bluuebunny/crossref_metadata_2025_split huggingface dataset:
Construction Strategy for an Internal Amplification Control for Real-Time Diagnostic Assays Using Nucleic Acid Sequence-Based Amplification: Development and Clinical Application
Authors: Rodríguez-Lázaro David , D'Agostino Martin , Pla Maria , Cook Nigel 
Published: December 2004

ABSTRACT

An important analytical control in molecular amplification-based methods is an internal amplification control (IAC), which should be included in each reaction mixture. An IAC is a nontarget nucleic acid sequence which is coamplified simultaneously with the target sequence. With negative results for the target nucleic acid, the absence of an IAC signal indicates that amplification has failed. A general strategy for the construction of an IAC for inclusion in molecular beacon-based real-time nucleic acid sequence-based amplification (NASBA) assays is presented. Construction proceeds in two phases. In the first phase, a double-stranded DNA molecule that contains nontarget sequences flanked by target sequences complementary to the NASBA primers is produced. At the 5′ end of this DNA molecule is a T7 RNA polymerase binding sequence. In the second phase of construction, RNA transcripts are produced from the DNA by T7 RNA polymerase. This RNA is the IAC; it is amplified by the target NASBA primers and is detected by a molecular beacon probe complementary to the internal nontarget sequences. As a practical example, an IAC for use in an assay for the detection of
Mycobacterium avium
subsp.
paratuberculosis
is described, its incorporation and optimization within the assay are detailed, and its application to spiked and natural clinical samples is shown to illustrate the correct interpretation of the diagnostic results.
Evaluation of Multiplex PCRs for Diagnosis of Infection with Diarrheagenic
Escherichia coli
and
Shigella
spp
Authors: Aranda K. R. S. , Fagundes-Neto U. , Scaletsky I. C. A. 
Published: December 2004

ABSTRACT

We have developed two multiplex PCR assays that detect typical and atypical enteropathogenic
Escherichia coli
(EPEC) isolates, enteroaggregative
E. coli
(EAEC) isolates, enterotoxigenic
E. coli
(ETEC) isolates, enteroinvasive
E. coli
(EIEC) isolates, Shiga toxin-producing
E. coli
(STEC) isolates, and
Shigella
spp. The targets selected for each group were
eae
and
bfpA
for EPEC isolates, the target of probe CVD432 for EAEC isolates, the genes encoding heat-labile and heat-stable toxins for ETEC isolates,
stx
1
and
stx
2
for STEC isolates, and
ipaH
for EIEC isolates and
Shigella
spp. These PCRs were specific and sensitive for rapid detection of target isolates in stools. Among 150 stool specimens from the acute diarrhea tested, 9 samples (6%) had atypical EPEC, 9 (6%) had typical EPEC, 7 (4.7%) had EAEC, 3 (2%) had EIEC, 3 (2%) had
Shigella
spp., and 1 (0.7%) had an O26 STEC strain; we also detected mixed infections, 2 (1.3%) with EAEC and
Shigella
spp., 1 (0.7%) with atypical and typical EPEC strains, and another with atypical EPEC and EAEC strains. One of the multiplex PCRs directly applied to 36 stool specimens correctly identified 100% of EPEC and EAEC isolates.
Detection of the
van
Alphabet and Identification of Enterococci and Staphylococci at the Species Level by Multiplex PCR
Authors: Depardieu Florence , Perichon Bruno , Courvalin Patrice 
Published: December 2004

ABSTRACT

A multiplex PCR assay was developed for detection of the six types of glycopeptide resistance characterized in enterococci and for identification of
Enterococcus faecium
,
Enterococcus faecalis
,
Staphylococcus aureus
, and
Staphylococcus epidermidis
at the species level. Primers targeting the genes
vanA
,
vanB
,
vanC
,
vanD
,
vanE
,
vanG
, and
ddl
of
E. faecium
and
E. faecalis
and
nuc
of
S. aureus
and a chromosomal portion specific to
S. epidermidis
were designed to allow amplification of fragments with various sizes. This specific and sensitive technique allows detection of glycopeptide-resistant strains, in particular methicillin-resistant
S. aureus
, that may escape phenotype-based automated rapid methods.
Evaluation of Decontamination Methods and Growth Media for Primary Isolation of
Mycobacterium ulcerans
from Surgical Specimens
Authors: Yeboah-Manu Dorothy , Bodmer Thomas , Mensah-Quainoo Ernestina , Owusu Samuel , Ofori-Adjei David , Pluschke Gerd 
Published: December 2004

