bluuebunny/crossref_metadata_2025 huggingface dataset:
Heterostructures for achieving large responsivity in InAs/GaAs quantum dot infrared photodetectors
Authors: Chakrabarti S. , Stiff-Roberts A. D. , Bhattacharya P. , Kennerly S. W.
Published: July 2004

Quantum dot infrared photodetectors (QDIPs) offer normal-incidence detection, low dark current, high operating temperature, and multi-wavelength detection. In an effort to get better responsivity in QDIPs at higher operating temperatures, we have grown, fabricated, and characterized two different QDIP heterostructures. The first is a device with 70 quantum dot layers. As a result, the peak responsivity is measured to be 0.12 A/W for Vbias=2.0 V and T=175 K. The second device has ten layers of InAs quantum dots embedded in an AlAs/GaAs superlattice. As a result of the subsequent large dot density and increased carrier confinement, the peak responsivity is measured to be 2.5 A/W for Vbias=−1.5 V and T=78 K.
Analysis of atomic structure in InAs quantum dashes grown on AlGaAsSb metamorphic buffers
Authors: Balakrishnan Ganesh , Huang Shenghong , Dawson L. R. , Huffaker D. L.
Published: July 2004

We describe structural analysis at the atomic level of InAs quantum dashes (Qdashes) grown on a GaAs substrate using an AlGaAsSb metamorphic buffer. Such characterization methods reveal the strain distribution within the quantum feature and provide insight into formation processes. The Qdashes are studied using high-resolution transmission electron microscopy that allows real-space measurement of the atomic lattice constant, a0. The variation of the lattice constant is measured along both the [110] and the [1-10] directions in the asymmetric Qdash and indicates a highly tensile core surrounded by a compressively strained matrix. The study involves both capped and uncapped dashes to observe the effect of the encapsulating matrix on the lattice constants in the dash.
Reduced interface reaction during the epitaxial Fe growth on InAs for high efficiency spin injection
Authors: Yoh Kanji , Ohno Hiroshi , Sueoka Kazuhisa , Ramsteiner Manfred E.
Published: July 2004

We have investigated Fe/InAs interfaces for two different growth temperatures of Fe and the effect on the spin injection properties through an Fe/InAs junction. Secondary ion mass spectroscopy and transmission electron microscopy studies of the Fe/InAs interfaces revealed that Fe films grown at 175 °C clearly suffer from increased reaction and out-diffusion of semiconductor constituents compared to those grown at 23 °C. The lower temperature samples showed an increased degree of spin polarization of 18%–20% which translates to 36%–40% of spin injection efficiency assuming selection rules between heavy and light holes in the p-type InAs substrate. It is close to the spin polarization of 40%–45% in an Fe spin injector itself.
ZnO-based thin-film transistors of optimal device performance
Authors: Bae H. S. , Im Seongil
Published: July 2004

We report on ZnO-based thin-film transistors (TFTs) fabricated using SiO2/p-Si substrates on which their ZnO channel layers have been deposited by rf sputtering at various temperatures: room temperature (RT), 100, and 200 °C. When they went through rapid thermal annealing in forming gas ambient (H2:N2=1:10) for n-type doping, the highest field effect mobility of ∼1.93 cm2/V s was achieved from ZnO-TFTs prepared using the deposition temperature of 200 °C while a low mobility (∼0.2 cm2/V s) was from TFTs using RT-deposited ZnO. However, the TFT sample using 200 °C deposited ZnO, unlike the others, revealed a very large amount of off-state current resulting in the on/off current ratio of only ∼102. It is because the ZnO layer deposited at 200 °C is too conductive to act as an optimum TFT channel. It is concluded that optimal ZnO-based TFTs are obtained using the medium deposition temperature of 100 °C.
Cyclic AMP-Induced Repair of Zebrafish Spinal Circuits
Authors: Bhatt Dimple H. , Otto Stefanie J. , Depoister Brett , Fetcho Joseph R.
Published: July 2004

