bluuebunny/crossref_metadata_2025_split huggingface dataset:
Use of Highly Discriminatory Fingerprinting to Analyze Clusters of
Clostridium difficile
Infection Cases Due to Epidemic Ribotype 027 Strains
Authors: Fawley W. N. , Freeman J. , Smith C. , Harmanus C. , van den Berg R. J. , Kuijper E. J. , Wilcox M. H. 
Published: January 2008

ABSTRACT

We compared multilocus variable-number tandem-repeat analysis (MLVA) and macrorestriction endonuclease analysis using pulsed-field gel electrophoresis (PFGE) to determine their utility to identify clusters of
Clostridium difficile
infection (CDI) among 91 isolates of PCR ribotype 027 (NAP1, for
N
orth
A
merican
p
ulsed-field type
1
) from nine hospitals (and 10 general practitioners associated with one institution) in England. We also examined whether mortality in CDI cases was associated with specific MLVA subtypes. PFGE discriminated between ribotype 027 strains at >98% similarity, identifying five pulsovars (I to V) of 1 to 53 isolates. MLVA was markedly more discriminatory, identifying 23 types of 1 to 15 isolates (>71% similarity). PFGE pulsovars I and IV contained 14 and 8 MLVA types, respectively. Twenty-one of twenty-three (91%) of MLVA types were specific to individual PFGE pulsovars. Four CDI clusters were identified in institution A by conventional epidemiological analysis. MLVA typing identified two enlarged and two additional clusters. Thirty of forty-four (68%) patients in institution A with CDI caused by ribotype 027 strains were assigned to seven distinct clusters by a combination of MLVA typing and epidemiological records. Of 33 patients, comprising 14 different MLVA types, nine (27%) died by day 30 (early deaths). Eight of nine (89%) were associated with PFGE type IV
C. difficile
ribotype 027. Five of nine early deaths were associated with MLVA type 16, which was the dominant type in this cohort (10/33 cases); 4 other distinct MLVA types accounted for the other early deaths. MLVA was far superior to PFGE for analyzing clusters of CDI both within and between institutions. Further study is needed to examine whether subtypes of
C. difficile
ribotype 027 affect outcome.
Selection and Validation of a Multilocus Variable-Number Tandem-Repeat Analysis Panel for Typing
Shigella
spp
Authors: Gorgé Olivier , Lopez Stéphanie , Hilaire Valérie , Lisanti Olivier , Ramisse Vincent , Vergnaud Gilles 
Published: January 2008

ABSTRACT

The
Shigella
genus has historically been separated into four species, based on biochemical assays. The classification within each species relies on serotyping. Recently, genome sequencing and DNA assays, in particular the multilocus sequence typing (MLST) approach, greatly improved the current knowledge of the origin and phylogenetic evolution of
Shigella
spp. The
Shigella
and
Escherichia
genera are now considered to belong to a unique genomospecies. Multilocus variable-number tandem-repeat (VNTR) analysis (MLVA) provides valuable polymorphic markers for genotyping and performing phylogenetic analyses of highly homogeneous bacterial pathogens. Here, we assess the capability of MLVA for
Shigella
typing. Thirty-two potentially polymorphic VNTRs were selected by analyzing in silico five
Shigella
genomic sequences and subsequently evaluated. Eventually, a panel of 15 VNTRs was selected (i.e., MLVA15 analysis). MLVA15 analysis of 78 strains or genome sequences of
Shigella
spp. and 11 strains or genome sequences of
Escherichia coli
distinguished 83 genotypes.
Shigella
population cluster analysis gave consistent results compared to MLST. MLVA15 analysis showed capabilities for
E. coli
typing, providing classification among pathogenic and nonpathogenic
E. coli
strains included in the study. The resulting data can be queried on our genotyping webpage (
http://mlva.u-psud.fr
). The MLVA15 assay is rapid, highly discriminatory, and reproducible for
Shigella
and
Escherichia
strains, suggesting that it could significantly contribute to epidemiological trace-back analysis of
Shigella
infections and pathogenic
Escherichia
outbreaks. Typing was performed on strains obtained mostly from collections. Further studies should include strains of much more diverse origins, including all pathogenic
E. coli
types.
Amino Acid Mutation N348I in the Connection Subdomain of Human Immunodeficiency Virus Type 1 Reverse Transcriptase Confers Multiclass Resistance to Nucleoside and Nonnucleoside Reverse Transcriptase Inhibitors
Authors: Hachiya Atsuko , Kodama Eiichi N. , Sarafianos Stefan G. , Schuckmann Matthew M. , Sakagami Yasuko , Matsuoka Masao , Takiguchi Masafumi , Gatanaga Hiroyuki , Oka Shinichi 
Published: January 2008

