bluuebunny/crossref_metadata_2025_split huggingface dataset:
Relationship between Glycolysis and Exopolysaccharide Biosynthesis in
Lactococcus lactis
Authors: Ramos Ana , Boels Ingeborg C. , de Vos Willem M. , Santos Helena 
Published: July 2002

ABSTRACT

The relationships between glucose metabolism and exopolysaccharide (EPS) production in a
Lactococcus lactis
strain containing the EPS gene cluster (Eps
+
) and in nonproducer strain MG5267 (Eps
−
) were characterized. The concentrations of relevant phosphorylated intermediates in EPS and cell wall biosynthetic pathways or glycolysis were determined by
31
P nuclear magnetic resonance. The concentrations of two EPS precursors, UDP-glucose and UDP-galactose, were significantly lower in the Eps
+
strain than in the Eps
−
strain. The precursors of the peptidoglycan pathway, UDP-
N
-acetylglucosamine and UDP-
N
-acetylmuramoyl-pentapeptide, were the major UDP-sugar derivatives detected in the two strains examined, but the concentration of the latter was greater in the Eps
+
strain, indicating that there is competition between EPS synthesis and cell growth. An intermediate in biosynthesis of histidine and nucleotides, 5-phosphorylribose 1-pyrophosphate, accumulated at concentrations in the millimolar range, showing that the pentose phosphate pathway was operating. Fructose 1,6-bisphosphate and glucose 6-phosphate were the prominent glycolytic intermediates during exponential growth of both strains, whereas in the stationary phase the main metabolites were 3-phosphoglyceric acid, 2-phosphoglyceric acid, and phosphoenolpyruvate. The activities of relevant enzymes, such as phosphoglucose isomerase, α-phosphoglucomutase, and UDP-glucose pyrophosphorylase, were identical in the two strains.
13
C enrichment on the sugar moieties of pure EPS showed that glucose 6-phosphate is the key metabolite at the branch point between glycolysis and EPS biosynthesis and ruled out involvement of the triose phosphate pool. This study provided clues for ways to enhance EPS production by genetic manipulation.
Mutations in the Listerial
proB
Gene Leading to Proline Overproduction: Effects on Salt Tolerance and Murine Infection
Authors: Sleator Roy D. , Gahan Cormac G. M. , Hill Colin 
Published: July 2002

ABSTRACT

The observed sensitivity of
Listeria monocytogenes
to the toxic proline analogue
l
-azetidine-2-carboxylic acid (AZ) suggested that proline synthesis in
Listeria
may be regulated by feedback inhibition of γ-glutamyl kinase (GK), the first enzyme of the proline biosynthesis pathway, encoded by the
proB
gene. Taking advantage of the
Epicurian coli
mutator strain XL1-Red, we performed random mutagenesis of the recently described
proBA
operon and generated three independent mutations in the listerial
proB
homologue, leading to proline overproduction and salt tolerance when expressed in an
E. coli
(Δ
proBA
) background. While each of the mutations (located within a conserved 26-amino-acid region of GK) was shown to confer AZ resistance (AZ
r
) on an
L. monocytogenes proBA
mutant, listerial transformants failed to exhibit the salt-tolerant phenotype observed in
E. coli
. Since proline accumulation has previously been linked to the virulence potential of a number of pathogenic bacteria, we analyzed the effect of proline overproduction on
Listeria
pathogenesis. However, our results suggest that as previously described for proline auxotrophy, proline hyperproduction has no apparent impact on the virulence potential of
Listeria
.
Electropositive Filter Membrane as an Alternative for the Elimination of PCR Inhibitors from Sewage and Water Samples
Authors: Queiroz A. P. S. , Santos F. M. , Sassaroli A. , Hársi C. M. , Monezi T. A. , Mehnert D. U. 
Published: July 2002