ABSTRACT

We evaluated four decontamination methods and one nondecontamination procedure in combination with four egg-based media for the primary isolation of
Mycobacterium ulcerans
from tissue specimens. With mycobacterial recovery and contamination rates of 75.6 and 2.4%, respectively, the combination of the oxalic acid decontamination method with Lowenstein-Jensen medium supplemented with glycerol yielded the best results.
Vancomycin Resistance,
esp
, and Strain Relatedness: a 1-Year Study of Enterococcal Bacteremia
Authors: Harrington S. M. , Ross T. L. , Gebo K. A. , Merz W. G. 
Published: December 2004

ABSTRACT

The prevalence of
esp,
a gene associated with infection-derived and outbreak strains, in enterococcal blood isolates from 2002 was determined. Fifty-five of 137 (40.1%)
Enterococcus faecalis
isolates, 30 of 58 (51.7%)
E. faecium
isolates, 1 of 1
E. raffinosus
isolate, 0 of 4
E. gallinarum
isolates, and 0 of 1
E. casseliflavus
isolate were positive.
esp
wasn't associated with vancomycin resistance (VR) or clinical service. VR
E. faecium
isolates were less genetically diverse than vancomycin-susceptible strains. A large cluster of VR isolates, belonging to
esp
-positive
E. faecium
, was revealed. These data support the hypothesis that
esp
and VR may contribute to dissemination of particular clones.
Study of Molecular Epidemiology of Candidiasis in Portugal by PCR Fingerprinting of
Candida
Clinical Isolates
Authors: Correia Alexandra , Sampaio Paula , Almeida Judite , Pais Célia 
Published: December 2004

ABSTRACT

PCR fingerprinting was used to type 177 yeast isolates obtained from two medical institutions.
Candida albicans
was the predominant species found, followed by
C. tropicalis
,
C. glabrata
,
C. parapsilosis
,
C. guilliermondii
, and
C. krusei
, which accounted for over 20% of the strains isolated. This survey represents the first study of molecular epidemiology of candidiasis in Portugal.
Identification of a Novel Mycoplasma Species from an Oriental White-Backed Vulture (
Gyps bengalensis
)
Authors: Oaks J. Lindsay , Donahoe Shannon L. , Rurangirwa Fred R. , Rideout Bruce A. , Gilbert Martin , Virani Munir Z. 
Published: December 2004

ABSTRACT

An intracellular organism was isolated from the tissues of an Oriental white-backed vulture (
Gyps bengalensis
) in chicken embryo fibroblast cell cultures. Biochemical and physical properties, ultrastructural features, and 16S ribosomal DNA sequencing classified this organism as a new taxon of mycoplasma, for which the name “
Mycoplasma vulturii
” is proposed.
Histidine Decarboxylase in
Enterobacteriaceae
Revisited
Authors: Wauters Georges , Avesani Véronique , Charlier Jacqueline , Janssens Michèle , Delmée Michel 
Published: December 2004

ABSTRACT

With a modification of Taylor's decarboxylation broth, histidine decarboxylase was detected in
Enterobacter aerogenes
,
Morganella morganii
,
Raoultella ornithinolytica
, and some strains of
Citrobacter youngae
and
Raoultella planticola
. This method provides a useful confirmatory test for identification of
E. aerogenes
strains.
Development and Evaluation of Fluorogenic TaqMan Reverse Transcriptase PCR Assays for Detection of Dengue Virus Types 1 to 4
Authors: Ito Mikako , Takasaki Tomohiko , Yamada Ken-Ichiro , Nerome Reiko , Tajima Shigeru , Kurane Ichiro 
Published: December 2004

ABSTRACT
The fluorogenic TaqMan reverse transcriptase PCR (RT-PCR) assay was developed for detecting each of the dengue virus (DV) types 1 to 4. DV genome was detected in all the 35 serum samples from confirmed dengue cases by the TaqMan RT-PCR, although it was not detected in 13 and 21% by conventional type-specific and cross-reactive RT-PCR, respectively.
Molecular Characterization of Fluconazole Resistance in a Case of
Candida albicans
Ocular Infection
Authors: Pancholi Preeti , Park Steven , Perlin David , Kubin Christine , Della-Latta Phyllis 
Published: December 2004

