25 



report (p. 11) that, when he was working with meningococci, 

 he plated out a culture and examined 40 colonies which were 

 all identical in appearance, but one differed from the rest in failing 

 to lower the titre of the serum for the parent strain, i.e., it showed 

 •' a diminution of antigenic complexity." Schiitze,* again, 

 found that he could get " substrains." i.e., strains in which it 

 was shown by absorption that the " antigen mosaic " was 

 " defective," by simply making single cell cultures from a normal 

 strain. In both these examples, as in Andrewes' work, the 

 variants were obtained from young cultures and the only differ- 

 ences between the variant and the normal were those which 

 could be demonstrated by serological tests. 



Andrewes' data are a particularly good example of what I con- 

 sider to be the earliest stage of spontaneous variation. It is found 

 in young cultures and consists of only a slight irregularity in the 

 synthesis of bacterial protoplasm, characterised by one feature 

 only — irregularity in the disappearance and reappearance of a 

 particular antigenic constituent. 



I propose now to call attention to a more advanced stage of 

 spontaneous variation. Examples are to be found in the work of 

 Arkwright, Benians and Mary Cowan, to which F. Griffith has 

 made reference above (p. 1). In the present connection the 

 following points are worth noting. The production of the changes 

 noted by Arkwright seems to require a more prolonged influence 

 than is the case with Andrewes' variants ; old cultures have been 

 found especially useful for plating out colonies exhibiting the 

 characteristic modifications. And the changes in the variants 

 are more profound and more stable. Their colonies are distin- 

 guished by their " rough " or " smooth " appearance. The 

 source of the former, the " R " variants, " has been almost 



invariably old broth or agar cultures. They may be obtained 

 " from most strains if a culture which has been kept at room tem- 

 " perature or in the incubator for a month or more is plated out." 

 And, though he found that " R " forms might sometimes be made 

 to revert to " S " by daily subculture, the two types were much 

 less liable to reversion than Andrewes' varieties. Like the latter, 

 the " S " and " R " forms " differ very decidedly in their agglu- 

 " tinating, antigenic and absorbing properties with specific sera," 

 but a new feature is found in association with the " R " form ; 



it agglutinates in 0*85 per cent, solution of sodium chloride 

 and in broth cultures it forms a deposit leaving the liquid clear." 

 With regard to his " S " and " R " forms of a Shiga strain, Ark- 

 wright stated that both " were fatal to rabbits when injected 

 " subcutaneously or intravenously in very small doses, but 

 " no attempt was made to find the M.L.D." 



Benians' somewhat earlier observations are worth bringing 

 into line with Arkwright's, as the results are more or less similar. 

 He inoculated a guinea-pig subcutaneously with a mixture of 

 a typical (a) Shiga bacillus and mucilage of tragacanth; the 



* Journ. Hyg., XX., p. 333., 1921. 



