- Volume 36, Issue 1, 1977

In order to clucidate the properties of an inhibitor of rabies virus haemagglutinin in normal animal sera, experiments were made with the HEP Flury strain and calf serum which contains the inhibitor. The results of physico-chemical treatment, gelfiltration and density analysis suggested lipoprotein involvement. When inhibitor and haemagglutinin were mixed, the separate activities could be recovered from the mixture by centrifuging on a sucrose density gradient. By contrast, neither haemagglutinin nor inhibitor could be recovered by this treatment when the inhibitor was added at the start of virus growth. The binding of inhibitor with rabies virus during virus growth seems irreversible and different from the binding of inhibitor with pre-formed rabies haemagglutinin.

Using a plot of genome dry mass against particle dry mass for 58 virus groups, the utility of minimum convex polygons for delineating possible viral taxa has been examined.

Mice challenged with rabies virus were vaccinated 24 h later with one of two types of rabies vaccine. Vaccine prepared from a BHK cell substrate resulted in the production of serum interferon and neutralizing antibody, whereas vaccine prepared from a human diploid cell substrate gave rise to neutralizing antibody but no interferon. Only the first vaccine was effective in reducing mortality.

Various combinations of mouse hyperimmune antiserum, purified IgG, complement and RNase were administered to other groups of mice challenged with rabies, but these had no significant effect on their survival.

Human embryonic kidney cells have been transformed by exposing cells to sheared fragments of adenovirus type 5 DNA. The transformed cells (designated 293 cells) exhibited many of the characteristics of transformation including the elaboration of a virus-specific tumour antigen. Analysis of the polypeptides synthesized in the 293 cells by labelling with 35S-methionine and SDS PAGE showed a variable pattern of synthesis, different in a number of respects from that seen in other human cells. On labelling the surface of cells by lactoperoxidase catalysed radio-iodination, the absence of a labelled polypeptide analogous to the 250 K (LETS) glycoprotein was noted. Hybridization of labelled cellular RNA with restriction fragments of adenovirus type 5 DNA indicated transcription of a portion of the adenovirus genome at the conventional left hand end.

The structure of the iridescent virus type I was examined using negative staining technique. After storage for 1 to 2 months at 4 °C or treatment with chloroform, the icosahedral capsid breaks up into its structural elements — pentagons and triangles, each consisting of 31 and 55 subunits respectively. When such a preparation is centrifuged through a linear sucrose gradient (10 to 40%), 5 zones are revealed. An electron microscopic analysis showed the following distribution of the material (from top to bottom of the tube): (1) free triangles; (2) associated triangles, ‘core’; (3) partially disrupted virions; (4) free intact virus particles; (5) virus aggregates.

A model for iridescent virus type I capsid is proposed, consisting of 12 pentagons (372 subunits) and 20 triangles (1100 subunits). The total number of subunits is 1472.

TSWV nucleic acid, extracted from purified virus with phenol-SDS, was non-infectious. Following electrophoresis of the extracted nucleic acid in 2% poly-acrylamide gels containing 0.5% agarose, 3 major and 2 minor bands were observed. The amount of material contained in the different bands varied with the season. Bands 1, 2, 3 and 4 were sensitive to RNases and resistant to DNase. This was a strong indication of the presence of single-stranded RNA in these bands. Precipitation in 2 m-LiCl, effect of heating and mobility in gels containing an increasing percentage of acrylamide confirmed the single-stranded character. Band 1a was resistant to the action of RNases and stained with Coomassie brilliant blue. The mol. wt. of RNAs 1, 2, 3 and 4 (corresponding to bands 1 to 4) were estimated under both non-denaturing and denaturing conditions and appeared to be 2.5 to 2.7 × 106, 1.9 × 106, 1.7 × 106 and 1.3 × 106, respectively. Following dissociation of the virus with Nonidet P 40, an infectious zone could be isolated after sedimentation in an 8 to 43% glycerol gradient.

The physicochemical and serological properties of a virus isolated from the bivalve mollusc, Tellina tenuis, have been examined. The virus has a diam. of 59 nm, sediments at 430S in sucrose gradients and bands at a density of 1.32 g/ml in CsCl. The virus contains RNA with a mol. wt. about 2.8 × 106 as estimated by poly-acrylamide gel electrophoresis but in sucrose gradients the RNA sediments at 14S. The virus RNA is resistant to ribonuclease under conditions in which ribosomal RNA and the single stranded Mengo virus RNA are completely hydrolysed. Two major polypeptides, mol. wt. 67 and 40 × 103, and one minor polypeptide, mol. wt. 110 × 103, are present in the virus particle. These properties are similar to those found for different serotypes of infectious pancreatic necrosis (IPN) virus. Although there was only a very low level of cross-neutralization between Tellina virus and IPN virus, there was some cross-reaction in immune electron microscopy tests and in immunofluorescence tests with infected tissue culture cells. This cross reaction, together with the close similarity in morphology and physicochemical properties, suggests that Tellina virus and IPN virus belong to the same virus group.

