bluuebunny/crossref_metadata_2025_split huggingface dataset:
Nucleotide Sequence-Based Multitarget Identification
Authors: Vinayagamoorthy T. , Mulatz Kirk , Hodkinson Roger 
Published: July 2003

ABSTRACTMULTIGEN technology (T. Vinayagamoorthy, U.S. patent 6,197,510, March 2001) is a modification of conventional sequencing technology that generates a single electropherogram consisting of short nucleotide sequences from a mixture of known DNA targets. The target sequences may be present on the same or different nucleic acid molecules. For example, when two DNA targets are sequenced, the first and second sequencing primers are annealed to their respective target sequences, and then a polymerase causes chain extension by the addition of new deoxyribose nucleotides. Since the electrophoretic separation depends on the relative molecular weights of the truncated molecules, the molecular weight of the second sequencing primer was specifically designed to be higher than the combined molecular weight of the first sequencing primer plus the molecular weight of the largest truncated molecule generated from the first target sequence. Thus, the series of truncated molecules produced by the second sequencing primer will have higher molecular weights than those produced by the first sequencing primer. Hence, the truncated molecules produced by these two sequencing primers can be effectively separated in a single lane by standard gel electrophoresis in a single electropherogram without any overlapping of the nucleotide sequences. By using sequencing primers with progressively higher molecular weights, multiple short DNA sequences from a variety of targets can be determined simultaneously. We describe here the basic concept of MULTIGEN technology and three applications: detection of sexually transmitted pathogens (Neisseria gonorrhoeae,Chlamydia trachomatis, andUreaplasma urealyticum), detection of contaminants in meat samples (coliforms, fecal coliforms, andEscherichia coliO157:H7), and detection of single-nucleotide polymorphisms in the humanN-acetyltransferase (NAT1) gene (S. Fronhoffs et al., Carcinogenesis 22:1405-1412, 2001).
Rapid Detection of
Candida albicans
in Clinical Blood Samples by Using a TaqMan-Based PCR Assay
Authors: Maaroufi Younes , Heymans Corine , De Bruyne Jean-Marc , Duchateau Valerie , Rodriguez-Villalobos Hector , Aoun Michel , Crokaert Françoise 
Published: July 2003

ABSTRACT

We describe a rapid and reproducible PCR assay for quantitation of the
Candida albicans
ribosomal DNA (rDNA) in clinical blood samples based on the TaqMan principle (Applied Biosystems), in which a signal is generated by cleavage of a template-specific probe during amplification. We used two fluorogenic probes based on universal, fungus-specific primers, one for the detection of
C. albicans
species DNA and one for the detection of all
Candida
genus DNA.
C. albicans
blastoconidia mixed with whole blood in a titration experiment yielded a linear PCR signal over a range of 3 orders of magnitude. The TaqMan-based PCR assay for
C. albicans
exhibited a low limit of detection (5 CFU/ml of blood) and an excellent reproducibility (96 to 99%). While the
C. albicans
species-specific probe had 100% specificity for
C. albicans
, all
Candida
genus-specific probes cross-reacted with other organisms likely to coinfect patients with
C. albicans
infections. On the basis of these data, we determined the
C. albicans
loads with a species-specific probe from 122 blood samples from 61 hematology or oncology patients with clinically proven or suspected systemic
Candida
infections. Eleven positive samples exhibited a wide range of
C. albicans
loads, extending from 5 to 100,475 CFU/ml of blood. The sensitivity and specificity of the present assay were 100 and 97%, respectively, compared with the results of blood culture. These data indicate that the TaqMan-based PCR assay for quantitation of
C. albicans
with a species-specific probe provides an attractive alternative for the identification and quantitation of
C. albicans
rDNA in pure cultures and blood samples.
Rapid Detection of Human Pathogenic Orthobunyaviruses
Authors: Weidmann Manfred , Rudaz Veronique , Nunes Marcio R. T. , Vasconcelos Pedro F. C. , Hufert Frank T. 
Published: July 2003

