- Volume 68, Issue 9, 1987
Volume 68, Issue 9, 1987
- Animal
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Evidence for the Expression of Protein IX in Some Rat Cells Transformed with Adenovirus Type 12 Early Region 1 DNA
More LessSummarySera were produced by injection of a particular rat cell line (HLBRK EcoC1), produced by transformation of baby rat kidney cells with adenovirus 12 early region 1 DNA, into the syngeneic host. Antibodies to a protein of M r 15200 as determined by SDS-PAGE, were detected in the sera from two rats bearing tumours. The 15K protein was identified as the viral structural component polypeptide IX by a number of criteria: (i) the protein was only observed in adenovirus 12-infected cells, (ii) it was produced at intermediate and late times after infection, (iii) it was immunoprecipitated from purified adenovirus 12 particles and (iv) it was immunoprecipitated from a purified protein preparation.
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Simian Virus 40 Infection Is Not Mediated by Lysosomal Activation
More LessSummaryThe uptake of simian virus 40 (SV40) virions to the nucleus, the site of viral replication, proceeds via engulfment at the cytoplasmic membrane and transport in monopinocytotic vesicles through the cytoplasm to the nuclear membrane. In the case of Semliki Forest virus and poliovirus which undergo primary endocytosis in a similar manner, neutralization of the acid pH in these vesicles abolishes viral infectivity. We have examined the effects of the lysosomotropic agents chloroquine and ammonium chloride on the uptake of SV40 and find that neutralization of the acid pH in cellular organelles has no effect on the progress of SV40 infection. Although the initial endocytotic pathway appears similar for the viruses, the vesicular transport of SV40 to the nucleus proceeds, therefore, via an alternative endocytotic compartment which is not inhibited by increasing the endosomal pH.
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Quantitative Immunocytochemical Assay for Infectious Avian Retroviruses
More LessSummaryA simple and accurate immunocytochemical focus assay is described, whereby both transforming and non-transforming avian retroviruses can be enumerated. After virus infection of chick embryo fibroblasts, an agar overlay is applied; foci of infected cells (expression foci) are detected immunocytochemically after 5 to 7 days. The primary antibodies are monoclonal sera directed against either viral p 19gag or pp60v-src. Detection of expression foci after transfection of cells with cloned viral DNA is also demonstrated.
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Properties of a Hepatitis A Virus Candidate Vaccine Strain
More LessSummaryThis paper describes the biophysical and biochemical properties as well as electron microscopical studies of a candidate hepatitis A vaccine strain propagated in human fibroblast cells. Our results indicated that, in CsCl, the density of hepatitis A virus (HAV) from cell culture supernatant and of HAV extracted from infected cells was influenced by the quantity of lipid material associated with HAV. Antigenicity of untreated HAV, therefore, was detected primarily in low density CsCl fractions (1·11 g/ml, 1·21 g/ml). After lipid reduction with NP40 detergent or chloroform/Genetron, antigenicity and infectivity were primarily detected in high density CsCl fractions (1·31 g/ml). Electron microscopy demonstrated a strong association between membranous material and virus particles of low density in CsCl as well as virus-like particles in ultrathin sections of HAV-infected human fibroblast cells. The uncovered virus particles banded in the 1·31 g/ml dense CsCl fraction lacked lipid material. The s value was 79 for 1·19 g/ml to 1·22 g/ml dense HAV and 147 for 1·29 g/ml to 1·33 g/ml HAV. Autoradiography of the radioiodinated dense HAV revealed some proteins with high M r (120K to 67K) and others with low M r (37K to 15K).
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Enterovirus Replication in Porcine Ileal Explants
More LessSummaryOrgan explants of porcine ileum were cultured in different media for up to 48 h. Tissue preservation was evaluated by light microscopy and by transmission and scanning electron microscopy. Cellular structure was well maintained after incubation for 48 h in CMRL-1066 supplemented with insulin and cortisone. Explants of absorptive or lymphoid tissue from young or adult pigs were incubated with either coxsackievirus B5 (which is infectious for swine) or human poliovirus type 1 (which served as a control) for 24 h at 37 °C. Progeny virus was detected by plaque assay. Replication was most evident in the absorptive tissue explants from young pigs. In tissues from adults, replication occurred equally well in absorptive and lymphoid tissues. Infection in explants was inefficient, and the yield of progeny virus was low.
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Mumps Virus-induced Alterations in Cellular Excitability During Persistent Infections
More LessSummaryPersistent mumps virus infections were established in rat pheochromocytoma (PC- 12) and human medulloblastoma (TE-671) continuous cell lines. Significant amounts of infectious virus were produced by the PC-12 cells; infectious virus production by the TE-671 cells was limited. This restricted replication may be due to decreased production of viral envelope glycoproteins by TE-671 cells. The presence of virus changed the distribution of stimulus-evoked electrical responsiveness of both cell lines from responsiveness composed primarily of normal, rapidly rising, all-or-nothing action potentials to one dominated by abnormal, slowly rising, graded responses or by no response at all. Such changes have the potential to disrupt neural integration within the nervous system, and suggest a new mechanism by which persistent virus infections might play a role in chronic neurological and/or mental disease.