Four cell lines biochemically transformed by u.v.-irradiated herpes simplex virus contain virus DNA fragments ranging from 3 to 22% of the HSV genome. Of five revertant clones selected for 3H-TdR or BrdUrd resistance, four had lost all detectable virus DNA while the fifth, selected for BrdUrd resistance, retained the entire virus fragment but there was a reduction of virus copies per cell from 5 to 1. Three ‘supertransformed’ revertant cell lines contained virus DNA fragments ranging from 12 to 28%. The number of virus DNA fragments per cell ranged from 1 to 5 and clearly indicated that a single copy of the virus thymidine kinase gene is adequate for biochemical transformation. The determination of the base composition of the transforming virus DNA fragment indicated that the transforming DNA has a base composition approximately the same as the HSV genome and does not constitute a low GC virus DNA region. Cross hybridization between HSV-1 transformed cells and HSV-2 DNA is very slight, indicating that the DNA found in clone 139 is not entirely composed of the HSV-1 and HSV-2 common sequences.

A number of chemical and physical agents were screened to determine their effectiveness in inducing simian virus (SV40) production in a virogenic clone of SV40-transformed Chinese hamster cells. Mitomycin C (MC) was the most effective inducing agent, and MC induction was further characterized. It was found that levels of infectious SV40 DNA were increased above control levels as early as 6 h after addition of MC to the culture medium and reached maximum levels by 48 h. Virus capsid (V) antigen and virions followed with a lag of about 24 h. V antigen production was sensitive to hydroxyurea, suggesting a dependence on virus DNA synthesis. The proportion of virus-producing cells (infectious centres) and the virus burst per cell were both stimulated by MC. Studies of 3H-thymidine incorporation demonstrated that the rate of SV40 DNA synthesis was maximal at 48 h post-induction, at which time cellular DNA synthesis was almost abolished. Caffeine, at doses not toxic to non-induced cells, strongly inhibited SV40 production in both non-induced and induced cells, suggesting some role for DNA repair mechanisms.

Chinese hamster kidney cells are semi-permissive to simian virus 40 (SV40). Exposure to mitomycin C (MC) of Chinese hamster kidney cells infected with SV40 DNA enhanced the yield of infectious virus 10- to 100-fold. This stimulation occurred whether the treatment was performed before or after infection. A simultaneous increase in the number of V antigen-synthesizing cells and virus-producing cells, as well as the virus burst size, was observed upon MC pretreatment, whereas the proportion of T antigen-synthesizing cells remained unchanged. MC pretreatment clearly stimulated virus DNA replication in SV40 virus-infected cells. Cells treated with MC exhibited an unbalanced growth pattern, with continuing protein synthesis in the absence of cell division and a markedly reduced ability to replicate the cellular DNA. These results suggest that MC enhances the permissiveness of Chinese hamster kidney cells by inducing the synthesis of a specific cellular factor(s) required for SV40 replication in these cells. Exposure to ultraviolet light also enhanced infectious virus production in Chinese hamster kidney cells.

Egg-grown virus of an influenza A strain (virus N), produces highly infectious particles in this host with its haemagglutinin glycoprotein present in the cleaved form. It also contains relatively large amounts of mucopolysaccharide. This substance cannot be detected in virus derived from cultures of chick embryo cells which has uncleaved haemagglutinin and reduced infectivity. These observations indicate that the host-dependent differences in infectivity cannot be attributed to the presence of mucopolysaccharide as a masking substance at the virus surface, and further substantiate the essential role of the cleavage of the haemagglutinin for activation of infectivity.

Cleavage of the precursor HA into fragments HA1 and HA2 can be accomplished by a variety of proteases. However, only cleavage by trypsin or a trypsin-like enzyme results in formation of highly infectious virus. Activation of infectivity therefore requires cleavage of a specific peptide bond with arginine or lysine in carboxyl linkage.

Double infection experiments demonstrate that virus N displays neither cleavage of the haemagglutinin nor formation of highly infectious virus under conditions where both phenomena are observed with fowl plague virus, another influenza A strain. This observation demonstrates that the relative resistance of the haemagglutinin to cleavage, and thus the resistance of the virus to activation, is a genuine structural property of this glycoprotein rather than the consequence of a low level of proteolysis in infected cells.

