bluuebunny/crossref_metadata_2025_split huggingface dataset:
Transcription from Fusion Promoters Generated during Transposition of Transposon Tn
4652
Is Positively Affected by Integration Host Factor in
Pseudomonas putida
Authors: Teras Riho , Hõrak Rita , Kivisaar Maia 
Published: July 2002

ABSTRACT

We have previously shown that both ends of the Tn
3
family transposon Tn
4652
contain integration host factor (IHF) binding sites and that IHF positively regulates expression of the Tn
4652
transposase gene
tnpA
in
Pseudomonas putida
(R. Hõrak, and M. Kivisaar, J. Bacteriol. 180:2822–2829, 1998). Tn
4652
can activate silent genes by creating fusion promoters during the transposition. The promoters are created as fusions between the −35 hexamer provided by the terminal inverted repeats of Tn
4652
and the −10 hexamers in the target DNA. Two fusion promoters, PRA1 and PLA1, that contain sequences of the right and left termini of Tn
4652
, respectively, were chosen for the study of mechanisms of transcription activation. Gel mobility shift analysis using crude extracts from
P. putida
cells allowed us to detect specific binding of
P. putida
IHF to the ends of the transposon Tn
4652
. We found that the rate of transcription from the fusion promoter PRA1 is enhanced by IHF. Notably, the positive effect of IHF on transcription from the promoter PRA1 appeared only when cells of
P. putida
reached the stationary growth phase. We speculate that the intracellular concentration of IHF might be critical for the in vivo effect of IHF on transcription from the fusion promoters in
P. putida
. In the case of PLA1, the mechanism of transcription modulation by IHF is different than that observed for PRA1. Our results demonstrate that transcription of neighboring genes from outwardly directed promoters at the ends of a mobile DNA element could be influenced by the same factors that control transposition of the element.
Expression of a New Operon from
Bacillus subtilis
,
ykzB-ykoL
, under the Control of the TnrA and PhoP-PhoR Global Regulators
Authors: Robichon Denis , Arnaud Maryvonne , Gardan Rozenn , Pragai Zoltan , O'Reilly Mary , Rapoport Georges , Débarbouillé Michel 
Published: July 2002

ABSTRACT

The
ykzB
and
ykoL
genes encode two peptides, of 51 and 60 amino acids, the functions of which are unknown. The
ykzB
and
tnrA
genes are contiguous and transcribed divergently. Expression of
ykzB
and
ykoL
is induced by glutamate and is under the control of the TnrA global regulator of nitrogen utilization. TnrA regulated its own synthesis in glutamate minimal medium. Two DNA sequences (TnrAB1 and TnrAB2) homologous to the TnrA binding site are present in the region between
tnrA
and
ykzB
. Deletion mapping indicated that the TnrAB2 binding site was involved in activation of the
ykzB
promoter. In addition, transcription of
tnrA
depends on the presence of the TnrAB1 binding site. The
ykzB
and
ykoL
genes are probably in the same transcriptional unit. A single promoter involved in transcription in the presence of glutamate was mapped by primer extension.
ykoL
expression was induced by phosphate limitation and depended on the PhoP-PhoR two-component regulatory system. Its promoter was mapped to the region between
ykoL
and
ykzB
. Four boxes similar to the PhoP binding site are present upstream from the
ykoL
promoter. These boxes are probably recognized by PhoP∼P during the activation of transcription in phosphate limitation conditions.
Differential Distribution of Novel Restriction-Modification Systems in Clonal Lineages of
Neisseria meningitidis
Authors: Claus Heike , Friedrich Alexander , Frosch Matthias , Vogel Ulrich 
Published: July 2002

