104 



body of the fly may depend to some extent on the period of 

 time elapsing between their development into the final stage 

 and their escape or removal from the proboscis. 



Observations on the worm embryos suggest that these 

 normally, do not leave the egg-membrane, and their role is a 

 passive one. 



Animal Experimentation. 



Preliminary experiments carried out in 1916 and 1917 

 with the final larval stage of both Habronema musca and 

 H. rruegastoma had proved somewhat disappointing. The 

 object of the experiment was to determine if possible which 

 species of larva was responsible for the production of the 

 granulomata. As the escape of the larvae from the proboscis 

 of the fly appeared to be largely a matter of chance, it was 

 decided to inoculate the larvae into the subcutaneous tissues 

 of an animal. The larvae were obtained by dissection of the 

 heads and proboscides of flies, and placed in a sealed pipette 

 held vertically and filled with normal saline solution. When 

 the larvae had gravitated to the end of the column of saline 

 at the capillary end of the pipette the saline was removed, 

 except for a small drop which contained the larvae. An 

 incision was made through the skin of a horse and the larvae 

 inoculated into the subcutaneous tissues. In each case a very 

 small granuloma resulted, which on microscopic examination 

 showed an infiltration of the tissues with eosinophiles, some 

 hyperplasia of the fixed cells, and the formation of multi- 

 nucleated cells. No necrotic areas were produced. 



It was conceivable that keeping the larvae in saline 

 solution for one or two hours before inoculation had rendered 

 them more vulnerable to the activity of the tissue cells and 

 fluids, and that their more rapid destruction in the body had 

 prevented the occurrence of necrosis. In later experiments 

 larvae were either allowed to escape from the proboscis directly 

 into the tissues of experimental animals, or a mixture of equal 

 parts of normal serum and saline was used as a medium of 

 inoculation. The larvae were only allowed to remain in this 

 fluid for about thirty minutes before being used. 



The preliminary experiments made in the latter part of 

 1916 and the early part of 1917 were seven in number, and 

 the animal used was the one designated as pony in the later 

 experiments outlined below. These preliminary experiments 

 were made as follows : — 



Experiments with Habronema muscae : — 



(1) Embryos of H . muscae were placed in moist sawdust and 



the mass applied to the shaved skin of a horse. 



(2) Six larvae from the proboscis of a fly were placed on the 



shaved skin of the animal, the site being moistened 

 with saline solution. 



