bluuebunny/crossref_metadata_2025 huggingface dataset:
Is flow cytometry accurate enough to screen platelet autoantibodies?
Authors: Hézard Nathalie , Simon Gérard , Macé Catherine , Jallu Vincent , Kaplan Cécile , Nguyen Philippe
Published: February 2008

BACKGROUND: The diagnosis of immune thrombocytopenic purpura (ITP) is a diagnosis of exclusion, as stated by international guidelines. Nevertheless, the assessment of platelet (PLT) antibodies has been reported as helpful for the diagnosis and the follow‐up of ITP patients. PLT antibodies are detected by highly specialized assays, such as monoclonal antibody–specific immobilization of PLT antigen (MAIPA) test. Flow cytometry for PLT‐associated immunoglobulin G (PAIgG) detection has been described more recently. This study was meant to evaluate the utility of flow cytometry to screen accurately patients needing further MAIPA testing.

STUDY DESIGN AND METHODS: PAIgG, PAIgM, and PAIgA were determined in 107 consecutive patients and in 147 healthy controls in parallel. MAIPA testing was performed in all patients. The accuracy of flow cytometry was assessed with a receiver operating characteristics (ROC) curve analysis versus MAIPA.

RESULTS: MAIPA assay found PLT‐specific IgG in 27 patients (25%). The ROC curve analysis showed that no false‐negative result in flow cytometry was obtained for a mean fluorescence intensity (MFI) cutoff of 0.2. With this cutoff, PAIgG were positive in 61 patients (57%). In this series, MAIPA was unnecessary in 42 percent of patients (corresponding to true‐negative results). When MAIPA was positive, PAIgM values ranged from 0.1 to 1.0, and PAIgA from 0.1 to 2.

CONCLUSION: Flow cytometry for PAIgG assessment may be used to accurately decide whether or not MAIPA must be subsequently performed. In this series, MAIPA was unnecessary in 42 percent of patients. Moreover, PAIgM results suggested that its determination combined with PAIgG may be of interest in ITP investigation.
Enhancing attitudes and intentions in prospective blood donors: evaluation of a new donor recruitment brochure
Authors: France Christopher R. , Montalva Roen , France Janis L. , Trost Zina
Published: February 2008

BACKGROUND: Although little empiric evidence has been published concerning the efficacy of blood donor recruitment materials, research suggests that simple attempts to enhance knowledge may not be sufficient to motivate donation. In contrast, recent donor motivation studies highlight the importance of anxiety, attitudes, and perceived ability to cope with donation (i.e., self‐efficacy) as crucial determinants of donation intention. Therefore, recruitment materials that specifically address these constructs have the potential to outperform traditional educational brochures.

STUDY DESIGN AND METHODS: Participants were randomly assigned to read one of three brochures: 1) a new brochure addressing common donor concerns and suggesting specific coping strategies, 2) a standard blood center brochure, or 3) a control brochure on healthy eating and exercise. Standardized questionnaires were completed before and after the brochures to assess change in blood donation anxiety, attitude, self‐efficacy, and intention.

RESULTS: Although no significant changes were noted for the control brochure, after reading the new brochure participants reported significant improvements in attitude, anxiety, self‐efficacy, and donation intention. The standard donation brochure had an intermediate effect.

CONCLUSION: Efforts to address common donor fears and to provide useful coping suggestions may improve the effectiveness of blood donation recruitment materials.
Immunomagnetic selection of progenitor cells from peripheral blood after thawing with an automatic system in a pediatric patient with a neuroblastoma
Authors: Zinno Francesco , Landi Fabiola , Aureli Viviana , Donfrancesco Alberto , Isacchi Giancarlo
Published: February 2008

BACKGROUND: Immunomagnetic selection of peripheral blood progenitor cells (PBPCs) in patients with tumoral infiltration in marrow makes it possible to reduce contamination of cellular concentrates, but this procedure cannot always be used, mainly because of the low cellular count in apheresis concentrates.

STUDY DESIGN AND METHODS: In this case two cellular concentrates taken separately at two different times were selected and cryopreserved; they were thawed with an automatic instrument.

RESULTS: After manipulation, a selected concentrate containing 24.16 × 106 CD34+ cells with a purity of 90.15 percent was obtained; vitality after thawing and selection was 88 and 96 percent, respectively. The engraftment was achieved on Day +17 from the infusion of the previously selected PBPCs, as the literature also shows us.

CONCLUSION: The time passed between the infusion and the engraftment gives us evidence of the efficacy of immunomagnetic selection carried out after thawing 2 cell units that were collected at different times from the same patient. In this way, it has been possible to perform an autologous transplant in a patient in which CD34+ cells transplant is recommended, but from whom the number of collected cells after a single mobilization cycle would not have been sufficient for the engraftment.
Buffy‐coat platelet variables and metabolism during storage in additive solutions or plasma
Authors: Zhang Jerry G. , Carter Cedric J. , Culibrk Brankica , Devine Dana V. , Levin Elena , Scammell Kenneth , Weiss Sandra , Gyongyossy‐Issa Maria I.C.
Published: February 2008

BACKGROUND: Buffy‐coat processing allows for the use of platelet additive solutions (PASs). PASs reduce plasma‐associated transfusion reactions and conserve plasma for transfusion or fractionation. Platelet (PLT) storage in plasma was compared to storage in three commercially available PASs compared to assess their influence on in vitro laboratory variables.

STUDY DESIGN AND METHODS: Platelet concentrates (PCs) were prepared from leukoreduced pools of four buffy coats (BCPs) suspended in autologous plasma or one of PASs (Composol, Fresenius‐Kabi; T‐Sol, Baxter Corp.; or SSP+, MacoPharma). On Days 1, 2, 3, 5, and 7 of storage, samples were tested for PLT concentration, mean PLT volume (MPV), CD62P, morphology, pO2, pCO2, glucose, lactate and total protein concentration, pH, extent of shape change (ESC), and hypotonic shock response (HSR). Data were analyzed by analysis of variance (ANOVA) with repeated measures and t tests.