Neurons in the human central nervous system (CNS) are unable to regenerate, as a result of both an inhibitory environment and their inherent inability to regrow. In contrast, the CNS environment in fish is permissive for growth, yet some neurons still cannot regenerate. Fish thus offer an opportunity to study molecules that might surmount the intrinsic limitations they share with mammals, without the complication of an inhibitory environment. We show by in vivo imaging in zebrafish that post-injury application of cyclic adenosine monophosphate can transform severed CNS neurons into ones that regenerate and restore function, thus overcoming intrinsic limitations to regeneration in a vertebrate.
Frataxin Acts as an Iron Chaperone Protein to Modulate Mitochondrial Aconitase Activity
Authors: Bulteau Anne-Laure , O'Neill Heather A. , Kennedy Mary Claire , Ikeda-Saito Masao , Isaya Grazia , Szweda Luke I.
Published: July 2004

Numerous degenerative disorders are associated with elevated levels of prooxidants and declines in mitochondrial aconitase activity. Deficiency in the mitochondrial iron-binding protein frataxin results in diminished activity of various mitochondrial iron-sulfur proteins including aconitase. We found that aconitase can undergo reversible citrate-dependent modulation in activity in response to pro-oxidants. Frataxin interacted with aconitase in a citrate-dependent fashion, reduced the level of oxidant-induced inactivation, and converted inactive [3Fe-4S]
1+
enzyme to the active [4Fe-4S]
2+
form of the protein. Thus, frataxin is an iron chaperone protein that protects the aconitase [4Fe-4S]
2+
cluster from disassembly and promotes enzyme reactivation.
Immune Therapy for Autoimmune Diseases
Authors: Steinman Lawrence
Published: July 2004

Our increasing understanding of the pathophysiology of autoimmune disease has revealed a number of checkpoints that can be targeted with immune therapy, including key mediators of lymphocyte adhesion and migration, destructive cytokines involved in tissue damage, and the complex of molecules critical in the presentation of self-antigen and the activation of autoaggressive T lymphocytes. In many organ-specific autoimmune diseases, the identity of the molecules attacked by T cells and autoantibodies is known and attempts are under way to tolerize the immune system with a high level of specificity to these targets.
Evolution of Vaccinia Virus-Specific CD8
+
Cytotoxic T-Lymphocyte Responses in Primary Vaccinees and Revaccinees
Authors: Tenorio Allan R. , Peeples Mark E. , Patri Manokiran , Rezai Katayoun , Trenholme Gordon M.
Published: July 2004

Determination of successful vaccination with vaccinia virus is based on visual confirmation of a dermal response (take). Some revaccinees do not manifest a take, which may be due to a preexisting immunity rather than to poor technique or inadequate virus. Cytotoxic T-lymphocyte (CTL) response appears to be the most important immune defense in limiting response to vaccination. We evaluated vaccinia virus-specific CTL responses in revaccinees. Subjects with and without takes displayed comparable CTL responses. Vaccinia virus-specific CD8
+
CTL responses may be useful in interpreting the response to vaccination, particularly in individuals who are revaccinated and have difficult-to-interpret visual takes.
Novel Hairpin-Shaped Primer Assay To Study the Association of the −44 Single-Nucleotide Polymorphism of theDEFB1Gene with Early-Onset Periodontal Disease
Authors: Boniotto Michele , Hazbón Manzour Hernando , Jordan William James , Lennon Greig Patrick , Eskdale Joyce , Alland David , Gallagher Grant
Published: July 2004

A powerful, cost-effective new method for studying single-nucleotide polymorphisms (SNPs) is described. This method is based on the use of hairpin-shaped primers (HP), which give a sensitive and specific PCR amplification of each specific allele, without the use of costly fluorophore-labeled probes and any post-PCR manipulation. The amplification is monitored in real-time using SYBR Green I dye and takes only 2 h to yield results. The HP assay has a simple design and utilizes a conventional real-time PCR apparatus. The −44 C→G transversion in theDEFB1gene (which encodes human β-defensin 1) has been previously associated withCandidacarriage in oral epithelia. In this study, we analyzed the association between early-onset periodontal disease (EOP) and the −44 SNP. We used an HP assay to study the distribution of the −44 SNP in 264 human DNAs obtained from two cohorts of EOP patients and healthy controls from different ethnic backgrounds. The results indicate that the −44 SNP has a similar distribution between EOP and healthy patients, suggesting that it is not associated with the disease.
Impact of Serological Methodology on Assessment of the Link between
Chlamydia pneumoniae
and Vascular Diseases
Authors: Maraha Boulos , den Heijer Martin , Kluytmans Jan , Peeters Marcel
Published: July 2004