ABSTRACT
PCR is the best method for the detection of enteric viruses present at low concentrations in environmental samples. However, some organic and inorganic compounds present in these samples can interfere in the reaction. Many of these substances are cytotoxic, too. The ZP60S filter membranes used in addition to fluorpentane treatment are quite efficient for virus concentration and simultaneous elimination of cytotoxicity from environmental samples. In this study, both procedures were used to promote the elimination of reverse transcriptase PCR (RT-PCR) inhibitors from sewage and sewage-polluted creek water. Samples were subjected separately to each of the following procedures: filtration through electropositive filter membranes (ZP60S), organic extraction with Vertrel XF, and filtration through ZP60S followed by organic extraction. Afterwards, aliquots were experimentally inoculated with rotavirus SA-11 RNA and subjected to RT-seminested PCR for amplification of the VP7 gene. Results showed that the ZP60S membranes efficiently eliminated the RT-PCR inhibitors from water samples. The sample processing method was also applied to 31 in natura sewage and creek water samples for detection of naturally occurring rotavirus. A duplex seminested PCR was used for the quick detection of couples of the four rotavirus genotypes (G1 to G4). Eight samples (25.8%) were positive, and rotavirus sequences were not detected in 23 (74.2%). Results were confirmed by direct immunoperoxidase method. In summary, the use of electropositive filter membrane is appropriate for the elimination of substances that can interfere with RT-PCR, obviating additional sample purification methods.
Availability of Glutamate and Arginine during Acid Challenge Determines Cell Density-Dependent Survival Phenotype of
Escherichia coli
Strains
Authors: Cui Shenghui , Meng Jianghong , Bhagwat Arvind A. 
Published: July 2002

ABSTRACT

The cell density-dependent acid sensitivity phenotypes of
Escherichia coli
strains K-12 and O157:H7 were examined with reference to three possible mechanisms of acid resistance. There was no evidence of any diffusible substance released from dead cells which could influence the cell density-dependent acid survival phenotype. Instead, cell density-dependent acid survival phenotype was associated with induction of glutamate- and arginine-decarboxylase acid survival pathways and concomitant availability of glutamate and arginine during acid challenge.
Lyme Disease
Borrelia
spp. in Ticks and Rodents from Northwestern China
Authors: Takada Nobuhiro , Masuzawa Toshiyuki , Ishiguro Fubito , Fujita Hiromi , Kudeken Midori , Mitani Harumi , Fukunaga Masahito , Tsuchiya Kimiyuki , Yano Yasuhiro , Ma Xiao-Hang 
Published: July 2002

ABSTRACT

In May 1999, field surveys of Lyme disease spirochetes were conducted around the Tianshan Mountains in Xinjiang Uygur Autonomous Region in northwestern People's Republic of China.
Ixodes persulcatus
ticks were obtained in a Tianchi Lake valley with primary forest, while the tick fauna was poor in the semidesert or at higher altitudes in this region. Species identities were confirmed by molecular analysis in which an internal transcribed spacer sequence was used. Of 55 adult ticks, 22 (40%) were positive for spirochetes as determined by Barbour-Stoenner-Kelly culture passages. In addition, some rodents, including
Apodemus uralensis
(5 of 14 animals) and
Cricetulus longicaudatus
(the only animal examined), and some immature stages of
I. persulcatus
(4 of 11 ticks) that had fed on
A. uralensis
were positive for spirochetes. Based on 5S-23S rRNA intergenic spacer restriction fragment length polymorphism analysis and reactivity with monoclonal antibodies, 35 cultures (including double isolation cultures) were identified as
Borrelia garinii
(20 isolates, including 9 Eurasian pattern B isolates and 11 Asian pattern C isolates),
Borrelia afzelii
(10 pattern D isolates), and mixed cultures (5 cultures, including isolates that produced
B. garinii
patterns B and C plus
B. afzelii
pattern D). These findings revealed that Lyme disease pathogens are distributed in the mountainous areas in northwestern China even though it is an arid region, and they also confirmed the specific relationship between
I. persulcatus
and genetic patterns of
Borrelia
spp. on the Asian continent.
Gene Cassette PCR: Sequence-Independent Recovery of Entire Genes from Environmental DNA
Authors: Stokes H. W. , Holmes Andrew J. , Nield Blair S. , Holley Marita P. , Nevalainen K. M. Helena , Mabbutt Bridget C. , Gillings Michael R. 
Published: July 2002

ABSTRACT
The vast majority of bacteria in the environment have yet to be cultured. Consequently, a major proportion of both genetic diversity within known gene families and an unknown number of novel gene families reside in these uncultured organisms. Isolation of these genes is limited by lack of sequence information. Where such sequence data exist, PCR directed at conserved sequence motifs recovers only partial genes. Here we outline a strategy for recovering complete open reading frames from environmental DNA samples. PCR assays were designed to target the 59-base element family of recombination sites that flank gene cassettes associated with integrons. Using such assays, diverse gene cassettes could be amplified from the vast majority of environmental DNA samples tested. These gene cassettes contained complete open reading frames, the majority of which were associated with ribosome binding sites. Novel genes with clear homologies to phosphotransferase, DNA glycosylase, methyl transferase, and thiotransferase genes were identified. However, the majority of amplified gene cassettes contained open reading frames with no identifiable homologues in databases. Accumulation analysis of the gene cassettes amplified from soil samples showed no signs of saturation, and soil samples taken at 1-m intervals along transects demonstrated different amplification profiles. Taken together, the genetic novelty, steep accumulation curves, and spatial heterogeneity of genes recovered show that this method taps into a vast pool of unexploited genetic diversity. The success of this approach indicates that mobile gene cassettes and, by inference, integrons are widespread in natural environments and are likely to contribute significantly to bacterial diversity.
Detection of the
isiA
Gene across Cyanobacterial Strains: Potential for Probing Iron Deficiency
Authors: Geiß U. , Vinnemeier J. , Kunert A. , Lindner I. , Gemmer B. , Lorenz M. , Hagemann M. , Schoor A. 
Published: July 2002

