Vegetative Mycelium of Saprolegnia. 65 
it was found that in the case of the vegetative hyphae many sections were 
lost through imperfect adhesion to the slide. A much better idea of the 
state of the hyphae was obtained by mounting them whole, and no difficulty 
was found in examining them. Three methods of staining were used : 
Flemming’s triple stain, iron-haematoxylin, and Delafield’s haematoxylin, 
though it was found necessary to alter the usual times recommended for the 
first of these. The slides were removed from water to a saturated solution 
of aniline-water-safranin, where they were left for three to six hours. They 
were then treated thus: water three minutes, gentian violet (aqueous) 
thirty minutes, water one minute, orange G (aqueous) one minute, 90 per 
cent, alcohol one minute, 100 per cent, alcohol one minute, clove oil five 
minutes, benzol, Canada balsam. Although the process of taking the 
material from water to 90 per cent, alcohol may seem very drastic, in only 
a few cases did shrinkage of the cytoplasm take place to any appreciable 
extent, and, since the stain was only used for the nuclear figures, this did 
not matter. Xylol as a clearing agent seemed to cause too great a shrinkage 
of the cytoplasm, and so benzol was substituted. 
Cytological Observations. 
In the early stages the floating hyphae are slender and contain a mass 
of dense cytoplasm with small vacuoles. They grow rapidly in breadth 
until they have reached the normal thickness, by which time the cytoplasm 
has been reduced to a thin peripheral layer. The intramatrical hyphae, on 
the other hand, are very poor in cytoplasm. 
Protein granules constitute the main portion of the cytoplasm, and show 
very distinctly in stained preparations. In living material they are visible 
under high-power objectives, and seem to be constantly moving, both 
in streaming and in Brownian movement. Cytoplasmic streaming is con¬ 
tinuous, chiefly in the direction of the apex, though slight back currents 
occur. It appears that the tendency is to form zoosporangia, for, when the 
accumulation of cytoplasm at the apex has become sufficiently dense, 
a septum is formed and a zoosporangium cut off. As soon as the cyto¬ 
plasmic continuity is thus broken, fresh cytoplasm accumulates below the 
septum, so that, by the time the zoospores have formed and escaped, 
sufficient Q'toplasm and nuclei have accumulated below the septum to fill 
the empty zoosporangium case completely. All that remains is for the 
septum to grow upwards into the old sporangium and the cytoplasm 
immediately below follows. 
In one instance a count of nuclei was made in a preparation treated with 
Flemming’s stain, in which the zoospores were about to escape from the 
sporangium. The cytoplasm for a distance below, of about the length 
of a sporangium, was practically as dense as that in the zoospores. The 
