bluuebunny/crossref_metadata_2025_split huggingface dataset:
Involvement of Nitrate Reductase and Pyoverdine in Competitiveness of
Pseudomonas fluorescens
Strain C7R12 in Soil
Authors: Mirleau Pascal , Philippot Laurent , Corberand Thérèse , Lemanceau Philippe 
Published: July 2002

ABSTRACT

Involvement of nitrate reductase and pyoverdine in the competitiveness of the biocontrol strain
Pseudomonas fluorescens
C7R12 was determined, under gnotobiotic conditions, in two soil compartments (bulk and rhizosphere soil), with the soil being kept at two different values of matric potential (−1 and −10 kPa). Three mutants affected in the synthesis of either the nitrate reductase (Nar
−
), the pyoverdine (Pvd
−
), or both (Nar
−
Pvd
−
) were used. The Nar
−
and Nar
−
Pvd
−
mutants were obtained by site-directed mutagenesis of the wild-type strain and of the Pvd
−
mutant, respectively. The selective advantage given by nitrate reductase and pyoverdine to the wild-type strain was assessed by measuring the dynamic of each mutant-to-total-inoculant (wild-type strain plus mutant) ratio. All three mutants showed a lower competitiveness than the wild-type strain, indicating that both nitrate reductase and pyoverdine are involved in the fitness of
P. fluorescens
C7R12. The double mutant presented the lowest competitiveness. Overall, the competitive advantages given to C7R12 by nitrate reductase and pyoverdine were similar. However, the selective advantage given by nitrate reductase was more strongly expressed under conditions of lower aeration (−1 kPa). In contrast, the selective advantage given by nitrate reductase and pyoverdine did not differ in bulk and rhizosphere soil, indicating that these bacterial traits are not specifically involved in the rhizosphere competence but rather in the saprophytic ability of C7R12 in soil environments.
Inoculation onto Solid Surfaces ProtectsSalmonellaspp. during Acid Challenge: a Model Study Using Polyethersulfone Membranes
Authors: Gawande Purushottam V. , Bhagwat Arvind A. 
Published: July 2002

ABSTRACTSalmonellae are the most frequently reported cause of outbreaks of food-borne gastroenteritis in the United States. In clinical trials, the oral infective dose (ID) for healthy volunteers was estimated to be approximately 1 million cells. However, in reports from various outbreaks, the ID ofSalmonellaspecies associated with solid foods was estimated to be as few as 100 cells. We found that fresh-cut produce surfaces not only provided suitable solid support for pathogen attachment but also played a critical role in increasing the acid tolerance of the pathogen. However the acidic nature of certain produce played no role in making salmonellae resistant to stomach acidity. Inoculation onto fresh-cut produce surfaces, as well as onto inert surfaces, such as polyethersulfone membranes and tissue paper, increased the survival of salmonellae during acid challenge (50 mM Na-citrate, pH 3.0; 37°C; 2 h) by 4 to 5 log units. Acid challenge experiments using cells inoculated onto polyethersulfone membranes provided a model system suitable for studying the underlying fundamentals of the protection that occurs whenSalmonellastrains are associated with solid foods. The surface-associated acid protection, which was observed in severalSalmonellastrains, required de novo protein synthesis and was independent of stationary-phase sigma transcription factor.
Higher Abundance of Bacteria than of Viruses in Deep Mediterranean Sediments
Authors: Danovaro Roberto , Manini Elena , Dell'Anno Antonio 
Published: July 2002

ABSTRACT

The interactions between viral abundance and bacterial density, biomass, and production were investigated along a longitudinal transect consisting of nine deep-sea stations encompassing the entire Mediterranean basin. The numbers of viruses were very low (range, 3.6 × 10
7
to 12.0 × 10
7
viruses g
−1
) and decreased eastward. The virus-to-bacterium ratio was always < 1.0, indicating that the deep-sea sediments of the Mediterranean Sea are the first example of a marine ecosystem not numerically dominated by viruses. The lowest virus numbers were found where the lowest bacterial metabolism and turnover rates and the largest cell size were observed, suggesting that bacterial doubling time might play an important role in benthic virus development.
Cre-
loxP
Recombination System for Large Genome Rearrangements in
Lactococcus lactis
Authors: Campo Nathalie , Daveran-Mingot Marie-Line , Leenhouts Kees , Ritzenthaler Paul , Le Bourgeois Pascal 
Published: July 2002

