bluuebunny/crossref_metadata_2025_split huggingface dataset:
Validation of Reference Genes in Cervical Cell Samples from Human Papillomavirus-Infected and -Uninfected Women for Quantitative Reverse Transcription-PCR Assays
Authors: Daud Ibrahim I. , Scott Mark E. 
Published: July 2008

ABSTRACTReference genes for quantitative reverse transcription-PCR (qRT-PCR) studies must be validated for the cell type studied and should be stable between the groups that represent the independent variable in an experimental design. We sought to identify the reference genes in cervical cell specimens showing the most stable expression between human papillomavirus (HPV)-infected and -uninfected women without high-grade cervical intraepithelial neoplasia. Using endocervical cells collected by cytology brush and Sybr green-based qRT-PCR, eight candidate genes were screened for amplification efficiency, specificity, and overall stability (by use of geNorm software). The five most stable genes were then further evaluated both for overall stability (geNorm) and intergroup stability (by use of NormFinder software) in specimens from HPV-negative and HPV-positive women. The combination of the glyceraldehyde-3-phosphate dehydrogenase gene (GAPDH) andRPLP0was the most stable overall, with a geNorm stability measure of 0.603. The intergroup analysis showedGAPDHto be the most stable single gene andRPLP0to be second most stable and also showed that these genes represent the most stable two-gene combination, with a NormFinder stability value of 0.130. The fact that these two distinct approaches identified the same pair of genes provides added confidence that, when the focus is on HPV infection, a normalization factor derived from these two genes is likely to be appropriate.
Validation of a Competitive Enzyme-Linked Immunosorbent Assay for Detection ofBabesia bigeminaAntibodies in Cattle
Authors: Goff Will L. , Johnson Wendell C. , Molloy John B. , Jorgensen Wayne K. , Waldron Susan J. , Figueroa Julio V. , Matthee Olivier , Adams D. Scott , McGuire Travis C. , Pino Ignacio , Mosqueda Juan , Palmer Guy H. , Suarez Carlos E. , Knowles Donald P. , McElwain Terry F. 
Published: July 2008

ABSTRACTA competitive enzyme-linked immunosorbent assay (cELISA) based on a broadly conserved, species-specific, B-cell epitope within the C terminus ofBabesia bigeminarhoptry-associated protein 1a was validated for international use. Receiver operating characteristic analysis revealed 16% inhibition as the threshold for a negative result, with an associated specificity of 98.3% and sensitivity of 94.7%. Increasing the threshold to 21% increased the specificity to 100% but modestly decreased the sensitivity to 87.2%. By using 21% inhibition, the positive predictive values ranged from 90.7% (10% prevalence) to 100% (95% prevalence) and the negative predictive values ranged from 97.0% (10% prevalence) to 48.2% (95% prevalence). The assay was able to detect serum antibody as early as 7 days after intravenous inoculation. The cELISA was distributed to five different laboratories along with a reference set of 100 defined bovine serum samples, including known positive, known negative, and field samples. The pairwise concordance among the five laboratories ranged from 100% to 97%, and all kappa values were above 0.8, indicating a high degree of reliability. Overall, the cELISA appears to have the attributes necessary for international application.
Oral Immunization with RecombinantLactobacillus plantarumInduces a Protective Immune Response in Mice with Lyme Disease
Authors: del Rio Beatriz , Dattwyler Raymond J. , Aroso Miguel , Neves Vera , Meirelles Luciana , Seegers Jos F. M. L. , Gomes-Solecki Maria 
Published: July 2008

ABSTRACTMucosal immunization is advantageous over other routes of antigen delivery because it can induce both mucosal and systemic immune responses. Our goal was to develop a mucosal delivery vehicle based on bacteria generally regarded as safe, such asLactobacillusspp. In this study, we used the Lyme disease mouse model as a proof of concept. We demonstrate that an oral vaccine based on live recombinantLactobacillus plantarumprotects mice from tick-transmittedBorrelia burgdorferiinfection. Our method of expressing vaccine antigens inL. plantaruminduces both systemic and mucosal immunity after oral administration. This platform technology can be applied to design oral vaccine delivery vehicles against several microbial pathogens.
Characterization of Vancomycin-Heteroresistant
Staphylococcus aureus
from the Metropolitan Area of Detroit, Michigan, over a 22-Year Period (1986 to 2007)
Authors: Rybak Michael J. , Leonard Steve N. , Rossi Kerri L. , Cheung Chrissy M. , Sadar Helio S. , Jones Ronald N. 
Published: July 2008

