bluuebunny/crossref_metadata_2025 huggingface dataset:
Toward Molecular Parasitologic Diagnosis: Enhanced Diagnostic Sensitivity for Filarial Infections in Mobile Populations
Authors: Fink Doran L. , Fahle Gary A. , Fischer Steven , Fedorko Daniel F. , Nutman Thomas B.
Published: October 2010

The diagnosis of filarial infections among individuals residing in areas where the disease is not endemic requires both strong clinical suspicion and expert training in infrequently practiced parasitological methods. Recently developed filarial molecular diagnostic assays are highly sensitive and specific but have limited availability and have not been closely evaluated for clinical use outside populations residing in areas of endemicity. In this study, we assessed the performance of a panel of real-time PCR assays for the four most common human filarial pathogens among blood and tissue samples collected from a cohort of patients undergoing evaluation for suspected filarial infections. Compared to blood filtration, real-time PCR was equally sensitive for the detection of microfilaremia due to
Wuchereria
bancrofti
(2 of 46 samples positive by both blood filtration and PCR with no discordant results) and
L
oa
loa
(24 of 208 samples positive by both blood filtration and PCR, 4 samples positive by PCR only, and 3 samples positive by blood filtration only). Real-time PCR of skin snip samples was significantly more sensitive than microscopic examination for the detection of
Onchocerca
volvulus
microfiladermia (2 of 218 samples positive by both microscopy and PCR and 12 samples positive by PCR only). The molecular assays required smaller amounts of blood and tissue than conventional methods and could be performed by laboratory personnel without specialized parasitology training. Taken together, these data demonstrate the utility of the molecular diagnosis of filarial infections in mobile populations.
Comparison of Real-Time PCR Techniques to Cytotoxigenic Culture Methods for Diagnosing
Clostridium difficile
Infection
Authors: Knetsch C. W. , Bakker D. , de Boer R. F. , Sanders I. , Hofs S. , Kooistra-Smid A. M. D. , Corver J. , Eastwood K. , Wilcox M. H. , Kuijper E. J.
Published: October 2010

In the past decade, the incidence of
Clostridium difficile
infections (CDI) with a more severe course has increased in Europe and North America. Assays that are capable of rapidly diagnosing CDI are essential. Two real-time PCRs (LUMC and LvI) targeting
C. difficile
toxin genes (
tcdB
, and
tcdA
and
tcdB
, respectively) were compared with the BD GeneOhm PCR (targeting the
tcdB
gene), using cytotoxigenic culture as a gold standard. In addition, a real-time PCR targeting the
tcdC
frameshift mutation at position 117 (Δ117 PCR) was evaluated for detecting toxigenic
C. difficile
and the presence of PCR ribotype 027 in stool samples. In total, 526 diarrheal samples were prospectively collected and included in the study. Compared with those for cytotoxigenic culture, sensitivity, specificity, positive predicted value (PPV), and negative predicted value (NPV) were for PCR LUMC 96.0%, 88.0%, 66.0%, and 98.9%, for PCR LvI 100.0%, 89.4%, 69.7%, and 100.0%, for PCR Δ117 98.0%, 90.7%, 71.9%, and 99.5%, and for PCR BD GeneOhm 88.3%, 96.9%, 86.5%, and 97.4%. Compared to those with feces samples cultured positive for
C. difficile
type 027, the sensitivity, specificity, PPV, and NPV of the Δ117 PCR were 95.2%, 96.2%, 87.0%, and 98.7%. We conclude that all real-time PCRs can be applied as a first screening test in an algorithm for diagnosing CDI. However, the low PPVs hinder the use of the assays as stand-alone tests. Furthermore, the Δ117 PCR may provide valuable information for minimizing the spread of the epidemic
C. difficile
PCR ribotype 027.
National Surveillance of Fungemia in Denmark (2004 to 2009)
Authors: Arendrup Maiken Cavling , Bruun Brita , Christensen Jens Jørgen , Fuursted Kurt , Johansen Helle Krogh , Kjældgaard Poul , Knudsen Jenny Dahl , Kristensen Lise , Møller Jens , Nielsen Lene , Rosenvinge Flemming Schønning , Røder Bent , Schønheyder Henrik Carl , Thomsen Marianne K. , Truberg Kjeld
Published: October 2010

A 6-year nationwide study of fungemia in Denmark was performed using data from an active fungemia surveillance program and from laboratory information systems in nonparticipating regions. A total of 2,820 episodes of fungemia were recorded. The incidence increased from 2004 to 2007 (7.7 to 9.6/100,000) and decreased slightly from 2008 to 2009 (8.7 to 8.6/100,000). The highest incidences were seen at the extremes of age (i.e., 11.3 and 37.1/100,000 for those <1 and 70 to 79 years old, respectively). The rate was higher for males than for females (10.1 versus 7.6/100,000,
P
= 0.003), with the largest difference observed for patients >50 years of age. The species distribution varied significantly by both age and gender.
Candida
species accounted for 98% of the pathogens, and
C. albicans
was predominant, although the proportion decreased (64.4% to 53.2%,
P
< 0.0001).
C. glabrata
ranked second, and the proportion increased (16.5% to 25.9%,
P
= 0.003).
C. glabrata
was more common in adults and females than in children and males, whereas
C. tropicalis
was more common in males (
P
= 0.020)
. C. krusei
was a rare isolate (4.1%) except at one university hospital. Acquired resistance to amphotericin and echinocandins was rare. However, resistance to fluconazole (MIC of >4 μg/ml) occurred in
C. albicans
(7/1,183 [0.6%]),
C. dubliniensis
(2/65 [3.1%]),
C. parapsilosis
(5/83 [6.0%]), and
C. tropicalis
(7/104 [6.7%]). Overall, 70.8% of fungemia isolates were fully fluconazole susceptible, but the proportion decreased (79.7% to 68.9%,
P
= 0.02). The study confirmed an incidence rate of fungemia in Denmark three times higher than those in other Nordic countries and identified marked differences related to age and gender. Decreased susceptibility to fluconazole was frequent and increasing.
Are Nasal Carriers of
Staphylococcus aureus
More Likely To Become Colonized or Infected with Methicillin-Resistant
Staphylococcus aureus
on Admission to a Hospital?
Authors: Krebes Juliane , Al-Ghusein Hasan , Feasey Nick , Breathnach Aodhan , Lindsay Jodi A.
Published: October 2010

