bluuebunny/crossref_metadata_2025_split huggingface dataset:
Expression of the 1-Aminocyclopropane-1-Carboxylic Acid Deaminase Gene Requires Symbiotic Nitrogen-Fixing Regulator GenenifA2inMesorhizobium lotiMAFF303099
Authors: Nukui Noriyuki , Minamisawa Kiwamu , Ayabe Shin-Ichi , Aoki Toshio 
Published: July 2006

ABSTRACTMany soil bacteria contain 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase, which degrades ACC, a precursor of the phytohormone ethylene. In order to examine the regulation of theacdSgene encoding ACC deaminase inMesorhizobium lotiMAFF303099 during symbiosis with the host legumeLotus japonicus, we introduced the β-glucuronidase (GUS) gene intoacdSso that GUS was expressed under control of theacdSpromoter, and we also generated disruption mutants with mutations in a nitrogen fixation regulator gene,nifA. The histochemical GUS assay showed that there was exclusive expression ofacdSin mature root nodules. Two homologousnifAgenes, mll5857 and mll5837, were found in the symbiosis island ofM. lotiand were designatednifA1andnifA2, respectively. Quantitative reverse transcription-PCR demonstrated thatnifA2disruption resulted in considerably diminished expression ofacdS,nifH, andnifA1in bacteroid cells. In contrast,nifA1disruption slightly enhanced expression of theacdStranscripts and suppressednifHto some extent. These results indicate that theacdSgene and other symbiotic genes are positively regulated by the NifA2 protein, but not by the NifA1 protein, inM. loti. The mode of gene expression suggests thatM. loti acdSparticipates in the establishment and/or maintenance of mature nodules by interfering with the production of ethylene, which induces negative regulation of nodulation.
Molecular and Insecticidal Characterization of a Cry1I Protein Toxic to Insects of the Families Noctuidae, Tortricidae, Plutellidae, and Chrysomelidae
Authors: Ruiz de Escudero Iñigo , Estela Anna , Porcar Manuel , Martínez Clara , Oguiza José A. , Escriche Baltasar , Ferré Juan , Caballero Primitivo 
Published: July 2006

ABSTRACT

The most notable characteristic of
Bacillus thuringiensis
is its ability to produce insecticidal proteins. More than 300 different proteins have been described with specific activity against insect species. We report the molecular and insecticidal characterization of a novel
cry
gene encoding a protein of the Cry1I group with toxic activity towards insects of the families Noctuidae, Tortricidae, Plutellidae, and Chrysomelidae. PCR analysis detected a DNA sequence with an open reading frame of 2.2 kb which encodes a protein with a molecular mass of 80.9 kDa. Trypsin digestion of this protein resulted in a fragment of ca. 60 kDa, typical of activated Cry1 proteins. The deduced sequence of the protein has homologies of 96.1% with Cry1Ia1, 92.8% with Cry1Ib1, and 89.6% with Cry1Ic1. According to the Cry protein classification criteria, this protein was named Cry1Ia7. The expression of the gene in
Escherichia coli
resulted in a protein that was water soluble and toxic to several insect species. The 50% lethal concentrations for larvae of
Earias insulana
,
Lobesia botrana
,
Plutella xylostella
, and
Leptinotarsa decemlineata
were 21.1, 8.6, 12.3, and 10.0 μg/ml, respectively. Binding assays with biotinylated toxins to
E. insulana
and
L. botrana
midgut membrane vesicles revealed that Cry1Ia7 does not share binding sites with Cry1Ab or Cry1Ac proteins, which are commonly present in
B. thuringiensis
-treated crops and commercial
B. thuringiensis
-based bioinsecticides. We discuss the potential of Cry1Ia7 as an active ingredient which can be used in combination with Cry1Ab or Cry1Ac in pest control and the management of resistance to
B. thuringiensis
toxins.
Comparison of Virulence Gene Profiles of
Escherichia coli
Strains Isolated from Healthy and Diarrheic Swine
Authors: Chapman Toni A. , Wu Xi-Yang , Barchia Idris , Bettelheim Karl A. , Driesen Steven , Trott Darren , Wilson Mark , Chin James J.-C. 
Published: July 2006