ABSTRACT

We screened for heteroresistant, vancomycin-intermediate
Staphylococcus aureus
(hVISA) among clinical isolates of methicillin-resistant
S. aureus
collected from three hospitals (two urban teaching hospitals and one community hospital) in the Detroit metropolitan area over a 22-year period. The Macro Etest method was used to screen all available isolates. Confirmation of hVISA-positive screens were confirmed by population-area under the concentration-time curve (AUC) analysis. A total of 1,499 isolates revealed hVISA/VISA rates of 2.2/0.4% (
n
= 225; 1986 to 1993), 7.6/2.3% (
n
= 356; 1994 to 2002), and 8.3/0.3% (
n
= 917; 2003 to 2007). Population-AUC analysis confirmed 92.6% of the hVISA-positive strains determined by the Macro Etest method. For the isolates with known sources (1,208), the predominant source of hVISA was blood (60%), followed by lung (21%), skin and wound infections (14%), abscess (1%), and other (4%). The percentage of hVISA-positive strains appeared to increase as a function of the vancomycin MIC. Staphylococcal cassette chromosome
mec
(SCC
mec
) typing revealed that the majority (56.9%) of the hVISA strains were SCC
mec
type II and 39.4% were type IV; the majority of these strains were collected from 2000 to 2007. Our data indicate that the prevalence of hVISA may be increasing. Based on the association of vancomycin treatment failure in patients with hVISA, surveillance of hVISA strains is warranted.
Detection and Characterization of Group C Rotaviruses in Asymptomatic Piglets in Ireland
Authors: Collins P. J. , Martella V. , O'Shea H. 
Published: July 2008

ABSTRACTGroup C rotaviruses are important human enteric pathogens that have also been detected in a variety of mammalian species, including pigs. Group C rotaviruses have been identified in piglets with diarrhea, but their ecology remains to be elucidated. By screening of 292 fecal samples collected from 4- to 5-week-old asymptomatic pigs from four herds in Ireland between 2005 and 2007, 13 (4.4%) samples tested positive by reverse transcription-PCR for group C rotavirus. Group A rotaviruses were also detected in 19 samples but not in conjunction with group C viruses. The gene encoding the major group C neutralization antigen, the outer capsid protein VP7, was sequenced. The majority of the strains were very closely related to each other (>99% amino acid [aa] identity) and were characterized as genogroup G1 since they were genetically related to the prototype porcine strain Cowden (92.6% aa identity). Conversely, two strains (1GA/05/Cork/Ire and 281/07/Dublin/Ire) were characterized as genogroup G6 since they displayed the highest identity (89.2 to 94.0% aa) to porcine G6 strains (43/06-22-like). Unexpectedly, one such G6 strain, 1GA/05/Cork/Ire, lacked the 4-aa insertion in the VP7 variable region VR8 found in all the other G6 group C rotaviruses. This study provides evidence that porcine group C rotavirus may be detected not infrequently in asymptomatic piglets. In addition, it provides evidence that, unlike the human viruses, porcine group C rotaviruses display broad genetic heterogeneity, which may pose a challenge for the development of prophylactic tools.
Genetic Lineages of Community-Associated Methicillin-Resistant
Staphylococcus aureus
in Kuwait Hospitals
Authors: Udo E. E. , O'Brien F. G. , Al-Sweih N. , Noronha Bobby , Matthew B. , Grubb W. B. 
Published: July 2008

ABSTRACT

Twenty-six community-associated methicillin-resistant
Staphylococcus aureus
(CAMSRA) isolates were characterized by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) and screened for accessory gene regulator (agr), capsular polysaccharide (cap), and Panton-Valentine leucocidin (PVL) genes. They exhibited five PFGE patterns (types A to E). The majority were PFGE type A (12 isolates) or type B (8 isolates). MLST showed that PFGE type A isolates belonged to sequence type 80 (ST80), while the PFGE type B isolates were ST30. The ST80 and ST30 clones contained agr allotype 3, cap type 8, and PVL. The results showed that two internationally recognized CAMRSA clones are dominant in Kuwait hospitals.
Llama-Derived Single-Chain Antibody Fragments Directed to Rotavirus VP6 Protein Possess Broad Neutralizing Activity In Vitro and Confer Protection against Diarrhea in Mice
Authors: Garaicoechea Lorena , Olichon Aurelien , Marcoppido Gisela , Wigdorovitz Andrés , Mozgovoj Marina , Saif Linda , Surrey Thomas , Parreño Viviana 
Published: July 2008

