bluuebunny/crossref_metadata_2025_split huggingface dataset:
Matrix-Assisted Laser Desorption Ionization–Time of Flight and Comparative Genomic Analysis of M-18 Group A Streptococcus Strains Associated with an Acute Rheumatic Fever Outbreak in Northeast Italy in 2012 and 2013
Authors: Gaibani Paolo , Scaltriti Erika , Foschi Claudio , Baggio Enrico , Tamburini Maria Vittoria , Creti Roberta , Pascucci Maria Grazia , Fagioni Marco , Ambretti Simone , Comandatore Francesco , Pongolini Stefano , Landini Maria Paola 
Published: March 2015

ABSTRACT

Acute rheumatic fever (ARF) is a postsuppurative sequela caused by
Streptococcus pyogenes
infections affecting school-age children. We describe here the occurrence of an ARF outbreak that occurred in Bologna province, northeastern Italy, between November 2012 and May 2013. Molecular analysis revealed that ARF-related group A
Streptococcus
(GAS) strains belonged to the M-18 serotype, including subtypes
emm18.29
and
emm18.32
. All M-18 GAS strains shared the same antigenic profile, including SpeA, SpeB, SpeC, SpeL, SpeM, and SmeZ. Matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) analysis revealed that M-18 GAS strains grouped separately from other serotypes, suggesting a different
S. pyogenes
lineage. Single nucleotide polymorphisms and phylogenetic analysis based on whole-genome sequencing showed that
emm18.29
and
emm18.32
GAS strains clustered in two distinct groups, highlighting genetic variations between these subtypes. Comparative analysis revealed a similar genome architecture between
emm18.29
and
emm18.32
strains that differed from noninvasive
emm18.0
strains. The major sources of differences between M-18 genomes were attributable to the prophage elements. Prophage regions contained several virulence factors that could have contributed to the pathogenic potential of
emm18.29
and
emm18.32
strains. Notably, phage ΦSPBO.1 carried erythrogenic toxin A gene (
speA1
) in six ARF-related M-18 GAS strains but not in
emm18.0
strains. In addition, a phage-encoded hyaluronidase gene (
hylP.2
) presented different variants among M-18 GAS strains by showing internal deletions located in the α-helical and TSβH regions. In conclusion, our study yielded insights into the genome structure of M-18 GAS strains responsible for the ARF outbreak in Italy, thus expanding our knowledge of this serotype.
Assessing the Public Health Risk of Shiga Toxin-Producing Escherichia coli by Use of a Rapid Diagnostic Screening Algorithm
Authors: de Boer Richard F. , Ferdous Mithila , Ott Alewijn , Scheper Henk R. , Wisselink Guido J. , Heck Max E. , Rossen John W. , Kooistra-Smid Anna M. D. 
Published: March 2015

ABSTRACT

Shiga toxin-producing
Escherichia coli
(STEC) is an enteropathogen of public health concern because of its ability to cause serious illness and outbreaks. In this prospective study, a diagnostic screening algorithm to categorize STEC infections into risk groups was evaluated. The algorithm consists of prescreening stool specimens with real-time PCR (qPCR) for the presence of
stx
genes. The qPCR-positive stool samples were cultured in enrichment broth and again screened for
stx
genes and additional virulence factors (
escV
,
aggR
,
aat
,
bfpA
) and O serogroups (O26, O103, O104, O111, O121, O145, O157). Also, PCR-guided culture was performed with sorbitol MacConkey agar (SMAC) and CHROMagar STEC medium. The presence of virulence factors and O serogroups was used for presumptive pathotype (PT) categorization in four PT groups. The potential risk for severe disease was categorized from high risk for PT group I to low risk for PT group III, whereas PT group IV consists of unconfirmed
stx
qPCR-positive samples. In total, 5,022 stool samples of patients with gastrointestinal symptoms were included. The qPCR detected
stx
genes in 1.8% of samples. Extensive screening for virulence factors and O serogroups was performed on 73 samples. After enrichment, the presence of
stx
genes was confirmed in 65 samples (89%). By culture on selective media, STEC was isolated in 36% (26/73 samples). Threshold cycle (

