bluuebunny/crossref_metadata_2025_split huggingface dataset:
Single Rapid Real-Time Monitored Isothermal RNA Amplification Assay for Quantification of Human Immunodeficiency Virus Type 1 Isolates from Groups M, N, and O
Authors: de Baar Michel P. , van Dooren Maaike W. , de Rooij Esther , Bakker Margreet , van Gemen Bob , Goudsmit Jaap , de Ronde Anthony 
Published: July 2002

ABSTRACT

Because human immunodeficiency virus type 1 (HIV-1) subtypes and circulating recombinant forms (CRFs) are spreading rapidly worldwide and are becoming less confined to a geographical area, RNA assays that can detect and quantify all HIV-1 isolates reliably are in demand. We have developed a fast, real-time monitored RNA assay based on an isothermal nucleic acid sequence-based amplification technology that amplifies a part of the long terminal repeat region of the HIV-1 genome. Real-time detection was possible due to the addition of molecular beacons to the amplification reaction that was monitored in a fluorimeter with a thermostat. The lower level of detection of the assay was 10 HIV-1 RNA molecules per reaction, and the lower level of quantification was 100 copies of HIV-1 RNA with a dynamic range of linear quantification between 10
2
and 10
7
RNA molecules. All HIV-1 groups, subtypes, and CRFs could be detected and quantified with equal efficiency, including the group N isolate YBF30 and the group O isolate ANT70. To test the clinical utility of the assay, a series of 62 serum samples containing viruses that encompassed subtypes A through G and CRFs AE and AG of HIV-1 group M were analyzed, and these results were compared to the results of a commercially available assay. This comparison showed that the quantification results correlated highly (
R
2
= 0.735) for those subtypes that could be well quantified by both assays (subtypes B, C, D, and F), whereas improved quantification was obtained for subtypes A and G and CRFs AE and AG. A retrospective study with six individuals infected with either a subtype A, B, C, or D or an AG isolate of HIV-1 group M, who were treated with highly active antiretroviral therapy, revealed that the assay was well suited to the monitoring of therapy effects. In conclusion, the newly developed real-time monitored HIV-1 assay is a fast and sensitive assay with a large dynamic range of quantification and is suitable for quantification of most if not all subtypes and groups of HIV-1.
Genotypic Characterization of
Toxoplasma gondii
Strains Associated with Human Toxoplasmosis in Spain: Direct Analysis from Clinical Samples
Authors: Fuentes Isabel , Rubio Jose M. , Ramı́rez Carmen , Alvar Jorge 
Published: July 2002

ABSTRACT

Genetic analysis of the
SAG2
locus was performed to determine the prevalence of the different genotypes of
Toxoplasma gondii
(strain types I, II, and III) associated with human toxoplasmosis in Spain. This determination was made directly from primary clinical samples, obviating the previous process of isolation in mice or cell culture. A total of 34 isolates of
T. gondii
, collected from immunocompromised patients and congenital infection cases, were analyzed. Restriction fragment length polymorphism in PCR-amplified
SAG2
products was used to group strains into one of the three genotypes of
T. gondii
. Complete characterization of the
SAG2
gene was successful in 76.5% of the cases, demonstrating the feasibility of direct genotype analysis from clinical samples of different origins. Strains of
T. gondii
type II were the most prevalent in immunocompromised patients, with 52% of cases, while strains of type I were present in 75% of the congenital infection cases. These data differ from previous reports that show type II strains to be mostly associated with all kinds of human toxoplasmosis. These differences might be an effect of selection in the process of culture and isolation of the samples performed by other researchers prior to strain characterization.
Relevance of Reactivity in Commercially Available Hepatitis C Virus Antibody Assays
Authors: Polywka Susanne , Schröter Matthias , Feucht Heinz-Hubert , Zöllner Bernhard , Laufs Rainer 
Published: July 2002