RESULTS: PLT recoveries from BCPs were higher (p < 0.05) with plasma than any PAS. Storage medium and duration did not affect PLT concentration or MPV over time. CD62P expression and morphology were significantly different among PCs pooled with different media. ANOVA showed (p < 0.05) differences among the rates of change of pCO2, pH, glucose consumption, lactate production, and ESC; PASs such as Composol and SSP+ offered excellent maintenance of pH and low rates of glucose consumption. PAS performed poorly in ESC and HSR compared to plasma. Correlation studies reveal far more significant correlations between variables of PLTs in PAS than in plasma.

CONCLUSION: Newer PASs, for example, SSP+ and Composol, can maintain PLT integrity and moderate metabolism similarly to plasma but offer consistently lower PLT recoveries and limited osmotic balance.
Isolation and phytotoxicity of a metabolite from Curvularia eragrostidis and characterisation of its modes of action
Authors: Jiang S.‐J. , Qiang S. , Zhu Y.‐Z. , Dong Y.‐F.
Published: February 2008

Digitaria sanguinalis is a widespread troublesome weed distributed all over the world. Curvularia eragrostidis QZ‐2000 is a potential candidate for biocontrol of D. sanguinalis. A phytotoxic metabolite from the culture filtrate of this fungus was extracted by ethyl acetate, isolated by bioassay‐guided column chromatography and thin layer chromatography on silica gel, characterised by ultra violet, infrared ray (IR), mass spectrometry (MS), 1H‐nuclear magnetic resonance (NMR) and 13C‐NMR spectral data analyses and identified as α,β‐dehydrocurvularin. The phytotoxin significantly inhibited seed germination of D. sanguinalis from 43 to 688 μM. The EC50 value of seed germination was 152 μM. The EC50 values of elongation of radicle and coleoptile were 102 and 172 μM, respectively. α,β‐dehydrocurvularin caused extensive necrosis on leaves of many notorious weeds at 688 μM, while maize and soybean were insensitive to it. Therefore, α,β‐dehydrocurvularin was regarded as a non‐host‐selective phytotoxin. At concentrations of 172–688 μM, α,β‐dehydrocurvularin caused a decrease in chlorophyll content. α,β‐dehydrocurvularin had stronger impacts on chlorophyll A fluorescence, photophosphorylation and Mg2+‐ATPase activity at higher concentrations. These results suggest that α,β‐dehydrocurvularin affected the photosynthetic capacity. In the present study, α,β‐dehydrocurvularin significantly inhibited mitosis of root tip cells. It supports that the α,β‐dehydrocurvularin has potential for development as a natural bioherbicide.
Successional Change and Resilience of a Very Dry Tropical Deciduous Forest Following Shifting Agriculture
Authors: Lebrija‐Trejos Edwin , Bongers Frans , Pérez‐García Eduardo A. , Meave Jorge A.
Published: February 2008

We analyzed successional patterns in a very dry tropical deciduous forest by using 15 plots differing in age after abandonment and contrasted them to secondary successions elsewhere in the tropics. We used multivariate ordination and nonlinear models to examine changes in composition and structure and to estimate forest recovery rates and resilience. A shrub phase characterized early succession (0–3 yr); afterwards, the tree Mimosa acantholoba became dominant. Below its canopy, sprouts and seed‐regenerated individuals of mature forest species slowly accumulated. Canopy height, plant density, and crown cover stabilized in less than 15 yr, whereas species richness, diversity, and basal area continued to increase. The pioneer species group has very low diversity and the long‐lived pioneer phase typical of humid forests is absent; species composition may therefore recover soon as suggested by convergence toward mature forest species composition. The time trend of plant density also differed from humid forests for it lacked its characteristic density decline, presumably because of differences in regeneration mechanisms between very dry and other less water‐stressed forest types. As opposed to the prevailing hypothesis, resilience was not higher than in moister forests, and thus factors other than structure relative simplicity must be accounted for when assessing resilience.
Conservation Genetics ofJacquemontia reclinata(Convolvulaceae), an Endangered Species from Southern Florida: Implications for Restoration Management
Authors: Thornton Hannah E. B. , Roncal Julissa , Lewis Carl E. , Maschinski Joyce , Francisco‐Ortega Javier
Published: February 2008

Guidelines designed to aid in the restoration of rare species have been previously proposed using two primary strategies to select individuals for augmentation and reintroduction: mixing progeny from different populations or separating individuals from different populations. Understanding the genetic structure and diversity of an endangered species can offer insights into conservation management strategies. We used random amplified polymorphic DNA markers to assess the genetic structure and diversity ofJacquemontia reclinata, a federally endangered species endemic to Southeastern Florida. We sampled 20 percent of total number of individuals from eight of the ten known wild populations. Across individuals high levels of polymorphic loci (94.7%) were found and larger populations had greater genetic diversity. Cluster and ordination analyses found that one population was genetically differentiated from all the others; this population grows in a unique habitat. Most genetic variation (77.5%) was found within populations, and genetic distances between populations were not explained by their geographic distances. We recommend the use of two management units in restoration programs forJ. reclinata, one consisting of the genetically differentiated population and the second consisting of the other seven populations sampled.
Potential and Pitfalls of Applying Theory to the Practice of Financial Education
Authors: LYONS ANGELA C. , NEELAKANTAN URVI
Published: February 2008

