bluuebunny/crossref_metadata_2025 huggingface dataset:
Evaluation of the Automated Phoenix System for Potential Routine Use in the Clinical Microbiology Laboratory
Authors: Donay J.-L. , Mathieu D. , Fernandes P. , Prégermain C. , Bruel P. , Wargnier A. , Casin I. , Weill F. X. , Lagrange P. H. , Herrmann J. L.
Published: April 2004

A comparative study was designed to evaluate the identification (ID) and antimicrobial susceptibility testing (AST) performances of the BD Phoenix Automated Microbiology System (Becton Dickinson Diagnostic Systems [BD], Pont de Claix, France). A total of 305 single clinical isolates were collected, and comparisons were made with routine manual methods in use in our microbiology laboratories. The percentages of correct IDs were 93.3, 89.4, 91.8, and 85.7% for enterobacteria, nonfermenting gram-negative bacilli, staphylococci, and streptococci-enterococci, respectively. The median ID time was 3 h, and the median time for AST was 10 h 30 min. AST results showed variable percentages of errors for the different antibiotics. None of the enterobacteria and 0.3% of
Pseudomonas aeruginosa
isolates showed a very major error (VME). Only one strain of
Staphylococcus aureus
showed a VME with oxacillin. We demonstrate here the efficiency of the Phoenix system, which can be used for the majority of strains encountered in a university-based laboratory, for ID and AST.
Practical Strategies for Performance Optimization of the Enhanced Gen-Probe Amplified Mycobacterium Tuberculosis Direct Test
Authors: Sloutsky A. , Han L. L. , Werner B. G.
Published: April 2004

The enhanced Gen-Probe Amplified Mycobacterium Tuberculosis Direct (MTD) test was evaluated using a combined set of 338 acid-fast smear-positive and smear-negative, respiratory and nonrespiratory clinical specimens received by the Massachusetts State Tuberculosis Laboratory from September 1999 through March 2002. Microbiological culture was used as the reference method; therefore, the sensitivity and specificity of the MTD test were calculated for culture-positive specimens only. The initial assessment indicated that the overall sensitivity, specificity, and positive and negative predictive values of the MTD test for all specimens grouped together were 62, 98, 99, and 68%, respectively. A detailed discrepancy analysis revealed that two major factors causing negative MTD results in specimens that were culture positive for
M. tuberculosis
complex were patient treatment with antituberculosis drugs prior to testing and the presence of inhibitory substances in the specimen. Based on these findings, a protocol for optimizing MTD test performance in this setting is proposed in which (i) specimens from patients taking antituberculosis medications are excluded from testing and (ii) all initially MTD-negative or MTD-equivocal specimens are subjected to testing for inhibitors. If this strategy was followed, the MTD test sensitivity would be at least 91%, a significant improvement over the initial sensitivity of 62%. Accordingly, the negative predictive value would increase from 68 to 91%.
Multiplex PCR Genotyping Assay That Distinguishes between Isolates ofClostridium perfringensType A Carrying a Chromosomal Enterotoxin Gene (cpe) Locus, a PlasmidcpeLocus with an IS1470-Like Sequence, or a PlasmidcpeLocus with an IS1151Sequence
Authors: Miyamoto Kazuaki , Wen Qiyi , McClane Bruce A.
Published: April 2004

Clostridium perfringenstype A isolates carrying the enterotoxin (cpe) gene are important causes of both food poisoning and non-food-borne diarrheas in humans. In North America and Europe, food poisoning isolates were previously shown to carry a chromosomalcpegene, while non-food-borne gastrointestinal (GI) disease isolates from those two geographic locations were found to have a plasmidcpegene. In this report, we describe the development of an economical multiplex PCRcpegenotyping assay that works with culture lysates to distinguish among type A isolates carrying a chromosomalcpegene, a plasmidcpegene with a downstream IS1470-like sequence, or a plasmidcpegene with a downstream IS1151sequence. When this multiplex PCR assay was applied in molecular epidemiologic studies, it was found that (i) all 57 examined type A isolates with a plasmidcpegene have either IS1470-like or IS1151sequences downstream of the plasmidcpegene; (ii) an IS1470-like sequence, rather than an IS1151sequence, is more commonly present downstream of the plasmidcpegene (particularly in North American non-food-borne human GI disease isolates); and (iii) as previously shown in the United States and Europe, isolates carrying the chromosomalcpegene also appear to be the major cause ofC. perfringensfood poisoning in Japan. The superiority of this new multiplex PCR assay over existingcpegenotyping approaches should facilitate further molecular epidemiologic investigations ofC. perfringensenterotoxin-associated GI illnesses and their associatedcpe-positive type A isolates.
Direct Detection of Rifampin- and Isoniazid-Resistant
Mycobacterium tuberculosis
in Auramine-Rhodamine-Positive Sputum Specimens by Real-Time PCR
Authors: Ruiz Maite , Torres Maria J. , Llanos Ana C. , Arroyo Aurelio , Palomares Jose C. , Aznar Javier
Published: April 2004

