- Volume 68, Issue 9, 1987
Volume 68, Issue 9, 1987
- Bacterial
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Identification of a Temperature-resistant Bacteriophage φX174 Mutant
More LessSummaryBacteriophage ϕX174am3trD a high temperature-resistant mutant of ϕX174am3, was 104 times more stable than 0 ϕX174am3 as judged by its survival ratio after heat treatment at 54 °C for 120 min. Complementation tests showed an involvement of gene G. Sequence analysis of this gene revealed three mutation sites, one transition and two insertions. The first was a silent mutation and the others brought about a change in one amino acid and an addition of another, respectively, in the gene G protein. These changes in amino acid sequence resulted in a change in the secondary structure of the protein. A β-turn region in part of the gene G protein of ϕX174am3 was changed to an α-helix in ϕX174am3trD These results indicate that the temperature resistance of ϕX174am3trD may be caused by elevated hydrophobicity in the mutated region or by strong interaction between the mutated gene G protein and other capsid proteins.
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- Animal
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Identification of a Synthetic Peptide as Part of a Major Neutralization Epitope of Respiratory Syncytial Virus
More LessSummaryA 7000 M r cleavage fragment of the F1 subunit that carries the major neutralization epitope has been identified by chemical and enzymatic cleavage of the fusion protein of respiratory syncytial (RS) virus (Long strain) with an efficient RS virus-neutralizing monoclonal antibody. Based on the published mRNA-deduced sequence of the A2 strain, coupled to the hydropathicity profile and prediction of protein conformation, the neutralization epitope has tentatively been localized on the first third of the F1 protein N-terminal, probably in the region of amino acids 215Ser to 236Glu. Analysis of three peptides covering different portions of the 212Cys to 236Glu region of the FI fusion protein identified a peptide (Cys-216Asn to 236Glu) that reacted strongly with the neutralizing monoclonal antibody and that was efficient in blocking neutralization and in plaque-reducing assays, confirming that the neutralization epitope was localized in that region. Further analysis with two other synthetic peptides (212Cys to 222Glu and Cys-221Ile to 236Glu) indicated that the dodecapeptide Ile-Glu-Phe-Gln-Lys- Asn-Asn-Arg-Leu-Leu-Glu mimicked either the whole or a major part of the neutralization epitope. This opens a promising avenue for the simple design of a synthetic peptide vaccine to control RS virus infection.
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Preparation and Properties of Immune-stimulating Complexes Containing Hepatitis B Virus Surface Antigen
More LessSummaryImmune-stimulating complexes (iscoms) have been prepared containing the major S gene products (HBsAg) of the hepatitis B virus genome. Immunization of B ALB/c mice with a single dose of hepatitis B iscoms in saline resulted in a high titre antibody response to HBsAg. In contrast, the original HBsAg preparation required an adjuvant to produce equivalent amounts of antibody. Analysis of sera from mice immunized with hepatitis B iscoms revealed antibodies directed against the major a determinants of HBsAg. High secondary antibody responses were observed in immunized animals previously inoculated with a sub-immunogenic dose of HBsAg indicating that hepatitis B iscoms may represent a suitable immunogen for use in individuals in whom a course of immunization with currently licensed hepatitis B vaccines has failed to produce a significant anti-HBs response.
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Expression of the Infectious Bronchitis Virus Spike Protein by Recombinant Vaccinia Virus and Induction of Neutralizing Antibodies in Vaccinated Mice
SummaryA cDNA clone of the infectious bronchitis virus (IBV) spike protein gene has been recombined into vaccinia virus. Cells infected with the recombinant virus synthesized IBV spike antigen which was recognized by antibody raised against purified spike protein. Immunofluorescence showed that the IBV spike antigen was transported to the infected cell surface membrane and immunoprecipitation showed the presence of the glycosylated 180K mol. wt. polypeptide precursor of the two spike subunits S1 and S2 that comigrated with this antigen from IBV-infected cells. Vaccinated mice produced antibody that recognized the IBV spike antigen by ELISA and which neutralized IBV infectivity as shown by ciliostasis tests on tracheal organ cultures.
