Villous: Beset with short, undivided, hair-like extensions (Fig. 123, d). Plumose: A delicate feathery growth. Arborescent: Branched or tree-like, beset with branched hair-like extensions (Fig. 123, e). Fig. 124. Stab cultivations-types of growth: f, Crateriform; g, saccate; h, infundibuliform; j, napiform; k, fusiform; /, stratiform. (C) Area of Liquefaction (if present).— Crateriform: A saucer-shaped liquefaction of the gelatine (Fig. 124, f). Saccate: Shape of an elongated sack, tubular, cylindrical (Fig. 124, 9). Infundibuliform: Shape of a funnel, conical (Fig. 124, h). Napiform: Shape of a turnip (Fig. 124, j). Fusiform: Outline of a parsnip, narrow at either end, broadest below the surface (Fig. 124, k). Stratiform: Liquefaction extending to the walls of the tube and downwards horizontally (Fig. 124, 1). (D) Character of the Liquefied Gelatine. 1. Pellicle on surface. 2. Uniformly turbid. 3. Granular. 4. Mainly clear, but containing flocculi. 1. Presence or absence of liquefaction. 3. Bulk of growth at the surface-aerobic. 4. Bulk of growth in depths—anaerobic. Fluid Media. 1. Surface of the Liquid. Presence or absence of froth due to gas bubbles. Presence or absence of pellicle formation. Character of pellicle. 2. Body of the Liquid. Uniformly turbid. Flocculi in suspension. Granules in suspension. Clear, with precipitate at bottom of tube. Colouration of fluid, presence or absence of. 3. Precipitate.— Character. Amount. Colour. Litmus Milk Cultivations. Unaltered. 1. Reaction: Acid. . Alkaline. 2. Odour. 3. Formation of gas. Unaltered. 4. Consistency: Digested (? character of solution). 5. Clot: Character Coagulated. hard: solid. soft: flocculent. (a) Coagulum undissolved. (b) Coagulum finally digested, completely: incompletely. 6. Whey: Resulting solution, clear: turbid. Abundant. Scanty. Clear. BY MICROSCOPICAL METHODS. Preparations must be made from the cultivations at intervals of, say, twenty-four hours, during the period they are under observation, and examined— (A) Living.-1. In hanging drop, to determine motility or non-motility. In this connection it must be remembered that under certain conditions as to environment (e. g., cold, heat, light, unsuitable medium, etc.) motile bacilli may fail to exhibit activity. No organism, therefore, should be recorded as non-motile from one observation only; a series of observations at different ages and under varying conditions should form the basis of an opinion as to the absence of true locomotion. Size. In the case of non-motile or sluggishly motile organisms, endeavour to measure several individuals in each hanging drop by means of the eyepiece micrometer, and average the results. If the organism is one which forms spores, ob serve (a) Spore Formation.-Prepare hanging-drop cultivations (vide page 69) from vegetative forms of the organism, adding a trace of magenta solution (0.5 per cent.) to the drop, on the point of the platinum needle, to facilitate the observation of the phenomenon by rendering the bacilli more distinct. Place the preparation on the stage of the microscope; if necessary, using a warm stage. Arrange illumination, etc., and select a solitary bacillus for observation, by the help of the -inch lens. Substitute the -inch oil-immersion lens for the sixth, and observe the formation of the spore; if possible, measure any alteration in size which may occur by means of the Ramsden micrometer. (b) Spore Germination. In a similar manner prepare hanging-drop cultivations from old cultivations in which no living vegetative forms are present, and observe the process of germination. The comfort of the microscopist is largely enhanced in those cases where the period of observation is at all lengthy, by the use of some form of eye screen before the unemployed eye, such as is figured on page 59 (Fig. 41). If it is impossible to carry out the method suggested above, proceed as follows: (a) Spore Formation.-Plant the organism in broth and incubate under optimum conditions. At regular intervals, say every thirty minutes, remove a loopful of the cultivation and prepare a coverslip film preparation. Fix, while still wet, in the corrosive sublimate fixing solution. Stain with aniline gentian violet, and partially decolourise with 2 per cent. acetic acid. Mount and number consecutively. (b) Spore Germination.-Expose a thick emulsion of the spores to a temperature of 80° C. for ten minutes in the differential steriliser (vide page 202). Transfer the emulsion to a tube of sterile nutrient 'broth and incubate. Remove specimens from the tube culture at intervals of, say, five minutes. Fix, stain, etc., as under (a), and examine. (a) Prepare cover-slip film preparations at the various ages, and fix by exposure to a temperature of 115° C. for twenty minutes (vide page 75). (b) Stain the preparations by Gram's method (if applicable) or with dilute carbol-fuchsin, and mount in the usual way. (c) Measure (vide page 63) some twenty-five individuals in each film by means of the Ramsden's or the stage micrometer and average the result. Pleomorphism: If noted, record The predominant character of the variant forms. On what medium or media they are observed. (b) To demonstrate details of structure: Flagella: If noted, record Method of staining (vide page 87). Position and arrangement (vide page 115). Number. Spores: If noted, record Method of staining. Shape. Size. Position within the parent cell. Condition, as to shape, of the parent cell (vide page 74). On what medium they are best observed.. Age of medium. |