bluuebunny/crossref_metadata_2025_split huggingface dataset:
Development of an Internal Positive Control for Rapid Diagnosis of Avian Influenza Virus Infections by Real-Time Reverse Transcription-PCR with Lyophilized Reagents
Authors: Das Amaresh , Spackman Erica , Senne Dennis , Pedersen Jan , Suarez David L. 
Published: September 2006

ABSTRACTWe developed an internal positive control (IPC) RNA to help ensure the accuracy of the detection of avian influenza virus (AIV) RNA by reverse transcription (RT)-PCR and real-time RT-PCR (RRT-PCR). The IPC was designed to have the same binding sites for the forward and reverse primers of the AIV matrix gene as the target amplicon, but it had a unique internal sequence used for the probe site. The amplification of the viral RNA and the IPC by RRT-PCR were monitored with two different fluorescent probes in a multiplex format, one specific for the AIV matrix gene and the other for the IPC. The RRT-PCR test was further simplified with the use of lyophilized bead reagents for the detection of AIV RNA. The RRT-PCR with the bead reagents was more sensitive than the conventional wet reagents for the detection of AIV RNA. The IPC-based RRT-PCR detected inhibitors in blood, kidney, lungs, spleen, intestine, and cloacal swabs, but not allantoic fluid, serum, or tracheal swabs The accuracy of RRT-PCR test results with the lyophilized beads was tested on cloacal and tracheal swabs from experimental birds inoculated with AIV and compared with virus isolation (VI) on embryonating chicken eggs. There was 97 to 100% agreement of the RRT-PCR test results with VI for tracheal swabs and 81% agreement with VI for cloacal swabs, indicating a high level of accuracy of the RRT-PCR assay. The same IPC in the form of armored RNA was also used to monitor the extraction of viral RNA and subsequent detection by RRT-PCR.
An Outbreak of Keratitis Caused by
Mycobacterium immunogenum
Authors: Sampaio Jorge Luiz Mello , Junior Doraldo Nassar , de Freitas Denise , Höfling-Lima Ana Luisa , Miyashiro Kozue , Alberto Fernando Lopes , Leão Sylvia Cardoso 
Published: September 2006

ABSTRACT

From 8 October to 12 November 2003, 36 patients underwent surgical correction of myopia in a São Paulo, Brazil, clinic. Five patients had clinical signs of infectious keratitis, and a
Mycobacterium
species with previously unreported patterns determined by PCR restriction enzyme analysis of the
hsp65
gene and PCR restriction enzyme analysis of the 16S-23S rRNA internal transcribed spacer (ITS) was isolated from corneal scrapings from four of these patients. Subsequent evaluation by phenotypic tests and partial sequencing of the
hsp65
,
sodA
,
rpoB
, and 16S rRNA genes and the ITS supported the species identification as a variant of
Mycobacterium immunogenum
. The source of infection was not determined. The outbreak was caused by a single clone, as evidenced by identical pulsed-field gel electrophoresis and enterobacterial repetitive intergenic consensus-PCR profiles. This is the first report of an outbreak where this species was isolated from infected tissues.
Multicenter Evaluation of the Etest and Disk Diffusion Methods for Differentiating Daptomycin-Susceptible from Non-Daptomycin-Susceptible
Staphylococcus aureus
Isolates
Authors: Jevitt Laura A. , Thorne Grace M. , Traczewski Maria M. , Jones Ronald N. , McGowan John E. , Tenover Fred C. , Brown Steven D. 
Published: September 2006

