Discussion 77 



Lederberg: That would suggest one of two possibilities: either (1) that 

 during the course of growth in broth you accumulate a higher frequency 

 of alteration, depending on the nuclear configuration of the cell that you 

 begin with and that shows up either in both or in only one of the first 

 daughter progeny; or (2) that the noxious stimulus is not agar but the 

 switch from broth to agar, which may cause some imbalance in the 

 relative rate of formation of cell wall or cell substance. Once the cells 

 start to grow on agar they might become well adjusted to those particu- 

 lar circumstances. Have you got the observational evidence that would 

 bear on this question? 



Hughes: I don't think so. I have always thought that there was a 

 mutation involved in this, and that in broth it was no disadvantage to 

 have this particular character, and that the population built up in broth 

 would be detected when transferred to solid medium. ^Vhether it is the 

 shock of transfer to solid medium, or something else, I don't know. 



Lederberg : Is there any indication that the time of cultivation in broth 

 has an influence on the proportion of lethals which appear when the 

 clones are transferred back on agar ? This would be one line of evidence 

 relative to the question of accumulation of these mutations in broth. 



Hughes: My impression was that a short subculture, i.e. building up 

 a microcolony of perhaps 200 or 300 in broth, gave much the same per- 

 centages as an overnight broth culture. I would like to repeat that. 



Lederberg: That would favour the conception that it is the sharp 

 change of medium that is the noxious stimulus. 



Ultraviolet light induces a possibly analogous abnormality. After a 

 week of exposure of Esch. coli K12 to ultraviolet 60-80 per cent of the 

 cells may divide to give one normal, rapidly dividing offspring, and 

 another abnormal, swollen and probably in viable one. This is almost 

 certainly a reflection of the multinucleate structure of the cells (Leder- 

 berg, J., et al. (1951), Cold Spr. Harb. Symp. quant. Biol., 16, 413). 



lerusalimsky : Dr. Hughes mentioned in his interesting report a very 

 important point on the inequality of bacteriological cells arising from 

 division. This question has not been investigated sufficiently. It is not 

 known whether inequality of cells is random and non-regular or whether 

 one of them always resembles the mother and the other the daughter. 

 Malek in Czechoslovakia and Streshinsky in the USSR support the 

 second point of view, and our experiments also tend to support it. Our 

 experiments are directed towards explaining the causes of the varying 

 resistance of individual cells in a population, and discovering the relation 

 between the degree of resistance and the physiological age of cells. 

 Unless these individual differences are taken into consideration, it is very 

 difficult to distinguish adaptive phj^siological changes from mutations. 

 It seems to me that Dr. Hughes' film has proved that resistance to peni- 

 cillin arises mostly in young daughter cells, and therefore this pheno- 

 menon is probably a physiological adaptation. 