ABSTRACT

Ocular yeast infections in diabetics are a therapeutic challenge. Drug resistance and reduced azole susceptibility are major concerns. The case we describe characterizes a
Candida albicans
strain from a vitrectomy specimen that was susceptible to fluconazole by in vitro testing but recalcitrant to therapy. Molecular studies revealed transient overexpression of
CDR1
and
ERG11
mRNA in the presence of fluconazole that may have contributed to poor clinical response in this patient.
Early Reinfection with Human Metapneumovirus in an Infant
Authors: Ebihara Takashi , Endo Rika , Ishiguro Nobuhisa , Nakayama Tsuguyo , Sawada Hiroyuki , Kikuta Hideaki 
Published: December 2004

ABSTRACT
Human metapneumovirus (hMPV) is one of the major pathogens of respiratory illness. Reinfection with hMPV occurs frequently throughout life. We describe an infant who was infected with two different hMPV strains during a period of only 1 month.
Diagnosis based injury severity scaling: investigation of a method using Australian and New Zealand hospitalisations
Authors: Stephenson S , Henley G , Harrison J E , Langley J D 
Published: December 2004

Objective:To assess the performance of theInternational Classification of Diseases(ICD) based injury severity score, ICISS, when applied to two versions of the 10th edition of ICD, ICD-10 and ICD-10-AM.Design:ICISS was assessed on its ability to predict threat to life using logistic regression modelling. Models used ICISS and age as predictors and survival as the outcome.Setting:Australia and New Zealand.Patients or subjects:Hospitalisations with an ICD-10-AM principal diagnosis in the range S00–T89 from 1 July 1999 to 30 June 2001 (Australia) or 1 July 1999 to 31 December 2001 (New Zealand).Interventions:None.Main outcome measures:The models were assessed in terms of their discrimination, measured by the concordance score, and calibration, measured using calibration curves and the Hosmer-Lemeshow statistic.Results:523 633 Australian and 124 767 New Zealand hospitalisations were selected, including 7230 and 1565 deaths respectively. Discrimination was high in all the fitted models with concordance scores of 0.885 to 0.910. Calibration results were also promising with all calibration curves being close to linear, though ICISS appeared to underestimate mortality somewhat for cases with an ICISS score less than 0.6. Overall ICISS performed better when applied to the Australian than the New Zealand hospitalisations. Australian and New Zealand hospitalisations were very similar. ICISS was also only a little more successful when ICD-10-AM rather than mapped ICD-10 was used.Conclusions:ICISS appears to be a reasonable way to estimate severity for databases using ICD-10 or ICD-10-AM. It is also likely to work well for other clinical variants of ICD-10.
Child and family safety device affordability by country income level: an 18 country comparison
Authors: Hendrie D , Miller T R , Orlando M , Spicer R S , Taft C , Consunji R , Zaloshnja E 
Published: December 2004

Objective:To compare availability, urban price, and affordability of child/family safety devices between 18 economically diverse countries.Design:Descriptive: urban price surveys by local safety organisations or shoppers.Setting:Retail stores and internet vendors.Main outcome measures:Prices expressed in US dollars, and affordability measured by hours of factory work needed to buy a child safety seat, a belt-positioning booster seat, a child bicycle helmet, and a smoke alarm.Results:Prices of child and family safety devices varied widely between countries but the variation for child safety seats and bicycle helmets did not relate strongly to country income. Safety devices were expensive, often prohibitively so, in lower income countries. Far more hours of factory work were required to earn a child safety device in lower income than middle income, and middle income than higher income, countries. A bicycle helmet, for example, cost 10 hours of factory work in lower income countries but less than an hour in higher income countries. Smoke alarms and booster seats were not available in many lower income countries.Conclusions:Bicycles and two-axle motor vehicles were numerous in lower and middle income countries, but corresponding child safety devices were often unaffordable and sometimes not readily available. The apparent market distortions and their causes merit investigation. Advocacy, social marketing, local device production, lowering of tariffs, and mandatory use legislation might stimulate market growth. Arguably, a moral obligation exists to offer subsidies that give all children a fair chance of surviving to adulthood.
Epidemiology of lower limb fractures in general practice in the United Kingdom
Authors: Kaye J A , Jick H 
Published: December 2004

