bluuebunny/crossref_metadata_2025_split huggingface dataset:
Phylogenetic and Ecological Analysis of Novel Marine Stramenopiles
Authors: Massana Ramon , Castresana Jose , Balagué Vanessa , Guillou Laure , Romari Khadidja , Groisillier Agnès , Valentin Klaus , Pedrós-Alió Carlos 
Published: June 2004

ABSTRACTCulture-independent molecular analyses of open-sea microorganisms have revealed the existence and apparent abundance of novel eukaryotic lineages, opening new avenues for phylogenetic, evolutionary, and ecological research. Novel marine stramenopiles, identified by 18S ribosomal DNA sequences within the basal part of the stramenopile radiation but unrelated to any previously known group, constituted one of the most important novel lineages in these open-sea samples. Here we carry out a comparative analysis of novel stramenopiles, including new sequences from coastal genetic libraries presented here and sequences from recent reports from the open ocean and marine anoxic sites. Novel stramenopiles were found in all major habitats, generally accounting for a significant proportion of clones in genetic libraries. Phylogenetic analyses indicated the existence of 12 independent clusters. Some of these were restricted to anoxic or deep-sea environments, but the majority were typical components of coastal and open-sea waters. We specifically identified four clusters that were well represented in most marine surface waters (together they accounted for 74% of the novel stramenopile clones) and are the obvious targets for future research. Many sequences were retrieved from geographically distant regions, indicating that some organisms were cosmopolitan. Our study expands our knowledge on the phylogenetic diversity and distribution of novel marine stramenopiles and confirms that they are fundamental members of the marine eukaryotic picoplankton.
Spatial Distribution of Bacterial Communities and Phenanthrene Degradation in the Rhizosphere of
Lolium perenne
L
Authors: Corgié S. C. , Beguiristain T. , Leyval C. 
Published: June 2004

ABSTRACT
Rhizodegradation of organic pollutants, such as polycyclic aromatic hydrocarbons, is based on the effect of root-produced compounds, known as exudates. These exudates constitute an important and constant carbon source that selects microbial populations in the plant rhizosphere, modifying global as well as specific microbial activities. We conducted an experiment in two-compartment devices to show the selection of bacterial communities by root exudates and phenanthrene as a function of distance to roots. Using direct DNA extraction, PCR amplification, and thermal gradient gel electrophoresis screening, bacterial population profiles were analyzed in parallel to bacterial counts and quantification of phenanthrene biodegradation in three layers (0 to 3, 3 to 6, and 6 to 9 mm from root mat) of unplanted-polluted (phenanthrene), planted-polluted, and planted-unpolluted treatments. Bacterial community differed as a function of the distance to roots, in both the presence and the absence of phenanthrene. In the planted and polluted treatment, biodegradation rates showed a strong gradient with higher values near the roots. In the nonplanted treatment, bacterial communities were comparable in the three layers and phenanthrene biodegradation was high. Surprisingly, no biodegradation was detected in the section of planted polluted treatment farthest from the roots, where the bacterial community structure was similar to those of the nonplanted treatment. We conclude that root exudates and phenanthrene induce modifications of bacterial communities in polluted environments and spatially modify the activity of degrading bacteria.
The
Aspergillus parasiticus estA
-Encoded Esterase Converts Versiconal Hemiacetal Acetate to Versiconal and Versiconol Acetate to Versiconol in Aflatoxin Biosynthesis
Authors: Chang Perng-Kuang , Yabe Kimiko , Yu Jiujiang 
Published: June 2004

