bluuebunny/crossref_metadata_2025_split huggingface dataset:
Validation of Roche COBAS Amplicor Assay for Detection of
Chlamydia trachomatis
in Rectal and Pharyngeal Specimens by an
omp1
PCR Assay
Authors: Lister N. A. , Tabrizi S. N. , Fairley C. K. , Garland S. 
Published: January 2004

ABSTRACT

Screening guidelines for men who have sex with men (MSM) recommend testing of extragenital sites (pharyngeal and rectal) for gonorrhoea and chlamydia. Testing of specimens from these sites is not validated by most commercial nucleic amplification tests, such as the COBAS Amplicor assay. To investigate the utility of the COBAS Amplicor assay for detection of
Chlamydia trachomatis
in extragenital specimens, this study developed and evaluated confirmatory tests using the
omp1
gene as an alternative target for amplification by PCR. Of anal and throat swabs collected from men in male-only saunas, 52 swabs that tested
C. trachomatis
positive by COBAS Amplicor and 30 swabs that tested as negative were included for confirmatory
omp1
PCR testing. A total of 49 (94%) COBAS Amplicor-positive samples were confirmed by the
omp1
PCR. A substantial proportion of specimens were confirmed by using a nested
omp1
PCR (27%). Not confirmed by any
omp1
PCR were three anal swabs (6%). It is most probable that these samples contained lower bacterial levels that were near or below the detection level of the
omp1
PCR assays. The findings of this study support the confident reporting of
C. trachomatis
detected by COBAS Amplicor in extragenital specimens and support the utility of this assay as a screening test for MSM.
Surveillance Network for Herpes Simplex Virus Resistance to Antiviral Drugs: 3-Year Follow-Up
Authors: Danve-Szatanek C. , Aymard M. , Thouvenot D. , Morfin F. , Agius G. , Bertin I. , Billaudel S. , Chanzy B. , Coste-Burel M. , Finkielsztejn L. , Fleury H. , Hadou T. , Henquell C. , Lafeuille H. , Lafon M. E. , Le Faou A. , Legrand M. C. , Maille L. , Mengelle C. , Morand P. , Morinet F. , Nicand E. , Omar S. , Picard B. , Pozzetto B. , Puel J. , Raoult D. , Scieux C. , Segondy M. , Seigneurin J. M. , Teyssou R. , Zandotti C. 
Published: January 2004

ABSTRACT
Herpes simplex virus (HSV) infections are very common in the general population and among immunocompromised patients. Acyclovir (ACV) is an effective treatment which is widely used. We deemed it essential to conduct a wide and coordinated survey of the emergence of ACV-resistant HSV strains . We have formed a network of 15 virology laboratories which have isolated and identified, between May 1999 and April 2002, HSV type 1 (HSV-1) and HSV-2 strains among hospitalized subjects. The sensitivity of each isolate to ACV was evaluated by a colorimetric test (C. Danve, F. Morfin, D. Thouvenot, and M. Aymard, J. Virol. Methods 105:207-217, 2002). During this study, 3,900 isolated strains among 3,357 patients were collected; 55% of the patients were immunocompetent. Only six immunocompetent patients excreted ACV-resistant HSV strains (0.32%), including one female patient not treated with ACV who was infected primary by an ACV-resistant strain. Among the 54 immunocompromised patients from whom ACV-resistant HSV strains were isolated (3.5%), the bone marrow transplantation patients showed the highest prevalence of resistance (10.9%), whereas among patients infected by human immunodeficiency virus, the prevalence was 4.2%. In 38% of the cases, the patients who excreted the ACV-resistant strains were treated with foscarnet (PFA), and 61% of them developed resistance to PFA. The collection of a large number of isolates enabled an evaluation of the prevalence of resistance of HSV strains to antiviral drugs to be made. This prevalence has remained stable over the last 10 years, as much among immunocompetent patients as among immunocompromised patients.
Serotype 14 Variants of the France 9V
−3
Clone from Baltimore, Maryland, Can Be Differentiated by the
cpsB
Gene
Authors: McEllistrem M. Catherine , Noller Anna C. , Visweswaran Shyam , Adams Jennifer M. , Harrison Lee H. 
Published: January 2004