Researchers are increasingly using interdisciplinary theory to bring rigor to the practice of financial education. Practitioners often do not see the value of the theory because it does not coincide with their observations of how people behave, and researchers do not yet have enough experience with interdisciplinary theory to demonstrate its usefulness to practitioners. If carefully applied, theory can be used to set appropriate financial goals and to positively change consumers’ financial behaviors. Better communication can bridge the gap between theory and practice to the benefit of the consumer.
Measuring What Really Matters to Consumers
Authors: MAYER ROBERT N.
Published: February 2008

Virtually every major interest group except consumers has developed a ranking system or report card that can be used to galvanize the public and pressure policy makers for change. This article discusses the potential value of consumer‐relevant indices that permit international comparisons and addresses some of the challenges to constructing these measures.
A Different Perspective to Study the Effect of Freeze, Air, and Osmotic Drying on Oil Absorption during Potato Frying
Authors: Moreno M.C. , Bouchon P.
Published: February 2008

ABSTRACT:  The objective of this article is to assess the effect of different dehydration pretreatments on oil absorption and illustrate how the different ways results can be reported may even drive to opposite conclusions. To do so, potato cylinders were blanched in hot water and dried until a moisture content of 62% (w.b.) by either freeze‐drying, air drying, and osmotic drying with a sucrose solution or osmotic drying with a NaCl solution. Control (blanched) and dried potatoes were deep‐fat fried at 170 °C for time periods between 1 and 5 min. Water removal and total oil uptake were determined and 2 oil fractions were distinguished: superficial oil and penetrated oil. Compared to the control, freeze dried samples increased oil uptake in 15.4% (d.b.) whereas air‐dried samples reduced it in 11.2% (d.b.). Similarly, osmotic dehydrated samples showed a high reduction in oil uptake compared to the control (up to 27%[d.b.] when using a sucrose solution). However, this high decrease in oil absorption was attributed to the increase in solids content occurring during the osmotic dehydration process rather than a reduction in the amount of oil taken up. In fact, when the amount of oil absorbed per cylinder was determined, it was verified that oil uptake of osmotically dehydrated samples was even higher than the control, as opposed to what has been previously reported in the literature. These results highlight the importance of selecting an adequate basis to carry out comparisons properly.
Sex differences in contractile properties and fatigue resistance of human skeletal muscle
Authors: Wüst Rob C. I. , Morse Christopher I. , De Haan Arnold , Jones David A. , Degens Hans
Published: February 2008

To explore the cause of higher skeletal muscle fatigue resistance in women than men, we used electrically evoked contractions (1 s on, 1 s off, 30 Hz, 2 min), which circumvent motivational bias and allow examination of contractile properties. We compared 29 men [26.5 (7.0) years old; mean (s.d.)] with 35 women [25.4 (7.6) years old]. Strength of the quadriceps muscle was higher in men than women (P < 0.001). The lower maximal rate of relaxation in women (P= 0.002) indicates that their muscles were slower than those of men. The torque declined less in women than in men [37.7 (10.7) versus 29.9 (10.0)%; P= 0.002], and was not related to muscle strength or size, as determined with magnetic resonance imaging. The sex difference in fatigability was also seen when the circulation to the leg was occluded [torque declined 76.9 (10.8) versus 59.5 (16.9)% in men versus women, respectively; P= 0.008]. The maximal rate of relaxation correlated with the fatigability of the muscle under all conditions (correlations ranging from 0.34 to 0.51, P < 0.02). We conclude that the sex‐related difference in skeletal muscle fatigue resistance is not explicable by differences in motivation, muscle size, oxidative capacity and/or blood flow between sexes, but might be related to differences in fibre type composition.
Intact Membrane Lipids of “
Candidatus
Nitrosopumilus maritimus,” a Cultivated Representative of the Cosmopolitan Mesophilic Group I Crenarchaeota
Authors: Schouten Stefan , Hopmans Ellen C. , Baas Marianne , Boumann Henry , Standfest Sonja , Könneke Martin , Stahl David A. , Sinninghe Damsté Jaap S.
Published: February 2008

In this study we analyzed the membrane lipid composition of “
Candidatus
Nitrosopumilus maritimus,” the only cultivated representative of the cosmopolitan group I crenarchaeota and the only mesophilic isolate of the phylum
Crenarchaeota
. The core lipids of “
Ca
. Nitrosopumilus maritimus” consisted of glycerol dialkyl glycerol tetraethers (GDGTs) with zero to four cyclopentyl moieties. Crenarchaeol, a unique GDGT containing a cyclohexyl moiety in addition to four cyclopentyl moieties, was the most abundant GDGT. This confirms unambiguously that crenarchaeol is synthesized by species belonging to the group I.1a crenarchaeota. Intact polar lipid analysis revealed that the GDGTs have hexose, dihexose, and/or phosphohexose head groups. Similar polar lipids were previously found in deeply buried sediments from the Peru margin, suggesting that they were in part synthesized by group I crenarchaeota.
Improved Methodology for Identification of Protists and Microalgae from Plankton Samples Preserved in Lugol's Iodine Solution: Combining Microscopic Analysis with Single-Cell PCR
Authors: Auinger Barbara M. , Pfandl Karin , Boenigk Jens
Published: February 2008

