SYNCHRONOUS GROWTH FOR SPORE PRODUCTION 15 



problem of spore production. I am sure you will want to question him 

 about his procedures and results, so my discussion of his paper will be 

 short. 



In order to study spore germination it is necessary to produce free un- 

 germinated spores and separate them from the vegetative cells, germinated 

 spores, and the sporangia that contain spores. Mr. Collier has presented an 

 approach to the problem that has several merits. With some anaerobes an 

 incubation period of 2 to 3 weeks is reconmiended for optimum sporula- 

 tion. He has been able to produce spores from CI. roseum in a matter of 

 5 to 7 hours. It is also interesting to note that the percentage sporulation 

 is quite high if the culture is examined prior to the so-called "recycHng" 

 process. 



He was able to lyse the vegetative cells by storage at 4°C. for 1 to 2 

 weeks. With another anaerobe, PA 3679, we were not successful in lysing 

 the cells at 4°C., but we did use lysozyme to remove the vegetative cells 

 from the harvested spore crop. This procedure was very successful. 



From the information presented, it seems that there was a gradual pro- 

 duction of spores, and the combination of aeration with penicillin blocked 

 or arrested further sporulation of the culture. Aeration alone or penicillin 

 alone will also block sporulation. This suggests a similarity between these 

 results and the endotrophic sporulation of aerobic bacilli. 



Hardwick and Foster (1953) studied the enzymatic changes during sporo- 

 genesis in some aerobic bacteria and advanced the hpyothesis that sporo- 

 genesis occurs at the expense of proteins (or enzymes) pre-existing in the 

 vegetative cell. The procedure was similar to that of Halvorson and Spie- 

 gelman (1952) who established, by means of amino acid analogues, amino 

 acid involvement in adaptive enzyme formation in yeast. Additional work 

 may show a similiar type of mechanism in the sporulation of anaerobic 

 bacteria. 



The Michigan State workers also report that a synchronized technique will 

 work for the production of PA 3679 spores. This is a new procedure and 

 I feel that is a fruitful field of endeavor. 



Informal Discussion 



Rayman: In the commercial butyl fermentation, the culture must be trans- 

 ferred five to six times to get good production of solvents. The yield of 

 butyl alcohol and acetone is poor for the first and second transfers, but 

 improves up to the fifth. After that, the yield drops so that by the eighth 

 or ninth transfer it is practically nothing. Mr. Collier, I am wondering if 

 you are getting a sporulation-germination cycle that is equating itself about 

 the fifth transfer. This is my explanation of why the yield of solvents is 



