Rayner.—Obligate Symbiosis in Calluna vulgaris. 119 
the indirect results of invasion of leaves by parasitic Fungi ( 43 , 44 ). The 
discussion of hypotheses to account for the storage of a calcium salt in the 
plant tissues may profitably be postponed until more data are available as to 
the nutritive preferences of the Fungus. 
The Stem. The presence of mycelium on the outside of the young 
stem and the penetration of branches from it into the underlying tissues 
have already been noted (p. 116). Evidence of the presence of mycelium 
may be found in most tissues of the stem, e. g. in the parenchyma of the 
cortex, and in that associated with the vascular tissues ; the cells of the 
pith, also, probably contain hyphae in a vestigial condition. 
The tissues of the pith and of the cortex likewise contain quantities 
of calcium oxalate in the form of large crystals and crystal-aggregates. 
Untreated sections are often rendered absolutely opaque by the presence in 
every cell of these crystals. In rare cases small fragments of hyphae which 
stain deeply with cotton blue or aniline blue can be identified in the neigh¬ 
bourhood of crystals; more frequently the hyphae are present in an 
extremely attenuated condition, are very transparent, refuse to take up 
the characteristic stains, and can only be identified with difficulty. 
The most satisfactory method of obtaining conclusive proof of the 
presence of mycelium in such tissues is by slow maceration in sulphuric 
acid, or by prolonged treatment with clearing agents, such as chloral 
hydrate, eau de Javelle, or potash, before staining. 
4. The Isolation of the Fungus. 
During the past three years repeated and fruitless attempts have been 
made to isolate the mycorrhizal Fungus from the root-cells. 
The method first adopted was to remove clean transparent young roots 
from the outside of a ball of soil surrounding the roots of a pot plant. 
These roots were washed in fast-running water for twenty-four hours, rinsed 
repeatedly in sterilized distilled water, after which small pieces were planted 
out on agar plates. (Direct sterilization of the surface of the root, by heat 
or momentary immersion in weak solutions of mercuric chloride, was , 
found to be impracticable.) 
Using this method, a number of fungal species were isolated, none 
of which, however, satisfied the test of successful inoculation into sterile 
seedlings. 
Among those constantly found were species of Cladospormm , Peni - 
cillium , Citromyces^ Fusarium , and Alternaria , all of which are apparently 
constant or fairly constant members of the epiphytic microflora of the roots. 
A species of Cladosporium was invariably present in my cultures, and is 
always dominant if unsterilized seedlings are planted in agar. 
The hanging-drop method of culture was tried with the same lack 
of success. Pieces of washed root were transferred to hanging drops of 
sterile water, and to similar drops of various solutions, which it was 
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