bluuebunny/crossref_metadata_2025_split huggingface dataset:
High-Throughput Method for Detecting Genomic-Deletion Polymorphisms
Authors: Goguet de la Salmonière Yves-Olivier Luc , Kim C. C. , Tsolaki A. G. , Pym A. S. , Siegrist M. S. , Small Peter M. 
Published: July 2004

ABSTRACT

DNA microarrays have been successfully used with different microorganisms, including
Mycobacterium tuberculosis
, to detect genomic deletions relative to a reference strain. However, the cost and complexity of the microarray system are obstacles to its widespread use in large-scale studies. In order to evaluate the extent and role of large sequence polymorphisms (LSPs) or insertion-deletion events in bacterial populations, we developed a technique, termed deligotyping, which hybridizes multiplex-PCR products to membrane-bound, highly specific oligonucleotide probes. The approach has the benefits of being low cost and capable of simultaneously interrogating more than 40 bacterial strains for the presence of 43 genomic regions. The deletions represented on the membrane were selected from previous comparative genomic studies and ongoing microarray experiments. Highly specific probes for these deletions were designed and attached to a membrane for hybridization with strain-derived targets. The targets were generated by multiplex PCR, allowing simultaneous amplifications of 43 different genomic loci in a single reaction. To validate our approach, 100 strains that had been analyzed with a high-density microarray were analyzed. The membrane accurately detected the deletions identified by the microarray approach, with a sensitivity of 99.9% and a specificity of 98.0%. The deligotyping technique allows the rapid and reliable screening of large numbers of
M. tuberculosis
isolates for LSPs. This technique can be used to provide insights into the epidemiology, genomic evolution, and population structure of
M. tuberculosis
and can be adapted for the study of other organisms.
Risk Factors for Sporadic Cryptosporidiosis among Immunocompetent Persons in the United States from 1999 to 2001
Authors: Roy Sharon L. , DeLong Stephanie M. , Stenzel Sara A. , Shiferaw Beletshachew , Roberts Jacquelin M. , Khalakdina Asheena , Marcus Ruthanne , Segler Suzanne D. , Shah Dipti D. , Thomas Stephanie , Vugia Duc J. , Zansky Shelley M. , Dietz Vance , Beach Michael J. 
Published: July 2004

ABSTRACT

Many studies have evaluated the role of
Cryptosporidium
spp. in outbreaks of enteric illness, but few studies have evaluated sporadic cryptosporidiosis in the United States. To assess the risk factors for sporadic cryptosporidiosis among immunocompetent persons, a matched case-control study was conducted in seven sites of the Foodborne Diseases Active Surveillance Network (FoodNet) involving 282 persons with laboratory-identified cryptosporidiosis and 490 age-matched and geographically matched controls. Risk factors included international travel (odds ratio [OR] = 7.7; 95% confidence interval [95% CI] = 2.7 to 22.0), contact with cattle (OR = 3.5; 95% CI = 1.8 to 6.8), contact with persons >2 to 11 years of age with diarrhea (OR = 3.0; 95% CI = 1.5 to 6.2), and freshwater swimming (OR = 1.9; 95% CI = 1.049 to 3.5). Eating raw vegetables was protective (OR = 0.5; 95% CI = 0.3 to 0.7). This study underscores the need for ongoing public health education to prevent cryptosporidiosis, particularly among travelers, animal handlers, child caregivers, and swimmers, and the need for further assessment of the role of raw vegetables in cryptosporidiosis.
Dissociation of Serum and Liver Hepatitis C Virus RNA Levels in Patients Coinfected with Human Immunodeficiency Virus and Treated with Antiretroviral Drugs
Authors: Furione Milena , Maserati Renato , Gatti Marta , Baldanti Fausto , Cividini Agostino , Bruno Raffaele , Gerna Giuseppe , Mondelli Mario U. 
Published: July 2004

