- Volume 35, Issue 1, 1977
Volume 35, Issue 1, 1977
- Announcement
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- Review Article
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- Articles
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Genetic Control of Resistance to Subgroup A and Subgroup C Tumour Viruses in Rhode Island Red Fowl: Evidence for Linkage between the Tumour Virus a (tva) and Tumour Virus c (tvc) Loci
More LessSUMMARYA study, using the Rhode Island Red (RIR) strain of fowl maintained at Houghton Poultry Research Station, was made to investigate the genetic control of cellular response to infection with viruses of subgroups A and C. Family matings within the RIR strain and test-crosses between the RIR parents and White Leghorn (WL) parents of known ararcrcr genotype were set up to ascertain linkage between the tumour virus a (tva) and tumour virus c (tvc) loci.
The results confirmed that in this RIR strain, the two loci, tva and tvc, control the cellular response to viruses of subgroups A and C, respectively, as reported in other breeds of fowl (WL and New Hampshire). As in WL fowl, the two loci are linked. The linkage value of 0.22 in the male sex agreed well with that reported in the WL male sex, indicating that the two loci are located in the same sites in homologous chromosomes in the two breeds. However, in the RIR strain, no sex difference in crossing over between the two linked loci was found, contrary to that reported in WL fowl where the absence of crossing over between the two loci was observed in the heterogametic female sex.
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Production of RNA and Artificial Top Component from Parsley Carrot-leaf Virus Heated In vitro
More LessSummaryThe extinction-temperature profile of parsley carrot-leaf virus in 0.02 m-(Na-K) phosphate buffer, pH 7.2, containing 0.1 m-NaCl, was determined. At the Td (dissociation temperature = 65 °C), the point at which E 260 begins to increase, the virus particles apparently dissociate to form RNA and empty protein shells (top component). At the Tf (temperature at the inflexion point of the curve = 70 °C), corresponding to half the maximum increase in E 260, the protein denatures and precipitates. Temperatures below Td have little effect on the virus, whereas temperatures higher than Tf also degrade the RNA. The results of the present work seem to suggest that the increase in E 260 of PCLV preparations upon heating is mostly attributable to turbidity caused by coat protein denaturation rather than to effects on RNA.
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A Variant of Tobacco Rattle Virus: Evidence for a Second Gene in RNA-2
More LessSUMMARYThe isolation of YS, a spontaneous variant of the CAM strain of tobacco rattle virus, is described. Unlike CAM, YS produced yellow symptoms in several species of host plant. In experiments with mixtures of virus particles from the two strains, this character was shown to be controlled by RNA-2. However, the coat proteins of YS and CAM were indistinguishable by serology, by electrophoresis in polyacrylamide gels or by tryptic peptide mapping. It is concluded that the mutation responsible for the yellow symptoms is not in the coat protein gene, but elsewhere in RNA-2.
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Effect of Chaotropic Salts and Protein Denaturants on the Thermal Stability of Mouse Fibroblast Interferon
More LessSUMMARYAltering the aqueous environment, especially with agents that affect hydrogen bonds, markedly affects the stability of mouse L cell interferon. Low pH stabilizes interferon whereas high pH labilizes it; heavy water further enhances interferon thermostability at pH 2 but not at pH 9. Exposure to the protein denaturants, 4 m-guanidine hydrochloride and 6 m-urea, significantly decreases the activity of interferon at pH 2 and pH 9; however, the residual interferon activity is relatively thermostable. Certain chaotropic salts protect interferon against thermal destruction, and in terms of effectiveness, their sequence is in the order SCN− > I− ⩾ Cl− = ClO4 − = Br− > NO3 −. Interferon becomes more stable to heat as the NaSCN concentration is increased from 0.25 m to 2.0 m. Molecular sieve chromatography of interferon in the presence of 1.5 m-NaSCN at pH 7 shows a shift in its apparent mol. wt. from 25000 to 42000. Unlike most proteins, the unfolded conformation of interferon appears to be more stable to heat than the molecule with a smaller Stokes’ radius.
