bluuebunny/crossref_metadata_2025_split huggingface dataset:
Evaluation of the COBAS Amplicor HBV Monitor Assay and Comparison with the Ultrasensitive HBV Hybrid Capture 2 Assay for Quantification of Hepatitis B Virus DNA
Authors: Konnick Eric Q. , Erali Maria , Ashwood Edward R. , Hillyard David R. 
Published: February 2005

ABSTRACTPerformance characteristics of the COBAS Amplicor HBV Monitor test (Roche Diagnostics), which measures hepatitis B virus (HBV) DNA quantitatively, were evaluated and compared with the Ultrasensitive HBV Hybrid Capture 2 (HC2; Digene Corporation) assay. Linearity and within-run precision were assessed for both methods by using eight HBV DNA-positive samples serially diluted to obtain a range of <100 to 500,000 HBV DNA copies/ml and run in triplicate. Agreement between the methods was studied with 100 clinical samples. HC2 assay performance near the limit of detection was investigated through repeat testing of 149 samples with HC2 and testing of 37 samples with HC2 results of <4,700 HBV DNA copies/ml by Amplicor assay and a qualitative PCR assay. The linearity experiment for Amplicor had regression of observed values compared to expected values (y= 1.073x− 0.247;R2= 0.993,n= 32; for HC2,y= 0.855x+ 0.759,R2= 0.729,n= 18). Within-run standard deviation of log HBV DNA copies/ml ranged from 0.003 to 0.348 (Amplicor) and 0.027 to 0.253 (HC2). Agreement assessed by Deming regression was poor [Amplicor = 1.197(HC2) − 0.961;R2= 0.799, standard error of the estimate (SEE) = 0.710,n= 94]. Near the lower limit of detection, 32 of 149 repeat HC2 results were <4,700 HBV DNA copies/ml. Of the 37 samples with HC2 results of <4,700 HBV DNA copies/ml, HBV DNA was not detected in 15 samples, while HBV DNA was detected by at least one PCR method in 12 samples. Amplicor is linear from 200 to 200,000 HBV DNA copies/ml with undiluted samples, and this range can be expanded through dilution. Inconsistent HC2 results near the limit of detection justify use of a grey zone.
Molecular Identification of a Novel Deltaproteobacterium as the Etiologic Agent of Epizootic Bovine Abortion (Foothill Abortion)
Authors: King Donald P. , Chen Ching-I , Blanchard Myra T. , Aldridge Brian M. , Anderson Mark , Walker Richard , Maas John , Hanks Don , Hall Mark , Stott Jeffrey L. 
Published: February 2005

ABSTRACTEpizootic bovine abortion (EBA) is endemic in California's coastal range and the foothill regions of the Sierra Nevada, where it has been the primary diagnosed cause of abortion in beef cattle for >50 years. Investigation of these losses has defined a specific fetal syndrome characterized by late-term abortion or birth of weak or dead calves. Although the unusual clinical presentation and unique fetal pathology associated with EBA have been recognized since the 1950s, the identity of the etiologic agent is unknown. In this study, suppression-hybridization PCR was used to identify a fragment of the 16S rRNA gene of a previously undescribed bacterium in thymus tissue derived from affected fetuses. Phylogenetic analysis revealed that this pathogen was a deltaproteobacterium closely related to members of the orderMyxococcales. A specific PCR was subsequently developed to detect the presence of this bacterium in DNA extracted from fetal thymuses. Using histopathology as the definitive diagnosis for EBA, this PCR demonstrated 100% specificity and 88% sensitivity. The bacterium was also detected in the argasid tickOrnithodoros coriaceus, which is the recognized vector of EBA. These data imply a close association between this novel agent and the etiology of EBA.
Use of the MicroSeq 500 16S rRNA Gene-Based Sequencing for Identification of Bacterial Isolates That Commercial Automated Systems Failed To Identify Correctly
Authors: Fontana Carla , Favaro Marco , Pelliccioni Marco , Pistoia Enrico Salvatore , Favalli Cartesio 
Published: February 2005