ABSTRACT

We identified clinical isolates with phenotypic resistance to nevirapine (NVP) in the absence of known nonnucleoside reverse transcriptase inhibitor (NNRTI) mutations. This resistance is caused by N348I, a mutation at the connection subdomain of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT). Virologic analysis showed that N348I conferred multiclass resistance to NNRTIs (NVP and delavirdine) and to nucleoside reverse transcriptase inhibitors (zidovudine [AZT] and didanosine [ddI]). N348I impaired HIV-1 replication in a cell-type-dependent manner. Acquisition of N348I was frequently observed in AZT- and/or ddI-containing therapy (12.5%;
n
= 48;
P
< 0.0001) and was accompanied with thymidine analogue-associated mutations, e.g., T215Y (
n
= 5/6) and the lamivudine resistance mutation M184V (
n
= 1/6) in a Japanese cohort. Molecular modeling analysis shows that residue 348 is proximal to the NNRTI-binding pocket and to a flexible hinge region at the base of the p66 thumb that may be affected by the N348I mutation. Our results further highlight the role of connection subdomain residues in drug resistance.
Cell Cycle Association of the Retinoblastoma Protein Rb and the Histone Demethylase LSD1 with the Epstein-Barr Virus Latency Promoter Cp
Authors: Chau Charles M. , Deng Zhong , Kang Hyojueng , Lieberman Paul M. 
Published: January 2008

ABSTRACT

The Epstein-Barr virus C promoter (Cp) regulates the major multicistronic transcript encoding the EBNA-LP, 1, 2, and 3 genes required for B-cell proliferation during latency. The growth-transforming potential of these viral genes suggests that they must be tightly regulated with the host cell cycle and differentiation process. To better understand Cp regulation, we used DNA affinity purification to identify cellular and viral proteins that bind to Cp in latently infected cells. Several previously unknown factors were identified, including the cell cycle regulatory proteins E2F1 and Rb. E2F1 bound to a specific site in Cp located in the core Cp region 3′ of the known EBNA2-responsive RBP-Jk (CSL, CBF1) binding site. The histone H3 K4 demethylase LSD1 (BCC110) was also identified by DNA affinity and was shown to form a stable complex with Rb. Coimmunoprecipitation assays demonstrated that E2F1, Rb, and LSD1 bind to Cp in a cell cycle-dependent manner. Rb and LSD1 binding to Cp increased after the S phase, corresponding to a decrease in histone H3 K4 methylation and Cp transcription. Coimmunoprecipitation and immunofluorescence assays reveal that LSD1 interacts with Rb. Surprisingly, LSD1 did not coimmunoprecipitate with E2F1, suggesting that it associates with Rb independently of E2F1. Depletion of LSD1 by small interfering RNAs inhibited Cp basal transcription levels, and overexpression of LSD1 altered the cell cycle profile in p53-positive (p53
+
), but not p53-negative (p53
−
), HCT cells. These findings indicate that Cp is a cell cycle-regulated promoter that is under the control of Rb and the histone demethylase LSD1 in multiple latency types.
The Human Cytomegalovirus Fc Receptor gp68 Binds the Fc CH2-CH3 Interface of Immunoglobulin G
Authors: Sprague Elizabeth R. , Reinhard Henrike , Cheung Evelyn J. , Farley Alexander H. , Trujillo Robin Deis , Hengel Hartmut , Bjorkman Pamela J. 
Published: January 2008

ABSTRACTRecognition of immunoglobulin G (IgG) by surface receptors for the Fc domain of immunoglobulin G (Fcγ), FcγRs, can trigger both humoral and cellular immune responses. Two human cytomegalovirus (HCMV)-encoded type I transmembrane receptors with Fcγ-binding properties (vFcγRs), gp34 and gp68, have been identified on the surface of HCMV-infected cells and are assumed to confer protection against IgG-mediated immunity. Here we show that Fcγ recognition by both vFcγRs occurs independently of N-linked glycosylation of Fcγ, in contrast with the properties of host FcγRs. To gain further insight into the interaction with Fcγ, truncation mutants of the vFcγR gp68 ectodomain were probed for Fcγ binding, resulting in localization of the Fcγ binding site on gp68 to residues 71 to 289, a region including an immunoglobulin-like domain. Gel filtration and biosensor binding experiments revealed that, unlike host FcγRs but similar to the herpes simplex virus type 1 (HSV-1) Fc receptor gE-gI, gp68 binds to the CH2-CH3 interdomain interface of the Fcγ dimer with a nanomolar affinity and a 2:1 stoichiometry. Unlike gE-gI, which binds Fcγ at the slightly basic pH of the extracellular milieu but not at the acidic pH of endosomes, the gp68/Fcγ complex is stable at pH values from 5.6 to pH 8.1. These data indicate that the mechanistic details of Fc binding by HCMV gp68 differ from those of host FcγRs and from that of HSV-1 gE-gI, suggesting distinct functional and recognition properties.
Single-Cell, Phosphoepitope-Specific Analysis Demonstrates Cell Type- and Pathway-Specific Dysregulation of Jak/STAT and MAPK Signaling Associated with In Vivo Human Immunodeficiency Virus Type 1 Infection
Authors: Lee Andrew W. , Sharp Elizabeth R. , O'Mahony Alison , Rosenberg Michael G. , Israelski Dennis M. , Nolan Garry P. , Nixon Douglas F. 
Published: January 2008