ABSTRACT

European serotype 14 variants of the France 9V
−3
clone, which have arisen through recombination events involving the penicillin binding protein 1a (
pbp1a
) gene, have
cpsB
sequences distinct from those of the 9V
−3
clone. Serotype 14 variants of the 9V
−3
clone have not been compared to genetically diverse serotype 14 strains isolated from an entire metropolitan area in the United States. All serotype 14 non-penicillin-susceptible
Streptococcus pneumoniae
strains causing invasive disease in Baltimore, Md., from 1995 to 1996 were compared by using pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST),
pbp1a
PCR restriction profiles, and
cpsB
and
pbp1a
sequences. The
cpsB
genes from strains of 13 serotypes also were analyzed to assess the correlation with serotype. Twenty-seven percent (3 of 11) of the serotype 14 strains were related by PFGE and MLST to the 9V
−3
clone. The serotype 14 variants from Baltimore, unlike the European variants, were related neither to the 9V
−3
clone nor to the R6 strain from positions 1498 to 1710 of the
pbp1a
gene. All serotype 14 strains had
cpsB
sequences that differed by ≤1% (0 to 5 of 476 bp) from each other and that were ≥16% (78 to 83 of 476 bp) divergent from that of the 9V
−3
clone. Allowing for a 2-bp difference in the
cpsB
sequence resulted in the highest correlation between the
cpsB
gene and serotype. Overall, 95% (84 of 88) of the strains were classified correctly by serotype with the
cpsB
sequence. The distal recombination site of the Baltimore serotype 14 variants of the 9V
−3
clone was not identical to that of the European serotype 14 variants. The
cpsB
gene was serotype specific regardless of whether capsular switching occurred. Although the correlation between serotype and the
cpsB
sequence was high, the overall diversity of the
cpsB
gene within a serotype likely will limit the role of this gene in a sequence-based serotyping method.
Real-Time Reverse Transcription Loop-Mediated Isothermal Amplification for Rapid Detection of West Nile Virus
Authors: Parida Manmohan , Posadas Guillermo , Inoue Shingo , Hasebe Futoshi , Morita Kouichi 
Published: January 2004

ABSTRACT

A one-step, single tube, real-time accelerated reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for detecting the envelope gene of West Nile (WN) virus. The RT-LAMP assay is a novel method of gene amplification that amplifies nucleic acid with high specificity, efficiency, and rapidity under isothermal conditions with a set of six specially designed primers that recognize eight distinct sequences of the target. The whole procedure is very simple and rapid, and amplification can be obtained in less than 1 h by incubating all of the reagents in a single tube with reverse transcriptase and
Bst
DNA polymerase at 63°C. Detection of gene amplification could be accomplished by agarose gel electrophoresis, as well as by real-time monitoring in an inexpensive turbidimeter. When the sensitivity of the RT-LAMP assay was compared to that of conventional RT-PCR, it was found that the RT-LAMP assay demonstrated 10-fold higher sensitivity compared to RT-PCR, with a detection limit of 0.1 PFU of virus. By using real-time monitoring, 10
4
PFU of virus could be detected in as little as 17 min. The specificity of the RT-LAMP assay was validated by the absence of any cross-reaction with other, closely related, members of the
Flavivirus
group, followed by restriction digestion and nucleotide sequencing of the amplified product. These results indicate that the RT-LAMP assay is extremely rapid, cost-effective, highly sensitive, and specific and has potential usefulness for rapid, comprehensive WN virus surveillance along with virus isolation and/or serology.
Prevalence of
Bartonella henselae
in Italian Stray Cats: Evaluation of Serology To Assess the Risk of Transmission of
Bartonella
to Humans
Authors: Fabbi Massimo , De Giuli Luciana , Tranquillo Massimo , Bragoni Roldano , Casiraghi Maurizio , Genchi Claudio 
Published: January 2004