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Effect of in vitro Mutations in a Vaccinia Virus Early Promoter Region Monitored by Herpes Simplex Virus Thymidine Kinase Expression in Recombinant Vaccinia Virus
More LessSummaryThe location of a promoter (PF) in the HindIII F region of the vaccinia virus genome was mapped by introducing deletions into this region of the DNA. Modified promoters were fused to the herpes simplex virus (HSV) thymidine kinase (TK) gene in plasmids facilitating the construction of recombinant vaccinia viruses, and promoter function was monitored by the ability of such plasmids to rescue TK+ vaccinia viruses from cells infected with TK− virus. Deletions from the 3′ end of the promoter region produced mutants for which function was either not inhibited or abolished, allowing the 3′ promoter boundary to be defined to within 13 nucleotides. As indicated by the presence of the PF transcript in early RNA and the kinetics of HSV TK expression in recombinant vaccinia viruses, transcription from PF occurred primarily at early times during infection. The major transcript was initiated at a site within 20 nucleotides of the 3′ end of the promoter and nine bases upstream of the probable translation initiation codon. In one mutant for which a small but reproducible increase in promoter function was detected, the transcription start site was deleted. Nevertheless, transcription still appeared to begin at the equivalent position with respect to the promoter, despite the altered nucleotide sequence. The location of the start site for the PF transcript indicated that the HSV TK gene, inserted at the BamHI site following the promoter, was preceded by an initiation codon which could potentially attenuate expression of the inserted gene. Conversion of this ATG codon to TAG did not significantly improve HSV TK expression.
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Avirulent Rotavirus Infections Protect Calves from Disease with and without Inducing High Levels of Neutralizing Antibody
More LessSummaryTwo bovine rotaviruses, C3-160 and 17/4, which multiplied in calves without inducing disease, were studied for their ability to protect against disease caused by a virulent rotavirus, CP-1. Rotavirus 17/4 and the virulent virus CP-1 cross-neutralized poorly and, on the basis of 20-fold differences in neutralization titres, belonged to different serotypes. Rotaviruses C3-160 and CP-1 were more closely related: neutralization of the CP-1 virus by C3-160 antisera was within 20-fold of the homologous titre although neutralization of the C3-160 virus by CP-1 antisera was not. Nine gnotobiotic calves infected with either C3-160 or 17/4, had rotavirus antibody in their faeces and/or serum 21 days after oral inoculation as detected by indirect immunofluorescence and IgG, IgM and/or IgA antibodies by ELISA. As expected from the antigenic relationships between the viruses, the sera and faeces from the four calves infected with C3-160 contained moderate levels of neutralizing antibody to the virulent virus CP-1 and the sera and most of the faeces from the five calves infected with 17/4 contained undetectable or low levels. When challenged with CP-1 on day 21, four age-matched controls developed disease whereas all of four calves previously infected with C3-160 and four of five calves previously infected with 17/4 were protected from disease. It was concluded that avirulent rotavirus infection provided protection against disease caused by a virulent rotavirus even when one of the avirulent viruses was poorly related to the virulent virus by neutralization. Mechanisms other than neutralizing antibody appeared to be important in protection.
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Mixed Infection of Culicoides variipennis with Bluetongue Virus Serotypes 10 and 17: Evidence for High Frequency Reassortment in the Vector
More LessSummaryThe primary vector species for bluetongue virus (BTV) in the United States, Culicoides variipennis, was orally infected with BTV serotype 10, BTV serotype 17, or a mixture of the two viruses. The recovery of virus from the infected flies was low following a period of extrinsic incubation. Electrophoretic analysis of progeny virus from singly infected flies revealed that only the parental electropherotype could be isolated from those flies. In contrast, electrophoretic analysis of virus from mixedly infected flies revealed that eight of the 11 virus-positive flies produced virus progeny with reassortant electropherotypes. The proportion of reassortant progeny varied from 7 to 78% (mean 42%), depending on the individual fly. Analysis of segregation of the parental origin of genome segments in the reassortant progeny virus suggested that, while reassortment of most segments was random, selection for genome segment 8 from the type 17 parent may have occurred. Analysis of segregation in individual mixedly infected flies showed that each fly yielded a relatively unique set of reassortants, but that specific electropherotypes were isolated repeatedly from individual flies. These data indicated that the vector species C. variipennis was a permissive host for high frequency reassortment of genome segments of BTV.