ABSTRACT

A combination of uni- and multiplex PCR assays targeting 58 virulence genes (VGs) associated with
Escherichia coli
strains causing intestinal and extraintestinal disease in humans and other mammals was used to analyze the VG repertoire of 23 commensal
E. coli
isolates from healthy pigs and 52 clinical isolates associated with porcine neonatal diarrhea (ND) and postweaning diarrhea (PWD). The relationship between the presence and absence of VGs was interrogated using three statistical methods. According to the generalized linear model, 17 of 58 VGs were found to be significant (
P
< 0.05) in distinguishing between commensal and clinical isolates. Nine of the 17 genes represented by
iha
,
hlyA
,
aidA
,
east1
,
aah
,
fimH
,

iroN
E. coli

,
traT
, and
saa
have not been previously identified as important VGs in clinical porcine isolates in Australia. The remaining eight VGs code for fimbriae (F4, F5, F18, and F41) and toxins (STa, STb, LT, and Stx2), normally associated with porcine enterotoxigenic
E. coli
. Agglomerative hierarchical algorithm analysis grouped
E. coli
strains into subclusters based primarily on their serogroup. Multivariate analyses of clonal relationships based on the 17 VGs were collapsed into two-dimensional space by principal coordinate analysis. PWD clones were distributed in two quadrants, separated from ND and commensal clones, which tended to cluster within one quadrant. Clonal subclusters within quadrants were highly correlated with serogroups. These methods of analysis provide different perspectives in our attempts to understand how commensal and clinical porcine enterotoxigenic
E. coli
strains have evolved and are engaged in the dynamic process of losing or acquiring VGs within the pig population.
Identification of an Abscisic Acid Gene Cluster in the Grey Mold
Botrytis cinerea
Authors: Siewers Verena , Kokkelink Leonie , Smedsgaard Jørn , Tudzynski Paul 
Published: July 2006

ABSTRACT

Like several other phytopathogenic fungi, the ascomycete
Botrytis cinerea
is known to produce the plant hormone abscisic acid (ABA) in axenic culture. Recently,
bcaba1
, the first fungal gene involved in ABA biosynthesis, was identified. Neighborhood analysis of
bcaba1
revealed three further candidate genes of this pathway: a putative P450 monooxygenase-encoding gene (
bcaba2
), an open reading frame without significant similarities (
bcaba3
), and a gene probably coding for a short-chain dehydrogenase/reductase (
bcaba4
). Targeted inactivation of the genes proved the involvement of BcABA2 and BcABA3 in ABA biosynthesis and suggested a contribution of BcABA4. The close linkage of at least three ABA biosynthetic genes is strong evidence for the presence of an abscisic acid gene cluster in
B. cinerea
.
A DNA Microarray Platform Based on Direct Detection of rRNA for Characterization of Freshwater Sediment-Related Prokaryotic Communities
Authors: Peplies Jörg , Lachmund Christine , Glöckner Frank Oliver , Manz Werner 
Published: July 2006