ABSTRACT

Using representational difference analysis, we isolated novel meningococcal restriction-modification (R-M) systems.
Nme
BI, which is a homologue of the R-M system
Hga
I of
Pasteurella volantium
, was present in meningococci of the ET-5 complex and of lineage III.
Nme
AI was found in serogroup A, ET-37 complex, and cluster A4 meningococci.
Nme
DI was harbored by meningococci of the ET-37 complex and of cluster A4, but not by serogroup A meningococci. Two of the R-M systems,
Nme
BI and
Nme
DI, were located at homologous positions between the phenylalanyl-tRNA synthetase genes
pheS
and
pheT
, which appeared to be a preferential target for the insertion of foreign DNA in meningococci. The distribution of the three R-M systems was tested with 103 meningococcal strains comprising 49 sequence types. The vast majority of the strains had either
Nme
BI,
Nme
AI, or both
Nme
AI and
Nme
DI. Using cocultivation experiments, we could demonstrate that
Nme
BI, which was present in ET-5 complex meningococci, was responsible for a partial restriction of DNA transfer from meningococci of the ET-37 complex to meningococci of the ET-5 complex.
Identification of a
UPC2
Homolog in
Saccharomyces cerevisiae
and Its Involvement in Aerobic Sterol Uptake
Authors: Shianna Kevin V. , Dotson W. David , Tove Shirley , Parks Leo W. 
Published: July 2002

ABSTRACT

Saccharomyces cerevisiae
normally will not take up sterols from the environment under aerobic conditions. A specific mutant,
upc2
-
1
, of the predicted transcriptional activator
UPC2
(YDR213w) has been recognized as a strain that allows a high level of aerobic sterol uptake. Another predicted transcriptional activator, the YLR228c gene product, is highly homologous to Upc2p. In fact, at the carboxy terminus 130 of the last 139 amino acids are similar between the two proteins. Since these proteins are very similar, the effect of mutations in the YLR228c open reading frame (ORF) was compared with like alterations in
UPC2
. First, the YLR228c ORF was insertionally inactivated and crossed with various
UPC2
constructs. Deletion of YLR228c and
UPC2
in combination resulted in nonviability, suggesting that the two proteins have some essential overlapping function. The
upc2
-
1
point mutation responsible for aerobic sterol uptake was duplicated in the homologous carboxy region of the YLR228c ORF using site-directed mutagenesis. This mutation on a high-copy vector resulted in an increase in sterol uptake compared to an isogenic wild-type strain. The combination of both point mutations resulted in the greatest level of aerobic sterol uptake. When the YLR228c point mutation was expressed from a low-copy vector there was little if any effect on sterol uptake. Gas chromatographic analysis of the nonsaponifiable fractions of the various strains showed that the major sterol for all YLR228c and
UPC2
combinations was ergosterol, the consensus yeast sterol.
Bacillus anthracis
pXO1 Plasmid Sequence Conservation among Closely Related Bacterial Species
Authors: Pannucci James , Okinaka Richard T. , Sabin Robert , Kuske Cheryl R. 
Published: July 2002

ABSTRACT

The complete sequencing and annotation of the 181.7-kb
Bacillus anthracis
virulence plasmid pXO1 predicted 143 genes but could only assign putative functions to 45. Hybridization assays, PCR amplification, and DNA sequencing were used to determine whether pXO1 open reading frame (ORF) sequences were present in other bacilli and more distantly related bacterial genera. Eighteen
Bacillus
species isolates and four other bacterial species were tested for the presence of 106 pXO1 ORFs. Three ORFs were conserved in most of the bacteria tested. Many of the pXO1 ORFs were detected in closely related
Bacillus
species, and some were detected only in
B. anthracis
isolates. Three isolates,
Bacillus cereus
D-17,
B. cereus
43881, and
Bacillus thuringiensis
33679, contained sequences that were similar to more than one-half of the pXO1 ORF sequences examined. The majority of the DNA fragments that were amplified by PCR from these organisms had DNA sequences between 80 and 98% similar to that of pXO1. Pulsed-field gel electrophoresis revealed large potential plasmids present in both
B. cereus
43881 (341 kb) and
B. thuringiensis
ATCC 33679 (327 kb) that hybridized with a DNA probe composed of six pXO1 ORFs.
Anaerobic Respiration Using Fe
3+
, S
0
, and H
2
in the Chemolithoautotrophic Bacterium
Acidithiobacillus ferrooxidans
Authors: Ohmura Naoya , Sasaki Kazuhiro , Matsumoto Norio , Saiki Hiroshi 
Published: July 2002