Of 840 patients at hospital admission, 2.7% were positive for methicillin-resistant
Staphylococcus aureus
(MRSA) and 22.3% were positive for methicillin-susceptible
S. aureus
(MSSA). During the next 8 months, 4.8% of the MSSA-positive patients acquired MRSA with no lineage association. A total of 5.2% of noncarriers acquired MRSA. We find no evidence that colonized hosts are more susceptible to acquiring MRSA.
Poliovirus RNA Is Released from the Capsid near a Twofold Symmetry Axis
Authors: Bostina Mihnea , Levy Hazel , Filman David J. , Hogle James M.
Published: October 2010

After recognizing and binding to its host cell, poliovirus (like other nonenveloped viruses) faces the challenge of translocating its genome across a cellular membrane and into the cytoplasm. To avoid entanglement with the capsid, the RNA must exit via a single site on the virion surface. However, the mechanism by which a single site is selected (from among 60 equivalents) is unknown; and until now, even its location on the virion surface has been controversial. To help to elucidate the mechanism of infection, we have used single-particle cryo-electron microscopy and tomography to reconstruct conformationally altered intermediates that are formed by the poliovirion at various stages of the poliovirus infection process. Recently, we reported icosahedrally symmetric structures for two forms of the end-state 80S empty capsid particle. Surprisingly, RNA was frequently visible near the capsid; and in a subset of the virions, RNA was seen on both the inside and outside of the capsid, caught in the act of exiting. To visualize RNA exiting, we have now determined asymmetric reconstructions from that subset, using both single-particle cryo-electron microscopy and cryo-electron tomographic methods, producing independent reconstructions at ∼50-Å resolution. Contrary to predictions in the literature, the footprint of RNA on the capsid surface is located close to a viral 2-fold axis, covering a slot-shaped area of reduced density that is present in both of the symmetrized 80S reconstructions and which extends by about 20 Å away from the 2-fold axis toward each neighboring 5-fold axis.
Domain Structure of Lassa Virus L Protein
Authors: Brunotte Linda , Lelke Michaela , Hass Meike , Kleinsteuber Katja , Becker-Ziaja Beate , Günther Stephan
Published: October 2010

The 200-kDa L protein of arenaviruses plays a central role in viral genome replication and transcription. This study aimed at providing evidence for the domain structure of L protein by combining bioinformatics with a stepwise mutagenesis approach using the Lassa virus minireplicon system. Potential interdomain linkers were predicted using various algorithms. The prediction was challenged by insertion of flexible sequences into the predicted linkers. Insertion of 5 or 10 amino acid residues was tolerated at seven sites (S407, G446, G467, G774, G939, S1952, and V2074 in Lassa virus AV). At two of these sites, G467 and G939, L protein could be split into an N-terminal and a C-terminal part, which were able to
trans
-complement each other and reconstitute a functional complex upon coexpression. Coimmunoprecipitation studies revealed physical interaction between the N- and C-terminal domains, irrespective of whether L protein was split at G467 or G939. In confocal immunofluorescence microscopy, the N-terminal domains showed a dot-like, sometimes perinuclear, cytoplasmic distribution similar to that of full-length L protein, while the C-terminal domains were homogenously distributed in cytoplasm. The latter were redistributed into the dot-like structures upon coexpression with the corresponding N-terminal domain. In conclusion, this study demonstrates two interdomain linkers in Lassa virus L protein, at G467 and G939, suggesting that L protein is composed of at least three structural domains spanning residues 1 to 467, 467 to 939, and 939 to 2220. The first domain seems to mediate accumulation of L protein into cytoplasmic dot-like structures.
A Recombinant Measles Virus Unable To Antagonize STAT1 Function Cannot Control Inflammation and Is Attenuated in Rhesus Monkeys
Authors: Devaux Patricia , Hudacek Andrew W. , Hodge Gregory , Reyes-del Valle Jorge , McChesney Michael B. , Cattaneo Roberto
Published: October 2010

Measles remains a leading cause of death worldwide among children because it suppresses immune function. The measles virus (MV) P gene encodes three proteins (P, V, and C) that interfere with innate immunity, controlling STAT1, STAT2, mda5, and perhaps other key regulators of immune function. We identified here three residues in the shared domain of the P and V proteins—tyrosine 110, valine 112, and histidine 115—that function to retain STAT1 in the cytoplasm and inhibit interferon transcription. This information was used to generate a recombinant measles virus unable to antagonize STAT1 function (STAT1-blind MV) differing only in these three residues from a wild-type strain of well-defined virulence. This virus was used to assess the relevance of P and V interactions with STAT1 for virulence in primates. When a group of six rhesus monkeys (
Macaca mulatta
) was inoculated intranasally with STAT1-blind MV, viremia was short-lived, and the skin rash and other clinical signs observed with wild-type MV were absent. The STAT1-blind virus less efficiently controlled the inflammatory response, as measured by enhanced transcription of interleukin-6 and tumor necrosis factor alpha in peripheral blood mononuclear cells from infected hosts. Importantly, neutralizing antibody titers and MV-specific T-cell responses were equivalent in hosts infected with either virus. These findings indicate that efficient MV interactions with STAT1 are required to sustain virulence in a natural host by controlling the inflammatory response against the virus. They also suggest that selectively STAT1-blind MV may have utility as vectors for targeted oncolysis and vaccination.
Differential Regulation of Human Papillomavirus Type 8 by Interferon Regulatory Factors 3 and 7
Authors: Oldak Monika , Tolzmann Liv , Wnorowski Artur , Podgórska Marta Justyna , Silling Steffi , Lin Rongtuan , Hiscott John , Müller Cornelia Sigrid Lissi , Vogt Thomas , Smola Hans , Smola Sigrun
Published: October 2010

