bluuebunny/crossref_metadata_2025_split huggingface dataset:
Quantitative Tracing, byTaqNuclease Assays, of aSynechococcusEcotype in a Highly Diversified Natural Population
Authors: Becker Sven , Fahrbach Michael , Böger Peter , Ernst Anneliese 
Published: August 2002

ABSTRACTQuantitativeTaqnuclease assays (TNAs) (TaqMan PCR), nested PCR in combination with denaturing gradient gel electrophoresis (DGGE), and epifluorescence microscopy were used to analyze the autotrophic picoplankton (APP) of Lake Constance. Microscopic analysis revealed dominance of phycoerythrin (PE)-richSynechococcusspp. in the pelagic zone of this lake. Cells passing a 3-μm-pore-size filter were collected during the growth period of the years 1999 and 2000. The diversity of PE-richSynechococcusspp. was examined using DGGE to analyze GC-clamped amplicons of a noncoding section of the 16S-23S intergenic spacer in the ribosomal operon. In both years, genotypes represented by three closely related PE-richSynechococcusstrains of our culture collection dominated the population, while other isolates were traced sporadically or were not detected in their original habitat by this method. For TNAs, primer-probe combinations for two taxonomic levels were used, one to quantify genomes of all knownSynechococcus-type cyanobacteria in the APP of Lake Constance and one to enumerate genomes of a single ecotype represented by the PE-rich isolateSynechococcussp. strain BO 8807. During the growth period, genome numbers of knownSynechococcusspp. varied by 2 orders of magnitude (2.9 × 103to 3.1 × 105genomes per ml). The ecotypeSynechococcussp. strain BO 8807 was detected in every sample at concentrations between 1.6 × 101and 1.3 × 104genomes per ml, contributing 0.02 to 5.7% of the quantified cyanobacterial picoplankton. Although the quantitative approach taken in this study has disclosed several shortcomings in the sampling and detection methods, this study demonstrated for the first time the extensive internal dynamics that lie beneath the seemingly arbitrary variations of a population of microbial photoautotrophs in the pelagic habitat.
Evidence for Natural Horizontal Transfer of thepcpBGene in the Evolution of Polychlorophenol-Degrading Sphingomonads
Authors: Tiirola Marja A. , Wang Hong , Paulin Lars , Kulomaa Markku S. 
Published: August 2002

ABSTRACTThe chlorophenol degradation pathway inSphingobium chlorophenolicumis initiated by thepcpBgene product, pentachlorophenol-4-monooxygenase. The distribution of the gene was studied in a phylogenetically diverse group of polychlorophenol-degrading bacteria isolated from contaminated groundwater in Kärkölä, Finland. All the sphingomonads isolated were shown to sharepcpBgene homologs with 98.9 to 100% sequence identity. The gene product was expressed when the strains were induced by 2,3,4,6-tetrachlorophenol. A comparative analysis of the 16S rDNA andpcpBgene trees suggested that a recent horizontal transfer of thepcpBgene was involved in the evolution of the catabolic pathway in the Kärkölä sphingomonads. The full-length KärköläpcpBgene allele had approximately 70% identity with the threepcpBgenes previously sequenced from sphingomonads. It was very closely related to the environmental clones obtained from chlorophenol-enriched soil samples (M. Beaulieu, V. Becaert, L. Deschenes, and R. Villemur, Microbiol. Ecol.40:345-355, 2000). The gene was not present in polychlorophenol-degrading nonsphingomonads isolated from the Kärkölä source.
Rhamnolipid Stimulates Uptake of Hydrophobic Compounds by
Pseudomonas aeruginosa
Authors: Noordman Wouter H. , Janssen Dick B. 
Published: August 2002