ABSTRACT

Modern detection and identification tools can help to provide answers to urgent questions about the incidence, prevalence, and epidemiology of currently emerging diseases. We developed highly sensitive one-step TaqMan reverse transcription-PCR assays with sensitivities ranging from 10
4
to 10
1
molecules for 11 human pathogens of the orthobunyaviruses. We compared the performances of these assays on three currently available cyclers (ABI-PRISM 7700, LightCycler, and SmartCycler). The assay for Oropouche virus (OROV) was tested using sera collected from days 1 to 5 after onset of OROV disease and was found to be greatly superior to an established nested PCR system. A mean copy number of 1.31 × 10
7
OROV RNA/ml of serum was detected. Diagnostic RNA detection can be used as early as day 1 after onset of OROV disease. The use of a mobile SmartCycler and a hands-on time of less than 3 h could help to intensify outbreak surveillance and control, especially in field studies.
Fluorescent Dye Terminator Sequencing Methods for Quantitative Determination of Replication Fitness of Human Immunodeficiency Virus Type 1 Containing the Codon 74 and 184 Mutations in Reverse Transcriptase
Authors: Nurpeisov Viktoria , Hurwitz Selwyn J. , Sharma Prem L. 
Published: July 2003

ABSTRACT
The fluorescent dye-labeled dideoxynucleotide automated DNA sequencing system has been routinely used for monitoring the development of resistance mutations in human immunodeficiency virus type 1 reverse transcriptase (RT) and protease genes during therapy. This system has provided information regarding the presence of mixtures of nucleotides in the clinical samples but has not previously been validated for the quantitative determination between peak heights and relative DNA concentration. We evaluated this system by using various ratios of wild-type and mutated DNA fragments and by performing sequencing reactions at actual melting temperatures of specific primers. Several different ratios of purified DNA fragments containing mixtures of L74/V74 and M184/V184 were sequenced, and peak heights were measured. Regression analysis between ratios of peak heights and DNA concentration demonstrated a statistically significant linear correlation, suggesting that the quantification of two different species of DNA in a mixture could be achieved with the fluorescent dye-labeled dideoxynucleotide system. These strategies have broader implications for the quantification of replication fitness of viruses, particularly those containing RT mutations at codons 74 and 184.
Novel Selective Medium for Isolation of
Burkholderia pseudomallei
Authors: Howard K. , Inglis T. J. J. 
Published: July 2003

ABSTRACT

Isolation of
Burkholderia pseudomallei
currently relies on the use of Ashdown's selective agar (ASA). We designed a new selective agar (
Burkholderia pseudomallei
selective agar [BPSA]) to improve recovery of the more easily inhibited strains of
B. pseudomallei. B. pseudomallei
,
Burkholderia cepacia
, and
Pseudomonas aeruginosa
were used to determine the selectivity and sensitivity of BPSA. BPSA was more inhibitory to
P. aeruginosa
and
B. cepacia
and should make recognition of
Burkholderia
species easier due to distinctive colony morphology. BPSA also inhibited
Enterococcus
,
Escherichia
,
Staphylococcus
, and
Streptococcus
. These results indicate that BPSA is a potential replacement for ASA.
First Isolation and Identification of
Rickettsia conorii
from Ticks Collected in the Region of Fokida in Central Greece
Authors: Psaroulaki Anna , Spyridaki Ioanna , Ioannidis Alexandros , Babalis Thomas , Gikas Achilleas , Tselentis Yannis 
Published: July 2003

ABSTRACT

Three different spotted-fever group rickettsiae—
Rickettsia conorii
,
R. massiliae
, and
R. rhipicephali
—were detected and identified by PCR-restriction fragment length polymorphism analysis in
Rhipicephalus
ticks collected from domestic animals in the Fokida region of Greece, where a high seroprevalence of antibodies to
R. conorii
was previously demonstrated. The infection rate of ticks was 1.6%. Moreover,
R. conorii
was isolated from one
Rhipicephalus sanguineus
tick.
Legionella pneumophila
Serogroup 1 Strain Paris: Endemic Distribution throughout France
Authors: Aurell Helena , Etienne Jerome , Forey Françoise , Reyrolle Monique , Girardo Pascale , Farge Pierre , Decludt Bénédicte , Campese Christine , Vandenesch François , Jarraud Sophie 
Published: July 2003