The genus β human papillomavirus (HPV) type 8 is associated with nonmelanoma skin cancer in patients with epidermodysplasia verruciformis, and evidence for its protumorigenic potential in the general population increases. To date, strategies to suppress genus β HPV infections are limited. Interferon regulatory factors IRF-3 and IRF-7 play key roles in the activation of the innate immune response to viral infections. In this study, we show for the first time that both IRF-3 and IRF-7 regulate transcription of a papillomavirus, but with opposing effects. IRF-7, expressed in the suprabasal layers of human epidermis, increased HPV8 late promoter activity via direct binding to viral DNA. UV-B light-induced activation of the HPV8 promoter involved IRF-7 as a downstream effector. In contrast, IRF-3, expressed in all layers of human epidermis, induced strong HPV8 suppression in primary keratinocytes. IRF-3-mediated suppression prevailed over IRF-7-induced HPV8 transcription. Unlike the E6 oncoprotein of the mucosal high-risk HPV16, the HPV8 E6 protein did not bind to IRF-3 and only weakly antagonized its activity. Strong antiviral activity was also observed, when keratinocytes were treated with potent IRF-3 activators, poly(I:C) or RNA bearing 5′ phosphates. In conclusion, we show that IRF-3 activation induces a state of cell-autonomous immunity against HPV in primary human keratinocytes. Our study suggests that local application of IRF-3-activating compounds might constitute an attractive novel therapeutic strategy against HPV8-associated diseases, particularly in epidermodysplasia verruciformis patients.
Activation of Plasmacytoid Dendritic Cells by Kaposi's Sarcoma-Associated Herpesvirus
Authors: West John A. , Gregory Sean M. , Sivaraman Vijay , Su Lishan , Damania Blossom
Published: October 2010

Kaposi's sarcoma-associated herpesvirus (KSHV) is associated with multiple human malignancies, including Kaposi's sarcoma, primary effusion lymphoma, and multicentric Castleman's disease. Following primary infection, KSHV typically goes through a brief period of lytic replication prior to the establishment of latency. Plasmacytoid dendritic cells (pDCs) are the major producers of type 1 interferon (IFN), primarily in response to virus infection. Toll-like receptors (TLRs) are key components of the innate immune system, and they serve as pathogen recognition receptors that stimulate the host antiviral response. pDCs express exclusively TLR7 and TLR9, and it is through these TLRs that the type 1 interferon response is activated in pDCs. Currently, it is not known whether KSHV is recognized by pDCs and whether activation of pDCs occurs in response to KSHV infection. We now report evidence that KSHV can infect human pDCs and that pDCs are activated upon KSHV infection, as measured by upregulation of CD83 and CD86 and by IFN-α secretion. We further show that induction of IFN-α occurs through activation of TLR9 signaling and that a TLR9 inhibitor diminishes the production and secretion of IFN-α by KSHV-infected pDCs.
A Vaccinia Virus Deletion Mutant Reveals the Presence of Additional Inhibitors of NF-κB
Authors: Fagan-Garcia Katharine , Barry Michele
Published: October 2010

The classical nuclear factor kappa B (NF-κB) signaling pathway is an important regulator of inflammation and innate immunity that is activated by a wide variety of stimuli, including virus infection, tumor necrosis factor alpha (TNF-α), and interleukin 1β (IL-1β). Poxviruses, including vaccinia virus (VV) and ectromelia virus, encode multiple proteins that function in immune evasion. Recently, a growing number of genes encoded by poxviruses have been shown to target and disrupt the NF-κB signaling pathway. To determine if additional gene products that interfere with NF-κB signaling existed, we used a vaccinia virus deletion mutant, VV811, which is missing 55 open reading frames lacking all known inhibitors of TNF-α-induced NF-κB activation. Immunofluorescence analysis of HeLa cells treated with TNF-α and IL-1β revealed that NF-κB translocation to the nucleus was inhibited in VV811-infected cells. This was further confirmed through Western blotting of cytoplasmic and nuclear extracts for NF-κB. Additionally, VV811 infection inhibited TNF-α-induced IκBα degradation. In contrast to vaccinia virus strain Copenhagen (VVCop)-infected cells, VV811 infection resulted in the dramatic accumulation of phosphorylated IκBα. Correspondingly, coimmunoprecipitation assays demonstrated that the NF-κB-inhibitory IκBα-p65-p50 complex was intact in VV811-infected cells. Significantly, cells treated with 1-β-
d
-arabinofuranosylcytosine, an inhibitor of poxvirus late gene expression, demonstrated that an additional vaccinia virus late gene was involved in the stabilization of IκBα. Overall, this work indicates that unidentified inhibitors of NF-κB exist in vaccinia virus. The complex inhibition of NF-κB by vaccinia virus illustrates the importance of NF-κB activation in the antiviral response.
Norovirus GII.4 Strain Antigenic Variation
Authors: Lindesmith Lisa C. , Donaldson Eric F. , Baric Ralph S.
Published: October 2010