ABSTRACT

The chemolithoautotrophic bacterium
Acidithiobacillus ferrooxidans
has been known as an aerobe that respires on iron and sulfur. Here we show that the bacterium could chemolithoautotrophically grow not only on H
2
/O
2
under aerobic conditions but also on H
2
/Fe
3+
, H
2
/S
0
, or S
0
/Fe
3+
under anaerobic conditions. Anaerobic respiration using Fe
3+
or S
0
as an electron acceptor and H
2
or S
0
as an electron donor serves as a primary energy source of the bacterium. Anaerobic respiration based on reduction of Fe
3+
induced the bacterium to synthesize significant amounts of a
c-
type cytochrome that was purified as an acid-stable and soluble 28-kDa monomer. The purified cytochrome in the oxidized form was reduced in the presence of the crude extract, and the reduced cytochrome was reoxidized by Fe
3+
. Respiration based on reduction of Fe
3+
coupled to oxidation of a
c-
type cytochrome may be involved in the primary mechanism of energy production in the bacterium on anaerobic iron respiration.
Distinct Roles of P
II
-Like Signal Transmitter Proteins and
amtB
in Regulation of
nif
Gene Expression, Nitrogenase Activity, and Posttranslational Modification of NifH in
Azoarcus
sp. Strain BH72
Authors: Martin Dietmar E. , Reinhold-Hurek Barbara 
Published: July 2002

ABSTRACT

P
II
-like signal transmitter proteins, found in
Bacteria
,
Archaea
, and plants, are known to mediate control of carbon and nitrogen assimilation. They indirectly regulate the activity of key metabolic enzymes and transcription factors by protein-protein interactions with signal transduction proteins. Many
Proteobacteria
harbor two paralogous P
II
-like proteins, GlnB and GlnK, whereas a novel third P
II
paralogue (GlnY) was recently identified in
Azoarcus
sp. strain BH72, a diazotrophic endophyte of grasses. In the present study, evidence was obtained that the P
II
-like proteins have distinct roles in mediating nitrogen and oxygen control of
nif
gene transcription and nitrogenase activity. Full repression of
nif
gene transcription in the presence of a combined nitrogen source or high oxygen concentrations was observed in wild-type and
glnB
and
glnK
knockout mutants, revealing that GlnB and GlnK can complement each other in mediating the repression. In contrast, in a
glnBK
double mutant strain in the presence of only GlnY,
nif
gene transcription was still detectable, albeit at a lower level, on nitrate or 20% oxygen. As another level of control, nitrogenase activity was regulated by at least three types of mechanisms in strain BH72: covalent modification of dinitrogenase reductase (NifH), probably by ADP-ribosylation, and two other, unknown means. Functional inactivation upon ammonium addition (switch-off) required the putative high-affinity ammonium transporter AmtB and GlnK, but not GlnB or GlnY. Functional inactivation in response to anaerobiosis did not depend on AmtB, GlnK, or GlnB. In contrast, covalent modification of NifH required both GlnB and GlnK and AmtB as response to ammonium addition, whereas it required either GlnB or GlnK and not AmtB when cells were shifted to anaerobiosis. In a
glnBK
double mutant expressing only GlnY, NifH modification was completely abolished, further revealing functional differences between the three P
II
paralogues.
Transmission Dynamics and Molecular Characterization of
Mycobacterium tuberculosis
Isolates with Low Copy Numbers of IS
6110
Authors: Soini Hanna , Pan Xi , Teeter Larry , Musser James M. , Graviss Edward A. 
Published: July 2002