ABSTRACT

An analysis of 691 French clinical
Legionella
isolates showed that the endemic
L. pneumophila
serogroup 1 strain Paris was responsible for 12.2% of all cases of legionellosis and had a specific pulsed-field gel electrophoresis pattern. We also demonstrated the presence of this endemic clone throughout Europe.
Multilocus Sequence Typing of
Staphylococcus aureus
with DNA Array Technology
Authors: van Leeuwen Willem B. , Jay Corinne , Snijders Susan , Durin Nathalia , Lacroix Bruno , Verbrugh Henry A. , Enright Mark C. , Troesch Alain , van Belkum Alex 
Published: July 2003

ABSTRACT

A newly developed oligonucleotide array suited for multilocus sequence typing (MLST) of
Staphylococcus aureus
strains was analyzed with two strain collections in a two-center study. MLST allele identification for the first strain collection fully agreed with conventional strain typing. Analysis of strains from the second collection revealed that chip-defined MLST was concordant with conventional MLST. Array-mediated MLST data were reproducible, exchangeable, and epidemiologically concordant.
Detection of
Legionella pneumophila
by Real-Time PCR for the
mip
Gene
Authors: Wilson Deborah A. , Yen-Lieberman Belinda , Reischl Udo , Gordon Steve M. , Procop Gary W. 
Published: July 2003

ABSTRACT

A real-time PCR assay for the
mip
gene of
Legionella pneumophila
was tested with 27 isolates of
L. pneumophila
, 20 isolates of 14 other
Legionella
species, and 103 non-
Legionella
bacteria. Eight culture-positive and 40 culture-negative clinical specimens were tested. This assay was 100% sensitive and 100% specific for
L. pneumophila
.
Characterization of a Vancomycin-Resistant
Enterococcus faecalis
(VREF) Isolate from a Dog with Mastitis: Further Evidence of a Clonal Lineage of VREF in New Zealand
Authors: Manson Janet M. , Keis Stefanie , Smith John M. B. , Cook Gregory M. 
Published: July 2003

ABSTRACT

We report here on the characterization of a vancomycin-resistant
Enterococcus faecalis
(VREF) isolated from a dog with mastitis. The isolate was positive for the
vanA
,
ermB
, and
tet
(M) genes, with
vanA
and
ermB
carried on the same transferable plasmid. Comparison of this isolate with VREF from poultry and human sources in New Zealand demonstrated identical
Sma
I macrorestriction patterns and Tn
1546
-like elements. This is further evidence of a clonal lineage of VREF in New Zealand.
Evaluation of [
13
C]Urea Breath Test and
Helicobacter pylori
Stool Antigen Test for Diagnosis of
H. pylori
Infection in Children from a Developing Country
Authors: Cardinali Luciana de Carvalho Costa , Rocha Gifone Aguiar , Rocha Andreia Maria Camargos , de Moura Sílvia Beleza , de Figueiredo Soares Taciana , Esteves Ana Maria Braz , Nogueira Ana Margarida Miguel Ferreira , Cabral Mônica Maria Demas Álvares , de Carvalho Anfrisina Sales Teles , Bitencourt Paulo , Ferreira Alexandre , Queiroz Dulciene Maria Magalhães 
Published: July 2003

ABSTRACT

The [
13
C]urea breath test (
13
C-UBT) and
Helicobacter pylori
stool antigen test (HpSA) for the diagnosis of
H. pylori
infection in children were validated. The sensitivity, specificity, and positive and negative predictive values were 93.8, 99.1, 97.8, and 98.0%, respectively, for the
13
C-UBT and 96.9, 100, 100, and 98.0%, respectively, for HpSA. Both tests are appropriate for diagnosing
H. pylori
infection in children.
Comparison of Genotyping
Helicobacter pylori
Directly from Biopsy Specimens and Genotyping from Bacterial Cultures
Authors: Park Chang-Young , Kwak Minjung , Gutierrez Oscar , Graham David Y. , Yamaoka Yoshio 
Published: July 2003