Here we introduce a method for quantitative analysis of planktonic protists and microalgae from preserved field samples combining morphological and small-subunit (SSU) rRNA gene sequence analysis. We linked a microscopic screening with PCR of single cells using field samples preserved with Lugol's iodine solution. Cells possessing a rigid cell wall were incubated with Viscozyme and subsequently with proteinase K for cell disruption; this was unnecessary for fragile cells. The addition of sodium thiosulfate to the PCR tube considerably decreased the inhibiting effect of the fixative (iodine) on the PCR and thus allowed for successful single-cell PCR even of long DNA fragments (up to as many as 3,000 base pairs). We further applied the protocol to investigate the dominant SSU rRNA genotypes in distinct flagellate morphospecies originating from different samples. We hypothesized that despite the morphological similarity, protist morphospecies in different habitats or sampled during different seasons are represented by different genotypes. Our results indicate species-specific differences: the two species
Ochromonas
sp. and
Dinobryon divergens
were represented by several different genotypes each, and for the latter species, the dominating genotype differed with habitat. In contrast,
Dinobryon pediforme
,
Dinobryon bavaricum
, and
Synura sphagnicola
were exclusively represented by a single genotype each, and the respective genotype was the same in different samples. In summary, our results highlight the significance of molecular variation within protist morphospecies.
The Genomes of the Non-Clearing-Zone-Forming and Natural-Rubber- Degrading SpeciesGordonia polyisoprenivoransandGordonia westfalicaHarbor Genes Expressing Lcp Activity inStreptomycesStrains
Authors: Bröker Daniel , Dietz David , Arenskötter Matthias , Steinbüchel Alexander
Published: February 2008

The latex-clearing protein (LcpK30) from the rubber-degrading bacteriumStreptomycessp. strain K30 is involved in the cleavage of poly(cis-1,4-isoprene), yielding isoprenoid aldehydes and ketones. Lcp homologues have so far been detected in all investigated clearing-zone-forming rubber-degrading bacteria. Internal degenerated oligonucleotides derived fromlcpgenes ofStreptomycessp. strain K30 (lcpK30),Streptomyces coelicolorstrain A3(2), andNocardia farcinicastrains IFM10152 and E1 were applied in PCR to investigate whetherlcphomologues occur also in the non-clearing-zone-forming rubber-utilizing bacteriaGordonia polyisoprenivoransstrains VH2 and Y2K,Gordonia alkanivoransstrain 44187, andGordonia westfalicastrain Kb1, which grow adhesively on rubber. The 1,230- and 1,224-bplcp-homologous genes fromG. polyisoprenivoransstrain VH2 (lcpVH2) andG. westfalicastrain Kb1 (lcpKb1) were obtained after screening genomic libraries by degenerated PCR amplification, and their translational products exhibited 50 and 52% amino acid identity, respectively, to LcpK30. RecombinantlcpVH2andlcpKb1harboring cells of the non-rubber-degradingStreptomyces lividansstrain TK23 were able to form clearing zones and aldehydes on latex overlay-agar plates, thus indicating thatlcpVH2andlcpKb1encode functionally active proteins. Analysis by gel permeation chromatography demonstrated lower polymer concentrations and molecular weights of the remaining polyisoprenoid molecules after incubation with these recombinantS. lividansstrains. Reverse transcription-PCR analysis demonstrated thatlcpVH2was transcribed in cells ofG. polyisoprenivoransstrain VH2 cultivated in the presence of poly(cis-1,4-isoprene) but not in the presence of sodium acetate. Anti-LcpK30immunoglobulin Gs, which were raised in this study, were rather specific for LcpK30and did not cross-react with LcpVH2and LcpKb1. AlcpVH2disruption mutant was still able to grow with poly(cis-1,4-isoprene) as sole carbon source; therefore,lcpVH2seems not to be essential for rubber degradation inG. polyisoprenivorans.
Critical Evaluation of Two Primers Commonly Used for Amplification of Bacterial 16S rRNA Genes
Authors: Frank Jeremy A. , Reich Claudia I. , Sharma Shobha , Weisbaum Jon S. , Wilson Brenda A. , Olsen Gary J.
Published: February 2008

rRNA-based studies, which have become the most common method for assessing microbial communities, rely upon faithful amplification of the corresponding genes from the original DNA sample. We report here an analysis and reevaluation of commonly used primers for amplifying the DNA between positions 27 and 1492 of bacterial 16S rRNA genes (numbered according to the
Escherichia coli
rRNA). We propose a formulation for a forward primer (27f) that includes three sequences not usually present. We compare our proposed formulation to two common alternatives by using linear amplification—providing an assessment that is independent of a reverse primer—and in combination with the 1492 reverse primer (1492r) under the PCR conditions appropriate for making community rRNA gene clone libraries. For analyses of DNA from human vaginal samples, our formulation was better at maintaining the original rRNA gene ratio of
Lactobacillus
spp. to
Gardnerella
spp., particularly under stringent amplification conditions. Because our 27f formulation remains relatively simple, having seven distinct primer sequences, there is minimal loss of overall amplification efficiency and specificity.
Single-Cell and Population Lag Times as a Function of Cell Age
Authors: Pin Carmen , Baranyi József
Published: February 2008

After inoculation, the times to the first divisions are longer and more widely distributed for those
Escherichia coli
single cells that spent more time in the stationary phase prior to inoculation. The second generation times are still longer than the typical generation times in the exponential phase, and this extended the apparent lag time of the cell population. The greater the variability of the single-cell interdivision intervals, the shorter are both the lag time and the doubling time of the population.
Rapid Mineralization of the Phenylurea Herbicide Diuron by
Variovorax
sp. Strain SRS16 in Pure Culture and within a Two-Member Consortium
Authors: Sørensen Sebastian R. , Albers Christian N. , Aamand Jens
Published: February 2008