We assessed the impact of five serologic tests on the link between
Chlamydia pneumoniae
and abdominal aortic aneurysms (AAA). The results of the tests were inconsistent. Agreement among the five tests was generally poor. Detection of the link between
C. pneumoniae
and AAA depends on the serologic methodology chosen.
Evaluation of
Clonorchis sinensis
Recombinant 7-Kilodalton Antigen for Serodiagnosis of Clonorchiasis
Authors: Zhao Qin-Ping , Moon Sung-Ung , Lee Hyeong-Woo , Na Byoung-Kuk , Cho Seung-Yull , Kong Yoon , Jiang Ming-Sen , Li Ai-Hua , Kim Tong-Soo
Published: July 2004

The diagnostic applicability of the
Clonorchis sinensis
recombinant 7-kDa protein was evaluated. In enzyme-linked immunosorbent assays and immunoblots, the protein showed high sensitivities (81.3 and 71.9%, respectively) and specificities (92.6 and 89.7%, respectively) for sera obtained from various helminthic infections. Some paragonimiasis sera showed cross-reactions. The antigen might be valuable in the serodiagnosis of human clonorchiasis.
Bacteremia in an Immunocompromised Patient Caused by a Commensal
Neisseria meningitidis
Strain Harboring the Capsule Null Locus (
cnl
)
Authors: Vogel Ulrich , Claus Heike , von Müller Lutz , Bunjes Donald , Elias Johannes , Frosch Matthias
Published: July 2004

We recently described the capsule null locus (
cnl
) of constitutively unencapsulated
Neisseria meningitidis
clonal lineages.
cnl
meningococci were recovered from healthy carriers at high frequency. We here report on the first case of invasive disease caused by
cnl
meningococci in a severely immunosuppressed patient with chronic graft-versus-host disease after allogeneic peripheral blood stem cell transplantation. The sequence type 845 strain was extensively typed and, furthermore, shown to be sensitive to serum bactericidal activity.
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

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

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

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

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

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

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

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

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

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

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

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

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

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.
Antigenic Evidence of Prevalence and Diversity ofMycobacterium tuberculosisArabinomannan
Authors: Glatman-Freedman Aharona , Casadevall Arturo , Dai Zongdong , Jacobs William R. , Li Anping , Morris Sheldon L. , Navoa Josepine Anne D. , Piperdi Sajida , Robbins John B. , Schneerson Rachel , Schwebach J. Reid , Shapiro Michael
Published: July 2004

Arabinomannan (AM) is a polysaccharide of the mycobacterial capsule. The capsular polysaccharides of various microorganisms are diverse, and this diversity is important for classification of organisms into serotypes and vaccine development. In the present study we examined the prevalence and diversity of AM amongMycobacterium tuberculosisstrains using four AM-binding monoclonal antibodies (MAbs). One of these MAbs, MAb 9d8, is known to bind to AM specifically. By whole-cell enzyme-linked immunosorbent assay (ELISA), the AM recognized by MAb 9d8 was detected on the surfaces of 9 of 11 strains, while 2 strains showed no reactivity with MAb 9d8. However, the AM recognized by MAb 9d8 was found in the culture supernatants of all 11M. tuberculosisstrains tested, as demonstrated by capture ELISA. Other AM-binding MAbs reacted both with the surfaces and with the culture supernatants of all 11 strains. Mice immunized with an experimental AM-recombinantPseudomonas aeruginosaexoprotein A (rEPA) conjugate vaccine had an increased antibody response to AM and a moderate reduction in the numbers of CFU in their organs 7 days after challenge. Our results indicate that AM was detected in allM. tuberculosisstrains tested, with differences in epitope distributions of certain strains. In addition, our results suggest that an experimental AM-rEPA vaccine has a moderate effect on the numbers of CFU in organs early after infection.
Detailed Protocol for Purification of
Chlamydia pneumoniae
Elementary Bodies
Authors: Mukhopadhyay Sanghamitra , Clark Alejandra P. , Sullivan Erin D. , Miller Richard D. , Summersgill James T.
Published: July 2004