ABSTRACT
Sera from 2,148 patients were tested with a third-generation microparticle enzyme immunoassay (MEIA), a confirmatory assay, and a reverse transcription-PCR. Overall, 85.6% of reactivities were confirmed, 13.2% were shown to be unspecifically reactive, and 1.2% were indeterminate. The rate of confirmed MEIA reactivities clearly depended on the strength of the reactivity.
Use of Recombinant Mitogillin for Improved Serodiagnosis ofAspergillus fumigatus-Associated Diseases
Authors: Weig Michael , Frosch Matthias , Tintelnot Kathrin , Haas Andrea , Groß Uwe , Linsmeier Bernd , Heesemann Jürgen 
Published: July 2002

ABSTRACTDuring human infection,Aspergillus fumigatussecretes a 18-kDa protein that can be detected as an immunodominant antigen in the urine of infected patients. Recently, this protein was shown to be mitogillin, a ribotoxin that cleaves a single phosphodiester bond of the 29S rRNA of eukaryotic ribosomes. We proved the immunogenic capacity of mitogillin in a rabbit animal model, indicating its usefulness as an antigen for serological diagnosis of invasive aspergillosis. The mitogillin gene fromA. fumigatuswas transferred from plasmid pMIT+ to expression vector pQE30 and expressed inEscherichia colias a fusion protein. Purified recombinant mitogillin was recognized by serum immunoglobulin G (IgG) of polyclonal rabbit sera that were obtained by immunization with purified native mitogillin. Consequently, we developed an enzyme-linked immunosorbent assay for detection of IgG, IgM, and IgA antibodies to recombinant mitogillin. In serum samples of patients suffering from aspergilloma (AO;n= 32), invasive pulmonary aspergillosis (IPA;n= 42), or invasive disseminated aspergillosis (IDA;n= 40), a good correlation of production of IgG antibody against mitogillin and clinical disease was observed (for patients with AO, 100% [32 of 32] were positive; for patients with IPA, 64% [31 of 42] were positive; for patients with IDA, 60% [24 of 40] were positive). In contrast, positive titers for serum IgG and IgM antibodies against mitogillin were found in only 1.3% of the serum samples of healthy volunteers and positive titers for IgA antibody were found in only 1.0% of the serum samples of healthy volunteers (n= 307; specificity = 95.4%). These results indicate that recombinant mitogillin expressed inE. colican be used for improvement of the serodiagnosis ofA. fumigatus-associated diseases.
iceA
Genotypes of
Helicobacter pylori
Strains Isolated from Brazilian Children and Adults
Authors: Ashour Abdussalam Ali Ramadan , Birchal Collares Guilherme , Nogueira Mendes Edilberto , Ribeiro de Gusmão Valquı́ria , Queiroz Dulciene Maria de Magalhães , Prazeres Magalhães Paula , Teles de Carvalho Anfrisina Sales , de Oliveira Celso Affonso , Ferreira Nogueira Ana Margarida Miguel , Aguiar Rocha Gifone , Rocha Andreia Maria Camargos 
Published: July 2002

ABSTRACT

Data concerning the geographic distribution of
iceA
alleles are scarce, and information on the association of the gene with the disease is rare and still controversial. Furthermore, no such study has been developed in Brazil, where duodenal ulcer and gastric adenocarcinoma are very common. We investigated, by PCR, the frequency of
iceA
alleles and
cagA
status in
Helicobacter pylori
strains isolated from 142 patients (62 children and 80 adults; 66 female; mean age, 30.0 years; age range, 3 to 78 years) with gastritis, duodenal ulcer, or gastric adenocarcinoma.
iceA
was identified in bacterium samples obtained from all patients. Eleven (7.7%) of them were infected with multiple strains. Among the patients with nonmixed infection,
iceA2
allele was detected in 118 (90.1%).
iceA2
allele was associated with ulcer (
P
= 0.02) and with carcinoma (
P
= 0.001).
iceA2
amplicons of 229, 334, or 549 bp were detected, but none of them was associated with the patient's disorder.
iceA2
strains were more frequent in patients older than 7 years (
P
= 0.001). The gene was also more frequent in strains obtained from males (
P
= 0.02).
cagA
was more common in strains obtained from carcinoma (
P
= 0.0008) and ulcer patients (
P
< 0.006).
cagA
-positive strains were more frequent in children older than 7 years (
P
< 0.003). No association between
cagA
status and sex was found (
P
= 0.28). In conclusion, we think
iceA
should not be used as a reliable marker for predicting the clinical outcome of
H. pylori
infection.
High Prevalence of VanB2 Vancomycin-Resistant
Enterococcus faecium
in Taiwan
Authors: Lu Jang-Jih , Perng Cherng-Lih , Ho Ming-Fa , Chiueh Tzong-Shi , Lee Wei-Hwa 
Published: July 2002