The phenylurea herbicide diuron [
N
-(3,4-dichlorophenyl)-
N
,
N
-dimethylurea] is widely used in a broad range of herbicide formulations, and consequently, it is frequently detected as a major water contaminant in areas where there is extensive use. We constructed a linuron [
N
-(3,4-dichlorophenyl)-
N
-methoxy-
N
-methylurea]- and diuron-mineralizing two-member consortium by combining the cooperative degradation capacities of the diuron-degrading organism
Arthrobacter globiformis
strain D47 and the linuron-mineralizing organism
Variovorax
sp. strain SRS16. Neither of the strains mineralized diuron alone in a mineral medium, but combined, the two strains mineralized 31 to 62% of the added [
ring
-U-
14
C]diuron to
14
CO
2
, depending on the initial diuron concentration and the cultivation conditions. The constructed consortium was used to initiate the degradation and mineralization of diuron in soil without natural attenuation potential. This approach led to the unexpected finding that
Variovorax
sp. strain SRS16 was able to mineralize diuron in a pure culture when it was supplemented with appropriate growth substrates, making this strain the first known bacterium capable of mineralizing diuron and representatives of both the
N
,
N
-dimethyl- and
N
-methoxy-
N
-methyl-substituted phenylurea herbicides. The ability of the coculture to mineralize microgram-per-liter levels of diuron was compared to the ability of strain SRS16 alone, which revealed the greater extent of mineralization by the two-member consortium (31 to 33% of the added [
ring
-U-
14
C]diuron was mineralized to
14
CO
2
when 15.5 to 38.9 μg liter
−1
diuron was used). These results suggest that the consortium consisting of strains SRS16 and D47 could be a promising candidate for remediation of soil and water contaminated with diuron and linuron and their shared metabolite 3,4-dichloroaniline.
Identification of Siderophore Biosynthesis Genes Essential for Growth of
Aeromonas salmonicida
under Iron Limitation Conditions
Authors: Najimi Mohsen , Lemos Manuel L. , Osorio Carlos R.
Published: February 2008

Aeromonas salmonicida
subsp.
salmonicida
, the etiological agent of furunculosis in fish, produces a catechol-type siderophore under iron-limiting conditions. In this study, the Fur titration assay (FURTA) was used to identify a cluster of six genes,
asbG, asbF, asbD, asbC, asbB
, and
asbI
, encoding proteins similar to components of the siderophore biosynthetic machinery in other bacteria. Reverse transcriptase PCR analyses showed that this cluster consists of four iron-regulated transcriptional units. Mutants with deletions in either
asbD
(encoding a multidomain nonribosomal peptide synthetase),
asbG
(encoding a histidine decarboxylase), or
asbC
(encoding a predicted histamine monooxygenase) did not grow under iron-limiting conditions and did not produce siderophores. Growth of the Δ
asbG
strain under iron starvation conditions was restored by addition of histamine, suggesting that the siderophore in this species could contain a histamine-derived moiety. None of the mutants could grow in the presence of transferrin, indicating that
A. salmonicida
uses the catechol-type siderophore for removal of iron from transferrin rather than relying on a receptor for this iron-binding protein. All 18
A. salmonicida
strains analyzed by DNA probe hybridization were positive in tests for the presence of the
asbD
gene, and all of them promoted the growth of
asbD, asbG
, and
asbC
mutants, suggesting that this siderophore-mediated iron uptake system is conserved among
A. salmonicida
isolates. This study provides the first description of siderophore biosynthesis genes in this fish pathogen, and the results demonstrate that the
asbD, asbG
, and
asbC
genes are necessary for the production of a catecholate siderophore that is essential for the growth of
A. salmonicida
under iron limitation conditions.
Phenotype of a Mechanosensitive Channel Mutant,
mid
-
1
, in a Filamentous Fungus,
Neurospora crassa
Authors: Lew Roger R. , Abbas Zohaib , Anderca Marinela I. , Free Stephen J.
Published: February 2008

In the yeast
Saccharomyces cerevisiae
, the
MID1
(
m
ating-
i
nduced
d
eath) gene encodes a stretch-activated channel which is required for successful mating; the mutant phenotype is rescued by elevated extracellular calcium. Homologs of the
MID1
gene are found in fungi that are morphologically complex compared to yeast, both
Basidiomycetes
and
Ascomycetes
. We explored the phenotype of a
mid
-
1
knockout mutant in the filamentous ascomycete
Neurospora crassa
. The mutant exhibits lower growth vigor than the wild type (which is not rescued by replete calcium) and mates successfully. Thus, the role of the MID-1 protein differs from that of the homologous gene product in yeast. Hyphal cytology, growth on diverse carbon sources, turgor regulation, and circadian rhythms of the
mid-1
mutant are all similar to those of the wild type. However, basal turgor is lower than wild type, as is the activity of the plasma membrane H
+
-ATPase (measured by cyanide [CN
−
]-induced depolarization of the energy-dependent component of the membrane potential). In addition, the mutant is unable to grow at low extracellular Ca
2+
levels or when cytoplasmic Ca
2+
is elevated with the Ca
2+
ionophore A23187. We conclude that the MID-1 protein plays a role in regulation of ion transport via Ca
2+
homeostasis and signaling. In the absence of normal ion transport activity, the mutant exhibits poorer growth.
Cell Cycle Synchrony in
Giardia intestinalis
Cultures Achieved by Using Nocodazole and Aphidicolin
Authors: Poxleitner Marianne K. , Dawson Scott C. , Cande W. Zacheus
Published: February 2008

Giardia intestinalis
is a ubiquitous intestinal protozoan parasite and has been proposed to represent the earliest diverging lineage of extant eukaryotes. Despite the importance of
Giardia
as a model organism, research on
Giardia
has been hampered by an inability to achieve cell cycle synchrony for in vitro cultures. This report details successful methods for attaining cell cycle synchrony in
Giardia
cultures. The research presented here demonstrates reversible cell cycle arrest in G
1
/S and G
2
/M with aphidicolin and nocodazole, respectively. Following synchronization, cells were able to recover completely from drug treatment and remained viable and maintained synchronous growth for 6 h. These techniques were used to synchronize
Giardia
cultures to increase the percentages of mitotic spindles in the cultures. This method of synchronization will enhance our ability to study cell cycle-dependent processes in
G. intestinalis
.
Complete Genome Sequence of the Mosquitocidal Bacterium
Bacillus sphaericus
C3-41 and Comparison with Those of Closely Related Bacillus Species
Authors: Hu Xiaomin , Fan Wei , Han Bei , Liu Haizhou , Zheng Dasheng , Li Qibin , Dong Wei , Yan Jianping , Gao Meiying , Berry Colin , Yuan Zhiming
Published: February 2008