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Heterotypic Exclusion Between Vesicular Stomatitis Viruses of the New Jersey and Indiana Serotypes
More LessSUMMARYCo-infection of cells with vesicular stomatitis viruses of the Indiana and New Jersey serotypes results in interference. Using specifically-labelled immunofluorescent antibodies, it was demonstrated that within any one co-infected cell, one virus serotype replicated to the relative exclusion of the other serotype. This result was further substantiated by an examination of the virus serotypes released by infectious centres co-infected with both viruses.
Dominance of one serotype over the other was shown to be a function of the relative multiplicity of the two viruses. Superinfection by the second serotype at a higher multiplicity resulted in dominance by the second virus during the early period (up to 1.5 h) post-infection. After this time, the minority virus was able to overcome this dominance. Dominance of the majority virus was also abolished by u.v. inactivation.
Cell protein synthesis appeared to be less affected in cells infected with both serotypes than when infection was with a single serotype.
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Characterization of Human Papovavirus RFV: Comparison with SV40 and BKV
More LessSummaryHuman papovavirus, RFV, isolated from urine of a renal transplant patient was compared with two strains of SV40 and with the prototype human papovavirus, BKV. Neutralization tests showed that RFV and BKV are indistinguishable, while large-plaque (LP) and small-plaque (SP) isolates of SV40 gave a low but significant level of cross-reaction with rabbit or human antisera against RFV.
DNA reassociation saturation tests using 125I-labelled RFV DNA show that BKV has 88% homology, and SP-SV40 has 29% homology to RFV. We conclude that RFV and BKV are nearly, if not totally, identical and are not SV40 variants.
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An Examination of Permeation Chromatography on Columns of Controlled Pore Glass for Routine Purification of Plant Viruses
More LessSUMMARYPermeation chromatography on controlled pore glass (CPG) was compared with sucrose density gradient centrifugation for the purification of fifteen widely differing plant viruses. CPG chromatography gave excellent recovery of nearly all the viruses as evaluated by E 260, serological titre and infectivity assay and proved as good as or better than sucrose density gradient centrifugation. Both methods gave similar separations of virus from host material and acceptably pure virus preparations. CPG columns accepted virus in high or low concentration; non-specific adsorption was not observed using CPG pre-treated with poly(ethylene glycol) of mol. wt. 20000.
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The Role of Defective Interfering Particles in Persistent Infection of Vero Cells by Measles Virus
More LessSUMMARYPersistent infections by measles virus were rapidly established in the majority of Vero cells when monolayers were infected with virus stocks that had been passed three to five times from an undiluted inoculum. These virus stocks had low infectivity titres but normal haemagglutinin titres and were able to cause interference. The ability of such virus stocks to establish persistent infections seems to be due to the presence of defective interfering particles rather than of virus mutants. Measles virus released from a persistently infected Vero cell line at the 93rd passage had properties similar to the undiluted passage virus that generated persistent infections.
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Requirement of the Bacteriophage T7 0.7 Gene for Phage Growth in the Presence of the Col 1b Factor
More LessSUMMARYA variety of colicinogenic factors were examined for their effect on the growth of phages related to T7. Col B1 and Col B4 inhibited the growth of every phage tested. Col B2, however, did not interfere with T3 and H. Col E2 interfered only with growth of T3, while Col Ib produced abortive infection with some T7 mutants. The abortive infection associated with Col Ib seems to be correlated with the absence of the virus phosphokinase; mixed infection with T7 wild type and a phosphokinase deficient mutant was productive but the phage yield was depressed. The abortively infected cells lysed.
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Further Studies on the Biology of an Antiviral Factor (AVF) From Virus-Infected Plants and Its Association With the N-Gene of Nicotiana Species
Y. Antignus, I. Sela and I. HarpazSUMMARYThe antiviral factor (AVF) from virus-infected plants, purified on polyacrylamide gels, could be labelled with radioactive phosphorus and its activity could be eluted from the gels. The radioactivity and the antiviral activity were co-purified and thus co-electrophoresed; hence, the previously reported radioactive zone (Antignus, Sela & Harpaz, 1975) can be regarded as AVF. The production of AVF requires both the presence of the N-gene in the plant as well as virus infection. AVF production is inhibited by actinomycin D, but its activity is not affected by this drug. AVF production is correlated in time with the development of virus resistance in a local-lesion host. AVF inhibits TMV multiplication in infected leaves and suppresses virus synthesis almost totally in a local-lesion host. Some AVF is also produced when Nicotiana glutinosa L. is infected with a non-localized virus, but to a much lesser extent and at a later stage of infection. The production of AVF in N. glutinosa is not blocked at 30 °C, even though TMV is no longer localized at this temperature.