ABSTRACTDespite extensive evidence of cell signaling alterations induced by human immunodeficiency virus type 1 (HIV-1) in vitro, the relevance of these changes to the clinical and/or immunologic status of HIV-1-infected individuals is often unclear. As such, mapping the details of cell type-specific degradation of immune function as a consequence of changes to signaling network responses has not been readily accessible. We used a flow cytometric-based assay of signaling to determine Janus kinase/signal transducers and activators of transcription (Jak/STAT) signaling changes at the single-cell level within distinct cell subsets from the primary immune cells of HIV-1-infected donors. We identified a specific defect in granulocyte-macrophage colony-stimulating factor (GM-CSF)-driven Stat5 phosphorylation in the monocytes of HIV-1+ donors. This inhibition was statistically significant in a cohort of treated and untreated individuals. Ex vivo Stat5 phosphorylation levels varied among HIV-1+ donors but did not correlate with CD4+T-cell counts or HIV-1 plasma viral load. Low Stat5 activation occurred in HIV-1-infected donors despite normal GM-CSF receptor levels. Investigation of mitogen-activated protein kinase (MAPK) pathways, also stimulated by GM-CSF, led to the observation that lipopolysaccharide-stimulated extracellular signal-regulated kinase phosphorylation is enhanced in monocytes. Thus, we have identified a specific, imbalanced monocyte signaling profile, with inhibition of STAT and enhancement of MAPK signaling, associated with HIV-1 infection. This understanding of altered monocyte signaling responses that contribute to defective antigen presentation during HIV-1 infection could lead to immunotherapeutic approaches that compensate for the deficiency.
Proteolytic Cleavage of VP1-2 Is Required for Release of Herpes Simplex Virus 1 DNA into the Nucleus
Authors: Jovasevic Vladimir , Liang Li , Roizman Bernard 
Published: January 2008

ABSTRACT

In this report we propose a model in which after the herpes simplex virus (HSV) capsid docks at the nuclear pore, the tegument protein attached to the capsid must be cleaved by a serine or a cysteine protease in order for the DNA to be released into the nucleus. In support of the model are the following results. (i) Exposure of cells at the time of or before infection to
l
-(tosylamido-2-phenyl) ethyl chloromethyl ketone (TPCK), a serine-cysteine protease inhibitor, prevents the release of viral DNA or expression of viral genes. TPCK does not block viral gene expression after entry of viral DNA into the nucleus. (ii) The tegument protein VP1-2, the product of the U
L
36 gene, is cleaved shortly after the entry of the HSV 1 (HSV-1) virion into the cell. (iii) The proteolytic cleavage of VP1-2 does not occur in cells that are infected with HSV-1 under conditions that prevent the release of the viral DNA into the nucleus. (iv) The proteolytic cleavage of VP1-2 occurs only after the capsid is attached to the nuclear pore. Thus, TPCK prevented the release of HSV-1 DNA into the nucleus when added to medium 1 hour after infection with
ts
B7 at 39.5°C followed by a shift down to the permissive temperature. The
ts
lesion maps in the U
L
36 gene. At the nonpermissive temperature, the capsids accumulate at the nuclear pore but the DNA is not released into the nucleus.
Identification of Hot Spots in the Variola Virus Complement Inhibitor (SPICE) for Human Complement Regulation
Authors: Yadav Viveka Nand , Pyaram Kalyani , Mullick Jayati , Sahu Arvind 
Published: January 2008

