierre , Mayrand Denis
Published: July 2002

Three tetracyclines (tetracycline, doxycycline, and minocycline) were found to possess iron-chelating activity in a colorimetric siderophore assay. Determination of MICs indicated that the activity of doxycycline against the periodontopathogen
Actinobacillus actinomycetemcomitans
was only slightly influenced by the presence of an excess of iron that likely saturates the antibiotic. On the other hand, the MICs of doxycycline and minocycline were significantly lower for
A. actinomycetemcomitans
cultivated under iron-poor conditions than under iron-rich conditions.
Pharmacokinetics of a Fluoronaphthyridone, Trovafloxacin (CP 99,219), in Infants and Children following Administration of a Single Intravenous Dose of Alatrofloxacin
Authors: Bradley John S. , Kearns Gregory L. , Reed Michael D. , Capparelli Edmund V. , Vincent John ,   The Pediatric Pharmacology Research Unit Network
Published: July 2002

The pharmacokinetics of trovafloxacin following administration of a single intravenous dose of alatrofloxacin, equivalent to 4 mg of trovafloxacin per kg of body weight, were determined in 6 infants (ages 3 to 12 months) and 14 children (ages, 2 to 12 years). There was rapid conversion of alatrofloxacin to trovafloxacin, with an average ± standard deviation (SD) peak trovafloxacin concentration determined at the end of the infusion of 4.3 ± 1.4 μg/ml. The primary pharmacokinetic parameters (average ± SD) analyzed were volume of distribution at steady state (1.6 ± 0.6 liters/kg), clearance (151 ± 82 ml/h/kg), and half-life (9.8 ± 2.9 h). The drug was well tolerated by all children. There were no age-related differences in any of the pharmacokinetic parameters studied. Less than 5% of the administered dose was excreted in the urine over 24 h. On the basis of the mean area under the concentration-time curve of 30.5 ± 10.1 μg · h/ml and the susceptibility (≤0.5 μg/ml) of common pediatric bacterial pathogens to trovafloxacin, dosing of 4 mg/kg/day once or twice daily should be appropriate.
Novel β-Lactamase Genes from Two Environmental Isolates of
Vibrio harveyi
Authors: Teo Jeanette W. P. , Suwanto Antonius , Poh Chit Laa
Published: July 2002

Two ampicillin-resistant (Amp
r
) isolates of
Vibrio harveyi
, W3B and HB3, were obtained from the coastal waters of the Indonesian island of Java. Strain W3B was isolated from marine water near a shrimp farm in North Java while HB3 was from pristine seawater in South Java. In this study, novel β-lactamase genes from W3B (
bla
VHW-1
) and HB3 (
bla
VHH-1
) were cloned and their nucleotide sequences were determined. An open reading frame (ORF) of 870 bp encoding a deduced protein of 290 amino acids (VHW-1) was revealed for the
bla
gene of strain W3B while an ORF of 849 bp encoding a 283-amino-acid protein (VHH-1) was deduced for
bla
VHH-1
. At the DNA level, genes for VHW-1 and VHH-1 have a 97% homology, while at the protein level they have a 91% homology of amino acid sequences. Neither gene sequence showed homology to any other β-lactamases in the databases. The deduced proteins were found to be class A β-lactamases bearing low levels of homology (<50%) to other β-lactamases of the same class. The highest level of identity was obtained with β-lactamases from
Pseudomonas aeruginosa
, i.e., PSE-1, PSE-4, and CARB-3, and
Vibrio cholerae
CARB-6. Our study showed that both strains W3B and HB3 possess an endogenous plasmid of approximately 60 kb in size. However, Southern hybridization analysis employing
bla
VHW-1
as a gene probe demonstrated that the
bla
gene was not located in the plasmid. A total of nine ampicillin-resistant
V. harveyi
strains, including W3B and HB3, were examined by pulsed-field gel electrophoresis of
Not
I-digested genomic DNA. Despite a high level of intrastrain genetic diversity, the
bla
VHW-1
probe hybridized only to an 80- or 160-kb
Not
I genomic fragment in different isolates.
Sublethal Concentrations of Pleurocidin-Derived Antimicrobial Peptides Inhibit Macromolecular Synthesis in
Escherichia coli
Authors: Patrzykat Aleksander , Friedrich Carol L. , Zhang Lijuan , Mendoza Valentina , Hancock Robert E. W.
Published: July 2002

