132 INFLUENCE OF TEMPERATURE ON BIOLOGICAL SYSTEMS 



faster than that found normally at 37°. The culture eventually grows 

 exponentially at the normal 37° rate and the extrapolated exponential 

 growth curves meet at the time where the temperature shift occurred. 

 Thus, a single temperature shift in the S. typhimurium system does re- 

 sult in a small amount of synchronization. The characteristic behavior in 

 this experiment is reproducible only if the culture has grown exponentially 

 at 25° for several generations. It was also shown (10) that a 37° culture 

 just entering the exponential phase, and exposed for a limited time to 

 25° after which it was returned to 37°, shows a more pronounced syn- 

 chronization effect than is observed in the single temperature transfer 

 experiment. The best synchronization effect resulting from the two tem- 

 perature shifs occurs if the culture is exposed for about 30 minutes to 

 the lower temperature, and it is probably significant that this time is 

 less than, but of the order of magnitude of, the 45-minute generation time 

 at 25°. Nevertheless, even with two temperature transfers, only a part 

 of the cell population is synchronized as evidenced by approximately a 

 50% jump in colony counts occurring 5 to 10 minutes after the second 

 temperature transfer. The most complete synchronization is achieved by 

 means of repeated temperature transfers between 25° and 37°. The pre- 

 cise cycling conditions are different for dilute cultures and for cultures at 

 densities of the order of 3 to 6 X 10" cells per ml, and synchronization has 

 not been effected at densities higher than this. To synchronize cultures 

 containing 3 to 6 X 10' cells per ml the following procedure is used. An 

 overnight 37° broth culture is diluted 100-fold in broth and aerated at 

 37° for 75 minutes at w^hich time it is just coming into the exponential 

 phase of growth. The temperature is then alternately lowered to 25° for 

 28 minutes and raised to 37° again for 8 minutes. The temperature changes 

 are made suddenly by diluting with hot or cold broth in such a way that 

 the density of the culture remains wdthin the limits given above. This 

 temperature cycling is continued for four, five or six cycles and usually 

 several cycles of synchronized growth are obtained in this way. If a cul- 

 ture is so synchronized and then left at 37° the growth rapidly becomes 

 exponential, suggesting that the distribution of division times is very 

 broad. 



Hotchkiss (6) obtained synchronously dividing cultures of Pneumo- 

 coccus by reducing the temperature of a culture growing in the late ex- 

 ponential phase in complex media from 37° to 25° for 15 minutes and then 

 raising the temperature to 37° again. It was shown by means of viable 

 counts that following the second temperature transfer two or three cycles 

 of synchronous growth ensued with a gradual loss of synchrony. A cyclical 

 fluctuation in the susceptibility to transformation also resulted, although 

 not with quite the same period of the synchronous growth curve. 



