- Volume 46, Issue 2, 1980

Electron micrographs are presented which illustrate ultrastructural relationships between the polyhedron macromolecular lattice, polyhedron membrane and condensed fibrous sheets in Spodoptera frugiperda cells infected with Autographa californica nuclear polyhedrosis virus. A fibrous material appears to be implicated in the production of all three structures in the infected nuclei. Fibres within an unorganized network were considered to bind together to form a matrix which became ordered into a macromolecular lattice. Accumulations of fibrous masses seen at the periphery of the polyhedron apparently condensed to form the polyhedron membrane. Single and double condensed fibrous sheets are also found in the nucleus; the ultrastructural studies indicate that they too may have arisen by condensation of the fibrous material.

Ultraviolet-irradiated herpes simplex virus (u.v.-HSV) induced endogenous xenotropic type C virus from A1–2 cells, derived from the BALB/c mouse, as determined by infectious centre focus-forming assay on permissive normal rat kidney (NRK) cells. The number of cells induced to release type C virus by irradiated HSV was dependent on the level of u.v. exposure received by the HSV. Optimal induction occurred when cells were infected with irradiated HSV during their exponential growth phase. Virus induction decreased under conditions of simultaneous cellular exposure to hydroxyurea or actinomycin D, inhibitors of DNA and RNA synthesis, respectively, with actinomycin D having a greater inhibitory effect. This suggests that both DNA and RNA synthesis are required for irradiated HSV induction of murine xenotropic virus. Hydroxyurea decreased induction in the first few hours after infection of A1–2 cells with irradiated HSV, suggesting that the biological events involving DNA synthesis which are required for induction by u.v.-HSV occur shortly after infection.

Semliki Forest, Sindbis and rubella viruses can fuse erythrocytes from several different species. Large fusion vesicles consisting of tens to hundreds of red blood cells were seen under optimal conditions. For the haemagglutination and cell fusion to occur the adsorption of virus and further incubation had to be carried out at pH 5.8. Haemagglutination took place over a wide temperature range (0 to 40 °C) whereas fusion required temperatures between 37 and 42 °C. Haemolysis of red blood cells induced by togaviruses also required initial incubation at pH 5.8 to enable attachment of the virus to occur after which the pH of the buffer could be raised to neutrality without inhibiting the haemolysis. The amount of togaviruses and Sendai virus required to fuse red blood cells was about the same [1 haemagglutinating unit (HAU)/ml] but different ionic conditions were required for fusion.

Herpes simplex type 1 Angelotti (HSV-1 ANG) virions were shown to contain two major acid-soluble proteins, BP1 and 2, which by size and charge analysis were also found to be associated with chromatin isolated from HSV-1 ANG-infected African green monkey kidney cells (HSV-chromatin). BP1 and 2 proved to exist in a phosphorylated state both in virions and in HSV-chromatin. BP1 consisted of a single polypeptide of 38 K mol. wt. which was correlated to the tegument protein VP22. In SDS-polyacrylamide gels BP2 migrated as a single polypeptide band with an apparent mol. wt. of 12 K. Urea gel analysis revealed that BP2 consisted of three components, BP2a, b and c, of different phosphate contents. Arguments were provided that these components probably represent different polypeptides of similar mol. wt. HSV-chromatin, in addition to BP1, BP2a, b and c contained a further major virus-induced basic phosphopolypeptide of mol. wt. 65 K which was not detected in acid-extracts of mature virions.

A new and rapid extraction procedure is described for the isolation and partial purification of low mol. wt. RNA as well as DNA from 500 g batches of chrysanthemum stunt viroid-infected chrysanthemum. The chrysanthemum stunt viroid was then purified to homogeneity from this extract by one cycle of non-denaturing polyacrylamide slab gel electrophoresis followed by one cycle of denaturing slab gel electrophoresis. The covalently closed circular form and the linear form of the viroid co-migrated on the first gel but were well separated on the second. The yield of circular chrysanthemum stunt viroid was 200 µg/kg of infected chrysanthemum shoots and of the linear form was 35 µg/kg.

32P-labelled complementary DNA prepared against the circular form was used to show, by hybridization analysis, that the circular and linear forms contained identical nucleotide sequences. The circular and linear forms were equally infectious when inoculated onto Gynura aurantiaca. The linear viroid contained all four nucleotides at the 5′-terminus, about 72% of which had a 5′-phosphate.

We have isolated a new filamentous phage from Xanthomonas citri, a pathogenic bacterium of citrus canker. This phage, designated as Cf, measures 10075 ± 985 × 10−1 nm in length and forms an unusually clear but tiny plaque on the host within 12 h p.i. Infection by Cf neither killed nor drastically prevented host cells from propagation. However, the rate of host cell division was severely retarded. This phage exhibited an exceedingly narrow host range and was unstable in conventional tris buffer and synthetic medium routinely used for the preparation of other filamentous phages. Based on nucleotide composition analysis (which also revealed that Cf contains an unusual nucleotide), thermal denaturation characteristics and the hydroxyapatite column elution pattern, the genome of Cf was shown to be single-stranded (ss) DNA. During phage multiplication in host cells single-stranded virus DNA, replicative form I and replicative form II were detected. As shown by acrylamide gel electrophoresis, the size and conformation of the virus, RFI and RFII DNA species, were the same as their counterparts of another filamentous phage Xf.

Enzyme activities of particulate and solubilized RNA-dependent RNA polymerase preparations from tobacco plants infected with 10 ts mutants of alfalfa mosaic virus (AMV), having their mutations in each of the three parts of the virus genome, were found to behave similarly with respect to temperature as compared to preparations obtained from plants infected with wild type AMV or from uninfected plants. Thus it is very unlikely that a virus-specific protein is involved in these in vitro RNA polymerase activities.