ABSTRACT

Population-based analysis of
Mycobacterium tuberculosis
transmission in Houston, Tex., over 5 years identified 377 patients infected with an isolate containing one to four copies of IS
6110
. The isolates were analyzed by spoligotyping and assigned to one of three major genetic groups based on nucleotide polymorphisms in codons
katG
463 and
gyrA
95. Prospectively obtained patient interviews were reviewed to assess epidemiologic links between apparently clustered patients. A total of 13 groups of isolates with the same IS
6110
profile were identified, representing 326 of the 377 patients (86.5%; range 2 to 113 patients). In contrast, 28 groups of isolates containing 334 patients (88.6%) had the same spoligotype (range, 2 to 143 patients). Combination of IS
6110
profile and spoligotype data identified 31 clusters with 300 patients (79.6%; range, 2 to 82 patients). All 377 isolates belonged to major genetic group 1 (77 patients) or genetic group 2 (300 patients); no major genetic group 3 isolates were identified. Among the 228 patients interviewed, 33 patients (14.5%) were directly linked to another patient in the same cluster. Possible epidemiologic links were also found among 11 patients. Moreover, many clusters consisted of individuals with the same ethnicity. In conclusion, we confirmed that IS
6110
profiling and spoligotyping together provide enhanced molecular discrimination of
M. tuberculosis
isolates with low copy numbers of IS
6110
. Identification of epidemiologic links among some of the patients verified that the combination of these two methods reliably indexes tuberculosis transmission.
Rapid Identification of Virulent Type I Strains of the Protozoan Pathogen
Toxoplasma gondii
by PCR-Restriction Fragment Length Polymorphism Analysis at the
B1
Gene
Authors: Grigg Michael E. , Boothroyd John C. 
Published: July 2002

ABSTRACT

Sequence analysis at the 35-fold-repetitive
B1
locus identified three restriction sites capable of discriminating type I (mouse-virulent) from type II or III (mouse-avirulent) strains of
Toxoplasma gondii. B1
PCR-restriction fragment length polymorphism analysis of 8 type I, 17 type II, and 8 type III strains confirms the specificity of the assay. It should now be possible to ask whether strain genotype affects the severity and type of clinical disease in humans.
Quantification of Proviral Load of Human Immunodeficiency Virus Type 2 Subtypes A and B Using Real-Time PCR
Authors: Damond Florence , Descamps Diane , Farfara Isabelle , Telles Jean Noël , Puyeo Sophie , Campa Pauline , Leprêtre Annie , Matheron Sophie , Brun-Vezinet Françoise , Simon François 
Published: July 2002

ABSTRACT

We have developed and evaluated a new method to quantify human immunodeficiency virus type 2 (HIV-2) proviral DNA based on LightCycler real-time PCR. The assay has a detection limit of 5 copies/10
5
peripheral blood mononuclear cells (PBMC) and is insensitive to HIV-2 strain variability: HIV-2 subtypes A and B are both recognized and quantified. The intra- and interassay coefficients of variation range from 16 to 40% for high provirus concentrations (5 × 10
5
copies) and from 41 to 39% for low concentrations (5 copies). We used this method to compare the proviral DNA load and viral RNA load in plasma with clinical and immunological status for 29 patients infected by HIV-2 (subtype A in 17 and subtype B in 12). The proviral load (median, 201 copies/10
5
PBMC) was similar to that reported for HIV-1 infection. The median proviral loads did not correlate with the CD4
+
cell count categories and were as follows for CD4
+
cell counts of >400, 200 to 400, and <200 cells/mm
3
, respectively: 121 copies/10
5
PBMC (
n
= 8; range, <5 to 712 copies/10
5
PBMC); 114 copies/10
5
PBMC (
n
= 9; range, <5 to 1,907 copies/10
5
PBMC); and 285 copies/10
5
PBMC (
n
= 12; range, 53 to 2,524 copies/10
5
PBMC). Proviral load did not correlate with plasma HIV-2 RNA positivity. As HIV-2 is considered to replicate less efficiently than HIV-1, these high proviral loads might be explained by the proliferation of infected cells.
Viral Load of Human Herpesvirus 8 in Peripheral Blood of Human Immunodeficiency Virus-Infected Patients with Kaposi's Sarcoma
Authors: Tedeschi Rosamaria , Enbom Malin , Bidoli Ettore , Linde Annika , De Paoli Paolo , Dillner Joakim 
Published: July 2002

