bluuebunny/crossref_metadata_2025_split huggingface dataset:
Aetiology of Opuntia ficus‐indica malformations and stunting disease
Authors: Granata G. , Paltrinieri S. , Botti S. , Bertaccini A. 
Published: November 2006

AbstractThe phytoplasma aetiology of epidemic stunting of cladodes and stunted growth observed in a cactus pear plantation in Carlentini (Sicily, Italy) was investigated with graft inoculation trials and PCR/restriction fragment length polymorphism analyses of 16S ribosomal RNA (rRNA) gene and S3 ribosomal protein gene. After sequencing of the 1525‐bp from 16S rRNA gene of both naturally infected and graft‐inoculated symptomatic cactus pear samples, phytoplasma TS belonging to ribosomal subgroup 16SrII‐C were identified as aetiological agents of this worldwide spread disease.
Mutations in Penicillin-Binding Protein (PBP) Genes and in Non-PBP Genes during Selection of Penicillin-Resistant
Streptococcus gordonii
Authors: Haenni Marisa , Moreillon Philippe 
Published: November 2006

ABSTRACT

Penicillin resistance in
Streptococcus
spp. involves multiple mutations in both penicillin-binding proteins (PBPs) and non-PBP genes. Here, we studied the development of penicillin resistance in the oral commensal
Streptococcus gordonii
. Cyclic exposure of bacteria to twofold-increasing penicillin concentrations selected for a progressive 250- to 500-fold MIC increase (from 0.008 to between 2 and 4 μg/ml). The major MIC increase (≥35-fold) was related to non-PBP mutations, whereas PBP mutations accounted only for a 4- to 8-fold additional increase. PBP mutations occurred in class B PBPs 2X and 2B, which carry a transpeptidase domain, but not in class A PBP 1A, 1B, or 2A, which carry an additional transglycosylase domain. Therefore, we tested whether inactivation of class A PBPs affected resistance development in spite of the absence of mutations. Deletion of PBP 1A or 2A profoundly slowed down resistance development but only moderately affected resistance in already highly resistant mutants (MIC = 2 to 4 μg/ml). Thus, class A PBPs might facilitate early development of resistance by stabilizing penicillin-altered peptidoglycan via transglycosylation, whereas they might be less indispensable in highly resistant mutants which have reestablished a penicillin-insensitive cell wall-building machinery. The contribution of PBP and non-PBP mutations alone could be individualized in DNA transformation. Both PBP and non-PBP mutations conferred some level of intrinsic resistance, but combining the mutations synergized them to ensure high-level resistance (≥2 μg/ml). The results underline the complexity of penicillin resistance development and suggest that inhibition of transglycosylase might be an as yet underestimated way to interfere with early resistance development.
qnrA
in CTX-M-Producing
Escherichia coli
Isolates from France
Authors: Lavigne Jean-Philippe , Marchandin Hélène , Delmas Julien , Bouziges Nicole , Lecaillon Evelyne , Cavalie Laurent , Jean-Pierre Hélène , Bonnet Richard , Sotto Albert 
Published: November 2006

ABSTRACT

By PCR, we screened for
qnr
genes 112 clinical isolates of extended-spectrum β-lactamase-producing
Escherichia coli
collected from hospitals in France during 2004. For the first time, 7.7% of CTX-M-producing
E. coli
isolates presented a plasmid-mediated resistance to quinolones. All strains harbored a
qnrA
gene located on a
sul1
-type class 1 integron with similar structure to the In
36
integron.
A
trans
-Golgi Network Resident Protein, golgin-97, Accumulates in Viral Factories and Incorporates into Virions during Poxvirus Infection
Authors: Alzhanova Dina , Hruby Dennis E. 
Published: November 2006