Bacillus sphaericus
strain C3-41 is an aerobic, mesophilic, spore-forming bacterium that has been used with great success in mosquito control programs worldwide. Genome sequencing revealed that the complete genome of this entomopathogenic bacterium is composed of a chromosomal replicon of 4,639,821 bp and a plasmid replicon of 177,642 bp, containing 4,786 and 186 potential protein-coding sequences, respectively. Comparison of the genome with other published sequences indicated that the
B. sphaericus
C3-41 chromosome is most similar to that of
Bacillus
sp. strain NRRL B-14905, a marine species that, like
B. sphaericus
, is unable to metabolize polysaccharides. The lack of key enzymes and sugar transport systems in the two bacteria appears to be the main reason for this inability, and the abundance of proteolytic enzymes and transport systems may endow these bacteria with exclusive metabolic pathways for a wide variety of organic compounds and amino acids. The genes shared between
B. sphaericus
C3-41 and
Bacillus
sp. strain NRRL B-14905, including mobile genetic elements, membrane-associated proteins, and transport systems, demonstrated that these two species are a biologically and phylogenetically divergent group. Knowledge of the genome sequence of
B. sphaericus
C3-41 thus increases our understanding of the bacilli and may also offer prospects for future genetic improvement of this important biological control agent.
Hjm/Hel308A DNA Helicase fromSulfolobus tokodaiiPromotes Replication Fork Regression and Interacts with Hjc Endonuclease In Vitro
Authors: Li Zhuo , Lu Shuhong , Hou Guihua , Ma Xiaoqing , Sheng Duohong , Ni Jinfeng , Shen Yulong
Published: February 2008

Hjm and Hel308a are novel, RecQ-like DNA helicases recently identified in the euryarchaeotesPyrococcus furiosusandMethanothermobacter thermautotrophicus, respectively. In this study, an Hjm/Hel308 homologue (designated StoHjm) fromSulfolobus tokodaii, a hyperthermophilic archaeon belonging to theCrenarchaeotasubdomain of archaea, was cloned, purified, and characterized. Unlike Hjm and Hel308a, which unwind DNA in a 3′-to-5′ direction, StoHjm unwound DNA in both 3′-to-5′ and 5′-to-3′ directions. Remarkably, StoHjm exhibited structure-specific single-stranded-DNA-annealing and fork regression activities in vitro. In addition, gel filtration, affinity pulldown, and yeast two-hybrid analyses revealed that StoHjm physically interacted with StoHjc, the Holliday junction-specific endonuclease fromS. tokodaii. This interaction may have functional significance, because the unwinding activity of StoHjm was inhibited by StoHjc in vitro. These results may suggest that the Hjm/Hel308 family helicases, in association with Hjc endonucleases, are involved in processing of stalled replication forks.
YjgF Is Required for Isoleucine Biosynthesis whenSalmonella entericaIs Grown on Pyruvate Medium
Authors: Christopherson Melissa R. , Schmitz G. E. , Downs Diana M.
Published: February 2008

The YjgF/YER057c/UK114 family of proteins is conserved across the three domains of life, yet no biochemical function has been clearly defined for any member of this family. InSalmonella enterica, a deletion ofyjgFresults in a requirement for isoleucine when the mutant strain is grown in glucose-serine or pyruvate medium. Feedback inhibition of IlvA is required for the curative effect of isoleucine on glucose-serine medium. On pyruvate medium,yjgFmutants are unable to synthesize enough isoleucine for growth. From this study, we conclude that the isoleucine requirement of ayjgFmutant on pyruvate is a consequence of the decreased transaminase B (IlvE) activity that has previously been characterized in these mutants.
Nitrate Enhances the Survival of
Mycobacterium tuberculosis
during Inhibition of Respiration
Authors: Sohaskey Charles D.
Published: February 2008

When oxygen is slowly depleted from growing cultures of
Mycobacterium tuberculosis
, they enter a state of nonreplicating persistence that resembles the dormant state seen with latent tuberculosis. In this hypoxic state, nitrate reductase activity is strongly induced. Nitrate in the medium had no effect on long-term persistence during gradual oxygen depletion (Wayne model) for up to 46 days, but significantly enhanced survival during sudden anaerobiosis. This enhancement required a functional nitrate reductase. Thioridazine is a member of the class of phenothiazines that act, in part, by inhibiting respiration. Thioridazine was toxic to both actively growing and nonreplicating cultures of
M. tuberculosis
. At a sublethal concentration of thioridazine, nitrate in the medium improved the growth. At lethal concentrations of thioridazine, nitrate increased survival during aerobic incubation as well as in microaerobic cultures that had just entered nonreplicating persistence (NRP-1). In contrast, the survival of anaerobic persistent (NRP-2) cultures exposed to thioridazine was not increased by the addition of nitrate. Nitrate reduction is proposed to play a role during the sudden interruption of aerobic respiration due to causes such as hypoxia, thioridazine, or nitric oxide.
Transposable Modules Generated by a Single Copy of Insertion Sequence IS
Pme1
and Their Influence on Structure and Evolution of Natural Plasmids of
Paracoccus methylutens
DM12
Authors: Bartosik Dariusz , Putyrski Mateusz , Dziewit Lukasz , Malewska Edyta , Szymanik Michal , Jagiello Ewa , Lukasik Jacek , Baj Jadwiga
Published: February 2008