Study objective:To estimate the incidence of lower limb fractures in the United Kingdom and assess the relative importance of various risk factors for lower limb fractures.Design:Cohort analysis and matched case-control study.Setting:General practices contributing information to the General Practice Research Database.Subjects:Individuals registered with these general practices who were at risk for a first time lower limb fracture from 1 January1990 to 31 December 2001.Main outcome measures:Age, sex, and fracture site specific incidence rates; relative risks and population attributable risks for various medical risk factors.Results:Overall, the risk of lower limb fracture was 17% higher in women then in men. Within age groups, men and women had generally similar proportions of fractures at specific sites in the lower limb. Among the risk factors evaluated, road collisions were associated with the highest relative risk for lower limb fracture, but only accounted for 3.1% or less of the population attributable risk for specific fracture types in any age group. The relative risk for lower limb fracture associated with a diagnosis of dementia was 2.3 (95% confidence interval 2.0 to 2.6), while relative risk estimates for other medical diagnoses were less than 2. Fracture risk was increased among current users of corticosteroids, antipsychotics, antidepressants, and hypnotic/sedatives, but the population attributable risks for each of these drug classes within fracture and age specific strata were only 3.0% or less.Conclusions:Many risk factors for lower limb fracture have been identified, but population attributable risk estimates for various risk factors are small. These findings suggest that multifactorial prevention programs are needed to decrease the incidence of lower limb fractures in the general population.
Mechanical testing of recombinant human bone morphogenetic protein-7 regenerated bone in sheep mandibles
Authors: Kontaxis A , Abu-Serriah M , Ayoub A F , Barbenel J C 
Published: December 2004

A new method was developed in this study for testing excised sheep mandibles as a cantilever. The method was used to determine the strength and stiffness of sheep hemi-mandibles including a 35 mm defect bridged by regenerated bone. Recombinant human bone morphogenetic protein-7 (rhBMP-7) in a bovine collagen type-I carrier was used for the bone regeneration. Initial tests on ten intact sheep mandibles confirmed that the strength, stiffness and area beneath the load-deformation curves of the right and left hemi-mandibles were not significantly different, confirming the validity of using the contra-lateral hemi-mandible as a control side. Complete bone regeneration occurred in six hemi-mandibles treated with rhBMP, but the quality and mechanical properties of the bone were very variable. The new bone in three samples contained fibrous tissue and was weaker and less stiff than the contra-lateral side (strength, 10-20 per cent; stiffness, 6-15 per cent). The other half had better-quality bone and was significantly stiffer and stronger ( p < 0.05), with strength 45-63 per cent and stiffness 35-46 per cent of the contra-lateral side. Hemi-mandibles treated with collagen alone had no regenerated bone bridge suggesting that 35 mm is a critical-size bone defect.
Sentinel node biopsy to evaluate the metastatic dissemination of oesophageal adenocarcinoma
Authors: Lamb P J , Griffin S M , Burt A D , Lloyd J , Karat D , Hayes N 
Published: December 2004

Abstract

Background
The aim of this study was to determine the feasibility and accuracy of sentinel lymph node (SLN) biopsy for oesophageal adenocarcinoma.

Methods
Fifty-seven patients with adenocarcinoma of the lower oesophagus (n = 40) or gastric cardia (n = 17) underwent endoscopic peritumoral injection of 99mTc-radiolabelled nanocolloid before en bloc resection with extended lymphadenectomy. SLNs were identified during surgery using a handheld γ probe and the pattern of radioactive uptake was quantified after operation. All 1667 resected lymph nodes were examined immunohistochemically for micrometastases.

Results
SLNs were identified in all 57 patients. They contained metastases (n = 32) or micrometastases (n = 3) in 35 of 37 node-positive patients and there were two false-negative studies. The overall accuracy of SLN biopsy was 96 per cent and SLNs were more likely to contain tumour than other lymph nodes (P &lt; 0·001). Tumour-infiltrated nodal stations had a higher proportion of radioactive uptake (P &lt; 0·001). Lower oesophageal tumours had a greater proportion of SLNs (P = 0·018), radioactive uptake (P &lt; 0·001) and malignant nodes (P = 0·004) in the mediastinum than gastric cardia tumours.

Conclusion
The sentinel node concept is applicable to oesophageal adenocarcinoma and could be used to tailor the extent of lymphadenectomy. There is a close relationship between patterns of radioactive uptake and lymphatic tumour dissemination, which differ for lower oesophageal and gastric cardia tumours.
Determination of therapeutic threshold in sacral nerve modulation for faecal incontinence
Authors: Koch S M P , van Gemert W G , Baeten C G M I 
Published: December 2004

Abstract

Background
The aim of the study was to determine the therapeutic stimulation threshold in patients with successful sacral nerve modulation for faecal incontinence.