ABSTRACT
Reliable automated identification and susceptibility testing of clinically relevant bacteria is an essential routine for microbiology laboratories, thus improving patient care. Examples of automated identification systems include the Phoenix (Becton Dickinson) and the VITEK 2 (bioMérieux). However, more and more frequently, microbiologists must isolate “difficult” strains that automated systems often fail to identify. An alternative approach could be the genetic identification of isolates; this is based on 16S rRNA gene sequencing and analysis. The aim of the present study was to evaluate the possible use of MicroSeq 500 (Applera) for sequencing the 16S rRNA gene to identify isolates whose identification is unobtainable by conventional systems. We analyzed 83 “difficult” clinical isolates: 25 gram-positive and 58 gram-negative strains that were contemporaneously identified by both systems—VITEK 2 and Phoenix—while genetic identification was performed by using the MicroSeq 500 system. The results showed that phenotypic identifications by VITEK 2 and Phoenix were remarkably similar: 74% for gram-negative strains (43 of 58) and 80% for gram-positive strains were concordant by both systems and also concordant with genetic characterization. The exceptions were the 15 gram-negative and 9 gram-positive isolates whose phenotypic identifications were contrasting or inconclusive. For these, the use of MicroSeq 500 was fundamental to achieving species identification. In clinical microbiology the use of MicroSeq 500, particularly for strains with ambiguous biochemical profiles (including slow-growing strains), identifies strains more easily than do conventional systems. Moreover, MicroSeq 500 is easy to use and cost-effective, making it applicable also in the clinical laboratory.
Development of Colorimetric Microtiter Plate Assay for Assessment of Antimicrobials against
Acanthamoeba
Authors: McBride James , Ingram Paul R. , Henriquez Fiona L. , Roberts Craig W. 
Published: February 2005

ABSTRACT

We have developed and optimized a 96-well microtiter plate assay, based on the reduction of alamarBlue, to assess the efficacies of much needed new antimicrobials against
Acanthamoeba
species. This assay has been optimized for determination of drug efficacy against two potentially pathogenic species,
Acanthamoeba castellanii
and
Acanthamoeba polyphaga
, and has been validated by comparison of their relative susceptibilities to chlorhexidine, a drug widely used to treat
Acanthamoeba
keratitis. The results demonstrate that the assay is comparable to a manual counting assay and that
A. polyphaga
is more resistant to chlorhexidine than
A. castellanii
. Thus, by use of the manual counting assay, 3.125 μM chlorohexidine was almost completely effective against
A. castellanii
, whereas this concentration was less than 20% effective against
A. polyphaga
. Similar results were obtained by the alamarBlue assay. The new assay was used to determine the relative susceptibilities of
A. castellanii
and
A. polyphaga
to the alkylphosphocholines (APCs) hexadecylphosphocholine (hexadecyl-PC; miltefosine) and octadecylphosphocholine (octadecyl-PC) as well as an alkylgycerolphosphocholine, edelfosine. Both APCs studied were equally effective against
A. castellanii
, but octadecyl-PC was less effective than hexadecyl-PC against
A. polyphaga
. Both APCs were more effective than edelfosine against both
Acanthamoeba
species.
A. polyphaga
was found to be significantly less susceptible to each of the phosphocholine analogues. The newly described assay offers a number of advantages over those described previously. It is less labor-intensive than previously described assays and is sensitive and rapid, and the results can be read in a nonsubjective manner. As it is based on a standard 96-well, microtiter plate, it is amenable to automation and high throughput.
Characterization of Cytolethal Distending Toxin ofCampylobacterSpecies Isolated from Captive Macaque Monkeys
Authors: Dassanayake Rohana P. , Zhou You , Hinkley Susanne , Stryker Cynthia J. , Plauche Gail , Borda Juan T. , Sestak Karol , Duhamel Gerald E. 
Published: February 2005