ABSTRACT

PCR for
vacA
and
cagA
genotypes of
Helicobacter pylori
using DNA isolated from infected gastric biopsy specimens was approximately equal to genotyping using bacterial DNA from cultures. Inconsistent results were associated with low
H. pylori
density in biopsies. A higher proportion of mixed infection was found when biopsies were used.
Comparison of Four Antimicrobial Susceptibility Testing Methods To Determine the In Vitro Activities of Piperacillin and Piperacillin-Tazobactam against Clinical Isolates of
Enterobacteriaceae
and
Pseudomonas aeruginosa
Authors: Karlowsky James A. , Weaver Mellany K. , Thornsberry Clyde , Dowzicky Michael J. , Jones Mark E. , Sahm Daniel F. 
Published: July 2003

ABSTRACT

Susceptibility to piperacillin was similar to that to piperacillin-tazobactam (<1% difference) for 6,938 isolates of
Enterobacter aerogenes
and 13,954 isolates of
Enterobacter cloacae
tested using a Vitek system; for the same species, in contrast, susceptibility rates to piperacillin-tazobactam were 5.9 to 13.9% higher than to piperacillin using disk diffusion, MicroScan, and Vitek 2 testing. Unprecedented phenotypes (piperacillin susceptible and piperacillin-tazobactam intermediate; piperacillin intermediate and piperacillin-tazobactam resistant; piperacillin susceptible and piperacillin-tazobactam resistant) accounted for 6.1% of the results for
E. aerogenes
isolates and 6.0% of the results for
E. cloacae
isolates tested with the Vitek system.
Strain Variation among
Bordetella pertussis
Isolates from Québec and Alberta Provinces of Canada from 1985 to 1994
Authors: Peppler Mark S. , Kuny Sharee , Nevesinjac Anna , Rogers Christina , de Moissac Yvon R. , Knowles Kathleen , Lorange Manon , De Serres Gaston , Talbot James 
Published: July 2003

ABSTRACT

Pulsed-field gel electrophoresis and gene typing were able to differentiate among 3,597
Bordetella pertussis
isolates circulating in Alberta and Québec Provinces, Canada, from 1985 to 1994 and distinguish them from the strains used in vaccine production. This study provides a baseline for continued surveillance of prevalent and emerging strains of
B. pertussis
in Canada.
Involvement of Adherence and Adhesion
Staphylococcus epidermidis
Genes in Pacemaker Lead-Associated Infections
Authors: Klug Didier , Wallet Frédéric , Kacet Salem , Courcol René J. 
Published: July 2003

ABSTRACT

We explored three genes of attachment (
fbe
and
atlE
) and adhesion (
ica
) in 27 and 10
Staphylococcus epidermidis
strains involved in pacemaker-related infections (PMI) and intravascular-catheter-related infections (IVCI), respectively, and in 25 saprophytic strains. The detection rates of
fbe
and
atlE
were identical in PMI and IVCI strains, but
ica
detection rates were identical in PMI and saprophytic strains.
Evaluation of Use of Recombinant Em18 and Affinity-Purified Em18 for Serological Differentiation of Alveolar Echinococcosis from Cystic Echinococcosis and Other Parasitic Infections
Authors: Xiao Ning , Mamuti Wulamu , Yamasaki Hiroshi , Sako Yasuhito , Nakao Minoru , Nakaya Kazuhiro , Gottstein Bruno , Schantz Peter M. , Lightowlers Marshall W. , Craig Philip S. , Ito Akira 
Published: July 2003

ABSTRACT
To further evaluate recombinant Em18 antigen (rEm18) for immunodiagnosis of human alveolar echinococcosis, 208 serum samples were examined by enzyme-linked immunosorbent assay (ELISA). To comparatively assess the results of rEm18-ELISA, ELISA and immunoblot analysis with two affinity-purified native antigens were also performed with 45 selected serum samples. The results indicate that rEm18 is highly useful for serodiagnosis.
Candida dubliniensis
Infections in a Pediatric Population: Retrospective Identification from Clinical Laboratory Isolates of
Candida albicans
Authors: Kim Jean O. , Garofalo Lucille , Blecker-Shelly Deborah , McGowan Karin L. 
Published: July 2003

