and Laboratory Methods. 2215 



filaments would tend to prove it (Fig. 9). The hyphal bodies are often found 

 attached to one another in considerable masses, each with a distinct cell wall or 

 as a " bud," the separation not being complete. Both hyphal bodies and rest- 

 ing spores were found in the same grasshopper. In all probability, if the hyphal 

 bodies do not germinate and produce conidia on the outside of the body, they grad- 

 ually become spherical, put on an extra cell wall and become resting spores. As 

 yet I have found no resting spores in caterpillars corresponding exactly to those 

 found in grasshoppers. There were some so much like resting spores in size 

 and shape that it was a difficult matter to tell which was which when they were 

 lying side by side. Both are spherical, had two heavy cell walls, and the inter- 

 nal structure was similar (Figs. 13, 14). The one essential difference was that 

 the cell wall of those obtained from the grasshoppers was slightly colored. The 

 thick walled bodies (Figs. 9-13) in the caterpillars were not formed until after 

 they had been kept in the laboratory for over four months ; they were of various 

 sizes and shapes, from spherical to cylindrical and irregularly curved ; they 

 occurred singly in filaments (Fig. 9) and in masses loosely joined together, sep- 

 aration having apparently taken place by either budding or fission (Fig. 12). 

 Whether these irregular shaped bodies will in time become spherical remains to 

 be determined, also whether they still retain their vitality and are capable of ger- 

 mination. I have not yet been able to germinate the resting spores from grass- 

 hoppers, but I feel confident that it can be done by exercising a little ingenuity 

 and patience. When it is time for them to germinate they will undoubtedly do 

 so readily. Considerable difficulty has sometimes been experienced in attempt- 

 ing to germinate certain seeds during the autumn and winter, yet they germinated 

 when it was time for them to do so naturally. A " resting spore " may not be 

 resting, but on the other hand doing something — maturing — and will germinate 

 only after it has matured and under the proper conditions. The hyphal bodies 

 and conidia when fresh germinated after being in water for a short time. 



CULTURES. 



Cultures from Hyphal Bodies. When I found out that the conidia and hyphal 

 bodies would begin to germinate and develop hypha almost as soon as put in 

 water, I began to wonder if it might not be possible to grow this E^npusa on the 

 same culture media that I had been growing the South African Mucor, although 

 I was well aware that Empusa is considered as an obligatory parasite, parasitic 

 upon certain species of insects. 



Nevertheless, hyphal bodies were removed from both grasshoppers and cater- 

 pillars and cultures made from them in " poured plates " of bouillon-agar and by 

 " planting " them in plates of agar by means of an inoculating needle. It was 

 found to be a difficult matter to obtain pure cultures by either method. The 

 hyphal bodies do not separate readily ; they were already contaminated when 

 removed from the body cavity with the filaments of other fungi and bacteria. 

 The contaminations were removed in part by washing. The amount of contam- 

 ination varied considerably in different insects ; those that had recently died of 

 course containing much less than those that had been dead for some time. 