ABSTRACTAn association between certainCampylobacterspecies and enterocolitis in humans and nonhuman primates is well established, but the association between cytolethal distending toxin and disease is incompletely understood. The purpose of the present study was to examineCampylobacterspecies isolated from captive conventionally raised macaque monkeys for the presence of thecdtBgene and for cytolethal distending toxin activity. The identity of each isolate was confirmed on the basis of phenotypic and genotypic analyses. The presence of cytolethal distending toxin was confirmed on the basis of characteristic morphological changes in HeLa cells incubated with filter-sterilized whole-cell lysates of reference and monkeyCampylobacterisolates and examinations by light microscopy, confocal microscopy, and flow cytometry. AlthoughcdtBgene sequences were found in bothCampylobacter jejuniandCampylobacter coli, the production of cytolethal distending toxin correlated positively (P< 0.0001) only withC. jejuni. We concluded that cytolethal distending toxin activity is a characteristic ofC. jejuni. OurC. jejuni cdtBgene-specific PCR assay might be of assistance for differentiating toxigenicC. jejunifromC. coliin clinical laboratories.
Seroprevalence of Noroviruses in Swine
Authors: Farkas Tibor , Nakajima Setsuko , Sugieda Masaaki , Deng Xiaoyun , Zhong Weiming , Jiang Xi 
Published: February 2005

ABSTRACT
Noroviruses (NVs) are important human pathogens that cause acute gastroenteritis. Genetically related animal enteric NVs have also been described, but there is no evidence of interspecies transmission of NVs. In this study we characterized antibody prevalence among domestic pigs by using recombinant capsid antigens of two human NVs (Norwalk and Hawaii) and one swine NV (SW918) that is genetically related to GII human NVs. Recombinant SW918 capsid protein expressed in baculovirus self-assembled into virus-like particles (VLPs) that were detected by antibodies against GII (Hawaii and Mexico), but not GI (Norwalk and VA115), human NVs. NVs recognize human histo-blood group antigens as receptors, but SW918 VLPs did not bind to human saliva samples with major histo-blood group types. Seventy-eight of 110 (71%) pig serum samples from the United States and 95 of 266 (36%) pig serum samples from Japan possessed antibodies against SW918. Serum samples from pigs in the United States were also tested for antibodies against human NVs; 63% were positive for Norwalk virus (GI) and 52% for Hawaii virus (GII). These results indicate that NV infections are common among domestic pigs; the finding of antigenic relationships between SW918 and human NVs and the detection of antibodies against both GI and GII human NVs in domestic animals highlights the importance of further studies on NV gastroenteritis as a possible zoonotic disease.
Use of an Immunochromatographic Assay for Rapid Detection of
Trichomonas vaginalis
in Vaginal Specimens
Authors: Huppert Jill S. , Batteiger Byron E. , Braslins Phillip , Feldman James A. , Hobbs Marcia M. , Sankey Heather Z. , Sena Arlene C. , Wendel Karen A. 
Published: February 2005