ABSTRACT

We have used a new genetic strategy based on the Cre-
loxP
recombination system to generate large chromosomal rearrangements in
Lactococcus lactis
. Two
loxP
sites were sequentially integrated in inverse order into the chromosome either at random locations by transposition or at fixed points by homologous recombination. The recombination between the two chromosomal
loxP
sites was highly efficient (approximately 1 × 10
−1
/cell) when the Cre recombinase was provided in
trans
, and parental- or inverted-type chromosomal structures were isolated after removal of the Cre recombinase. The usefulness of this approach was demonstrated by creating three large inversions of 500, 1,115, and 1,160 kb in size that modified the lactococcal genome organization to different extents. The Cre-
loxP
recombination system described can potentially be used for other gram-positive bacteria without further modification.
Identification of Novel Hexapeptides Bioactive against Phytopathogenic Fungi through Screening of a Synthetic Peptide Combinatorial Library
Authors: López-García Belén , Pérez-Payá Enrique , Marcos Jose F. 
Published: July 2002

ABSTRACTThe purpose of the present study was to improve the antifungal activity against selected phytopathogenic fungi of the previously identified hexapeptide PAF19. We describe some properties of a set of novel synthetic hexapeptides whosed-amino acid sequences were obtained through screening of a synthetic peptide combinatorial library in a positional scanning format. As a result of the screening, 12 putative bioactive peptides were identified, synthesized, and assayed. The peptides PAF26 (Ac-rkkwfw-NH2), PAF32 (Ac-rkwhfw-NH2), and PAF34 (Ac-rkwlfw-NH2) showed stronger activity than PAF19 against isolates ofPenicillium digitatum,Penicillium italicum, andBotrytis cinerea. PAF26 and PAF32, but not PAF34, were also active againstFusarium oxysporum. Penicillium expansumwas less susceptible to all four PAF peptides, and only PAF34 showed weak activity against it. Assays were also conducted on nontarget organisms, and PAF26 and PAF32 showed much-reduced toxicity toEscherichia coliandSaccharomyces cerevisiae,demonstrating selectivity towards certain filamentous fungi. Thus, the data showed distinct activity profiles for peptides differentiated by just one or two residue substitutions. Our conclusion from this observation is that a specificity factor is involved in the activity of these short peptides. Furthermore, PAF26 and PAF32 displayed activities againstP. digitatum,P. italicum, andB. cinereasimilar to that of the hemolytic 26-amino acid melittin, but they did not show the high toxicity of melittin towards bacteria and yeasts. The four peptides acted additively, with no synergistic interactions among them, and PAF26 was shown to have improved activity over PAF19 in in vivo orange fruit decay experiments.
Alterations in
Vibrio fischeri
Motility Correlate with a Delay in Symbiosis Initiation and Are Associated with Additional Symbiotic Colonization Defects
Authors: Millikan Deborah S. , Ruby Edward G. 
Published: July 2002

ABSTRACT

Motility is required for
Vibrio fischeri
cells to interact with and specifically colonize the light-emitting organ of their host, the squid
Euprymna scolopes
. To investigate the influence of motility on the expression of the symbiotic phenotype, we isolated mutants of the squid symbiont
V. fischeri
ES114 that had altered migration abilities. Spontaneous hyperswimmer (HS) mutants, which migrated more rapidly in soft agar and were hyperflagellated relative to the wild type, were isolated and grouped into three phenotypic classes. All of the HS strains tested, regardless of class, were delayed in symbiosis initiation. This result suggested that the hypermotile phenotype alone contributes to an inability to colonize squid normally. Class III HS strains showed the greatest colonization defect: they colonized squid to a level that was only 0.1 to 10% that achieved by ES114. In addition, class III strains were defective in two capabilities, hemagglutination and luminescence, that have been previously described as colonization factors in
V. fischeri
. Class II and III mutants also share a mucoid colony morphology; however, class II mutants can colonize
E. scolopes
to a level that was 40% of that achieved by ES114. Thus, the mucoid phenotype alone does not contribute to the greater defect exhibited by class III strains. When squid were exposed to ES114 and any one of the HS mutant strains as a coinoculation, the parent strain dominated the resulting symbiotic light-organ population. To further investigate the colonization defects of the HS strains, we used confocal laser-scanning microscopy to visualize
V. fischeri
cells in their initial interaction with
E. scolopes
tissue. Compared to ES114, HS strains from all three classes were delayed in two behaviors involved in colonization: (i) aggregation on host-derived mucus structures and (ii) migration to the crypts. These results suggest that, while motility is required to initiate colonization, the presence of multiple flagella may actually interfere with normal aggregation and attachment behavior. Furthermore, the pleiotropic nature of class III HS strains provides evidence that motility is coregulated with other symbiotic determinants in
V. fischeri
.
Molecular Identification of Family 38 α-Mannosidase ofBacillussp. Strain GL1, Responsible for Complete Depolymerization of Xanthan
Authors: Nankai Hirokazu , Hashimoto Wataru , Murata Kousaku 
Published: July 2002