ABSTRACT

Daptomycin is a novel cyclic lipopeptide that is approved by the U.S. Food and Drug Administration for the treatment of complicated skin and skin structure infections associated with
Staphylococcus aureus
and other gram-positive pathogens and also staphylococcal bacteremia, including right-sided endocarditis. The Clinical and Laboratory Standards Institute (CLSI) established “susceptible-only” interpretive criteria for broth microdilution (BMD) and disk diffusion (DD) testing of daptomycin in 2005. However, a series of
S. aureus
isolates have been recovered with daptomycin MICs in the nonsusceptible range (i.e., MICs of >1 μg/ml). The objective of this study was to determine the ability of the Etest and DD methods to differentiate daptomycin-susceptible from nonsusceptible isolates of
S. aureus
compared to the results of the CLSI BMD reference method. There was a good correlation between Etest MIC results and the results of BMD among laboratories (
r
= 0.86 to 0.88), with 95.3% of the Etest MICs within a ±1 log
2
dilution of the BMD MIC result. A total of 92 of 102 (90.2%) non-daptomycin-susceptible isolates of
S. aureus
identified by BMD in two participating laboratories were also classified as nonsusceptible by Etest. However, the very major and major error rates reported by one of the participating laboratories were 13.5 and 4.0%, respectively, primarily due to the absence of an intermediate category. The DD method, however, did not reliably differentiate daptomycin-susceptible from non-daptomycin-susceptible isolates. In 2005, daptomycin disks were voluntarily removed from the market by Cubist Pharmaceuticals. The disk diffusion breakpoints were subsequently removed from the CLSI M100 standard in 2006.
Performance of Human Immunodeficiency Virus Type 1 gp41 Assays for Detecting Enfuvirtide (T-20) Resistance Mutations
Authors: Marcial Michelle , Lu Jing , Deeks Steven G. , Ziermann Rainer , Kuritzkes Daniel R. 
Published: September 2006

ABSTRACT
The performance of a gp41 assay, created using reagents designed for use with the OpenGene DNA Sequencing System, was evaluated using a panel of plasma samples obtained from enfuvirtide-naive and -experienced human immunodeficiency virus type 1-infected subjects. Resulting sequence data were highly accurate compared to a “home brew” assay and clonal sequence analysis.
Diagnosis of Gastric Syphilis by Direct Immunofluorescence Staining and Real-Time PCR Testing
Authors: Chen Cheng-Yen , Chi Kai-Hua , George Robert W. , Cox David L. , Srivastava Amitabh , Rui Silva Mário , Carneiro Fátima , Lauwers Gregory Y. , Ballard Ronald C. 
Published: September 2006

ABSTRACT
We report on a case of gastric syphilis in a patient with chronic dyspepsia. The diagnosis was established by serology and the demonstration of spirochetes in diffusely inflammed gastric mucosa by staining with a fluorescent monoclonal antibody specific for pathogenic treponemes and by the detection of specific treponemal DNA sequences by a real-time PCR.
Clinical Significance of Coagulase-Negative Staphylococci Recovered from Nonsterile Sites
Authors: Tan Thean Yen , Ng Siew Yong , Ng Wan Xing 
Published: September 2006

ABSTRACT

Laboratory criteria were used to select coagulase-negative staphylococci isolated from nonsterile body sites for further investigation. Fifty-seven percent of the study isolates were clinically significant, predominantly causing community-acquired soft tissue infections. There were species-related differences in virulence, with 91% of
Staphylococcus lugdunensis
isolates clinically significant compared with 11% of
S. haemolyticus
isolates.
Differential Detection of
Entamoeba histolytica
,
Entamoeba dispar
, and
Entamoeba moshkovskii
by a Single-Round PCR Assay
Authors: Hamzah Zulhainan , Petmitr Songsak , Mungthin Mathirut , Leelayoova Saovanee , Chavalitshewinkoon-Petmitr Porntip 
Published: September 2006

