bluuebunny/crossref_metadata_2025_split huggingface dataset:
Detection and Genotyping of Human Rotavirus VP4 and VP7 Genes by Reverse Transcriptase PCR and Reverse Hybridization
Authors: van Doorn Leen-Jan , Kleter Bernhard , Hoefnagel Evert , Stainier Isabelle , Poliszczak Annick , Colau Brigitte , Quint Wim 
Published: June 2009

ABSTRACT
Rotavirus infections can be diagnosed in stool samples by serological and molecular methods. We developed a novel reverse transcriptase PCR (RT-PCR) method for the amplification of rotavirus RNA and a reverse hybridization assay on a strip to detect amplimers and identify the specific G and P genotypes present in human stool specimens. An additional aim was to permit specific identification of the rotavirus G1P[8] strain, used in the Rotarix vaccine. Novel broad-spectrum PCR primers were developed for both VP4 and VP7, permitting the amplification of a wide range of rotavirus genotypes. Primer sets comprise mixtures of defined primer sequences. For the identification of G and P genotypes, two reverse hybridization strip assays were developed. Both the VP4 and the VP7 strip contain universal probes for the detection of VP4 and VP7 sequences, irrespective of the G or P genotype. The VP4 strip contains type-specific probes for P[4], P[6], P[8], P[9], and P[10]. The VP7 strip contains type-specific probes for G1, G2, G3, G4, G5, G6, G8, and G9. In addition, probes to distinguish between wild-type G1 and G1 vaccine strain sequences were present. Testing by analysis of multiple reference strains confirmed that both RT-PCR methods allowed the detection of a broad spectrum of genotypes. RT-PCR for VP7 was more sensitive than RT-PCR for VP4, but all samples identified as positive for rotavirus antigen by an enzyme-linked immunosorbent assay (ELISA) were also positive for both VP4 and VP7. The high specificity of the reverse hybridization method was confirmed by sequence analysis as well as by type-specific PCR, and the vaccine strain could also be specifically identified. The reverse hybridization method permits accurate identification of mixed infections with different genotypes. Rotavirus genotypes for which no type-specific probes were present on the strip were adequately identified by the universal detection probes. The assay was formally validated by analyses of specificity, sensitivity, precision, accuracy, and robustness. In a panel of 149 ELISA-positive stool samples, comparison with conventional type-specific RT-PCR methods revealed the superiority of the novel method, mainly in cases of mixed rotavirus infections. This novel method permits highly accurate detection and identification of human rotavirus infections in stool samples. This validated assay could be useful for large-scale epidemiological and clinical trials.
Plasmid-Borne Virulence-Associated Genes Have a Conserved Organization in Virulent Strains of Avian Pathogenic
Escherichia coli
Authors: Tivendale Kelly A. , Allen Joanne L. , Browning Glenn F. 
Published: June 2009

ABSTRACT

Avian pathogenic
Escherichia coli
(APEC) is an important respiratory pathogen of poultry. Various virulence factors are responsible for determining the pathogenicity of these strains, and it is commonly believed they are encoded on large plasmids the strains carry. This study examined a series of strains, the pathogenicity of which had previously been determined by aerosol exposure, for possession of large plasmids and found all isolates carried at least one large plasmid, regardless of the level of virulence. Virulence-associated genes carried on these plasmids were also examined, and it was shown that highly virulent strains carried at least four virulence-associated genes on their largest plasmid. Two of the virulence-associated genes were shown to be chromosomally located in a strain of intermediate virulence, while no virulence-associated genes were carried by the low-virulence strain. The organization of the virulence-associated genes was shown to be highly conserved among APEC isolates of high virulence, supporting the concept of a conserved portion of the putative virulence region that contributes to the pathogenicity of APEC strains.
Use of a New Single Multiplex PCR-Based Assay for Direct Simultaneous Characterization of Six
Neisseria meningitidis
Serogroups
Authors: Fraisier Christophe , Stor Richard , Tenebray Bernard , Sanson Yannick , Nicolas Pierre 
Published: June 2009