Our objective was to evaluate the feasibility of a molecular assay based on a real-time PCR technique, carried out with a LightCycler instrument (Roche Biochemicals), to identify
Mycobacterium tuberculosis
bacilli and to detect rifampin and isoniazid resistance in DNA extracts from sputum samples. We studied three genes:
rpoB
, which is associated with rifampin resistance, and
katG
and
inhA
, which are associated with isoniazid resistance. A total of 205 sputum samples collected from 108 patients diagnosed with pulmonary tuberculosis with positive auramine-rhodamine-staining (AR) sputum samples, were tested. The sensitivities of the LightCycler PCR assay for the positive AR specimens was 97.5% (200 of 205) for
rpoB
and
inhA
genes and 96.5% (198 of 205) for the
katG
gene. For the total number of patients tested, the sensitivity was 100% (108 of 108 patients) for rifampin, whereas the sensitivity was 98.1% (106 of 108 patients) for isoniazid. Full agreement was found with the Bactec MGIT 960 method and the genotype inferred from the LightCycler data for rifampin. The phenotypic method for isoniazid reported 13 resistant strains (≥0.1 μg/ml). In seven (53.8%) strains there was a concordance between both methods, but we found that six (46.2%) strains reported as resistant by the phenotypic method were determined to be susceptible by real-time PCR. For the 75 strains reported as susceptible by the phenotypic method, the concordance with the LightCycler data was 100%. Our results demonstrate that rifampin-resistant
M. tuberculosis
could be detected in DNA extracted from auramine-rhodamine-positive sputum samples in a single-tube assay that took less than 3 h to perform for a collection of auramine-rhodamine-positive specimens obtained from patients with culture-documented pulmonary tuberculosis. Similarly, this occurs in half of the isoniazid-resistant
M. tuberculosis
DNA extracted from auramine-rhodamine-positive specimens.
Endocarditis with Ruptured Cerebral Aneurysm Caused by
Cardiobacterium valvarum
sp. nov
Authors: Han Xiang Y. , Meltzer Michelle C. , Woods Joan T. , Fainstein Victor
Published: April 2004

A fastidious gram-negative bacterium was isolated from the blood of a 37-year-old man who had insidious endocarditis with a sudden rupture of a cerebral aneurysm. Characterization of the organism through phylogenetic and phenotypic analyses revealed a novel species of
Cardiobacterium
, for which the name
Cardiobacterium valvarum
sp. nov. is proposed.
C. valvarum
will supplement the current sole species
Cardiobacterium hominis
, a known cause of endocarditis. Surgeries and antibiotic treatment cured the patient's infection and associated complications. During cardiac surgery, a congenital bicuspid aortic valve was found to be the predisposing factor for his endocarditis.
Chromatography Paper Strip Method for Collection, Transportation, and Storage of Rotavirus RNA in Stool Samples
Authors: Rahman Mustafizur , Goegebuer Truus , De Leener Karolien , Maes Piet , Matthijnssens Jelle , Podder Goutam , Azim Tasnim , Van Ranst Marc
Published: April 2004

We developed a novel method that uses sodium dodecyl sulfate-EDTA-treated chromatography paper strips to collect unconcentrated fresh stool samples. After the paper strips were stored for 4 months at room temperature, rotavirus RNA could be successfully amplified by using reverse transcriptase PCR. The use of filter paper strips as a specimen support allows (self-)collection of stool samples by untrained persons. Diarrheal stool samples from remote areas can be stored and transported to a central diagnostic laboratory without the need for freezers or special shipping conditions. This convenient and inexpensive rotavirus sample collection system can be of use in epidemiological surveillance studies and vaccine trials.
Diversity of Group A Human Rotavirus Types Circulating over a 4-Year Period in Madrid, Spain
Authors: Sánchez-Fauquier Alicia , Wilhelmi Isabel , Colomina Javier , Cubero Eusebio , Roman Enriqueta
Published: April 2004

The incidence and distribution of human rotavirus G types among children under 5 years old with acute gastroenteritis were determined over a 4-year period (1998 to 2002) by using monoclonal antibodies and reverse transcription-PCR methods. Rotavirus was detected in 1,155 (31%) of 3,760 specimens tested. Rotavirus was studied in every month of the 48-month survey period. Rotavirus activity occurred mainly (51%) in the typically cooler months in Spain (November to February). The age distribution of rotavirus-positive cases showed that 90% of patients (1,038 of 1,155) were under 2 years old. Rotavirus types were determined for 576 of 1,155 patients (50%). G1 was the main genotype detected (53%), and the second most common was G4 (24%). The G2, G9, and G3 rotavirus types were detected in 14, 6, and 2% of the cases, respectively. Dual infections were detected in only 0.6%. The seasonal distribution of genotypes showed a significant genotypic shift: whereas G4 strains predominated (57%) during the 1998 to 2000 seasons, the G1 gradually increased to account for 75% in the 2000 to 2002 seasons. In addition, the present study reports the first detection of the G9 genotype in human fecal samples in Spain. Therefore, additional types may be required for vaccine development strategies that currently target only types G1 to G4.
Simple Algorithm for Identification of
Bordetella pertussis
Pertactin Gene Variants
Authors: Muyldermans Gaëtan , Piérard Denis , Hoebrekx Nathalie , Advani Reza , Van Amersfoorth Shirley , De Schutter Iris , Soetens Oriane , Eeckhout Leo , Malfroot Anne , Lauwers Sabine
Published: April 2004

Studies performed in several countries have demonstrated the recent emergence and subsequent dominance of circulating
Bordetella pertussis
strains harboring pertactin and pertussis toxin variants not included in pertussis vaccines. Determination of the pertactin gene variants is commonly performed using a time-consuming and expensive sequence analysis. We developed a simple and reliable pertactin typing algorithm suitable for large-scale screening. The assay correctly identified all pertactin alleles in representative strains. The typing of 231 clinical strains of
B. pertussis
routinely isolated in Belgium showed that this algorithm was adequate to identify less-frequent
prn
types like
prn9
and
prn11
.
Lack of Longitudinal Intrapatient Correlation between p24 Antigenemia and Levels of Human Immunodeficiency Virus (HIV) Type 1 RNA in Patients with Chronic HIV Infection during Structured Treatment Interruptions
Authors: Prado Julia G. , Shintani Ayumi , Bofill Margarita , Clotet Bonaventura , Ruiz Lidia , Martinez-Picado Javier
Published: April 2004