ABSTRACT

The biodegradation of hexadecane by five biosurfactant-producing bacterial strains (
Pseudomonas aeruginosa
UG2
, Acinetobacter calcoaceticus
RAG1,
Rhodococcus erythropolis
DSM 43066,
R. erythropolis
ATCC 19558, and strain BCG112) was determined in the presence and absence of exogenously added biosurfactants. The degradation of hexadecane by
P. aeruginosa
was stimulated only by the rhamnolipid biosurfactant produced by the same organism. This rhamnolipid did not stimulate the biodegradation of hexadecane by the four other strains to the same extent, nor was degradation of hexadecane by these strains stimulated by addition of their own biosurfactants. This suggests that
P. aeruginosa
has a mode of hexadecane uptake different from those of the other organisms. Rhamnolipid also enhanced the rate of epoxidation of the aliphatic hydrocarbon α,ω-tetradecadiene by a cell suspension of
P. aeruginosa
. Furthermore, the uptake of the hydrophobic probe 1-naphthylphenylamine by cells of
P. aeruginosa
was enhanced by rhamnolipid, as indicated by stopped-flow fluorescence experiments. Rhamnolipid did not stimulate the uptake rate of this probe in de-energized cells. These results indicate that an energy-dependent system is present in
P. aeruginosa
strain UG2 that mediates fast uptake of hydrophobic compounds in the presence of rhamnolipid.
Lipopeptide Production in
Pseudomonas
sp. Strain DSS73 Is Regulated by Components of Sugar Beet Seed Exudate via the Gac Two-Component Regulatory System
Authors: Koch Birgit , Nielsen Tommy H. , Sørensen Dan , Andersen Jens Bo , Christophersen Carsten , Molin Søren , Givskov Michael , Sørensen Jan , Nybroe Ole 
Published: August 2002

ABSTRACT

Pseudomonas
sp. strain DSS73 isolated from the sugar beet rhizosphere produces the cyclic lipopeptide amphisin, which inhibits the growth of plant-pathogenic fungi. By Tn
5
::
luxAB
mutagenesis, we obtained two nonproducing mutant strains, DSS73-15C2 and DSS73-12H8. The gene interrupted by the transposon in strain DSS73-15C2 (
amsY
) encoded a protein with homology to peptide synthetases that was designated amphisin synthetase. DSS73-12H8 carried the transposon in a regulatory gene encoding a protein with homology to the sensor kinase GacS. Growth of strain DSS73-15C2 (
amsY
) was impaired during the transition to stationary phase in a minimal medium amended with an exudate of sugar beet seeds. This growth phenotype could be complemented by purified amphisin. Seed exudate further induced expression of bioluminescence from the
amsY
::
luxAB
reporter during the transition to stationary phase. This agreed with an increase in amphisin production by the DSS73 wild-type strain during early stationary phase. Amphisin synthesis in DSS73 was strictly dependent on GacS, and even induction by seed exudate depended on a functional
gacS
locus. Hence, a signal triggering the GacS/GacA two-component system appeared to be present in the seed exudate.
Construction of Yeast Strains with High Cell Surface Lipase Activity by Using Novel Display Systems Based on the Flo1p Flocculation Functional Domain
Authors: Matsumoto Takeshi , Fukuda Hideki , Ueda Mitsuyoshi , Tanaka Atsuo , Kondo Akihiko 
Published: August 2002

ABSTRACT

We constructed a novel cell-surface display system, using as a new type of cell-wall anchor 3,297 or 4,341 bp of the 3′ region of the FLO1 gene (
FS
or
FL
gene, respectively), which encodes the flocculation functional domain of Flo1p. In this system, the N terminus of the target protein was fused to the FS or FL protein and the fusion proteins were expressed under the control of the inducible promoter
UPR-ICL
(5′ upstream region of the isocitrate lyase of
Candida tropicalis
). Using this new system, recombinant lipase with a pro sequence from
Rhizopus oryzae
(rProROL), which has its active site near the C terminus, was displayed on the cell surface. Cell-surface display of the FSProROL and FLProROL fusion proteins was confirmed by immunofluorescence microscopy and immunoblotting. Lipase activity reached 145 IU/liter (61.3 IU/g [dry cell weight]) on the surface of the yeast cells, which successfully catalyzed the methanolysis reaction. Using these whole-cell biocatalysts, methylesters synthesized from triglyceride and methanol reached 78.3% after 72 h of reaction. To our knowledge, this is the first example of cell-surface display of lipase with high activity. Interestingly, the yeast cells displaying the FLProROL protein showed strong flocculation, even though the glycosylphosphatidylinositol anchor attachment signal and cell-membrane-anchoring region of Flo1p had been deleted from this gene. The cell-surface display system based on FL thus endows the yeast strain with both novel enzyme display and strong flocculation ability.
Environmental Factors Influencing Human Viral Pathogens and Their Potential Indicator Organisms in the Blue Mussel,
Mytilus edulis
: the First Scandinavian Report
Authors: Hernroth Bodil E. , Conden-Hansson Ann-Christine , Rehnstam-Holm Ann-Sofi , Girones Rosina , Allard Annika K. 
Published: August 2002