ABSTRACT

We developed a new
Neisseria meningitidis
multiplex PCR to determine six serogroups, including X-specific primers, and to allow direct W135/Y discrimination. This assay offers a simple and low-cost method for serogrouping
N
.
meningitidis
from cerebrospinal fluid that could be useful in Africa.
The Ubiquitin-Specific Peptidase USP15 Regulates Human Papillomavirus Type 16 E6 Protein Stability
Authors: Vos Robin M. , Altreuter Jennifer , White Elizabeth A. , Howley Peter M. 
Published: June 2009

ABSTRACT
Proteomic identification of human papillomavirus type 16 (HPV16) E6-interacting proteins revealed several proteins involved in ubiquitin-mediated proteolysis. In addition to the well-characterized E6AP ubiquitin-protein ligase, a second HECT domain protein (HERC2) and a deubiquitylating enzyme (USP15) were identified by tandem affinity purification of HPV16 E6-associated proteins. This study focuses on the functional consequences of the interaction of E6 with USP15. Overexpression of USP15 resulted in increased levels of the E6 protein, and the small interfering RNA-mediated knockdown of USP15 decreased E6 protein levels. These results implicate USP15 directly in the regulation of E6 protein stability and suggest that ubiquitylated E6 could be a substrate for USP15 ubiquitin peptidase activity. It remains possible that E6 could affect the activity of USP15 on specific cellular substrates, a hypothesis that can be tested as more is learned about the substrates and pathways controlled by USP15.
Virus Entry via the Alternative Coreceptors CCR3 and FPRL1 Differs by Human Immunodeficiency Virus Type 1 Subtype
Authors: Nedellec R. , Coetzer M. , Shimizu N. , Hoshino H. , Polonis V. R. , Morris L. , Mårtensson U. E. A. , Binley J. , Overbaugh J. , Mosier D. E. 
Published: June 2009

ABSTRACT
Human immunodeficiency virus type 1 (HIV-1) infects target cells by binding to CD4 and a chemokine receptor, most commonly CCR5. CXCR4 is a frequent alternative coreceptor (CoR) in subtype B and D HIV-1 infection, but the importance of many other alternative CoRs remains elusive. We have analyzed HIV-1 envelope (Env) proteins from 66 individuals infected with the major subtypes of HIV-1 to determine if virus entry into highly permissive NP-2 cell lines expressing most known alternative CoRs differed by HIV-1 subtype. We also performed linear regression analysis to determine if virus entry via the major CoR CCR5 correlated with use of any alternative CoR and if this correlation differed by subtype. Virus pseudotyped with subtype B Env showed robust entry via CCR3 that was highly correlated with CCR5 entry efficiency. By contrast, viruses pseudotyped with subtype A and C Env proteins were able to use the recently described alternative CoR FPRL1 more efficiently than CCR3, and use of FPRL1 was correlated with CCR5 entry. Subtype D Env was unable to use either CCR3 or FPRL1 efficiently, a unique pattern of alternative CoR use. These results suggest that each subtype of circulating HIV-1 may be subject to somewhat different selective pressures for Env-mediated entry into target cells and suggest that CCR3 may be used as a surrogate CoR by subtype B while FPRL1 may be used as a surrogate CoR by subtypes A and C. These data may provide insight into development of resistance to CCR5-targeted entry inhibitors and alternative entry pathways for each HIV-1 subtype.
Subtype-Specific Differences in the Human Immunodeficiency Virus Type 1 Reverse Transcriptase Connection Subdomain of CRF01_AE Are Associated with Higher Levels of Resistance to 3′-Azido-3′-Deoxythymidine
Authors: Delviks-Frankenberry Krista A. , Nikolenko Galina N. , Maldarelli Frank , Hase Saiki , Takebe Yutaka , Pathak Vinay K. 
Published: June 2009