ABSTRACT
A DNA microarray platform for the characterization of bacterial communities in freshwater sediments based on a heterogeneous set of 70 16S rRNA-targeted oligonucleotide probes and directly labeled environmental RNA was developed and evaluated. Application of a simple protocol for the efficient background blocking of aminosilane-coated slides resulted in an improved signal-to-noise ratio and a detection limit of 10 ng for particular 16S rRNA targets. An initial specificity test of the system using RNA from pure cultures of different phylogenetic lineages showed a fraction of false-positive signals of ∼5% after protocol optimization and a marginal loss of correct positive signals. Subsequent microarray analysis of sediment-related community RNA from four different German river sites suggested low diversity for the groups targeted but indicated distinct differences in community composition. The results were supported by parallel fluorescence in situ hybridization in combination with sensitive catalyzed reporter deposition (CARD-FISH). In comparisons of the data of different sampling sites, specific detection of populations with relative cellular abundances down to 2% as well as a correlation of microarray signal intensities and population size is suggested. Our results demonstrate that DNA microarray technology allows for the fast and efficient precharacterization of complex bacterial communities by the use of standard single-cell hybridization probes and the direct detection of environmental rRNA, also in methodological challenging habitats such as heterogeneous lotic freshwater sediments.
13
C-Labeled Gluconate Tracing as a Direct and Accurate Method for Determining the Pentose Phosphate Pathway Split Ratio in
Penicillium chrysogenum
Authors: Kleijn Roelco J. , van Winden Wouter A. , Ras Cor , van Gulik Walter M. , Schipper Dick , Heijnen Joseph J. 
Published: July 2006

ABSTRACT

In this study we developed a new method for accurately determining the pentose phosphate pathway (PPP) split ratio, an important metabolic parameter in the primary metabolism of a cell. This method is based on simultaneous feeding of unlabeled glucose and trace amounts of [U-
13
C]gluconate, followed by measurement of the mass isotopomers of the intracellular metabolites surrounding the 6-phosphogluconate node. The gluconate tracer method was used with a penicillin G-producing chemostat culture of the filamentous fungus
Penicillium chrysogenum
. For comparison, a
13
C-labeling-based metabolic flux analysis (MFA) was performed for glycolysis and the PPP of
P. chrysogenum
. For the first time mass isotopomer measurements of
13
C-labeled primary metabolites are reported for
P. chrysogenum
and used for a
13
C-based MFA. Estimation of the PPP split ratio of
P. chrysogenum
at a growth rate of 0.02 h
−1
yielded comparable values for the gluconate tracer method and the
13
C-based MFA method, 51.8% and 51.1%, respectively. A sensitivity analysis of the estimated PPP split ratios showed that the 95% confidence interval was almost threefold smaller for the gluconate tracer method than for the
13
C-based MFA method (40.0 to 63.5% and 46.0 to 56.5%, respectively). From these results we concluded that the gluconate tracer method permits accurate determination of the PPP split ratio but provides no information about the remaining cellular metabolism, while the
13
C-based MFA method permits estimation of multiple fluxes but provides a less accurate estimate of the PPP split ratio.
Greengenes, a Chimera-Checked 16S rRNA Gene Database and Workbench Compatible with ARB
Authors: DeSantis T. Z. , Hugenholtz P. , Larsen N. , Rojas M. , Brodie E. L. , Keller K. , Huber T. , Dalevi D. , Hu P. , Andersen G. L. 
Published: July 2006

ABSTRACT

A 16S rRNA gene database (
http://greengenes.lbl.gov
) addresses limitations of public repositories by providing chimera screening, standard alignment, and taxonomic classification using multiple published taxonomies. It was found that there is incongruent taxonomic nomenclature among curators even at the phylum level. Putative chimeras were identified in 3% of environmental sequences and in 0.2% of records derived from isolates. Environmental sequences were classified into 100 phylum-level lineages in the
Archaea
and
Bacteria
.
Toll-Like Receptor 2 Polymorphisms and Nontuberculous Mycobacterial Lung Diseases
Authors: Ryu Yon Ju , Kim Eun Joo , Koh Won-Jung , Kim Hojoong , Kwon O Jung , Chang Jung Hyun 
Published: July 2006

ABSTRACT
To investigate the occurrence of the Toll-like receptor 2 (TLR2) polymorphisms in patients with pulmonary disease caused by nontuberculous mycobacteria (NTM), TLR2 Arg677Trp and Arg753Gln polymorphisms were examined. TLR2 polymorphisms do not appear to be responsible for host susceptibility to NTM lung disease, at least in the Korean population.
Epitope-Blocking Enzyme-Linked Immunosorbent Assay for Detection of Antibodies to Ross River Virus in Vertebrate Sera
Authors: Oliveira Nidia M. M. , Broom Annette K. , Mackenzie John S. , Smith David W. , Lindsay Michael D. A. , Kay Brian H. , Hall Roy A. 
Published: July 2006