Noroviruses are the principal cause of epidemic gastroenteritis worldwide. Multiple reports have concluded that the major capsid proteins of GII.4 strains, which cause 80% of norovirus infections worldwide, are evolving rapidly, resulting in new epidemic strains. Surrogate neutralization assays using sera from outbreaks and from immunized mice suggest that, as with influenza virus, antigenic variation maintains GII.4 persistence in the face of human population herd immunity. To test this hypothesis, mice were hyperimmunized with virus-like particles (VLPs) representing an early (GII.4-1987) and a contemporary (GII.4-2006) GII.4 strain. Anti-GII.4-1987 IgG monoclonal antibodies (MAbs) strongly reacted with GII.4 VLPs derived between only 1987 and 2002. Ligand binding blockade was more efficient with GII.4-1987 and GII.4-1997 VLPs than with GII.4-2002. Anti-GII.4-2006 IgG MAbs recognized either a broad panel of GII.4 VLPs (1987 to 2006) or a subset of contemporary (2004 to 2006) VLPs. Most 2006 antibodies did not recognize or only poorly recognized GII.4 VLPs of 2007 or 2008, documenting rapid antigenic evolution of GII.4 capsids. Generally, 2006 MAbs blocked homotypic VLP-ligand binding but were unable to block VLPs representing strains primarily circulating during or earlier than 2002. These analyses demonstrate that both subtle and significant evolutionary change has occurred within antibody epitopes between epidemic strains, providing direct evidence that the GII.4 noroviruses are undergoing antigenic variation, likely in response to herd immunity. As with influenza virus, HIV, and hepatitis C virus, norovirus antigenic variation will significantly influence the design of efficacious vaccines and immunotherapeutics against these important human pathogens.
A New Model of Epstein-Barr Virus Infection Reveals an Important Role for Early Lytic Viral Protein Expression in the Development of Lymphomas
Authors: Ma Shi-Dong , Hegde Subramanya , Young Ken H. , Sullivan Ruth , Rajesh Deepika , Zhou Ying , Jankowska-Gan Ewa , Burlingham William J. , Sun Xiaoping , Gulley Margaret L. , Tang Weihua , Gumperz Jenny E. , Kenney Shannon C.
Published: October 2010

Epstein-Barr virus (EBV) infects cells in latent or lytic forms, but the role of lytic infection in EBV-induced lymphomas is unclear. Here, we have used a new humanized mouse model, in which both human fetal CD34
+
hematopoietic stem cells and thymus/liver tissue are transplanted, to compare EBV pathogenesis and lymphoma formation following infection with a lytic replication-defective BZLF1-deleted (Z-KO) virus or a lytically active BZLF1
+
control. Both the control and Z-KO viruses established long-term viral latency in all infected animals. The infection appeared well controlled in some animals, but others eventually developed CD20
+
diffuse large B cell lymphomas (DLBCL). Animals infected with the control virus developed tumors more frequently than Z-KO virus-infected animals. Specific immune responses against EBV-infected B cells were generated in mice infected with either the control virus or the Z-KO virus. In both cases, forms of viral latency (type I and type IIB) were observed that are less immunogenic than the highly transforming form (type III) commonly found in tumors of immunocompromised hosts, suggesting that immune pressure contributed to the outcome of the infection. These results point to an important role for lytic EBV infection in the development of B cell lymphomas in the context of an active host immune response.
The 2009 Pandemic H1N1 and Triple-Reassortant Swine H1N1 Influenza Viruses Replicate Efficiently but Elicit an Attenuated Inflammatory Response in Polarized Human Bronchial Epithelial Cells
Authors: Zeng Hui , Pappas Claudia , Katz Jacqueline M. , Tumpey Terrence M.
Published: November 2010

The pandemic H1N1 virus of 2009 (2009 H1N1) produced a spectrum of disease ranging from mild illness to severe illness and death. Respiratory symptoms were frequently associated with virus infection, with relatively high rate of gastrointestinal symptoms reported. To better understand 2009 H1N1 virus pathogenesis in humans, we studied virus and host responses following infection of two cell types: polarized bronchial and pharyngeal epithelial cells, which exhibit many features of the human airway epithelium, and colon epithelial cells to serve as a human intestinal cell model. Selected 2009 H1N1 viruses were compared to both seasonal H1N1 and triple-reassortant swine H1N1 influenza viruses that have circulated among North American pigs since before the 2009 pandemic. All H1N1 viruses replicated productively in airway cells; however, in contrast to seasonal H1N1 virus infection, infection with the 2009 H1N1 and triple-reassortant swine H1N1 viruses resulted in an attenuated inflammatory response, a weaker interferon response, and reduced cell death. Additionally, the H1N1 viruses of swine origin replicated less efficiently at the temperature of the human proximal airways (33°C). We also observed that the 2009 H1N1 viruses replicated to significantly higher titers than seasonal H1N1 virus in polarized colon epithelial cells. These studies reveal that in comparison to seasonal influenza virus, H1N1 viruses of swine origin poorly activate multiple aspects of the human innate response, which may contribute to the virulence of these viruses. In addition, their less efficient replication at human upper airway temperatures has implications for the understanding of pandemic H1N1 virus adaptation to humans.
Dominant Inhibition of Akt/Protein Kinase B Signaling by the Matrix Protein of a Negative-Strand RNA Virus
Authors: Dunn Ewan F. , Connor John H.
Published: October 2010