C
T

) values for
stx
genes were significantly lower after enrichment compared to direct qPCR (
P
< 0.001). In total, 11 (15%), 19 (26%), 35 (48%), and 8 (11%) samples were categorized into PT groups I, II, III, and IV, respectively. Several virulence factors (
stx
2
,
stx
2a
,
stx
2f
,
toxB
,
eae
,
efa1
,
cif
,
espA
,
tccP
,
espP
,
nleA
and/or
nleB
,
tir
cluster) were associated with PT groups I and II, while others (
stx
1
,
eaaA
,
mch
cluster,
ireA
) were associated with PT group III. Furthermore, the number of virulence factors differed between PT groups (analysis of variance,
P
< 0.0001). In conclusion, a diagnostic algorithm enables fast discrimination of STEC infections associated with a high to moderate risk for severe disease (PT groups I and II) from less-virulent STEC (PT group III).
Detection of Enterovirus D68 in Canadian Laboratories
Authors: Hatchette Todd F. , Drews Steven J. , Grudeski Elsie , Booth Tim , Martineau Christine , Dust Kerry , Garceau Richard , Gubbay Jonathan , Karnauchow Tim , Krajden Mel , Levett Paul N. , Mazzulli Tony , McDonald Ryan R. , McNabb Alan , Mubareka Samira , Needle Robert , Petrich Astrid , Richardson Susan , Rutherford Candy , Smieja Marek , Tellier Raymond , Tipples Graham , LeBlanc Jason J. 
Published: March 2015

ABSTRACT
The recent emergence of a severe respiratory disease caused by enterovirus D68 prompted investigation into whether Canadian hospital and provincial laboratories can detect this virus using commercial and laboratory-developed assays. This study demonstrated analytical sensitivity differences between commercial and laboratory-developed assays for the detection of enterovirus D68.
A Metagenomics and Case-Control Study To Identify Viruses Associated with Bovine Respiratory Disease
Authors: Ng Terry Fei Fan , Kondov Nikola O. , Deng Xutao , Van Eenennaam Alison , Neibergs Holly L. , Delwart Eric 
Published: March 2015

ABSTRACT

Bovine respiratory disease (BRD) is a common health problem for both dairy and beef cattle, resulting in significant economic loses. In order to identify viruses associated with BRD, we used a metagenomics approach to enrich and sequence viral nucleic acids in the nasal swabs of 50 young dairy cattle with symptoms of BRD. Following deep sequencing,
de novo
assembly, and translated protein sequence similarity searches, numerous known and previously uncharacterized viruses were identified. Bovine adenovirus 3, bovine adeno-associated virus, bovine influenza D virus, bovine parvovirus 2, bovine herpesvirus 6, bovine rhinitis A virus, and multiple genotypes of bovine rhinitis B virus were identified. The genomes of a previously uncharacterized astrovirus and picobirnaviruses were also partially or fully sequenced. Using real-time PCR, the rates of detection of the eight viruses that generated the most reads were compared for the nasal secretions of 50 animals with BRD versus 50 location-matched healthy control animals. Viruses were detected in 68% of BRD-affected animals versus 16% of healthy control animals. Thirty-eight percent of sick animals versus 8% of controls were infected with multiple respiratory viruses. Significantly associated with BRD were bovine adenovirus 3 (
P
< 0.0001), bovine rhinitis A virus (
P
= 0.005), and the recently described bovine influenza D virus (
P
= 0.006), which were detected either alone or in combination in 62% of animals with BRD. A metagenomics and real-time PCR detection approach in carefully matched cases and controls can provide a rapid means to identify viruses associated with a complex disease, paving the way for further confirmatory tests and ultimately to effective intervention strategies.

IMPORTANCE
Bovine respiratory disease is the most economically important disease affecting the cattle industry, whose complex root causes include environmental, genetics, and infectious factors. Using an unbiased metagenomics approach, we characterized the viruses in respiratory secretions from BRD cases and identified known and previously uncharacterized viruses belonging to seven viral families. Using a case-control format with location-matched animals, we compared the rates of viral detection and identified 3 viruses associated with severe BRD signs. Combining a metagenomics and case-control format can provide candidate pathogens associated with complex infectious diseases and inform further studies aimed at reducing their impact.
Increased Levels of Macrophage Inflammatory Proteins Result in Resistance to R5-Tropic HIV-1 in a Subset of Elite Controllers
Authors: Walker Wendy E. , Kurscheid Sebastian , Joshi Samit , Lopez Charlie A. , Goh Gerald , Choi Murim , Barakat Lydia , Francis John , Fisher Ann , Kozal Michael , Zapata Heidi , Shaw Albert , Lifton Richard , Sutton Richard E. , Fikrig Erol 
Published: March 2015