A detailed protocol for the growth and harvest of purified elementary bodies of
Chlamydia pneumoniae
is presented. This procedure utilizes a flask-to-flask passage scheme designed to achieve high bacterial titers in a short period of time.
Phenotypic and Molecular Characterization of Erythromycin Resistance in Four Isolates of
Streptococcus
-Like Gram-Positive Cocci Causing Bacteremia
Authors: Woo Patrick C. Y. , To Amanda P. C. , Lau Susanna K. P. , Fung Ami M. Y. , Yuen Kwok-yung
Published: July 2004

Among nine patients with bacteremia caused by
Granulicatella
or
Gemella
in a 6-year period (July 1995 to June 2001), three had bacteremia caused by erythromycin-resistant
Granulicatella adiacens
and one had bacteremia caused by erythromycin-resistant
Gemella haemolysans
. All four isolates possessed
mef
genes, whereas none possessed
ermT
,
ermTR
, or
ermB
genes.
Real-Time PCR Assay for Detection and Genotype Differentiation of
Giardia lamblia
in Stool Specimens
Authors: Guy Rebecca A. , Xiao Chongxie , Horgen Paul A.
Published: July 2004

Real-time PCR, using dual-labeled fluorescent probes targeting the β-giardin gene, was used to detect
Giardia lamblia
in human stool specimens and to discriminate between isolates from the two major genetic assemblages of
G. lamblia
infective to humans, assemblages A and B.
Evaluation of Indirect Fluorescence Antibody Assay for Detection of
Bartonella clarridgeiae
and Seroprevalence of
B. clarridgeiae
among Patients with Suspected Cat Scratch Disease
Authors: Tsuneoka Hidehiro , Umeda Akiko , Tsukahara Masato , Sasaki Kohsuke
Published: July 2004

The possibility of
Bartonella clarridgeiae
being a causative agent of cat scratch disease (CSD) was investigated by using indirect fluorescence antibody assays with 288 suspected CSD patients. Immunoglobulin G antibody to noncocultivated
B. clarridgeiae
was suitable only for detection of
B. clarridgeiae
antibody. Significant cross-reactivity between
Bartonella henselae
and
B. clarridgeiae
was noted, and no CSD case caused by
B. clarridgeiae
was detected.
Use of Two Sensitive and Specific Immunoblot Markers, Em70 and Em90, for Diagnosis of Alveolar Echinococcosis
Authors: Korkmaz Metin , Inceboz Tonay , Celebi Fehmi , Babaoglu Aylin , Uner Ahmet
Published: July 2004

Antibodies against
Echinococcus multilocularis
metacestodes were screened by immunoblotting sera from patients with alveolar echinococcosis (
n
= 39), cystic echinococcosis (
n
= 109), or other parasitic infections (
n
= 66) and healthy individuals (
n
= 32). Two antigens, approximately 70 and 90 kDa, are found to be valuable for confirmatory diagnosis, with a sensitivity and specificity of 100 and 99.51%, respectively.
Quality Control Guidelines for BAL9141 (Ro 63-9141), an Investigational Cephalosporin, When Reference MIC and Standardized Disk Diffusion Susceptibility Test Methods Are Used
Authors: Anderegg T. R. , Jones R. N. , Sader H. S.
Published: July 2004

