Waterhouse. — Studies in the Physiology of Parasitism. VII. 559 
The infected shoots were then enclosed in glass or mica cylinders plugged 
with cotton-wool. Better germinations and more abundant infections were 
obtained in this way than by sowing teleutospores on the leaves, although 
infections were obtained by the latter means. Attempts to get infection of 
mature leaves invariably failed. 
Small pieces of the infected leaves were fixed at intervals in acetic 
alcohol (absolute alcohol three parts, glacial acetic acid one part), and in 
Flemming’s strong solution diluted with an equal volume of water. Both 
gave good results. The material was embedded in the usual way and was 
cut into sections 4 n thick. In the clearing process, satisfactory results 
were obtained from the cedar-wood oil method, in which the dehydrated 
material was placed in absolute alcohol floating on cedar-wood oil, whence 
it gradually passed into the latter liquid under the action of gravity. 
For demonstrating the early stages of infection the best details were 
given by iron-alum-haematoxylin followed by Sudan III. Such sections 
mounted in glycerin jelly showed the cuticle very sharply stained. Scharlach 
red and Congo red in place of the Sudan III gave almost as good results. 
Sections stained with Congo red alone and mounted in euparal also showed 
up well. For the latter stages of infection, the safranin-gentian violet- 
orange G combination gave the best results. 
Observations. 
Germinations on the slide . Germination of the teleutospores was 
rapid. Promycelia were to be seen after 5 to 6 hours and sporidia were 
obtained on the slide after 12 to 15 hours. These at once germinated by 
pushing out a germ tube (Fig. 1) ; sometimes two (Fig. 2) or even three 
(Fig. 3) such tubes were produced. It was early found that the germinating 
sporidia adhered readily to the slide and could be stained, washed, & c., 
without loss. The adhesion is due to the fact that the germ tubes possess 
a mucilaginous sheath similar to that of Botrytis cinerea. This stains 
faintly with dilute aqueous gentian violet. At the edges of the sheath, at 
a distance from the main wall of the germ tube, aggregations of particles 
further emphasize the presence of the sheath (Figs. 1, 2, and 3). The germ 
tube may continue to elongate, and becomes septate ; in one case it attained 
a length of over 1 mm. Frequently, after elongating slightly, the germ 
tube swells into a vesicle (Figs. 4 and 5). In some cases the vesicle was 
large and in contact with the glass slide and was therefore of the nature of an 
appressorium ; around such an appressorium a mucilaginous investment 
may or may not be observed (Fig. 5). From the vesicle may arise a second 
germ tube (Fig. 6). It was generally found that the production of long 
germ tubes was most frequent when the sporidia were in water. In the 
nutrient media the production of vesicles was more common, and in the 
barberry extract these vesicles were often very irregular in shape. 
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