NUTRITIONAL AND ENVIRONMENTAL CONDITIONS 23 



Table I 



Sulfur requirement for sporulation of 



Bacillus coagulans [43-P] 



Percent sporulation 



1. Complete medium 75-90 



2. [1] minus S-amino acids 5-10 



3. [2] plus inorganic SO4 70-90 



appeared to be unaffected. However, we did one experiment too many. We 

 re-examined the composition of our medium and noted that, when sulfur- 

 containing amino acids were left out, the only sulfur in the medium other 

 than the unknown amount which might be contributed by the agar was 

 that contributed by the 10 ppm MnS04 which we had added to the medium 

 to satisfy the manganese requirement. When we fortified the medium with 

 inorganic sulfate and left out the methionine and cystine, the organism 

 sporulated equally as well as in the complete medium. The complete data 

 suggest that instead of a methionine requirement the organism actually has 

 a sulfur requirement which is greater for sporulation than for growth. I 

 think that these particular data serve to emphasize a more important point, 

 namely, that when we investigate sporulation we are studying a phenome- 

 non of the interdependent relationships and we must be cautious in drawing 

 conclusions unless we know these relationships. 



The role of alanine in stimulating or suppressing sporulation is con- 

 troversial. In our laboratory, both in shake culture and in replacement cul- 

 ture, L-alanine exhibited some stimulatory effect towards the sporulation of 

 B. coagulans, even though this compound is an active germination stimu- 

 lant for spores of this organism. Foster and Heligman (1949a), using B. 

 cereus cultured in a glutamate-salts medium, reported that the addition of 

 L-alanine, DL-alanine, or /^-alanine suppressed the stimulatory effect of glu- 

 cose towards sporulation. This suppression of sporulation could be re- 

 versed by the addition of yeast or liver extract, leucine or isoleucine. Gre- 

 let (1955), working with different strains of B. cereus cultured in a glu- 

 cose-amino acid-salt medium, demonstrated the following : ( 1 ) his strains 

 were exacting to both glutamate and alanine while the other three amino 

 acids — valine, leucine, and isoleucine — stimulated growth; (2) good spor- 

 ulation always occurred when alanine in the complete medium became limit- 

 ing; and (3) good sporulation was also provoked when the valine, leucine, 

 or isoleucine became limiting even though excess alanine was still present 

 in the' medium. 



(c) Nutrilites. The only data previously presented on the effect of a 

 specific growth factor on sporulation is that of Williams and Harper (1951), 