ABSTRACT

Poxviruses are the only DNA viruses known to replicate and assemble in the cytoplasm of infected cells. Poxvirus morphogenesis is a complicated process in which four distinct infectious forms of the virus are produced: intracellular mature virus, intracellular enveloped virus, cell-associated enveloped virus, and extracellular enveloped virus. The source of primary membrane wrapping the intracellular mature virus, the first infectious form, is still unknown. Although the membrane was suggested to originate from the endoplasmic reticulum-Golgi intermediate compartment, none of the marker proteins from this or any other cell compartments has been found in the intracellular mature virus. Thus, it was hypothesized that the membrane is either extensively modified by the virus or synthesized de novo. In the work described here, we demonstrate that a host cell protein residing in the
trans
-Golgi network membrane, golgin-97, is transported to the sites of virus replication and assembly and becomes incorporated into the virions during poxvirus infection. Inside the virion, golgin-97 is associated with the insoluble core protein fraction. Being able to adopt a long rod-like structure, the protein apparently extends through the virion envelope and protrudes from its surface. Here we discuss the potential role and functions of golgin-97 in poxvirus replication and propose two working models.
Translational Inhibition and Increased Interferon Induction in Cells Infected with C Protein-Deficient Measles Virus
Authors: Nakatsu Yuichiro , Takeda Makoto , Ohno Shinji , Koga Ritsuko , Yanagi Yusuke 
Published: November 2006

ABSTRACT
In addition to the phosphoprotein, the P gene of measles virus (MV) also encodes the V and C proteins by an RNA editing process and by alternative initiation of translation in a different reading frame, respectively. Although the MV C protein is required for efficient MV replication in vivo and in some cultured cells, its exact functions in virus infection are currently unclear. Here, we report that a recombinant MV lacking the C protein (MVΔC) grew poorly in a human cell line possessing the intact interferon (IFN) pathway and that this growth defect was associated with reduced viral translation and genome replication. The translational inhibition was correlated with phosphorylation of the alpha subunit of eukaryotic translation initiation factor 2. Moreover, increased IFN induction was observed in MVΔC-infected cells. The NS1 protein of influenza virus, which binds to double-stranded RNA (dsRNA) and consequently inhibits IFN induction and dsRNA-dependent protein kinase activation, complemented the growth defect of MVΔC. These results indicate that the MV C protein inhibits IFN induction and modulates host antiviral responses, thereby ensuring MV growth in host cells.
Prevalence and Phylogeny of Kakugo Virus, a Novel Insect Picorna-Like Virus That Infects the Honeybee (
Apis mellifera
L.), under Various Colony Conditions
Authors: Fujiyuki Tomoko , Ohka Seii , Takeuchi Hideaki , Ono Masato , Nomoto Akio , Kubo Takeo 
Published: November 2006

ABSTRACT

We previously identified a novel insect picorna-like virus, termed Kakugo virus (KV), from the brains of aggressive worker honeybees that had counterattacked a giant hornet. To survey the prevalence of KV in worker populations engaged in various labors, we quantified KV genomic RNA. KV was detected specifically from aggressive workers in some colonies, while it was also detected from other worker populations in other colonies where the amount of KV detected in the workers was relatively high, suggesting that KV can infect various worker populations in the honeybee colonies. To investigate whether the KV strains detected were identical, phylogenetic analysis was performed. There was less than a 2% difference in the RNA-dependent RNA polymerase (RdRp) sequences between KV strains from aggressive workers and those from other worker populations, suggesting that all of the viruses detected were virtually the same KV. We also found that some of the KV-infected colonies were parasitized by
Varroa
mites, and the sequences of the KV strains detected from the mites were the same as those detected from the workers of the same colonies, suggesting that the mites mediate KV prevalence in the honeybee colonies. KV strains had approximately 6% and 15% sequence differences in the RdRp region from deformed wing virus and Varroa destructor virus 1, respectively, suggesting that KV represents a viral strain closely related to, but distinct from, these two viruses.
A Helper-Dependent Capsid-Modified Adenovirus Vector Expressing Adeno-Associated Virus Rep78 Mediates Site-Specific Integration of a 27-Kilobase Transgene Cassette
Authors: Wang Hongjie , Lieber André 
Published: November 2006

