bluuebunny/crossref_metadata_2025_split huggingface dataset:
Genetic and Antigenic Diversity of the Surface Protective Antigen Proteins of
Erysipelothrix rhusiopathiae
Authors: To Ho , Nagai Shinya 
Published: May 2007

ABSTRACT

The surface protective antigen (Spa) protein of
Erysipelothrix rhusiopathiae
has been shown to be highly immunogenic and is a potential candidate for a new vaccine against erysipelas. In this study, we cloned and sequenced
spa
genes from all
E. rhusiopathiae
serovar reference strains as well as from a serovar 18 strain which was not classified as any species in the genus
Erysipelothrix.
Sequence analysis revealed that the Spa proteins could be classified into three molecular species, including SpaA, which was previously found in serovars 1a and 2, and the newly designated SpaB and SpaC proteins. The SpaA protein is produced by
E. rhusiopathiae
serovars 1a, 1b, 2, 5, 8, 9, 12, 15, 16, 17, and N, the SpaB protein is produced by
E. rhusiopathiae
serovars 4, 6, 11, 19, and 21, and the SpaC protein is produced only by serovar 18. The amino acid sequence similarity was high among members of each Spa type (96 to 99%) but low between different Spa types (∼60%). The greatest diversity in Spa proteins was found in the N-terminal half of the molecule (50 to 57% similarity), which was shown to be involved in immunoprotection. Coinciding with this, immunoblot analysis revealed that rabbit antisera specific to each Spa reacted strongly with the homologous Spa protein but weakly with heterologous Spa proteins. A mouse cross-protection study showed that the three recombinant Spa (rSpa) proteins elicited complete protection against challenge with homologous strains but that the level of protection against challenge with heterologous strains varied depending on the rSpa protein used for immunization. Our study is the first to demonstrate sequence and antigenic diversity in Spa proteins and to indicate that rSpaC may be the most promising antigen for use as a vaccine component because of its broad cross-protectiveness.
Detection of Mutations Conferring Resistance to Linezolid in
Enterococcus
spp. by Fluorescence In Situ Hybridization
Authors: Werner Guido , Bartel Melanie , Wellinghausen Nele , Essig Andreas , Klare Ingo , Witte Wolfgang , Poppert Sven 
Published: May 2007

ABSTRACT

A fluorescence in situ hybridization (FISH) assay was established to detect linezolid resistance (conferred by the mutation 2576G>T in the gene coding for the 23 string of the ribosomal RNA) in enterococci. The assay was evaluated with 106
Enterococcus
isolates; it showed a sensitivity of 100% for the detection of phenotypic resistance and was even able to identify a single mutated allele in phenotypically linezolid-susceptible isolates.
Detection and Identification ofLeishmaniaSpecies from Clinical Specimens by Using a Real-Time PCR Assay and Sequencing of the CytochromebGene
Authors: Foulet Françoise , Botterel Françoise , Buffet Pierre , Morizot Gloria , Rivollet Danièle , Deniau Michèle , Pratlong Francine , Costa Jean-Marc , Bretagne Stéphane 
Published: May 2007

ABSTRACTVisceral and cutaneous leishmaniases are heterogenous entities. TheLeishmaniaspecies that a given patient harbors usually cannot be determined clinically, and this identification is essential to prescribe the best species-specific therapeutic regimen. Our diagnosis procedure includes a real-time PCR assay targeted at the 18S rRNA gene, which detects allLeishmaniaspecies but which is not specific for a givenLeishmaniaspecies. We developed a species identification based on sequencing of the cytochromeb(cytb) gene directly from the DNA extracted from the clinical specimen. The sequences were analyzed using the Sequence Analysis/Seqscape v2.1 software (Applied Biosystems). This software is designed to automatically identify the closest sequences from a reference library after analysis of all known or unknown polymorphic positions. The library was built with theLeishmaniacytbgene sequences available in GenBank. Fifty-three consecutive real-time PCR-positive specimens were studied for species identification. The cytbgene was amplified in the 53 specimens. Sequencing resulted in the identification of six different species with ≥99% identity with the reference sequences over 872 nucleotides. The identification was obtained in two working days and was in accordance with the multilocus enzyme electrophoresis identification when available. Real-time PCR followed by sequencing of the cytbgene confirmed the diagnosis of leishmaniasis and rapidly determined the infecting species directly from the clinical specimen without the need for the isolation of parasites. This technique has the potential to significantly accelerate species-adapted therapeutic decisions regarding treatment of leishmaniasis.
Entry of
Rice Dwarf Virus
into Cultured Cells of Its Insect Vector Involves Clathrin-Mediated Endocytosis
Authors: Wei Taiyun , Chen Hongyan , Ichiki-Uehara Tamaki , Hibino Hiroyuki , Omura Toshihiro 
Published: May 2007