ABSTRACT
Group A rotavirus is one of the most common causes of severe diarrhea in human infants and newborn animals. Rotavirus virions are triple-layered particles. The outer capsid proteins VP4 and VP7 are highly variable and represent the major neutralizing antigens. The inner capsid protein VP6 is conserved among group A rotaviruses, is highly immunogenic, and is the target antigen of most immunodiagnosis tests. Llama-derived single-chain antibody fragments (VHH) are the smallest molecules with antigen-binding capacity and can therefore be expected to have properties different from conventional antibodies. In this study a library containing the VHH genes of a llama immunized with recombinant inner capsid protein VP6 was generated. Binders directed to VP6, in its native conformation within the viral particle, were selected and characterized. Four selected VHH directed to conformational epitopes of VP6 recognized all human and animal rotavirus strains tested and could be engineered for their use in immunodiagnostic tests for group A rotavirus detection. Three of the four VHH neutralized rotavirus in vivo independently of the strain serotype. Furthermore, this result was confirmed by in vivo partial protection against rotavirus challenge in a neonatal mouse model. The present study demonstrates for the first time a broad neutralization activity of VP6 specific VHH in vitro and in vivo. Neutralizing VHH directed to VP6 promise to become an essential tool for the prevention and treatment of rotavirus diarrhea.
Genomic Analysis Reveals Age-Dependent Innate Immune Responses to Severe Acute Respiratory Syndrome Coronavirus
Authors: Baas Tracey , Roberts Anjeanette , Teal Thomas H. , Vogel Leatrice , Chen Jun , Tumpey Terrence M. , Katze Michael G. , Subbarao Kanta 
Published: July 2008

ABSTRACT
The relationship between immunosenescence and the host response to virus infection is poorly understood at the molecular level. Two different patterns of pulmonary host responses to virus were observed when gene expression profiles from severe acute respiratory syndrome coronavirus (SARS-CoV)-infected young mice that show minimal disease were compared to those from SARS-CoV-infected aged mice that develop pneumonitis. In young mice, genes related to cellular development, cell growth, and cell cycle were downregulated during peak viral replication, and these transcripts returned to basal levels as virus was cleared. In contrast, aged mice had a greater number of upregulated immune response and cell-to-cell signaling genes, and the expression of many genes was sustained even after viral clearance, suggesting an exacerbated host response to virus. Interestingly, in SARS-CoV-infected aged mice, a subset of genes, including Tnfa, Il6, Ccl2, Ccl3, Cxcl10, and Ifng, was induced in a biphasic pattern that correlated with peak viral replication and a subsequent influx of lymphocytes and severe histopathologic changes in the lungs. We provide insight into gene expression profiles and molecular signatures underlying immunosenescence in the context of the host response to viral infection.
The Interaction between Human Papillomavirus Type 16 and FADD Is Mediated by a Novel E6 Binding Domain
Authors: Tungteakkhun Sandy S. , Filippova Maria , Neidigh Jonathan W. , Fodor Nadja , Duerksen-Hughes Penelope J. 
Published: July 2008

ABSTRACTHigh-risk strains of human papillomavirus, such as types 16 and 18, have been etiologically linked to cervical cancer. Most cervical cancer tissues are positive for both the E6 and E7 oncoproteins, since it is their cooperation that results in successful transformation and immortalization of infected cells. We have reported that E6 binds to tumor necrosis factor receptor 1 and to Fas-associated death domain (FADD) and, in doing so, prevents E6-expressing cells from responding to apoptotic stimuli. The binding site of E6 to FADD localizes to the first 23 amino acids of FADD and has now been further characterized by the use of deletion and site-directed mutants of FADD in pull-down and functional assays. The results from these experiments revealed that mutations of serine 16, serine 18, and leucine 20 obstruct FADD binding to E6, suggesting that these residues are part of the E6 binding domain on FADD. Because FADD does not contain the two previously identified E6 binding motifs, the LxxφLsh motif, and the PDZ motif, a novel binding domain for E6 has been identified on FADD. Furthermore, peptides that correspond to this region can block E6/FADD binding in vitro and can resensitize E6-expressing cells to apoptotic stimuli in vivo. These results demonstrate the existence of a novel E6 binding domain.
The Crystal Structure of Coxsackievirus B3 RNA-Dependent RNA Polymerase in Complex with Its Protein Primer VPg Confirms the Existence of a Second VPg Binding Site on
Picornaviridae
Polymerases
Authors: Gruez Arnaud , Selisko Barbara , Roberts Michael , Bricogne Gérard , Bussetta Cécile , Jabafi Ilham , Coutard Bruno , De Palma Armando M. , Neyts Johan , Canard Bruno 
Published: July 2008