Methods
Patients who had undergone successful permanent sacral nerve modulator implantation and had been followed up for a minimum of 3 months were included. The sensitivity threshold and motor threshold were determined and correlated with therapeutic response. Patients went home with the stimulator set at 0·6 V below the sensitivity threshold. Each week the voltage was increased by 0·2 V until the sensitivity threshold was reached. The effects on anorectal physiology and continence were recorded.

Results
Eight patients (seven women) with a median age of 58·5 years were included. The median follow-up was 6·3 months. The median sensibility threshold volume of rectal sensation was 50 ml, the median urge threshold volume was 140 ml and the median maximum tolerated rectal volume 240 ml. The median number of incontinence episodes and days per week affected by incontinence decreased from 5·0 and 3·8 before operation to 0·7 and 0·7 respectively after follow-up for 3 months. At anorectal manometry the median resting and stimulation anal canal pressures were 57 and 85 mmHg respectively, and remained constant over time. The therapeutic response threshold was significantly lower than the sensitivity threshold (median 1·6 versus 1·7 V; P = 0·042). The median motor threshold was 2·1 V, significantly higher than the sensitivity threshold (P = 0·009). The stimulation threshold for suboptimal therapeutic response was 1·4 V. In five of the eight patients the therapeutic response threshold was the same as the sensitivity threshold.

Conclusion
Sacral nerve modulation can produce a therapeutic effect below the sensitivity threshold. A lower stimulation voltage increases the lifespan of the pulse generator.
Transient transfection of epidermal growth factor receptor gene into MCF7 breast ductal carcinoma cell line
Authors: Alokail Majed S. 
Published: December 2004

AbstractEpidermal growth factor receptor (EGFR) is activated by autocrine growth factors in many types of tumours, including breast tumours. This receptor has been linked to a poor prognosis in breast cancer and may promote proliferation, migration, invasion, and cell survival as well as inhibition of apoptosis. Human breast ductal carcinoma MCF7 cells were transfected using FuGENE™ 6 with 1 μg of pcDNA3‐EGFR containing the full‐length human EGFR promoter or 1 μg of the vectors alone (pcDNA3). The transfected cells were transferred into a 25‐cm2 flask containing growth medium and G418. Confluent cultures were lysed, total protein levels measured and electrophoresed. The electrophoresed samples were transferred to nitrocellulose and incubated overnight at 4°C with either anti‐EGFR or anti‐phospho‐ERK and immunoreactive bands were visualized using HRP‐linked secondary antibody. We created a model system of EGFR overexpression in MCF7 clones with stably transfected pcDNA3/EGFR plasmid. These cells have been shown to promote substantial phosphorylation of both ERK1 and ERK2. The high level of EGFR and ERK1/2 phosphorylation was not seen in the pcDNA3 vector control cells or in non‐transfected cells. In this article we describe successful transient transfection experiments on MCF7 cells using the FuGENE™ 6 Transfection Reagent. The overexpression of EGFR could be a mediated stress response and a survival signal that involves ERK1 and ERK2 phosphorylation. Copyright © 2004 John Wiley & Sons, Ltd.
Cytological features of epithelioid mesenchymal neoplasms: A study of 21 cases
Authors: Lin Oscar , Olgac Semra , Zakowski Maureen 
Published: December 2004

AbstractEpithelioid mesenchymal neoplasms (EMNs) are rare tumors that share cytological, histological, and immunohistochemical features with epithelial tumors. It is important to distinguish EMNs from epithelial tumors in cytology specimens due to their different clinical management and prognosis. The cytomorphological features of histologically confirmed EMN were reviewed. Twenty‐one cytological specimens of EMN were evaluated and characterized by polygonal cells with moderate to abundant dense cytoplasm, prominent nucleoli, and pleomorphism. Additional findings included the presence of a distinct population of spindle cells, hemosiderin‐containing cells, multinucleated cells, and granuloma‐like structures in selected cases. Cytokeratin immunoreactivity was seen in two cases and was negative in one case. This study shows that the cytological features of EMNs and epithelial tumors overlap; nonetheless, some features are more helpful in suggesting EMN. A panel of immunocytochemical studies must include specific mesenchymal markers to avoid a misdiagnosis of carcinoma in cases of cytokeratin‐positive EMN. Diagn. Cytopathol. 2005;32:5–10. © 2005 Wiley‐Liss, Inc.