ABSTRACT

The use of
isiA
expression to monitor the iron status of cyanobacteria was investigated. Studies of laboratory cultures of the cyanobacterium
Synechocystis
sp. strain PCC 6803 showed that
isiA
expression is dependent on the organism's response to iron deficiency;
isiA
expression starts as soon as a decline in the rate of growth begins.
isiA
expression is switched on at concentrations of iron citrate of less than 0.7 μM. A PCR method was developed for the specific amplification of the iron-regulated
isiA
gene from a variety of cyanobacteria. After we developed degenerate primers, 15 new internal
isiA
fragments (840 bp) were amplified, cloned, and sequenced from strains obtained from algal collections, from new isolates, and from enriched field samples. Furthermore,
isiA
expression could be detected by means of reverse transcription-PCR when enriched field samples were exposed to restricted iron availability. These results imply that determining the level of iron-regulated
isiA
expression can serve to indicate iron deficiency in cyanobacterial samples of differing origins from the field.
Enterotoxigenic Potential ofStaphylococcus intermedius
Authors: Becker Karsten , Keller Birgit , von Eiff Christof , Brück Michaela , Lubritz Gabriele , Etienne Jerome , Peters Georg 
Published: July 2002

ABSTRACTStaphylococcal food poisoning (SFP) caused by enterotoxigenic staphylococci is one of the main food-borne diseases. In contrast toStaphylococcus aureus, a systematic screening for the enterotoxins has not yet been performed on the genomic level for the coagulase-positive speciesS.intermedius. Therefore, the enterotoxigenic potential of 281 different veterinary (canine,n= 247; equine,n= 23; feline,n= 9; other,n= 2) and 11 human isolates ofS.intermediuswas tested by using a multiplex PCR DNA-enzyme immunoassay system targeting the staphylococcal enterotoxin genessea,seb,sec,sed, andsee. Molecular results were compared by in vitro testing of enterotoxin production by two immunoassays. A total of 33 (11.3%)S.intermediusisolates, including 31 (12.6%) canine isolates, 1 equine isolate, and 1 human isolate, tested positive for thesecgene. In vitro production of the respective enterotoxins was detected in 30 (90.9%) of these isolates by using immunological tests. In contrast, none of 65 veterinary specimen-derived isolates additionally tested and comprising 13 (sub)species of coagulase-negative staphylococci were found to be enterotoxigenic. This study shows on both molecular and immunological levels that a substantial number ofS.intermediusisolates harbor the potential for enterotoxin production. Since evidence for noninvasive zoonotic transmission ofS.intermediusfrom animal hosts to humans has been documented, an enterotoxigenic role of this microorganism in SFP via contamination of food products may be assumed.
Role of β-Oxidation Enzymes in γ-Decalactone Production by the Yeast
Yarrowia lipolytica
Authors: Waché Yves , Aguedo Mario , Choquet Armelle , Gatfield Ian L. , Nicaud Jean-Marc , Belin Jean-Marc 
Published: July 2002

ABSTRACT

Some microorganisms can transform methyl ricinoleate into γ-decalactone, a valuable aroma compound, but yields of the bioconversion are low due to (i) incomplete conversion of ricinoleate (C
18
) to the C
10
precursor of γ-decalactone, (ii) accumulation of other lactones (3-hydroxy-γ-decalactone and 2- and 3-decen-4-olide), and (iii) γ-decalactone reconsumption. We evaluated acyl coenzyme A (acyl-CoA) oxidase activity (encoded by the
POX1
through
POX5
genes) in
Yarrowia lipolytica
in lactone accumulation and γ-decalactone reconsumption in
POX
mutants. Mutants with no acyl-CoA oxidase activity could not reconsume γ-decalactone, and mutants with a disruption of
pox3
, which encodes the short-chain acyl-CoA oxidase, reconsumed it more slowly. 3-Hydroxy-γ-decalactone accumulation during transformation of methyl ricinoleate suggests that, in wild-type strains, β-oxidation is controlled by 3-hydroxyacyl-CoA dehydrogenase. In mutants with low acyl-CoA oxidase activity, however, the acyl-CoA oxidase controls the β-oxidation flux. We also identified mutant strains that produced 26 times more γ-decalactone than the wild-type parents.
Application of Digital Image Analysis and Flow Cytometry To Enumerate Marine Viruses Stained with SYBR Gold
Authors: Chen Feng , Lu Jing-rang , Binder Brian J. , Liu Ying-chun , Hodson Robert E. 
Published: July 2002