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In vivo Reassortment of Thogoto Virus (a Tick-borne Influenza-like Virus) following Oral Infection of Rhipicephalus appendiculatus Ticks
More LessSummaryArboviruses with segmented genomes have the potential to reassort in both their vertebrate hosts and arthropod vectors. Reassortment of Thogoto virus, a tick-borne orthomyxo-like virus, has been demonstrated following dual infection of hamsters by temperature-sensitive mutants. To investigate whether similar events can occur in ticks, Rhipicephalus appendiculatus larvae and nymphs were dually infected by interrupted feeding on viraemic hamsters. Wild-type reassortant virus was isolated from the ticks 12 to 15 days after engorgement. Following moulting, nymphs and adults transmitted reassortant virus to uninfected hamsters. This is the first reported evidence that a tick-borne arbovirus can reassort in vivo in a naturally infected arthropod vector. The relative roles of vector and vertebrate host in generating and perpetuating reassortant viruses in nature are discussed.
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The Ultrastructure of Cell Cultures Infected with Border Disease and Bovine Virus Diarrhoea Viruses
More LessSummaryThe morphology of border disease virus and bovine virus diarrhoea virus in infected bovine embryonic testis cells was examined by electron microscopy. Particles which appeared to be mature virions of both viruses were similar, being roughly circular and approximately 46 nm in diameter with a 20 to 25 nm core. Virus replication took place totally within the cytoplasm in association with structures formed from modified endoplasmic reticulum.
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Nucleotide Sequence of the Genes Coding for the Membrane Glycoproteins E1 and E2 of Rubella Virus
More LessSummaryThe complete nucleotide sequence of the genes coding for the two membrane glycoproteins E1 and E2 of rubella virus has been determined from cloned cDNA derived from the 40S genomic RNA. A sequence of 2451 nucleotides extending from the poly (A) tract towards the 5′ end is presented. Within one continuous open reading frame E2 is located amino-terminally followed by E1 and a 58 residue long untranslated 3′ region preceding the poly(A) tract. The coding regions of E2 and E1 are unusually G/C rich, 71·4% and 66·4% respectively. At the carboxy-terminal end of the coding region of E1, there is an inverted complementary nucleotide sequence, which could form a 13 base pair hairpin structure. Mature E2 and E1 are both preceded by a stretch of uncharged mainly non-polar amino acids, 21 and 20 residues in length, respectively. These could serve as signal peptides that mediate the membrane translocation of the proteins. At the carboxy termini of both proteins there are stretches of hydrophobic amino acids, 19 and 27 residues in length, which probably represent the transmembrane anchors of the proteins. The size of mature E1 is 481 amino acids (mol. wt. 51502), whereas the exact size of mature E2 could not be established as its carboxy- terminal end could not be located in the sequence. A maximum size of 262 amino acids (mol. wt. 28277) is, however, suggested. Between the E2 and E1 genes, there is a stretch of seven amino acids, five of which are arginines, which may serve as cleavage sites for a trypsin-like protease. E1 contains three and E2 four potential sites for asparagine- linked glycosylation. Both proteins are cysteine-rich (5 %). Comparison of the rubella virus amino acid sequence to those of several alphaviruses indicated no sequence homology.
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Characterization of Membrane Components of the Erythrocyte Involved in Vesicular Stomatitis Virus Attachment and Fusion at Acidic pH
More LessSummaryGoose erythrocyte membranes were isolated and tested for their ability to compete with red cell receptors for vesicular stomatitis virus (VSV) attachment and fusion at acidic pH. Crude membranes, solubilized with Triton X-100, Tween 80 and octyl-β-d- glucopyranoside, showed a dose-dependent inhibitory effect on virus binding and haemolysis. The chemical nature of the active molecules was investigated by enzyme digestion and by separation of purified components. Only the lipid moiety, specifically phospholipid and glycolipid, was found to inhibit VSV attachment; a more detailed analysis of these molecules showed that phosphatidylinositol, phosphatidylserine and GM3 ganglioside were responsible for the inhibitory activity and could therefore represent VSV binding sites on goose erythrocyte membranes. Removal of negatively charged groups from these molecules by enzymic treatment significantly reduced their activity, suggesting that electrostatic interactions play an important role in the binding of VSV to the cell surface. Enzymic digestion of whole erythrocytes confirmed the involvement of membrane lipid molecules in the cell surface receptor for VSV.