ABSTRACT

A single-round PCR assay was developed for detection and differential diagnosis of the three
Entamoeba
species found in humans,
Entamoeba moshkovskii
,
Entamoeba histolytica
, and
Entamoeba dispar
, that are morphologically identical as both cysts and trophozoites. A conserved forward primer was derived from the middle of the small-subunit rRNA gene, and reverse primers were designed from signature sequences specific to each of these three
Entamoeba
species. PCR generates a 166-bp product with
E. histolytica
DNA, a 752-bp product with
E. dispar
DNA, and a 580-bp product with
E. moshkovskii
DNA. Thirty clinical specimens were examined, and the species present were successfully detected and differentiated using this assay. It was possible to detect as little as 10 pg of
E. moshkovskii
and
E. histolytica
DNA, while for
E. dispar
the sensitivity was about 20 pg of DNA. Testing with DNA from different pathogens, including bacteria and other protozoa, confirmed the high specificity of the assay. We propose the use of this PCR assay as an accurate, rapid, and effective diagnostic method for the detection and discrimination of these three morphologically indistinguishable
Entamoeba
species in both routine diagnosis of amoebiasis and epidemiological surveys.
Fourier-Transform Infrared Spectroscopic Analysis Is a Powerful Tool for Studying the Dynamic Changes in
Staphylococcus aureus
Small-Colony Variants
Authors: Becker Karsten , Laham Nahed Al , Fegeler Wolfgang , Proctor Richard A. , Peters Georg , von Eiff Christof 
Published: September 2006

ABSTRACT

Infections due to small-colony variants (SCVs) of
Staphylococcus aureus
in patients with chronic and recurrent infections are an emerging problem; however, studies with this subpopulation are hampered by the fact that SCVs may exhibit unstable phenotypes, making them difficult to study, particularly in broth media. In this study, two
S. aureus
sets comprising the (i) normal and the (ii) SCV phenotype (clonal with normal phenotype) recovered from clinical specimens, as well as (iii) corresponding site-directed mutants displaying the SCV phenotype (knockout of
hemB
) and (iv) their complemented mutants were examined by Fourier-transform infrared (FTIR) spectroscopy. Phenotypes were defined on solid and in broth media. Using first-derivative infrared spectra to calculate spectral distances, hierarchical clustering based on spectral information resulted in a dendrogram with clear discrimination between SCV and normal phenotypes. The SCVs gave an FTIR fingerprint that was easily recognizable and that was much closer to other SCVs than to their parent strains. This technique offers for the first time a noninvasive approach to investigate dynamic processes of reversion of SCVs to the normal phenotype and vice versa. Thus, FTIR spectroscopy allowed a rapid and reproducible tool for the examination of different subpopulations of
S. aureus
on solid and in broth media for diagnostic and research purposes.
Detection of Hepatitis B Virus (HBV) Genotype E Carried—Even in the Presence of High Titers of Anti-HBs Antibodies—by an Argentinean Patient of African Descent Who Had Received Vaccination against HBV
Authors: Mathet Verónica L. , Cuestas María L. , Ruiz Vanesa , Minassian María L. , Rivero Cintia , Trinks Julieta , Daleoso Graciela , León Liliana M. , Sala Andrea , Libellara Beatriz , Corach Daniel , Oubiña José R. 
Published: September 2006

ABSTRACT
Genotype E hepatitis B virus (HBV) was detected in two Argentine sisters exhibiting an African mitochondrial lineage. One of them (who had been vaccinated against HBV) exhibited anti-HBs cocirculating antibodies without HBsAg escape mutants, while her unvaccinated sister showed a D144A HBsAg escape mutant without anti-HBs antibodies. Both sisters carried an unusual L209V substitution within HBsAg.
Phenotypic Detection of Carbapenem-Susceptible Metallo-β-Lactamase-Producing Gram-Negative Bacilli in the Clinical Laboratory
Authors: Franklin Clare , Liolios Lisa , Peleg Anton Y. 
Published: September 2006