ABSTRACT

In aflatoxin biosynthesis, the pathway for the conversion of 1-hydroxyversicolorone to versiconal hemiacetal acetate (VHA) to versiconal (VHOH) is part of a metabolic grid. In the grid, the steps from VHA to VHOH and from versiconol acetate (VOAc) to versiconol (VOH) may be catalyzed by the same esterase. Several esterase activities are associated with the conversion of VHA to VHOH, but only one esterase gene (
estA
) is present in the complete aflatoxin gene cluster of
Aspergillus parasiticus
. We deleted the
estA
gene from
A. parasiticus
SRRC 2043, an
O
-methylsterigmatocystin (OMST)-accumulating strain. The
estA-
deleted mutants were pigmented and accumulated mainly VHA and versicolorin A (VA). A small amount of VOAc and other downstream aflatoxin intermediates, including VHOH, versicolorin B, and OMST, also were accumulated. In contrast, a VA-accumulating mutant, NIAH-9, accumulated VA exclusively and neither VHA nor VOAc were produced. Addition of the esterase inhibitor dichlorvos (dimethyl 2,2-dichlorovinylphosphate) to the transformation recipient strain RHN1, an
estA
-deleted mutant, or NIAH-9 resulted in the accumulation of only VHA and VOAc. In in vitro enzyme assays, the levels of the esterase activities catalyzing the conversion of VHA to VHOH in the cell extracts of two
estA
-deleted mutants were decreased to approximately 10% of that seen with RHN1. Similar decreases in the esterase activities catalyzing the conversion of VOAc to VOH were also obtained. Thus, the
estA
-encoded esterase catalyzes the conversion of both VHA to VHOH and VOAc to VOH during aflatoxin biosynthesis.
Inactivation of the Flagellin Gene
flaA
in
Magnetospirillum gryphiswaldense
Results in Nonmagnetotactic Mutants Lacking Flagellar Filaments
Authors: Schultheiss Daniel , Kube Michael , Schüler Dirk 
Published: June 2004

ABSTRACT

Magnetotactic bacteria synthesize magnetosomes, which cause them to orient and migrate along magnetic field lines. The analysis of magnetotaxis and magnetosome biomineralization at the molecular level has been hindered by the unavailability of genetic methods, namely the lack of a means to introduce directed gene-specific mutations. Here we report a method for knockout mutagenesis by homologous recombination in
Magnetospirillum gryphiswaldense
. Multiple flagellin genes, which are unlinked in the genome, were identified in
M. gryphiswaldense
. The targeted disruption of the flagellin gene
flaA
was shown to eliminate flagella formation, motility, and magnetotaxis. The techniques described in this paper will make it possible to take full advantage of the forthcoming genome sequences of
M. gryphiswaldense
and other magnetotactic bacteria.
Comparison of Total Culturable Virus Assay and Multiplex Integrated Cell Culture-PCR for Reliability of Waterborne Virus Detection
Authors: Lee Hwa Kyung , Jeong Yong Seok 
Published: June 2004

ABSTRACT
The total culturable virus assay (TCVA) and an integrated cell culture-PCR (ICC-PCR) were compared in parallel to evaluate their detection reliability. Source, finished, and tap water samples from three drinking water treatment plant systems were analyzed by TCVA, and every cell culture dish was subsequently examined by reverse transcription (RT) multiplex PCR using enterovirus- and adenovirus-specific primers. Twenty-seven of 180 (15%) inoculated dishes exhibited cytopathic effects (CPE). Virus concentrations for source water ranged from 3.3 to 21.0 most probable numbers of infectious units (MPN) per 100 liters. No finished or tap water samples were positive. On the other hand, 38 (21%) of the dishes were positive in multiplex ICC-PCR. Virus concentrations ranged from 4.5 to 10.2 MPN/100 liters for source water and 0 to 0.9 MPN/100 liters for finished and tap water. In spite of its superior sensitivity, the ICC-PCR assay resulted in lower virus concentration values than the TCVA for two of the source water sites. Retest of the CPE-positive dishes using reovirus-specific RT-PCR revealed that 24 of the 27 (89%) dishes were also positive for reoviruses. These observations suggested that the detection reliability of ICC-PCR is restricted by the primer sets that are integrated in the reaction mixture. The observation of an uneven distribution of PCR-positive culture dishes in a given sample raises an additional caution that simple extrapolation of the ICC-PCR result from the analysis of a limited fraction of collected samples should be avoided to minimize possible over- and underestimation of the amount of virus.
Comparison of Coliforms and Coliphages as Tools for Assessment of Viral Contamination in River Water
Authors: Skraber S. , Gassilloud B. , Gantzer C. 
Published: June 2004

