bluuebunny/crossref_metadata_2025_split huggingface dataset:
An Important Proportion of Genital Samples Submitted for
Chlamydia trachomatis
Detection by PCR Contain Small Amounts of Cellular DNA as Measured by β-Globin Gene Amplification
Authors: Coutlée François , de Ladurantaye Manon , Tremblay Carole , Vincelette Jean , Labrecque Louise , Roger Michel 
Published: December 2019

ABSTRACT

We assessed the quality of genital samples submitted for
Chlamydia trachomatis
detection by PCR by a second PCR assay for the presence of human β-globin DNA. Endocervical and urethral samples were first tested by the COBAS AMPLICOR
C. trachomatis
assay (Roche Diagnostic Systems) with an internal control and were then amplified for the presence of β-globin DNA with primers PC04 and GH20. Samples that contained inhibitors were retested after dilution 1:10. A total of 407 genital samples (311 endocervical swabs from 311 women and 96 urethral swabs from 95 men and 1 woman) collected over a 1-month period were evaluated. The internal control could not be amplified, despite dilution, from 3 of 23 samples that were retested after dilution because of inhibition, leaving 404 samples that could be analyzed by PCR. Eleven samples tested positive for
C. trachomatis
. Thirty (7.4%) of the 404 samples were negative for β-globin. Twelve of the 23 undiluted samples that contained inhibitors tested positive for β-globin DNA. Amplification of β-globin DNA in samples submitted for
C. trachomatis
detection by the COBAS AMPLICOR
C. trachomatis
assay demonstrated that an important proportion of the samples did not contain cellular DNA. Assessment of the quality of the samples for PCR analysis by β-globin amplification is feasible but cannot replace use of the internal control.
A 5′ Nuclease PCR (TaqMan) High-Throughput Assay for Detection of the
mecA
Gene in Staphylococci
Authors: Killgore George E. , Holloway Brian , Tenover Fred C. 
Published: December 2019

ABSTRACT

In an effort to find a rapid, efficient, and reliable method of screening large numbers of bacterial isolates for specific antimicrobial resistance genes, we compared conventional PCR results to the results generated using the TaqMan 5′ nuclease PCR kit in conjunction with an ABI Prism 7700 Sequence Detector for detecting the
mecA
gene in various species of staphylococci. DNA was extracted using two techniques. The first used a high-salt extraction method suitable for conventional PCR but resulted in a 7.2% rate of PCR inhibition with the TaqMan technique. PCR inhibition could be overcome by diluting samples 1:5 prior to testing. The second method used the Qiagen QIAamp Tissue Kit; no instances of PCR inhibition were encountered with this method. A total of 197 (96%) of the 206 samples with no inhibition showed agreement between the two methods. Eight of the nine disagreements were likely the result of low-level DNA cross contamination caused by frequent specimen handling. Target DNA in all eight of these samples was first detected in the initial tests only after >30 PCR cycles, and all were negative upon repeat testing even after 40 PCR cycles using freshly extracted DNA. Among those positive samples in agreement, target DNA was invariably detected before 30 PCR cycles. The TaqMan assay eliminated the need to load, run, stain, and read agarose gels and provided the advantage of instant detection of PCR product by laser-activated fluorescence. Thus, final results were obtained 2 h after PCR was initiated, as opposed to a requirement of 2 days to examine 96 samples by agarose gel electrophoresis.
Real-Time Automated PCR for Early Diagnosis and Monitoring of Cytomegalovirus Infection after Bone Marrow Transplantation
Authors: Machida Utako , Kami Masahiro , Fukui Takafumi , Kazuyama Yukumasa , Kinoshita Moritoshi , Tanaka Yuji , Kanda Yoshinobu , Ogawa Seishi , Honda Hiroaki , Chiba Shigeru , Mitani Kinuko , Muto Yoshitomo , Osumi Kazuoki , Kimura Satoshi , Hirai Hisamaru 
Published: December 2019