ABSTRACT

Thirty-six VanB glycopeptide-resistant
Enterococcus faecium
isolates were collected from patients in five different hospitals in Taiwan. The vancomycin resistance genes were amplified by the long
vanB
PCR, which amplifies the 6,373-bp
vanB
gene cluster including the

vanR
B2
, vanS
B2
, vanY
B2
, vanW
B2
, vanH
B2
, vanB2

, and

vanX
B2

genes. The deduced amino acid sequences were found to be 95 to 98% homologous to those of the
vanB1
gene cluster: VanR
B1
, 97%; VanS
B1
, 97%; VanY
B1
, 96%; VanH
B1
, 95%; VanB1, 96%; and VanX
B1
, 98%. Restriction enzyme analysis of the long
vanB
PCR products revealed that all 36 isolates had the same
vanB2
-specific pattern. DNA sequence analysis of the
vanB2
gene, which is a
d
-Ala–
d
-Lac ligase gene, revealed that none of the 36 sequences were identical to the previously published
vanB2
sequence. Thirty-one isolates had 1 nucleotide different from the published
vanB2
sequence. The sequences of the other five isolates differed from the published
vanB2
sequence by 2 or 3 nucleotides. Four isolates with a low or moderate resistance to vancomycin (MIC = 4 to 32 μg/ml) were found to have the same leucine-to-methionine change at amino acid position 308 of the
vanB2
gene. The genomic DNAs of all 36 isolates were digested with
Sma
I and then typed by pulsed-field gel electrophoresis (PFGE). Eight different PFGE types (I to VIII) were observed, and type I was found to be prevalent in all hospitals examined in this study. This result suggests that intra- and interhospital dissemination of this
E. faecium
strain has occurred in Taiwan.
Frequent Detection of the Human Herpesvirus 6-Specific Genomes in the Livers of Children with Various Liver Diseases
Authors: Ozaki Yoshikazu , Tajiri Hitoshi , Tanaka-Taya Keiko , Mushiake Sotaro , Kimoto Akihito , Yamanishi Koichi , Okada Shintaro 
Published: July 2002

ABSTRACT
This study was performed to investigate the frequency of human herpesvirus 6 (HHV-6) infection of the liver in children with a variety of liver diseases and to evaluate the role of HHV-6 infection in pediatric patients with prolonged non-B non-C hepatitis. Detection of the HHV-6 genomes in liver, in peripheral blood mononuclear cells (PBMC), and in plasma was performed by PCR or by in situ hybridization. Liver biopsy materials from 48 patients, in whom HHV-6 infection was serologically confirmed, were available for PCR analysis. Sequences of the HHV-6B genome were detectable in the livers of 36 of 48 patients (75%). The presence of the genome was not associated with serum transaminase activities. The genome was detectable in PBMC of 22 of 31 (71%) patients tested. In these 31 patients HHV-6 was detected in both the livers and PBMC of 20, was detected in PBMC but not in the livers of 2, was detected in the livers but not in PBMC of 3, and was detected in neither of samples of 6. In situ hybridization of the livers of six patients showed the presence of the HHV-6B genome in the nuclei of hepatocytes. The anti-HHV-6 immunoglobulin M antibody was detectable in 2 of 9 of the non-B non-C hepatitis patients, whereas none of the 22 patients with etiology-defined liver diseases tested positive. Cell-free viral DNA was not detectable in either group of patients. Our results showed that HHV-6B is frequently present in the livers of children with a variety of liver diseases but do not support the assumption that HHV-6B infection of the liver is associated with prolonged non-B non-C hepatitis.
Isolation and Characterization of
Borrelia burgdorferi
Sensu Lato Strains in an Area of Italy Where Lyme Borreliosis Is Endemic
Authors: Ciceroni Lorenzo , Ciarrochi Simonetta , Ciervo Alessandra , Mondarini Valeria , Guzzo Francesco , Caruso Giuseppe , Murgia Rossella , Cinco Marina 
Published: July 2002