Cationic bactericidal peptides are components of natural host defenses against infections. While the mode of antibacterial action of cationic peptides remains controversial, several targets, including the cytoplasmic membrane and macromolecular synthesis, have been identified for peptides acting at high concentrations. The present study identified peptide effects at lower, near-lethal inhibitory concentrations. An amidated hybrid of the flounder pleurocidin and the frog dermaseptin (P-Der), two other pleurocidin derivatives, and pleurocidin itself were studied. At 2 μg/ml, the MIC, P-Der inhibited
Escherichia coli
growth in a broth dilution assay but did not cause bacterial death within 30 min, as estimated by viable count analysis. Consistent with this, P-Der demonstrated a weak ability to permeabilize membranes but was able to translocate across the lipid bilayer of unilamellar liposomes. Doses of 20 μg/ml or more reduced bacterial viable counts by about 2 log orders of magnitude within 5 min after peptide treatment. Abrupt loss of cell membrane potential, observed with a fluorescent dye, dipropylthiacarbocyanine, paralleled bacterial death but did not occur at the sublethal, inhibitory concentrations. Both lethal and sublethal concentrations of P-Der affected macromolecular synthesis within 5 min, as demonstrated by incorporation of [
3
H]thymidine, [
3
H]uridine, and [
3
H]histidine, but the effects were qualitatively distinct at the two concentrations. Variations of the inhibition pattern described above were observed for pleurocidin and two other derivatives. Our results indicate that peptides at their lowest inhibitory concentrations may be less capable of damaging cell membranes, while they maintain their ability to inhibit macromolecular synthesis. Better understanding of the effects of peptides acting at their MICs will contribute to the design of new peptides effective at lower, less toxic concentrations.
Control of Expression of Divergent
Pseudomonas putida put
Promoters for Proline Catabolism
Authors: Vı́lchez Susana , Manzanera Maximino , Ramos Juan L.
Published: July 2002

Pseudomonas putida
KT2440 uses proline as the sole C and N source. Utilization of this amino acid involves its uptake, which is mediated by the PutP protein, and its conversion into glutamate, mediated by the PutA protein. Sequence analysis revealed that the
putA
and
putP
genes are transcribed divergently. Expression from the
putP
and
putA
genes was analyzed at the mRNA level in different host backgrounds in the absence and presence of proline. Expression from the
put
promoters was induced by proline. The transcription initiation points of the
putP
and
putA
genes were precisely mapped via primer extension, and sequence analysis of the upstream DNA region showed well-separated promoters for these two genes. The PutA protein acts as a repressor of
put
gene expression in
P. putida
because expression from the
put
promoters is constitutive in a host background with a knockout
putA
gene. This regulatory activity is independent of the catabolic activity of PutA, because we show that a point mutation (Glu896→Lys) that prevents catalytic activity allowed the protein to retain its regulatory activity. Expression from the
put
promoters in the presence of proline in a
putA
-proficient background requires a positive regulatory protein, still unidentified, whose expression seems to be ς
54
dependent because the
put
genes were not expressed in a ς
54
-deficient background. Expression of the
putA
and
putP
genes was equally high in the presence of proline in ς
38
- and
ihf
-deficient
P. putida
backgrounds.
Knockout of thep-Coumarate Decarboxylase Gene fromLactobacillus plantarumReveals the Existence of Two Other Inducible Enzymatic Activities Involved in Phenolic Acid Metabolism
Authors: Barthelmebs Lise , Divies Charles , Cavin Jean-François
Published: July 2002

Lactobacillus plantarumNC8 contains apdcgene coding forp-coumaric acid decarboxylase activity (PDC). A food grade mutant, designated LPD1, in which the chromosomalpdcgene was replaced with the deletedpdcgene copy, was obtained by a two-step homologous recombination process using an unstable replicative vector. The LPD1 mutant strain remained able to weakly metabolizep-coumaric and ferulic acids into vinyl derivatives or into substituted phenyl propionic acids. We have shown thatL. plantarumhas a second acid phenol decarboxylase enzyme, better induced with ferulic acid than withp-coumaric acid, which also displays inducible acid phenol reductase activity that is mostly active when glucose is added. Those two enzymatic activities are in competition forp-coumaric and ferulic acid degradation, and the ratio of the corresponding derivatives depends on induction conditions. Moreover, PDC appeared to decarboxylate ferulic acid in vitro with a specific activity of about 10 nmol · min−1· mg−1in the presence of ammonium sulfate. Finally, PDC activity was shown to confer a selective advantage on LPNC8 grown in acidic media supplemented withp-coumaric acid, compared to the LPD1 mutant devoid of PDC activity.
Input of Protein to Lake Water Microcosms Affects Expression of Proteolytic Enzymes and the Dynamics ofPseudomonasspp
Authors: Worm Jakob , Nybroe Ole
Published: July 2002