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Immunoaffinity Purification of a 72K Early Antigen of Human Cytomegalovirus: Analysis of Humoral and Cell-mediated Immunity to the Purified Polypeptide
More LessSummaryAn early antigen of human cytomegalovirus (HCMV) was purified from infected cells as a 72K polypeptide by immunoaffinity chromatography using a monoclonal antibody. It was located in both nucleus and cytoplasm, and was non-glycosylated and undetectable on the surface of infected cells. Known seropositive subjects had antibody against the purified protein and it elicited proliferative T cell responses in 10 of 16 subjects. Five of 14 T cell lines established in response to the purified protein were predominantly CD8+ and of these two showed major histocompatibility complex- restricted cytotoxicity against HCMV-infected cells. This provides further evidence that antigens expressed at early times may be targets for the immune response during persistent HCMV infection.
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Delayed-type Hypersensitivity Responses to Murine Cytomegalovirus in Genetically Resistant and Susceptible Strains of Mice
More LessSummaryThe delayed-type hypersensitivity (DTH) response in mice infected with murine cytomegalovirus (MCMV) was measured by ear swelling following a challenge with heat-treated MCMV. DTH was dose-dependent and could be detected as early as 3 days post-infection with peak responses occurring between days 15 and 21 postinfection. The DTH response was found to be specific for MCMV since it could not be elicited by either herpes simplex virus type 1 or influenza A virus in MCMV-primed mice. The elicited DTH response was greater in mice primed with attenuated compared with virulent MCMV. The DTH response was shown to correlate positively with the genetically determined resistance status of mouse strains to this virus. Previous research has shown that resistance to lethal infection with MCMV is controlled by H-2-linked genes since mice having the k haplotype are more resistant than mice having the b or d haplotype of the H-2 complex. Also, non-H-2-linked genes in CBA, C3H, C57BL/10 and probably other strains confer resistance. Resistant strains (C3H [H-2k], CBA [H-2k]) developed greater DTH responses than those of susceptible strains (BALB/c [H-2d], C57BL/10 [H-2b]) inoculated with the same dose of virus. In addition, the genetically resistant mouse strains BIO. BR [H-2k] and BALB. K [H-2k] gave a significantly greater DTH response than that of the corresponding congenic strains C57BL/10 [H-2b], BALB/c [H-2d] and BALB. B [H-2b] which are genetically susceptible to the virus. Also, the DTH response of C57BL/10 [H-2b] was significantly higher than that of BALB. B [H-2b] which correlates with their relative genetic resistance to MCMV, indicating the importance of non-H-2-linked genes. Furthermore, in addition to the response of greater magnitude, resistant strains (CBA, C3H, B10.BR) produced DTH responses to MCMV by day 3 compared with day 5 post-infection for susceptible BALB/c mice. These findings indicate that the magnitude and the time of appearance of the DTH response correlates positively with the genetically determined resistance status, although the role of DTH responses in controlling MCMV infections remains to be determined.
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Biological Basis for Virulence of Three Strains of Herpes Simplex Virus Type 1
More LessSummaryHerpes simplex virus type 1 (HSV-1) strains F, HF and HFEM were studied with respect to pathogenicity in mice and growth characteristics in vivo and in vitro compared to the neurovirulent HSV-1 strains 17 syn+ and KOS. All three viruses demonstrated reduced virulence in mouse brains and were completely avirulent after footpad inoculation. They were shown to express high levels of thymidine kinase activity. Investigations concerning the virulence phenotype indicated that the defect(s) in strains F, HF and HFEM related to general replication deficiency in mouse cells. It was also shown that although the replication restriction observed for strains F and HF was specific for murine cells, strain HFEM did not replicate well in any cell type tested. Additional studies indicated that the avirulence phenotype which followed peripheral inoculation was related to different genotypes, since strain F complemented HF and HFEM and, as expected, the latter two agents did not complement each other. All three agents were found to complement the non-neuroinvasive strain KOS. Finally, the data also show that two herpesviruses which are highly restricted in murine cells (e.g. strains F and HF) can still interact in the animal and produce a lethal infection.
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Abnormal Forms of the Herpes Simplex Virus Immediate Early Polypeptide Vmw175 Induce the Cellular Stress Response
More LessSummaryInduction of the major stress response in chick embryo fibroblasts, which follows infection at 38·5 °C with the herpes simplex virus mutant isK, was investigated. Synthesis of cellular stress proteins occurred only when the mutant form of an immediate early polypeptide, Vmwl75, was overproduced. Infection with mutant inl411, which has an amber (TAG) termination signal inserted between codons 83 and 84 of the gene encoding Vmwl75 and therefore specifies a truncated portion of the polypeptide, failed to stimulate stress protein synthesis. The results suggested that the presence of abnormal forms of Vmwl75 at high concentrations was the signal for induction of the stress response in tsK-infected cells.