ABSTRACT

Between 1993 and 1998, we isolated
Borrelia burgdorferi
sensu lato from 55 of the 119 patients with clinically diagnosed Lyme borreliosis who were admitted to “San Martino” Hospital in Belluno, Veneto, an Adriatic region in northeastern Italy where Lyme borreliosis is endemic. Upon hospitalization, all patients presented erythema migrans. Isolates were typed using ribosomal DNA PCR-restriction fragment length polymorphism (RFLP) analysis of the
rrfA-rrlB
intergenic spacer. Of the 41 isolates typed, 37 belonged to
Borrelia afzelii
, 2 to
Borrelia garinii
, and 2 to
B. burgdorferi
sensu stricto. Pulsed-field gel electrophoresis, performed on 21 strains (13 new isolates and 8 controls), revealed different RFLP patterns within the
B. garinii
and
B. afzelii
strains; among the five
B. garinii
strains and the 12
B. afzelii
strains, three or two different RFLP patterns were identified, according to the restriction enzyme used. The protein patterns of the new isolates confirmed their genotypic classification and revealed the level of expression of some immunodominant proteins like OspA and other characteristic Osps. These findings constitute the first report of such a high recovery rate of
B. burgdorferi
from patients in a very restricted area in Italy; they also indicate the predominance of the genospecies
B. afzelii
in the study area and the heterogeneity of the circulating strains.
Nomenclature of Major Antimicrobial-Resistant Clones of
Streptococcus pneumoniae
Defined by the Pneumococcal Molecular Epidemiology Network
Authors: McGee L. , McDougal L. , Zhou J. , Spratt B. G. , Tenover F. C. , George R. , Hakenbeck R. , Hryniewicz W. , Lefévre J. C. , Tomasz A. , Klugman K. P. 
Published: July 2002

ABSTRACT
The emergence of disease caused by penicillin-resistant and multidrug-resistant pneumococci has become a global concern, necessitating the identification of the epidemiological spread of such strains. The Pneumococcal Molecular Epidemiology Network was established in 1997 under the auspices of the International Union of Microbiological Societies with the aim of characterizing, standardizing, naming, and classifying antimicrobial agent-resistant pneumococcal clones. Here we describe the nomenclature for 16 pneumococcal clones that have contributed to the increase in antimicrobial resistance worldwide. Guidelines for the recognition of these clones using molecular typing procedures (pulsed-field gel electrophoresis, BOX-PCR, and multilocus sequence typing) are presented, as are the penicillin-binding profiles and macrolide resistance determinants for the 16 clones. This network can serve as a prototype for the collaboration of scientists in identifying clones of important human pathogens and as a model for the development of other networks.
Direct Detection of
Legionella
Species from Bronchoalveolar Lavage and Open Lung Biopsy Specimens: Comparison of LightCycler PCR, In Situ Hybridization, Direct Fluorescence Antigen Detection, and Culture
Authors: Hayden R. T. , Uhl J. R. , Qian X. , Hopkins M. K. , Aubry M. C. , Limper A. H. , Lloyd R. V. , Cockerill F. R. 
Published: July 2002