Structured treatment interruptions (STIs) have been proposed as a potential treatment strategy during human immunodeficiency virus type 1 (HIV-1) antiretroviral therapy. This still-experimental intervention requires a close monitoring of patients' plasma viremia and CD4
+
-T-cell counts during the treatment interruption phase. By using signal amplification of a heat-dissociated p24 antigen (p24Ag) assay, we compared p24Ag levels with levels of HIV RNA in plasma. One hundred seventy-four plasma samples were obtained from 51 chronically HIV-infected patients: 117 from patients who underwent STIs and 57 from patients who did not. Partial immune complex dissociation and clearance of those complexes by the erythrocytes were also investigated. A significant association between the two assays was observed (β = 0.23, 95% confidence interval = 0.18, 0.28;
P
< 0.0001), but the association was smaller in the subset of samples from patients undergoing STIs. Moreover, discordant results and lack of longitudinal intrapatient correlation between levels of p24Ag and HIV-1 RNA were higher in this group. Incomplete immune complex dissociation and binding of those complexes to erythrocytes could be contributing factors involved in the diminished detection of p24Ag. Therefore, signal amplification of a heat-dissociated p24Ag had a positive association with current HIV RNA assays in a population-based analysis. However, it might not be sensitive enough to monitor longitudinal intrapatient viremia during STIs in patients with high CD4
+
-T-cell counts potentially due to the production of high-affinity anti-p24 antibodies and clearance of immune complexes by erythrocytes.
Analysis of
Helicobacter pylori
Genotypes and Correlation with Clinical Outcome in Turkey
Authors: Saribasak Huseyin , Salih Barik A. , Yamaoka Yoshio , Sander Ersan
Published: April 2004

The predominant
Helicobacter pylori
strains circulating among geographic locations differ in regard to genomic structure. The association of the
cagA
-positive,
vacA
s1 genotypes with peptic ulcer disease (PUD) and gastric cancer was reported in Western countries but not in East Asian countries. Strains from Western countries predominantly possessed
cagA
type 2a,
vacA
s1a or s1b/m1a, or
vacA
m2a genotypes, whereas strains from East Asia possessed
cagA
type 1a,
vacA
s1c/m1b, or
vacA
m2b genotypes. Whether the Turkish strains possessed such genotypes was investigated and correlated with the disease outcome. Seventy-three patients from Turkey were enrolled.
H. pylori
was detected in 65 (89%) patients (22 with gastritis, 33 with PUD, and 10 with gastric cancer) by any of the following tests:
Campylobacter
-like organism test, culture, or PCR. Among the
H. pylori-
positive patients, presence of the
cagA
gene (78%) was significantly associated with PUD (
P
< 0.00001), gastric cancer (
P
< 0.001), and
vacA
s1a genotypes (
P
< 0.0001). Multiple
vacA
genotypes were more prevalent in PUD and gastric cancer patients than in patients with gastritis. Restriction fragment length polymorphism analysis of the
cagA
gene revealed three different patterns with no significant association with clinical outcome. Turkish strains examined predominantly possessed
cagA
type 2a,
vacA
s1a/m1a, or
vacA
m2a genotypes, which were typical genotypes in strains from Western countries. This fact might be one of the reasons for the low prevalence of severe gastroduodenal diseases in Turkey compared to the East Asian countries.
Evaluation of Performance and Potential Clinical Impact of ProSpecT Shiga Toxin
Escherichia coli
Microplate Assay for Detection of Shiga Toxin-Producing
E. coli
in Stool Samples
Authors: Gavin Patrick J. , Peterson Lance R. , Pasquariello Anna C. , Blackburn Joanna , Hamming Mark G. , Kuo Kuo J. , Thomson Richard B.
Published: April 2004

Shiga toxin-producing
Escherichia coli
bacteria (STEC) are emerging pathogens capable of producing sporadic and epidemic diarrhea, hemorrhagic colitis, and potentially life-threatening hemolytic-uremic syndrome. Although the presence of
E. coli
O157 can be readily detected in stool by sorbitol-MacConkey agar culture (SMAC), STEC non-O157 serotypes cannot. In contrast to culture, testing for the presence of Shiga toxins 1 and 2 in stool detects both O157 and non-O157 STEC serotypes capable of causing disease. Over two consecutive summers, we evaluated the performance of the ProSpecT Shiga toxin
E. coli
Microplate assay (Alexon-Trend, Ramsey, Minn.), an enzyme immunoassay for the detection of Shiga toxins 1 and 2, on all stools submitted for culture of enteric pathogens, and the potential clinical impact of Shiga toxin detection. Twenty-nine stool specimens were STEC positive by ProSpecT assay. Twenty-seven of 29 STEC-positive isolates were confirmed by SMAC and serotyping or by a second enzyme immunoassay and PCR (positive predictive value, 93%). Thirteen of 27 confirmed Shiga toxin-producing strains were serotype O157. The remaining 14 strains represented 8 other serotypes. The ProSpecT assay was 100% sensitive and specific for detection of
E. coli
O157 in stool (7 of 7) compared to SMAC. In addition, the ProSpecT assay detected twice as many STEC as SMAC. Fifty-two percent of confirmed STEC-positive stools were nonbloody. Thus, in our population, screening strategies that test only visibly bloody stools for STEC would miss a majority of cases. Eleven (41%) STEC-positive patients were hospitalized, and eight (30%) developed severe disease (two developed hemolytic-uremic syndrome, and six developed hemorrhagic colitis). Prior to detection of STEC infection, seven (26%) and eight patients (30%) underwent unnecessary diagnostic procedures or received potentially deleterious empirical treatment, respectively. We propose that establishing a specific diagnosis of STEC may have prevented these potentially harmful interventions. We conclude that the ProSpecT assay is sensitive and specific for the detection of Shiga toxins 1 and 2 in stool and has potentially significant clinical impact for the individual patient and public health. Shiga toxin assays should be considered for routine use in settings where prevalence of STEC disease warrants testing.
Multiple Polymorphic Loci for Molecular Typing of Strains of
Mycobacterium leprae
Authors: Groathouse Nathan A. , Rivoire Becky , Kim Hansuk , Lee Hyeyoung , Cho Sang-Nae , Brennan Patrick J. , Vissa Varalakshmi D.
Published: April 2004