ABSTRACT

Bartonella henselae
is the major etiological agent of cat scratch disease in humans. Cats act as the natural reservoir of
B. henselae
and can transmit the infection to humans by a bite or scratch. The prevalence of
B. henselae
in cat populations was evaluated by serological and bacteriological tests. A total of 769 stray cats from three urban and three rural areas in northern Italy were sampled between January 1999 and December 2000. The positive and the negative predictive values of serological tests with respect to bacteremic status were evaluated. Tests of a total of 140 cats (18%) resulted in detection of bacteremia. A total of 540 cats were tested by serology; 207 (38%) were seropositive. Of the 531 cats tested by both methods, the results for 65 (12.2%) showed both bacteremia detection and seropositivity. The molecular typing of the isolates showed that 20.6% of bacteremic cats were infected with
B. henselae
type I strain, 61.1% were infected with
B. henselae
type II, and 18.3% were coinfected with both. A statistically significant difference in antibody and bacteremia prevalences among geographical areas was detected. Statistical analysis showed no association between characteristics such as seroprevalence-bacteremic status, sex, general health status, and the presence of ectoparasites. The negative predictive value of serological test was 84.7%, and the positive predictive value was 31.8%. Receiving operator characteristic analysis of the data showed that serological tests had a low predictive value in relation to the bacteremic status of a cat; in surveys aimed at assessing the real risk of
B. henselae
infection in a human population, therefore, we suggest the use of blood culture as the reference test. Nevertheless, both blood culture assays and serological tests for
Bartonella
infection should be performed for a complete evaluation of the health status of cats.
Effects of Inoculum and β-Lactamase Activity in AmpC- and Extended-Spectrum β-Lactamase (ESBL)-Producing
Escherichia coli
and
Klebsiella pneumoniae
Clinical Isolates Tested by Using NCCLS ESBL Methodology
Authors: Queenan Anne Marie , Foleno Barbara , Gownley Colleen , Wira Ellyn , Bush Karen 
Published: January 2004

ABSTRACT

Escherichia coli
and
Klebsiella pneumoniae
isolates with extended-spectrum β-lactamases (ESBLs) or AmpC cephalosporinases generally respond as predicted to NCCLS tests for ESBL production. However, inoculum size may affect MICs. The effect of inoculum level in clinical isolates expressing β-lactamases were studied at inocula within 0.5 log unit of the standard inoculum, using broth microdilution methodology with ceftazidime, cefotaxime, cefepime, cefpodoxime, and aztreonam. Strains with TEM-1 or no β-lactamases gave consistent MIC results with inocula of 10
5
and 10
6
CFU/ml. When the bacteria were screened for ESBL production and the lower inoculum was used, several strains with ESBLs, including CTX-M-10, TEM-3, TEM-10, TEM-12, TEM-6, SHV-18, and K1, gave false-negative results for one or more antimicrobial agents (MICs below the NCCLS screening concentration for detecting suspected ESBLs). When the higher inoculum was used, MICs of at least one antimicrobial agent increased at least fourfold in strains producing TEM-3, TEM-10, TEM-28, TEM-43, SHV-5, SHV-18, and K1. All antimicrobial agents showed an inoculum effect with at least one ESBL producer. Confirmatory clavulanate effects were seen for both inocula for all ESBL-producing strains with all antimicrobial agents tested, except for the CTX-M-10-producing
E. coli
with ceftazidime and the SHV-18-producing
K. pneumoniae
with cefotaxime. In kinetic studies, cefpodoxime and cefepime were hydrolyzed by ESBLs in a manner similar to that of cefotaxime. When total β-lactamase activity and hydrolysis parameters were evaluated, however, no single factor was predictive of inoculum effects. These results indicate that the NCCLS screening and confirmation tests are generally predictive of ESBL production, but false-negative results can arise when a lower inoculum is used in testing.
Comparative Analysis of Multilocus Sequence Typing and Pulsed-Field Gel Electrophoresis for Characterizing
Listeria monocytogenes
Strains Isolated from Environmental and Clinical Sources
Authors: Revazishvili Tamara , Kotetishvili Mamuka , Stine O. Colin , Kreger Arnold S. , Morris J. Glenn , Sulakvelidze Alexander 
Published: January 2004