The objective of this study was to determine how an input of protein to lake water affects expression of a proteolytic potential and influences the abundance and composition of a specific group of bacteria.Pseudomonasspp. were chosen as a target group that can be recovered on selective growth media and contain both proteolytic and nonproteolytic strains. Amendment with 2 mg of casein per liter increased total proteinase activity (hydrolysis of [3H]casein) by 74%, leucine-aminopeptidase activity (hydrolysis of leucine-methyl-coumarinylamide) by 133%, bacterial abundance by 44%, and phytoplankton biomass (chlorophylla) by 39%. The casein amendment also increased the abundance of culturablePseudomonasspp. by fivefold relative to control microcosms but did not select for proteolytic isolates. Soluble proteins immunochemically related to thePseudomonas fluorescensalkaline proteinase, AprX, were detected in amended microcosms but not in the controls. The expression of this class of proteinase was confirmed exclusively for proteolyticPseudomonasisolates from the microcosms. The population structure ofPseudomonasisolates was determined from genomic fingerprints generated by universally primed PCR, and the analysis indicated that casein amendment led to only minor shifts in population structure. The appearance of AprX-like proteinases in the lake water might thus reflect a general induction of enzyme expression rather than pronounced shifts in thePseudomonaspopulation structure. The limited effect of casein amendment onPseudomonaspopulation structure might be due to the availability of casein hydrolysates to bacteria independent of their proteinase expression. In the lake water, 44% of the total proteinase activity was recovered in 0.22-μm-pore-size filtrates and thus without a direct association with the bacteria providing the extracellular enzyme activity. Since allPseudomonasisolates expressed leucine-aminopeptidase in pure culture, proteolytic as well as nonproteolytic pseudomonads were likely members of the bacterial consortium that metabolized protein in the lake water.
In Situ Detection, Isolation, and Physiological Properties of a Thin Filamentous Microorganism Abundant in Methanogenic Granular Sludges: a Novel Isolate Affiliated with a Clone Cluster, the Green Non-Sulfur Bacteria, Subdivision I
Authors: Sekiguchi Yuji , Takahashi Hiroki , Kamagata Yoichi , Ohashi Akiyoshi , Harada Hideki
Published: July 2002

We previously showed that very thin filamentous bacteria affiliated with the division green non-sulfur bacteria were abundant in the outermost layer of thermophilic methanogenic sludge granules fed with sucrose and several low-molecular-weight fatty acids (Y. Sekiguchi, Y. Kamagata, K. Nakamura, A. Ohashi, H. Harada, Appl. Environ. Microbiol. 65:1280–1288, 1999). Further 16S ribosomal DNA (rDNA) cloning-based analysis revealed that the microbes were classified within a unique clade, green non-sulfur bacteria (GNSB) subdivision I, which contains a number of 16S rDNA clone sequences from various environmental samples but no cultured representatives. To investigate their function in the community and physiological traits, we attempted to isolate the yet-to-be-cultured microbes from the original granular sludge. The first attempt at isolation from the granules was, however, not successful. In the other thermophilic reactor that had been treating fried soybean curd-manufacturing wastewater, we found filamentous microorganisms to outgrow, resulting in the formation of projection-like structures on the surface of granules, making the granules look like sea urchins. 16S rDNA-cloning analysis combined with fluorescent in situ hybridization revealed that the projections were comprised of the uncultured filamentous cells affiliated with the GNSB subdivision I and
Methanothermobacter
-like cells and the very ends of the projections were comprised solely of the filamentous cells. By using the tip of the projection as the inoculum for primary enrichment, a thermophilic, strictly anaerobic, filamentous bacterium, designated strain UNI-1, was successfully isolated with a medium supplemented with sucrose and yeast extract. The strain was a very slow growing bacterium which is capable of utilizing only a limited range of carbohydrates in the presence of yeast extract and produced hydrogen from these substrates. The growth was found to be significantly stimulated when the strain was cocultured with a hydrogen-utilizing methanogen,
Methanothermobacter thermautotrophicus
, suggesting that the strain is a sugar-fermenting bacterium, the growth of which is dependent on hydrogen consumers in the granules.
Phylogenetic and Functional Heterogeneity of Sediment Biofilms along Environmental Gradients in a Glacial Stream
Authors: Battin Tom J. , Wille Anton , Sattler Birgit , Psenner Roland
Published: July 2002

