74 H. HALVORSON et al. 



1 % DPA, viability decreases and a higher dependence of heat 

 resistance on DPA is also evident. Black, Hashimoto and 

 Gerhardt^, employing the technique of endotropic sporulation 

 in distilled water, produced low DPA spores which were found 

 to be relatively susceptible to heat. This has recently been 

 extended to show a correlation between the Ca2+ content of the 

 sporulation medium and DPA biosynthesis^"*. The presence of 

 these two may be manditorily coupled such that Ca2+ is required 

 for DPA synthesis and the latter as a means of chelating and 

 maintaining high levels of Ca-+ or other divalent cations. 



RESPIRATORY ACTIVITY OF DORMANT SPORES 



The second biochemical description of spores concerns their 

 apparent physiological inertness. Consider, for example, the 

 aerobic sporeformers which require oxidative reactions for the 

 supply of both energy and building materials. The respiratory 

 rate (Qo.) on glucose for vegetative cells ranges from 60-100. 

 Reports on spores have varied considerably. Levinson and 

 Hyatt^^ reported a Qo, value of 2.4 in a preparation of B. 

 megatherium spores containing approximately 10% germinated 

 forms. Crook^^, in examining a well washed suspension of 

 B. subtilis spores, observed a Qo, value of 0.3 by means of a 

 microrespirometer. Unfortunately he did not report the per- 

 centage of germinated forms. In any such studies the cleanliness 

 of the spores and the absence of germinated forms are essential 

 to measurements of the metabolic activity of the dormant forms 

 themselves. 



We have reexamined this point by extensively washing fresh 

 spores of B. cereus 12-16 times until they are free of detectible 

 levels of germinated forms^". When dormant spores were tested 

 at very high densities (30 mg/ Warburg cup) there was no 

 detectible O2 uptake after 60 min in the presence of glucose, 

 whereas heat-shocked spores took up 60 //I O2 and aged heat- 

 shocked spores 300 [A O2. In the latter case extensive germina- 

 tion is associated with respiratory activity. Based on the limita- 

 tions of the manometric technique employed, the Qo, of the 