The need for molecular tools for the differentiation of isolates of
Mycobacterium leprae
, the organism that causes leprosy, is urgent in view of the continuing high levels of new case detection, despite years of aggressive chemotherapy and the consequent reduction in the prevalence of leprosy. The slow onset of leprosy and the reliance on physical examination for detection of disease have restricted the epidemiological tracking necessary to understand and control transmission. Two genetic loci in several isolates of
M. leprae
have previously been demonstrated to contain variable-number tandem repeats (VNTRs). On the basis of these reports and the availability of the full genome sequence, multiple-locus VNTR analysis for strain typing has been undertaken. A panel of 11 short tandem repeat (STR) loci with repeat units of 1, 2, 3, 6, 12, 18, 21, and 27 bp from four clinical isolates of
M. leprae
propagated in armadillo hosts were screened by PCR. Fragment length polymorphisms were detected at 9 of the 11 loci by agarose gel electrophoresis. Sequencing of representative DNA products confirmed the presence of VNTRs between isolates. The application of nine new polymorphic STRs in conjunction with automated methods for electrophoresis and size determination allows greater discrimination between isolates of
M. leprae
and enhances the potential of this technique to track the transmission of leprosy.
New Approach for Serological Testing for Leptospirosis by Using Detection of
Leptospira
Agglutination by Flow Cytometry Light Scatter Analysis
Authors: Yitzhaki S. , Barnea A. , Keysary A. , Zahavy E.
Published: April 2004

Leptospirosis is considered an important reemerging infectious disease worldwide. The standard and most widespread method for the diagnosis of leptospirosis is the microscopic agglutination test (MAT). This test is laborious and time-consuming, and the interpretation of the results is subjective. In the present work we describe an application of flow cytometry (FCM) as a tool for the serological diagnosis of leptospirosis. The analysis is based on the sensitivity of FCM to the size and shape of the bacteria analyzed by measurement of light scatter parameters: forward scatter (FSC) and side scatter (SSC). The addition of positive serum to an infecting leptospiral serovar results in a shift of the light scatter parameter to a different location with higher FSC and SSC values, indicating the formation of leptospiral aggregates. By using immunofluorescent staining, we have shown that the large particles formed are the agglutinated leptospires. Quantification of the agglutination process has been achieved by calculating an agglutination factor (

A
f

), based on changes in the light scatter parameters measured by FCM.

A
f

enables us to determine the specificity of the serological reaction of the patient serum with each leptospiral serovar. In this work, 27 serum samples from 18 leptospirosis patients were tested by both the MAT and the FCM techniques, in which each serum sample was tested against 13 serovars. Twenty-six human serum samples derived from patients with a variety of other defined illnesses were used as negative controls and enabled us to define the

A
f

threshold value as <9.3 for negative patients, while any value higher than that would be a positive result for leptospirosis. Compared to MAT, the FCM technique was found to be more specific and sensitive, especially in identifying the serogroup in the acute phase of the disease. The whole process was found to be rapid and took less than 1.5 h. Moreover, FCM analysis is objective and can be automated for the handling of large numbers of samples.
Molecular Analysis of Adenovirus Isolates from Vaccinated and Unvaccinated Young Adults
Authors: Blasiole Daniel A. , Metzgar David , Daum Luke T. , Ryan Margaret A. K. , Wu Jianguo , Wills Christopher , Le Charles T. , Freed Nikki E. , Hansen Christian J. , Gray Gregory C. , Russell Kevin L.
Published: April 2004

Infections of adenovirus type 4 (Ad4) and Ad7 were discovered among previously vaccinated individuals through febrile respiratory illness surveillance at military recruit camps. Genetic analysis was performed on these isolates and a sample of adenovirus isolates from unvaccinated patients. Antigenic regions of the adenovirus hexon gene from 21 vaccinated and 31 unvaccinated patients were sequenced and compared to homologous regions of Ad4 and Ad7 vaccine strains and of other representative hexon sequences archived in GenBank. The phylogenetic distribution of sequences from vaccinated individuals closely resembled those from unvaccinated individuals. The most common Ad7 strain was the Ad7d2 hexon genotype, and the most common Ad4 strain was a genotype nearly identical to the recently discovered Z-G 95-873 Ad4 variant. Near exclusive isolation of Ad4 since 1999 indicates that the Ad4 variant is currently responsible for the vast majority of adenovirus morbidity in military recruit camps. Different ratios of nonsynonymous to synonymous nucleotide substitution rates in known antigenic regions compared to nonantigenic regions indicated positive selection for diversity in the antigenic regions and purifying selection in the nonantigenic regions.
Dual Toxin-Producing Strain of
Clostridium botulinum
Type Bf Isolated from a California Patient with Infant Botulism
Authors: Barash Jason R. , Arnon Stephen S.
Published: April 2004

A retrospective study of
Clostridium botulinum
strains isolated from patients from California with infant botulism identified the fourth known
C. botulinum
strain that produces both type B and type F botulinum toxins. This unique strain represented 0.12% of the California infant botulism case isolates from 1976 to 2003. The relative concentrations of type B and F toxins produced were temperature dependent.
Quality Control Limits for Voriconazole Disk Susceptibility Tests on Mueller-Hinton Agar with Glucose and Methylene Blue
Authors: Pfaller M. A. , Barry A. , Bille J. , Brown S. , Ellis D. , Meis J. F. , Rennie R. , Rinaldi M. , Rogers T. , Traczewski M.
Published: April 2004

An international collaborative study was performed in order to propose quality control limits for voriconazole disk diffusion tests on Mueller-Hinton agar with 2% glucose and 0.5 μg of methylene blue per ml. The supplement may be added to the agar before autoclaving, or Mueller-Hinton agar plates may be flooded with a glucose-methylene blue solution. Replicate tests on both types of agar plates with 1-μg voriconazole disks generated data to propose zone size limits for tests of
Candida parapsilosis
ATCC 22019 (28 to 37 mm),
Candida albicans
ATCC 90028 (31 to 42 mm), and
Candida krusei
ATCC 6258 (16 to 25 mm).
Candida tropicalis
ATCC 750 was not useful for this purpose.
Comparison of Two Widely Used PCR Primer Systems for Detection of
Toxoplasma
in Amniotic Fluid, Blood, and Tissues
Authors: Chabbert Elisabeth , Lachaud Laurence , Crobu Lucien , Bastien Patrick
Published: April 2004