ABSTRACT

Elite controllers (ECs) are a rare group of HIV seropositive individuals who are able to control viral replication without antiretroviral therapy. The mechanisms responsible for this phenotype, however, have not been fully elucidated. In this study, we examined CD4
+
T cell resistance to HIV in a cohort of elite controllers and explored transcriptional signatures associated with cellular resistance. We demonstrate that a subgroup of elite controllers possess CD4
+
T cells that are specifically resistant to R5-tropic HIV while remaining fully susceptible to X4-tropic and vesicular stomatitis virus G (VSV-G)-pseudotyped viruses. Transcriptome analysis revealed 17 genes that were differentially regulated in resistant elite controllers relative to healthy controls. Notably, the genes encoding macrophage inflammatory protein 1α (MIP-1α),
CCL3
and
CCL3L1
, were found to be upregulated. The MIP-1α, MIP-1β, and RANTES chemokines are natural ligands of CCR5 and are known to interfere with HIV replication. For three elite controllers, we observed increased production of MIP-1α and/or MIP-1β at the protein level. The supernatant from resistant EC cells contained MIP-1α and MIP-1β and was sufficient to confer R5-tropic resistance to susceptible CD4
+
T cells. Additionally, this effect was reversed by using inhibitory anti-MIP antibodies. These results suggest that the T cells of these particular elite controllers may be naturally resistant to HIV infection by blocking R5-tropic viral entry.

IMPORTANCE
HIV is a pandemic health problem, and the majority of seropositive individuals will eventually progress to AIDS unless antiretroviral therapy (ART) is administered. However, rare patients, termed elite controllers, have a natural ability to control HIV infection in the absence of ART, but the mechanisms by which they achieve this phenotype have not been fully explored. This paper identifies one mechanism that may contribute to this natural resistance: some elite controllers have CD4
+
T cells that produce high levels of MIP chemokines, which block R5-tropic HIV entry. This mechanism could potentially be exploited to achieve a therapeutic effect in other HIV-seropositive individuals.
Retroviral Cyclin Controls Cyclin-Dependent Kinase 8-Mediated Transcription Elongation and Reinitiation
Authors: Birkenheuer Claire H. , Brewster Connie D. , Quackenbush Sandra L. , Rovnak Joel 
Published: March 2015

ABSTRACT

Walleye dermal sarcoma virus (WDSV) infection is associated with the seasonal development and regression of walleye dermal sarcoma. Previous work showed that the retroviral cyclin (RV-cyclin), encoded by WDSV, has separable cyclin box and transcription activation domains. It binds to cyclin-dependent kinase 8 (CDK8) and enhances its kinase activity. CDK8 is evolutionarily conserved and is frequently overexpressed in human cancers. It is normally activated by cyclin C and is required for transcription elongation of the serum response genes (immediate early genes [IEGs])
FOS
,
EGR1
, and
cJUN
. The IEGs drive cell proliferation, and their expression is brief and highly regulated. Here we show that constitutive expression of RV-cyclin in the HCT116 colon cancer cell line significantly increases the level of IEG expression in response to serum stimulation. Quantitative reverse transcription-PCR (RT-PCR) and nuclear run-on assays provide evidence that RV-cyclin does not alter the initiation of IEG transcription but does enhance the overall rate of transcription elongation and maintains transcription reinitiation. RV-cyclin does not increase activating phosphorylation events in the mitogen-activated protein kinase pathway and does not inhibit decay of IEG mRNAs. At the
EGR1
gene locus, RV-cyclin increases and maintains RNA polymerase II (Pol II) occupancy after serum stimulation, in conjunction with increased and extended
EGR1
gene expression. The RV-cyclin increases CDK8 occupancy at the
EGR1
gene locus before and after serum stimulation. Both of RV-cyclin's functional domains, i.e., the cyclin box and the activation domain, are necessary for the overall enhancement of IEG expression. RV-cyclin presents a novel and ancient mechanism of retrovirus-induced oncogenesis.

IMPORTANCE
The data reported here are important to both virology and cancer biology. The novel mechanism pinpoints CDK8 in the development of walleye dermal sarcoma and sheds light on CDK8's role in many human cancers. CDK8 controls expression from highly regulated genes, including the interferon-stimulated genes. Its function is likely the target of many viral interferon-resistance mechanisms. CDK8 also controls cellular responses to metabolic stimuli, stress, and hypoxia, in addition to the serum response. The retroviral cyclin (RV-cyclin) represents a highly selected probe of CDK8 function. RV-cyclin does not control CDK8 specificity but instead enhances CDK8's effects on regulated genes, an important distinction for its use to delineate natural CDK8 targets. The outcomes of this research are applicable to investigations of normal and abnormal CDK8 functions. The mechanisms defined here will contribute directly to the dermal sarcoma model in fish and clarify an important path for oncogenesis and innate resistance to viruses.
Protease Inhibitors Block Multiple Functions of the NS3/4A Protease-Helicase during the Hepatitis C Virus Life Cycle
Authors: McGivern David R. , Masaki Takahiro , Lovell William , Hamlett Chris , Saalau-Bethell Susanne , Graham Brent 
Published: March 2015