ABSTRACT
Rapid detection of metallo-β-lactamase (MBL)-producing gram-negative pathogens is critical to prevent their widespread dissemination. Thus far, no standardized phenotypic method is available, and previously reported techniques have poor sensitivity for detecting carbapenem-susceptible MBL-carrying isolates, an increasingly described phenomenon. We developed a phenotypic detection method using both a double-disk synergy test and a combined-disk test with imipenem and 292 μg EDTA on one agar plate. Genotypic confirmation was used for validation. Of the 134 clinical isolates, 84 were confirmed to carry an MBL. Of these, 51 (61%) were susceptible to at least one carbapenem, and 22 (26%) were isolated from blood. The phenotypic method correctly differentiated all MBL-producing isolates (sensitivity, 100%). Fifty-one of the 52 MBL-negative isolates were correctly differentiated (specificity, 98%). This study reports the validation of a simple and accurate MBL detection method that can be easily incorporated into the daily routine of a clinical laboratory. Early detection of MBL-carrying organisms, including those with susceptibility to carbapenems, is of paramount clinical importance, as it allows rapid initiation of strict infection control practices as well as therapeutic guidance for confirmed infection.
Real-Time PCR Assays for Detection of Bocavirus in Human Specimens
Authors: Lu Xiaoyan , Chittaganpitch Malinee , Olsen Sonja J. , Mackay Ian M. , Sloots Theo P. , Fry Alicia M. , Erdman Dean D. 
Published: September 2006

ABSTRACT

The recently discovered human bocavirus (HBoV) is the first member of the family
Parvoviridae
, genus
Bocavirus
, to be potentially associated with human disease. Several studies have identified HBoV in respiratory specimens from children with acute respiratory disease, but the full spectrum of clinical disease and the epidemiology of HBoV infection remain unclear. The availability of rapid and reliable molecular diagnostics would therefore aid future studies of this novel virus. To address this, we developed two sensitive and specific real-time TaqMan PCR assays that target the HBoV NS1 and NP-1 genes. Both assays could reproducibly detect 10 copies of a recombinant DNA plasmid containing a partial region of the HBoV genome, with a dynamic range of 8 log units (10
1
to 10
8
copies). Eight blinded clinical specimen extracts positive for HBoV by an independent PCR assay were positive by both real-time assays. Among 1,178 NP swabs collected from hospitalized pneumonia patients in Sa Kaeo Province, Thailand, 53 (4.5%) were reproducibly positive for HBoV by one or both targets. Our data confirm the possible association of HBoV infection with pneumonia and demonstrate the utility of these real-time PCR assays for HBoV detection.
Comparison of API 20E and
invA
PCR for Identification of
Salmonella enterica
Isolates from Swine Production Units
Authors: Nucera Daniele M. , Maddox Carol W. , Hoien-Dalen Patricia , Weigel Ronald M. 
Published: September 2006

ABSTRACT

API 20E and
invA
PCR were evaluated for the identification of
Salmonella enterica
isolates from swine farms. API 20E had the highest agreement with other tests at the 99.9% likelihood level. Both tests had 100% sensitivity and 96% specificity compared to 16S rRNA sequencing. Compared to serotyping, both tests had 96% sensitivity; specificity was 86% for API 20E and 79% for
invA
PCR.
Quantitative Assessment of Serum NV-F Virus DNA Concentrations in Samples from Patients Coinfected with Hepatitis B or C Virus
Authors: Hsu Chao-Wei , Cheng Ju-Chien , Yeh Chau-Ting 
Published: September 2006