ABSTRACT
We describe the development of an epitope-blocking enzyme-linked immunosorbent assay (ELISA) for the sensitive and rapid detection of antibodies to Ross River virus (RRV) in human sera and known vertebrate host species. This ELISA provides an alternative method for the serodiagnosis of RRV infections.
Understanding the Immune Responses to the Meningococcal Strain-Specific Vaccine MeNZB Measured in Studies of Infants
Authors: Ruijne N. , Lea R. A. , O'Hallahan J. , Oster P. , Martin D. 
Published: July 2006

ABSTRACT

Vaccine trials with infants enrolled between 6 and 10 weeks of age (young infants) and 6 and 8 months of age (older infants) provided an opportunity to evaluate immunoglobulin G (IgG) isotype distribution and avidity maturation as indicators of antibody function and immunologic memory. Following vaccination with a strain-specific outer membrane vaccine, MeNZB, pre- and postvaccination sera were used to determine IgG isotype responses and avidity indices (AI) in subsets of vaccinated subjects. Measurements of IgG isotypes involved 100 infants from each trial. AI were measured in 50 infants from the young infant trial who received a fourth vaccine dose and in 40 older infants from whom serum was collected 7 months after the primary vaccination course. IgG1 and IgG3 dominated the responses to the vaccine. A modest linear correlation (
P
< 0.001) occurred between serum bactericidal antibody (SBAb) titers and the total IgG or the IgG1 antibody units in older infants. The young infants showed a modest linear correlation between SBAb and total IgG (
P
= 0.005) and a weak linear correlation between SBAb and IgG1 (
P
= 0.003). Increased avidity with age was demonstrated in both groups. The AI in the young infants increased from 51.5% (95% confidence interval [CI], 47.7 to 54.7) postvaccination to 68.7% (95% CI, 65.5 to 71.9%) following the fourth dose of vaccine (
P
< 0.001). The mean avidity of the older infants increased significantly (
P
= 0.00012) from 42.4% (95% CI, 39.1 to 45.3%) postvaccination to 50.4% (95% CI, 47.2 to 53.6%) 4 months later. A fourth dose of MeNZB is now being given to young infants at 10 months of age.
Innate Immune Response toAnaplasma phagocytophilumContributes to Hepatic Injury
Authors: Scorpio Diana G. , von Loewenich Friederike D. , Göbel Heike , Bogdan Christian , Dumler J. Stephen 
Published: July 2006

ABSTRACTIn mice,Anaplasma phagocytophilumcontrol is independent of phagocyte oxidase (phox), inducible NO synthase (NOS2), tumor necrosis factor (TNF), and MyD88 Toll-like receptor signaling. We show that despite evasion of these host responses, phox, NOS2, TNF, and MyD88 are activated and contribute to inflammation and hepatic injury more thanA. phagocytophilumitself.
Quantifying Specific Antibody Concentrations by Enzyme-Linked Immunosorbent Assay Using Slope Correction
Authors: Barrette Roger W. , Urbonas Jessica , Silbart Lawrence K. 
Published: July 2006