ABSTRACT

Candida dubliniensis
is a recently described species that shares many phenotypic and morphological features with
Candida albicans
. The clinical significance of isolating
C
.
dubliniensis
from the pediatric population is not clear, as most clinical isolates have been recovered from the oral cavities or bloodstreams of adults infected with human immunodeficiency virus. In order to understand further the epidemiology of
C
.
dubliniensis
in our pediatric population, we identified
C
.
dubliniensis
isolates from clinical isolates previously identified in the laboratory as
C
.
albicans
and conducted a retrospective chart review of cases of
C
.
dubliniensis
infections. A total of 205 isolates from 183 patients were tested, and only 14 (6.8%) were identified as
C
.
dubliniensis
. In 5 of the 14 positive cultures,
C
.
dubliniensis
was the sole organism isolated (two respiratory tract specimens, one tongue specimen, one vaginal specimen, and one skin specimen). A case review showed that there were no adverse outcomes for any of the patients, and only one of the patients with cultures positive for
C
.
dubliniensis
was immunocompromised. In our pediatric population, the distinction of
C
.
dubliniensis
from
C
.
albicans
did not prove to have significant clinical relevance. Data from further investigations may help to define better the role of
C
.
dubliniensis
as a potential pathogen in children.
Rapid Molecular Diagnosis of Posttraumatic Keratitis and Endophthalmitis Caused byAlternaria infectoria
Authors: Ferrer Consuelo , Montero Javier , Alió Jorge L. , Abad José L. , Ruiz-Moreno José M. , Colom Francisca 
Published: July 2003

ABSTRACTThe first case ofAlternaria infectoriaocular infection is reported. Keratitis and endophthalmitis developed after eye-perforating trauma from a lemon tree branch. Two months after surgery and empirical steroid and antibiotic treatment, diagnosis by molecular methods was performed. PCR amplification was positive for a fungus after 4 h. Antifungal treatment with amphotericin B and fluconazole was initiated immediately. DNA sequence analysis showedAlternaria infectoriato be the causal agent. After topical and systemic administration of antifungal treatment, ocular inflammation disappeared and visual acuity improved. DNA typing was found to be a useful tool to achieve early identification of the causal agent.
Evaluation of the Clinical Sensitivities of Three Viral Load Assays with Plasma Samples from a Pediatric Population Predominantly Infected with Human Immunodeficiency Virus Type 1 Subtype G and BG Recombinant Forms
Authors: Antunes Rute , Figueiredo Sofia , Bártolo Inês , Pinheiro Manuel , Rosado Lino , Soares Isabel , Lourenço Helena , Taveira Nuno 
Published: July 2003

ABSTRACT
The viral load assays AMPLICOR HIV-1 Monitor Test 1.5, Nuclisens HIV-1 QT, and Quantiplex HIV RNA 3.0 (bDNA) were evaluated for their abilities to quantify human immunodeficiency virus type 1 (HIV-1) RNA in 64 plasma samples from 21 children infected in Portugal. The children were infected with HIV-1 subtypes A1, B, F1, G, and BG recombinant virus. AMPLICOR v1.5 and Quantiplex v3.0 detected all samples, and there was a good correlation of results between the two kits. Thirty-eight specimens containing HIV-1 subtype B, G, or recombinant BG, could not be detected by Nuclisens HIV-1 QT. We also evaluated the new Retina HIV-1 assay on 21 samples that were HIV-1 positive; Retina HIV-1 failed to detect 5 of 11 subtype G specimens. AMPLICOR v1.5 and Quantiplex v3.0 assays may be used for HIV-1 RNA quantification in Portugal, whereas an improvement in sensitivity for subtype G and recombinant BG is required for Nuclisens HIV-1 QT and Retina HIV-1.
Tsukamurella
Conjunctivitis: a Novel Clinical Syndrome
Authors: Woo Patrick C. Y. , Ngan Antonio H. Y. , Lau Susanna K. P. , Yuen Kwok-Yung 
Published: July 2003