ABSTRACT

A novel hepatotropic virus, named NV-F virus, was recently identified. The clinical information for this virus is still scarce. Using PCR assay, NV-F viral DNA (NV-F-DNA) was detected in 12 of 50 (24%) hepatitis C virus (HCV)-infected patients (HCV-coinfected [HCVCI] group), 34 of 250 (13.6%) hepatitis B virus (HBV)-infected patients (HBV-coinfected [HBVCI] group), and 28 of 100 (28%) non-A-to-E (NAE) hepatitis patients. Basic clinical parameters were not significantly different among the three groups. By use of a PCR-based quantitative assay, the NV-F-DNA concentration was found to be above the detection limit (1.2 × 10
5
copies/ml) in 12/12 (100%) HCVCI patients, 14/34 (41.2%) HBVCI patients, and 4/28 (14.3%) NAE patients. The median serum NV-F-DNA concentration was 9.3 × 10
5
copies/ml in HCVCI patients, but it was below the detection limit in HBVCI and NAE patients (
P
values were 0.0045 and 0.0001, respectively). Stepwise multiple regression analysis identified the presence of anti-HCV as an independent factor for NV-F-DNA concentrations (β = 6.2 × 10
9
;
P
= 0.0245). In HBVCI patients, the NV-F-DNA concentration was inversely correlated with the HBV DNA concentration. The median NV-F-DNA concentration was below the detection limit in patients with HBV DNA concentrations above 1.4 × 10
5
copies/ml, but it was 1.58 × 10
6
copies/ml in patients with HBV DNA concentrations below 1.4 × 10
5
copies/ml (
P
= 0.030). In conclusion, NV-F-DNA concentrations were higher in HCVCI patients. A reciprocal relationship was found between NV-F-DNA and HBV DNA concentrations in HBVCI patients, indicating the presence of viral interference between these two DNA viruses.
Ventilator-Associated Pneumonia Caused by
Dolosigranulum pigrum
Authors: Hoedemaekers A. , Schülin T. , Tonk B. , Melchers W. J. G. , Sturm P. D. J. 
Published: September 2006

ABSTRACT

Dolosigranulum pigrum
is an unusual gram-positive catalase-negative coccus. It was isolated, only after prolonged incubation, from bronchial secretions from a patient with ventilator-associated pneumonia. The patient responded well to antimicrobial therapy. Identification was done by 16S rRNA DNA sequence analysis, but it can be done with relatively simple phenotypic tests.
Comparison of Tests To Detect Oxacillin Resistance in
Staphylococcus intermedius
,
Staphylococcus schleiferi
, and
Staphylococcus aureus
Isolates from Canine Hosts
Authors: Bemis David A. , Jones Rebekah D. , Hiatt Lauren E. , Ofori Edward D. , Rohrbach Barton W. , Frank Linda A. , Kania Stephen A. 
Published: September 2006

ABSTRACT

Multiple tests were compared to the reference standard PBP2a latex agglutination test for detection of
mecA
-mediated oxacillin resistance in canine staphylococci. Cefoxitin disk diffusion, using breakpoints for human isolates of coagulase-negative
Staphylococcus
spp., had low sensitivity for detection of oxacillin resistance in members of the
Staphylococcus intermedius
group.
The Key Role of Pulsed-Field Gel Electrophoresis in Investigation of a Large Multiserotype and Multistate Food-Borne Outbreak ofSalmonellaInfections Centered in Pennsylvania
Authors: Sandt Carol H. , Krouse Donna A. , Cook Charles R. , Hackman Amy L. , Chmielecki Wayne A. , Warren Nancy G. 
Published: September 2006

ABSTRACTFive different serotypes ofSalmonella entericawere implicated in a large outbreak linked to fresh Roma tomatoes served at gas station deli counters in Pennsylvania and nearby states during July 2004:S. entericaserotypes Javiana, Anatum, Thompson, Typhimurium, and Muenchen. One of these serotypes, Anatum, was isolated from both tomatoes and patients. Pulsed-field gel electrophoresis (PFGE) played a key role in identifying the outbreak-associated isolates and distinguishing them from unrelated sporadic isolates. It also demonstrated that the genetic fingerprints of serotype Anatum isolates derived from patients were indistinguishable from those derived from tomatoes. Rapid communication of PFGE fingerprints with other public health laboratories through the Centers for Disease Control and Prevention's PulseNet USA national molecular surveillance network for bacterial food-borne pathogens facilitated the tracking of this outbreak in other states. The work described in this report emphasizes the laboratory's role in core public health functions and services, thereby providing a highly visible example of public health in action.
Mycobacterium tuberculosis
Complex and Mycobacterial Heat Shock Proteins in Lymph Node Tissue from Patients with Pulmonary Sarcoidosis
Authors: Dubaniewicz Anna , Dubaniewicz-Wybieralska Mirosława , Sternau Adam , Zwolska Zofia , Iżycka-Świeszewska Ewa , Augustynowicz-Kopeć Ewa , Skokowski Jarosław , Singh Mahavir , Zimnoch Lech 
Published: September 2006