ABSTRACT

Viral load is an important marker of activity of viral diseases for a number of viruses. We wished to evaluate whether the viral load of human herpesvirus 8 (HHV-8) in peripheral blood was a consistent feature of Kaposi's sarcoma (KS) patients and whether the viral load correlated with human immunodeficiency virus (HIV) RNA levels, CD4 counts, and/or the HHV-8 seroreactivity. Fifty-four consecutive plasma samples from 14 patients with KS were evaluated for HHV-8 viral load by quantitative real-time PCR. Samples were analyzed at the start of highly active antiretroviral therapy (HAART) and at different intervals during treatments. The median HHV-8 DNA load before HAART treatment was 8,998 (ranging from 170 to 40,100) copies/ml and 12,270 (ranging from 40 to 142,575) copies/ml during HAART. There were both increasing and decreasing trends. There was an association between HHV-8 DNA and HIV RNA viral loads (odds ratio [OR] = 5.40; 95% confidence interval [95% CI], 1.54 to 18.98) and between HHV-8 viral load and CD4 cell counts (OR = 7.24; 95% CI, 1.30 to 40.35). High HHV-8 viral load was also correlated with the titers of antibodies to the lytic HHV-8 antigen detected with immunofluorescence (
P
< 0.01), but not with antibodies to the latent HHV-8 antigen. In conclusion, we found that HHV-8 viremia in KS is associated with HIV viral load, CD4 cell counts, and lytic HHV-8 serological reactivity. HHV-8 viral load monitored by real time PCR might be useful for determination HHV-8 viral load during the follow-up of KS patients.
Molecular Investigation of the Aum Shinrikyo Anthrax Release in Kameido, Japan
Authors: Keim Paul , Smith Kimothy L. , Keys Christine , Takahashi Hiroshi , Kurata Takeshi , Kaufmann Arnold 
Published: July 2002

ABSTRACT

In 1993, the Aum Shinrikyo cult aerosolized
Bacillus anthracis
spores over Kameido, Japan. Spore samples were obtained from the release site, cultured, and characterized by molecular genetic typing. The isolates were consistent with strain Sterne 34F2, which is used in Japan for animal prophylaxis against anthrax.
Phenotypic and Genotypic Approaches to Characterization of Isolates of
Neisseria meningitidis
from Patients and Their Close Family Contacts
Authors: Tzanakaki G. , Urwin R. , Musilek M. , Kriz P. , Kremastinou J. , Pangalis A. , Blackwell C. C. , Maiden M. C. J. 
Published: July 2002

ABSTRACT

Characterization of isolates of
Neisseria meningitidis
obtained from patients with meningococcal disease or from pharyngeal swabs of asymptomatic carriers can be achieved by several approaches which provide different levels of discrimination. A total of 45 gram negative, oxidase-positive diplococcus strains isolated from 15 individuals with meningococcal disease and 30 of their family contacts were examined by three approaches: serological typing, multilocus enzyme electrophoresis (MLEE), and multilocus sequence typing (MLST). For 10 of the 15 patient and contact groups, all of the isolates were confirmed as meningococci, and the bacteria obtained from the patients and contacts, including their mother or principal caregiver in the case of children, were indistinguishable by all three methods. In the remaining five groups the isolates from the patients were distinct from those recovered from the contacts, and in three examples, in two separate groups, the contacts were shown by MLST to be carrying strains of
Neisseria lactamica
. The data obtained from the three techniques were consistent, although complete serological typing was possible for only a minority of isolates. Both MLEE and MLST established the genetic relationships of the isolates and identified members of known hypervirulent lineages, but MLST was faster than MLEE and had the additional advantages that it could be performed on noninfective material distributed by mail and that the results from different laboratories could be compared via the internet (
http://mlst.zoo.ox.ac.uk
).
Effects of Storage and Type of Blood Collection Tubes on Hepatitis C Virus Level in Whole Blood Samples
Authors: Kessler Harald H. , Stelzl Evelyn , Raggam Reinhard B. , Haas Josef , Kirchmeir Franz , Hegenbarth Karin , Daghofer Elisabeth , Santner Brigitte I. , Marth Egon , Stauber Rudolf E. 
Published: July 2002