ABSTRACT

We examined hepatitis C virus (HCV) RNA levels in serum, peripheral blood mononuclear cells (PBMC), and the liver for 135 patients with chronic HCV infections, 44 of whom were human immunodeficiency virus (HIV) positive and treated with highly active antiretroviral therapy (group A), 66 of whom were HIV negative (group B), with abnormal serum alanine aminotransferase (ALT) values, and 25 of whom were HIV negative, with ALT values of ≤1.5 times the normal value (group C). Patients had not been treated with interferon, with or without ribavirin, at the time of the study. A statistically significant correlation between HCV RNA levels in the liver and serum was reproducibly documented, whereas this was inconsistent for serum and PBMC. A comparative evaluation of HCV RNA levels in the liver and PBMC showed significantly lower values for group A than for groups B and C (
P
< 0.01 and
P
< 0.0001, respectively). In contrast, HCV RNA levels in serum were significantly higher for group A than for group B (
P
< 0.001). A dissociation between HCV RNA levels in serum and the liver was found for patients with HIV-HCV coinfections. Although the relative contribution of extrahepatic reservoirs, including lymphoid cells, to HCV RNA levels in serum is unclear, it may be speculated that a low intrahepatic HCV burden is caused by restored immunocompetence after successful antiretroviral therapy in coinfected patients.
High Prevalence of Hepatitis C Virus Type 5 in Central France Evidenced by a Prospective Study from 1996 to 2002
Authors: Henquell Cécile , Cartau Carole , Abergel Armand , Laurichesse Henri , Regagnon Christel , De Champs Christophe , Bailly Jean-Luc , Peigue-Lafeuille Hélène 
Published: July 2004

ABSTRACT

From 1996 to 2002, hepatitis C virus (HCV) typing was prospectively performed for 1,281 unselected HCV-infected and viremic patients, irrespective of their clinical status. Eighty-three patients (6.5%) were coinfected with human immunodeficiency virus (HIV) and HCV. A total of 1,195 strains were identified by a serotype screening (Murex HCV Serotyping 1-6 assay) and/or genotyping (Inno-LiPA HCV II) test. The distribution of HCV types showed an unusually high rate of type 5 (14.2%) that was stable over time and was the third most frequent type, after type 1 (59.1%) and type 3 (15.1%). HCV type 5 was more frequent in patients who were older than 50 (
P
= 10
−6
), but its frequency did not differ significantly by gender (
P
= 0.21). Serotyping was performed for 1,160 strains but failed for 30.2% of them. The efficiency depended on HIV status (for HCV-HIV-coinfected patients, half of the strains were untypeable) and HCV type. Genotyping was performed for 428 samples, with an overall efficiency of 99.3%. It failed in three cases, which were subsequently identified as HCV type 2. Serotyping and genotyping results for 39 patients showed discrepancies between the two methods for 4 patients, who had HCV type 2, type 6, or mixed infections. Thus, HCV type 5 may also be encountered frequently in Western countries. Its apparent confinement to a restricted area raises the question of how it emerged and underscores the need for further studies of HCV type prevalence, routes of transmission, pathogenicity, and responses to treatment.
Molecular Epidemiology of
Mycobacterium tuberculosis
in Western Sweden
Authors: Brudey Karine , Gordon Max , Moström Peter , Svensson Liselott , Jonsson Bodil , Sola Christophe , Ridell Malin , Rastogi Nalin 
Published: July 2004

ABSTRACT

The genetic diversity of
Mycobacterium tuberculosis
isolates among patients from Sweden was determined by a combination of two PCR-based techniques (spoligotyping and variable number of tandem repeats analysis). It resulted in a clustering of 23.6% of the isolates and a rate of recent transmission of 14.1%. The clustered isolates mainly belonged to the Haarlem family (23.2%), followed by the Beijing (9.8%), Latin American and Mediterranean (LAM; 8%), and East African-Indian (EAI; 6.2%) families. A comparison of the spoligotypes with those in the international spoligotyping database showed that 62.5% of the clustered isolates and 36.6% of all isolates typed were grouped into six major shared types. A comparison of the spoligotypes with those in databases for Scandinavian countries showed that 33% of the isolates belonged to an ill-defined T family, followed by the EAI (22%), Haarlem (20%), LAM (11%), Central Asian (5%), X (5%), and Beijing (4%) families. Both the highest number of cases and the proportion of clustered cases were observed in patients ages 15 to 39 years. Nearly 10% of the isolates were resistant to one or more drugs (essentially limited to isoniazid monoresistance). However, none of the strains were multidrug resistant. Data on the geographic origins of the patients showed that more than two-thirds of the clustered patients with tuberculosis were foreign-born individuals or refugees. These results are explained on the basis of both the historical links within specific countries and recently imported cases of tuberculosis into Sweden.
Use of In Vivo-Generated Biofilms from Hemodialysis Catheters To Test the Efficacy of a Novel Antimicrobial Catheter Lock for Biofilm Eradication In Vitro
Authors: Kite P. , Eastwood K. , Sugden S. , Percival S. L. 
Published: July 2004