The PCR diagnosis of toxoplasmosis suffers from lack of standardization. Interlaboratory comparative studies of PCR methods have been performed, but intralaboratory comparisons are scarce. Here, we optimized and compared the technical performances of two PCR primer systems widely used for
Toxoplasma
detection. The differences between the two methods were visible only at low parasite concentrations (≤1
Toxoplasma
genome per reaction tube). Nevertheless, when clinical samples were tested, both methods significantly differed in their technical sensitivities and specificities. Only one method appeared adequate for samples containing blood or tissue.
Genotyping of Enteric Adenoviruses by Using Single-Stranded Conformation Polymorphism Analysis and Heteroduplex Mobility Assay
Authors: Soares Caroline C. , Volotão Eduardo M. , Albuquerque Maria Carolina M. , Nozawa Carlos M. , Linhares Rosa Elisa C. , Volokhov Dmitriy , Chizhikov Vladimir , Lu Xiaoyan , Erdman Dean , Santos Norma
Published: April 2004

Single-stranded conformation polymorphism (SSCP) analysis and heteroduplex mobility assays (HMAs) were used to identify and genotype enteric adenoviruses (EAd). The results were compared to those of restriction endonuclease assays, species-specific PCRs, and direct nucleotide sequence analyses. Of the 31 stool samples tested, 15 isolates were identified as EAd and 7 were identified as nonenteric Ad by all methods. An agreement of 100% was found between the SSCP and HMA results.
Development of a Multiplex PCR Technique for Detection and Epidemiological Typing of
Salmonella
in Human Clinical Samples
Authors: Alvarez Juan , Sota Mertxe , Vivanco Ana Belén , Perales Ildefonso , Cisterna Ramón , Rementeria Aitor , Garaizar Javier
Published: April 2004

We have developed a multiplex PCR assay for
Salmonella
detection and epidemiological typing. Six sets of primers were designed to detect the major
Salmonella
serotypes and phage types in Spain. An internal amplification control was designed in order to detect PCR inhibition. The different amplification profiles obtained allowed us to detect
Salmonella
bacteria and to distinguish the clinically prevalent
Salmonella enterica
serotypes Enteritidis, Typhimurium and subspecies I serotype 4,5,12:i:−. Using this method, we could detect a specific band for DT104 and U302 phage types in
Salmonella
serotype Typhimurium.
Salmonella enterica
serotype Hadar and other C2 serogroup strains showed two specific band profiles. In the validation stage, the assay was reproducible for all serotypes studied, apart from some C2 serogroup strains. When the technique was applied to clinical stool specimens, the prevalent serotypes Enteritidis and Typhimurium were detected with a sensitivity of 93%, specificity of 100%, and efficiency of 98%. Also, a low PCR inhibition rate (8%) was obtained. The overall agreement of the multiplex PCR with conventional culture-based techniques was 95% for
Salmonella
typing using Cohen's kappa index.
DNA Polymorphism Assay Distinguishes Isolates ofLeishmania donovaniThat Cause Kala-Azar from Those That Cause Post-Kala-Azar Dermal Leishmaniasis in Humans
Authors: Sreenivas Gannavaram , Subba Raju B. V. , Singh Ruchi , Selvapandiyan Angamuthu , Duncan Robert , Sarkar Dwijen , Nakhasi Hira L. , Salotra Poonam
Published: April 2004

Leishmania donovaniin India causes visceral infection (kala-azar) and dermal infection (post-kala-azar dermal leishmaniasis). We report here the identification of polymorphism in a well-defined genetic locus among theLeishmaniaparasites causing the visceral and dermal manifestations, in a comparison of 15 post-kala-azar dermal leishmaniasis and 12 kala-azar patient isolates.
Multilocus Restriction Typing Method To Predict the Sequence Type of Meningococci
Authors: Dyet Kristin H. , Simmonds Robin S. , Martin Diana R.
Published: April 2004

A multilocus restriction typing (MLRT) method was developed to reduce the number of sequencing reactions required to determine the clonal relationships among serogroup B meningococci causing an epidemic in New Zealand. MLRT was a rapid, simple, and inexpensive method, and the results had an excellent correlation with multilocus sequence typing results.
Unique Variations of
pbp2b
Sequences in Penicillin-Nonsusceptible
Streptococcus pneumoniae
Isolates from Korea
Authors: Baek Jin Yang , Ko Kwan Soo , Oh Won Sup , Jung Sook-In , Kim Yeon Sook , Chang Hyun-Ha , Lee Hyuck , Kim Shin Woo , Peck Kyong Ran , Lee Nam Yong , Song Jae-Hoon
Published: April 2004

pbp2b
gene alterations were analyzed in 102 clinical isolates of
Streptococcus pneumoniae
(30 penicillin susceptible, 23 intermediate, and 49 resistant) from Korea. On the basis of PBP2B amino acid sequences, penicillin-nonsusceptible isolates of
S. pneumoniae
belonged to six groups, and 76% of the isolates in groups I to IV showed the same divergent block of amino acid alterations. Thirteen isolates (group II) also possessed a divergent block that was identical to that of
Streptococcus oralis
. The
pbp2b
genes of most Korean isolates showed novel mosaic mutations due to horizontal gene transfer. The Thr252→Ala substitution, previously thought to be associated only with penicillin-nonsusceptible strains, was also found in three penicillin-susceptible strains. On the basis of their
pbp2b
nucleotide sequences, all penicillin-nonsusceptible isolates can be detected by multiplex PCR, which can be used as a novel method for detection of antibiotic-resistant pneumococcal strains in clinical specimens.
First International Quality Assurance Study on the Rapid Detection of Viral Agents of Bioterrorism
Authors: Niedrig Matthias , Schmitz Herbert , Becker Stephan , Günther Stephan , ter Meulen Jan , Meyer Herman , Ellerbrok Heinz , Nitsche Andreas , Gelderblom Hans R. , Drosten Christian
Published: April 2004