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Identification of Sequences in Epstein-Barr Virus DNA Required for the Expression of the Second Epstein-Barr Virus-determined Nuclear Antigen in COS-1 Cells
More LessSummaryThe BamHI W YH region of the Epstein-Barr virus (EB V) genome encodes a protein localized to the nucleus of the infected cell, the EBV-determined nuclear antigen EBNA2. We have constructed a series of recombinant vectors that carried the complete EBNA2 gene, or the gene modified so as to contain defined deletions involving presumed exons and regulatory elements of the gene. The recombinant vectors were transfected into COS-1 cells which permit the replication of simian virus 40 origin-containing plasmids to a high copy number, and the transient expression of EBNA2 was analysed. A recombinant plasmid that carried a BglII-NotI subfragment of the BamHI WYH region (nucleotides 44664 to 50628) contained all the information necessary for inducing the expression of a full length EBNA2 polypeptide. Moreover, EBV DNA sequences between nucleotides 45442 and 48337 could be deleted without interfering with the ability of the vectors to induce EBNA2. On the other hand the loss of the left one-third of the BglII-NotI fragment completely abolished the EBNA2- inducing capacity of the vector. A rightward promoter consensus sequence in the BamHI W part of the BglII-NotI fragment was functional in COS-1 cells expressing EBNA2 and essential for EBV-specific RNA synthesis. The results indicated that transcription of the EBNA2 gene was initiated in the BamHI W fragment, that the transcript was spliced and that all of EBNA2 was encoded within the continuous long open reading frame in the BamHI Y and H fragments.
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Humoral and Cellular Immunity to Varicella-Zoster Virus Glycoprotein gpI and to a Non-glycosylated Protein, p170, in the Strain 2 Guinea-pig
More LessSummaryStrain 2 guinea-pigs were inoculated with infectious varicella-zoster virus (VZV) or with immunoaffinity-purified proteins of VZV. Monoclonal antibodies to the VZV gpI (90000/58000 complex) and to a non-glycosylated protein, pl70, were used to prepare the polypeptide antigens. Humoral and cell-mediated immune responses to the infectious virus were compared with those elicited by the gpI and pl70 proteins. Both VZV IgG antibody production and T lymphocyte proliferation to VZV were detected after immunization with infectious VZV and with VZV proteins. The antibody and T lymphocyte responses waned after protein immunization in comparison with the responses induced by infectious VZV but were detected again immediately after reimmunization with gpI or p170.
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Genetic and Environmental Determination of Herpes Simplex Virus Type 1 Penetration into Non-permissive Rat XC Cells
More LessSummaryWe have tested a number of herpes simplex virus type 1 (HSV-1) populations for their ability to enter and express virus polypeptides in non-permissive rat XC cells. The viruses tested included 40 intratypic F(MP) recombinants and different batches of virus, belonging to the same strains, that had been produced in two different permissive systems (HEp-2 or Vero cells). Our results indicated that the ability to infect XC cells was determined in part by genetic elements of the virus genome and in part by phenotypic characteristics conferred by the permissive cell that had produced the virus: a virus strain like HSV-1 MP which, when produced in HEp-2 cells, was able to infect XC cells, lost this ability when produced in Vero cells. Working only with viruses produced in HEp-2 cells we showed that the ability to enter XC cells could be transferred from the MP strain to the F strain (which does not normally infect XC cells efficiently) by transfer of the cloned BamHI B (map units 0·745 to 0·81) or BamHI L (map units 0·706 to 0·745) fragments isolated from MP DNA. The implicated locus or loci seemed to segregate, however, from loci controlling gC synthesis and cell fusion, which have been described as mapping in the same region.
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The Influence of Androgens on Paralysis in Mice Following Intravenous Inoculation of Herpes Simplex Virus
More LessSummaryIn mice, intravenous inoculation of relatively avirulent strains of herpes simplex virus [e.g. P2C6, a mutant of strain CL(101), deficient in thymidine kinase] produced infection in the adrenal gland and mid-spinal cord followed by hind limb paralysis without death. Male mice were less susceptible to paralysis than female mice. Castration of male mice before inoculation increased their susceptibility to that of female animals; treatment with testosterone reversed this change. The differences in susceptibility to paralysis in the various categories of animal were not reflected in differences in growth of virus in the adrenal gland or spinal cord.