ABSTRACT

Electron microscopy revealed that the entry of
Rice dwarf virus
(RDV) into insect vector cells involved endocytosis via coated pits. The treatment of cells with drugs that block receptor-mediated or clathrin-mediated endocytosis significantly reduced RDV infectivity. However, the drug that blocks caveola-mediated endocytosis had a negligible effect on such infection. Infection was also inhibited when cells had been pretreated with bafilomycin A1, which interferes with acidification of endosomes. Moreover, immunofluorescence staining indicated that the virus is internalized into early endosomes. Together, our data indicate that RDV enters insect vector cells through receptor-mediated, clathrin-dependent endocytosis and is sequestered in early endosomes.
The F Gene of Rodent Brain-Adapted Mumps Virus Is a Major Determinant of Neurovirulence
Authors: Lemon Ken , Rima Bertus K. , McQuaid Stephen , Allen Ingrid V. , Duprex W. Paul 
Published: May 2007

ABSTRACTPrior to the introduction of live-attenuated vaccines, mumps virus (MuV) was the leading cause of virus-induced meningitis. Although vaccination has been effective at controlling the disease, the use of insufficiently attenuated strains has been associated with high rates of aseptic meningitis in vaccinees. The molecular basis of MuV attenuation is poorly understood, and no reliable molecular markers of virulence have been identified. In this study, reverse genetics has been used to identify molecular determinants of MuV neuropathogenesis. Recombinant viruses, containing the envelope-associated genes from the Kilham (MuVKH) rodent brain-adapted strain of MuV, were generated in the Jeryl Lynn 5 (MuVJL5) vaccine strain background. The syncytium phenotypes of the recombinant viruses on Vero cells differed depending on the source of the fusion (F) and hemagglutinin-neuraminidase (HN) glycoproteins, with heterologous combinations showing either an increase or a decrease in the level of cell fusion compared to that of the homologous parental combinations. This was confirmed by transiently cotransfecting eukaryotic F and HN glycoprotein expression constructs. A Lewis rat model that discriminates between neurovirulent and nonneurovirulent MuV strains based on the extent of hydrocephalus induced in the rat brain after intracerebral inoculation was used to assess the phenotype of the recombinant viruses. Expression of the matrix (M), small hydrophobic (SH), or HN gene in isolation did not confer a neurovirulent phenotype. Expression of the F gene of the neurovirulent strain alone was sufficient to induce significant levels of hydrocephalus. Coexpression of the homologous HN gene led to a marginal increase in the level of hydrocephalus.
Tumor Necrosis Factor Activates a Conserved Innate Antiviral Response to Hepatitis B Virus That Destabilizes Nucleocapsids and Reduces Nuclear Viral DNA
Authors: Puro Robyn , Schneider Robert J. 
Published: May 2007

ABSTRACTTumor necrosis factor (TNF) is critical for the control of hepatitis B virus (HBV) in the clinical setting and in model systems. TNF induces noncytopathic suppression and clearance of HBV in animal models, possibly through reduction of viral nucleocapsids, but the mechanism is not well described. Here, we demonstrate the molecular mechanism and broad host range for TNF action against HBV. We show that TNF rapidly blocks HBV replication by promoting destabilization of preexisting cytoplasmic viral nucleocapsids containing viral RNA and DNA, as well as empty nucleocapsids. TNF destabilized human HBV nucleocapsids in a variety of human hepatocytic cell lines and in primary rat hepatocytes and also destabilized duck HBV (DHBV) nucleocapsids in chicken hepatocytic cells. Lysates from TNF-treated uninfected cells also destabilized HBV nucleocapsids in vitro. Moreover, inhibition of DHBV DNA replication by TNF blocks nuclear accumulation of the viral transcription template, maintenance of which is essential for the establishment and maintenance of chronic infection. We show that TNF destabilization of HBV nucleocapsids does not involve ubiquitination or methylation of the viral core protein and is not mediated by the nitric oxide free radical arm of the TNF pathway. These results define a novel antiviral mechanism mediated by TNF against multiple types of HBVs in different species.
Bovine Papillomavirus Type 1 Infection Is Mediated by SNARE Syntaxin 18
Authors: Laniosz Valerie , Nguyen Kha C. , Meneses Patricio I. 
Published: May 2007