We have conducted an international quality assurance study of filovirus, Lassa virus, and orthopox virus PCR with 24 participants. Of the participating laboratories, 45.8 and 66.7% detected virus in all plasma samples, which contained ≥5,000 and ≥100,000 copies per ml, respectively. Sensitivity levels were not significantly different between viruses. False-negative results were attributable to a lack of sensitivity.
Dissemination of a Clone of Unusual Phenotype of Pandrug-Resistant
Acinetobacter baumannii
at a University Hospital in Taiwan
Authors: Kuo Lu-Cheng , Teng Lee-Jene , Yu Chong-Jen , Ho Shen-Wu , Hsueh Po-Ren
Published: April 2004

From December 2002 to February 2003, 15 isolates of pandrug-resistant unidentified
Acinetobacter
species were recovered from seven patients treated on different wards or intensive care units. Both 16S-23S rRNA intergenic spacer PCR-restriction fragment length polymorphism profiles and sequence analysis of these isolates identified them as
Acinetobacter baumannii
. This pandrug-resistant
A. baumannii
strain with an unusual phenotype could persist in humans for long periods and was widely disseminated throughout the hospital.
PCR-Based Assay for Detection of
Neisseria meningitidis
Capsular Serogroups 29E, X, and Z
Authors: Bennett Désirée E. , Mulhall Robert M. , Cafferkey Mary T.
Published: April 2004

PCR-based assays for the identification of
Neisseria meningitidis
serogroups 29E, X, and Z by detection of specific regions of the
ctrA
gene are described. The specificities of these assays were confirmed using serogroups A, B, C, 29E, H, W135, X, Y, and Z and nongroupable meningococcal isolates.
Development and Comparison of Enzyme Immunoassays for Diagnosis of Chagas' Disease Using Fixed Forms of
Trypanosoma cruzi
(Epimastigotes, Amastigotes, and Trypomastigotes) and Assessment of Antigen Stability for the Three Assays
Authors: Berrizbietia Mariolga , Ndao Momar , Gottschalk Marcelo , Aché Alberto , Vásquez Fabio , Lacouture Sonia , Medina Mehudy , Ward Brian J.
Published: April 2004

Three enzyme immunoassays (EIAs) for diagnosis of Chagas' disease were developed with fixed forms of
Trypanosoma cruzi
using a panel of 435 sera from the following groups: Venezuelan subjects positive by immunofluorescence (
n
= 70), Venezuelan healthy controls (
n
= 85), healthy Canadians (
n
= 166), and subjects with other parasitic diseases (
n
= 114). All assays achieved 100% sensitivity and reasonable specificity for amastigotes (97.6%), epimastigotes (98.3%), and trypomastigotes (99.3%). The fixed-trypomastigote assay was stable over 4 months at 4°C and room temperature. These data suggest that a fixed-trypomastigote EIA may be a suitable candidate for blood bank screening.
Characterization of an Anonymous Molecular Marker Strongly Linked to
Escherichia coli
Strains Causing Neonatal Meningitis
Authors: Clermont Olivier , Bonacorsi Stéphane , Bingen Edouard
Published: April 2004

An anonymous 14.9-kb
rrn
-containing HindIII fragment is strongly linked to
Escherichia coli
strains causing neonatal meningitis. We show in this report that this fragment does not encode new virulence factors but lacks
arpA
, a gene common in avirulent
E. coli
strains, and we developed a PCR test to detect this fragment.
Detection of theEscherichia coliGroup 2 Polysaccharide Capsule Synthesis GenekpsMby a Rapid and Specific PCR-Based Assay
Authors: Johnson James R. , O'Bryan Timothy T.
Published: April 2004

A rapid and simple PCR-based assay for detection of the group 2 capsule synthesis genekpsMofEscherichia coliwas designed and validated. When combined with the published group 2 primers (kpsIIf, 5′-GCGCATTTGCTGATACTGTTG-3′; kpsIIr, 5′-CATCCAGACGATAAGCATGAGCA-3′), the new primers (the kpsIIf primer and a new reverse primer K2r, 5′-AGGTAGTTCAGACTCACACCT-3′) allowed specific identification by exclusion of the heretofore elusive K2kpsMvariant. The primers yielded the predicted amplicon when multiplexed with other primers and used under varied assay conditions, including a range of concentrations of individual reaction mixture ingredients and of annealing temperatures (from 54 to 64°C).
Usefulness of
Toxoplasma gondii
-Specific Recombinant Antigens in Serodiagnosis of Human Toxoplasmosis
Authors: Pietkiewicz H. , Hiszczyńska-Sawicka E. , Kur J. , Petersen E. , Nielsen H. V. , Stankiewicz M. , Andrzejewska I. , Myjak P.
Published: April 2004

Toxoplasma gondii
SAG1, GRA1, and GRA7 recombinant antigens may be regarded as tools for the detection of
T. gondii
immunoglobulin G antibodies in persons with chronic and acute toxoplasmosis. GRA7 is more correlated with acute toxoplasmosis. A combination of these antigens will increase the sensitivity of enzyme-linked immunosorbent assays.
Use of an Immunoglobulin G Avidity Test To Discriminate between Primary and Secondary Dengue Virus Infections
Authors: de Souza Vanda Akico Ueda Fick , Fernandes Silvana , Araújo Evaldo Stanislau , Tateno Adriana Fumie , Oliveira Olímpia M. N. P. F. , Oliveira Renato dos Reis , Pannuti Cláudio Sérgio
Published: April 2004