BAL9141 is a novel cephalosporin with a broad spectrum of activity, including activity against methicillin-resistant staphylococci. This multicenter study was performed to establish quality control (QC) guidelines for susceptibility testing of BAL9141 in phase 3 clinical trials and after U.S. Food and Drug Administration approval. The proposed 3 or 4 log
2
dilution MIC ranges encompass 97.8 to 100.0% of reported results, while the proposed 7- to 9-mm-zone-diameter QC ranges included 95.2 to 99.4% of the participant-reported disk diffusion results.
Inhibition of CFTR channels by a peptide toxin of scorpion venom
Authors: Fuller Matthew D. , Zhang Zhi-Ren , Cui Guiying , Kubanek Julia , McCarty Nael A.
Published: July 2004

Peptide toxins have been valuable probes in efforts to identify amino acid residues that line the permeation pathway of cation-selective channels. However, no peptide toxins have been identified that interact with known anion-selective channels such as the cystic fibrosis transmembrane conductance regulator (CFTR). CFTR channels are expressed in epithelial cells and are associated with several genetic disorders, including cystic fibrosis and polycystic kidney disease. Several organic inhibitors have been used to investigate the structure of the Cl− permeation pathway in CFTR. However, investigations of the wider cytoplasmic vestibule have been hindered by the lack of a high-affinity blocker that interacts with residues in this area. In this study we show that venom of the scorpion Leiurus quinquestriatus hebraeus reversibly inhibits CFTR, in a voltage-independent manner, by decreasing single-channel mean burst duration and open probability only when applied to the cytoplasmic surface of phosphorylated channels. Venom was able to decrease burst duration and open probability even when CFTR channels were locked open by treatment with either vanadate or adenosine 5′-(β,γ-imido)triphosphate, and block was strengthened on reduction of extracellular Cl− concentration, suggesting inhibition by a pore-block mechanism. Venom had no effect on ATP-dependent macroscopic opening rate in channels studied by inside-out macropatches. Interestingly, the inhibitory activity was abolished by proteinase treatment. We conclude that a peptide toxin contained in the scorpion venom inhibits CFTR channels by a pore-block mechanism; these experiments provide the first step toward isolation of the active component, which would be highly valuable as a probe for CFTR structure and function.
Novel method for measuring junctional proton permeation in isolated ventricular myocyte cell pairs
Authors: Swietach Pawel , Vaughan-Jones Richard D.
Published: July 2004

Partial exposure of single ventricular myocytes to membrane-permeant weak acids or bases, using a dual-microperfusion technique, generates large and stable intracellular pH (pHi) gradients. In this study, we have investigated the feasibility of using the technique to estimate junctional proton permeability. This was done by recording the pHi gradient developed across the junctional region of a pair of conjoined ventricular myocytes, isolated enzymically from a guinea pig heart when one of the cells was partially exposed to acetate or ammonium. We show that under HEPES-buffered conditions, the junctional discontinuity in the pHi profile can be used to derive an apparent proton permeability coefficient ( PHapp). The mean PHapp obtained was 4.45 ± 0.21·10−4 cm/s ( n = 43) at an average junctional pHi of 7.04 ± 0.02. In the presence of the junctional inhibitor α-glycyrrhetinic acid, exposure of the proximal cell to weak acid or base produced no pHi change in the distal cell, confirming that distal changes were normally caused by acid-base flux through connexons assembled into junctional channels. The validity of the dual-microperfusion method was tested further by using a diffusion-permeation-reaction model for intracellular protons, designed to highlight possible errors in the estimates of PHapp. Our technique for measuring PHapp provides a useful alternative to the previous, more invasive technique of locally loading acid through a cell-attached patch pipette. The technique may provide a simple method for investigating the factors regulating cell-to-cell proton transmission.
Endocannabinoid-Dependent Neocortical Layer-5 LTD in the Absence of Postsynaptic Spiking
Authors: Sjöström Per Jesper , Turrigiano Gina G. , Nelson Sacha B.
Published: July 2004