ABSTRACT

Events that lead to viral infections include the binding of the virus to the target cells, internalization of the virus into the cells, and the ability of the viral genome to be expressed. These steps are mediated by cellular and viral proteins and are temporally regulated. The papillomavirus capsid consists of two virally encoded capsid proteins, L1 and L2. Much is known about the role of the major capsid protein L1 compared to what is known of the role of the L2 protein. We identified the interaction of the L2 protein with SNARE protein syntaxin 18, which mediates the trafficking of vesicles and their cargo between the endoplasmic reticulum, the
cis
-Golgi compartment, and possibly the plasma membrane. Mutations of L2 residues 41 to 44 prevented the interaction of L2 protein with syntaxin 18 in cotransfection experiments and resulted in noninfectious pseudovirions. In this paper, we describe that syntaxin 18 colocalizes with infectious bovine papillomavirus type 1 (BPV1) pseudovirions during infection but does not colocalize with the noninfectious BPV1 pseudovirions made with an L2 mutant at residues 41 to 44. We show that an antibody against BPV1 L2 residues 36 to 49 (αL2 36-49) binds to in vitro-generated BPV1 pseudoviral capsids and does not coimmunoprecipitate syntaxin 18- and BPV1 L2-transfected proteins. αL2 36-49 was able to partially or completely neutralize infection of BPV1 pseudovirions and genuine virions. These results support the dependence of syntaxin 18 during BPV1 infection and the ability to interfere with infection by targeting the L2-syntaxin 18 interaction and further define the infectious route of BPV1 mediated by the L2 protein.
Identification of a Ca
2+
-Binding Domain in the Rubella Virus Nonstructural Protease
Authors: Zhou Yubin , Tzeng Wen-Pin , Yang Wei , Zhou Yumei , Ye Yiming , Lee Hsiau-wei , Frey Teryl K. , Yang Jenny 
Published: May 2007

ABSTRACT

The rubella virus (RUB) nonstructural protein (NS) open reading frame (ORF) encodes a polypeptide precursor that is proteolytically self cleaved into two replicase components involved in viral RNA replication. A putative EF-hand Ca
2+
-binding motif that was conserved across different genotypes of RUB was predicted within the nonstructural protease that cleaves the precursor by using bioinformatics tools. To probe the metal-binding properties of this motif, we used an established grafting approach and engineered the 12-residue Ca
2+
-coordinating loop into a non-Ca
2+
-binding scaffold protein, CD2. The grafted EF-loop bound to Ca
2+
and its trivalent analogs Tb
3+
and La
3+
with
K

d

s of 214, 47, and 14 μM, respectively. Mutations (D1210A and D1217A) of two of the potential Ca
2+
-coordinating ligands in the EF-loop led to the elimination of Tb
3+
binding. Inductive coupled plasma mass spectrometry was used to confirm the presence of Ca
2+
([Ca
2+
]/[protein] = 0.7 ± 0.2) in an NS protease minimal metal-binding domain, RUBCa, that spans the EF-hand motif. Conformational studies on RUBCa revealed that Ca
2+
binding induced local conformational changes and increased thermal stability (Δ
T

m

= 4.1°C). The infectivity of an RUB infectious cDNA clone containing the mutations D1210A/D1217A was decreased by ∼20-fold in comparison to the wild-type (wt) clone, and these mutations rapidly reverted to the wt sequence. The NS protease containing these mutations was less efficient at precursor cleavage than the wt NS protease at 35°C, and the mutant NS protease was temperature sensitive at 39°C, confirming that the Ca
2+
-binding loop played a structural role in the NS protease and was specifically required for optimal stability under physiological conditions.
Dendritic Cells Infected withvpr-Positive Human Immunodeficiency Virus Type 1 Induce CD8+T-Cell Apoptosis via Upregulation of Tumor Necrosis Factor Alpha
Authors: Majumder Biswanath , Venkatachari Narasimhan J. , Schafer Elizabeth A. , Janket Michelle L. , Ayyavoo Velpandi 
Published: May 2007