Vesicular stomatitis virus (VSV) is a rhabdovirus that alters host nuclear and cytoplasmic function upon infection. We have investigated the effect of VSV infection on cellular signaling through the phosphatidylinositol-3 kinase (PI3k)/Akt signaling pathway. Akt phosphorylation at both threonine 308 (Thr308) and serine 473 (Ser473) was inhibited in cells infected with VSV. This inhibition was rapid (beginning within the first 2 to 3 h postinfection) and correlated with the dephosphorylation of downstream effectors of Akt, such as glycogen synthase kinase 3β (GSK3β) and mammalian target of rapamycin (mTOR). The dephosphorylation of Akt occurred in the presence of growth factor stimulation and was not overcome through constitutive membrane targeting of Akt or high levels of phosphatidylinositol-3,4,5-triphosphate (PIP3) accumulation in the membrane. Akt dephosphorylation was not a result of alterations in PDK1 phosphorylation or activity, changes in phosphatase and tensin homologue deleted on chromosome 10 (PTEN) levels, or the downregulation of PI3k signaling. Inactivation of Akt was caused by the expression of the viral M protein in the absence of other viral components, and an M protein mutant that does not inhibit RNA polymerase II (Pol II) transcription and nuclear/cytoplasmic transport was also defective in inhibiting Akt phosphorylation. These data illustrate that VSV utilizes a novel mechanism to alter this central player in cell signaling and oncogenesis. It also suggests an inside-out model of signal transduction where VSV interruption of nuclear events has a rapid and significant effect on membrane signaling events.
Function of the Small Hydrophobic Protein of J Paramyxovirus
Authors: Li Zhuo , Xu Jie , Patel Jui , Fuentes Sandra , Lin Yuan , Anderson Danielle , Sakamoto Kaori , Wang Lin-Fa , He Biao
Published: October 2010

At 18,954 nucleotides, the J paramyxovirus (JPV) genome is one of the largest in the family
Paramyxoviridae
, consisting of eight genes in the order 3′-N-P/V/C-M-F-SH-TM-G-L-5′. To study the function of novel paramyxovirus genes in JPV, a plasmid containing a full-length cDNA clone of the genome of JPV was constructed. In this study, the function of the small hydrophobic (SH) protein of JPV was examined by generating a recombinant JPV lacking the coding sequence of the SH protein (rJPVΔSH). rJPVΔSH was viable and had no growth defect in tissue culture cells. However, more tumor necrosis factor alpha (TNF-α) was produced during rJPVΔSH infection, suggesting that SH plays a role in inhibiting TNF-α production. rJPVΔSH induced more apoptosis in tissue culture cells than rJPV. Virus-induced apoptosis was inhibited by neutralizing antibody against TNF-α, suggesting that TNF-α contributes to JPV-induced apoptosis
in vitro
. The expression of JPV SH protein inhibited TNF-α-induced NF-κB activation in a reporter gene assay, suggesting that JPV SH protein can inhibit TNF-α signaling
in vitro
. Furthermore, infection of mice with rJPVΔSH induced more TNF-α expression, indicating that SH plays a role in blocking TNF-α expression
in vivo
.
Determinants That Specify the Integration Pattern of Retrotransposon Tf1 in the
fbp1
Promoter of
Schizosaccharomyces pombe
Authors: Majumdar Anasuya , Chatterjee Atreyi Ghatak , Ripmaster Tracy L. , Levin Henry L.
Published: October 2010

Long terminal repeat (LTR) retrotransposons are closely related to retroviruses and, as such, are important models for the study of viral integration and target site selection. The transposon Tf1 of
Schizosaccharomyces pombe
integrates with a strong preference for the promoters of polymerase II (Pol II)-transcribed genes. Previous work
in vivo
with plasmid-based targets revealed that the patterns of insertion were promoter specific and highly reproducible. To determine which features of promoters are recognized by Tf1, we studied integration in a promoter that has been characterized. The promoter of
fbp1
has two upstream activating sequences, UAS1 and UAS2. We found that integration was targeted to two windows, one 180 nucleotides (nt) upstream and the other 30 to 40 nt downstream of UAS1. A series of deletions in the promoter showed that the integration activities of these two regions functioned autonomously. Integration assays of UAS2 and of a synthetic promoter demonstrated that strong promoter activity alone was not sufficient to direct integration. The factors that modulate the transcription activities of UAS1 and UAS2 include the activators Atf1p, Pcr1p, and Rst2p as well as the repressors Tup11p, Tup12p, and Pka1p. Strains lacking each of these proteins revealed that Atf1p alone mediated the sites of integration. These data indicate that Atf1p plays a direct and specific role in targeting integration in the promoter of
fbp1
.
Structure-Based Analysis of the Interaction between the Simian Virus 40 T-Antigen Origin Binding Domain and Single-Stranded DNA
Authors: Meinke Gretchen , Phelan Paul J. , Fradet-Turcotte Amélie , Bohm Andrew , Archambault Jacques , Bullock Peter A.
Published: October 2010