ABSTRACT
Assessing the magnitude of an antibody response is important to many research and clinical endeavors; however, there are considerable differences in the experimental approaches used to achieve this end. Although the time-honored approach of end point titration has merit, the titer can often be misleading due to differences in how it is calculated or when samples contain high concentrations of low-avidity antibodies. One frequently employed alternative is to adapt commercially available enzyme-linked immunosorbent assay kits, designed to measure total antibody concentrations, to estimate antigen-specific antibody concentrations. This is accomplished by coating the specific antigen of interest in place of the capture antibody provided with the kit and then using the kit's standard curve to quantify the specific antibody concentration. This approach introduces considerable imprecision, due primarily to its reliance on a single sample dilution. This “single-point” approach fails to address differences in the slope of the sample titration curve compared to that of the standard curve. Here, we describe a general approach for estimating the effective concentration of specific antibodies, using antisera against foot-and-mouth disease virus VP1 peptide. This was accomplished by initially calculating the slope of the sample titration curve and then mathematically correcting the slope to that of a corresponding standard curve. A significantly higher degree of precision was attained using this approach rather than the single-point method.
Sequence-Based Methods for Identifying Epidemiologically Linked Herpes Simplex Virus Type 2 Strains
Authors: Martin Emily Toth , Koelle David M. , Byrd Benjamin , Huang Meei-Li , Vieira Jeffrey , Corey Lawrence , Wald Anna 
Published: July 2006

ABSTRACT
Traditional methods for confirming the identity of herpes simplex virus (HSV) isolates use restriction fragment length polymorphism (RFLP). However, RFLP is less amenable to high-throughput analyses of many samples, and the extent to which small differences in RFLP patterns distinguish between different viral strains remains unclear. Viral HSV type 2 (HSV-2) DNA isolates from 14 persons experiencing a primary HSV-2 infection and from their sexual partners were analyzed by RFLP and heteroduplex mobility assays. We also compared the HSV-2 sequences from seven regions, including noncoding regions between UL19 and UL20, UL24 and UL25, UL37 and UL38, and UL41 and UL42 and coding segments of the gC, gB, and gG genes. Although the resulting RFLP patterns of the couples were almost identical, minor banding differences existed between the source and susceptible partners in five couples. Heteroduplex mobility assays were unable to distinguish between unrelated strains. Overall, 22 sites of sequence variation were found in 1,482 bp of analyzed sequence. The DNA sequences differentiated between all unrelated infections, and epidemiologically related isolates had identical sequences in all but two pairs. Our results suggest that a multilocus assay based on several DNA sequences has the potential to be an informative tool for identifying epidemiologically related HSV-2 strains.
Use of Genotype MTBDR Assay for Molecular Detection of Rifampin and Isoniazid Resistance in
Mycobacterium tuberculosis
Clinical Strains Isolated in Italy
Authors: Miotto Paolo , Piana Federica , Penati Valeria , Canducci Filippo , Migliori Giovanni Battista , Cirillo Daniela Maria 
Published: July 2006

ABSTRACT

Mycobacterium tuberculosis
is one of the leading causes of death worldwide, and multidrug-resistant tuberculosis (MDR-TB) is associated with a high case fatality rate. Rapid identification of resistant strains is crucial for the early administration of appropriate therapy, for prevention of development of further resistance, and to curtail the spread of MDR strains. The Genotype MTBDR (Hain Lifescience, Nehren, Germany) is a reverse hybridization line probe assay designed for the rapid detection of
rpoB
and
katG
gene mutations in clinical isolates. The ability of this technique to correctly identify resistant and MDR-TB strains was tested on 206 isolates from the Italian drug resistance surveillance system. This panel included the majority of MDR strains isolated in Italy in the past 3 years. The results of the test were compared to conventional drug susceptibility test performed on isolated strains and verified by sequencing the regions of interest of the bacterial genome. The rate of concordance between the results of the MTBDR and those obtained with “in vitro” sensitivity was 91.5% (130 of 142) for rifampin and 67.1% (116 of 173) for isoniazid. We also applied this test directly to a panel of 36 clinical specimens collected from patients with active TB. The MTBDR correctly identified the two cases of MDR-TB included in the panel. These results show that the MTBDR test is useful in the detection and management of tuberculosis when MDR disease is suspected.
Sensitivities of Four New Commercial Hepatitis B Virus Surface Antigen (HBsAg) Assays in Detection of HBsAg Mutant Forms
Authors: Ly Thoai Duong , Servant-Delmas Annabelle , Bagot Sébastien , Gonzalo Sylvie , Férey Marie-Pierre , Ebel Anne , Dussaix Elisabeth , Laperche Syria , Roque-Afonso Anne-Marie 
Published: July 2006