ABSTRACT
A novel nucleic acid stain, SYBR Gold, was used to stain marine viral particles in various types of samples. Viral particles stained with SYBR Gold yielded bright and stable fluorescent signals that could be detected by a cooled charge-coupled device camera or by flow cytometry. The fluorescent signal strength of SYBR Gold-stained viruses was about twice that of SYBR Green I-stained viruses. Digital images of SYBR Gold-stained viral particles were processed to enumerate the concentration of viral particles by using digital image analysis software. Estimates of viral concentration based on digitized images were 1.3 times higher than those based on direct counting by epifluorescence microscopy. Direct epifluorescence counts of SYBR Gold-stained viral particles were in turn about 1.34 times higher than those estimated by the transmission electron microscope method. Bacteriophage lysates stained with SYBR Gold formed a distinct population in flow cytometric signatures. Flow cytometric analysis revealed at least four viral subpopulations for a Lake Erie sample and two subpopulations for a Georgia coastal sample. Flow cytometry-based viral counts for various types of samples averaged 1.1 times higher than direct epifluorescence microscopic counts. The potential application of digital image analysis and flow cytometry for rapid and accurate measurement of viral abundance in aquatic environments is discussed.
Molecular Characterization and Identification of
Bacillus clausii
Strains Marketed for Use in Oral Bacteriotherapy
Authors: Senesi Sonia , Celandroni Francesco , Tavanti Arianna , Ghelardi Emilia 
Published: July 2002

ABSTRACT

A substantial number of
Bacillus
species have been marketed for use in oral bacteriotherapy because of their purported ability to prevent or treat various gastrointestinal disorders. Recently, some of the
Bacillus
strains in Enterogermina, which is made up of aqueous suspensions of viable
Bacillus
spores, have been partially characterized and aligned with members of the
Bacillus alcalophilus
subgroup rather than with
Bacillus subtilis
, as previously reported. With a view toward verifying the original taxonomic position of the Enterogermina strains, we catalogued both phenotypic and genotypic traits exhibited by the four
Bacillus
strains isolated from the spore mixtures found in original commercial preparations dated 1975 and 1984 and commercial preparations now being propagated industrially. Analyses of physiological and biochemical traits, complete 16S rRNA gene sequences, DNA-DNA reassociation, tRNA intergenic spacer length polymorphism, single-strand conformation polymorphism of PCR-amplified spacer regions of tRNA genes, and randomly amplified polymorphic DNA led to the finding that all of the Enterogermina strains belong to a unique genospecies, which is unequivocally identified as the alkalitolerant species
Bacillus clausii
. Moreover, we provide evidence that in contrast to several reference strains of
B. clausii
, the strains constituting Enterogermina are characterized by a notable low level of intraspecific genome diversity and that each strain has remained the same for the last 25 years.
Congruent Phylogenies of Most Common Small-Subunit rRNA and Dissimilatory Sulfite Reductase Gene Sequences Retrieved from Estuarine Sediments
Authors: Joulian Catherine , Ramsing Niels B. , Ingvorsen Kjeld 
Published: July 2002

ABSTRACT

The diversity of sulfate-reducing bacteria (SRB) in brackish sediment was investigated using small-subunit rRNA and dissimilatory sulfite reductase (DSR) gene clone libraries and cultivation. The phylogenetic affiliation of the most commonly retrieved clones for both genes was strikingly similar and produced
Desulfosarcina variabilis
-like sequences from the inoculum but
Desulfomicrobium baculatum
-like sequences from a high dilution in natural media. Related organisms were subsequently cultivated from the site. PCR bias appear to be limited (or very similar) for the two primersets and target genes. However, the DSR primers showed a much higher phylogenetic specificity. DSR gene analysis is thus a promising and specific approach for investigating SRB diversity in complex habitats.
Three Glycoproteins with Antimutagenic Activity Identified in
Lactobacillus plantarum
KLAB21
Authors: Rhee Chang-Ho , Park Heui-Dong 
Published: July 2002