ABSTRACT

This study was carried out in order to investigate human enteric virus contaminants in mussels from three sites on the west coast of Sweden, representing a gradient of anthropogenic influence. Mussels were sampled monthly during the period from February 2000 to July 2001 and analyzed for adeno-, entero-, Norwalk-like, and hepatitis A viruses as well as the potential viral indicator organisms somatic coliphages, F-specific RNA bacteriophages, bacteriophages infecting
Bacteroides fragilis
, and
Escherichia coli
. The influence of environmental factors such as water temperature, salinity, and land runoff on the occurrence of these microbes was also included in this study. Enteric viruses were found in 50 to 60% of the mussel samples, and there were no pronounced differences between the samples from the three sites.
E. coli
counts exceeded the limit for category A for shellfish sanitary safety in 40% of the samples from the sites situated in fjords. However, at the site in the outer archipelago, this limit was exceeded only once, in March 2001, when extremely high levels of atypical indole-negative strains of
E. coli
were registered at all three sites. The environmental factors influenced the occurrence of viruses and phages differently, and therefore, it was hard to find a coexistence between them. This study shows that, for risk assessment, separate modeling should be done for every specific area, with special emphasis on environmental factors such as temperature and land runoff. The present standard for human fecal contamination,
E. coli
, seems to be an acceptable indicator of only local sanitary contamination; it is not a reliable indicator of viral contaminants in mussels. To protect consumers and get verification of “clean” mussels, it seems necessary to analyze for viruses as well. The use of a molecular index of the human contamination of Swedish shellfish underscores the need for reference laboratories with high-technology facilities.
Fumarate-Mediated Inhibition of Erythrose Reductase, a Key Enzyme for Erythritol Production by
Torula corallina
Authors: Lee Jung-Kul , Koo Bong-Seong , Kim Sang-Yong 
Published: August 2002

ABSTRACT

Torula corallina
, a strain presently being used for the industrial production of erythritol, has the highest erythritol yield ever reported for an erythritol-producing microorganism. The increased production of erythritol by
Torula corallina
with trace elements such as Cu
2+
has been thoroughly reported, but the mechanism by which Cu
2+
increases the production of erythritol has not been studied. This study demonstrated that supplemental Cu
2+
enhanced the production of erythritol, while it significantly decreased the production of a major by-product that accumulates during erythritol fermentation, which was identified as fumarate by instrumental analyses. Erythrose reductase, a key enzyme that converts erythrose to erythritol in
T. corallina
, was purified to homogeneity by chromatographic methods, including ion-exchange and affinity chromatography. In vitro, purified erythrose reductase was significantly inhibited noncompetitively by increasing the fumarate concentration. In contrast, the enzyme activity remained almost constant regardless of Cu
2+
concentration. This suggests that supplemental Cu
2+
reduced the production of fumarate, a strong inhibitor of erythrose reductase, which led to less inhibition of erythrose reductase and a high yield of erythritol. This is the first report that suggests catabolite repression by a tricarboxylic acid cycle intermediate in
T. corallina
.
The Bacterivorous Soil Flagellate
Heteromita globosa
Reduces Bacterial Clogging under Denitrifying Conditions in Sand-Filled Aquifer Columns
Authors: Mattison Richard G. , Taki Hironori , Harayama Shigeaki 
Published: August 2002

ABSTRACT

An exopolymer (slime)-producing soil bacterium
Pseudomonas
sp. (strain PS+) rapidly clogged sand-filled columns supplied with air-saturated artificial groundwater containing glucose (500 mg liter
−1
) as a sole carbon source and nitrate (300 mg liter
−1
) as an alternative electron acceptor. After 80 days of operation under denitrifying conditions, the effective porosity and saturated hydraulic conductivity (permeability) of sand in these columns had fallen by 2.5- and 26-fold, respectively. Bacterial biofilms appeared to induce clogging by occluding pore spaces with secreted exopolymer, although there may also have been a contribution from biogas generated during denitrification. The bacterivorous soil flagellate
Heteromita globosa
minimized reductions in effective porosity (1.6-fold) and permeability (13-fold), presumably due to grazing control of biofilms. Grazing may have limited growth of bacterial biomass and hence the rate of exopolymer and biogas secretion into pore spaces. Evidence for reduction in biogas production is suggested by increased nitrite efflux from columns containing flagellates, without a concomitant increase in nitrate consumption. There was no evidence that flagellates could improve flow conditions if added once clogging had occurred (60 days). Presumably, bacterial biofilms and their secretions were well established at that time. Nevertheless, this study provides evidence that bacterivorous flagellates may play a positive role in maintaining permeability in aquifers undergoing remediation treatments.
Enzymatic Properties and Intracellular Localization of the Novel
Trichoderma reesei
β-Glucosidase BGLII (Cel1A)
Authors: Saloheimo Markku , Kuja-Panula Juha , Ylösmäki Erkko , Ward Michael , Penttilä Merja 
Published: August 2002