Mammalian ribonucleotide reductase is a complex enzyme modified in its activity by a complex regulatory system involving adenosine triphosphate (ATP) and deoxyribonucleoside triphosphates. Infection of KB cells with herpes simplex virus (HSV) type 1 or 2 induces the formation of an altered ribonucleotide reductase. The properties of partially purified reductase from uninfected KB cells have been compared with the enzymes obtained from HSV-1 and HSV-2 infected KB cells. We found that the virus-induced enzymes are similar to the KB enzyme in some properties but differed significantly from the host enzyme in three respects: (1) virus induced reductase was not inhibited significantly by deoxythymidine triphosphate regardless of ATP concentration, (2) magnesium was not required for virus enzyme activity although 2 mm-Mg2+ did stimulate the reaction, and (3) magnesium concentration required for optimal activity was different for virus and host enzymes. These changes are evidence that the enzyme molecules present after infection by HSV-1 or HSV-2 differ from those present before infection.

Judged by the yield of infective virus, good infection of cowpea mesophyll protoplasts with alfalfa mosaic virus is obtained when protoplasts are resuspended at 0 °C in 0.5 m-mannitol, 0.01 m-potassium phosphate, pH 5.6, containing 1 µg/ml poly-l-ornithine and about 2 µg/ml virus. At 25°C, virus infectivity is first detectable 12 h after inoculation and increases exponentially in the next 12 h. Forty hours after inoculation about 1 to 5 × 106 virus particles are produced per living protoplast. As with intact leaves, a mixture of bottom component, middle component and top component b nucleoprotein is required for the infection of protoplasts.

Virus multiplication is not sensitive to chloramphenicol (200 µg/ml) but is completely inhibited by cycloheximide (10 µg/ml) applied at any point in the growth cycle. Actinomycin D (10 µg/ml) interferes with an early step in virus replication.

In two distantly serologically related isolates (A and G12) of tomato black ring virus that each produce two RNA species, RNA-1 was found only in bottom component (B) particles and RNA-2 only in middle component (M) particles. Preparations of separated M and B particles were each barely infective, but produced 8 to 30 times more lesions when mixed, indicating that both kinds of particle are needed for infection, presumably because they contain different parts of the genome. Infectivity was not enhanced when M particles of isolate G12 were mixed with B particles of isolate A, but it was increased when M particles of isolate A were mixed with B particles of isolate G12. The lesions thus produced were abnormally small; isolates cultured from some of them were slower than the parental isolates to produce systemic symptoms in Chenopodium quinoa and had serological properties indicating that their coat protein cistron is in RNA-2. These properties were stable on subculture. Pseudo-recombinant isolates were also produced when RNA-2 of isolate A was mixed with RNA-1 of isolate G12, but not with the converse combination.

An adenine-rich fraction has been extracted from the mycovirus infecting the Phycomycete Allomyces arbuscula. This fraction which accounts for approximately 8% of the total RNA is heterogeneous and contains tracts of 25 to 45 nucleotides in length. The majority of the poly(A) tracts have a mol. wt. of 1.2 × 104 and a sedimentation value of 2 to 3S.

The multiple microtrephination technique was used in the rabbit cornea to compare the activity against herpes simplex virus (HSV) of adenine arabinoside (ara-A), ara-A 5′ monophosphate (ara-AMP) and hypoxanthine arabinoside (ara-Hx), and to determine the effect of addition of adenosine deaminase inhibitor (ADAI) to each. The greatest antiviral activity was shown by ara-AMP, and the least by ara-Hx. ADAI increased the activity of ara-A but had no effect on ara-AMP or ara-Hx.

Crossed immunoelectrophoresis was applied for the characterization of five specific antigens in African swine fever (ASF) virus infected cells. Monospecific antisera were prepared against three individual antigens in the precipitation arcs.

The content and composition of carbohydrate in hepatitis B surface antigen HB8Ag) were clarified by gas chromatography. A value of 75.8 µg carbohydrate per mg of protein was obtained. The main components were N-acetylglucosamine, mannose, galactose and sialic acid and the minor one was fucose. No N-acetyl-galactosamine was detected. The fact that no sugar was detected in lipid fractions suggests that the sugar in HB8Ag exists almost exclusively in the form of glycoprotein and there is no glycolipid.

Five isolates of Mycoplasmatales virus-laidlawii 2 (MV-L2) derived from bovine strains of Acholeplasma laidlawii were shown to differ in host range, plaque morphology and neutralization tests with MV-L2 antiserum. Cross-testing using virus resistant clones of A. laidlawii confirmed the heterogeneity of this group. Adaptation of viruses to sub-optimal hosts was demonstrated.