An enzyme-linked immunosorbent assay-based immunoglobulin G (IgG) antibody avidity test was evaluated by using sera from 57 patients with acute dengue infection. Overall, 55 of 57 patients were correctly classified (27 of 27 with primary dengue and 28 of 30 with secondary dengue). We conclude that the IgG avidity test can be useful for differentiating between acute, primary, and secondary dengue infections.
Emergence of
vanA
Genotype Vancomycin-Resistant Enterococci with Low or Moderate Levels of Teicoplanin Resistance in Korea
Authors: Eom Joong-Sik , Hwang In-Sook , Hwang Byung-Yoen , Lee Jae-Gab , Lee Yeon-Joo , Cheong Hee-Jin , Park Yong-Ho , Park Seung-Chul , Kim Woo-Joo
Published: April 2004

In Korea, vancomycin-resistant enterococci have become important nosocomial pathogens since the late 1990s, and most vancomycin-resistant enterococcal isolates have been VanA phenotype-
vanA
genotype strains. In 2001, we experienced an outbreak of VanB phenotype-
vanA
genotype vancomycin-resistant enterococci at a university hospital. This is the first report of VanB-
vanA
vancomycin-resistant enterococci from humans in Korea.
Multiplex PCR for Diagnosis of Enteric Infections Associated with Diarrheagenic
Escherichia coli
Authors: Vidal Roberto , Vidal Maricel , Lagos Rossana , Levine Myron , Prado Valeria
Published: April 2004

A multiplex PCR for detection of three categories of diarrheagenic
Escherichia coli
was developed. With this method, enterohemorrhagic
E. coli
, enteropathogenic
E. coli
, and enterotoxigenic
E. coli
were identified in fecal samples from patients with hemorrhagic colitis, watery diarrhea, or hemolytic-uremic syndrome and from food-borne outbreaks.
Modified Colorimetric Assay for Susceptibility Testing of Azole Antifungal Drugs against
Candida
Species
Authors: Chen Jian , Wan Zhe , Li Ruoyu
Published: April 2004

We modified a rapid susceptibility assay (RSA) for antifungal susceptibility testing of azoles based upon glucose utilization. This modified RSA method provides quantitative endpoint readings in 6 h. In this study, the modified RSA and the National Committee for Clinical Laboratory Standards M27-A methods were used to determine the MICs of fluconazole and itraconazole for 118
Candida
isolates. Yeast nitrogen base containing 0.12 g of glucose per liter was used for the modified RSA method. For fluconazole, agreement among assays within one or two twofold dilutions was 72.9 and 88.1%, respectively; for itraconazole, agreement within one or two twofold dilutions was 82.2 and 89.8%, respectively. These data suggest that the modified RSA method is a reliable and rapid method for azole antifungal susceptibility testing against
Candida
species.
Establishment of an Active Laboratory-Based Surveillance for Bacterial Meningitis in Croatia and Molecular Characterization of
Neisseria meningitidis
Isolates Causing Meningococcal Disease That Were Collected in the Year 2000, the First Year of Activity
Authors: Boras A. , Jeren T. , Sacchi C. T. , Schmink S. , Bozinovic D. , Barsic B. , Rosenstein N. E. , Popovic T.
Published: April 2004

In 2000, 23
Neisseria meningitidis
(meningococcal [Men]) isolates were collected in Croatia through an active laboratory-based surveillance for bacterial meningitis (17 Men serogroup B [MenB], 4 MenC, 1 MenW135, and 1 nongroupable isolate). Molecular characterization revealed a substantial level of diversity with only six isolates belonging to electrophoretic type 5 (ET-5) and ET-37 hypervirulent complexes.
Furunculosis Due to
Mycobacterium mageritense
Associated with Footbaths at a Nail Salon
Authors: Gira Amy K. , Reisenauer Amy H. , Hammock Lauren , Nadiminti Uma , Macy Jonathan T. , Reeves Ariane , Burnett Cindy , Yakrus Mitchell A. , Toney Sean , Jensen Bette J. , Blumberg Henry M. , Caughman S. Wright , Nolte Frederick S.
Published: April 2004

We report two cases of lower-extremity furunculosis caused by
Mycobacterium mageritense
. Both patients were patrons of the same nail salon, where they received footbaths prior to pedicures.
M. mageritense
bacteria isolated from two whirlpool footbaths were determined to be closely related to the patient isolates by pulsed-field gel electrophoresis.
Intraocular Detection of
Bartonella henselae
in a Patient with HLA-B27 Uveitis
Authors: Drancourt Michel , Bodaghi Bahram , Lepidi Hubert , Le Hoang Phuc , Raoult Didier
Published: April 2004

Bartonella henselae
uveitis was diagnosed in a 40-year-old woman with underlying HLA-B27 uveitis on the basis of immunodetection and molecular detection of the organism, a Marseilles genogroup and a CAL-1 genotype strain, in the vitreous fluid. This case illustrates that
B. henselae
should be included in the differential diagnosis of uveitis and the usefulness of immunodetection for rapid and specific diagnosis.
Mycotic Aneurysm Caused byStreptococcus constellatussubsp.constellatus
Authors: Chiang Wen-Chu , Tsai Jui-Chang , Chen Shey-Ying , Hsu Chiung-Yuan , Wu Chen-Tu , Teng Lee-Jene , Hsueh Po-Ren
Published: April 2004

An infected mycotic aneurysm due toStreptococcus constellatussubsp.constellatushas not previously been reported. We report on this condition in an 87-year-old woman who had aggravating abdominal pain and a large fusiform aneurysm over the thoracic-abdominal aorta with mural thrombus. Isolates from two sets of blood cultures and the debrided tissue were identified asS. constellatussubsp.constellatusby their biochemical reaction profiles, compatible 16S rRNA gene sequencing results, and sequencing results for the partialgroESLgene and the 16S-23S intergenic spacer region.
Detection of
Atopobium vaginae
in Postmenopausal Women by Cultivation-Independent Methods Warrants Further Investigation
Authors: Burton Jeremy P. , Devillard Estelle , Cadieux Peter A. , Hammond Jo-Anne , Reid Gregor
Published: April 2004

