66 Wager and Penis ton.—Cytological Observations on 
which have been kept in a quiescent condition until volutin has disappeared 
from most of the cells (about three weeks), that in some cells a large quantity 
is still present. Such cells, however, present the well-known characteristic 
microscopic appearance of disorganized cells. Further, in preparations of 
such yeast we find a number of granules which give the volutin reaction 
with methylene blue lying outside the cells. It seems probable that these 
are volutin granules which have escaped from disorganized cells. 
Under active metabolic conditions volutin disappears and reappears 
with remarkable ease, and for this reason is extremely difficult of observa¬ 
tion. If the temperature is lowered so as to impede but not stop 
fermentation, when a large quantity of volutin is present, it may disappear 
completely from every cell within 1-5 hours and reappear in the same time 
when the original activity is resumed. 
When the volutin granules first make their appearance they are 
extremely small, and occur in the immediate neighbourhood of the 
chromatin network, but on the outside (PI. X). Each granule appears to 
originate independently, and in the early stages of their formation it is 
common to see several minute granules at remote intervals immediately 
outside the chromatin network. It is significant that in these early stages 
they are never found in the vicinity of the nucleolus. 
As soon as the granules become numerous they tend to aggregate, as 
Guilliermond points out, and apparently fuse to form large granules, often 
forming a single granule of enormous size (Figs. 171-3). They may then 
take up a position in the neighbourhood of the nucleolus, and it is common 
to find a number of comparatively large granules or a single enormous 
granule resting on the nucleolus, frequently completely obscuring it (Figs. 
171, 180). Sometimes they increase in number without aggregating, and 
remain round the outside of the chromatic network, where they often 
exhibit a very regular arrangement, resembling a reticulum (Fig. 175), 
especially when stained in haemalum, and occasionally in methylene blue 
preparations. It seems possible that this is what Hieronymus saw, and 
regarded as similar to a structure described by him for the Cyanophyceae. 
(See Wager, ’ 98 , p. 503.) 
When the volutin granules begin to disappear, they first break up into 
smaller granules, and then, as Guilliermond states, there can be no doubt 
that they become dissolved in the cytoplasm, which becomes intensely 
stained with methylene blue just at this stage. It is interesting to note 
in this connexion, that in preparations stained with methylene blue, or the 
fuchsin-methyl green mixture, or similar combinations, these granules are 
often seen, each surrounded by a less deeply stained area in the cyto¬ 
plasm, which gives them the appearance of being contained within small 
vacuoles. 
It may here be pointed out that the intense reaction of the cytoplasm 