ABSTRACT

Trichomonas vaginalis
infection is estimated to be the most widely prevalent nonviral sexually transmitted infection in the world. Wet-mount microscopy is the most common diagnostic method, although it is less sensitive than culture. The OSOM Trichomonas Rapid Test (Genzyme Diagnostics, Cambridge, Mass.) (referred to here as OSOM) is a new point-of-care diagnostic assay for
T. vaginalis
that uses an immunochromatographic capillary flow (dipstick) assay and provides results in 10 min. The purpose of this study was to determine the test characteristics of OSOM compared to those of a composite reference standard (CRS) comprised of wet-mount microscopy and
T. vaginalis
culture. This multicenter cross-sectional study enrolled sexually active women ≥18 years of age who presented with symptoms of vaginitis, exposure to
T. vaginalis
, or multiple sexual partners. Vaginal-swab specimens were obtained for
T. vaginalis
culture, wet mount, and rapid testing. The prevalence of
T. vaginalis
in this sample was 23.4% (105 of 449) by the CRS. The sensitivity and specificity of OSOM vaginal-swab specimens were 83.3 and 98.8%, respectively, while wet mount had a sensitivity and specificity of 71.4 and 100%, respectively, compared to the CRS. OSOM performed significantly better than wet mount (
P
= 0.004) and detected
T. vaginalis
in samples that required 48 to 72 h of incubation prior to becoming culture positive. The performance of the rapid test was not affected by the presence of coinfections with chlamydia and gonorrhea. The OSOM Trichomonas Rapid Test is a simple, objective test that can be expected to improve the diagnosis of
T. vaginalis
, especially where microscopy and culture are unavailable.
Evaluation of a Two-Step Approach for Large-Scale, Prospective Genotyping ofMycobacterium tuberculosisIsolates in the United States
Authors: Cowan Lauren S. , Diem Lois , Monson Timothy , Wand Philip , Temporado David , Oemig Tanya V. , Crawford Jack T. 
Published: February 2005

ABSTRACTGenotyping ofMycobacterium tuberculosisisolates is useful in tuberculosis control for confirming suspected transmission links, identifying unsuspected transmission, and detecting or confirming possible false-positive cultures. The value is greatly increased by reducing the turnaround time from positive culture to genotyping result and by increasing the proportion of cases for which results are available. Although IS6110fingerprinting provides the highest discrimination, amplification-based methods allow rapid, high-throughput processing and yield digital results that can be readily analyzed and thus are better suited for large-scale genotyping.M. tuberculosisisolates (n= 259) representing 99% of culture-positive cases of tuberculosis diagnosed in Wisconsin in the years 2000 to 2003 were genotyped by using spoligotyping, mycobacterial interspersed repetitive unit (MIRU) typing, and IS6110fingerprinting. Spoligotyping clustered 64.1% of the isolates, MIRU typing clustered 46.7% of the isolates, and IS6110fingerprinting clustered 29.7% of the isolates. The combination of spoligotyping and MIRU typing yielded 184 unique isolates and 26 clusters containing 75 isolates (29.0%). The addition of IS6110fingerprinting reduced the number of clustered isolates to 30 (11.6%) if an exact pattern match was required or to 44 (17.0%) if the definition of a matching IS6110fingerprint was expanded to include patterns that differed by the addition of a single band. Regardless of the genotyping method chosen, the addition of a second or third method decreased clustering. Our results indicate that using spoligotyping and MIRU typing together provides adequate discrimination in most cases. IS6110fingerprinting can then be used as a secondary typing method to type the clustered isolates when additional discrimination is needed.
Sequence Analysis of the
Escherichia coli
O15 Antigen Gene Cluster and Development of a PCR Assay for Rapid Detection of Intestinal and Extraintestinal Pathogenic
E. coli
O15 Strains
Authors: Beutin Lothar , Tao Jiang , Feng Lu , Krause Gladys , Zimmermann Sonja , Gleier Kerstin , Xia Qiuyu , Wang Lei 
Published: February 2005

ABSTRACT

A collection of 33
Escherichia coli
serogroup O15 strains was studied with regard to O:H serotypes and virulence markers and for detection of the O-antigen-specific genes
wzx
and
wzy
. The strains were from nine different countries, originated from healthy or diseased humans and animals and from food, and were isolated between 1941 and 2003. On the basis of virulence markers and clinical data the strains could be split into different pathogroups, such as uropathogenic
E. coli
, enteropathogenic
E. coli
, Shiga toxin-producing
E. coli
, and enteroaggregative
E. coli
. H serotyping and genotyping of the flagellin (
fliC
) gene revealed 11 different H types and a close association between certain H types, virulence markers, and pathogroups was found. Nucleotide sequence analysis of the O-antigen gene cluster revealed putative genes for biosynthesis of O15 antigen. PCR assays were developed for sensitive and specific detection of the O15-antigen-specific genes
wzx
and
wzy.
The high pathotype diversity found in the collection of 33 O15 strains contrasted with the high level of similarity found in the genes specific to the O15 antigen. This might indicate that the O15 determinant has been spread by horizontal gene transfer to a number of genetically unrelated strains of
E. coli.
Diagnosis of Congenital Toxoplasmosis by Two-Dimensional Immunoblot Differentiation of Mother and Child Immunoglobulin G Profiles
Authors: Nielsen Henrik Vedel , Schmidt Dorte Remmer , Petersen Eskild 
Published: February 2005