ABSTRACT

One hundred seventy-five
Listeria monocytogenes
strains were characterized by serotyping, pulsed-field gel electrophoresis (PFGE), and multilocus sequence typing (MLST) based on loci in
actA
,
betL
,
hlyA
,
gyrB
,
pgm
, and
recA.
One hundred twenty-two sequence types (STs) were identified by MLST based on allelic profiles of the four housekeeping genes (
betL
,
gyrB
,
pgm
, and
recA
), and 34 and 38 alleles were identified for
hlyA
and
actA
, respectively. Several
actA
and
hlyA
alleles appeared to be predominantly associated with clinical isolates. MLST differentiated most of the
L. monocytogenes
strains better than did PFGE, and the discriminating ability of PFGE was better than that of serotyping. Several strains with different serotypes were found, by MLST and PFGE, to have very closely related genetic backgrounds, which suggested possible “antigen switching” among them. MLST can be a useful typing tool for differentiating
L. monocytogenes
strains (including strains undistinguishable by PFGE typing and serotyping), and it may be of value during investigations of food-borne outbreaks of listeriosis.
Assessment of Agreement between the AMPLICOR HIV-1 MONITOR Test Versions 1.0 and 1.5
Authors: Hill Charles E. , Green Alicia M. , Ingersoll Jessica , Easley Kirk A. , Nolte Frederick S. , Caliendo Angela M. 
Published: January 2004

ABSTRACT

The agreement of the microwell plate AMPLICOR HIV-1 MONITOR version 1.0 (MWP 1.0), the microwell plate AMPLICOR HIV-1 MONITOR version 1.5 (MWP 1.5), and the COBAS AMPLICOR HIV-1 MONITOR version 1.5 (COBAS 1.5) tests was evaluated using clinical specimens and well-characterized control material. Two hundred patient plasma specimens and a panel of known human immunodeficiency virus type 1 (HIV-1) subtypes were tested. All data were log
10
transformed prior to analysis. The 95% limits of agreement for the three tests at the average of 3.66 log
10
copies/ml were ± 0.28 log
10
, ± 0.34 log
10
, and ± 0.34 log
10
copies/ml for MWP 1.0-MWP 1.5, MWP 1.0-COBAS 1.5, and MWP 1.5-COBAS 1.5, respectively. Ten specimens (6.1%) had differences exceeding the limits of agreement for the MWP 1.0 and MWP 1.5 tests. Correlation coefficients among the three tests were high (
r
≥ 0.96). The viral-load values obtained with the MWP 1.0 test were only 2.1% higher on average than those measured with the MWP 1.5 test and 1.6% higher than those seen with the COBAS 1.5 test. The MWP 1.5 test values were 0.8% higher than the COBAS 1.5 test values. Overall, there was less agreement among the different tests for viral-load values near the lower limit of quantification. The MWP 1.0 test underquantified subtypes A, E, F, G, and H by 1.0 to 2.0 log
10
copies/ml; this problem was not observed with the MWP 1.5 test. The close agreement among the results obtained with the different test versions and formats suggests that it is not necessary to reestablish a baseline viral load when changing AMPLICOR HIV-1 MONITOR tests, unless the patient is known to be infected with a non-B subtype.
Isolation of a Fastidious
Bergeyella
Species Associated with Cellulitis after a Cat Bite and a Phylogenetic Comparison with
Bergeyella zoohelcum
Strains
Authors: Shukla Sanjay K. , Paustian David L. , Stockwell Patrick J. , Morey Roger E. , Jordan Jean G. , Levett Paul N. , Frank Daniel N. , Reed Kurt D. 
Published: January 2004