We demonstrated that a single copy of insertion sequence IS
Pme1
can mobilize adjacent segments of genomic DNA of
Paracoccus methylutens
DM12, which leads to the generation of diverse transposable elements of various size and DNA contents. All elements (named transposable modules [TMos]) contain IS
Pme1
(placed at the 5′ ends of the elements) and have variable 3′-end regions of between 0.5 and 5 kb. IS
Pme1
was shown to encode an outwardly oriented promoter, which may activate the transcription of genes transposed within TMos in evolutionarily distinct hosts. TMos may therefore be considered to be natural systems enabling gene capture, expression, and spread. However, unless these elements have been inserted into a highly conserved genetic context to enable a precise definition of their termini, it is extremely difficult or even impossible to identify them in bacterial genomes by in silico sequence analysis. We showed that TMos are present in the chromosome and plasmids of strain DM12. Sequence analysis of plasmid pMTH1 (32 kb) revealed that four TMos, previously identified with a trap vector, pMEC1, comprise 87% of its genome. Repeated TMos within pMTH1 may stimulate other structural rearrangements resulting from homologous recombination between long repeat sequences. This illustrates that TMos may play a significant role in shaping the structure of natural plasmids, which consequently may have a great impact on the evolution of plasmid genomes.
Regulation of the Synthesis of the Angucyclinone Antibiotic Alpomycin inStreptomyces ambofaciensby the Autoregulator Receptor AlpZ and Its Specific Ligand
Authors: Bunet Robert , Mendes Marta V. , Rouhier Nicolas , Pang Xiuhua , Hotel Laurence , Leblond Pierre , Aigle Bertrand
Published: February 2008

Streptomyces ambofaciensproduces an orange pigment and the antibiotic alpomycin, both of which are products of a type II polyketide synthase gene cluster identified in each of the terminal inverted repeats of the linear chromosome. Five regulatory genes encodingStreptomycesantibiotic regulatory proteins (alpV, previously shown to be an essential activator gene;alpT; andalpU) and TetR family receptors (alpZandalpW) were detected in this cluster. Here, we demonstrate that AlpZ, which shows high similarity to γ-butyrolactone receptors, is at the top of a pathway-specific regulatory hierarchy that prevents synthesis of thealppolyketide products. Deletion of the two copies ofalpZresulted in the precocious production of both alpomycin and the orange pigment, suggesting a repressor role for AlpZ. Consistent with this, expression of the fivealp-located regulatory genes and of two representative biosynthetic structural genes (alpAandalpR) was induced earlier in thealpZdeletion strain. Furthermore, recombinant AlpZ was shown to bind to specific DNA sequences within the promoter regions ofalpZ,alpV, andalpXW, suggesting direct transcriptional control of these genes by AlpZ. Analysis of solvent extracts ofS. ambofacienscultures identified the existence of a factor which induces precocious production of alpomycin and pigment in the wild-type strain and which can disrupt the binding of AlpZ to its DNA targets. This activity is reminiscent of γ-butyrolactone-type molecules. However, the AlpZ-interacting molecule(s) was shown to be resistant to an alkali treatment capable of inactivating γ-butyrolactones, suggesting that the AlpZ ligand(s) does not possess a lactone functional group.
Benefits of long-term β-blockade in experimental chronic aortic regurgitation
Authors: Plante Eric , Lachance Dominic , Champetier Serge , Drolet Marie-Claude , Roussel Élise , Arsenault Marie , Couet Jacques
Published: February 2008

The objective of this study was to assess the long-term effects of β-blockade on survival and left ventricular (LV) remodeling in rats with aortic valve regurgitation (AR). The pharmacological management of chronic AR remains controversial. No drug has been definitively proven to delay the need for valve replacement or to affect morbidity and/or mortality. Our group has reported that the adrenergic system is activated in an animal model of AR and that adrenergic blockade may help maintain normal LV function. The effects of prolonged treatment with a β-blocker are unknown. Forty Wistar rats with severe AR were divided into 2 groups of 20 animals each and treated with metoprolol (Met, 25 mg·kg−1·day−1) or left untreated for 1 yr. LV remodeling was evaluated by echocardiography. Survival was assessed by Kaplan-Meir curves. Hearts were harvested for tissue analysis. All Met-treated animals were alive after 6 mo vs. 70% of untreated animals. After 1 yr, 60% of Met-treated animals were alive vs. 35% of untreated animals ( P = 0.028). All deaths, except one, were sudden. There were no differences in LV ejection fraction (all >50%) or LV dimensions. LV mass tended to be lower in the Met-treated group. There was less subendocardial fibrosis in this group, as well as lower LV filling pressures (LV end-diastolic pressure). β-Adrenergic receptor ratio (β1/β2) was improved. One year of treatment with Met was well tolerated. Met improved 1-yr survival, minimized LV hypertrophy, improved LV filling pressures, decreased LV subendocardial fibrosis, and helped restore the β-adrenergic receptor ratio.
Cyclooxygenase products sensitize muscle mechanoreceptors in humans with heart failure
Authors: Middlekauff Holly R. , Chiu Josephine , Hamilton Michele A. , Fonarow Gregg C. , MacLellan W. Robb , Hage Antoine , Moriguchi Jaime , Patel Jignesh
Published: February 2008