ABSTRACT

In this report, we describe the first three cases of
Tsukamurella
conjunctivitis in the literature. All three patients presented with congestion of one eye with small amounts of serous discharge for 1 to 2 days. All three recovered after 10 days of treatment with polymyxin B-neomycin or chloramphenicol eyedrops. Sequencing of the 16S rRNA genes of the three isolates recovered from the serous discharge of the three patients showed that they were all
Tsukamurella
species. The phenotypic characteristics of the isolate obtained from one patient best fit the phenotypic profile of
Tsukamurella pulmonis
, whereas those of the other two best fit that of
Tsukamurella tyrosinosolvens
.
Development of a Duplex PCR Assay for Detection of
Brachyspira hyodysenteriae
and
Brachyspira pilosicoli
in Pig Feces
Authors: La Tom , Phillips Nyree D. , Hampson David J. 
Published: July 2003

ABSTRACT

A duplex PCR (D-PCR) amplifying portions of the
Brachyspira hyodysenteriae
NADH oxidase gene and the
B. pilosicoli
16S rRNA gene was developed and then tested on DNA extracted from 178 porcine fecal samples. The feces also underwent anaerobic culture and species-specific PCRs. Fecal extraction-D-PCR detected seven additional samples containing
B. hyodysenteriae
and five more containing
B. pilosicoli
.
Differences in the Frequency of Resistance to Antiretroviral Drug Classes among Human Immunodeficiency Virus Type 1 Clinical Isolates
Authors: Campo Rafael E. , Lichtenberger Paola N. , Rosa Isabella , Suarez German , Rivera Fernando A. , Rodriguez Allan E. , Jayaweera Dushyantha T. , Wahlay Natalie A. , Kolber Michael A. 
Published: July 2003

ABSTRACTGenotypic resistance to all antiretroviral classes was widespread among human immunodeficiency virus type 1 isolates failing therapy. Resistance to nonnucleoside reverse transcriptase inhibitors was found most frequently and resistance to protease inhibitors was found least frequently, most likely due to differences in the number of enzymatic amino acid substitutions leading to resistance to each particular drug class.
Sequence of the
Escherichia coli
O121 O-Antigen Gene Cluster and Detection of Enterohemorrhagic
E. coli
O121 by PCR Amplification of the
wzx
and
wzy
Genes
Authors: Fratamico Pina M. , Briggs Connie E. , Needle Danielle , Chen Chin-Yi , DebRoy Chitrita 
Published: July 2003

ABSTRACT

The DNA sequence of the 15,155-bp O-antigen gene cluster of
Escherichia coli
O121 was determined, and 14 open reading frames were identified (all had the same transcriptional direction). Analyses of results indicated that the
wzx
(O-antigen flippase) and
wzy
(O-antigen polymerase) genes were
E. coli
O121 specific, so regions in these two genes were chosen for development of PCR assays. The PCR assays using DNA from 99
E. coli
O121 strains, strains representative of non-O121
E. coli
serogroups, and strains of other bacterial genera and PCR assays using DNA from seven enrichments of swine fecal samples naturally contaminated with
E. coli
O121 showed specificity for
E. coli
O121. Thus, the PCR assay can be employed to reliably identify
E. coli
O121 and to potentially detect the organism in food, fecal, and environmental samples.
Low-Stringency Single-Specific-Primer PCR as a Tool for Detection of Mutations in the
rpoB
Gene of Rifampin-Resistant
Mycobacterium tuberculosis
Authors: Carvalho Wania S. , Spindola de Miranda Silvana , Costa Kátia M. , Araújo José G.V.C. , Augusto Claudio J. , Pesquero João B. , Pesquero Jorge L. , Gomes Maria A. 
Published: July 2003

ABSTRACT

By the low-stringency single-specific-primer PCR technique, a highly sensitive and rapid method for diagnosis of rifampin resistance in
Mycobacterium tuberculosis
was established. Seven rifampin-resistant and five rifampin-susceptible specimens were analyzed. Rifampin resistance was determined by MIC measurement. A complex electrophoretic pattern consisting of many bands was obtained for both susceptible and rifampin-resistant isolates. The same pattern was obtained for all of the susceptible specimens, but differences between resistant and susceptible isolates were found. DNA sequencing showed that a particular mutation produces a specific electrophoretic pattern.