62 STUDYING MOLDS, YEASTS, AND ACTINOMYCETES 



will allow ascospore formation to be much more easily brought about 

 than heretofore. Those workers cultured a strain of Aspergillus niger, 

 subcultures of which may be obtained from the American Type Cul- 

 ture Museum. The medium, after some days of incubation, was 

 filtered and resterilized. When several species of Zygosaccharomyces 

 were inoculated into this medium, conjugation and subsequent "spore 

 formation took place much more rapidly than in ordinary media and 

 a much larger proportion of the cells conjugated. It was determined 

 that two substances responsible for this stimulation of sporulation 

 were also produced by Zygosaccharomyces itself, and it was believed 

 that these substances are ordinarily set free into a medium only as 

 the cells die and autolyze. Nickerson and Thimann furthermore 

 found that the inclusion of riboflavin and sodium glutarate in ordi- 

 nary media for yeasts gave as good results as the Aspergillus filtrate. 

 They did not prove that the sporulation-inducing substances of the 

 mold were the same as the chemicals used, but they did show that 

 the results were the same. This work merits much extension. 



A new method of Lindegren and Lindegren -^ for inducing spore 

 formation in yeasts is given here. It has not been tried by the 

 authors but in the hands of the Lindegrens it gave excellent results. 



Presporulation Media 



Beet leaves extract (1500 ml. H2O, 



450 grams beet leaves autoclaved) 100 ml. 



Beet root extract (1500 ml. H2O, 



150 grams beet roots autoclaved) 200 ml. 



Apricot juice (canned) 350 ml. 



Grape juice 165 ml. 



Dried yeast 20 grams 



Glycerol 25 ml. 



Agar 30 grams 



Calcium carbonate 10 grams 



Water is added to make 1 liter. The mixture is steamed until dis- 

 solved, tubed, and sterilized. Most strains will produce spores di- 

 rectly on the slants in a few weeks. If spores are needed sooner, the 

 mixture is transferred to plaster of Paris. Instead of plaster of 

 Paris blocks, the tubes of Graham and Hastings ^- are used. A mix- 

 ture of equal parts of plaster of Paris (CaS04 anh.) and water are 

 mixed and dispensed into test tubes and solidified in a slanting posi- 

 tion, dried at 50° C. for 24 hours and autoclaved. About 1 ml. of 

 sterilized water is poured over a 3-day growth of yeast on the pre- 

 sporulation medium and allowed to stand 10 minutes. A thick sus- 

 pension is made by stirring. Some of the suspension is poured over 