ABSTRACT

The aim of the study was to evaluate the presence of pathogenic viruses in the Moselle River and to compare the usefulness of thermotolerant coliforms and somatic coliphages as tools for river water quality assessment in terms of viral contamination. Thermotolerant coliforms and somatic coliphages were enumerated by standardized methods in 170 samples of river water drawn from five sampling sites along the Moselle River (eastern France). BGM cell culture and integrated cell culture-reverse transcription-PCR DNA enzyme immunoassay were used to determine the presence of pathogenic viral genome (
Enterovirus
and
Norovirus
genogroup II [GGII]) and infectious
Enterovirus
spp. in 90 1-liter samples. No infectious
Enterovirus
spp. were isolated, but
Enterovirus
and
Norovirus
GGII genomes were detected in 38% of the samples.
Norovirus
GGII genome was mostly detected in winter, whereas
Enterovirus
genome was mostly detected in summer and fall. Somatic coliphages appeared to be less sensitive to higher river water temperature than thermotolerant coliforms. Furthermore, the number of river water samples positive for pathogenic viral genome increased with increasing concentration of somatic coliphages, whereas coliform concentration was unrelated to viral genome contamination. Consequently somatic coliphages, which are less sensitive to environmental factors than thermotolerant coliforms in river water, would provide a promising tool for assessment of river water quality in terms of fecal and viral pollution.
Isolation, Characterization, and Identification of Bacterial Contaminants in Semifinal Gelatin Extracts
Authors: De Clerck E. , Vanhoutte T. , Hebb T. , Geerinck J. , Devos J. , De Vos P. 
Published: June 2004

ABSTRACTBacterial contamination of gelatin is of great concern. Indeed, this animal colloid has many industrial applications, mainly in food and pharmaceutical products. In a previous study (E. De Clerck and P. De Vos, Syst. Appl. Microbiol.25:611-618), contamination of a gelatin production process with a variety of gram-positive and gram-negative bacteria was demonstrated. In this study, bacterial contamination of semifinal gelatin extracts from several production plants was examined. Since these extracts are subjected to harsh conditions during production and a final ultrahigh-temperature treatment, the bacterial load at this stage is expected to be greatly reduced. In total, 1,129 isolates were obtained from a total of 73 gelatin batches originating from six different production plants. Each of these batches was suspected of having bacterial contamination based on quality control testing at the production plant from which it originated. For characterization and identification of the 1,129 bacterial isolates, repetitive-element PCR was used to obtain manageable groups. Representative strains were identified by means of 16S rRNA genesequencing, species-specificgyrBPCR, andgyrAandrpoBsequencing and were tested for gelatinase activity. The majority of isolates belonged to members ofBacillusor related endospore-forming genera. Representative strains were identified asBacillus cereus,Bacillus coagulans,Bacillus fumarioli,Bacillus amyloliquefaciens,Bacillus licheniformis,Bacillus pumilus,Bacillus sonorensis,Bacillus subtilis,Bacillus gelatini,Bacillus thermoamylovorans,Anoxybacillus contaminans,Anoxybacillus flavithermus,Brevibacillus agri,Brevibacillus borstelensis, andGeobacillus stearothermophilus. The majority of these species include strains exhibiting gelatinase activity. Moreover, some of these species have known pathogenic properties. These findings are of great concern with regard to the safety and quality of gelatin and its applications.
Identification of Differential Gene Expression in Bacteria Associated with Coral Black Band Disease by Using RNA-Arbitrarily Primed PCR
Authors: Frias-Lopez Jorge , Bonheyo George T. , Fouke Bruce W. 
Published: June 2004

ABSTRACT

RNA-arbitrarily primed PCR techniques have been applied for the first time to identify differential gene expression in black band disease (BBD), a virulent coral infection that affects reef ecosystems worldwide. The gene activity for the BBD mat on infected surfaces of the brain coral
Diploria strigosa
was compared with that for portions of the BBD mat that were removed from the coral and suspended nearby in the seawater column. The results obtained indicate that three genes (DD 95-2, DD 95-4, and DD 99-9) were up-regulated in the BBD bacterial mat on the coral surface compared to the transcript base levels observed in the BBD mat suspended in seawater. Clone DD 95-4 has homology with known amino acid ABC transporter systems in bacteria, while clone DD 99-9 exhibits homology with chlorophyll A apoprotein A1 in cyanobacteria. This protein is essential in the final conformation of photosystem I P700. DD 95-2, the only gene that was fully repressed in the BBD mat samples suspended in seawater, exhibited homology with the AraC-type DNA binding domain-containing proteins. These transcriptional activators coordinate the expression of genes essential for virulence in many species of gram-negative bacteria.
Transformation of
Leuconostoc carnosum
4010 and Evidence for Natural Competence of the Organism
Authors: Helmark Søren , Hansen Mette E. , Jelle Birthe , Sørensen Kim I. , Jensen Peter R. 
Published: June 2004