ABSTRACT

Differentiation between the specific immunoglobulin G (IgG) response to
Toxoplasma gondii
by a mother and her newborn child is helpful in the diagnosis of congenital infection with
T. gondii
in newborns without
T. gondii
-specific IgM and/or IgA antibodies at birth. Previous methods include immunoblotting and complexing
T. gondii
antigen with the sera from the mother and child and comparing the bands after electrophoresis. We developed a two-dimensional immunoblotting (2DIB) method with
T. gondii
RH strain tachyzoite antigen and validated the method with sera from 11 children identified through the neonatal screening program for congenital toxoplasmosis in Denmark. The children were identified by using
Toxoplasma
-specific IgM antibodies at the screening test, but the presence of
T. gondii
-specific IgM and/or IgA antibodies could not be confirmed at the subsequent serum sample tested. The children were monitored for at least 12 months, and in seven of eight patients monitored for 12 months the results of the 2DIB-predicted congenital infection were confirmed by the presence of persistent
Toxoplasma
-specific IgG antibodies. 2DIB is a sensitive technique that allows early differentiation between passively transferred maternal
T. gondii
-specific IgG antibodies and antibodies synthesized by the newborn child.
Five New Genome Types of Adenovirus Type 37 Caused Epidemic Keratoconjunctivitis in Sapporo, Japan, for More Than 10 Years
Authors: Ariga Toshihide , Shimada Yasushi , Shiratori Kenji , Ohgami Kazuhiro , Yamazaki Shudo , Tagawa Yoshitsugu , Kikuchi Masayuki , Miyakita Yoshie , Fujita Kozo , Ishiko Hiroaki , Aoki Koki , Ohno Shigeaki 
Published: February 2005

ABSTRACT
Human adenovirus type 37 (HAdV-37) is a major cause of epidemic keratoconjunctivitis and has recently been the largest causative agent of keratoconjunctivitis in Japan. To investigate the genetic characteristics of HAdV-37 strains isolated in Sapporo, we analyzed the genome types and genetic relationships of 51 strains isolated there from 1990 through 2001. By using DNA restriction analysis, eight genome types (HAdV-37/D1, HAdV-37/D3, and HAdV-37/D6 to HAdV-37/D11) were identified, including five new ones. The restriction fragments of these genome types shared more than 95% identity with those of the prototype strain. By DNA sequence analysis, five and three single nucleotide substitutions, respectively, were found in partial sequences of the hexon and fiber genes. The combinations of mutations resulted in four hexon and fiber types (hx1 to hx4 and f1 to f4) and six hexon/fiber pairs (hx1/f1, hx2/f1, hx1/f2, hx1/f3, hx3/f4, and hx4/f4). The six pairs correlated well with certain genome types. In all three epidemics of keratoconjunctivitis to strike Sapporo in the past 12 years, specific genome types and fiber types were usually isolated: in the first epidemic, HAdV-37/D1 (f1) and HAdV-37/D3 (f1); in the second, HAdV-37/D6 (f2) and HAdV-37/D8 (f3); and in the third, HAdV-37/D10 (f4) and HAdV-37/D11 (f4). We conclude that mutations in the adenovirus genome occurred chronologically and that certain mutations were correlated with the epidemics of adenoviral keratoconjunctivitis.
Comparison of Hepatitis C Virus NS5b and 5′ Noncoding Gene Sequencing Methods in a Multicenter Study
Authors: Laperche Syria , Lunel Françoise , Izopet Jacques , Alain Sophie , Dény Paul , Duverlie Gilles , Gaudy Catherine , Pawlotsky Jean-Michel , Plantier Jean-Christophe , Pozzetto Bruno , Thibault Vincent , Tosetti François , Lefrère Jean-Jacques 
Published: February 2005