ABSTRACTVariola virus, the causative agent of smallpox, encodes a soluble complement regulator named SPICE. Previously, SPICE has been shown to be much more potent in inactivating human complement than the vaccinia virus complement control protein (VCP), although they differ only in 11 amino acid residues. In the present study, we have expressed SPICE, VCP, and mutants of VCP by substituting each or more of the 11 non-variant VCP residues with the corresponding residue of SPICE to identify hot spots that impart functional advantage to SPICE over VCP. Our data indicate that (i) SPICE is ∼90-fold more potent than VCP in inactivating human C3b, and the residues Y98, Y103, K108 and K120 are predominantly responsible for its enhanced activity; (ii) SPICE is 5.4-fold more potent in inactivating human C4b, and residues Y98, Y103, K108, K120 and L193 mainly dictate this increase; (iii) the classical pathway decay-accelerating activity of activity is only twofold higher than that of VCP, and the 11 mutations in SPICE do not significantly affect this activity; (iv) SPICE possesses significantly greater binding ability to human C3b compared to VCP, although its binding to human C4b is lower than that of VCP; (v) residue N144 is largely responsible for the increased binding of SPICE to human C3b; and (vi) the human specificity of SPICE is dictated primarily by residues Y98, Y103, K108, and K120 since these are enough to formulate VCP as potent as SPICE. Together, these results suggest that principally 4 of the 11 residues that differ between SPICE and VCP partake in its enhanced function against human complement.
Functional Analysis of the Transmembrane (TM) Domain of the
Autographa californica
Multicapsid Nucleopolyhedrovirus GP64 Protein: Substitution of Heterologous TM Domains
Authors: Li Zhaofei , Blissard Gary W. 
Published: January 2008

ABSTRACT

GP64, the major envelope glycoprotein of the
Autographa californica
multicapsid nucleopolyhedrovirus (AcMNPV) budded virion, is important for host cell receptor binding and mediates low-pH-triggered membrane fusion during entry by endocytosis. In the current study, we examined the functional role of the AcMNPV GP64 transmembrane (TM) domain by replacing the 23-amino-acid GP64 TM domain with corresponding TM domain sequences from a range of viral and cellular type I membrane proteins, including
Orgyia pseudotsugata
MNPV (OpMNPV) GP64 and F, thogotovirus GP75,
Lymantria dispar
MNPV (LdMNPV) F, human immunodeficiency virus type 1 (HIV-1) GP41, human CD4 and glycophorin A (GpA), and influenza virus hemagglutinin (HA), and with a glycosylphosphatidylinositol (GPI) anchor addition sequence. In transient expression experiments with Sf9 cells, chimeric GP64 proteins containing either a GPI anchor or TM domains from LdMNPV F or HIV-1 GP41 failed to localize to the cell surface and thus appear to be incompatible with either GP64 structure or cell transport. All of the mutant constructs detected at the cell surface mediated hemifusion (outer leaflet merger) upon low-pH treatment, but only those containing TM domains from CD4, GpA, OpMNPV GP64, and thogotovirus GP75 mediated pore formation and complete membrane fusion activity. This supports a model in which partial fusion (hemifusion) proceeds by a mechanism that is independent of the TM domain and the TM domain participates in the enlargement or expansion of fusion pores after hemifusion. GP64 proteins containing heterologous TM domains mediated virion budding with dramatically differing levels of efficiency. In addition, chimeric GP64 proteins containing TM domains from CD4, GpA, HA, and OpMNPV F were incorporated into budded virions but were unable to rescue the infectivity of a
gp64
null virus, whereas those with TM domains from OpMNPV GP64 and thogotovirus GP75 rescued infectivity. These results show that in addition to its basic role in membrane anchoring, the GP64 TM domain is critically important for GP64 trafficking, membrane fusion, virion budding, and virus infectivity. These critical functions were replaced only by TM domains from related viral membrane proteins.
Regulation of Immune Response and Inflammatory Reactions against Viral Infection by VCAM-1
Authors: Ou Rong , Zhang Menghua , Huang Lei , Flavell Richard A. , Koni Pandelakis A. , Moskophidis Demetrius 
Published: January 2008