The origin-binding domain (OBD) of simian virus 40 (SV40) large T-antigen (T-Ag) is essential for many of T-Ag's interactions with DNA. Nevertheless, many important issues related to DNA binding, for example, how single-stranded DNA (ssDNA) transits along the T-Ag OBD, have yet to be established. Therefore, X-ray crystallography was used to determine the costructure of the T-Ag OBD bound to DNA substrates such as the single-stranded region of a forked oligonucleotide. A second structure of the T-Ag OBD crystallized in the presence of poly(dT)
12
is also reported. To test the conclusions derived from these structures, residues identified as being involved in binding to ssDNA by crystallography or by an earlier nuclear magnetic resonance study were mutated, and their binding to DNA was characterized via fluorescence anisotropy. In addition, these mutations were introduced into full-length T-Ag, and these mutants were tested for their ability to support replication. When considered in terms of additional homology-based sequence alignments, our studies refine our understanding of how the T-Ag OBDs encoded by the polyomavirus family interact with ssDNA, a critical step during the initiation of DNA replication.
Translational Control of the Abundance of Cytoplasmic Poly(A) Binding Protein in Human Cytomegalovirus-Infected Cells
Authors: Perez Cesar , McKinney Caleb , Chulunbaatar Uyanga , Mohr Ian
Published: October 2010

Irrespective of their effects on ongoing host protein synthesis, productive replication of the representative alphaherpesvirus herpes simplex virus type 1, the representative gammaherpesvirus Kaposi's sarcoma herpesvirus, and the representative betaherpesvirus human cytomegalovirus [HCMV] stimulates the assembly of the multisubunit, cap-binding translation factor eIF4F. However, only HCMV replication is associated with an increased abundance of eIF4F core components (eIF4E, eIF4G, eIF4A) and the eIF4F-associated factor poly(A) binding protein (PABP). Here, we demonstrate that the increase in translation factor concentration was readily detected in an asynchronous population of HCMV-infected primary human fibroblasts, abolished by prior UV inactivation of virus, and genetically dependent upon viral immediate-early genes. Strikingly, while increased mRNA steady-state levels accompanied the rise in eIF4E and eIF4G protein levels, the overall abundance of PABP mRNA, together with the half-life of the polypeptide it encodes, remained relatively unchanged by HCMV infection. Instead, HCMV-induced PABP accumulation resulted from new protein synthesis and was sensitive to the mTORC1-selective inhibitor rapamycin, which interferes with phosphorylation of the mTORC1 substrate p70 S6K and the translational repressor 4E-BP1. While virus-induced PABP accumulation did not require p70 S6K, it was inhibited by the expression of a dominant-acting 4E-BP1 variant unable to be inactivated by mTORC1. Finally, unlike the situation in alpha- or gammaherpesvirus-infected cells, where PABP is redistributed to nuclei, PABP accumulated in the cytoplasm of HCMV-infected cells. Thus, cytoplasmic PABP accumulation is translationally controlled in HCMV-infected cells via a mechanism requiring mTORC1-mediated inhibition of the cellular 4E-BP1 translational repressor.
Successful Vaccination Strategies That Protect Aged Mice from Lethal Challenge from Influenza Virus and Heterologous Severe Acute Respiratory Syndrome Coronavirus
Authors: Sheahan Timothy , Whitmore Alan , Long Kristin , Ferris Martin , Rockx Barry , Funkhouser William , Donaldson Eric , Gralinski Lisa , Collier Martha , Heise Mark , Davis Nancy , Johnston Robert , Baric Ralph S.
Published: October 2010

Newly emerging viruses often circulate as a heterogeneous swarm in wild animal reservoirs prior to their emergence in humans, and their antigenic identities are often unknown until an outbreak situation. The newly emerging severe acute respiratory syndrome coronavirus (SARS-CoV) and reemerging influenza virus cause disproportionate disease in the aged, who are also notoriously difficult to successfully vaccinate, likely due to immunosenescence. To protect against future emerging strains, vaccine platforms should induce broad cross-reactive immunity that is sufficient to protect from homologous and heterologous challenge in all ages. From initial studies, we hypothesized that attenuated Venezuelan equine encephalitis virus (VEE) replicon particle (VRP) vaccine glycoproteins mediated vaccine failure in the aged. We then compared the efficacies of vaccines bearing attenuated (VRP
3014
) or wild-type VEE glycoproteins (VRP
3000
) in young and aged mice within novel models of severe SARS-CoV pathogenesis. Aged animals receiving VRP
3000
-based vaccines were protected from SARS-CoV disease, while animals receiving the VRP
3014
-based vaccines were not. The superior protection for the aged observed with VRP
3000
-based vaccines was confirmed in a lethal influenza virus challenge model. While the VRP
3000
vaccine's immune responses in the aged were sufficient to protect against lethal homologous and heterologous challenge, our data suggest that innate defects within the VRP
3014
platform mediate vaccine failure. Exploration into the mechanism(s) of successful vaccination in the immunosenescent should aid in the development of successful vaccine strategies for other viral diseases disproportionately affecting the elderly, like West Nile virus, influenza virus, norovirus, or other emerging viruses of the future.
Persistent Friend Virus Replication and Disease inApobec3-Deficient Mice Expressing Functional B-Cell-Activating Factor Receptor
Authors: Santiago Mario L. , Smith Diana S. , Barrett Bradley S. , Montano Mauricio , Benitez Robert L. , Pelanda Roberta , Hasenkrug Kim J. , Greene Warner C.
Published: October 2010