ABSTRACT

Random integration of viral gene therapy vectors and subsequent activation or disruption of cellular genes poses safety risks. Major efforts in the field are aimed toward targeting vector integration to specific sites in the host genome. The adeno-associated virus (AAV) Rep78 protein is able to target AAV integration to a specific site on human chromosome 19, called AAVS1. We studied whether this ability could be harnessed to achieve site-specific integration of a 27-kb transgene cassette into a model cell line for human hematopoietic cells (Mo7e). To deliver
rep78
and the transgene to Mo7e cells, we used helper-dependent adenovirus (Ad) vectors containing Ad serotype 35 fiber knob domains (HD-Ad). An HD-Ad vector containing the
rep78
gene under the control of the globin locus control region (LCR) (Ad.LCR-rep78) conferred Rep78 expression on Mo7e cells. Upon coinfection of Ad.LCR-rep78 with an HD-Ad vector containing a 27-kb globin-LCR-green fluorescent protein (GFP) transgene cassette flanked by AAV inverted terminal repeats (ITRs) (Ad.AAV-LCR-GFP), transduced cells were cloned and expanded (without selection pressure), and vector integration was analyzed in clones with more than 30% GFP-positive cells. Vector integration into the AAVS1 region was seen in 30% of analyzed integration sites, and GFP expression from these integrants was stable over time. Of the remaining integration sites, 25% were within the genomic globin LCR. In almost 90% of sites, transgene integration occurred via the Ad ITR. This indicates that rescue of the AAV ITR-flanked transgene cassette from Ad.AAV-LCR-GFP is not required for Rep78-mediated integration into AAVS1 and that free ends within the vector genome can be created by breaks within the Ad ITRs, whose structure is apparently recognized by cellular “nicking” enzymes. The finding that 55% of all analyzed integration sites were either within the AAVS1 or globin LCR region demonstrates that a high frequency of targeted integration of a large transgene cassette can be achieved in human hematopoietic stem cell lines.
Mimivirus Giant Particles Incorporate a Large Fraction of Anonymous and Unique Gene Products
Authors: Renesto Patricia , Abergel Chantal , Decloquement Philippe , Moinier Danielle , Azza Saïd , Ogata Hiroyuki , Fourquet Patrick , Gorvel Jean-Pierre , Claverie Jean-Michel 
Published: November 2006

ABSTRACT

Acanthamoeba polyphaga mimivirus
is the largest known virus in both particle size and genome complexity. Its 1.2-Mb genome encodes 911 proteins, among which only 298 have predicted functions. The composition of purified isolated virions was analyzed by using a combined electrophoresis/mass spectrometry approach allowing the identification of 114 proteins. Besides the expected major structural components, the viral particle packages 12 proteins unambiguously associated with transcriptional machinery, 3 proteins associated with DNA repair, and 2 topoisomerases. Other main functional categories represented in the virion include oxidative pathways and protein modification. More than half of the identified virion-associated proteins correspond to anonymous genes of unknown function, including 45 “ORFans.” As demonstrated by both Western blotting and immunogold staining, some of these “ORFans,” which lack any convincing similarity in the sequence databases, are endowed with antigenic properties. Thus, anonymous and unique genes constituting the majority of the mimivirus gene complement encode bona fide proteins that are likely to participate in well-integrated processes.
Binding and Nuclear Relocalization of Protein Kinase R by Human Cytomegalovirus TRS1
Authors: Hakki Morgan , Marshall Emily E. , De Niro Katherine L. , Geballe Adam P. 
Published: November 2006

ABSTRACT
The human cytomegalovirus (HCMV) TRS1 and IRS1 genes block the phosphorylation of the alpha subunit of eukaryotic initiation factor 2 (eIF2α) and the consequent shutoff of cellular protein synthesis that occur during infection with vaccinia virus (VV) deleted of the double-stranded RNA binding protein gene E3L (VVΔE3L). To further define the underlying mechanism, we first evaluated the effect of pTRS1 on protein kinase R (PKR), the double-stranded RNA (dsRNA)-dependent eIF2α kinase. Immunoblot analyses revealed that pTRS1 expression in the context of a VVΔE3L recombinant decreased levels of PKR in the cytoplasm and increased its levels in the nucleus of infected cells, an effect not seen with wild-type VV or a VVΔE3L recombinant virus expressing E3L. This effect of pTRS1 was confirmed by visualizing the nuclear relocalization of PKR-EGFP expressed by transient transfection. PKR present in both the nuclear and cytoplasmic fractions was nonphosphorylated, indicating that it was unactivated when TRS1 was present. PKR also accumulated in the nucleus during HCMV infection as determined by indirect immunofluorescence and immunoblot analysis. Binding assays revealed that pTRS1 interacted with PKR in mammalian cells and in vitro. This interaction required the same carboxy-terminal region of pTRS1 that is necessary to rescue VVΔE3L replication in HeLa cells. The carboxy terminus of pIRS1 was also required for rescue of VVΔE3L and for mediating an interaction of pIRS1 with PKR. These results suggest that these HCMV genes directly interact with PKR and inhibit its activation by sequestering it in the nucleus, away from both its activator, cytoplasmic dsRNA, and its substrate, eIF2α.
Preferential Translation of Vesicular Stomatitis Virus mRNAs Is Conferred by Transcription from the Viral Genome
Authors: Whitlow Zackary W. , Connor John H. , Lyles Douglas S. 
Published: November 2006