ABSTRACT

This biofilm study was conducted to assess the in vitro activity of tetrasodium EDTA on catheters that had been routinely removed from hemodialysis patients at Leeds Teaching Hospitals Trust due to maturation of fistula. Catheters were screened by culture of through-catheter flush, and isolates were identified by standard methodologies; 20 isolates were found to be biofilm positive. Initial biofilm cell count levels averaged above 10
5
CFU/1-cm catheter section. Bacteria identified in the biofilms were gram-negative (1 isolate), gram-positive (11 isolates), or mixed species (8 isolates). After a 24-h lock, 40 mg of tetrasodium EDTA per ml was effective at eradicating the total biofilm viable count in almost all cases. The efficacy of tetrasodium EDTA as a catheter lock potentially shows that this agent could substantially reduce catheter-related infections and be used to treat patients with limited access.
Further Standardization of Broth Microdilution Methodology for In Vitro Susceptibility Testing of Caspofungin against
Candida
Species by Use of an International Collection of More than 3,000 Clinical Isolates
Authors: Pfaller M. A. , Messer S. A. , Boyken L. , Rice C. , Tendolkar S. , Hollis R. J. , Diekema D. J. 
Published: July 2004

ABSTRACT

The influence of test variables on in vitro susceptibility testing of caspofungin was examined with 694 isolates of
Candida albicans
including seven laboratory-derived glucan synthesis mutants. The conditions providing the greatest separation between the mutant strains and the clinical isolates were RPMI medium, MIC end point criterion of partial inhibition, and incubation for 24 h. These testing conditions were then applied to 3,322 isolates of
Candida
spp. (3,314 clinical isolates and eight glucan synthesis mutants). Among the 11 isolates for which caspofungin MICs were ≥2 μg/ml, eight were accounted for by the glucan synthesis mutants. The MICs for >99% of isolates were ≤1 μg/ml, and thus these isolates were differentiated from strains with reduced in vitro and in vivo susceptibilities to caspofungin.
Cross-Resistance between Fluconazole and Ravuconazole and the Use of Fluconazole as a Surrogate Marker To Predict Susceptibility and Resistance to Ravuconazole among 12,796 Clinical Isolates of
Candida
spp
Authors: Pfaller M. A. , Messer S. A. , Boyken L. , Rice C. , Tendolkar S. , Hollis R. J. , Diekema D. J. 
Published: July 2004

ABSTRACT

Cross-resistance within a class of antimicrobial agents is a problem that is often encountered with antibacterial agents, and it is also an issue with antifungal agents. A current example is ravuconazole, a new triazole antifungal with an expanded spectrum and potency against
Candida
spp.,
Aspergillus
spp., and other opportunistic fungal pathogens. The present study addresses the issue of cross-resistance between fluconazole and ravuconazole and the use of fluconazole as a surrogate marker to predict the susceptibility of
Candida
spp. to ravuconazole. Reference broth microdilution MIC results for 12,796 strains of
Candida
spp. isolated from more than 200 medical centers worldwide were used. Ravuconazole MICs and tentative interpretive categories (susceptible, ≤1 μg/ml; resistant, ≥2 μg/ml) were compared with those of fluconazole by using regression statistics and error rate bounding analyses. For all 12,796 isolates, the absolute categorical agreement rate was 92.5% (rate of false-susceptible results, or very major errors [VME], 0.1%). Ravuconazole was active (MIC, ≤1 μg/ml) against 99.9% of the fluconazole-susceptible isolates, 96% of the fluconazole-susceptible dose-dependent isolates, and 49% of the fluconazole-resistant isolates, including 99% of the
Candida krusei
isolates. Since ravuconazole is 16- to 32-fold more potent than fluconazole, the performance of fluconazole as a surrogate marker for ravuconazole susceptibility was improved by designating those isolates with fluconazole MICs of ≤32 μg/ml susceptible to ravuconazole, resulting in a categorical agreement rate of 98.3%, with a VME rate of 0.3% (99 and 0.4%, respectively, when
C. krusei
was omitted). Cross-resistance between fluconazole and ravuconazole applies most directly to fluconazole-resistant
Candida glabrata
and is variable among other species of
Candida
. Fluconazole may serve as a surrogate marker to predict the susceptibility of
Candida
spp. to ravuconazole.
Evaluation of the Hyplex BloodScreen Multiplex PCR-Enzyme-Linked Immunosorbent Assay System for Direct Identification of Gram-Positive Cocci and Gram-Negative Bacilli from Positive Blood Cultures
Authors: Wellinghausen Nele , Wirths Beate , Essig Andreas , Wassill Lars 
Published: July 2004