ABSTRACT

This paper describes the characterization of an intracellular β-glucosidase enzyme BGLII (Cel1a) and its gene (
bgl2
) from the cellulolytic fungus
Trichoderma reesei
(
Hypocrea jecorina
). The expression pattern of
bgl2
is similar to that of other cellulase genes known from this fungus, and the gene would appear to be under the control of carbon catabolite repression mediated by the
cre1
gene. The BGLII protein was produced in
Escherichia coli
, and its enzymatic properties were analyzed. It was shown to be a specific β-glucosidase, having no β-galactosidase side activity. It hydrolyzed both cellotriose and cellotetraose. BGLII exhibited transglycosylation activity, producing mainly cellotriose from cellobiose and sophorose and cellobiose from glucose. Antibodies raised against BGLII showed the presence of the enzyme in
T. reesei
cell lysates but not in the culture supernatant. Activity measurements and Western blot analysis of
T. reesei
strains expressing
bgl2
from a constitutive promoter further confirmed the intracellular localization of this β-glucosidase.
Unveiling the Organisms behind Novel Eukaryotic Ribosomal DNA Sequences from the Ocean
Authors: Massana Ramon , Guillou Laure , Díez Beatriz , Pedrós-Alió Carlos 
Published: August 2002

ABSTRACTDespite the fact that the smallest eukaryotes (cells less than 5 μm in diameter) play key roles in marine food webs, particularly in open oligotrophic areas, the study of their in situ diversity started just one year ago. Perhaps the most remarkable finding of the most recent studies has been the discovery of completely new phylogenetic lineages, such as novel clades belonging to the stramenopile and alveolate phyla. The two new groups account for a significant fraction of clones in genetic libraries from North Atlantic, equatorial Pacific, Antarctic, and Mediterranean Sea waters. However, the identities and ecological relevance of these organisms remain unknown. Here we investigate the phylogenetic relationships, morphology, in situ abundance, and ecological role of novel stramenopiles. They form at least eight independent clades within the stramenopile basal branches, indicating a large phylogenetic diversity within the group. Two lineages were visualized and enumerated in field samples and enrichments by fluorescent in situ hybridization using specific rRNA-targeted oligonucleotide probes. The targeted organisms were 2- to 3-μm-diameter, round-shaped, nonpigmented flagellates. Further, they were found to be bacterivorous. One lineage accounted for up to 46% (average during an annual cycle, 19%) of heterotrophic flagellates in a coastal environment, providing evidence that novel stramenopiles are important and unrecognized components of the total stock of bacterial grazers.
Rickettsia monacensis
sp. nov., a Spotted Fever Group Rickettsia, from Ticks (
Ixodes ricinus
) Collected in a European City Park
Authors: Simser Jason A. , Palmer Ann T. , Fingerle Volker , Wilske Bettina , Kurtti Timothy J. , Munderloh Ulrike G. 
Published: August 2002

ABSTRACT

We describe the isolation and characterization of
Rickettsia monacensis
sp. nov. (type strain, IrR/Munich
T
) from an
Ixodes ricinus
tick collected in a city park, the English Garden in Munich, Germany. Rickettsiae were propagated in vitro with
Ixodes scapularis
cell line ISE6. BLAST analysis of the 16S rRNA, the citrate synthase, and the partial 190-kDa rickettsial outer membrane protein A (rOmpA) gene sequences demonstrated that the isolate was a spotted fever group (SFG) rickettsia closely related to several yet-to-be-cultivated rickettsiae associated with
I. ricinus
. Phylogenetic analysis of partial
rompA
sequences demonstrated that the isolate was genotypically different from other validated species of SFG rickettsiae.
R. monacensis
also replicated in cell lines derived from the ticks
I. ricinus
(IRE11) and
Dermacentor andersoni
(DAE100) and in the mammalian cell lines L-929 and Vero, causing cell lysis. Transmission electron microscopy of infected ISE6 and Vero cells showed rickettsiae within the cytoplasm, pseudopodia, nuclei, and vacuoles. Hamsters inoculated with
R. monacensis
had immunoglobulin G antibody titers as high as 1:16,384, as determined by indirect immunofluorescence assay. Western blot analyses demonstrated that the hamster sera cross-reacted with peptides from other phylogenetically distinct rickettsiae, including rOmpA.
R. monacensis
induced actin tails in both tick and mammalian cells similar to those reported for
R. rickettsii. R. monacensis
joins a growing list of SFG rickettsiae that colonize ticks but whose infectivity and pathogenicity for vertebrates are unknown.
Identification and Isolation of Two Ascomycete Fungi from Spores of the Arbuscular Mycorrhizal Fungus
Scutellospora castanea
Authors: Hijri Mohamed , Redecker Dirk , Petetot Jean A. MacDonald-Comber , Voigt Kerstin , Wöstemeyer Johannes , Sanders Ian R. 
Published: August 2002

