- Volume 17, Issue 3, 1972

Intrinsic interference is not limited to Newcastle disease virus: vesicular stomatitis (VSV) and orthomyxoviruses can be similarly inhibited at the stage of RNA synthesis. Detailed investigations of the interference of fowl plague virus (FPV) by VSV show: (1) adsorption of the interfering virus is not responsible; (2) the synthesis of all FPV virus components is affected, if the interfering virus is present during the initial stages of FPV replication and (3) competition of the interfering RNA polymerase with heterologous templates can be excluded by experiments carried out in vitro.

Temperature-sensitive mutants of adenovirus type 5 have been examined by various serological techniques. Cells infected with these mutants at the non-permissive temperature responded in different ways and four main serological groups could be assigned on the basis of complement fixation tests, namely group I, which showed no major differences in the production of capsid antigens; Group II, which showed no production of any of the capsid antigens; Group III, which showed no production of fibre antigen; and Group IV, which showed limited production of hexon antigen and variable yields of penton base antigen. Fluorescent antibody tests further subdivided the latter group into mutants which showed no production of hexon antigen and those which showed production of hexon antigen, although with an abnormal distribution. These results were in good agreement with the complementation data although within most of the serological groupings there was more than one complementation group.

Inositol, sodium glutamate and calcium lactobionate were found to protect foot-and-mouth disease virus against inactivation during spraying and equilibration in the first 1s in aerosols. They also protected virus against inactivation during freeze-drying.

Dimethyl sulphoxide and glycerol both protected virus against the inactivation which occurred between 1s and 5 min in aerosols, but they did not protect during freeze-drying. Pre-humidification before sampling reduced the inactivation of virus, particularly at 40% r.h. The different protective mechanisms of these compounds and of pre-humidification are discussed in relation to the mechanisms of aerosol inactivation.

Isolated protoplasts from healthy tobacco mesophyll tissue were infected with a Rothamsted culture of the common strain of tobacco mosaic virus. The average yield of virus/infected protoplast was estimated at 1.4 to 5.8 × 106 particles, which was almost as much as found in intact plants. Virus titre was assessed by infectivity assay, electron microscopy, and serological techniques.

High concentrations of interfering component added to BHK cells infected with the Indiana strain of vesicular stomatitis virus (VSV) inhibited the synthesis of virus and interfering component. The inhibition was at an early stage in virus replication. Lower concentrations of interfering component led to the formation of interfering component and depression of virus synthesis. When virus replication was depressed with interfering component prepared from a second strain of VSV (Brazil strain) the RNA of the Brazil interfering component was also replicated at the expense of the Indiana virus. The proteins of the progeny interfering component particles were of the Indiana virus type. It is proposed that interference can take place at different stages during infection, depending upon the amount of interfering component present.

Antisera produced in rabbits against extracts of cells infected with type 1 or type 2 herpes simplex virus neutralized thymidine kinase activities of six type 1 and six type 2 herpes virus strains. There was intertypic neutralizing activity, suggesting some common antigenic determinants, but much greater intratypic activity, suggesting that the enzyme bears type-specific determinants.

The effects of some anti-type 1 sera were complicated since they also stabilized the type 2 enzyme activity.

When anti-type 1 and anti-type 2 sera were absorbed with extracts of cells infected with thymidine-kinase deficient mutants of homologous type they retained their power to neutralize thymidine kinase activity but completely lost their ability to neutralize infectivity. Each absorbed serum gave a single precipitin line against the parent strain and none against the parent of the other type. It was surmised that these precipitin lines were associated with type-specific thymidine kinase antigens.

Partial or total removal of the envelope of influenza virus by enzymes or chemicals displayed an internal body, which is referred to as ‘virus core’. This is supposed to consist of the nucleocapsid and a ‘core membrane’ seen on the core surface in ultrathin sections as a single track about 30 Å thick. The virus membrane isolated by means of Nonidet P-40 revealed a typical triple track profile of 25 Å each track. ‘Cores’ limited by a single-track membrane were found only after treatment of fixed virus particles with snake venom. Other agents such as saponin, Nonidet or chloroform-methanol were efficient in removing the virus membrane, but, unfortunately, they damaged the fine structure of the ‘cores’.

Negative staining of virus particles treated overnight with proteolytic enzymes revealed a ‘helical skeleton’ of 35 Å thick fibres which may have been a part of the ‘core membrane’.