ABSTRACT

Antimutagenic substances were purified from a culture supernatant of
Lactobacillus plantarum
KLAB21 cells isolated from kimchi, a Korean traditional fermented vegetable, and their characteristics were investigated. The antimutagenic substances were separated into two fractions by DEAE-cellulose ion-exchange column chromatography, which were designated the R1 and R2 fractions. The R1 fraction was then divided into two fractions again by Sephadex G200 gel filtration chromatography, and the fractions were designated R1-1 and R1-2. All three fractions were further purified using a Sepharose CL-6B gel filtration column. All the purified fractions were successfully stained with fuchsin as well as Coomassie brilliant blue, suggesting that they are glycoproteins. The purified fractions were confirmed to possess antimutagenic activity against
N
-methyl-
N
′-nitro-
N
-nitrosoguanidine on
Salmonella enterica
serovar Typhimurium TA100 cells. Their molecular masses were determined to be 16 (R1-1), 11 (R1-2), and 14 (R2) kDa on the Sepharose CL-6B column. Total sugar contents were 8.4% (R1-1), 7.3% (R1-2), and 9.4% (R2). The amino acid compositions of the fractions were different from each other; the major amino acids were glutamic acid (21.5%) and phenylalanine (17.1%) in the R1-1 fraction and glycine (41.3%) in the R1-2 fraction, but valine (31%) and phenylalanine (22.6%) were the major amino acids in the R2 fraction.
Quantitative Analysis of the Relative Transcript Levels of ABC Transporter
Atr
Genes in
Aspergillus nidulans
by Real-Time Reverse Transcription-PCR Assay
Authors: Semighini Camile Pizeta , Marins Mozart , Goldman Maria Helena S. , Goldman Gustavo Henrique 
Published: July 2002

ABSTRACT

The development of assays for quantitative analysis of the relative transcript levels of ABC transporter genes by real-time reverse transcription-PCR (RT-PCR) might provide important information about multidrug resistance in filamentous fungi. Here, we evaluate the potential of real-time RT-PCR to quantify the relative transcript levels of ABC transporter
Atr
genes from
Aspergillus nidulans
. The
AtrA
to
AtrD
genes showed different and higher levels in the presence of structurally unrelated drugs, such as camptothecin, imazalil, itraconazole, hygromycin, and 4-nitroquinoline oxide. We also verified the relative transcript levels of the
Atr
genes in the
A. nidulans
imazalil-resistant mutants. These genes displayed a very complex pattern in different
ima
genetic backgrounds. The
imaB
mutant has higher basal transcript levels of
AtrB
and -
D
than those of the wild-type strain. The levels of these two genes are comparable when the
imaB
mutant is grown in the presence and absence of imazalil. The
imaC
, -
D
, and -
H
mutants have higher basal levels of
AtrA
than that of the wild type. The same behavior is observed for the relative transcript levels of
AtrB
in the
imaG
mutant background.
Clonal Analysis of the Human B-Cell Repertoire Using a Heteromobility Assay
Authors: Tewari Deepanker 
Published: July 2002

ABSTRACTA heteromobility duplex tracking assay was developed to analyze B-cell clonality. The assay was based on the genetic variability of B-cell immunoglobulin (Ig) sequences. Binding of amplified (Ig) sequences to a single-stranded radiolabeled Ig DNA probe resulted in the formation of heteroduplexes. The mobilities of these heteroduplexes helped to distinguish clonal B cells.
Quantitative Fluorogenic PCR Assay for Measuring Ovine Herpesvirus 2 Replication in Sheep
Authors: Hüssy D. , Stäuber N. , Leutenegger C. M. , Rieder S. , Ackermann M. 
Published: July 2002

ABSTRACT
A fluorogenic PCR specific for ovine herpesvirus 2 (OvHV-2) DNA was developed and compared to a previously established conventional seminested PCR. Testing of a total of 152 blood samples from both positive and negative animals revealed that the results of both assays corresponded to each other in 100% of the cases. A second fluorogenic PCR for genomic sheep DNA was required to normalize the quantity of viral DNA in the sample. Separate standard curves had to be constructed for each PCR. The analytical sensitivity of the new PCRs ranged between at least 10 copies and sometimes even 1 copy of target DNA per reaction mixture. In dilution series of the target DNAs, linear decreases of the signals were observed over 7 orders of magnitude. Thus, it was possible to calculate the amounts of viral DNA in relation to the amounts of cellular DNA by normalizing the absolute quantity of OvHV-2 DNA with the amount of genomic sheep DNA. By this technique, it was possible for the first time to quantitatively characterize the course of OvHV-2 replication in naturally infected sheep.