106 G. G. KRISHNA MURTY 



slight carry-over of activity in the beginning. Tested with glucose and L- 

 alanine, the lag period was reduced, but there was no decrease in the rate 

 of oxidation. 



Murrell concluded that L-alanine had to be present for the oxidation of 

 glucose and that the rate of oxidation was a function of both the number of 

 spores and the concentration of L-alanine. 



Church and Halvorson (1955, 1956), working with aged spores of B. 

 cereus var. terminalis, found that when the spores were heated at 65° from 

 60 to 90 minutes, they rapidly oxidized glucose in the absence of any de- 

 tectable germination of the spores. They found that the spores so activated 

 could also oxidize gluconate, 2-ketogluconate and pyruvate, but I shall leave 

 the discussion of these to Dr. Church. However, freshly grown spores either 

 failed to oxidize glucose or oxidized glucose at an extremely low rate under 

 similar conditions. They further observed that adenosine had the same effect 

 as heat shock in aged spores. The heat activation decayed with time, but 

 activity could be regained by a second heat shock, although the degree of 

 restoration decreased with the time elapsed between the first and second heat 

 treatments. 



We have obtained essentially similar results. Freshly grown or aged spores 

 of B. terminalis germinated rapidly and completely, but failed to grow when 

 incubated with the germinating nutrients under anaerobic conditions, an ob- 

 servation independently made also by Roth with other aerobic spores. Dense 

 suspensions of spores of B. terminalis failed to germinate completely under 

 anaerobic conditions, even when incubated for a long period with large 

 amounts of the nutrients supplemented with glucose. The resulting mixture 

 of germinated and ungerminated spores was washed free of the nutrients, 

 and by differential centrifugation a suspension relatively rich in ungerminated 

 spores was obtained. When heat-shocked to kill the germinated spores, this 

 suspension would actively oxidize glucose. This led us to believe that it 

 might be possible to activate the glucose-oxidizing enzymes of the spores in 

 the absence of germination, by heating a thick suspension of the spores in 

 the presence of small amounts of the germinating nutrients. This was in- 

 deed found to be the case, and the relations between the amount of enzymes 

 so activated, the duration of heating, the number of spores, the concentra- 

 tion of nutrients, the time of addition of nutrients, etc., are given below. 



Results. The amount of enzymes activated seemed to be a function of the 

 duration of heat shock and the concentration of nutrients. With a low con- 

 centration of nutrients the limiting activity of the enzymes is reached in 

 7 hours of heating (Table I). 



With a fixed concentration of nutrients, the amount of enzymes activated 

 became a function of both the duration of heating and strength of the spore 