ABSTRACT

The purpose of this study was to assess the usefulness of real-time automated PCR as a quantitative, highly reproducible, and sensitive method to detect cytomegalovirus (CMV) DNA in blood specimens. Intra- and interassay precision rates were 0.89% (small number of copies [L]), 1.43% (middle number of copies [M]), and 1.12% (high number of copies [H]), and 4.46% (L), 1.51% (M), and 2.28% (H), respectively. The linearity of this assay was obtained between 10 and 10
7
copies/well, with a minimum detection limit of 20 copies/well. Specimens from 55 of 70 healthy subjects were found to be positive for CMV antibody, but CMV DNA was not detected in any of them. In the qualitative assessment of each specimen, the results of the CMV antigenemia assay and those of the real-time PCR assay agreed in 80% (plasma specimens), 79% (all nucleated cells), and 86% (blood) of the cases examined. For eight patients diagnosed as having CMV infection or disease, no sample was positive in the antigenemia assay earlier than in the real-time PCR assay. Furthermore, the results of this assay could be obtained within 8 h. We concluded that the real-time PCR assay is useful for rapid diagnosis of CMV infection and monitoring of clinical courses.
Comparison of the Serodia
Treponema pallidum
Particle Agglutination, Captia Syphilis-G, and SpiroTek Reagin II Tests with Standard Test Techniques for Diagnosis of Syphilis
Authors: Pope Victoria , Fears Martha B. , Morrill William E. , Castro Arnold , Kikkert Susan E. 
Published: December 2019

ABSTRACT

We compared the microhemagglutination assay for
Treponema pallidum
(MHA-TP), a treponemal test, with two other treponemal tests, the Serodia
Treponema pallidum
particle agglutination (TP-PA) assay and the Captia Syphilis-G enzyme immunoassay, using 390 clinical serum samples. We also compared two nontreponemal tests, the rapid plasma Reagin (RPR) card test and the SpiroTek Reagin II test. Agreements of the MHA-TP with the TP-PA test and the Syphilis-G test were 97.4 and 97.7%, respectively. There was 89.2% agreement between the RPR and Reagin II tests. The Reagin II test was more apt to be reactive if the treponemal test was also reactive. We conclude that either the Serodia TP-PA test or the Captia Syphilis-G test is an appropriate substitute for the MHA-TP and that the Spirotek Reagin II test could substitute for the RPR test as a screening test.
Stability of IS
6110
Restriction Fragment Length Polymorphism Patterns of
Mycobacterium tuberculosis
Strains in Actual Chains of Transmission
Authors: Niemann Stefan , Rüsch-Gerdes Sabine , Richter Elvira , Thielen Heiko , Heykes-Uden Helga , Diel Roland 
Published: December 2019

ABSTRACT

The stability of IS
6110
restriction fragment length polymorphism (RFLP) patterns of
Mycobacterium tuberculosis
strains in actual transmission chains has been assessed by analyzing the variability of IS
6110
RFLP patterns of strains in fingerprint clusters that have been confirmed by classical epidemiological data. Forty susceptible and 35 drug-resistant (including 17 multidrug-resistant)
M. tuberculosis
strains obtained from 75 patients living in Germany have been analyzed. The epidemiological relationship among strains within the fingerprint clusters has been verified by family contacts (14 clusters) or by contact tracing of the public health offices (7 clusters). The time spans between the first and the last isolate of one cluster ranged from less than 1 to 29 months. Of the 75 strains only 1 showed a one-band variation when compared to the other nine isolates grouped in the same cluster, corresponding with a rate of change of ∼1.9% per possible transmission (one index patient per cluster was subtracted from the total number of isolates). These results confirm a high degree of stability of IS
6110
RFLP patterns of transmitted
M. tuberculosis
strains. Furthermore, the data presented indicate that isolates with identical IS
6110
DNA fingerprint patterns are a good indicator for the rate of recent transmission in a study population.
Comparison of 16S rRNA Gene PCR and BACTEC 9240 for Detection of Neonatal Bacteremia
Authors: Jordan J. A. , Durso M. B. 
Published: December 2019

ABSTRACT
Ten percent of infants born in the United States are admitted to neonatal intensive care units (NICU) annually. Approximately one-half of these admissions are from term infants (>34 weeks of gestation) at risk for systemic infection. Most of the term infants are not infected but rather have symptoms consistent with other medical conditions that mimic sepsis. The current standard of care for evaluating bacterial sepsis in the newborn is performing blood culturing and providing antibiotic therapy while awaiting the 48-h preliminary result of culture. Implementing a more rapid means of ruling out sepsis in term newborns could result in shorter NICU stays and less antibiotic usage. The purpose of this feasibility study was to compare the utility of PCR to that of conventional culture. To this end, a total of 548 paired blood samples collected from infants admitted to the NICU for suspected sepsis were analyzed for bacterial growth using the BACTEC 9240 instrument and for the bacterial 16S rRNA gene using a PCR assay which included a 5-h preamplification culturing step. The positivity rates by culture and PCR were 25 (4.6%) and 27 (4.9%) positive specimens out of a total of 548 specimens, respectively. The comparison revealed sensitivity, specificity, and positive and negative predictive values of 96.0, 99.4, 88.9, and 99.8%, respectively, for PCR. In summary, this PCR-based approach, requiring as little as 9 h of turnaround time and blood volumes as small as 200 μl, correlated well with conventional blood culture results obtained for neonates suspected of having bacterial sepsis.
Simultaneous Detection and Genotyping of “Norwalk-Like Viruses” by Oligonucleotide Array in a Reverse Line Blot Hybridization Format
Authors: Vinjé Jan , Koopmans Marion P. G. 
Published: December 2019

