The Mineral Requirements for Sporulation 



Harold R. Curran 



Dairy and Meat Lab., Eastern Utilization Research and Development Divi- 

 sion, Agricultural Research Service, U. S. Department of Agriculture, 

 Washington 25, D. C. 



ALTHOUGH scores of papers have been published on various aspects 

 of spore formation, relatively few have been concerned directly with 

 the role of minerals in sporogenesis. Salt effects on sporulation were 

 reported by Behring as early as 1889, and by Schrieber in 1896. However, 

 their real significance in the production of spores was not recognized until 

 many years later. Thus, Cook, in his comprehensive review "Bacterial 

 Spores" (1932), concluded "it does not appear that salts exert a direct in- 

 fluence on sporulation." The inability of early workers to devise chemically 

 defined media that would produce spores in quantity contributed to the 

 slow development of knowledge in this field. Cook (1931) and Tarr (1932) 

 were the first to overcome this difficulty, the latter by showing that good 

 sporulation by several aerobic species could be obtained by cultivation 

 in a mineral salts medium containing low concentrations of secondary am- 

 monium phosphate and sucrose. Roberts (1934) obtained 60-70% sporu- 

 lation of Bacillus subtilis with suitable minerals supplemented with aspar- 

 agine and levulose. Leifson (1931) studied the effects of inorganic salts on 

 the sporulation of Clostridium botulinum in 1% peptone. The basal med- 

 ium, which by itself produced no spores, was supplemented with a variety 

 of mineral salts used alone or in combination. Sporulation occurred only 

 when the supplement contained NH4+ and PO4 and to some extent 



SO4 ions. The Ca + + ion stimulated sporulation when added to NH4 + 

 and PO4 ions. 



Further evidence that peptone is deficient in minerals for sporulation was 

 supplied by Fabian and Bryan (1933), who observed greatly increased spor- 

 ulation of four mesophilic aerobes when the peptone solution was suitably 

 fortified with cations of the univalent chloride salts; di- and tri-valent ions 

 were without effect. Meat infusion tryptone glucose broth was shown by 

 Knaysi (1945) to be deficient in Mg++ for the sporulation of Bacillus my- 

 coides. The work of Foster and Heligman (1949) indicated that not only 

 peptone but most complex organic media, when used in the fluid state, do 

 not provide an adequate supply of minerals for the sporulation of Bacillus 

 cereus; the chief limiting factor in asporogenic media was shown to be an 



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