ABSTRACT
Mutations in hepatitis B virus surface antigen (HBsAg) involving amino acid substitution within the immunodominant “a” determinant may affect the performance of commercial HBsAg assays. The performances of four HBsAg assays that recently received Conformité Européene marking, Advia Centaur HBsAg (Bayer), Monolisa HBsAg Ultra (Bio-Rad), Liaison HBsAg (Dia Sorin), and Vidas HBsAg Ultra (bioMérieux), were compared with that of the routinely used HBsAg assay AxSYM HBsAg V2 (Abbott). Assays were evaluated for (i) analytical sensitivity performance with a national reference HBsAg panel (including 10 samples with calibrated HBsAg concentrations from 0.04 to 2.24 ng/ml) and (ii) the detection of HBsAg mutants by studying a panel of 35 HBsAg mutants (23 collected from patients and 12 recombinant mutants). The limits of detection of these assays were <0.15 ng/ml (from 0.089 to 0.121 ng/ml). The sensitivity performances for mutant virus detection varied, ranging from 37.1% to 91.4%. The lack of detection of these mutants by commercial assays was probably due to the epitope recognition of the anti-HBs assay reagents in the capture phase and in the conjugates. The prevalence and clinical impact of HBsAg mutants are under investigation. However, the manufacturers must be vigilant in the design of the assays in order to reduce the risk of missing a broad range of described S gene mutants.
Amplification of the Hyphal Wall Protein 1 Gene To Distinguish
Candida albicans
from
Candida dubliniensis
Authors: Romeo Orazio , Racco Cosimo , Criseo Giuseppe 
Published: July 2006

ABSTRACT

The authors developed a new, simple, and reliable PCR/restriction fragment length polymorphism technique, using amplification of the hyphal wall protein 1 gene of
Candida albicans
and its gene homologue in
Candida dubliniensis
, to differentiate the two species of
Candida
. Performed with a new primer set, CRR-f/CRR-r, PCR produced two different fragments: one of 1,180 bp for
C. albicans
, and one of 930 bp for
C. dubliniensis
.
Bacillus anthracis
Virulent Plasmid pX02 Genes Found in Large Plasmids of Two Other
Bacillus
Species
Authors: Luna Vicki A. , King Debra S. , Peak K. Kealy , Reeves Frank , Heberlein-Larson Lea , Veguilla William , Heller L. , Duncan Kathleen E. , Cannons Andrew C. , Amuso Philip , Cattani Jacqueline 
Published: July 2006

ABSTRACT

In order to cause the disease anthrax,
Bacillus anthracis
requires two plasmids, pX01 and pX02, which carry toxin and capsule genes, respectively, that are used as genetic targets in the laboratory detection of the bacterium. Clinical, forensic, and environmental samples that test positive by PCR protocols established by the Centers for Disease Control and Prevention for
B. anthracis
are considered to be potentially
B. anthracis
until confirmed by culture and a secondary battery of tests. We report the presence of 10 genes (
acpA
,
capA
,
capB
,
capC
,
capR
,
capD
, IS
1627
, ORF 48, ORF 61, and
repA
) and the sequence for the capsule promoter normally found on pX02 in
Bacillus circulans
and a
Bacillus
species closely related to
Bacillus luciferensis
. Tests revealed these sequences to be present on a large plasmid in each isolate. The 11 sequences consistently matched to
B. anthracis
plasmid pX02, GenBank accession numbers AF188935.1, AE011191.1, and AE017335.3. The percent nucleotide identities for
capD
and the capsule promoter were 99.9% and 99.7%, respectively, and for the remaining nine genes, the nucleotide identity was 100% for both isolates. The presence of these genes, which are usually associated with the pX02 plasmid, in two soil
Bacillus
species unrelated to
B. anthracis
alerts us to the necessity of identifying additional sequences that will signal the presence of
B. anthracis
in clinical, forensic, and environmental samples.