ABSTRACT
“Norwalk-like viruses” (NLVs) are the most common cause of outbreaks of nonbacterial gastroenteritis worldwide. To date, the method most widely used for typing of NLV strains is sequencing and subsequent phylogenetic analysis of reverse transcription (RT)-PCR products, which has revealed the existence of stable distinct lineages (genotypes). This typing method is rather costly, not routinely used in clinical laboratories, and not very suitable for the analysis of large numbers of samples. Therefore, we have developed a rapid and simple method for genotyping of NLVs. The method, designated reverse line blot hybridization, is based on the nucleotide divergence of a region of the gene for RNA polymerase which can be used to classify NLVs into genotypes. NLV RNA was amplified by RT-PCR and then hybridized to 18 different membrane-bound oligonucleotides that were able to discriminate among 13 NLV genotypes. Application of the method to a panel of 132 positive stool samples from 34 outbreaks and 20 sporadic cases of gastroenteritis collected in a 6-year period (1994 to 1999) resulted in successful genotyping of 124 samples (94%), as confirmed by phylogenetic analysis. The nucleotide sequences of the remaning eight strains (6%) from three outbreaks did not cluster with the known NLV genotypes. Phylogenetic analysis of the complete and partial open reading frame 2 (capsid gene) sequences of these strains revealed the existence of one novel genotype (Alphatron) and one potentially novel genotype (Amsterdam). This novel method, which allows simultaneous detection and genotyping of NLVs, is useful in the diagnosis and typing of NLVs obtained from outbreaks and in large-scale epidemiological studies.
Virulent Rough Filaments of
Listeria monocytogenes
from Clinical and Food Samples Secreting Wild-Type Levels of Cell-Free p60 Protein
Authors: Rowan Neil J. , Candlish Alan A. G. , Bubert Andreas , Anderson John G. , Kramer Karl , McLauchlin Jim 
Published: December 2019

ABSTRACT

Atypical rough cell filaments of
Listeria monocytogenes
(designated FR variants), isolated from clinical and food samples, form long filaments up to 96 μm in length and demonstrated wild-type levels of adherence, invasion, and cytotoxicity to human epithelial HEp-2, Caco-2, and HeLa cells. Unlike previously described avirulent rough mutants of
L. monocytogenes
that secrete diminished levels of the major extracellular protein p60 and that form long chains that consist of multiple cells of similar size (designated MCR variants), FR variants secreted wild-type or greater levels of p60. This study shows that virulent filamentous forms of
L. monocytogenes
occur in clinical and food environments and have atypical morphological characteristics compared to those of the wild-type form.
Assessment of Viral Loads in Patients with Chronic Hepatitis C with AMPLICOR HCV MONITOR Version 1.0, COBAS HCV MONITOR Version 2.0, and QUANTIPLEX HCV RNA Version 2.0 Assays
Authors: Martinot-Peignoux Michéle , Le Breton Véronique , Fritsch Sandrine , Le guludec Gaëlle , Labouret Nathalie , Keller Françoise , Marcellin Patrick 
Published: December 2019