Rfv3is an autosomal dominant gene that influences the recovery of resistant mice from Friend retrovirus (FV) infection by limiting viremia and promoting a more potent neutralizing antibody response. We previously reported thatRfv3is encoded byApobec3, an innate retrovirus restriction factor. However, it was recently suggested that theRfv3 susceptible phenotype of high viremia at 28 days postinfection (dpi) was more dominantly controlled by the B-cell-activating factor receptor (BAFF-R), a gene that is linked to but located outside the genetically mapped region containingRfv3. Although one prototypicalRfv3susceptible mouse strain, A/WySn, indeed contains a dysfunctional BAFF-R, two otherRfv3susceptible strains, BALB/c and A.BY, express functional BAFF-R genes, determined on the basis of genotyping and B-cell immunophenotyping. Furthermore, transcomplementation studies in (C57BL/6 [B6] × BALB/c)F1and (B6 × A.BY)F1mice revealed that the B6Apobec3gene significantly influences recovery from FV viremia, cellular infection, and disease at 28 dpi. Finally, theRfv3phenotypes of prototypic B6, A.BY, A/WySn, and BALB/c mouse strains correlate with reportedApobec3mRNA expression levels. Overall, these findings argue against the generality ofBAFF-Rpolymorphisms as a dominant mechanism to explain theRfv3recovery phenotype and further strengthen the evidence thatApobec3encodesRfv3.
B7-Mediated Costimulation of CD4 T Cells Constrains Cytomegalovirus Persistence
Authors: Arens Ramon , Loewendorf Andrea , Her Min J. , Schneider-Ohrum Kirsten , Shellam Geoffrey R. , Janssen Edith , Ware Carl F. , Schoenberger Stephen P. , Benedict Chris A.
Published: October 2010

Cytomegalovirus (CMV) utilizes multiple strategies to modulate immunity and promote lifelong, persistent/latent infection, including suppressing T cell activation pathways. Here we examined the role of B7 costimulatory ligands in establishing immune détente from both the host and virus perspectives. Mice lacking both B7.1 and B7.2 showed reduced early expansion of CMV-specific CD4 T cells, consequently allowing for enhanced levels of persistent virus replication. In turn, a CMV mutant lacking expression of the m138 and m147.5 gene products, which restrict B7.1 and B7.2 expression in infected antigen-presenting cells, induced a more robust CD4 T cell response and showed decreased persistence. Together, these data reveal a requirement for B7-mediated signaling in regulating the CMV-specific CD4 T cell response and establishing host-virus equilibrium.
The B Cell Response Is Redundant and Highly Focused on V1V2 during Early Subtype C Infection in a Zambian Seroconverter
Authors: Lynch Rebecca M. , Rong Rong , Boliar Saikat , Sethi Anurag , Li Bing , Mulenga Joseph , Allen Susan , Robinson James E. , Gnanakaran S. , Derdeyn Cynthia A.
Published: October 2010

High-titer autologous neutralizing antibody responses have been demonstrated during early subtype C human immunodeficiency virus type 1 (HIV-1) infection. However, characterization of this response against autologous virus at the monoclonal antibody (MAb) level has only recently begun to be elucidated. Here we describe five monoclonal antibodies derived from a subtype C-infected seroconverter and their neutralizing activities against pseudoviruses that carry envelope glycoproteins from 48 days (0 month), 2 months, and 8 months after the estimated time of infection. Sequence analysis indicated that the MAbs arose from three distinct B cell clones, and their pattern of neutralization compared to that in patient plasma suggested that they circulated between 2 and 8 months after infection. Neutralization by MAbs representative of each B cell clone was mapped to two residues: position 134 in V1 and position 189 in V2. Mutational analysis revealed cooperative effects between glycans and residues at these two positions, arguing that they contribute to a single epitope. Analysis of the cognate gp120 sequence through homology modeling places this potential epitope near the interface between the V1 and V2 loops. Additionally, the escape mutation R189S in V2, which conferred resistance against all three MAbs, had no detrimental effect on virus replication
in vitro
. Taken together, our data demonstrate that independent B cells repeatedly targeted a single structure in V1V2 during early infection. Despite this assault, a single amino acid change was sufficient to confer complete escape with minimal impact on replication fitness.
A controlled evaluation of the WHO Safe Communities model approach to injury prevention: increasing child restraint use in motor vehicles
Authors: Istre Gregory R , Stowe Martha , McCoy Mary A , Moore Billy J , Culica Dan , Womack Katie N , Anderson Ron J
Published: October 2010

Objective

To measure the effect of the WHO Safe Communities model approach to increasing child restraint use in motor vehicles.

Design

Pre- and post-intervention observations of restraint use in motor vehicles in several sites in the target area, and in a comparison area community.

Setting

Community; southeast Dallas, Texas, 2003–2005.

Interventions

A multifaceted approach to increasing use of child safety seats, booster seats and seat belts that included efforts in schools, day care centres, neighbourhoods and a local public clinic, along with child safety seat classes and a low-cost distribution programme.

Main outcome measures

Prevalence of restraint use among children 0–8 years old riding in motor vehicles.

Results

In the target area, the adjusted child restraint use increased by 23.9 percentage points versus 11.8 in the comparison area (difference 12.1; 95% CI 9.9 to 14.3), and adjusted driver seat belt use increased by 16.3 percentage points in the target area versus 4.9 in the comparison area (difference 11.4; 95% CI 11.0 to 11.7). Multivariable multilevel analysis showed that the increase in the target area was significantly greater than in the comparison area for child restraint use (OR 1.6; 95% CI 1.2 to 2.2), as well as for driver seat belt use and proportion of children riding in the back seat.

Conclusions

The Safe Communities approach was successful in promoting the use of child restraints in motor vehicles through a multifaceted intervention that included efforts in various community settings, instructional classes and child safety seat distribution.
The expression of EMP1 is downregulated in oral squamous cell carcinoma and possibly associated with tumour metastasis
Authors: Zhang Jun , Cao Wei , Xu Qin , Chen Wan-tao
Published: October 2010

Aims

To investigate the expression of EMP1 in oral squamous cell carcinoma (OSCC) tissues and its correlation with available clinical parameters of patients with OSCC.