ABSTRACTWhen cells ofBacillussp. strain GL1 were grown in a medium containing xanthan as a carbon source, α-mannosidase exhibiting activity towardp-nitrophenyl-α-d-mannopyranoside (pNP-α-d-Man) was produced intracellularly. The 350-kDa α-mannosidase purified from a cell extract of the bacterium was a trimer comprising three identical subunits, each with a molecular mass of 110 kDa. The enzyme hydrolyzedpNP-α-d-Man (Km= 0.49 mM) andd-mannosyl-(α-1,3)-d-glucose most efficiently at pH 7.5 to 9.0, indicating that the enzyme catalyzes the last step of the xanthan depolymerization pathway ofBacillussp. strain GL1. The gene for α-mannosidase cloned most by using N-terminal amino acid sequence information contained an open reading frame (3,144 bp) capable of coding for a polypeptide with a molecular weight of 119,239. The deduced amino acid sequence showed homology with the amino acid sequences of α-mannosidases belonging to glycoside hydrolase family 38.
Parasporin, a Human Leukemic Cell-Recognizing Parasporal Protein of
Bacillus thuringiensis
Authors: Mizuki Eiichi , Park Yu Shin , Saitoh Hiroyuki , Yamashita Satoko , Akao Tetsuyuki , Higuchi Kazuhiko , Ohba Michio 
Published: July 2002

ABSTRACT

An unusual property, human leukemic cell-recognizing activity, associated with parasporal inclusions of a noninsecticidal
Bacillus thuringiensis
soil isolate was investigated, and a protein (named parasporin in this study) responsible for the activity was cloned. The parasporin, encoded by a gene 2,169 bp long, was a polypeptide of 723 amino acid residues with a predicted molecular weight of 81,045. The sequence of parasporin contained the five conserved blocks commonly found in
B. thuringiensis
Cry proteins; however, only very low homologies (<25%) between parasporin and the existing classes of Cry and Cyt proteins were detected. Parasporin exhibited cytocidal activity only when degraded by proteases into smaller molecules of 40 to 60 kDa. Trypsin and proteinase K activated parasporin, while chymotrypsin did not. The activated parasporin showed strong cytocidal activity against human leukemic T cells (MOLT-4) and human uterus cervix cancer cells (HeLa) but not against normal T cells.
Use of Two-Dimensional Gel Electrophoresis To Measure Changes in Synovial Fluid Proteins from Patients with Rheumatoid Arthritis Treated with Antibody to CD4
Authors: Smith Marjorie A. , Bains Satbinder K. , Betts Joanna C. , Choy Ernest H. S. , Zanders Edward D. 
Published: July 2002

ABSTRACTSynovial fluid proteins from microliter volumes of synovial fluid were resolved by two-dimensional polyacrylamide gel electrophoresis and detected by silver staining to investigate the feasibility of using two-dimensional (2D) electrophoresis in the clinical research setting and provide global disease information of disease progression. Several hundred proteins could be resolved as spots, many of which displayed the characteristic pattern of plasma-derived glycoproteins. The lowest level of detection was approximately 0.2 ng from a total of 50 μg of protein loaded. Most of the proteins could be identified on the basis of pI and molecular weight when compared with plasma protein maps on the World Wide Web. Unknown proteins were characterized by mass spectrometry of tryptic digests and by comparison with peptide databases. Synovial fluids from patients with rheumatoid arthritis were analyzed using this technique. Each subject received a fixed dose of antibody to CD4 as part of a phase II clinical trial to determine the efficacy of this immunosuppressive treatment in modifying disease activity. Synovial fluid was removed at day 0, followed by administration of antibody. Subsequent removal of synovial fluid and additional administration of antibody were carried out at different times thereafter. Changes in levels of acute-phase proteins were quantified by densitometry of silver-stained 2D polyacrylamide gels. Other parameters of disease progression such as serum C-reactive protein and physician's global assessment of clinical condition were used for comparison. In this way, changes in acute-phase proteins towards normal levels, as measured by 2D polyacrylamide gel electrophoresis, could be correlated with clinical improvement and conventional clinical chemistry measurements. Thus, the system can be used for quantitative analysis of protein expression in sites of autoimmune disease activity such as the synovial fluid of rheumatoid arthritis patients.
Duration of Sample Storage Dramatically Alters Expression of the Human Immunodeficiency Virus Coreceptors CXCR4 and CCR5
Authors: Shalekoff Sharon , Tiemessen Caroline T. 
Published: July 2002