We used in situ hybridization with fluorescently labeled rRNA-targeted oligonucleotide probes concurrently with measurements of bacterial carbon production, biomass, and extracellular polymeric substances (EPS) to describe the bacterial community in sediments along a glacial stream. The abundance of sediment-associated
Archaea
, as detected with the ARCH915 probe, decreased downstream of the glacier snout, and a major storm increased their relative abundance by a factor of 5.5 to 7.9. Bacteria of the
Cytophaga-Flavobacterium
group were also sixfold to eightfold more abundant in the storm aftermath. Furthermore, elevated numbers of
Archaea
and members of the
Cytophaga-Flavobacterium
group characterized the phylogenetic composition of the supraglacial ice community. We postulate that glacial meltwaters constitute a possible source of allochthonous bacteria to the stream biofilms. Although stream water temperature increased dramatically from the glacier snout along the stream (3.5 km), sediment chlorophyll
a
was the best predictor for bacterial carbon production and specific growth rates along the stream. Concomitant with an increase in sediment chlorophyll
a
, the EPS carbohydrate-to-bacterial-cell ratio declined 11- to 15-fold along the stream prior to the storm, which is indicative of a larger biofilm matrix in upstream reaches. We assume that a larger biofilm matrix is required to assure prolonged transient storage and enzymatic processing of allochthonous macromolecules, which are likely the major substrate for microbial heterotrophs. Bacteria of the
Cytophaga-Flavobacterium
cluster, which are well known to degrade complex macromolecules, were most abundant in these stream reaches. Downstream, higher algal biomass continuously supplies heterotrophs with easily available exudates, therefore making a larger matrix unnecessary. As a result, bacterial carbon production and specific growth rates were higher in downstream reaches.
Biodegradation of
n
-Alkylcycloalkanes and
n
-Alkylbenzenes via New Pathways in
Alcanivorax
sp. Strain MBIC 4326
Authors: Dutta Tapan K. , Harayama Shigeaki
Published: July 2002

The degradation of long-chain
n
-alkylbenzenes and
n
-alkylcyclohexanes by
Alcanivorax
sp. strain MBIC 4326 was investigated. The alkyl side chain of these compounds was mainly processed by β-oxidation. In the degradation of
n
-alkylcyclohexanes, cyclohexanecarboxylic acid was formed as an intermediate. This compound was further transformed to benzoic acid via 1-cyclohexene-1-carboxylic acid.
Cyt1Ab1 and Cyt2Ba1 from
Bacillus thuringiensis
subsp.
medellin
and
B. thuringiensis
subsp
. israelensis
Synergize
Bacillus sphaericus
against
Aedes aegypti
and Resistant
Culex quinquefasciatus
(Diptera: Culicidae)
Authors: Wirth Margaret C. , Delécluse Armelle , Walton William E.
Published: July 2002

The interaction of two cytolytic toxins, Cyt1Ab from
Bacillus thuringiensis
subsp.
medellin
and Cyt2Ba from
Bacillus thuringiensis
subsp.
israelensis
, with
Bacillus sphaericus
was evaluated against susceptible and resistant
Culex quinquefasciatus
and the nonsensitive species
Aedes aegypti
. Mixtures of
B. sphaericus
with either cytolytic toxin were synergistic, and
B. sphaericus
resistance in
C. quinquefasciatus
was suppressed from >17,000- to 2-fold with a 3:1 mixture of
B. sphaericus
and Cyt1Ab. This trait may prove useful for combating insecticide resistance and for improving the activity of microbial insecticides.
Decrease in Cell Surface Galactose Residues of
Schizosaccharomyces pombe
Enhances Its Coflocculation with
Pediococcus damnosus
Authors: Peng Xuan , Sun Jun , Michiels Chris , Iserentant Dirk , Verachtert Hubert
Published: July 2002