ABSTRACT

We demonstrated that not whole
Mycobacterium tuberculosis
but its particular antigens, hsp70

Mtb

, hsp65

Mtb

, and hsp16

Mtb

, are present in lymph node tissues of patients with sarcoidosis (SA). hsp16

Mtb

occurs in the early stage of SA, whereas hsp70

Mtb

occurs in stage II of SA. hsp65

Mtb

is highly expressed in the capillary vessels in lymph node tissues in patients with SA.
Quinupristin-Dalfopristin Resistance inEnterococcus faeciumIsolates from Humans, Farm Animals, and Grocery Store Meat in the United States
Authors: Donabedian S. M. , Perri M. B. , Vager D. , Hershberger E. , Malani P. , Simjee S. , Chow J. , Vergis E. N. , Muder R. R. , Gay K. , Angulo F. J. , Bartlett P. , Zervos M. J. 
Published: September 2006

ABSTRACTThree hundred sixty-one quinupristin-dalfopristin (Q-D)-resistantEnterococcus faecium(QDREF) isolates were isolated from humans, turkeys, chickens, swine, dairy and beef cattle from farms, chicken carcasses, and ground pork from grocery stores in the United States from 1995 to 2003. These isolates were evaluated by pulsed-field gel electrophoresis (PFGE) to determine possible commonality between QDREF isolates from human and animal sources. PCR was performed to detect the streptogramin resistance genesvatD,vatE, andvgbAand the macrolide resistance geneermBto determine the genetic mechanism of resistance in these isolates. QDREF from humans did not have PFGE patterns similar to those from animal sources.vatEwas found in 35%, 26%, and 2% of QDREF isolates from turkeys, chickens, and humans, respectively, and was not found in QDREF isolates from other sources.ermBwas commonly found in QDREF isolates from all sources. Known streptogramin resistance genes were absent in the majority of isolates, suggesting the presence of other, as-yet-undetermined, mechanisms of Q-D resistance.
Temperature Gradient Gel Electrophoresis of Fecal 16S rRNA Reveals Active
Escherichia coli
in the Microbiota of Patients with Ulcerative Colitis
Authors: Sokol H. , Lepage P. , Seksik P. , Doré J. , Marteau P. 
Published: September 2006

ABSTRACT

Previous studies of the endogenous microbiota in patients with ulcerative colitis (UC) have not taken bacterial activity into account, yet bacteria with high transcriptional activity might have a more important pathophysiological role than inactive bacteria. We therefore analyzed the biodiversity of active bacteria in the fecal microbiota of UC patients, in comparison with that of healthy subjects. Feces were collected from nine patients with active UC and from nine healthy controls. Total DNA and RNA were extracted, and 16S ribosomal DNA and RNA were amplified by PCR and reverse transcription-PCR, respectively. Amplification products were compared by means of temporal temperature gradient gel electrophoresis (TTGE). Bands of interest were excised, sequenced, and identified by comparison with the GenBank database (NCBI). The dominant-species diversity based on RNA-derived TTGE profiles was significantly lower for UC patients than for healthy controls (
P
= 0.01). The mean similarity index between the “present” and “active” microbiota was 74% ± 18% for UC patients. Comparison of the individual “active” microbiota identified a band that was present for eight UC patients and only two controls (89% versus 22%;
P
= 0.008). The band was sequenced for 6 patients and always corresponded to
Escherichia coli
. The biodiversity of active bacteria in the dominant fecal microbiota of patients with UC is lower than that of healthy subjects.
E. coli
is more represented in the active microbiota of UC patients. The possible pathophysiological role of this difference remains to be determined.