ABSTRACTA national evaluation study was performed in 11 specialized laboratories with the objective of assessing their capacities to genotype hepatitis C virus (HCV) and define the applicability of a given genotyping method. The panel consisted of 14 samples positive for HCV RNA of different genotypes (including 3 samples with two different artificially mixed genotypes) and 1 HCV-negative sample. Seventeen sets of data were gathered from the 11 participating laboratories. The sensitivities ranged from 64.3 to 100% and from 42.7 to 85.7% for the methods that used sequencing of the NS5b region and the 5′ noncoding (5′ NC) region, respectively. When the data for the artificially mixed samples were excluded, NS5b genotyping gave correct results for 80% of the samples, 1.7% of the samples were misclassified, and 18.3% of the samples had false-negative results. By 5′ NC-region genotyping methods, 58.3% of the results were correct, 29.7% were incomplete, 8.3% were misclassifications, 1.2% were false positive, and 2.4% were false negative. Only two procedures based on NS5b sequencing correctly identified one of the three samples with mixtures of genotypes; the other methods identified the genotype corresponding to the strain with the highest viral load in the sample. Our results suggest that HCV 5′ NC-region genotyping methods give sufficient information for clinical purposes, in which the determination of the subtype is not essential, and that NS5b genotyping methods are more reliable for subtype determination, which is required in epidemiological studies.
A Novel
Shigella dysenteriae
Serovar Isolated in Canada
Authors: Melito P. L. , Woodward D. L. , Munro J. , Walsh J. , Foster R. , Tilley P. , Paccagnella A. , Isaac-Renton J. , Ismail J. , Ng L. K. 
Published: February 2005

ABSTRACT

The etiological agent most commonly associated with bacillary dysentery is
Shigella
. As part of its mandate, the Bacteriology and Enteric Disease Program of Health Canada identifies and serotypes unusual isolates of
Shigella
received from provincial laboratories of public health. In this report, six unusual isolates from three provinces were analyzed biochemically and serologically using slide and tube agglutinations and molecularly using standard pulsed-filed gel electrophoresis (PFGE), PCR, and PCR-restriction fragment length polymorphism (RFLP) techniques. All six isolates were identical. PFGE analysis grouped these strains; biochemically, they were mannitol negative and consistent with the profile of
Shigella
. Serologically, these strains produced weak reactions in
Shigella dysenteriae
serovars 4 and 16 and
Escherichia coli
O159 and O173 antisera. Molecular serotyping by PCR-RFLP of the
rfb
gene produced an
S. dysenteriae
serovar 2/
E. coli
O112ac pattern. They were positive by PCR for
ipaH
and
ial
enteroinvasive genes but negative for all other genes tested. Antiserum was prepared from one of the isolates and tested against
Shigella
and
E. coli
reference strains as well as the other isolates. The antiserum reacted with the five remaining isolates and showed cross-reactivity with
S. dysenteriae
serovars 1, 4, and 16;
Shigella flexneri
type 3; and
E. coli
O118, O159, O168, O172, and O173 antigens. Absorbing the sera with
E. coli
O159 and
S. dysenteriae
serovar 4 antigen removed all cross-reactions and only slightly reduced the homologous titer. Based on biochemical, molecular, and complete serological analysis, we propose that these six isolates represent a new provisional serovar of
S. dysenteriae
, type strain BEDP 02-5104.