ABSTRACT

Two filamentous fungi with different phenotypes were isolated from crushed healthy spores or perforated dead spores of the arbuscular mycorrhizal fungus (AMF)
Scutellospora castanea
. Based on comparative sequence analysis of 5.8S ribosomal DNA and internal transcribed spacer fragments, one isolate, obtained from perforated dead spores only, was assigned to the genus
Nectria
, and the second, obtained from both healthy and dead spores, was assigned to
Leptosphaeria
, a genus that also contains pathogens of plants in the Brassicaceae. PCR and randomly amplified polymorphic DNA-PCR analyses, however, did not indicate similarities between pathogens and the isolate. The presence of the two isolates in both healthy spores and perforated dead spores of
S. castanea
was finally confirmed by transmission electron microscopy by using distinctive characteristics of the isolates and
S. castanea
. The role of this fungus in
S. castanea
spores remains unclear, but the results serve as a strong warning that sequences obtained from apparently healthy AMF spores cannot be presumed to be of glomalean origin and that this could present problems for studies on AMF genes.
Isolation and Characterization of Differentially Expressed Genes in the Mycelium and Fruit Body of
Tuber borchii
Authors: Lacourt Isabelle , Duplessis Sébastien , Abbà Simona , Bonfante Paola , Martin Francis 
Published: August 2002

ABSTRACT

The transition from vegetative mycelium to fruit body in truffles requires differentiation processes which lead to edible fruit bodies (ascomata) consisting of different cell and tissue types. The identification of genes differentially expressed during these developmental processes can contribute greatly to a better understanding of truffle morphogenesis. A cDNA library was constructed from vegetative mycelium RNAs of the white truffle
Tuber borchii
, and 214 cDNAs were sequenced. Up to 58% of the expressed sequence tags corresponded to known genes. The majority of the identified sequences represented housekeeping proteins, i.e., proteins involved in gene or protein expression, cell wall formation, primary and secondary metabolism, and signaling pathways. We screened 171 arrayed cDNAs by using cDNA probes constructed from mRNAs of vegetative mycelium and ascomata to identify fruit body-regulated genes. Comparisons of signals from vegetative mycelium and fruit bodies bearing 15 or 70% mature spores revealed significant differences in the expression levels for up to 33% of the investigated genes. The expression levels for six highly regulated genes were confirmed by RNA blot analyses. The expression of glutamine synthetase, 5-aminolevulinic acid synthetase, isocitrate lyase, thioredoxin, glucan 1,3-β-glucosidase, and UDP-glucose:sterol glucosyl transferase was highly up-regulated, suggesting that amino acid biosynthesis, the glyoxylate cycle pathway, and cell wall synthesis are strikingly altered during morphogenesis.
Interaction of Gene-Cloned and Insect Cell-Expressed Aminopeptidase N of
Spodoptera litura
with Insecticidal Crystal Protein Cry1C
Authors: Agrawal Neema , Malhotra Pawan , Bhatnagar Raj K. 
Published: August 2002

ABSTRACT

Insecticidal toxins produced by
Bacillus thuringiensis
interact with specific receptors located in the midguts of susceptible larvae, and the interaction is followed by a series of biochemical events that lead to the death of the insect. In order to elucidate the mechanism of action of
B. thuringiensis
toxins, receptor protein-encoding genes from many insect species have been cloned and characterized. In this paper we report the cloning, expression, and characterization of Cry toxin-interacting aminopeptidase N (APN) isolated from the midgut of a polyphagous pest,
Spodoptera litura
. The
S. litura
APN cDNA was expressed in the Sf21 insect cell line by using a baculovirus expression system. Immunofluorescence staining of the cells revealed that the expressed APN was located at the surface of Sf21 cells. Treatment of Sf21 cells expressing
S. litura
APN with phosphatidylinositol-specific phospholipase C demonstrated that the APN was anchored in the membrane by a glycosylphosphatidylinositol moiety. Interaction of the expressed receptor with different Cry toxins was examined by immunofluorescence toxin binding studies and ligand blot and immunoprecipitation analyses. By these experiments we showed that the bioactive toxin, Cry1C, binds to the recombinant APN, while the nonbioactive toxin, Cry1Ac, showed no interaction.