FUNGOUS DISEASES OF THE HONEYBEE 11 



Isolations of single spores can be made as readilj from Czapek's 

 solution agar as from maltose agar, consequently it was frequently 

 used for this purpose. 



BEER-WOIKr Gwr.ATiw 



For the study of yeasts, beer wort was prepared from malted barley 

 and hops as follows : Two hundred grams of the finely ground barley 

 was soaked for an hour, with occasional stirring, in a liter of dis- 

 tilled water at 60° C. Four grams of hops was then added and the 

 mixture boiled for an hour during which time it was stirred at inter- 

 vals of a few minutes. Water was added from time to time to main- 

 tain the original volume. The barley meal was then separated from 

 the liquid by straining it through closely woven cloth, and the liquid 

 was cleared with the white of eggs. The quantity of maltose present 

 was determined, and water was added until the liquid beer wort con^ 

 tained approximately 3 per cent of this sugar. Gelatin was added 

 to the clear beer wort when a solid medium was desired. 



Potato Media 



Culture media for yeasts were prepared by cutting pieces of pota- 

 toes the proper size for the culture dish and sterilizing them after 

 stoppering or covering. Potato-broth agar was prepared at first for 

 the purpose of storing yeasts, but since tliere was no particular advan- 

 tage in its use, it was replaced with beer-wort gelatin. 



Liquid Nutrient Media 



Synthetic and natural liquid nutrient media were prepared accord- 

 ing to the formulas just given under solid media, except that agar and 

 gelatin were omitted. Honey, dextrose, and levulose were sometimes 

 substituted for the sugars given in these formulas. Beer wort was 

 used extensively in the study and identification of yeasts, since most 

 of the morphological and cultural characteristics of yeasts reported 

 in the past were made from growth in this nutrient medium. 



Sucrose, dextrose, levulose, maltose, galactose, dextrin, inulin, raf- 

 finose, d-mannose, and 1-arabinose were tested in liquid media to deter- 

 mine the ability of the yeasts listed on page 6 to ferment these 

 sugars. 



EXPERIMENTS WITH BEES 



PRELIMINARY INOCULATION WITH MOLDED COMBS 



In a previous study of the fungi associated with honeybees (5, 

 p. 63, 68) , it was observed that species of two groups, the Penicillia 

 and the Aspergilli, occur with greater frequency than do other forms. 

 These fungi, Penicillia and Aspergilli, were found within the bodies 

 of bees where the mycelium had permeated all of the softer tissues, 

 resulting in a characteristic mumification. Formation of spores of 

 these fungi was observed to occur both within the exoskeleton and 

 upon the exterior. After these observations a question arose as to the 

 relation of these organisms to the honeybee. Was the bee attacked 

 before death, or afterwards? If before, death might have resulted 

 from infection by the fungus. 