ABSTRACT

Host protein synthesis is inhibited in cells infected with vesicular stomatitis virus (VSV). It has been proposed that viral mRNAs are subjected to the same inhibition but are predominantly translated because of their abundance. To compare translation efficiencies of viral and host mRNAs during infection, we used an enhanced green fluorescent protein (EGFP) reporter expressed from a recombinant virus or from the host nucleus in stably transfected cells. Translation efficiency of host-derived EGFP mRNA was reduced more than threefold at eight hours postinfection, while viral-derived mRNA was translated around sevenfold more efficiently than host-derived EGFP mRNA in VSV-infected cells. To test whether mRNAs transcribed in the cytoplasm are resistant to shutoff of translation during VSV infection, HeLa cells were infected with a recombinant simian virus 5 (rSV5) that expressed GFP. Cells were then superinfected with VSV or mock superinfected. GFP mRNA transcribed by rSV5 was not resistant to translation inhibition during superinfection with VSV, indicating that transcription in the cytoplasm is not sufficient for preventing translation inhibition. To determine if
cis
-acting sequences in untranslated regions (UTRs) were involved in preferential translation of VSV mRNAs, we constructed EGFP reporters with VSV or control UTRs and measured the translation efficiency in mock-infected and VSV-infected cells. The presence of VSV UTRs did not affect mRNA translation efficiency in mock- or VSV-infected cells, indicating that VSV mRNAs do not contain
cis
-acting sequences that influence translation. However, we found that when EGFP mRNAs transcribed by VSV or by the host were translated in vitro, VSV-derived EGFP mRNA was translated 22 times more efficiently than host-derived EGFP mRNA. This indicated that VSV mRNAs do contain
cis
-acting structural elements (that are not sequence based), which enhance translation efficiency of viral mRNAs.
A Full-Length cDNA Infectious Clone of North American Type 1 Porcine Reproductive and Respiratory Syndrome Virus: Expression of Green Fluorescent Protein in the Nsp2 Region
Authors: Fang Ying , Rowland Raymond R. R. , Roof Michael , Lunney Joan K. , Christopher-Hennings Jane , Nelson Eric A. 
Published: November 2006

ABSTRACT
The recent emergence of a unique group of North American type 1 porcine reproductive and respiratory syndrome virus (PRRSV) in the United States presents new disease control problems for a swine industry that has already been impacted seriously by North American type 2 PRRSV. In this study, a full-length cDNA infectious clone was generated from a low-virulence North American type 1 PRRSV isolate, SD01-08. In vitro studies demonstrated that the cloned virus maintained growth properties similar to those of the parental virus. Virological, pathological, and immunological observations from animals challenged with cloned viruses were similar to those from animals challenged with the parental virus and a modified live virus vaccine. To further explore the potential use as a viral backbone for expressing foreign genes, the green fluorescent protein (GFP) was inserted into a unique deletion site located at amino acid positions 348 and 349 of the predicted Nsp2 region in the virus, and expression of the Nsp2-GFP fusion protein was visualized by fluorescent microscopy. The availability of this North American type 1 infectious clone provides an important research tool for further study of the basic viral biology and pathogenic mechanisms of this group of type 1 PRRSV in the United States.
Hemagglutinin (HA) Proteins from H1 and H3 Serotypes of Influenza A Viruses Require Different Antigen Designs for the Induction of Optimal Protective Antibody Responses as Studied by Codon-Optimized HA DNA Vaccines
Authors: Wang Shixia , Taaffe Jessica , Parker Christopher , Solórzano Alicia , Cao Hong , García-Sastre Adolfo , Lu Shan 
Published: November 2006

