64: R. MARKHAM 



was discovered by Bawden and Pirie (1937a), and is now known to be very- 

 important for virus activity, Bawden and Pirie used methods similar to those 

 used by Stanley, but employed other refinements, including the use of 

 enzymes for the removal of traces of plant antigens from the virus. 



Shortly after this, methods were developed for the isolation of the virus 

 by means of the newly developed high-speed, air-driven ultracentrifuges 

 (Bauer and Pickels, 1936), and these methods produced virus which was 

 much more uniform and appreciably more infectious than was that made 

 by salting-out techniques (Wyckoff et al., 1937). It is now known that plants 

 infected with tobacco mosaic virus contain more than one substance, and 

 that salting out tends to produce artifacts which have served to cause the 

 confusion which has existed and to some extent still exists in the field. 



B. The Virus in the Plant 



The tobacco mosaic virus is present in diseased tobacco plants (Fig. 5) to the 

 extent of several milligrams per gram of leaf tissue, the actual quantity being 

 dependent upon the age of the plants, strain of virus, and general cultural 

 conditions. In other hosts the yield is generally less, and may on occasion be 

 much less. In tobacco plants a substantial portion of the virus is present as 

 hexagonal plates (Fig. 1) which have been studied by a number of workers 

 (Bawden and Sheffield, 1939) and have been shown by Wilkins et al. (1950) 

 to consist very largely, if not entirely, of uniform length rods, some 300 m/M 

 in length, packed in hexagonal arrays. The structure of these crystals has been 

 investigated very thoroughly by these workers (Fig. 6), and their results 

 have done a lot to confirm the views put forward largely on electron micro- 

 scopic evidence (Oster and Stanley, 1946; Oster et al., 1947; Knight and 

 Oster, 1947; Williams and Steere, 1951; Steere, 1957) that the virus consists of 

 miiform length particles. 



Needle-like inclusions, resembhng Stanley's crystals, which are in fact 

 'paracrystals and only possess two-dimensional symmetry, are also to be 

 found, mainly in the terminal cells of the leaf hairs. The tissues also contain 

 a noninfectious protein (or proteins) related chemically and immunologically 

 to the virus, but which does not contain nucleic acid. The ability of the latter 

 material to disaggregate and reaggregate, and in doing so to engulf non- 

 specific nucleic acid, has been the cause of much confusion. Certainly the 

 early preparations of the virus must have contained much of this material, 

 which may be a precursor of the virus. 



Not all the virus material in plants is released when the plants are ground 

 up. Extra material related to the virus is obtained from the solid residues 

 if they are treated with cellulase or subjected to severe mechanical damage 

 (Bawden and Pirie, ] 945b). 