Methods

The mRNA levels of EMP1 were measured in 18 paired OSCC and corresponding adjacent normal tissues using RT-PCR. Another 45 pairs of OSCC samples were selected to detect the mRNA level of EMP1 using quantitative RT-PCR (qRT-PCR). The protein levels of EMP1 were also evaluated in 60 cases of patients with OSCC using immunohistochemical staining. The correlation between EMP1 expression and clinical parameters was analysed with non-parametric analysis.

Results

The results of RT-PCR and qRT-PCR showed that, compared with the paired normal tissues, the mRNA levels of EMP1 were significantly decreased in OSCC. The immunohistochemical results indicated that the EMP1 protein was also downregulated in OSCC (p=0.031). Decreased expression of EMP1 was significantly correlated with clinical stage (p=0.002) and lymph node metastasis (p=0.044) of patients with OSCC. Meanwhile, there was a significant difference between OSCCs at early and advanced stages (p=0.003), and between OSCCs with lymph node metastasis and no lymph node metastasis (p=0.045), respectively.

Conclusions

The results suggest that EMP1 may be a tumour suppressor and associated with lymph node metastasis in OSCC.
How reproductive and regenerative medicine meet in a Chinese fertility clinic. Interviews with women about the donation of embryos to stem cell research
Authors: Mitzkat Anika , Haimes Erica , Rehmann-Sutter Christoph
Published: October 2010

The social interface between reproductive medicine and embryonic stem cell research has been investigated in a pilot study at a large IVF clinic in central China. Methods included observation, interviews with hospital personnel, and five in-depth qualitative interviews with women who underwent IVF and who were asked for their consent to the donation of embryos for use in medical (in fact human embryonic stem cell) research. This paper reports, and discusses from an ethical perspective, the results of an analysis of these interviews. The participants talked of extreme social pressure to become pregnant. Once they had a baby, ‘spare’ embryos lost practical significance due to the Chinese one-child policy. In the context of decision making about donating embryos to research, the women used the clinical distinctions between ‘good and bad quality’ embryos and also between frozen and transferred embryos, as guiding moral distinctions. In the absence of concrete information about what sort of research their embryos should be used for, the women interviewed either refused consent (for fear that the embryo would be given to another couple) or accepted, expressing motives of solidarity with other women in a similar situation. This reveals that they filled the knowledge gap with an image of research improving fertility treatment.
OSCILLATIONS AND PHASE TRANSITION IN THE MEAN INFECTION RATE OF A FINITE POPULATION
Authors: WU QINGCHU , FU XINCHU , ZHANG HAIFENG , SMALL MICHAEL
Published: October 2010

We consider epidemic transmission in a finite population with both scale-free network structure and heterogeneous infection rates related to both susceptibility and infectiousness of individuals. The restriction to a finite population size and heterogeneity are included so that the model more closely matches reality. In particular, the arithmetic average behavior of heterogeneous infection rates with node-degree correlation is discussed. Through numerical simulations, we find that the average infection rate presents robust small-amplitude oscillations. Based on this property, we propose a statistical measure of the system dynamics — the epidemic spreading efficiency (ESE) — and discuss its phase transitions as a function of the various system parameters. The results show that the ESE presents a non-continuous phase transition and the ESE threshold is an extension of the epidemic threshold from the case of homogeneous infection rate (for homogeneous populations, the two quantities exhibit the same behavior). By comparing the ESE threshold and the epidemic threshold, we find that ESE threshold is larger than epidemic threshold for a sufficiently large network size. This implies that the traditional homogeneous assumption of infection rates in many epidemiological models overestimates the likelihood of epidemic disease survival.
SIMULATION OF NEWTONIAN AND NON-NEWTONIAN AXISYMMETRIC FLOW WITH AN AXISYMMETRIC LATTICE BOLTZMANN MODEL
Authors: LI X. F. , TANG G. H. , GAO T. Y. , TAO W. Q.
Published: October 2010

Axisymmetric flow is of both fundamental interest and practical significance. A recently derived axisymmetric lattice Boltzmann model [J. G. Zhou, Phys. Rev. E78, 036701 (2008)] is adopted for studying several typical axisymmetric flows. First, the Hagen–Poiseuille flow in circular pipes is validated and the Poiseuille flow in annular cylinders is studied under different values of the radius ratio. Second, pulsatile flow in an axisymmetric pipe with a sinusoidal pressure gradient is conducted. Third, flows through pipes with various constrictions or expansions are discussed. Finally, we extend the axisymmetric lattice Boltzmann method for non-Newtonian flow. It is found that the obtained numerical results agree well with available analytical solutions. It is also observed that constriction or expansion in a pipe influences the velocity distribution of the flow significantly. In addition, the results demonstrate that the modified axisymmetric lattice Boltzmann model is capable of handling non-Newtonian flow.
INVERSE STATISTICS AND ASSET ALLOCATION EFFICIENCY
Authors: BOLGORIAN MEYSAM
Published: October 2010

In this paper using inverse statistics analysis, the effect of investment horizon on the efficiency of portfolio selection is examined. Inverse statistics analysis is a general tool also known as probability distribution of exit time that is used for detecting the distribution of the time in which a stochastic process exits from a zone. This analysis was used in Refs. 1 and 2 for studying the financial returns time series. This distribution provides an optimal investment horizon which determines the most likely horizon for gaining a specific return. Using samples of stocks from Tehran Stock Exchange (TSE) as an emerging market and S&P 500 as a developed market, effect of optimal investment horizon in asset allocation is assessed. It is found that taking into account the optimal investment horizon in TSE leads to more efficiency for large size portfolios while for stocks selected from S&P 500, regardless of portfolio size, this strategy does not only not produce more efficient portfolios, but also longer investment horizons provides more efficiency.