ABSTRACTExpression of the chemokine receptors CXCR4 and CCR5 was monitored using EDTA-anticoagulated whole blood held for different time periods prior to fluorescent-antibody staining. When left overnight CXCR4 expression on leukocytes was substantially increased, whereas CCR5 expression was reduced. The results were similar when heparin and acid-citrate-dextrose were used as anticoagulants.
New Assay for Measuring Cell Surface Hydrophobicities of
Candida dubliniensis
and
Candida albicans
Authors: Jabra-Rizk M. A. , Falkler W. A. , Merz W. G. , Meiller T. F. 
Published: July 2002

ABSTRACT

Hydrophobic interactions, based on cell surface hydrophobicity (CSH), are among the many and varied mechanisms of adherence deployed by the pathogenic yeast
Candida albicans
. Recently it was shown that, unlike
C. albicans
,
C. dubliniensis
is a species that exhibits an outer fibrillar layer consistent with constant CSH. Previously,
C. dubliniensis
grown at 25 or 37°C was shown to coaggregate with the oral anaerobic bacterium
Fusobacterium nucleatum. C. albicans
, however, demonstrated similar coaggregation only when hydrophobic or grown at 25°C. This observation implied that coaggregation of
Candida
cells with
F. nucleatum
is associated with a hydrophobic yeast cell surface. To test this hypothesis, 42
C. albicans
and 40
C. dubliniensis
clinical isolates, including a
C. albicans
hydrophobic variant, were grown at 25 and 37°C and tested with the established hydrophobicity microsphere assay, which determines CSH levels based on the number of microspheres attached to the yeast cells. The coaggregation assay was performed in parallel experiments. All
C. dubliniensis
isolates grown at either temperature, hydrophobic 25°C-grown
C. albicans
isolates, and the
C. albicans
hydrophobic variant, unlike the 37°C-hydrophilic
C. albicans
isolates, exhibited hydrophobic CSH levels with the microsphere assay and simultaneously showed maximum, 4+, coaggregation with
F. nucleatum
. The parallel results obtained for
C. dubliniensis
using both assays support the use of the CoAg assay both as a rapid assay to determine CSH and to differentiate between
C. dubliniensis
and
C. albicans
.
Clinical and Serological Follow-Up of Patients with Human Granulocytic Ehrlichiosis in Slovenia
Authors: Lotrič-Furlan Stanka , Avsic-Zupanc T. , Petrovec M. , Nicholson W. L. , Sumner J. W. , Childs J. E. , Strle F. 
Published: July 2002

ABSTRACT
An evaluation of the clinical outcome and the duration of the antibody response of patients with human granulocytic ehrlichiosis (HGE) was undertaken in Slovenia. Adult patients with a febrile illness occurring within 6 weeks of a tick bite were classified as having probable or confirmed HGE based on the outcome of serological or PCR testing. Thirty patients (median age, 44 years) were enrolled, and clinical evaluations and serum collection were undertaken at initial presentation and at 14 days, 6 to 8 weeks, and 3 to 4, 6, 12, 18, and 24 months. An indirect immunofluorescence assay (IFA) was performed, and reciprocal titers of ≥128 were interpreted as positive. Patients presented a median of 4 days after the onset of fever and were febrile for a median of 7.5 days; four (13.3%) received doxycycline. Seroconversion was observed in 3 of 30 (10.0%) patients, and 25 (83.3%) showed >4-fold change in antibody titer. PCR results were positive in 2 of 3 (66.7%) seronegative patients but in none of 27 seropositive patients at the first presentation. IFA antibody titers of ≥128 were found in 14 of 29 (48.3%), 17 of 30 (56.7%), 13 of 30 (43.4%), and 12 of 30 (40.0%) patients 6, 12, 18, and 24 months after presentation, respectively. Patients reporting additional tick bites during the study had significantly higher antibody titers at most time points during follow-up. No long-term clinical consequences were found during follow-up.
Diagnosis of Babesiosis Using an Immunoblot Serologic Test
Authors: Ryan Raymond , Krause Peter J. , Radolf Justin , Freeman Kathy , Spielman Andrew , Lenz Ronald , Levin Andrew 
Published: July 2002