ABSTRACT

We developed a rapid thermocycling, real-time detection (also known as real-time PCR) method for the detection of
Legionella
species directly from clinical specimens. This method uses the LightCycler (Roche Molecular Biochemicals, Indianapolis, Ind.) and requires approximately 1 to 2 h to perform. Both a
Legionella
genus PCR assay and
Legionella pneumophila
species-specific PCR assay were designed. A total of 43 archived specimens from 35 patients were evaluated, including 19 bronchoalveolar lavage (BAL) specimens and 24 formalin-fixed, paraffin-embedded open lung biopsy specimens. Twenty-five of the specimens were culture-positive for Legionella (9 BAL specimens and 16 tissue specimens). BAL specimens were tested by LightCycler PCR (LC-PCR) methods and by a direct fluorescent antibody (DFA) assay, which detects
L. pneumophila
serogroups 1 to 6 and several other
Legionella
species. Tissue sections were tested by the two LC-PCR methods, by DFA, by an in situ hybridization (ISH) assay, specifically designed to detect
L. pneumophila
, and by Warthin-Starry (WS) staining. The results were compared to the “gold standard” method of bacterial culture. With BAL specimens the following assays yielded the indicated sensitivities and specificities, respectively:
Legionella
genus detection by
Legionella
genus LC-PCR, 100 and 100%;
Legionella
genus detection by DFA assay, 33 and 100%; and
L. pneumophila
detection by
L. pneumophila
species-specific LC-PCR, 100 and 100%. With open lung biopsy specimens the following assays yielded the indicated sensitivities and specificities, respectively:
Legionella
genus detection by LC-PCR 68.8 and 100%;
Legionella
genus detection by DFA assay, 44 and 100%;
Legionella
genus detection by WS staining, 63 and 100%;
L. pneumophila
species-specific detection by LC-PCR, 17 and 100%; and
L. pneumophila
species-specific detection by ISH, 100 and 100%. The analytical sensitivity of both LC-PCR assays was <10 CFU/reaction. LC-PCR is a reliable method for the direct detection of
Legionella
species from BAL specimens. The
Legionella
genus LC-PCR assay could be performed initially; if positive,
L. pneumophila
species-specific LC-PCR could then be performed (if species differentiation is desired). The speed with which the LC-PCR procedure can be performed offers significant advantages over both culture-based methods and conventional PCR techniques. In contrast, for the methods evaluated, culture was the best for detecting multiple
Legionella
species in lung tissue. WS staining,
Legionella
genus LC-PCR, and
L. pneumophila
species-specific ISH were useful as rapid tests with lung tissue.
Malaria Rapid Diagnostic Devices: Performance Characteristics of the
Para
Sight F Device Determined in a Multisite Field Study
Authors: Forney J. Russ , Magill Alan J. , Wongsrichanalai Chansuda , Sirichaisinthop Jeeraphat , Bautista Christian T. , Heppner D. Gray , Miller R. Scott , Ockenhouse Christian F. , Gubanov Alex , Shafer Robyn , DeWitt Caroline Cady , Quino-Ascurra Higinio A. , Kester Kent E. , Kain Kevin C. , Walsh Douglas S. , Ballou W. Ripley , Gasser Robert A. 
Published: July 2002