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Rifampicin-resistant Mutant supporting Bacteriophage Growth on Stationary Phase Achromobacter Cells
More LessSUMMARYA rifampicin-resistant Achromobacter mutant with an altered RNA polymerase was isolated. The mutant supports phage α3a growth in both log and stationary phase cells. Phage growth on stationary phase cells is sensitive to aeration and growth only occurs at oxygen concentrations of less than 5.2 p.p.m. The rifampicin-resistant mutant is similar to the spontaneous mutant strain 14 reported by Woods (1976) in that both mutants support stationary-phase phage growth under micro-aerophilic conditions. The isolation of the rifampicin-resistant mutant with an altered RNA polymerase suggests that the phenomenon of stationary phase phage growth could be due to a change in the template specificity of the Achromobacter RNA polymerase. Plaque morphology mutants which grow on log and/or stationary phase cells of the Achromobacter wild type, strain 14 and rifampicin-resistant strains are also described.
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Infection of Tobacco Mesophyll Protoplasts with Raspberry Ringspot Virus alone and together with Tobacco Rattle Virus
More LessSUMMARYRaspberry ringspot virus (RRV) infected tobacco mesophyll protoplasts best, judged by the yield of infective virus, when the inoculation mixture contained 0.04 to 5 µg/ml virus, 1.0 to 1.5 µg/ml poly-l-ornithine and phosphate buffer (0.006 to 0.025 m, pH 6.0 to 9.0); without poly-l-ornithine no infection occurred. The optimum temperature for accumulation of infective virus was 20 to 22 °C, and the virus content reached about 2 × 106 particles/protoplast in 3 days at 22 °C. On staining with fluorescent antibody to purified virus particles, infected protoplasts gave a faint generalized cytoplasmic fluorescence, with many also containing a more brightly fluorescing spot thought to correspond with the vesiculated inclusion body found by electron microscopy.
In contrast, many of the protoplasts inoculated with mixtures of RRV and the CAM strain of tobacco rattle virus (TRV-CAM) contained numerous discrete granules of RRV particle antigen throughout the cytoplasm. Infection with RRV had little effect on the distribution of TRV-CAM particle antigen or on the proportion of protoplasts infected with TRV-CAM. Up to 95% of protoplasts became infected with both viruses, which apparently infected independently. The aggregates of RRV antigen formed only in protoplasts also infected with TRV-CAM.
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The Interaction between Raspberry Ringspot and Tobacco Rattle Viruses in Doubly Infected Protoplasts
More LessSUMMARYTobacco mesophyll protoplasts infected with raspberry ringspot virus (RRV) show faint generalized staining when treated with fluorescent antibody to RRV, but many brightly staining granules develop after inoculation with a mixture of RRV and the CAM strain of tobacco rattle virus (TRV-CAM). The phenomenon occurred with three different strains of RRV but not when tobacco mosaic virus, or two other strains of tobacco rattle virus, were substituted for the CAM strain. The RRV antigen aggregates were produced only in protoplasts that synthesized TRV-CAM nucleoprotein, and they seemed not to result from enhanced accumulation of RRV. They were also produced in intact leaves of doubly infected Nicotiana benthamiana plants, and in protoplasts from singly infected plants that were superinfected with the second virus of the pair.
On mixing in vitro in a wide range of conditions, purified preparations of RRV and TRV-CAM formed aggregates containing particles of both viruses, and this reaction had the same virus and strain specificity as the production of RRV antigen aggregates in vivo. Aggregation seems to result from a reaction that increases hydrophobicity and not from electrostatic attraction of particles of opposite charge. It is concluded that the production of these aggregates in vivo results from a redistribution of RRV particles or coat protein within the protoplast or cell, and that this occurs because of the affinity between RRV particle antigen and TRV-CAM nucleoprotein, the longer particles of which accumulate predominantly on the surface of mitochondria.