ABSTRACT

Although the current indirect immunofluorescent assay (IFA) diagnostic antibody test for human babesiosis is sensitive and specific, an immunoblot antibody test may be easier to standardize and to perform. Our objective, therefore, was to determine the efficacy of and develop interpretive criteria for an immunoblot antibody test for diagnosing acute human babesiosis using a
Babesia microti
whole-cell lysate as the antigen. We compared the reactivity of sera to a
B. microti
immunoblot assay in 24 human subjects experiencing symptoms and expressing laboratory evidence of babesiosis, 28 subjects who experienced Lyme disease, 12 subjects who experienced human granulocytic ehrlichiosis, and 51 subjects who reported no history of any of these diseases and whose sera did not react against
B. microti
antigen in an IFA test. Immunoblot strips were impregnated with proteins derived from the GI strain of
B. microti
that had been electrophoresed in an acrylamide sodium dodecyl sulfate gel, followed by electroblotting onto nitrocellulose membranes. The sera of all subjects who experienced babesiosis reacted against the
B. microti
antigen in the IFA and against at least one of nine immunoblot protein bands specific to
B. microti
. In contrast, none of the sera from people who appeared not to have experienced this infection reacted against the
B. microti
antigen in the IFA (compared to 4% in the immunoblot assay). When two reactive bands were considered as definitive, immunoblot test sensitivity was 96%, while specificity was 99% and predictive positivity and predictive negativity were 96 and 99%, respectively. Our
B. microti
immunoblot procedure shows promise as a sensitive, specific, and reproducible assay for routine clinical diagnosis of acute babesiosis.
Circulating Inflammatory Mediators in Patients with Fever: Predicting Bloodstream Infection
Authors: Groeneveld A. B. Johan , Bossink Ailko W. J. , van Mierlo Gerard J. , Hack C. Erik 
Published: July 2002

ABSTRACT

The systemic host response to microbial infection involves clinical signs and symptoms of infection, including fever and elevated white blood cell (WBC) counts. In addition, inflammatory mediators are released, including activated complement product C3a, interleukin 6 (IL-6), and the acute-phase reactant secretory phospholipase A
2
(sPLA
2
). To compare the value of the latter with the former in predicting (the degree of) microbial infection at the bedside, we determined clinical variables and took blood samples daily for 3 consecutive days in 300 patients with a new fever (>38.0°C rectally or >38.3°C axillary). Microbiological culture results for 7 days after inclusion were collected. Patients were divided into clinical and microbial categories: those without and with a clinical focus of infection and those with negative cultures, with positive local cultures or specific stains for fungal (
n
= 13) or tuberculous infections (
n
= 1), and with positive blood cultures, including one patient with malaria parasitemia. The area under the curve (AUC) of the receiver operating characteristic (ROC) for prediction of positive cultures was 0.60 (
P
< 0.005) for peak temperature and 0.59 (
P
< 0.01) for peak WBC count, 0.60 (
P
< 0.005) for peak C3a, 0.63 (
P
< 0.001) for peak IL-6, and 0.61 (
P
< 0.001) for peak sPLA
2
. The AUC under the ROC curve for prediction of positive blood cultures was 0.68 (
P
< 0.001) for peak temperature and 0.56 for peak WBC count (
P
< 0.05). The AUC for peak C3a was 0.69, that for peak IL-6 was 0.70, and that for sPLA
2
was 0.67 (for all,
P
< 0.001). The degree of microbial invasion is thus a major determinant of the clinical and inflammatory host response in patients with fever. Moreover, circulating inflammatory mediators such as C3a and IL-6 may help to predict positive blood cultures, together with clinical signs and symptoms of the host response to microbial infection, even before culture results are available. This may help in the designing of entry criteria for therapeutic intervention studies.
Isolation and Characterization of Two European Strains ofEhrlichia phagocytophilaof Equine Origin
Authors: Bjöersdorff Anneli , Bagert Bodil , Massung Robert F. , Gusa Asiya , Eliasson Ingvar 
Published: July 2002

ABSTRACTWe report the isolation and partial genetic characterization of two equine strains of granulocyticEhrlichiaof the genogroupEhrlichia phagocytophila. Frozen whole-blood samples from two Swedish horses with laboratory-verified granulocytic ehrlichiosis were inoculated into HL-60 cell cultures. GranulocyticEhrlichiawas isolated and propagated from both horses. DNA extracts from the respective strains were amplified by PCR using primers directed towards the 16S rRNA gene, thegroESLheat shock operon gene, and theankgene. The amplified gene fragments were sequenced and compared to known sequences in the GenBank database. With respect to the 16S rRNA gene, thegroESLgene, and theankgene, the DNA sequences of the two equineEhrlichiaisolates were identical to sequences found in isolates from clinical cases of granulocytic ehrlichiosis in humans and domestic animals in Sweden. However, compared to amplified DNA from an AmericanEhrlichiastrain of theE.phagocytophilagenogroup, differences were found in thegroESLgene andankgene sequences.