Prior work in animals and humans suggests that muscle mechanoreceptor control of sympathetic activation [muscle sympathetic nerve activity (MSNA)] during exercise in heart failure (HF) patients is heightened compared with that of healthy humans and that muscle mechanoreceptors are sensitized by metabolic by-products. We sought to determine whether cyclooxygenase products and/or endogenous adenosine, two metabolites of ischemic exercise, sensitize muscle mechanoreceptors during rhythmic handgrip (RHG) exercise in HF patients. Indomethacin, which inhibits the production of prostaglandins, and saline control were infused in 12 HF patients. In a different protocol, aminophylline, which inhibits adenosine receptors, and saline control were infused in 12 different HF patients. MSNA was recorded (microneurography). During exercise following saline, MSNA increased in the first minute of exercise, consistent with baseline heightened mechanoreceptor sensitivity. MSNA continued to increase during 3 min of RHG, indicative that muscle mechanoreceptors are sensitized by ischemia metabolites. Indomethacin, but not aminophylline, markedly attenuated the increase in MSNA during the entire 3 min of low-level rhythmic exercise, consistent with the sensitization of muscle mechanoreceptors by cyclooxygenase products. Interestingly, even the early increase in MSNA was abolished by indomethacin infusion, indicative of the very early generation of cyclooxygenase products after the onset of exercise in HF patients. In conclusion, muscle mechanoreceptors mediate the increase in MSNA during low-level RHG exercise in HF. Cyclooxygenase products, but not endogenous adenosine, play a central role in muscle mechanoreceptor sensitization. Finally, muscle mechanoreceptors in patients with HF have heightened basal sensitivity to mechanical stimuli, which also appears to be mediated by the early generation of cyclooxygenase products, resulting in exaggerated early increases in MSNA.
Intermittent hypoxia modulates nitric oxide-dependent vasodilation and capillary perfusion during ischemia-reperfusion-induced damage
Authors: Bertuglia Silvia
Published: February 2008

The microvascular function of nitric oxide (NO) during ischemia-reperfusion (I/R) in intermittent hypoxia (IH)-pretreated hamsters was analyzed using 20 mg/kg of the nonselective NO inhibitor Nω-nitro-l-arginine methyl ester (l-NAME) and 5 mg/kg of the preferential inducible NO inhibitor S-methylisothiourea sulphate (SMT) injected before I/R. Studies were made in the hamster cheek pouch microcirculation (intravital fluorescence microscopy). IH consisted of 6 min of 8% O2breathing followed by 6 min of 21% O2for every 8 h for 21 days. Normoxia controls (NCs) were exposed to room air for the same period. The effects were characterized in terms of systemic hemodynamics, diameter, flow, wall shear stress in arterioles, capillary perfusion, and the concentrations of thiobarbituric acid-reactive substances (TBARS) and plasma NO, assessed as nitrite/nitrate (NOx) levels. IH did not change arterial blood pressure and increased hematocrit and shear stress. IH increased NOx and TBARS levels and reduced arterial diameter, blood flow, and capillary perfusion versus the NC. Conversely, TBARS and NOx were lower during I/R in IH-pretreated hamsters, resulting in vasodilation and the increase of capillary perfusion and shear stress. After IH, capillary perfusion was reduced by 24% (2.3%) and enhanced by 115% (1.7%) after I/R ( P < 0.05). Both modalities of NO blockade decreased NOx generation and increased TBARS versus IH. l-NAME and SMT induced a significant decrease in arteriolar diameter, blood flow, and capillary perfusion ( P < 0.05). l-NAME enhanced TBARS more than SMT and aggravated I/R damage. In conclusion, we demonstrated that preconditioning with IH greatly reduces oxidative stress and stimulates NO-induced vasodilation during I/R injury, thus maintaining capillary perfusion.
Inhalation of endothelin receptor blockers in pulmonary hypertension
Authors: Leuchte Hanno H. , Meis Tobias , El-Nounou Michal , Michalek Jens , Behr Jürgen
Published: February 2008

Endothelin 1 (ET-1) is a potent pulmonary vasoconstrictor and mediator of lung diseases. Antagonism of the ET-1-mediated effects has become an important therapeutic approach. ET-1 (A and B) receptors are differentially distributed in the lung vasculature. Whereas the ETAreceptors mainly mediate vasoconstriction, the endothelial ETBreceptor seems to have vasodilative properties. We sought to determine if antagonism of ET receptors can be achieved by inhalation of specific blockers in a model of ET-1-mediated pulmonary hypertension.
Lovastatin inhibits bronchial hyperresponsiveness by reducing RhoA signaling in rat allergic asthma
Authors: Chiba Yoshihiko , Arima Junko , Sakai Hiroyasu , Misawa Miwa
Published: February 2008

Recent studies revealed an importance of a monomeric GTP-binding protein, RhoA, in contraction of bronchial smooth muscle (BSM). RhoA and its downstream have been proposed as a new target for the treatment of airway hyperresponsiveness in asthma. Statins are known to inhibit the functional activation of RhoA via the depletion of geranylgeranylpyrophosphate. To determine the beneficial effects of statins on the airway hyperresponsiveness in allergic bronchial asthma, we investigated the effects of systemic treatment with lovastatin on the augmented BSM contraction and activation of RhoA in rats with allergic bronchial asthma. Rats were sensitized and repeatedly challenged with 2,4-dinitrophenylated Ascaris suum antigen. Animals were also treated with lovastatin (4 mg·kg−1·day−1ip) once a day before and during the antigen inhalation period. Repeated antigen inhalation caused a marked BSM hyperresponsiveness to ACh with the increased expression and translocation of RhoA. Lovastatin treatments significantly attenuated both the augmented contraction and RhoA translocation to the plasma membrane. Lovastatin also reduced the increased cell number in bronchoalveolar lavage fluids and histological changes induced by antigen exposure, whereas the levels of immunoglobulin E in sera and interleukins-4, -6, and -13 in bronchoalveolar lavage fluids were not significantly changed. These findings suggest that lovastatin ameliorates antigen-induced BSM hyperresponsiveness, an important factor of airway hyperresponsiveness in allergic asthmatics, probably by reducing the RhoA-mediated signaling.