ABSTRACT

Plasmid transformation in
Leuconostoc carnosum
4010 was analyzed. A successful transformation protocol for
L. carnosum
was established by modifying an existing protocol for
Lactococcus lactis
. Several parameters, including the number of generations that the cells had grown at the time of harvest, glycine concentration, the time of incubation for phenotypic expression, and the electrical field strength, were investigated and proved to have influence on the transformation frequency. Electrocompetence was found to be transient and to peak in the early exponential growth phase. Optimized conditions resulted in transformation frequencies of up to 6.7 × 10
5
transformants per microgram of plasmid DNA. A total of five plasmids in
L. carnosum
were successfully introduced and maintained. Interestingly, we discovered that DNA uptake was of a frequency of 3 × 10
−6
to 19 × 10
−6
transformants per CFU in the absence of an applied electrical field. We concluded that
L. carnosum
is naturally competent.
Isolation and Phylogenetic Analysis of Novel Viruses Infecting the Phytoplankton
Phaeocystis globosa
(Prymnesiophyceae)
Authors: Brussaard C. P. D. , Short S. M. , Frederickson C. M. , Suttle C. A. 
Published: June 2004

ABSTRACT

Viruses infecting the harmful bloom-causing alga
Phaeocystis globosa
(Prymnesiophyceae) were readily isolated from Dutch coastal waters (southern North Sea) in 2000 and 2001. Our data show a large increase in the abundance of putative
P. globosa
viruses during blooms of
P. globosa
, suggesting that viruses are an important source of mortality for this alga. In order to examine genetic relatedness among viruses infecting
P. globosa
and other phytoplankton, DNA polymerase gene (
pol
) fragments were amplified and the inferred amino acid sequences were phylogenetically analyzed. The results demonstrated that viruses infecting
P. globosa
formed a closely related monophyletic group within the family
Phycodnaviridae
, with at least 96.9% similarity to each other. The sequences grouped most closely with others from viruses that infect the prymnesiophyte algae
Chrysochromulina brevifilum
and
Chrysochromulina strobilus
. Whether the
P. globosa
viruses belong to the genus
Prymnesiovirus
or form a separate group needs further study. Our data suggest that, like their phytoplankton hosts, the
Chrysochromulina
and
Phaeocystis
viruses share a common ancestor and that these prymnesioviruses and their algal host have coevolved.
Use of
Sinorhizobium meliloti
as an Indicator for Specific Detection of Long-Chain
N
-Acyl Homoserine Lactones
Authors: Llamas Inmaculada , Keshavan Neela , González Juan E. 
Published: June 2004

ABSTRACT

Population-density-dependent gene expression in gram-negative bacteria involves the production of signal molecules characterized as
N
-acyl homoserine lactones (AHLs). The synthesis of AHLs by numerous microorganisms has been identified by using biosensor strains based on the
Agrobacterium tumefaciens
and
Chromobacterium violaceum
quorum-sensing systems. The symbiotic nitrogen-fixing bacterium
Sinorhizobium meliloti
is rapidly becoming a model organism for the study of quorum sensing. This organism harbors at least three different quorum-sensing systems (Sin, Mel, and Tra), which play a role in its symbiotic relationship with its host plant, alfalfa. The Sin system is distinguished among them for the production of long-chain AHLs, including C
18
-HL, the longest AHL reported so far. In this work, we show that construction of a
sinI
::
lacZ
transcriptional fusion results in a strain that detects long-chain AHLs with exquisite sensitivity. Overexpression of the SinR regulator protein from a vector promoter increases its sensitivity without loss of specificity. We also show that the resulting indicator strain can recognize long-chain AHLs produced by unrelated bacteria such as
Paracoccus denitrificans
and
Rhodobacter capsulatus
. This
S. meliloti
indicator strain should serve as a tool for the specific detection of long-chain AHLs in new systems.
Protein Expression by
Streptococcus mutans
during Initial Stage of Biofilm Formation
Authors: Welin J. , Wilkins J. C. , Beighton D. , Svensäter G. 
Published: June 2004