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Localization of an Arenavirus Protein in the Nuclei of Infected Cells
More LessSummaryHost cell nuclear involvement in an arenavirus infection was examined by immunofluorescence. Both polyclonal antisera and monoclonal antibodies specific for the major nucleocapsid (N) polypeptide revealed virus-specific nuclear inclusions in Pichinde virus-infected Vero cells. Immunoprecipitation of infected cell extracts with the anti-N monoclonal antibodies and subsequent analysis by SDS-PAGE, identified two N-related proteins with mol. wt. of 36000 (p36) and 28000 (p28) in addition to the N polypeptide. Only those monoclonal antibodies which precipitated p28 as well as N and p36 were found to produce nuclear as well as cytoplasmic fluorescence. These findings suggest that either the p28 protein itself or a conformational variant of N was the nuclear antigen detected.
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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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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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Volumes and issues
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Volume 105 (2024)
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Volume 104 (2023)
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Volume 103 (2022)
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Volume 102 (2021)
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Volume 101 (2020)
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Volume 100 (2019)
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Volume 99 (2018)
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Volume 98 (2017)
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Volume 97 (2016)
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Volume 96 (2015)
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Volume 95 (2014)
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Volume 94 (2013)
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Volume 93 (2012)
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Volume 92 (2011)
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Volume 91 (2010)
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Volume 90 (2009)
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Volume 89 (2008)
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Volume 88 (2007)
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Volume 87 (2006)
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Volume 86 (2005)
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Volume 85 (2004)
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Volume 84 (2003)
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Volume 83 (2002)
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Volume 82 (2001)
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Volume 81 (2000)
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Volume 80 (1999)
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Volume 79 (1998)
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Volume 78 (1997)
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Volume 77 (1996)
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Volume 76 (1995)
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Volume 75 (1994)
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Volume 74 (1993)
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Volume 73 (1992)
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Volume 72 (1991)
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Volume 71 (1990)
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Volume 70 (1989)
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Volume 69 (1988)
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Volume 68 (1987)
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Volume 67 (1986)
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Volume 66 (1985)
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Volume 65 (1984)
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Volume 64 (1983)
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Volume 63 (1982)
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Volume 62 (1982)
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Volume 61 (1982)
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Volume 60 (1982)
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Volume 59 (1982)
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Volume 58 (1982)
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Volume 57 (1981)
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Volume 56 (1981)
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Volume 55 (1981)
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Volume 54 (1981)
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Volume 53 (1981)
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Volume 52 (1981)
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Volume 51 (1980)
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Volume 50 (1980)
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Volume 49 (1980)
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Volume 48 (1980)
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Volume 47 (1980)
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Volume 46 (1980)
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Volume 45 (1979)
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Volume 44 (1979)
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Volume 43 (1979)
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Volume 42 (1979)
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Volume 41 (1978)
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Volume 40 (1978)
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Volume 39 (1978)
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Volume 38 (1978)
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Volume 37 (1977)
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Volume 36 (1977)
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Volume 35 (1977)
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Volume 34 (1977)
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Volume 33 (1976)
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Volume 32 (1976)
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Volume 31 (1976)
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Volume 30 (1976)
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Volume 29 (1975)
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Volume 28 (1975)
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Volume 27 (1975)
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Volume 26 (1975)
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Volume 25 (1974)
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Volume 24 (1974)
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Volume 23 (1974)
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Volume 22 (1974)
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Volume 21 (1973)
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Volume 20 (1973)
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Volume 19 (1973)
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Volume 18 (1973)
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Volume 17 (1972)
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Volume 16 (1972)
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Volume 15 (1972)
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Volume 14 (1972)
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Volume 13 (1971)
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Volume 12 (1971)
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Volume 11 (1971)
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Volume 10 (1971)
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Volume 9 (1970)
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Volume 8 (1970)
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Volume 7 (1970)
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Volume 6 (1970)
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Volume 5 (1969)
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Volume 4 (1969)
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Volume 3 (1968)
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Volume 2 (1968)
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Volume 1 (1967)