ABSTRACTHuman immunodeficiency virus type 1 (HIV-1) viral protein R (Vpr) plays a crucial role in viral replication and pathogenesis by inducing cell cycle arrest, apoptosis, translocation of preintegration complex, potentiation of glucocorticoid action, impairment of dendritic cell (DC) maturation, and T-cell activation. Recent studies involving the direct effects of Vpr on DCs and T cells indicated that HIV-1 containing Vpr selectively impairs phenotypic maturation, cytokine network, and antigen presentation in DCs and dysregulates costimulatory molecules and cytokine production in T cells. Here, we have further investigated the indirect effect of HIV-1 Vpr+virus-infected DCs on the bystander CD8+T-cell population. Our results indicate that HIV-1 Vpr+virus-infected DCs dysregulate CD8+T-cell proliferation and induce apoptosis. Vpr-containing virus-infected DC-mediated CD8+T-cell killing occurred in part through enhanced tumor necrosis factor alpha production by infected DCs and subsequent induction of death receptor signaling and activation of the caspase 8-dependent pathway in CD8+T cells. Collectively, these results provide evidence that Vpr could be one of the important contributors to the host immune escape by HIV-1 through its ability to dysregulate both directly and indirectly the DC biology and T-cell functions.
Ebola Virus Glycoprotein 1: Identification of Residues Important for Binding and Postbinding Events
Authors: Brindley Melinda A. , Hughes Laura , Ruiz Autumn , McCray Paul B. , Sanchez Anthony , Sanders David A. , Maury Wendy 
Published: May 2007

ABSTRACT
The filoviruses Ebola virus (EBOV) and Marburg virus (MARV) are responsible for devastating hemorrhagic fever outbreaks. No therapies are available against these viruses. An understanding of filoviral glycoprotein 1 (GP1) residues involved in entry events would facilitate the development of antivirals. Towards this end, we performed alanine scanning mutagenesis on selected residues in the amino terminus of GP1. Mutant GPs were evaluated for their incorporation onto feline immunodeficiency virus (FIV) particles, transduction efficiency, receptor binding, and ability to be cleaved by cathepsins L and B. FIV virions bearing 39 out of 63 mutant glycoproteins transduced cells efficiently, whereas virions bearing the other 24 had reduced levels of transduction. Virions pseudotyped with 23 of the poorly transducing GPs were characterized for their block in entry. Ten mutant GPs were very poorly incorporated onto viral particles. Nine additional mutant GPs (G87A/F88A, K114A/K115A, K140A, G143A, P146A/C147A, F153A/H154A, F159A, F160A, and Y162A) competed poorly with wild-type GP for binding to permissive cells. Four of these nine mutants (P146A/C147A, F153A/H154A, F159A, and F160A) were also inefficiently cleaved by cathepsins. An additional four mutant GPs (K84A, R134A, D150A, and E305/E306A) that were partially defective in transduction were found to compete effectively for receptor binding and were readily cleaved by cathepsins. This finding suggested that this latter group of mutants might be defective at a postbinding, cathepsin cleavage-independent step. In total, our study confirms the role of some GP1 residues in EBOV entry that had previously been recognized and identifies for the first time other residues that are important for productive entry.
Ultracentrifugation of Serum Samples Allows Detection of Hepatitis C Virus RNA in Patients with Occult Hepatitis C
Authors: Bartolomé Javier , López-Alcorocho Juan Manuel , Castillo Inmaculada , Rodríguez-Iñigo Elena , Quiroga Juan Antonio , Palacios Ricardo , Carreño Vicente 
Published: May 2007

ABSTRACTOccult hepatitis C virus (HCV) infection of patients with abnormal liver function tests of unknown origin who are anti-HCV and serum HCV RNA negative but who have HCV RNA in the liver has been described. As HCV replicates in the liver cells of these patients, it could be that the amount of circulating viral particles is under the detection limit of the most sensitive techniques. To prove this hypothesis, serum samples from 106 patients with occult HCV infection were analyzed. Two milliliters of serum was ultracentrifuged over a 10% sucrose cushion for 17 h at 100,000 ×gav, where av means average, and HCV RNA detection was performed by strand-specific real-time PCR. Out of the 106 patients, 62 (58.5%) had detectable serum HCV RNA levels after ultracentrifugation, with a median load of 70.5 copies/ml (range, 18 to 192). Iodixanol density gradient studies revealed that HCV RNA was positive at densities of 1.03 to 1.04 and from 1.08 to 1.19 g/ml, which were very similar to those found in the sera of patients with classical chronic HCV infection. Antigenomic HCV RNA was found in the livers of 56 of 62 (90.3%) patients with detectable serum HCV RNA levels after ultracentrifugation, compared to 27 of 44 (61.4%) negative patients (P< 0.001). No differences in the median loads of antigenomic HCV RNA between patients with an those without serum HCV RNA (4.5 × 104[range, 7.9 × 102to 1.0 × 106] versus 2.3 × 104[range, 4.0 × 102to 2.2 × 105]) were found. Alanine aminotransferase and gamma-glutamyl transpeptidase levels, liver necroinflammatory activity, and fibrosis did not differ between both groups. In conclusion, HCV RNA can be detected in the sera of patients with occult HCV infection after circulating viral particles are concentrated by ultracentrifugation.
Hydrogen Sulfide Induces Direct Radical-Associated DNA Damage
Authors: Attene-Ramos Matias S. , Wagner Elizabeth D. , Gaskins H. Rex , Plewa Michael J. 
Published: May 2007