ABSTRACT

Bergeyella zoohelcum
is an uncommon zoonotic pathogen typically associated with cat or dog bites. Previously, only five cases of
B. zoohelcum
infection have been reported. We report the isolation and characterization of a fastidious
Bergeyella
species from acute cellulitis in the upper extremity of a 60-year-old woman. The organism was too fastidious for identification and susceptibility testing with traditional culture methods. The isolate was characterized further by PCR amplification and sequencing of the 16S rRNA gene with broad-range eubacterial primers. Phylogenetic analysis of the 16S ribosomal DNA sequence indicated that this isolate was a member of the species
B. zoohelcum
(previously
Weeksella zoohelcum
), a gram-negative bacillus that is rarely associated with infections in humans. Despite sharing a close genetic relationship with other
B. zoohelcum
strains, this isolate was extremely fastidious in nature, raising the possibility that similar strains from cat or dog bite wound infections have been underreported.
Utility of NCCLS Guidelines for Identifying Extended-Spectrum β-Lactamases in Non-
Escherichia coli
and Non-
Klebsiella
spp. of
Enterobacteriaceae
Authors: Schwaber Mitchell J. , Raney Patti M. , Rasheed J. Kamile , Biddle James W. , Williams Portia , McGowan John E. , Tenover Fred C. 
Published: January 2004

ABSTRACT

NCCLS screening and confirmation methods for detecting extended-spectrum β-lactamases (ESBLs) apply only to
Escherichia coli
and
Klebsiella
spp., yet ESBLs have been found in other members of the family
Enterobacteriaceae
. We evaluated the effectiveness of NCCLS methods for detecting ESBLs in 690 gram-negative isolates of
Enterobacteriaceae
that excluded
E. coli
,
Klebsiella pneumoniae
, and
Klebsiella oxytoca.
Isolates were collected between January 1996 and June 1999 from 53 U.S. hospitals participating in Project ICARE (Intensive Care Antimicrobial Resistance Epidemiology). The antimicrobial susceptibility patterns of the isolates were determined by using the NCCLS broth microdilution method (BMD), and those isolates for which the MIC of ceftazidime, cefotaxime, ceftriaxone, or aztreonam was ≥2 μg/ml or the MIC of cefpodoxime was ≥8 μg/ml (positive ESBL screen test) were further tested for a clavulanic acid (CA) effect by BMD and the disk diffusion method (confirmation tests). Although 355 (51.4%) of the isolates were ESBL screen test positive, only 15 (2.2%) showed a CA effect. Since 3 of the 15 isolates were already highly resistant to the five NCCLS indicator drugs, ESBL detection would have an impact on the reporting of only 1.7% of the isolates in the study. Only 6 of the 15 isolates that showed a CA effect contained a
bla
TEM
,
bla
SHV
,
bla
CTX-M
, or
bla
OXA
β-lactamase gene as determined by PCR (with a corresponding isoelectric focusing pattern). Extension of the NCCLS guidelines for ESBL detection to
Enterobacteriaceae
other than
E. coli
and
Klebsiella
spp. does not appear to be warranted in the United States at present, since the test has poor specificity for this population and would result in changes in categorical interpretations for only 1.7% of
Enterobacteriaceae
tested.
Evaluation of a TaqMan PCR Assay To Detect Rabies Virus RNA: Influence of Sequence Variation and Application to Quantification of Viral Loads
Authors: Hughes G. J. , Smith J. S. , Hanlon C. A. , Rupprecht C. E. 
Published: January 2004