ABSTRACT

The RNA-dependent RNA polymerase (RdRp) is a central piece in the replication machinery of RNA viruses. In picornaviruses this essential RdRp activity also uridylates the VPg peptide, which then serves as a primer for RNA synthesis. Previous genetic, binding, and biochemical data have identified a VPg binding site on poliovirus RdRp and have shown that is was implicated in VPg uridylation. More recent structural studies have identified a topologically distinct site on the closely related foot-and-mouth disease virus RdRp supposed to be the actual VPg-primer-binding site. Here, we report the crystal structure at 2.5-Å resolution of active coxsackievirus B3 RdRp (also named 3D
pol
) in a complex with VPg and a pyrophosphate. The pyrophosphate is situated in the active-site cavity, occupying a putative binding site either for the coproduct of the reaction or an incoming NTP. VPg is bound at the base of the thumb subdomain, providing first structural evidence for the VPg binding site previously identified by genetic and biochemical methods. The binding mode of VPg to CVB3 3D
pol
at this site excludes its uridylation by the carrier 3D
pol
. We suggest that VPg at this position is either uridylated by another 3D
pol
molecule or that it plays a stabilizing role within the uridylation complex. The CVB3 3D
pol
/VPg complex structure is expected to contribute to the understanding of the multicomponent VPg-uridylation complex essential for the initiation of genome replication of picornaviruses.
ERK1/2-Mediated Phosphorylation of Small Hepatitis Delta Antigen at Serine 177 Enhances Hepatitis Delta Virus Antigenomic RNA Replication
Authors: Chen Yen-Shun , Huang Wen-Hung , Hong Shiao-Ya , Tsay Yeou-Guang , Chen Pei-Jer 
Published: July 2008

ABSTRACT
The small hepatitis delta virus (HDV) antigen (SHDAg) plays an essential role in HDV RNA double-rolling-circle replication. Several posttranslational modifications (PTMs) of HDAgs, including phosphorylation, acetylation, and methylation, have been characterized. Among the PTMs, the serine 177 residue of SHDAg is a phosphorylation site, and its mutation preferentially abolishes HDV RNA replication from antigenomic RNA to genomic RNA. Using coimmunoprecipitation analysis, the cellular kinases extracellular signal-related kinases 1 and 2 (ERK1/2) are found to be associated with the Flag-tagged SHDAg mutant (Ser-177 replaced with Cys-177). In an in vitro kinase assay, serine 177 of SHDAg was phosphorylated directly by either Flag-ERK1 or Flag-ERK2. Activation of endogenous ERK1/2 by a constitutively active MEK1 (hemagglutinin-AcMEK1) increased phosphorylation of SHDAg at Ser-177; this phosphorylation was confirmed by immunoblotting using an antibody against phosphorylated S177 and mass spectrometric analysis. Interestingly, we found an increase in the HDV replication from antigenomic RNA to genomic RNA but not in that from genomic RNA to antigenomic RNA. The Ser-177 residue was critical for SHDAg interaction with RNA polymerase II (RNAPII), the enzyme proposed to regulate antigenomic RNA replication. These results demonstrate the role of ERK1/2-mediated Ser-177 phosphorylation in modulating HDV antigenomic RNA replication, possibly through RNAPII regulation. The results may shed light on the mechanisms of HDV RNA replication.
Scrapie Resistance in ARQ Sheep
Authors: Laegreid W. W. , Clawson M. L. , Heaton M. P. , Green B. T. , O'Rourke K. I. , Knowles D. P. 
Published: July 2008

ABSTRACTVariation in the ovine prion protein amino acid sequence influences scrapie progression, with sheep homozygous for A136R154Q171considered susceptible. This study examined the association of survival time of scrapie-exposed ARQ sheep with variation elsewhere in the ovine prion gene. Four single nucleotide polymorphism alleles were associated with prolonged survival. One nonsynonymous allele (T112) was associated with an additional 687 days of survival for scrapie-exposed sheep compared to M112 sheep (odds ratio, 42.5;P= 0.00014). The only two sheep homozygous for T112 (TARQ) did not develop scrapie, suggesting that the allelic effect may be additive. These results provide evidence that TARQ sheep are genetically resistant to development of classical scrapie.
Differential Cholesterol Binding by Class II Fusion Proteins Determines Membrane Fusion Properties
Authors: Umashankar M. , Sánchez-San Martín Claudia , Liao Maofu , Reilly Brigid , Guo Alice , Taylor Gwen , Kielian Margaret 
Published: July 2008