ABSTRACT
Effective antibody responses provide crucial immunity against influenza virus infection. The hemagglutinin (HA) protein is the major target of protective antibody responses induced by viral infection and by vaccination with both inactivated and live-attenuated flu vaccines, but knowledge about the optimal designs of protective HA antigens from different flu serotypes is still limited. In this study, we have significantly improved the immunogenicity of HA-expressing DNA vaccines by using codon-optimized HA sequences for either an H1 serotype (A/NewCal/20/99) or an H3 serotype (A/Panama/2007/99) human influenza A virus and then used these constructs as model antigens to identify the optimal HA antigen designs to elicit high-level protective antibody responses. Two forms of HA antigen, a wild-type, full-length HA and a secreted form with transmembrane (TM) domain-truncated HA, were produced. Both forms of HA DNA vaccines, from either H1 or H3 serotypes, were able to elicit high levels of HA-specific immunoglobulin G responses in immunized rabbits as measured by enzyme-linked immunosorbent assay. Interestingly, the abilities of H1 HA and H3 HA antigens to elicit hemagglutination inhibition (HI) and neutralizing antibody (NAb) responses differ. For the H1 HA antigens, the full-length HA induced significantly higher HI and NAb responses than did the TM-truncated HA. For the H3 HA antigen, both the full-length HA and TM-truncated HA induced high levels of HI and NAb responses. These data indicate that H1 and H3 antigens have different expression requirements for the induction of an optimal protective antibody response and that the structure integrity of HA antigens is critical for eliciting type-specific protective antibody responses. Our findings will have an important impact on future subunit-based flu vaccine development.
Crystal Structure of the West Nile Virus Envelope Glycoprotein
Authors: Nybakken Grant E. , Nelson Christopher A. , Chen Beverly R. , Diamond Michael S. , Fremont Daved H. 
Published: November 2006

ABSTRACT
The envelope glycoprotein (E) of West Nile virus (WNV) undergoes a conformational rearrangement triggered by low pH that results in a class II fusion event required for viral entry. Herein we present the 3.0-Å crystal structure of the ectodomain of WNV E, which reveals insights into the flavivirus life cycle. We found that WNV E adopts a three-domain architecture that is shared by the E proteins from dengue and tick-borne encephalitis viruses and forms a rod-shaped configuration similar to that observed in immature flavivirus particles. Interestingly, the single N-linked glycosylation site on WNV E is displaced by a novel α-helix, which could potentially alter lectin-mediated attachment. The localization of histidines within the hinge regions of E implicates these residues in pH-induced conformational transitions. Most strikingly, the WNV E ectodomain crystallized as a monomer, in contrast to other flavivirus E proteins, which have crystallized as antiparallel dimers. WNV E assembles in a crystalline lattice of perpendicular molecules, with the fusion loop of one E protein buried in a hydrophobic pocket at the DI-DIII interface of another. Dimeric E proteins pack their fusion loops into analogous pockets at the dimer interface. We speculate that E proteins could pivot around the fusion loop-pocket junction, allowing virion conformational transitions while minimizing fusion loop exposure.
Double-Stranded RNA Binding of Influenza B Virus Nonstructural NS1 Protein Inhibits Protein Kinase R but Is Not Essential To Antagonize Production of Alpha/Beta Interferon
Authors: Dauber Bianca , Schneider Jana , Wolff Thorsten 
Published: November 2006

ABSTRACT
Expression of alpha/beta interferon (IFN-α/β) in virus-infected vertebrate cells is a key event in the establishment of a sustained antiviral response, which is triggered by double-stranded RNA (dsRNA) produced during viral replication. These antiviral cytokines initiate the expression of cellular proteins with activities that limit the replication and spread of the invading viruses. Within this response, the dsRNA-dependent protein kinase R (PKR) that is expressed at constitutive levels and upregulated by IFN-α/β acts as an important antiviral effector that can block the cellular translational machinery. We previously demonstrated that efficient replication of influenza B virus depends on the viral dsRNA-binding NS1 protein that inhibits the transcriptional activation of IFN-α/β genes. Here we tested the postulate that the viral NS1 protein counteracts antiviral responses through sequestering intracellular dsRNA by analyzing a collection of recombinant influenza B viruses. As expected, viruses expressing dsRNA-binding-defective NS1 proteins were strongly attenuated for replication in IFN-competent hosts. Interestingly, these virus mutants failed to prevent activation of PKR but could effectively limit IFN induction. Conversely, a mutant virus expressing the N-terminal dsRNA-binding domain of NS1 prevented PKR activation, but not IFN induction, suggesting an important role for the NS1 C-terminal part in silencing the activation route of IFN-α/β genes. Thus, our findings indicate an unexpected mechanistic dichotomy of the influenza B virus NS1 protein in the suppression of antiviral responses, which involves at least one activity that is largely separable from dsRNA binding.