ABSTRACT

Microscopic detection of parasites has been the reference standard for malaria diagnosis for decades. However, difficulty in maintaining required technical skills and infrastructure has spurred the development of several nonmicroscopic malaria rapid diagnostic devices based on the detection of malaria parasite antigen in whole blood. The
Para
Sight F test is one such device. It detects the presence of
Plasmodium falciparum
-specific histidine-rich protein 2 by using an antigen-capture immunochromatographic strip format. The present study was conducted at outpatient malaria clinics in Iquitos, Peru, and Maesod, Thailand. Duplicate, blinded, expert microscopy was employed as the reference standard for evaluating device performance. Of 2,988 eligible patients, microscopy showed that 547 (18%) had
P. falciparum
, 658 (22%) had
P. vivax
, 2 (0.07%) had
P. malariae
, and 1,750 (59%) were negative for
Plasmodium
. Mixed infections (
P. falciparum
and
P. vivax
) were identified in 31 patients (1%). The overall sensitivity of
Para
Sight F for
P. falciparum
was 95%. When stratified by magnitude of parasitemia (no. of asexual parasites per microliter of whole blood), sensitivities were 83% (>0 to 500 parasites/μl), 87% (501 to 1,000/μl), 98% (1,001 to 5,000/μl), and 98% (>5,000/μl). Device specificity was 86%.
Cariogenic Actinomyces Identified with a β-Glucosidase-Dependent Green Color Reaction toGardenia jasminoidesExtract
Authors: Chen Li , Ma Lili , Park No-Hee , Shi Wenyuan 
Published: July 2002

ABSTRACTThe oral bacteriaActinomyces naeslundiiandActinomyces viscosusare known to contribute to the initiation and progression of human dental caries, especially root caries. We report that bothA. naeslundiiandA. viscosusreact with a component in theGardenia jasminoidesextract to produce a distinct green product. This green color reaction was found to be dependent on the bacterial β-glucosidase. The reaction is specific for cariogenic actinomyces, and it can detect as few as 104cells ofA. naeslundiiandA. viscosusper ml.
Enzyme-Linked Immunosorbent Assay Specific to Dengue Virus Type 1 Nonstructural Protein NS1 Reveals Circulation of the Antigen in the Blood during the Acute Phase of Disease in Patients Experiencing Primary or Secondary Infections
Authors: Alcon Sophie , Talarmin Antoine , Debruyne Monique , Falconar Andrew , Deubel Vincent , Flamand Marie 
Published: July 2002

ABSTRACT
During flavivirus infection in vitro, nonstructural protein NS1 is released in a host-restricted fashion from infected mammalian cells but not vector-derived insect cells. In order to analyze the biological relevance of NS1 secretion in vivo, we developed a sensitive enzyme-linked immunosorbent assay (ELISA) to detect the protein in the sera of dengue virus-infected patients. The assay was based on serotype 1 NS1-specific mouse and rabbit polyclonal antibody preparations for antigen immunocapture and detection, respectively. With purified dengue virus type 1 NS1 as a protein standard, the sensitivity of our capture ELISA was less than 1 ng/ml. When a panel of patient sera was analyzed, the NS1 antigen was found circulating from the first day after the onset of fever up to day 9, once the clinical phase of the disease is over. The NS1 protein could be detected even when viral RNA was negative in reverse transcriptase-PCR or in the presence of immunoglobulin M antibodies. NS1 circulation levels varied among individuals during the course of the disease, ranging from several nanograms per milliliter to several micrograms per milliliter, and peaked in one case at 50 μg/ml of serum. Interestingly, NS1 concentrations did not differ significantly in serum specimens obtained from patients experiencing primary or secondary dengue virus infections. These findings indicate that NS1 protein detection may allow early diagnosis of infection. Furthermore, NS1 circulation in the bloodstream of patients during the clinical phase of the disease suggests a contribution of the nonstructural protein to dengue virus pathogenesis.
Catheter-Related Bacteremia Due to
Kocuria kristinae
in a Patient with Ovarian Cancer
Authors: Basaglia G. , Carretto E. , Barbarini D. , Moras L. , Scalone S. , Marone P. , De Paoli P. 
Published: July 2002

ABSTRACT

We report on the first case of a catheter-related recurrent bacteremia caused by
Kocuria kristinae
, a gram-positive microorganism belonging to the family
Micrococcaceae
, in a 51-year-old woman with ovarian cancer. This unusual pathogen may cause opportunistic infections in patients with severe underlying diseases.