The developmental sequence and subsequent release of the herpes simplex virus have been extensively studied by many investigators. Results by Morgan et al. (1959), Shipkey et al. (1967), Nii, Morgan & Rose (1968) have clearly shown that the naked nucleocapsids first appear in the nucleus of infected cells. Envelopment of the nucleocapsids occurred most frequently at the inner lamellae of the nuclear membrane and to a lesser extent at the membrane of cytoplasmic vacuoles (Nii et al. 1968). However, the mechanism of virus release is still controversial. Epstein (1962) described virus budding at the cell surface while Morgan et al. (1959) believed that viruses were released by the process of ‘reverse phagocytosis’. Recently, Schwartz & Roizman (1969) demonstrated the presence of cytoplasmic tubules which transported the enveloped nucleocapsids from perinuclear cisternae into the extracellular fluid. The entry of the infectious bovine rhinotracheitis (IBR) virus into the susceptible host cell has been previously reported (Zee & Talens, 1971).

In cells infected with paramyxovirus progeny 50 S RNA seems to be incorporated immediately into the nucleocapsid (Blair & Robinson, 1970). There has been no report of any attempt to estimate the time interval between the synthesis of a molecule of nucleocapsid protein and its incorporation into the structure of the nucleocapsid. In the experiments described in this paper the problem has been approached with the use of a pulse-chase, using protein precursors. The increase of label in the 200 S peak under chase conditions was used as a measure of the rate of nucleocapsid formation.

Stocks of the beaudette strain of Newcastle disease virus (NDV) were grown in eggs. Primary chick embryo cell cultures were inoculated as described by Kingsbury (1966), at a multiplicity of 10 to 20 p.f.u./cell and incubated at 37 °C. Lactalbumin hydrolysate (0.5%) in Hanks’s balanced salt solution (BSS) was used as maintenance medium.

The ionic strength and ionic composition of the medium can markedly affect certain aspects of virus replication or maturation in the case of some picornaviruses (Wallis & Melnick, 1962; Tolskoya et al. 1966; Fiala & Kenny, 1967; Blough et al. 1969; Agol et al. 1970) and in the case of Sindbis virus (SV), a group A togavirus (Waite & Pfefferkorn, 1970). In general, a lowered ionic strength appears to inhibit while a higher ionic strength increases virus yield. Blough et al. also reported enhanced crystallization of rhinovirus in HeLa cells after increasing the concentration of MgCl2 in the medium. Our observations described below show that the yield of dengue virus (DV), a group B togavirus, is also affected by the ionic strength of the medium. However, no clear-cut effect on the crystallization of DV in Vero cells (Matsumura, Stollar & Schlesinger, 1971) was noted after increasing the MgCl2 concentration.

Poly (A) segments form a part of mammalian mRNA (Darnell, et al. 1971; Edmonds, Vaughan & Nakazato, 1971; Mendecki, Lee & Brawerman, 1972), vaccinia virus mRNA (Kates, 1970) and the RNA of a number of RNA viruses (Armstrong et al. 1972; Gillespie, Marshall & Gallo, 1972; Johnston & Bose, 1972; Lai & Duesberg, 1972). The potential significance of the poly (A) segment in mRNA function is unknown, but it is of interest that the genomes of RNA tumour viruses contain a much greater proportion of poly (A) than the RNAs of other viruses and that poly (A) segments have not been detected in virus RNAs that do not function directly as mRNA, such as the RNAs of influenza virus, Newcastle disease virus and vesicular stomatitis virus (Gillespie et al. 1972; Lai & Duesberg, 1972). It has been reported that the poly (A) segment of poliovirus RNA represents 0.75% (Armstrong et al. 1972) or 0.25% (Gillespie et al. 1972) of the total nucleotide content of the virus genome.

The symptoms and histopathology of many virus diseases depend not so much on damage to the cells by virus but on the immunological reaction to the infection. The earliest (Traub, 1935) and still extensively studied (Oldstone & Dixon, 1970) infection of this type is murine lymphocytic choriomeningitis. From the in vivo work it has been commonly accepted that lymphocytic choriomengitis virus (LCMV) is intrinsically temperate, but there are numerous examples of LCMV-induced cytopathic effects (c.p.e.) in tissue culture (Pfau & Camyre, 1968). We have recently shown that c.p.e. in LCMV-infected L cells is largely regulated by an interference phenomenon (Welsh & Pfau, 1972). Cell culture-grown stocks of LCMV contain a transmissible component that specifically interferes with LCMV infective centre formation, synthesis and cytolytic activity; this interfering component sediments with the infectious virus particle, fails to pass through a Millipore filter of 25 nm pore size, and is inactivated by heat (60 °C), neutral red and LCMV-immune serum (Welsh & Pfau, 1970, 1972).