ABSTRACT
In this study, we compared serum hepatitis C virus (HCV) RNA concentrations with HCV RNA concentrations in whole blood collection tubes, including two different types of EDTA tubes and nucleic acid stabilization tubes (NASTs). We also investigated the impact of a processing delay on HCV RNA concentration in these tubes. In NASTs, the mean HCV RNA concentration was comparable to the mean serum HCV RNA concentration at “date zero.” In EDTA tubes, mean baseline HCV RNA concentrations were higher. Storage at room temperature up to 96 h did not result in a decline of HCV RNA concentration in any of the whole blood collection tubes. In NASTs, HCV RNA concentrations remained stable during the whole study period, whereas a significant increase of HCV RNA was observed in both types of EDTA tubes at 96 h compared to date zero. We concluded that HCV RNA remains stable in NASTs at room temperature for at least 96 h, allowing greater flexibility in sample collection and transport.
Characterization and Analysis of a Stable Serotype-Associated Membrane Protein (P30) of
Mycoplasma agalactiae
Authors: Fleury Bénédicte , Bergonier Dominique , Berthelot Xavier , Schlatter Yvonne , Frey Joachim , Vilei Edy M. 
Published: July 2002

ABSTRACT

The gene for a 30-kDa immunodominant antigen, P30, of
Mycoplasma agalactiae
was cloned from type strain PG2 and expressed in
Escherichia coli
. P30 is encoded on a monocistronic operon determined by two −10 boxes and a possible −35 region constituting the potential promoter, and a transcription termination site. The gene for the 266-amino-acid protein is preceded by a polypurine-rich region designed as the consensus sequence for a ribosome-binding site. Analysis of the amino acid sequence of P30 revealed the presence of a recognition site for a prokaryotic signal peptidase II at amino acid (aa) 24, indicating that P30 is a transmembrane protein. Moreover, Triton X-114 phase partitioning of
M. agalactiae
PG2 total antigen revealed that P30 is strongly hydrophobic and hence a possible membrane component. Immunoblot analysis using the monospecific polyclonal anti-P30-His serum indicated that P30 is specific to
M. agalactiae
. Furthermore, PCR amplification with specific primers for
p30
and Southern blot analysis revealed the presence of the gene in all
M. agalactiae
strains tested and its absence in the other mycoplasma species. Among 27 strains of
M. agalactiae
studied
,
20 strains belonging to the common serotypes A to D, including PG2, expressed P30 or part of it as detected by the monospecific polyclonal anti-P30 antibodies. The other seven strains belonging to the rarely isolated serotypes E to H were negative for P30. The
p30
gene was sequenced in 15 strains of
M. agalactiae
, 10 of which expressed P30 or at least part of it and 5 of which did not express P30. The negative strains carried mutations in both −10 boxes of the promoters. These mutations seem to be responsible for the lack of P30 expression in these strains. Analysis of sera from sheep that were experimentally infected with
M. agalactiae
revealed that P30 induced a strong and persistent immune response which was still very high two months after infection. In contrast, currently used enzyme-linked immunosorbent assay serology gave only low titers.
Atypical,
fljB
-Negative
Salmonella enterica
subsp.
enterica
Strain of Serovar 4,5,12:i:− Appears To Be a Monophasic Variant of Serovar Typhimurium
Authors: Echeita M. Aurora , Herrera Silvia , Usera Miguel A. 
Published: July 2002

ABSTRACT

An
fljB
-negative, multidrug-resistant
Salmonella enterica
serovar 4,5,12:i:− phage type DT U302 strain (resistant to ampicillin, chloramphenicol, sulfonamide, gentamicin, streptomycin, tetracycline, and sulfamethoxazole-trimethoprim) emerged and spread in Spain in 1997. Sequences specific for
Salmonella
serovar Typhimurium and phage type DT 104 and U302 were present in this atypical
Salmonella
strain, suggesting that it is a monophasic
Salmonella
serovar Typhimurium variant.