Pediococcus damnosus
can coflocculate with
Saccharomyces cerevisiae
and cause beer acidification that may or may not be desired. Similar coflocculations occur with other yeasts except for
Schizosaccharomyces pombe
which has galactose-rich cell walls. We compared coflocculation rates of
S. pombe
wild-type species TP4-1D, having a mannose-to-galactose ratio (Man:Gal) of 5 to 6 in the cell wall, with its glycosylation mutants
gms1-1
(Man:Gal = 5:1) and
gms1
Δ (Man:Gal = 1:0). These mutants coflocculated at a much higher level (30 to 45%) than that of the wild type (5%). Coflocculation of the mutants was inhibited by exogenous mannose but not by galactose
.
The
S. cerevisiae mnn2
mutant, with a mannan content similar to that of
gms1
Δ, also showed high coflocculation (35%) and was sensitive to mannose inhibition. Coflocculation of
P. damnosus
and
gms1
Δ (or
mnn2
) also could be inhibited by
gms1
Δ mannan (with unbranched α-1,6-linked mannose residues), concanavalin A (mannose and glucose specific), or NPA lectin (specific for α-1,6-linked mannosyl units). Protease treatment of the bacterial cells completely abolished coflocculation. From these results we conclude that mannose residues on the cell surface of
S. pombe
serve as receptors for a
P. damnosus
lectin but that these receptors are shielded by galactose residues in wild-type strains. Such interactions are important in the production of Belgian acid types of beers in which mixed cultures are used to improve flavor.
Isolation of Toxigenic
Nocardiopsis
Strains from Indoor Environments and Description of Two New
Nocardiopsis
Species,
N. exhalans
sp. nov. and
N. umidischolae
sp. nov
Authors: Peltola Joanna S. P. , Andersson Maria A. , Kämpfer Peter , Auling Georg , Kroppenstedt Reiner M. , Busse Hans-Jürgen , Salkinoja-Salonen Mirja S. , Rainey Frederick A.
Published: July 2002

Nocardiopsis
strains were isolated from water-damaged indoor environments. Two strains (
N. alba
subsp.
alba
704a and a strain representing a novel species, ES10.1) as well as strains of
N. prasina, N. lucentensis,
and
N. tropica
produced methanol-soluble toxins that paralyzed the motility of boar spermatozoa at <30 μg of crude extract (dry weight) ml
−1
.
N. prasina, N. lucentensis, N. tropica
, and strain ES10.1 caused cessation of motility by dissipating the mitochondrial membrane potential, Δψ, of the boar spermatozoa. Indoor strain 704a produced a substance that destroyed cell membrane barrier function and depleted the sperm cells of ATP. Indoor strain 64/93 was antagonistic towards
Corynebacterium renale
. Two indoor
Nocardiopsis
strains were xerotolerant, and all five utilized a wide range of substrates. This combined with the production of toxic substances suggests good survival and potential hazard to human health in water-damaged indoor environments. Two new species,
Nocardiopsis exhalans
sp. nov. (ES10.1
T
) and
Nocardiopsis umidischolae
sp. nov. (66/93
T
), are proposed based on morphology, chemotaxonomic and physiological characters, phylogenetic analysis, and DNA-DNA reassociations.
The Aquatic Budding Bacterium
Blastobacter denitrificans
Is a Nitrogen-Fixing Symbiont of
Aeschynomene indica
Authors: van Berkum Peter , Eardly Bertrand D.
Published: July 2002

Blastobacter
spp. are freshwater bacteria that form rosette structures by cellular attachment to a common base. Comparative analyses of ribosomal 16S rRNA gene and internally transcribed spacer region sequences indicated that
B. denitrificans
is a member of the α-subdivision of
Proteobacteria
. Among the α-
Proteobacteria
,
B. denitrificans
was related to a cluster of genera, including
Rhodopseudomonas palustris
,
Afipia felis
,
Nitrobacter hamburgensis
, and
Bradyrhizobium
spp. Although the precise phylogenetic relationships among these genera could not be established with a high degree of confidence, the sequences of
B. denitrificans
and several bradyrhizobial isolates from nodules of
Aeschynomene indica
were almost identical. Bradyrhizobia are bacteria that form nitrogen-fixing symbioses with legumes, including soybeans (
Glycine max
) and members of the genus
Aeschynomene
. From symbiotic infectiveness tests we demonstrated that the type strain for
B. denitrificans
, IFAM 1005, was capable of forming an effective nitrogen-fixing symbiosis with
A. indica
. Not only do these results reveal a previously unknown ecological adaptation of a relatively obscure aquatic bacterium, but they also demonstrate how evidence gathered from molecular systematic analyses can sometimes provide clues for predicting ecological behavior.
Double-Stranded RNA Mycovirus from
Fusarium graminearum
Authors: Chu Yeon-Mee , Jeon Jae-Jin , Yea Sang-Jin , Kim Yong-Ho , Yun Sung-Hwan , Lee Yin-Won , Kim Kook-Hyung
Published: July 2002

