Journal of General Virology - Volume 1, Issue 4, 1967

Proflavine inhibits DNA-dependent RNA synthesis in a way similar to actinomycin (1). It binds to DNA (2) and RNA (3, 4) and interferes with virus multiplication (4, 5). A dose-effect relationship has revealed that its binding capacity to DNA goes in parallel with the inhibition of cellular RNA synthesis and with the inhibition of the formation of fowl plague virus; while its binding capacity to RNA parallels the inhibition of protein synthesis and the inhibition of the formation of Newcastle disease virus (4). Therefore it has been suggested that proflavine inhibits fowl plague virus multiplication by the same mechanism as actinomycin does. The inhibition of Newcastle disease virus formation, however, should be due to an interference with an RNA template or t-RNA (6), inhibiting protein synthesis.

This investigation was concerned with the action of proflavine on the RNA-dependent RNA synthesis. The multiplication of Newcastle disease virus, strain italien, in primary chick fibroblast cells was chosen as a model.

The nutrition of ruminants is intimately connected with microbial digestion in the rumen. Because a substantial proportion of the diet ultimately utilized by the animal exists as bacterial substance for a time, the fate of the bacterial cell in the alimentary tract is of interest. We are at present investigating the possible significance of the bacterial cell wall in the nutrition of the sheep as such walls constitute from 10 to 30% of the dry weight of the cell and are known to contain carbohydrate (1). As a part of the investigation a general survey by electron microscopy of bacterial types present in rumen contents was made. Phage particles both free and attached to cells were found to be present. Many empty bacterial cell envelopes and fragments of cell walls were also found.

Recently Anderson and his associates have described a series of high-speed zonal centrifuge rotors designed to allow small animal viruses to be isolated from large volumes of culture fluid without pelleting (1, 2). In these rotors the culture fluid flows as a thin Sheet across the surface of a liquid density gradient. Particles suspended in the fluid are rapidly sedimented out of the stream and become trapped in the gradient solution. Further centrifugation causes the particles to band at, or near, their isopycnic points. The capabilities of this type of rotor have been explored with adenovirus and T3 phage (2). This paper describes the application of continuous flow zonal centrifugation to the problem of concentrating and purifying large quantities of polyoma virus and Semliki Forest virus from c. 51. of tissue culture fluid.

A Beckman model ZU ultracentrifuge and a BXVI continuous flow zonal rotor were used in these experiments.

When bacteria lysogenized by certain temperate phages like λ and p2 are superinfected by distinguishable mutants of their prophage, the superinfecting phage appears in the progeny released after induction of vegetative phage growth (1, 2). Phage p22 differs, however, for although bacteria lysogenized with it adsorb superinfecting phage p22, no superinfecting genetic markers like c 2 (clear plaque) or h (host range) appear in the phage obtained after induction, whether spontaneous or produced by u.v. irradiation. Superinfecting phage p22 is thus excluded and the wild-type prophage may be termed ‘excluding’ (x +). Non-excluding (x) mutants have now been isolated by taking advantage of the phenomenon of lysogenic conversion.

Salmonella typhimurium only forms somatic antigen 1 (01) if it is infected by phage p22 possessing the converting gene a1 + (3).

There is as yet no generally accepted theoretical basis for the classification of viruses. It has been suggested that this could be supplied by the theory that the properties of a virus are determined by the sequence of bases in the viral nucleic acid (1, 2). The classes which have to be constructed are then those of viruses whose nucleic acids have common base sequences and which are therefore partly equisemantic (2). Although base sequences cannot yet be directly determined, a preliminary general classification of viruses has been constructed by applying numerical taxonomic programs to attributes of viral nucleic acids (2), and could be tested by nucleic acid hybridization experiments. Discrepancies are likely to arise when intuitive ideas about virus interrelationships are compared with numerical classifications based on properties of the viral nucleic acids. Some of these may be due to faulty data, chance or distortion of relationships by the numerical program, but some may indicate relationships which were not previously considered.

The burst size of phage p22 in Salmonella typhimurium strain lt2 is 200 to 1000, whether a clear plaque mutant (1, 2) or a low multiplicity of wild-type (c +) phage is used. However, when vegetative phage growth is induced in strain lt2 lysogenized by c + phage by exposing it to ultraviolet light from a low pressure germicidal lamp, the burst size is far smaller and usually less than 40. This is so even when the prophage is one of the non-excluding mutants already described (3) and the bacteria are superinfected with virulent (c 2) phage just before irradiation, despite the appearance of both c + and c 2 phage in the burst. These observations suggested that the dose of u.v. radiation producing maximal induction damaged the bacteria to a degree that severely limited their capacity to support phage growth.

We recently reported that the two components from polyoma virus DNA which can be separated after preparative ultracentrifugation (1) show different chromatographic properties on hydroxyapatite (2). Analytical ultracentrifugation and electron microscopy established that the fast component (component I, 20 S) consists of twisted circular molecules only, while the slow component consists of a mixture of untwisted circular molecules (component II, 16 S) and linear molecules (component III, 14.5 S) in variable proportion depending on the DNA preparation examined (3). Since components II and III cannot be satisfactorily separated by preparative ultracentrifugation, the nature of the slow component we isolated by this method was still a matter of uncertainty and the basis for its chromatographic separation from the fast component had therefore to be questioned. We report here the results of experiments made to solve this problem.