ABSTRACT
We previously shown that mutations in the connection (CN) subdomain of human immunodeficiency virus type 1 (HIV-1) subtype B reverse transcriptase (RT) increase 3′-azido-3′-deoxythymidine (AZT) resistance in the context of thymidine analog mutations (TAMs) by affecting the balance between polymerization and RNase H activity. To determine whether this balance affects drug resistance in other HIV-1 subtypes, recombinant subtype CRF01_AE was analyzed. Interestingly, CRF01_AE containing TAMs exhibited 64-fold higher AZT resistance relative to wild-type B, whereas AZT resistance of subtype B containing the same TAMs was 13-fold higher, which in turn correlated with higher levels of AZT-monophosphate (AZTMP) excision on both RNA and DNA templates. The high level of AZT resistance exhibited by CRF01_AE was primarily associated with the T400 residue in wild-type subtype AE CN subdomain. An A400T substitution in subtype B enhanced AZT resistance, increased AZTMP excision on both RNA and DNA templates, and reduced RNase H cleavage. Replacing the T400 residue in CRF01_AE with alanine restored AZT sensitivity and reduced AZTMP excision on both RNA and DNA templates, suggesting that the T400 residue increases AZT resistance in CRF01_AE at least in part by directly increasing the efficiency of AZTMP excision. These results show for the first time that CRF01_AE exhibits higher levels of AZT resistance in the presence of TAMs and that this resistance is primarily associated with T400. Our results also show that mixing the RT polymerase, CN, and RNase H domains from different subtypes can underestimate AZT resistance levels, and they emphasize the need to develop subtype-specific genotypic and phenotypic assays to provide more accurate estimates of clinical drug resistance.
Measurement of Human Immunodeficiency Virus Type 1 Preintegration Transcription by Using Rev-Dependent Rev-CEM Cells Reveals a Sizable Transcribing DNA Population Comparable to That from Proviral Templates
Authors: Iyer Subashini R. , Yu Dongyang , Biancotto Angélique , Margolis Leonid B. , Wu Yuntao 
Published: June 2009

ABSTRACT
Preintegration transcription is an early process in human immunodeficiency virus type 1 infection and has been suggested to occur at a low level. The templates have also been suggested to represent a small population of nonintegrated viral DNA, particularly the two-long-terminal-repeat (2-LTR) circles. However, these determinations were made by either using PCR amplification of viral transcripts in bulk cell populations or utilizing the LTR-driving reporter cells that measure the synthesis of Tat. The intrinsic leakiness of LTR often makes the measurement of low-level viral transcription inaccurate. Since preintegration transcription also generates Rev, to eliminate the nonspecificity associated with the use of LTR alone we have developed a novel Rev-dependent indicator cell, Rev-CEM, to measure preintegration transcription based on the amount of Rev generated. In this report, using Rev-CEM cells, we demonstrate that preintegration transcription occurs on a much larger scale than expected. The transcribing population derived from nonintegrated viral DNA was comparable (at approximately 70%) to that derived from provirus in a productive viral replication cycle. Nevertheless, each nonintegrated viral DNA template exhibited a significant reduction in the level of transcriptional activity in the absence of integration. We also performed flow cytometry sorting of infected cells to identify viral templates. Surprisingly, our results suggest that the majority of 2-LTR circles are not active in directing transcription. It is likely that the nonintegrated templates are from the predominant DNA species, such as the full-length, linear DNA. Our results also suggest that a nonintegrating lentiviral vector can be as effective as an integrating vector in directing gene expression in nondividing cells, with the proper choice of an internal promoter.
Dexamethasone Treatment of Calves Latently Infected with Bovine Herpesvirus 1 Leads to Activation of the bICP0 Early Promoter, in Part by the Cellular Transcription Factor C/EBP-Alpha
Authors: Workman Aspen , Perez Sandra , Doster Alan , Jones Clinton 
Published: June 2009