Double-stranded RNA (dsRNA) viruses in some fungi are associated with hypovirulence and have been used or proposed as biological control agents. We isolated 7.5-kb dsRNAs from 13 of 286 field strains of
Fusarium graminearum
isolated from maize in Korea. One of these strains, DK21, was examined in more detail. This strain had pronounced morphological changes, including reduction in mycelial growth, increased pigmentation, reduced virulence towards wheat, and decreased (60-fold) production of trichothecene mycotoxins. The presence or absence of the 7.5-kb dsRNA was correlated with the changes in pathogenicity and morphology. The dsRNA could be transferred to virus-free strains by hyphal fusion, and the recipient strain acquired the virus-associated phenotype of the donor strain. The dsRNA was transmitted to approximately 50% of the conidia, and only colonies resulting from conidia carrying the mycovirus had the virus-associated phenotype. Partial nucleotide sequences of the purified dsRNA identify an RNA-dependent RNA polymerase sequence and an ATP-dependent helicase that are closely related to those of
Cryphonectria
hypovirus and
Barley yellow mosaic virus
. Collectively, these results suggest that this dsRNA isolated from
F. graminearum
encodes traits for hypovirulence.
Reversible and Irreversible Adhesion of Motile
Escherichia coli
Cells Analyzed by Total Internal Reflection Aqueous Fluorescence Microscopy
Authors: Vigeant Margot A.-S. , Ford Roseanne M. , Wagner Michael , Tamm Lukas K.
Published: July 2002

The initial events in bacterial adhesion are often explained as resulting from electrostatic and van der Waals forces between the cell and the surface, as described by DLVO theory (developed by Derjaguin, Landau, Verwey, and Overbeek). Such a theory predicts that negatively charged bacteria will experience greater attraction toward a negatively charged surface as the ionic strength of the medium is increased. In the present study we observed both smooth-swimming and nonmotile
Escherichia coli
bacteria close to plain, positively, and hydrophobically coated quartz surfaces in high- and low-ionic-strength media by using total internal reflection aqueous fluorescence microscopy. We found that reversibly adhering cells (cells which continue to swim along the surface for extended periods) are too distant from the surface for this behavior to be explained by DLVO-type forces. However, cells which had become immobilized on the surface did seem to be affected by electrostatic interactions. We propose that the “force” holding swimming cells near the surface is actually the result of a hydrodynamic effect, causing the cells to swim at an angle along the glass, and that DLVO-type forces are responsible only for the observed immobilization of irreversibly adhering cells. We explain our observations within the context of a conceptual model in which bacteria that are interacting with the surface may be thought of as occupying one of three compartments: bulk fluid, near-surface bulk, and near-surface constrained. A cell in these compartments feels either no effect of the surface, only the hydrodynamic effect of the surface, or both the hydrodynamic and the physicochemical effects of the surface, respectively.
New Automated Immunoassay Measuring Immunoglobulin A Antigliadin Antibodies for Prediction of Celiac Disease in Childhood
Authors: Grodzinsky E. , Ivarsson A. , Juto P. , Olcén P. , Fälth-Magnusson K. , Persson L. Å. , Hernell O.
Published: July 2002