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Identification of Bluetongue Virus-specific Immunoglobulin E in Cattle
More LessSummaryA modified passive cutaneous anaphylaxis test and an ELISA were used to identify IgE in calves vaccinated (sensitized) with chlorine dioxide-inactivated bluetongue virus (BTV) and in calves inoculated with infectious BTV. The levels of IgE were greatest in the vaccinated calves after challenge with infectious virus, which correlated with development of clinically apparent dermatitis and stomatitis. These findings suggest that some aspects of clinical bluetongue disease in cattle may have an immunopathological mechanism mediated by IgE (type I hypersensitivity).
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Identification of an Additional Sendai Virus Non-structural Protein Encoded by the P/C mRNA
More LessSummaryPeptide antiserum and monoclonal antibodies have detected a previously unrecognized small non-structural protein in Sendai virus-infected cells. This protein, designated X, appears to represent the C-terminal 95 amino acids of the P protein reading frame. The X protein appears to be almost as abundant as the P protein on a molar basis in vivo. No evidence of a precursor-product relationship between the X and P proteins could be found.
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Demonstration that Glycoprotein G Is the Attachment Protein of Respiratory Syncytial Virus
More LessSummaryTwo monospecific rabbit antisera to the G glycoprotein, one induced with purified G and the second with a recombinant vaccinia virus containing the gene for G, inhibit the attachment of purified [35S]methionine-labelled Long strain of respiratory syncytial virus to monolayers of HeLa cells. Attachment was not inhibited by monospecific rabbit antisera to glycoprotein F induced with either purified F or with a recombinant vaccinia virus containing the gene for F.
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- Plant
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Production of cDNA Clones from the MAV Isolate of Barley Yellow Dwarf Virus
More LessSummaryA library of cDNA clones was produced from the approximately 6 kb RNA of the MAV isolate of barley yellow dwarf virus (BYDV) in bacteriophage λgtl 1, by a method that involved random priming and cloning ds cDNAs of between 1·0 and 2·5 kbp. Screening with antiserum to the dissociated coat protein of the MAV isolate showed that approximately 2·5% of the recombinants were capable of expressing this protein. After subcloning some inserts into the plasmid pUC18, restriction endonuclease mapping showed that they collectively represented at least 85% (a total of 5·1 kbp) of the BYDV genome. We did not attempt to determine which, if any, of the immunologically positive clones expressed the entire coat protein, but of the nine examined, all shared a region of approximately 1000 bp, located between 750 bp and 1750 bp from the 3′ terminus of the restriction map. Sequence homology among different isolates of BYDV was examined by using selected MAV cDNA clones as probes in viral nucleic acid hybridization studies. Hybridization specificity varied according to the origin of the clones within the BYDV genome. Those from the putative coat protein-coding region hybridized well only to the homologous MAV isolate; those from elsewhere hybridized also with another isolate from the same subgroup (P-PAV). No clones hybridized significantly to a third isolate (RPV), which is in another subgroup of BYDV. The sensitivity of detection was related to probe size; the larger clones detected as little as 70 µg of purified virus (1·4 ng/ml in a 50 µl sample), and with these the sensitivity of virus detection in plant extracts by dot-blot hybridization was greater than that of ELISA.
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Movement and Intracellular Location of Sonchus Yellow Net Virus within Infected Nicotiana edwardsonii
More LessSummarySystemic movement of Sonchus yellow net virus to leaves and roots was first detected by ELISA 24 h after mechanical inoculation. Thereafter, virus levels rose to a maximum 10 days after inoculation; the highest levels were between 4·0 and 7·3 µg/g tissue, in leaves which were not yet fully expanded. Electron microscopy of tissue sections revealed that when the virus content of tissues was greatest, virtually all leaf and root cells were infected. Most of the virions were in the perinuclear space; only a few were scattered in the cytoplasm. Nuclei contained large viroplasms associated with viral nucleocapsids. Between 10 and 20 days after inoculation, levels of virus antigen and viral RNA fell to about 20% of their maximum. By 20 days after inoculation, no more than 10% of cells contained virus particles and almost all the virions were in the cytoplasm. These results suggest that this virus spreads systemically until most or all cells are infected. The plants then undergo a recovery phase during which virus disappears from the nuclei of infected cells and vesiculates into the cytoplasm.
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Translational Diffusion Coefficient of Tobacco Mosaic Virus Particles
More LessSummaryQuasi-elastic light scattering studies were made of tobacco mosaic virus (TMV) in 10 mm- or 100 mm-Tris-HCl buffer pH 7·2, or in 100 mm-sodium phosphate buffer pH 7·2. The translational diffusion coefficients, extrapolated to zero TMV concentration, were 3-75 × 10−8 cm2/s in phosphate buffer and 4·50 × 10−8 cm2/s in Tris buffer; the latter value is in good accord with that calculated for monodispersed TMV. Sedimentation studies showed that one sedimenting component was formed in Tris buffer, but two broad bands were formed in phosphate buffer. Sedimentation coefficients were 192 for monodispersed TMV and 214 for the second component, showing that it was an aggregate. These results indicate that the lower translational diffusion coefficient obtained in phosphate buffer is caused by the presence of TMV aggregates, and that Tris buffer is better than phosphate buffer for the preparation of monodispersed TMV solutions.
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