Abstract
Hydrogen sulfide (H2S) is produced by indigenous sulfate-reducing bacteria in the large intestine and represents an environmental insult to the colonic epithelium. Clinical studies have linked the presence of either sulfate-reducing bacteria or H2S in the colon with chronic disorders such as ulcerative colitis and colorectal cancer, although at this point, the evidence is circumstantial and underlying mechanisms remain undefined. We showed previously that sulfide at concentrations similar to those found in the human colon induced genomic DNA damage in mammalian cells. The present study addressed the nature of the DNA damage by determining if sulfide is directly genotoxic or if genotoxicity requires cellular metabolism. We also questioned if sulfide genotoxicity is mediated by free radicals and if DNA base oxidation is involved. Naked nuclei from untreated Chinese hamster ovary cells were treated with sulfide; DNA damage was induced by concentrations as low as 1 μmol/L. This damage was effectively quenched by cotreatment with butylhydroxyanisole. Furthermore, sulfide treatment increased the number of oxidized bases recognized by formamidopyrimidine [fapy]-DNA glycosylase. These results confirm the genotoxicity of sulfide and strongly implicate that this genotoxicity is mediated by free radicals. These observations highlight the possible role of sulfide as an environmental insult that, given a predisposing genetic background, may lead to genomic instability or the cumulative mutations characteristic of colorectal cancer. (Mol Cancer Res 2007;5(5):455–9)
Rendering unto Caesar?: The Politics of Church of England Clergy since 1980
Authors: Field Clive D. 
Published: May 2007

ABSTRACTThis is the first systematic attempt to chart the evolving political views of contemporary Church of England clergy. The article is based upon a comparative quantitative analysis and synthesis of eighteen national and four local surveys conducted between 1979 and 2004. Ministerial opinions on the state's influence on the Church and the Church's influence on the state are both considered. Ten specific conclusions are drawn. While the clergy generally cling to the concept of an Established Church, they are very critical of some of the traditional manifestations of that establishment. They also mostly think it highly appropriate for the Church to intervene in the world of party politics, and not simply on moral issues. In this they are positioned ahead of the thinking of many of the committed Anglican laity, for whom a degree of separation of religion and politics remains the ideal. The academic, ecclesiastical and political implications of these findings are briefly explored.
The human Shwachman-Diamond syndrome protein, SBDS, associates with ribosomal RNA
Authors: Ganapathi Karthik A. , Austin Karyn M. , Lee Chung-Sheng , Dias Anusha , Malsch Maggie M. , Reed Robin , Shimamura Akiko 
Published: May 2007

Abstract
Shwachman-Diamond syndrome (SDS) is an autosomal recessive disorder characterized by bone marrow failure, exocrine pancreatic dysfunction, and leukemia predisposition. Mutations in the SBDS gene are identified in most patients with SDS. SBDS encodes a highly conserved protein of unknown function. Data from SBDS orthologs suggest that SBDS may play a role in ribosome biogenesis or RNA processing. Human SBDS is enriched in the nucleolus, the major cellular site of ribosome biogenesis. Here we report that SBDS nucleolar localization is dependent on active rRNA transcription. Cells from patients with SDS or Diamond-Blackfan anemia are hypersensitive to low doses of actinomycin D, an inhibitor of rRNA transcription. The addition of wild-type SBDS complements the actinomycin D hypersensitivity of SDS patient cells. SBDS migrates together with the 60S large ribosomal subunit in sucrose gradients and coprecipitates with 28S ribosomal RNA (rRNA). Loss of SBDS is not associated with a discrete block in rRNA maturation or with decreased levels of the 60S ribosomal subunit. SBDS forms a protein complex with nucleophosmin, a multifunctional protein implicated in ribosome biogenesis and leukemogenesis. Our studies support the addition of SDS to the growing list of human bone marrow failure syndromes involving the ribosome.