ABSTRACT

Sensory neurons within trigeminal ganglia (TG) are the primary site for bovine herpesvirus 1 (BHV-1) latency. During latency, viral gene expression is restricted to the latency-related (LR) gene and the open reading frame ORF-E. We previously constructed an LR mutant virus that expresses LR RNA but not any of the known LR proteins. In contrast to calves latently infected with wild-type (wt) BHV-1 or the LR rescued virus, the LR mutant virus does not reactivate from latency following dexamethasone (DEX) treatment. In this study, we demonstrated that bICP0, but not bICP4, transcripts were consistently detected in TG of calves infected with the LR mutant or LR rescued virus following DEX treatment. Calves latently infected with the LR rescued virus but not the LR mutant virus expressed late transcripts, which correlated with shedding of infectious virus following DEX treatment. The bICP4 and bICP0 genes share a common immediate-early promoter, suggesting that this promoter was not consistently activated during DEX-induced reactivation from latency. The bICP0 gene also contains a novel early promoter that was activated by DEX in mouse neuroblastoma cells. Expression of a cellular transcription factor, C/EBP-alpha, was stimulated by DEX, and C/EBP-alpha expression was necessary for DEX induction of bICP0 early promoter activity. C/EBP-alpha directly interacted with bICP0 early promoter sequences that were necessary for
trans
activation by C/EBP-alpha. In summary, DEX treatment of latently infected calves induced cellular factors that stimulated bICP0 early promoter activity. Activation of bICP0 early promoter activity does not necessarily lead to late gene expression and virus shedding.
Tactile acuity and lumbopelvic motor control in patients with back pain and healthy controls
Authors: Luomajoki H , Moseley G L 
Published: June 2009

BackgroundVoluntary lumbopelvic control is compromised in patients with back pain. Loss of proprioceptive acuity is one contributor to decreased control. Several reasons for decreased proprioceptive acuity have been proposed, but the integrity of cortical body maps has been overlooked. We investigated whether tactile acuity, a clear clinical signature of primary sensory cortex organisation, relates to lumbopelvic control in people with back pain.MethodsForty-five patients with back pain and 45 age- and sex-matched healthy controls participated in this cross-sectional study. Tactile acuity at the back was assessed using two-point discrimination (TPD) threshold in vertical and horizontal directions. Voluntary motor control was assessed using an established battery of clinical tests.ResultsPatients performed worse on the voluntary lumbopelvic tasks than healthy controls did (p<0.001). TPD threshold was larger in patients (mean (SD)=61 (13) mm) than in healthy controls (44 (10) mm). Moreover, larger TPD threshold was positively related to worse performance on the voluntary lumbopelvic tasks (Pearson's r=0.49; p<0.001).DiscussionTactile acuity, a clear clinical signature of primary sensory cortex organisation, relates to voluntary lumbopelvic control. This relationship raises the possibility that the former contributes to the latter, in which case training tactile acuity may aid recovery and assist in achieving normal motor performance after back injury.
Traffic particles and occurrence of acute myocardial infarction: a case–control analysis
Authors: Tonne C , Yanosky J , Gryparis A , Melly S , Mittleman M , Goldberg R , von Klot S , Schwartz J 
Published: June 2009

Objectives:We modelled exposure to traffic particles using a latent variable approach and investigated whether long-term exposure to traffic particles is associated with an increase in the occurrence of acute myocardial infarction (AMI) using data from a population-based coronary disease registry.Methods:Cases of individually validated AMI were identified between 1995 and 2003 as part of the Worcester Heart Attack Study. Population controls were selected from Massachusetts, USA, resident lists. NO2and PM2.5filter absorbance were measured at 36 locations throughout the study area. The air pollution data were used to estimate exposure to traffic particles using a semiparametric latent variable regression model. Conditional logistic models were used to estimate the association between exposure to traffic particles and occurrence of AMI.Results:Modelled exposure to traffic particles was highest near the city of Worcester. Cases of AMI were more exposed to traffic and traffic particles compared to controls. An interquartile range increase in modelled traffic particles was associated with a 10% (95% CI 4% to 16%) increase in the odds of AMI. Accounting for spatial dependence at the census tract, but not block group, scale substantially attenuated this association.Conclusions:These results provide some support for an association between long-term exposure to traffic particles and risk of AMI. The results were sensitive to the scale selected for the analysis of spatial dependence, an issue that requires further investigation. The latent variable model captured variation in exposure, although on a relatively large spatial scale.
Accretionary orogens through Earth history
Authors: Cawood Peter A. , Kröner Alfred , Collins William J. , Kusky Timothy M. , Mooney Walter D. , Windley Brian F. 
Published: June 2009