ABSTRACT
Hepatitis C virus (HCV) NS3 is a multifunctional protein composed of a protease domain and a helicase domain linked by a flexible linker. Protease activity is required to generate viral nonstructural (NS) proteins involved in RNA replication. Helicase activity is required for RNA replication, and genetic evidence implicates the helicase domain in virus assembly. Binding of protease inhibitors (PIs) to the protease active site blocks NS3-dependent polyprotein processing but might impact other steps of the virus life cycle. Kinetic analyses of antiviral suppression of cell culture-infectious genotype 1a strain H77S.3 were performed using assays that measure different readouts of the viral life cycle. In addition to the active-site PI telaprevir, we examined an allosteric protease-helicase inhibitor (APHI) that binds a site in the interdomain interface. By measuring nucleotide incorporation into HCV genomes, we found that telaprevir inhibits RNA synthesis as early as 12 h at high but clinically relevant concentrations. Immunoblot analyses showed that NS5B abundance was not reduced until after 12 h, suggesting that telaprevir exerts a direct effect on RNA synthesis. In contrast, the APHI could partially inhibit RNA synthesis, suggesting that the allosteric site is not always available during RNA synthesis. The APHI and active-site PI were both able to block virus assembly soon (<12 h) after drug treatment, suggesting that they rapidly engage with and block a pool of NS3 involved in assembly. In conclusion, PIs and APHIs can block NS3 functions in RNA synthesis and virus assembly, in addition to inhibiting polyprotein processing.

IMPORTANCE
The NS3/4A protease of hepatitis C virus (HCV) is an important antiviral target. Currently, three PIs have been approved for therapy of chronic hepatitis C, and several others are in development. NS3-dependent cleavage of the HCV polyprotein is required to generate the mature nonstructural proteins that form the viral replicase. Inhibition of protease activity can block RNA replication by preventing expression of mature replicase components. Like many viral proteins, NS3 is multifunctional, but how PIs affect stages of the HCV life cycle beyond polyprotein processing has not been well studied. Using cell-based assays, we show here that PIs can directly inhibit viral RNA synthesis and also block a late stage in virus assembly/maturation at clinically relevant concentrations.
Mutational Analysis of Vaccinia Virus E3 Protein: the Biological Functions Do Not Correlate with Its Biochemical Capacity To Bind Double-Stranded RNA
Authors: Dueck Kevin J. , Hu YuanShen (Sandy) , Chen Peter , Deschambault Yvon , Lee Jocelyn , Varga Jessie , Cao Jingxin 
Published: March 2015

ABSTRACT
Vaccinia E3 protein has the biochemical capacity of binding to double-stranded RNA (dsRNA). The best characterized biological functions of the E3 protein include its host range function, suppression of cytokine expression, and inhibition of interferon (IFN)-induced antiviral activity. Currently, the role of the dsRNA binding capacity in the biological functions of the E3 protein is not clear. To further understand the mechanism of the E3 protein biological functions, we performed alanine scanning of the entire dsRNA binding domain of the E3 protein to examine the link between its biochemical capacity of dsRNA binding and biological functions. Of the 115 mutants examined, 20 were defective in dsRNA binding. Although the majority of the mutants defective in dsRNA binding also showed defective replication in HeLa cells, nine mutants (I105A, Y125A, E138A, F148A, F159A, K171A, L182A, L183A, and I187/188A) retained the host range function to various degrees. Further examination of a set of representative E3L mutants showed that residues essential for dsRNA binding are not essential for the biological functions of E3 protein, such as inhibition of protein kinase R (PKR) activation, suppression of cytokine expression, and apoptosis. Thus, data described in this communication strongly indicate the E3 protein performs its biological functions via a novel mechanism which does not correlate with its dsRNA binding activity.