Long-term depression (LTD) was induced in neocortical layer 5 pyramidal connections by pairing presynaptic firing with subthreshold postsynaptic depolarization (dLTD) or via a spike-timing protocol (tLTD). Like tLTD, dLTD reduced short-term depression and increased the coefficient of variation consistent with a presynaptic site of expression. Also like tLTD, dLTD was blocked by CB1 cannibinoid receptor blockade and required activation of presumably presynaptic NR2B-containing N-methyl-d-aspartate receptors. The two forms of LTD had identical magnitudes and time courses and occluded one another, and neither depended on frequency. Finally, dLTD shares with tLTD the asymmetric temporal window of induction. In conclusion, the types of LTD induced by these two protocols are indistinguishable, suggesting that the mechanism that underlies tLTD paradoxically does not require postsynaptic spiking: The subthreshold postsynaptic depolarizations of dLTD appear to fully substitute for postsynaptic spiking
Membrane and Firing Properties of Glutamatergic and GABAergic Neurons in the Rat Medial Vestibular Nucleus
Authors: Takazawa Tomonori , Saito Yasuhiko , Tsuzuki Keisuke , Ozawa Seiji
Published: July 2004

In previous studies, neurons in the medial vestibular nucleus (MVN) were classified mainly into 2 types according to their intrinsic membrane properties in in vitro slice preparations. However, it has not been determined whether the classified neurons are excitatory or inhibitory ones. In the present study, to clarify the relationship between the chemical and electrophysiological properties of MVN neurons, we explored mRNAs of cellular markers for GABAergic (glutamic acid decarboxylase 65, 67, and neuronal GABA transporter), glutamatergic (vesicular glutamate transporter 1 and 2), glycinergic (glycine transporter 2), and cholinergic neurons (choline acetyltransferase and vesicular acetylcholine transporter) expressed in electrophysiologically characterized MVN neurons in rat brain stem slice preparations. For this purpose, we combined whole cell patch-clamp recording analysis with single-cell reverse transcription–polymerase chain reaction (RT-PCR) analysis. We examined the membrane properties such as afterhyperpolarization (AHP), firing pattern, and response to hyperpolarizing current pulse to classify MVN neurons. From the single-cell RT-PCR analysis, we found that GABAergic neurons consisted of heterogeneous populations with different membrane properties. Comparison of the membrane properties of GABAergic neurons with those of other neurons revealed that AHPs without slow components and a firing pattern with delayed spike generation (late spiking) were preferential properties of GABAergic neurons. On the other hand, most glutamatergic neurons formed a homogeneous subclass of neurons exhibiting AHPs with slow components, repetitive firings with constant interspike intervals (continuous spiking), and time-dependent inward rectification in response to hyperpolarizing current pulses. We also found a small number of cholinergic neurons with various membrane properties. These findings clarify the electrophysiological properties of excitatory and inhibitory neurons in the MVN, and the information about the preferential membrane properties may be useful for identifying GABAergic and glutamatergic MVN neurons electrophysiologically.
Fracture Mechanics Analysis of Tilted Sandwich Debond (TSD) Specimen
Authors: Li Xiaoming , Carlsson Leif A.
Published: July 2004

The Tilted Sandwich Debond (TSD) specimen has recently been introduced for characterization of debond fracture of foam core sandwich panels. A precracked sandwich specimen is attached to an incline, and debonding is achieved by loading the partially debonded top face until further crack propagation occurs. In this paper we present finite element analysis of several configurations of the TSD specimen, viz. a homogeneous and isotropic specimen, interface crack specimen, and sub-interface crack specimen. Stress intensity factors are determined using various mesh refinements over a range of tilt angles, and the mode mixity is related to the potential for crack deflection (kinking) into the core. Irrespective of the tilt angle, it was found that cracks located at the face/core interface displayed amode mixity favorable for kinking into the core. Experiments on a sandwich with a H100 PVC foam core revealed kinking at an angle in reasonable agreement with the prediction. After kinking, analysis of sub-interface crack configurations showed that the crack loading becomes highly mode I dominated, which indicates crack propagation parallel to the face/core interface, again in agreement with experimental observations. Fracture mechanics analysis of foam materials, based on assumption of homogeneous material, however, becomes questionable when the singular domain becomes comparable to the cell size of the foam.