Whereas formerly only alcohol-forming yeast fungi were understood under the denomination "yeast," this term was extended later to include all budding fungi. After it became known that Ustilago, the smut found on grain, forms budding cells, it was asserted that the yeast cells of Ustilago, and the alcoholic fermentation fungi of practice, were morphologically identical. It was again inferred from this that yeast is derived from Ustilago. Although this opinion was never proved, it was yet strong enough to cause confusion of ideas. Even now the word " yeast" is frequently used in text-books to designate not only true saccharomycetes, but also all budding fungi.

There exists at the present time, meanwhile, no proof at all that saccharomycetes are a stage of development of other fungi. On the contrary, we must regard them as ascomycetes equally as independent as, e.g., the exoasceæ, the independence of which no one has doubted, and to which group they are closely connected, appearing in the same. forms (budding cells, endospores and mycelium) as the exoasceæ, and in these forms only.

2.-Structure and Shape of Yeast Cells.

A Saccharomyces growth always consists of budding cells, under certain conditions also of asci with endospores and of mycelium.

The budding cell consists of a membrane enclosing a mass of protoplasm in which there is a cell nucleus.

The Cell Contents. The cell nucleus, the existence of which was proved by Schmitz in 1879, is, as a rule, a spherical body; Hansen has, however, in some cases observed flat nuclei. In each yeast cell there is only one nucleus. To see this it is usually necessary to fix, harden and stain the preparation (see p. 89). Only in exceptional can they be seen without this preliminary treat

cases

ment. The cell nucleus has been studied in recent times by Jannsens, Leblanc and Wager. Essential points with regard to its structure and function have not yet been cleared up.

According to Jannsens and Leblanc the protoplasm of the yeast cell forms a fine network; during fermentation its appearance changes: vacuoles appear, the protoplasm becomes granular, and fat and oil particles may be formed. By vacuoles are understood cavities in the cells, which are filled with cell sap. A highly refractive body, the vacuolegranule, is usually seen in vacuoles, which is in constant motion, the so-called Brownian molecular movement. The number of these granules is usually one to three; only in exceptional cases are there more than three or none at all. According to Küster these bodies are decomposition products derived from the plasma; they form a plastic, semi-fluid mass. Using a weak aqueous solution of neutral red (1 to 5,000 or 1 to 10,000) they become intensely red in a few minutes, if the material is suitable, all other parts of the cell remaining colourless. Vacuole-granules coloured in this manner give up the colouring matter in a sugar solution, the latter gradually becoming red.

In the cell protoplasm may often be seen numerous granules usually of angular shape. They are easily stained by aniline dyes, especially methyl green (Casagrandi). They are soluble in alcohol, ether, chloroform, chloroform and ether, caustic potash, caustic soda, petroleum ether, etc.; sometimes they require several days to dissolve. According to Will they are of a fatty nature; he has further observed that when the oily substance had been removed by treatment with absolute alcohol, small faint bubbles appeared in the place of the granules, in the interior of which a network could be seen.

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If absolute alcohol is added to a preparation the cells be seen to shrink up, and are soon killed. In general

dead cells are distinguished from living ones, as we saw in Section II., by the greater ease with which they take up colouring matter.

The Cell Wall and its Gelatinous Formation. The membrane of the yeast cell is very thin in young individuals;

FIG. 63.-Gelatinous network. I., Network formation and yeast cells which are stained with methyl violet; most of the cells have disappeared; some (on the right) still lie in the meshes. II., It may be seen in a, b and d that the network forms complete walls; there are in a 3 cells, in b 1, and in c 2 cells lying in the meshes. (From Hansen's original drawing.)

in old cells, and in such as live under unfavourable conditions of nutrition, it may become tolerably thick, e.g., several micromillimetres. It can be shown distinctly by means of various reagents (e.g., dilute acid and alkalis). According to Will's and Casagrandi's experiments the membrane con

sists of two or more layers; this may be shown by treating the cells for a long time (days to weeks) with a 1 per cent. solution of osmic acid. The membrane dissolves easily in concentrated chromic acid, more slowly in concentrated sulphuric acid (Casagrandi).

The membrane of the cell gives off, under certain conditions, a mucilage which takes part in the formation of the gelatinous network described by Hansen (Fig. 63). After hardening an ordinary microscope preparation it shows itself in the form of strands and plates between which the cells are enclosed. The granulations originally present between the cells may be taken up into the substance of the network, which may be stained by this means. This formation may be readily obtained if a lump of thick yeast, as it usually occurs in breweries, is placed in a glass, covered, and put away for a short time until partial drying takes place. It usually occurs also in spore cultures on gypsum blocks, on gelatine and in yeast ring formations.

Will considers the network, which appears, e.g., during the drying up of beer yeast, to be different from that which occurs in film formations and in the yeast ring. In his opinion a gelatinisation of the cell membrane possibly takes part in bringing about the former, the albumen mixed up with the yeast, however, playing the chief part. The networks formed in the film and in the yeast ring may also, according to him, be of different constitution, since some forms are found which give the albumen reaction and some which do not. It is very difficult to distinguish here what, in the network formation, originates in the cell wall itself, what in the cell contents, and what in the surrounding medium; this is perhaps a problem which does not as yet admit of a solution.

Shape of the Cells.-Budding cells occur especially in or upon nutrient liquids, but are also found on solid substrata.

Their shape is exceedingly varied; they are spherical, eggshaped, oval, sausage-shaped, filament, dumb-bell, and lemonshaped, etc., besides occurring now and then in cultures in quite irregular and abnormal forms. It is a noteworthy fact that no definite cell shape is absolutely peculiar to one species; it is true that the majority of the cells of a species, under certain conditions of culture, occur in a certain shape; but if these conditions are altered, the shape will alter also. A Saccharomyces species can seldom or never be determined by microscopic observation alone. Hansen clearly demonstrated this multiplicity of form of the species and the defectiveness of the classification which had been employed before his time. Simultaneously he showed that, under definite conditions of culture, the shape of the cells affords a good group-characteristic for species. (Compare p. 233, “ Variation ".)

3.-Modes of Propagation.

Budding. Vegetative increase (the formation of new vegetative cells) takes place in all true saccharomycetes by means of budding. This takes place by a small outgrowth forming on the mother cell (Fig. 64), and gradually increasing in size.

The new cell may separate from the mother cell, or may remain connected with it; in the latter case large colonies of buds are formed (Fig. 64). In the genus Schizosaccharomyces vegetative increase takes place not by budding, but by splitting off. Near the middle of the mother cell is formed. a septum which splits up and thus sets the new cell free.

The budding of a single cell was studied by Mitscherlich in 1843; he distributed a little yeast in beer wort, so as to obtain one or two cells in an ordinary sealed up preparation, and gave a drawing of the various generations (Fig. 64). Under these circumstances thirteen hours passed