IMPORTANCE
dsRNAs produced during virus replication are important pathogen-associated molecular patterns (PAMPs) for inducing antiviral immune responses. One of the strategies used by many viruses to counteract such antiviral immune responses is achieved by producing dsRNA binding proteins, such as poxvirus E3 family proteins, influenza virus NS1, and Ebola virus V35 proteins. The most widely accepted model for the biological functions of this class of viral dsRNA binding proteins is that they bind to and sequester viral dsRNA PAMPs; thus, they suppress the related antiviral immune responses. However, no direct experimental data confirm such a model. In this study of vaccinia E3 protein, we found that the biological functions of the E3 protein are not necessarily linked to its biochemical capacity of dsRNA binding. Thus, our data strongly point to a new concept of virus modulation of cellular antiviral responses triggered by dsRNA PAMPs.
Flexibility of Bunyavirus Genomes: Creation of an Orthobunyavirus with an Ambisense S Segment
Authors: van Knippenberg Ingeborg , Elliott Richard M. 
Published: March 2015

ABSTRACT

The Bunyamwera (BUNV) orthobunyavirus NSs protein has proven a challenge to study in the context of viral infection. NSs is encoded in a reading frame that overlaps that of the viral nucleocapsid (N) protein thus limiting options for mutagenesis. In addition, NSs is poorly immunogenic, and antibodies only work in certain techniques while the protein itself is subject to proteasomal degradation. In order to generate a virus that expresses NSs independently of N, an ambisense S RNA segment was designed by mutating the 5′- and 3′-terminal nucleotide sequences. These mutations were previously shown to alter promoter activity so that both replication and transcription were promoted from both the genome and the antigenome RNAs (J. N. Barr et al., J. Virol.
79:
12602–12607, 2005). As proof of principle, a recombinant BUNV was created that expressed green fluorescent protein (GFP) in the ambisense orientation. GFP expression was detected throughout at least 10 passages. Recombinant BUNV encoding epitope-tagged versions of NSs in the ambisense orientation expressed NSs via a subgenomic mRNA, and two viruses grew to titers only modestly lower than parental rBUNdelNSs2 virus. The ambisense viruses were temperature sensitive, and NSs was shown to localize to both the nucleus and the cytoplasm during infection. These viruses will be useful in further studies on structure-function relationships of the orthobunyavirus NSs protein.

IMPORTANCE
Bunyamwera virus (BUNV) is the type species and model system for both the family
Bunyaviridae
and the genus
Orthobunyavirus
, a group that includes many significant human and animal pathogens. Studying the basic molecular biology of these viruses is of great importance to underpin research into vaccines and antivirals. We demonstrate here the plasticity of the BUNV genome by generating recombinant viruses where the normal negative-sense S segment has been converted into an ambisense segment, allowing independent expression of either a foreign gene (green fluorescent protein) or the viral nonstructural NSs protein. These new reagents will allow detailed investigation of NSs, the orthobunyavirus interferon antagonist.
Fatigue in rheumatoid arthritis; a persistent problem: a large longitudinal study
Authors: van Steenbergen Hanna W , Tsonaka Roula , Huizinga Tom W J , Boonen Annelies , van der Helm-van Mil Annette H M 
Published: March 2015

ObjectiveFatigue is prevalent and disabling in rheumatoid arthritis (RA). Surprisingly, the long-term course of fatigue is studied seldom and it is unclear to what extent it is influenced by inflammation. This study aimed to determine the course of fatigue during 8 years follow-up, its association with the severity of inflammation and the effect of improved treatment strategies.Methods626 patients with RA included in the Leiden Early Arthritis Clinic cohort were studied during 8 years. Fatigue severity, measured on a 0–100 mm scale, and other clinical variables were assessed yearly. Patients included in 1993–1995, 1996–1998 and 1999–2007 were treated with delayed treatment with disease-modifying antirheumatic drugs (DMARDs), early treatment with mild DMARDs and early treatment with methotrexate respectively. After multiple imputation, the serial measurements were analysed using linear quantile mixed models.ResultsMedian fatigue severity at baseline was 45 mm and remained, despite treatment, rather stable thereafter. Female gender (effect size=4.4 mm), younger age (0.2 mm less fatigue/year), higher swollen and tender joint counts (0.3 mm and 1.0 mm more fatigue/swollen or tender joint) and C reactive protein-levels (0.1 mm more fatigue per mg/L) were independently and significantly (p<0.05) associated with fatigue severity over 8 years. Although improved treatment strategies associated with less severe radiographic progression, there was no effect on fatigue severity (p=0.96).ConclusionsThis largest longitudinal study on fatigue so far demonstrated that the association between inflammation and fatigue is statistically significant but effect sizes are small, suggesting that non-inflammatory pathways mediate fatigue as well. Improved treatment strategies did not result in less severe fatigue. Therefore, fatigue in RA remains an ‘unmet need’.