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Immunogenic and Cell Attachment Sites of FMDV: Further Evidence for their Location in a Single Capsid Polypeptide
More LessSUMMARYChymotrypsin cleaves only one of the four major polypeptides of foot-and-mouth disease virus (FMDV serotype O) in situ. This polypeptide (VP1, mol. wt. 29 × 103) was first cleaved into fragments of mol. wt. 20 and 9 × 103 and further cleavage could be prevented by the addition of a large excess of bovine serum albumin. The infectivity of the virus particles at this stage was the same as that of the intact virus although the rate of attachment to BHK 21 cells was slower and the immunogenic activity was reduced. If hydrolysis was allowed to continue, VP1 was cleaved into fragments with mol. wt. r8 and < 9 × 103, similar to those obtained with trypsin, and the virus particles then had a greatly reduced infectivity and a lower immunogenicity. Treatment of strains from five other serotypes of the virus with the two enzymes cleaved only VP1 in each instance and there was a corresponding loss of infectivity. The results are discussed in relation to the location and biological activity of the virus polypeptides.
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Thymidine Transport in Herpesvirus hominis Type 1 and 2 Infected BHK 21 Cells
More LessSUMMARYIncrease of dThd-uptake 4 to 12 h after infection of BHK or primary rabbit kidney cells with Herpesvirus hominis of type 1 or 2 can be considered as an early function of the virus genome, because the presence of Cyd-Ara does not prevent the increase of uptake. However, increase of uptake can be prevented by addition of actinomycin D and cycloheximide early in the synthetic cycle.
Two modes of uptake have been differentiated by kinetic analysis: at low substrate concentration dThd is taken up by ‘facilitated transport’, whereas at high substrate concentration (above 2.5 µ m) simple diffusion takes place. The Km of transport of normal BHK or primary rabbit kidney cells (1.4 or 0.5 µ m respectively) is not changed after infection. Only the Vmax increases from 8 to 26.6 pmol in BHK cells or from 2.9 to 9.0 pmol in primary rabbit kidney cells. This indicates that ‘carrier sites’ with identical affinity for dThd-transport are responsible for the increase of transport after infection. This increase of transport is correlated with the induction of a virus coded thymidine kinase (TK) and not with different types of c.p.e. or cellular damage.
Transport of BdUrd increases in a similar manner to that of dThd after infection; transport of dCyd or dUrd increases only slightly, whereas the mechanism of dAdo or Urd uptake by infected cells is quite different.
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Highly Infectious RNA Isolated from Cowpea Chlorotic Mottle Virus with Low Specific Infectivity
More LessSUMMARYRecovery and specific infectivity of infectious RNA from cowpea chlorotic mottle virus of low specific infectivity (14 to 21 day infections) were greatly improved by using antioxidants during virus purification and RNA extraction, and by disrupting coat protein with pronase before phenol-SDS extraction. Total infectivity of RNA from virus of low infectivity was increased over 30 times. RNA profiles obtained using polyacrylamide gels were then similar for virus with high (4 to 7 day infections) or low specific infectivity. Low specific infectivity, therefore, seems to be caused by alteration of the coat protein or of the protein-RNA interaction in intact virus particles.
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Radioactive Labelling of Viruses: an Iodination Technique Preserving Biological Properties
More LessSUMMARYAn iodination procedure suitable for the radioactive labelling of viruses to be used in biological experiments is described. It is characterized by the addition of carrier protein to small amounts of virus before iodination with chloramine T, the use of low concentrations of chemicals, and a rapid purification of the labelled virus to minimize radiation inactivation. Using this procedure, polyoma virus was labelled to a specific activity 100 times greater than that which can be obtained with tritiated amino acids, while its sedimentation coefficient, buoyant density, decapsidation and haemagglutinating activity remained unaffected. Reduction in infectivity, possibly due to radiation inactivation, was slight. Similar results were obtained with adenovirus.
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Volume 105 (2024)
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