AbstractAccretionary orogens form at intraoceanic and continental margin convergent plate boundaries. They include the supra-subduction zone forearc, magmatic arc and back-arc components. Accretionary orogens can be grouped into retreating and advancing types, based on their kinematic framework and resulting geological character. Retreating orogens (e.g. modern western Pacific) are undergoing long-term extension in response to the site of subduction of the lower plate retreating with respect to the overriding plate and are characterized by back-arc basins. Advancing orogens (e.g. Andes) develop in an environment in which the overriding plate is advancing towards the downgoing plate, resulting in the development of foreland fold and thrust belts and crustal thickening. Cratonization of accretionary orogens occurs during continuing plate convergence and requires transient coupling across the plate boundary with strain concentrated in zones of mechanical and thermal weakening such as the magmatic arc and back-arc region. Potential driving mechanisms for coupling include accretion of buoyant lithosphere (terrane accretion), flat-slab subduction, and rapid absolute upper plate motion overriding the downgoing plate. Accretionary orogens have been active throughout Earth history, extending back until at least 3.2 Ga, and potentially earlier, and provide an important constraint on the initiation of horizontal motion of lithospheric plates on Earth. They have been responsible for major growth of the continental lithosphere through the addition of juvenile magmatic products but are also major sites of consumption and reworking of continental crust through time, through sediment subduction and subduction erosion. It is probable that the rates of crustal growth and destruction are roughly equal, implying that net growth since the Archaean is effectively zero.
Metamorphic patterns in orogenic systems and the geological record
Authors: Brown Michael 
Published: June 2009

Abstract

Regional metamorphism occurs in plate boundary zones. Accretionary orogenic systems form at subduction boundaries in the absence of continent collision, whereas collisional orogenic systems form where ocean basins close and subduction steps back and flips (arc collisions), simply steps back and continues with the same polarity (block and terrane collisions) or ultimately ceases (continental collisions). As a result, collisional orogenic systems may be superimposed on accretionary orogenic systems. Metamorphism associated with orogenesis provides a mineral record that may be inverted to yield apparent thermal gradients for different metamorphic belts, which in turn may be used to infer tectonic setting. Potentially, peak mineral assemblages are robust recorders of metamorphic
P
and
T
, particularly at high
P
–
T
conditions, because prograde dehydration and melting with melt loss produce nominally anhydrous mineral assemblages that are difficult to retrogress or overprint without fluid influx. Currently on Earth, lower thermal gradients are associated with subduction (and early stages of collision) whereas higher thermal gradients are characteristic of back-arcs and orogenic hinterlands. This duality of thermal regimes is the hallmark of asymmetric or one-sided subduction and plate tectonics on modern Earth, and a duality of metamorphic belts will be the characteristic imprint of asymmetric or one-sided subduction in the geological record. Accretionary orogenic systems may exhibit retreating trench–advancing trench cycles, associated with high (>750 °C GPa
−1
) thermal gradient type of metamorphism, or advancing trench–retreating trench cycles, associated with low (<350 °C GPa
−1
) to intermediate (350–750 °C GPa
−1
) thermal gradient types of metamorphism. Whether the subducting boundary advances or retreats determines the mode of evolution. Accretionary orogenic systems may involve accretion of allochthonous and/or para-autochthonous elements to continental margins at subduction boundaries. Paired metamorphic belts,
sensu
Miyashiro, comprising a low thermal gradient metamorphic belt outboard and a high thermal gradient metamorphic belt inboard, are characteristic and may record orogen-parallel terrane migration and juxtaposition by accretion of contemporary belts of contrasting type. A wider definition of ‘paired’ metamorphism is proposed to incorporate all types of dual metamorphic belts. An additional feature is ridge subduction, which may be reflected in the pattern of high d
T
/d
P
metamorphism and associated magmatism. Apparent thermal gradients derived from inversion of age-constrained metamorphic
P
–
T
data are used to identify tectonic settings of ancient metamorphism, to evaluate the age distribution of metamorphism in the rock record from the Neoarchaean Era to the Cenozoic Era, and to consider how this relates to the supercontinent cycle and the process of terrane export and accretion. In addition, I speculate about metamorphism and tectonics before the Mesoarchaean Era.