ABSTRACT

The correlation between response to antiviral therapy and pretreatment viral load in patients with chronic hepatitis C has prompted the development of quantitative assays to measure viral load. The aim of our study was to assess the clinical relevance of the newly developed semiautomated PCR system COBAS HCV MONITOR version 2.0 in comparison with (i) the AMPLICOR HCV MONITOR version 1.0 assay, which underestimates RNA concentration of hepatitis C virus (HCV) genotypes 2 to 6, and (ii) the QUANTIPLEX HCV RNA version 2.0 assay, which achieves equivalent quantification for each HCV genotype, with samples from 174 patients diagnosed with chronic hepatitis C before therapy. The level and range of quantification measured with AMPLICOR HCV MONITOR version 1.0 were 1 log lower than when measured with the COBAS HCV MONITOR version 2.0, at 0.261 × 10
6
RNA copies/ml (range, 0.001 × 10
6
to 2.50 × 10
6
RNA copies/ml) and 4.032 × 10
6
RNA copies/ml (range, 0.026 × 10
6
to 72.6 × 10
6
RNA copies/ml), respectively. The two assays showed a poor correlation (
r
2
= 0.175). The level and range of quantification were similar when measured with the COBAS HCV MONITOR version 2.0 and QUANTIPLEX HCV RNA version 2.0 assays, at 3.03 × 10
6
RNA copies/ml (range, 0.023 × 10
6
to 72.6 × 10
6
RNA copies/ml) and 4.91 Meq/ml (range, 0.200 to 49.5 Meq/ml), respectively. The two assays showed a strong correlation (
r
2
= 0.686) for each HCV genotype. The duration of treatment (6 or 12 months) is modulated according to HCV genotype and viral load. Our results indicate that COBAS HCV MONITOR version 2.0 and QUANTIPLEX HCV RNA version 2.0 assays showing an equal dynamic range for each HCV genotype are suitable tools to assess patients before therapy.
Early Virological Failure in Naive Human Immunodeficiency Virus Patients Receiving Saquinavir (Soft Gel Capsule)-Stavudine-Zalcitabine (MIKADO Trial) Is Not Associated with Mutations Conferring Viral Resistance
Authors: Mouroux M. , Yvon-Groussin A. , Peytavin G. , Delaugerre C. , Legrand M. , Bossi P. , Do B. , Trylesinski A. , Diquet B. , Dohin E. , Delfraissy J. F. , Katlama C. , Calvez V. ,   The MIKADO Study Group
Published: December 2019

ABSTRACTThe MIKADO trial was designed to evaluate the efficacy of stavudine-zalcitabine-saquinavir (soft gel capsule) [d4T-ddC-SQV(SGC)] in 36 naive patients (−3.3 log10units at week 24 [W24]). Among the 29 patients remaining on d4T-ddC-SQV(SGC) until W24, 10 harbored a virological failure (viral load of >200 copies/ml at W24) (group 1). To determine the reasons for therapeutic failure, genotypic and phenotypic resistance test results and SQV concentrations in plasma were analyzed and compared to those in successfully treated patients (viral load of <200 copies/ml at W24) (group 2). Reverse transcriptase and protease genotypic analyses in group 1 revealed the acquisition of only one SQV-associated mutation (L90M) in only two patients. There was no significant increase in the 50 or 90% inhibitory concentration of SQV in patients with or without the L90M mutation. However, the fact that two patients developed an L90M mutation only 4 weeks after relapse points to the need for genotypic resistance testing in the context of an initial failure of the antiretroviral regimen. At W24, the median SQV concentration in group 1 (71 ng/ml) was significantly lower than in group 2 (475 ng/ml), and the plasma SQV concentration was correlated with the viral load at W24 (r= −0.5;P< 0.05) and with the drop in viral load between day 0 and W24 (r= −0.5;P< 0.01). These results and the fact that the plasma SQV concentrations in the two groups prior to relapse (W12) were not significantly different strongly suggest that the early failure of this combination is not due to viral resistance but to a lack of compliance, pharmacological variability, and drug interactions or a combination of these factors.
Rapid Diagnosis of Legionnaires' Disease Using an Immunochromatographic Assay for
Legionella pneumophila
Serogroup 1 Antigen in Urine during an Outbreak in The Netherlands
Authors: Wever Peter C. , Yzerman Ed P. F. , Kuijper Ed J. , Speelman Peter , Dankert Jacob 
Published: December 2019

ABSTRACT

A new immunochromatographic assay for rapid qualitative detection of
Legionella pneumophila
serogroup 1 antigen in urine specimens was used during an outbreak of legionellosis in The Netherlands. The assay seems of the utmost value in providing a rapid diagnosis of Legionnaires' disease in patients with severe community-acquired pneumonia in an outbreak setting.
Rapid Differentiation of
Borrelia garinii
from
Borrelia afzelii
and
Borrelia burgdorferi
Sensu Stricto by LightCycler Fluorescence Melting Curve Analysis of a PCR Product of the
recA
Gene
Authors: Pietilä Johanna , He Qiushui , Oksi Jarmo , Viljanen Matti K. 
Published: December 2019

ABSTRACT

To differentiate the
Borrelia burgdorferi
sensu lato genospecies, LightCycler real-time PCR was used for the fluorescence (SYBR Green I) melting curve analysis of borrelial
recA
gene PCR products. The specific melting temperature analyzed is a function of the GC/AT ratio, length, and nucleotide sequence of the amplified product. A total of 32 DNA samples were tested. Of them three were isolated from
B. burgdorferi
reference strains and 16 were isolated from
B. burgdorferi
strains cultured from
Ixodes ricinus
ticks; 13 were directly isolated from nine human biopsy specimens and four
I. ricinus
tick midguts. The melting temperature of
B. garinii
was 2°C lower than that of
B. burgdorferi
sensu stricto and
B. afzelii
. Melting curve analysis offers a rapid alternative for identification and detection of
B. burgdorferi
sensu lato genospecies.
Immunoglobulin A1 Protease Activity inGemella haemolysans
Authors: Lomholt Jeanet A. , Kilian Mogens 
Published: December 2019