The prevalence of celiac disease (CD) in Sweden is about 4 cases per 1,000 people. Screening for CD with serological tests indicates similar high prevalences in many other countries. Between 1 November 1992 and 30 April 1995, 133 children (9 months to 16.7 years of age) with suspected CD were studied. The predictive value (PV) of immunoglobulin A antigliadin antibodies (IgA-AGA) in the serum as assayed with two new commercial automated immunoassays—the Pharmacia CAP System Gliadin IgA FEIA (CAP) and the UNICAP-100 (UNICAP)—and with three “in-house” methods was evaluated using assessment of the small intestinal mucosa morphology as the “gold standard.” All serum samples were analyzed for total serum IgA. At presentation the diagnostic sensitivities and specificities of the different tests varied from 0.72 to 0.88 and 0.67 to 0.87, respectively. All methods showed a higher sensitivity for CD in younger children. The area under each assay's receiver operating characteristic curve was calculated and varied between 0.82 and 0.89. The positive and negative PVs for the CAP and UNICAP, which were assays with a high sensitivity and a high specificity, respectively, were estimated. In the clinically selected population (prevalence of CD, 1 in 3) the positive PV was about 55%, and in the general population (prevalence, 1 in 250) it was about 1%. The negative PVs for both CAP and UNICAP were close to 100%; thus, when the AGA test was negative, the risk for CD was small. Interestingly, five children had serum IgA levels below the detection limit (<0.07 g/liter) when on a gluten-free diet, whereas they had normal levels at the time of the first biopsy. In conclusion, the automated immunoassays—based on ImmunoCAP technology—for analysis of IgA-AGA had a reliability comparable to that of the in-house methods.
Peptide Mimic of Phosphorylcholine, a Dominant Epitope Found onStreptococcus pneumoniae
Authors: Harris Shannon L. , Park Moon K. , Nahm Moon H. , Diamond Betty
Published: July 2002

Even in the age of antibiotics,Streptococcus pneumoniaecauses significant morbidity, especially in the young, the elderly, and the immunocompromised. While a carbohydrate-based vaccine exists, it is poorly immunogenic in the at-risk populations. In mice, antibodies directed against phosphorylcholine (PC), an epitope present on the cell wall C polysaccharide of all pneumococcal serotypes, protect against infection. However, PC itself is a poor vaccine candidate. We report here peptide mimics of PC based on the anti-idiotypic interaction of T15 anti-PC antibodies. T15 antibodies, the dominant and protective idiotype induced in mice by PC immunization, self-associate via a 24-amino-acid region in the PC binding site (ASRNKANDYTTEYSASVKGRFIVS; peptide 1). Peptide 1 has been shown to bind in the PC binding site. We demonstrated that amino acid sequences derived from peptide 1 starting at amino acid 9, 11, or 13 inhibit PC binding. Therefore, we immunized mice with bovine serum albumin (BSA) conjugates of peptide 1 or either of two selected 12-mers. The 12-mer peptides were not immunogenic. Mice immunized with peptide 1-BSA developed an anti-PC response consisting mainly immunoglobulin G1 and expressed the T15 heavy chain. Nonetheless, neither BALB/c nor CBA/N mice were protected from lethal pneumococcal infections by immunization with peptide 1-BSA. Preliminary data suggest that peptide 1-BSA is not able to elicit the canonical T15 light chain, explaining the absence of protection. This idiotype-derived mimotope of PC is a useful tool for understanding immunologic cross-reactivity and learning to design T-cell-dependent vaccines forS. pneumoniae.
Isolation and Characterization of a
Shigella flexneri
Invasin Complex Subunit Vaccine
Authors: Turbyfill K. Ross , Hartman Antoinette B. , Oaks Edwin V.
Published: July 2002

The invasiveness and virulence of
Shigella
spp. are largely due to the expression of plasmid-encoded virulence factors, among which are the invasion plasmid antigens (Ipa proteins). After infection, the host immune response is directed primarily against lipopolysaccharide (LPS) and the virulence proteins (IpaB, IpaC, and IpaD). Recent observations have indicated that the Ipa proteins (IpaB, IpaC, and possibly IpaD) form a multiprotein complex capable of inducing the phagocytic event which internalizes the bacterium. We have isolated a complex of invasins and LPS from water-extractable antigens of virulent shigellae by ion-exchange chromatography. Western blot analysis of the complex indicates that all of the major virulence antigens of
Shigella
, including IpaB, IpaC, and IpaD, and LPS are components of this macromolecular complex. Mice or guinea pigs immunized intranasally with purified invasin complex (invaplex), without any additional adjuvant, mounted a significant immunoglobulin G (IgG) and IgA antibody response against the
Shigella
virulence antigens and LPS. The virulence-specific response was very similar to that previously noted in primates infected with shigellae. Guinea pigs (keratoconjunctivitis model) or mice (lethal lung model) immunized intranasally on days 0, 14, and 28 and challenged 3 weeks later with virulent shigellae were protected from disease (
P
< 0.01 for both animal models).