ABSTRACTThe migration of activated antigen-specific immune cells to the target tissues of virus replication is controlled by the expression of adhesion molecules on the vascular endothelium that bind to ligands on circulating lymphocytes. Here, we demonstrate that the adhesion pathway mediated by vascular cell adhesion molecule 1 (VCAM-1) plays a role in regulating T-cell-mediated inflammation and pathology in nonlymphoid tissues, including the central nervous system (CNS) during viral infection. The ablation of VCAM-1 expression from endothelial and hematopoietic cells using aloxP-Crerecombination strategy had no major effect on the induction or overall tissue distribution of antigen-specific T cells during a systemic infection with lymphocytic choriomeningitis virus (LCMV), except in the case of lung tissue. However, enhanced resistance to lethal LCM and the significantly reduced magnitude and duration of footpad swelling observed in VCAM-1 mutant mice compared to B6 controls suggest a significant role for VCAM-1 in promoting successful local inflammatory reactions associated with efficient viral clearance and even life-threatening immunopathology under particular infection conditions. Interestingly, analysis of the infiltrating populations in the brains of intracerebrally infected mice revealed that VCAM-1 deletion significantly delayed migration into the CNS of antigen-presenting cells (macrophages and dendritic cells), which are critical for optimal stimulation of migrating virus-specific CD8+T cells initiating a pathological cascade. We propose that the impaired migration of these accessory cells in the brain may explain the improved clinical outcome of infection in VCAM-1 mutant mice. Thus, these results underscore the potential role of VCAM-1 in regulating the immune response and inflammatory reactions against viral infections.
A Single-Amino-Acid Mutation in Hepatitis C Virus NS5A Disrupting FKBP8 Interaction Impairs Viral Replication
Authors: Okamoto Toru , Omori Hiroko , Kaname Yuuki , Abe Takayuki , Nishimura Yorihiro , Suzuki Tetsuro , Miyamura Tatsuo , Yoshimori Tamotsu , Moriishi Kohji , Matsuura Yoshiharu 
Published: January 2008

ABSTRACT

Hepatitis C virus (HCV) nonstructural protein 5A (NS5A) regulates viral replication through its interaction with host and other viral proteins. We have previously shown that FK506-binding protein 8 (FKBP8) binds to NS5A and recruits Hsp90 to form a complex that participates in the replication of HCV. In this study, we examined the biochemical characteristics of the interaction and the intracellular localization of NS5A and FKBP8. Surface plasmon resonance analysis revealed that the dissociation constant of the interaction between the purified FKBP8 and NS5A expressed in bacteria was 82 nM. Mutational analyses of NS5A revealed that a single amino acid residue of Val or Ile at position 121, which is well conserved among all genotypes of HCV, is critical for the specific interaction with FKBP8. Substitution of the Val
121
to Ala drastically impaired the replication of HCV replicon cells, and the drug-resistant replicon cells emerging after drug selection were shown to have reverted to the original arrangement by replacing Ala
121
with Val. Examination of individual fields of the replicon cells by both fluorescence microscopy and electron microscopy (the correlative fluorescence microscopy-electron microscopy technique) revealed that FKBP8 is partially colocalized with NS5A in the cytoplasmic structure known as the membranous web. These results suggest that specific interaction of NS5A with FKBP8 in the cytoplasmic compartment plays a crucial role in the replication of HCV.
Hepatitis C Virus Inhibits Cell Surface Expression of HLA-DR, Prevents Dendritic Cell Maturation, and Induces Interleukin-10 Production
Authors: Saito Kousuke , Ait-Goughoulte Malika , Truscott Steven M. , Meyer Keith , Blazevic Azra , Abate Getahun , Ray Ratna B. , Hoft Daniel F. , Ray Ranjit 
Published: January 2008

ABSTRACTHepatitis C virus (HCV) chronic infection is characterized by low-level or undetectable cellular immune responses against HCV antigens. HCV proteins have been shown to affect various intracellular events and modulate immune responses, although the precise mechanisms used to mediate these effects are not fully understood. In this study, we have examined the effect of HCV proteins on the modulation of major histocompatibility complex (MHC) class II expression and other functions important for antigen presentation in humans. Expression of an HCV1-2962genomic clone (HCV-FL) in human fibrosarcoma cells (HT1080) inhibited gamma interferon (IFN-γ)-induced upregulation of human leukocyte antigen-DR (HLA-DR) cell surface expression. Furthermore, inhibition of promoter activities of MHC class II transactivator (CIITA), IFN-γ-activated site (GAS), and HLA-DR was observed in IFN-γ-inducible HT1080 cells expressing HCV-FL by in vitro reporter assays. Exposure of human monocyte-derived dendritic cells (DCs) to cell culture-grown HCV (HCVcc) genotype 1a (clone H77) or 2a (clone JFH1) significantly inhibited DC maturation and was associated with the production of IL-10. Furthermore, DCs exposed to HCVcc were impaired in their functional ability to stimulate antigen-specific CD4-positive (CD4+) and CD8+T-cell responses. Taken together, our results indicated that HCV can have direct and/or indirect inhibitory effects on antigen-presenting cells, resulting in reduction of antigen-specific T-cell activation. These effects may account for or contribute to the low overall level of immunogenicity of HCV observed in chronically infected patients.