WITH NOTES ON OTHER SPECIES. 97 



that the zoospores and oospores of the Saprolegniaceon are carried with the Algae to 

 a large extent. If it is desirable to avoid any possible infection from other sources 

 than the mass of Algse concerned, the water may be filtered, heated to boiling, and 

 then cooled, before the specimen is placed in it. DeBary found that, in practice, the 

 water supply of Strassburg never produced any of these fungi in cultures made with 

 water from its pipes alone ; and I have had the same experience in repeated trials 

 with that of Amherst. But water from the Cambridge pipes, and doubtless that from 

 others, will yield them at certain seasons, at least. The insects used may be freshly 

 killed, and their chitinous covering should be broken as little as possible ; but I have 

 found that, for winter cultures when fresh insects are not readily available, an excel- 

 lent substitute may be found in dead house-flies, collected in the fall and kept dry 

 and exposed to the air, but protected from dust. Since the dry surfaces of insects 

 are not readily wetted by water, it has proved useful to moisten them, whether fresh 

 or dried, with alcohol, and then to soak them in water for a few minutes to remove the 

 alcohol. They will then, when thrown into the culture vessel, sink until their bodies 

 are mostly below the surface and so present a much larger area to the swimming 

 zoospores of SaprolegniaceoR than if dry and floating largely above the surface. 



Since the zoospores depend for their activity on a sufficient supply of oxygen 

 we may expect them to be most abundant near the surface of the water, and since 

 they are chemotactic, being strongly attracted by nutrient substances, they must 

 readily reach the floating insects and germinate upon their bodies. An average time 

 for the appearance of the young hyphse is perhaps two days from the beginning of 

 the culture, but one day is ample time, as a rule? for the zoospores to have effected an 

 attachment to the substratum. The insects should now be transferred to a vessel of 

 fresh, clean water, and here the development of the fungus may be followed. The 

 water should be carefully changed daily or less often, as may be required, until the 

 maximum of vegetative activity is past. For superficial examination, the whole insect 

 with attached fungi may be floated upon a slide. For more thorough study, parts seen 

 by this preliminary method to be in the desired condition may be cut off" and mounted 

 under a cover, or used for a hanging drop culture. Rothert ('88) has pointed out that 

 well-grown filaments with reproductive organs continue to develop normally after 

 being cut off", until their protoplasm is exhausted. 



It is not easy, although it is usually possible, to obtain from a mixed culture of 

 several species, pure cultures of each. This may be accomplished by using sterilized 

 water, fresh, clean insects, well-soaked in alcohol and distilled water, and a very small 

 quantity of the fungus, preferably zoospores from a single sporangium. A few 

 attempts will give the desired result, if the first does not. The use of small portions 