We sequenced 16S rRNA genes from the vaginal swab contents of a postmenopausal woman with asymptomatic bacterial vaginosis (BV). Sequences from
Atopobium vaginae
were the most commonly detected. In a survey of 35 other postmenopausal women, this organism was detected in 44% with BV but not in any subjects deemed healthy.
Genotyping and Antifungal Susceptibility Profile of
Dipodascus capitatus
Isolates Causing Disseminated Infection in Seven Hematological Patients of a Tertiary Hospital
Authors: Gadea Ignacio , Cuenca-Estrella Manuel , Prieto Elena , Diaz-Guerra Teresa M. , Garcia-Cia Jose I. , Mellado Emilia , Tomas Jose F. , Rodriguez-Tudela Juan L.
Published: April 2004

Seven cases of disseminated infection due to
Dipodascus capitatus
are reported. Infections occurred in a hematological unit of a tertiary hospital during a period of 5 years. Five cases were refractory to antifungal therapy. Antifungal susceptibility testing of seven isolates was performed, and strains were typed by PCR fingerprinting with the core sequence of phage M13 and by random amplification of polymorphic DNA with two primers, Ap12h and W-80A. A very short range of MICs of each antifungal agent was observed. The MICs of amphotericin B ranged between 0.50 and 2 μg/ml. Strains were susceptible in vitro to flucytosine and susceptible (dose-dependent) to fluconazole and itraconazole. Voriconazole exhibited an activity in vitro comparable to that of itraconazole. Typing techniques allowed seven additional isolates of
D. capitatus
neither geographically nor temporally related to be classified into two different genomic patterns. The genomic type of the seven strains from the hematological unit was identical regardless of typing technique utilized. It would indicate that the seven cases of disseminated infection could be related epidemiologically.
Chryseomonas luteola
Identified as the Source of Serious Infections in a Moroccan University Hospital
Authors: Chihab Wafae , Alaoui Ahmed S. , Amar Mohamed
Published: April 2004

Chryseomonas luteola
has only rarely been reported as a human bacterial pathogen. It has been shown that this organism in particular affects patients with health or indwelling disorders. Most reported cases showed septicemia, meningitis, endocarditis, or peritonitis. Two
C. luteola
infections observed in Morocco are described in the present study.
Linezolid-Resistant
Enterococcus faecalis
Isolated from a Cord Blood Transplant Recipient
Authors: Dibo Imad , Pillai Satish K. , Gold Howard S. , Baer Maria R. , Wetzler Meir , Slack James L. , Hazamy Peggy A. , Ball Donna , Hsiao Chiu Bin , McCarthy Philip L. , Segal Brahm H.
Published: April 2004

We report a linezolid-resistant
Enterococcus faecalis
infection in a cord blood stem cell transplant recipient previously treated with linezolid for bloodstream infections by vancomycin-resistant enterococci. Sequencing showed a G2576U mutation in the 23S rRNA gene. Because of the important niche of linezolid in cancer treatment, linezolid-resistant
E. faecalis
is noteworthy.
Measurement and analysis of excitation functions for alpha-particle-induced reactions in rhenium
Authors: Singh N L , Gadkari M S
Published: April 2004

Excitation functions of the reactions 185Re[(α,n); (α,2n); (α,3n)] and 187Re[(α,n); (α,2n); (α,3n); (α,4n)] were investigated up to 50 MeV using the stacked-foil activation technique and high-purity germanium γ-ray spectroscopy method. Since the natural rhenium used as a target has two odd-mass stable isotopes of abundance 37.4% (185Re) and 62.6% (187Re), their activation in some cases gives the same residual nucleus through different reaction channels, but with very different Q values. In such cases, the individual reaction cross sections are separated with the help of the ratio of theoretical cross sections. The experimental cross sections were compared with the theoretical predictions considering equilibrium as well as pre-equilibrium contributions using code ALICE/90. It was found that the initial exciton configuration n0 = 4 (4p0h) appears to give a good fit to the experimental data. To the best of our knowledge, the excitation functions for 185Re[(α,n); (α,2n); (α,3n)] and 187Re[(α,3n); (α,4n)] reactions were measured for the first time. PACS No.: 25.55.e
Analysis of electron-atom collision processes involving strong coupling
Authors: Bougouffa S , Kamli A
Published: April 2004

Inelastic electron  sodium atom collisions are investigated in the two-channel approximation. The scattering potentials are calculated using hydrogen-like wave functions for the valence electron of sodium. The coupled differential equations governing the collision process are solved using the Numerov numerical technique. The results thus obtained are in good agreement with those obtained by other sophisticated models. PACS Nos.: 34.50.Fa, 34.80.Dp
Unified treatment of overlap integrals with integer and noninteger n Slater-type orbitals using translational and rotational transformations for spherical harmonics
Authors: Guseinov I I , Mamedov B A
Published: April 2004

A unified treatment of two-center overlap integrals over Slater-type orbitals (STO) with integer and noninteger values of the principal quantum numbers is described. Using translation and rotation formulas for spherical harmonics, the overlap integrals with integer and noninteger n Slater-type orbitals are expressed through the basic overlap integrals and spherical harmonics. The basic overlap integrals are calculated using auxiliary functions Aσ and Bk. The analytical relations obtained in this work are especially useful for the calculation of overlap integrals for large integer and noninteger principal quantum numbers. The formulas established in this study for overlap integrals can be used for the construction of series expansions based on addition theorems. PACS Nos.: 31.15.p, 31.20.Ej
Coulomb amplitude representation and nuclear diffraction and refraction
Authors: Wojciechowski H
Published: April 2004

A new, consistent with wave-optics, method of treating diffraction and refraction in nuclear elastic scattering, is proposed and discussed. The method is based on the decomposition of the total elastic-scattering amplitude, written in a different form than usual, into the amplitude describing pure diffraction, and the rest, i.e., refraction and reflection. It is shown that nuclear diffraction is due to incompleteness of the Coulomb-scattering amplitude and is described by it. It is show that diffraction can be regarded as a kind of ``background'' always present, irrespective of the system being studied. PACS Nos.: 24.10.Ht, 42.25.Fx, 42.25.Gy