ABSTRACT

Cells growing on surfaces in biofilms exhibit properties distinct from those of planktonic cells, such as increased resistance to biocides and antimicrobial agents. In spite of increased interest in biofilms, very little is known about alterations in cell physiology that occur upon attachment of cells to a surface. In this study we have investigated the changes induced in the protein synthesis by contact of
Streptococcus mutans
with a surface. Log-phase planktonic cells of
S. mutans
were allowed to adhere to a glass slide for 2 h in the presence of a
14
C-amino acid mixture. Nonadhered cells were washed away, and the adhered cells were removed by sonication. The proteins were extracted from the nonadhered planktonic and the adhered biofilm cells and separated by two-dimensional gel electrophoresis followed by autoradiography and image analysis. Image analysis revealed that the relative rate of synthesis of 25 proteins was enhanced and that of 8 proteins was diminished ≥1.3-fold in the biofilm cells. Proteins of interest were identified by mass spectrometry and computer-assisted protein sequence analysis. Of the 33 proteins associated with the adhesion response, all but 10 were identified by mass spectrometry and peptide mass fingerprinting. The most prominent change in adhered cells was the increase in relative synthesis of enzymes involved in carbohydrate catabolism indicating that a redirection in protein synthesis towards energy generation is an early response to contact with and adhesion to a surface.
Monitoring gp43 Antigenemia in Paracoccidioidomycosis Patients during Therapy
Authors: Marques da Silva Silvia Helena , Queiroz-Telles Flávio , Colombo Arnaldo Lopes , Blotta Maria Heloisa Souza Lima , Lopes José Daniel , Pires de Camargo Zoilo 
Published: June 2004

ABSTRACT

Paracoccidioidomycosis (PCM) is a systemic fungal disease that is particularly important among individuals living and working in rural areas of endemicity in Latin America. Detection of anti-
Paracoccidioides brasiliensis
antibodies is of limited value due to false-negative results. Detection of
P. brasiliensis
-gp43 circulating antigen is a practical approach for a specific diagnosis of the disease. In a previous study we described an inhibition enzyme-linked immunosorbent assay able to detect the 43-kDa
P. brasiliensis
antigen in sera of 100% of patients with the acute form of PCM and in 95.31 and 100% of patients with the chronic multifocal and unifocal forms of PCM. To investigate its potential application for the follow-up of PCM patients during treatment, antigen levels were monitored at regular intervals for up 8 to 12 months in serum samples from 23 patients. The results showed that treatment with itraconazole resulted in decreasing levels of circulating gp43 that were correlated with the reduction of anti-gp43 antibodies. It was also observed that by the end of 12 months of treatment gp43 levels were <5 μg/ml in all patients.
SHV-12-Like Extended-Spectrum-β-Lactamase-Producing Strains of
Salmonella enterica
Serotypes Babelsberg and Enteritidis Isolated in France among Infants Adopted from Mali
Authors: Weill François-Xavier , Demartin Marie , Tandé Didier , Espié Emmanuelle , Rakotoarivony Ignace , Grimont Patrick A. D. 
Published: June 2004

ABSTRACT

From December 2002 to June 2003, 14 cultures of
Salmonella enterica
serotype Babelsberg and 6 cultures of serotype Enteritidis, isolated in France from internationally adopted children, were identified at the French National Reference Center for
Salmonella
. All serotype Babelsberg isolates were related, as determined by pulsed-field gel electrophoresis, and all serotype Enteritidis strains displayed the same phage type. All serotype Enteritidis and seven serotype Babelsberg isolates produced an SHV-12-like extended-spectrum β-lactamase as determined by sequencing of PCR products and by isoelectrofocusing. Some serotype Enteritidis isolates exhibited additional antimicrobial resistance (aminoglycosides, tetracycline, chloramphenicol, sulfonamides, and trimethoprim). Our investigation indicated that these
Salmonella
isolates were certainly acquired in the same orphanage in Bamako, Mali, before the children were adopted by French families. An inappropriate use of ceftriaxone was probably the cause of the emergence of such strains. There is an urgent need to determine the origin of the contamination and to introduce adequate antibiotic protocols into this orphanage to prevent further transmission and dissemination. Screening for infections and follow-up, adapted to the origin of the internationally adopted children, should be recommended.
Molecular and Morphological Identification ofColletotrichumSpecies of Clinical Interest
Authors: Cano Josep , Guarro Josep , Gené Josepa 
Published: June 2004

ABSTRACTColletotrichumspecies have caused human infections in recent years. Because of the difficulties in recognizing them in vitro, we have designed a quick and unambiguous molecular test, based on the amplification of a specific fragment of the internal transcribed spacer 1 region, to distinguish anyColletotrichumisolate from other fungi, including the common pathogenic species. Analysis of the sequences of the ribosomal DNA (rDNA) fragment showed sufficient variability to clearly separate the five species ofColletotrichumthat are of clinical interest, i.e.,Colletotrichum coccodes,C. crassipes,C. dematium,C. gloeosporioides, andC. graminicola.Sequencing of the D1-D2 region of the large-subunit rDNA gene also supported these results. Additionally, we reviewed the most suitable morphological characteristics for the in vitro identification of these increasingly important opportunistic fungi.