ABSTRACTThe purpose of this study was to determine the occurrence and nature of immunoglobulin A1 (IgA1) protease activity in members of the genusGemellaand related taxa. Among a total of 22Gemellastrains belonging to the four speciesGemella haemolysans,Gemella morbillorum,Gemella sanguinis, andGemella bergeriaeand four reference strains of the speciesHelcococcus kunzii,Facklamia hominis, andGlobicatella sanguinis, IgA1 protease activity was an exclusive character of all nine isolates ofG. haemolysans. The IgA1 protease ofG. haemolysansappears to be a metallo-type IgA1 protease that cleaves the Pro227-Thr228peptide bond in the hinge region of the α1 chain like that of severalStreptococcusspecies. Phenotypic characterization of the isolates demonstrates that screening for IgA1 protease activity provides a valuable means for species differentiation in this group of bacteria.
Surveillance of Rotavirus Strains in the United States: Identification of Unusual Strains
Authors: Griffin D. D. , Kirkwood C. D. , Parashar U. D. , Woods P. A. , Bresee J. S. , Glass R. I. , Gentsch J. R. ,   the National Rotavirus Strain Surveillance System Collaborating Laboratories
Published: December 2019

ABSTRACT
Rotavirus strains from 964 fecal specimens collected from children at 11 U.S. hospital laboratories from November 1997 to March 1998 and from samples collected at 12 laboratories from November 1998 to March 1999 were typed for G and P proteins. Serotype G1 was the predominant serotype in 1997–1998 (88%), followed by G2 (6.2%), G9 (3.3%), and G3 (1.5%). This pattern was similar to that seen in 1998–1999: G1 (79%), G2 (15%), G9 (3.0%), G4 (1.6%), and G3 (0.3%). Novel P[9] strains were identified in both seasons, and analysis of a 364-nucleotide fragment from gene segment 4 of one of the strains demonstrated 97.3% nucleotide identity with the prototype P3[9],G3 strain, AU1, isolated in Japan. This is the first report of a human AU1-like strain in the United States. These results reinforce our initial findings that serotype G9 persists in the United States but has not become a predominant strain and that the common serotypes G1 to G4 account for almost 90% of strains in circulation. Other uncommon strains exist in the United States but may have been overlooked before because of their low prevalence and the use of inadequate diagnostic tools.
Rapid Automated Antimicrobial Susceptibility Testing of
Streptococcus pneumoniae
by Use of the bioMerieux VITEK 2
Authors: Jorgensen James H. , Barry Arthur L. , Traczewski M. M. , Sahm Daniel F. , McElmeel M. Leticia , Crawford Sharon A. 
Published: December 2019

ABSTRACT

The VITEK 2 is a new automated instrument for rapid organism identification and susceptibility testing. It has the capability of performing rapid susceptibility testing of
Streptococcus pneumoniae
with specially configured cards that contain enriched growth medium and antimicrobial agents relevant for this organism. The present study compared the results of testing of a group of 53 challenge strains of pneumococci with known resistance properties and a collection of clinical isolates examined in two study phases with a total of 402 and 416 isolates, respectively, with a prototype of the VITEK 2. Testing was conducted in three geographically separate laboratories; the challenge collection was tested by all three laboratories, and the unique clinical isolates were tested separately by the individual laboratories. The VITEK 2 results of tests with 10 antimicrobial agents were compared to the results generated by the National Committee for Clinical Laboratory Standards reference broth microdilution MIC test method. Excellent interlaboratory agreement was observed with the challenge strains. The overall agreement within a single twofold dilution of MICs defined by the VITEK 2 and reference method with the clinical isolates was 96.3%, although there were a number of off-scale MICs that could not be compared. The best agreement with the clinical isolates was achieved with ofloxacin and chloramphenicol (100%), and the lowest level of agreement among those drugs with sufficient on-scale MICs occurred with trimethoprim-sulfamethoxazole (89.7%). Overall there were 1.3% very major, 6.6% minor, and no major interpretive category errors encountered with the clinical isolates, although >80% of the minor interpretive errors involved only a single log
2
dilution difference. The mean time for generation of susceptibility results with the clinical isolates was 8.1 h. The VITEK 2 provided rapid, reliable susceptibility category determinations with both the challenge and clinical isolates examined in this study.