ABSTRACT

We evaluated the Hyplex BloodScreen PCR-enzyme-linked immunosorbent assay (ELISA) system (BAG, Lich, Germany), a new diagnostic test for the direct identification of gram-negative bacilli and gram-positive cocci from positive blood cultures, with 482 positive BACTEC 9240 blood culture bottles. The test involves amplification of the bacterial DNA by multiplex PCR and subsequent hybridization of the PCR product to specific oligonucleotide probes in an ELISA-based format. The available probes allow the separate detection of
Escherichia coli
,
Pseudomonas aeruginosa
,
Enterobacter aerogenes
,
Klebsiella
spp.,
Staphylococcus aureus
,
Staphylococcus epidermidis
,
Enterococcus faecalis
/
Enterococcus faecium
,
Streptococcus pyogenes
, and
Streptococcus pneumoniae
and the staphylococcal
mecA
gene. The Hyplex BloodScreen test showed an overall sensitivity of 100% for the identification of gram-negative bacilli and 96.6 to 100% for the identification of gram-positive cocci (
S. aureus
, 100%;
S. epidermidis
, 97.2%;
Enterococcus faecalis/Enterococcus faecium
, 96.6%; and
Streptococcus pneumoniae
, 100%). The specificities of the test modules ranged from 92.5 to 100% for gram-negative bacilli and 97.7 to 100% for gram-positive cocci (
Escherichia coli
, 92.5%;
Pseudomonas aeruginosa
, 98.5%;
Klebsiella
spp., 100%;
Enterobacter aerogenes
, 100%;
S. aureus
, 100%,
S. epidermidis
, 97.7%;
Enterococcus faecalis/Enterococcus faecium
, 99.6%;
Streptococcus pyogenes
, 100%; and
Streptococcus pneumoniae
, 99.3%). The result of the
mecA
gene detection module correlated with the result of the phenotypic oxacillin resistance testing in all 38 isolates of
Staphylococcus aureus
investigated. In conclusion, the Hyplex BloodScreen PCR-ELISA system is well suited for the direct and specific identification of the most common pathogenic bacteria and the direct detection of the
mecA
gene of
Staphylococcus aureus
in positive blood cultures.
Comparison of Different Methods of Isolation of DNA of Commonly Encountered
Candida
Species and Its Quantitation by Using a Real-Time PCR-Based Assay
Authors: Maaroufi Younes , Ahariz Naïma , Husson Mireille , Crokaert Françoise 
Published: July 2004

ABSTRACT

Molecular diagnosis based on genomic amplification methods such as real-time PCR assay has been reported as an alternative to conventional culture for early detection of invasive candidiasis. However, a major limitation of the molecular method is the difficulty associated with breaking fungal cell walls since the DNA extraction step still requires more than half of a working day. It has been suggested that PCR detection of free template DNA in serum is preferred over the use of whole blood for the diagnosis of systemic candidiasis. In this study, two conventional procedures (the first [the HLGT method] consists of boiling sera in an alkaline guanidine-phenol-Tris reagent, and the second [the PKPC method] uses proteinase K digestion, followed by organic extraction) and three commercially available kits for DNA isolation were evaluated for sensitivity, purity, cost, and use of template for most clinically important
Candida
species in a TaqMan-based PCR assay. To optimize these procedures, we evaluated the effect of adding 0.5% bovine serum albumin to DNA extracts and found that it decreased the effects of inhibitors. The QIAamp DNA blood kit did significantly shorten the duration of the DNA isolation but was among the most expensive procedures. Furthermore, the QIAamp DNA blood kit proved to be as sensitive as the HLGT DNA isolation method for PCR amplification from 52 serum samples from hematology or oncology patients with clinically proven or suspected systemic
Candida
infections. All PCR-positive samples showed approximately the same
Candida
species load by both procedures (100% correspondence), whereas one discordant result was obtained between PCR and blood culture.
Multiplex Real-Time PCR for Detection of
Anaplasma phagocytophilum
and
Borrelia burgdorferi
Authors: Courtney Joshua W. , Kostelnik Leah M. , Zeidner Nordin S. , Massung Robert F. 
Published: July 2004