ABSTRACT
The class II fusion proteins of the alphaviruses and flaviviruses mediate virus infection by driving the fusion of the virus membrane with that of the cell. These fusion proteins are triggered by low pH, and their structures are strikingly similar in both the prefusion dimer and the postfusion homotrimer conformations. Here we have compared cholesterol interactions during membrane fusion by these two groups of viruses. Using cholesterol-depleted insect cells, we showed that fusion and infection by the alphaviruses Semliki Forest virus (SFV) and Sindbis virus were strongly promoted by cholesterol, with similar sterol dependence in laboratory and field isolates and in viruses passaged in tissue culture. The E1 fusion protein from SFV bound cholesterol, as detected by labeling with photocholesterol and by cholesterol extraction studies. In contrast, fusion and infection by numerous strains of the flavivirus dengue virus (DV) and by yellow fever virus 17D were cholesterol independent, and the DV fusion protein did not show significant cholesterol binding. SFV E1 is the first virus fusion protein demonstrated to directly bind cholesterol. Taken together, our results reveal important functional differences conferred by the cholesterol-binding properties of class II fusion proteins.
Identification of an Arginine-Rich Motif in Human Papillomavirus Type 1 E1^E4 Protein Necessary for E4-Mediated Inhibition of Cellular DNA Synthesis In Vitro and in Cells
Authors: Roberts Sally , Kingsbury Sarah R. , Stoeber Kai , Knight Gillian L. , Gallimore Phillip H. , Williams Gareth H. 
Published: July 2008

ABSTRACT

Productive infections by human papillomaviruses (HPVs) are restricted to nondividing, differentiated keratinocytes. HPV early proteins E6 and E7 deregulate cell cycle progression and activate the host cell DNA replication machinery in these cells, changes essential for virus synthesis. Productive virus replication is accompanied by abundant expression of the HPV E4 protein. Expression of HPV1 E4 in cells is known to activate cell cycle checkpoints, inhibiting G
2
-to-M transition of the cell cycle and also suppressing entry of cells into S phase. We report here that the HPV1 E4 protein, in the presence of a soluble form of the replication-licensing factor (RLF) Cdc6, inhibits initiation of cellular DNA replication in a mammalian cell-free DNA replication system. Chromatin-binding studies show that E4 blocks replication initiation in vitro by preventing loading of the RLFs Mcm2 and Mcm7 onto chromatin. HPV1 E4-mediated replication inhibition in vitro and suppression of entry of HPV1 E4-expressing cells into S phase are both abrogated upon alanine replacement of arginine 45 in the full-length E4 protein (E1^E4), implying that these two HPV1 E4 functions are linked. We hypothesize that HPV1 E4 inhibits competing host cell DNA synthesis in replication-activated suprabasal keratinocytes by suppressing licensing of cellular replication origins, thus modifying the phenotype of the infected cell in favor of viral genome amplification.
SKELETON-BASED SHAPE CLASSIFICATION USING PATH SIMILARITY
Authors: BAI XIANG , YANG XINGWEI , YU DEGUANG , LATECKI LONGIN JAN 
Published: July 2008

Most of the traditional methods for shape classification are based on contour. They often encounter difficulties when dealing with classes that have large nonlinear variability, especially when the variability is structural or due to articulation. It is well-known that shape representation based on skeletons is superior to contour based representation in such situations. However, approaches to shape similarity based on skeletons suffer from the instability of skeletons, and matching of skeleton graphs is still an open problem. Using a new skeleton pruning method, we are able to obtain stable pruned skeletons even in the presence of significant contour distortions. We also propose a new method for matching of skeleton graphs. In contrast to most existing methods, it does not require converting of skeleton graphs to trees and it does not require any graph editing. Shape classification is done with Bayesian classifier. We present excellent classification results for complete shapes.
NONHYPERBOLIC BOUNDARY EQUILIBRIUM BIFURCATIONS IN PLANAR FILIPPOV SYSTEMS: A CASE STUDY APPROACH
Authors: DI BERNARDO MARIO , PAGANO DANIEL J. , PONCE ENRIQUE 
Published: July 2008

Boundary equilibrium bifurcations in piecewise smooth discontinuous systems are characterized by the collision of an equilibrium point with the discontinuity surface. Generically, these bifurcations are of codimension one, but there are scenarios where the phenomenon can be of higher codimension. Here, the possible collision of a nonhyperbolic equilibrium with the boundary in a two-parameter framework and the nonlinear phenomena associated with such collision are considered. By dealing with planar discontinuous (Filippov) systems, some of such phenomena are pointed out through specific representative cases. A methodology for obtaining the corresponding biparametric bifurcation sets is developed.