ABSTRACT

Published assays that use TaqMan PCR are consistently sensitive, rapid, and readily transferable. Here we describe a TaqMan PCR-based method for the detection of rabies virus (RV) RNA in tissue samples. We show that the method has an acceptable linear range, is both sensitive and specific, and, importantly, correlates with the concentration of infectious virus. In addition, the levels of RV-specific amplification are adjustable according to the levels of an endogenous control (β-actin mRNA), allowing the calculation of comparable quantities. We tested the capacity of this assay to cope with target sequence variations. The number of sequence mismatches between gene-specific oligonucleotides and the target sequence significantly affects amplification (
P
< 0.001), and point mutations at the center of the probe can result in false-negative results through the prevention of probe binding and subsequent fluorescence. This study demonstrates that the genetic heterogeneity of RVs may prove a serious obstacle in the development of a diagnostic assay based on TaqMan PCR; however, the quantification of RV levels may prove to be a valuable application of this assay.
Clonal and Horizontal Dissemination of
Klebsiella pneumoniae
Expressing SHV-5 Extended-Spectrum β-Lactamase in a Mexican Pediatric Hospital
Authors: Miranda Guadalupe , Castro Natividad , Leaños Blanca , Valenzuela Adriana , Garza-Ramos Ulises , Rojas Teresa , Solórzano Fortino , Chihu Lilia , Silva Jesús 
Published: January 2004

ABSTRACT

One hundred eighty-four clinical isolates of
Klebsiella pneumoniae
were recovered from August 1996 to October 1997 at the Pediatric Hospital of the Instituto Mexicano del Seguro Social in Mexico City, Mexico. Most of the isolates were collected from the neonatal intensive care unit and infant wards, which are located on the same floor of the hospital. Isolates were genotypically compared by pulsed-field gel electrophoresis with
Xba
I restriction of chromosomal DNA. Of 184 clinical isolates, 91 belonged to cluster A and comprised three subtypes (A1, A2, and A3), while 93 isolates, comprising two minor clones, B (10 isolates) and C (7 isolates), and 76 unique patterns, were considered unrelated isolates (URI). Susceptibility patterns were indistinguishable in both groups. Fifty extended-spectrum β-lactamase-producing isolates, including 34 from clone A and 16 from URI, were examined for further studies. Molecular and genetic analysis showed that 47 of 50 clinical isolates expressed the SHV-5 β-lactamase. This enzyme, in combination with TEM-1, was encoded in a ≥170-kb conjugative plasmid. Results indicate that dissemination of this resistance was due to clonal and horizontal spread.
Molecular Epidemiology of Clinical Isolates of Methicillin-Resistant
Staphylococcus aureus
in Taiwan
Authors: Huang Yhu-Chering , Su Lin-Hui , Wu Tsu-Lan , Liu Chun-Eng , Young Tzuu-Guang , Chen Po-Yen , Hseuh Po-Ren , Lin Tzou-Yien 
Published: January 2004

ABSTRACT

During July 2000 and October 2001, a total of 595 clinical isolates of methicillin-resistant
Staphylococcus aureus
(MRSA) were collected from six medical centers distributed in northern, central, and southern Taiwan. Specimen sources included blood (
n
= 279), pus (
n
= 173), sputum (
n
= 94), body fluids (
n
= 21), catheter tips (
n
= 20), and urine (
n
= 8). Pulsed-field gel electrophoresis (PFGE) with
Sma
I digestion was used to fingerprint these isolates. A total of 31 genotypes with 97 type-subtypes were identified. Subtypes could be identified in 7 genotypes. While there were 6 to 15 genotypes in each hospital, 433 isolates (73%) were shown to belong to a major type (genotype A, with 29 subtypes). This genotype was not only the type prevailing in all six hospitals but also the predominant clone in each hospital, accounting for 46 to 89% of all isolates in each hospital. Genotype C (16 subtypes) was the second dominant genotype, accounting for 9% of all isolates, and was distributed in five hospitals. Genotypes D (11 subtypes), E (5 subtypes), and B (6 subtypes) were distributed in five, four, and three hospitals, respectively. The other 26 types (30 type-subtypes) were minor. We conclude that the majority of MRSA clinical isolates shared a common PFGE pattern, indicating the presence of a single, epidemic MRSA clone prevailing in major hospitals in Taiwan.