ABSTRACT

A multiplex real-time PCR assay was developed for the simultaneous detection of
Anaplasma phagocytophilum
and
Borrelia burgdorferi
. The assay was tested on various
Anaplasma
,
Borrelia
,
Erhlichia
, and
Rickettsia
species, as well as on
Bartonella henselae
and
Escherichia coli
, and the assay was found to be highly specific for
A. phagocytophilum
and the
Borrelia
species tested (
B. burgdorferi
,
B. parkeri
,
B. andersonii
, and
B. bissettii
). The analytical sensitivity of the assay is comparable to that of previously described nested PCR assays (
A. phagocytophilum
, 16S rRNA;
B. burgdorferi
,
fla
gene), amplifying the equivalent of one-eighth of an
A. phagocytophilum
-infected cell and 50 borrelia spirochetes. The dynamic range of the assay for both
A. phagocytophilum
and
B. burgdorferi
was ≥4 logs of magnitude. Purified DNA from
A. phagocytophilum
and
B. burgdorferi
was spiked into DNA extracted from uninfected ticks and from negative control mouse and human bloods, and these background DNAs were shown to have no significant effect on sensitivity or specificity of the assay. The assay was tested on field-collected
Ixodes scapularis
ticks and shown to have 100% concordance compared to previously described non-probe-based PCR assays. To our knowledge, this is the first report of a real-time multiplex PCR assay that can be used for the simultaneous and rapid screening of samples for
A. phagocytophilum
and
Borrelia
species, two of the most common tick-borne infectious agents in the United States.
Multiplex PCR Assay for Detection of
Streptococcus suis
Species and Serotypes 2 and 1/2 in Tonsils of Live and Dead Pigs
Authors: Marois C. , Bougeard S. , Gottschalk M. , Kobisch M. 
Published: July 2004

ABSTRACT

A PCR assay was developed for the detection of
Streptococcus suis
serotypes 2 and 1/2. This multiplex PCR is based on the amplification of the gene coding for 16S rRNA of
S. suis
and on the amplification of the
cps2J
gene coding for the capsule of
S. suis
serotypes 2 and 1/2. An internal control was constructed and added in this test to monitor the efficiency of amplification in each reaction. To evaluate the specificity of the test, 31 strains of other bacterial species related to
S. suis
or isolated from pigs and 42 strains of
S. suis
serotypes 1 and 3 to 34 were analyzed. The detection threshold of the test was 28
S. suis
CFU/ml. The specificity and the sensitivity of the multiplex PCR test and the presence of an internal control allowed the analysis of biological samples without a culture step. The PCR assay was then applied to the detection of 14
S. suis
serotype 1/2 strains, 88
S. suis
serotype 2 strains isolated from pigs, and 25
S. suis
serotype 2 strains isolated from humans. This test was also applied to analyze tonsil samples of pigs experimentally infected and carrier pigs without any symptoms.
Comparison of Three Commercially Available Serologic Assays Used To Detect Human Parvovirus B19-Specific Immunoglobulin M (IgM) and IgG Antibodies in Sera of Pregnant Women
Authors: Butchko Allyson R. , Jordan Jeanne A. 
Published: July 2004

ABSTRACT
A split-sample study was conducted to evaluate the performances of three enzyme immunoassays (EIAs) utilizing one or more conformational antigens to detect human parvovirus B19 (B19V)-specific immunoglobulin M (IgM) or IgG in the sera of 198 pregnant women. We compared EIAs available from Biotrin International, Inc. (Dublin, Ireland), Medac Diagnostika (Wedel, Germany), and Mikrogen (Martinsried, Germany). Specimens with discordant results were analyzed further using an immunofluorescence assay (Biotrin). Equivocal data accounted for close to half of all the discrepant results for both IgM and IgG, with 7 of 15 discrepant results from the Medac and Mikrogen kits involving equivocal data and the Biotrin kit giving a single equivocal result. For each specimen, a consensus was established from the four test results if agreement occurred among at least three of four results. Overall, the highest percentage of agreement with the consensus results was seen when Biotrin kits were used; 194 (100%) of 194 and 194 (99.5%) of 195 results for IgM and IgG, respectively, agreed with the consensus results. When Medac kits were used, 189 (97.4%) of 194 and 191 (97.9%) of 195 results for IgM and IgG, respectively, agreed with the consensus, and when Mikrogen kits were used, 179 (92.3%) of 194 and 193 (99%) of 195 results for IgM and IgG, respectively, agreed with the consensus. Given the consensus results, the Medac EIA appeared to generate presumed false-positive results for IgM and the Mikrogen EIA appeared to generate presumed false-positive results for IgG and IgM. In summary, the Biotrin EIAs produced far fewer equivocal results than the other assays and results of the Biotrin EIAs agreed more often with the consensus results than did those of the other commercially available EIAs for detecting B19V-specific IgM and IgG antibodies.