I had the satisfaction of exhibiting several times to Mr. Reade, the upper and lower beads of the test-scales he brought with him, and we were both greatly pleased with the beautiful, I might say exquisite, colouring of the rows of beads or rouleaus, according as they were in the upper or lower plane. Not less remarkable were the prominent and enlarged appearance of every fourth or fifth bead of a vivid azure blue: a phenomenon capable, no doubt, of good physical demonstration.

The azure blue scales described in the paper are similar in structure to the Macrotoma Major; but very finely marked, more delicate and transparent.

Altogether this new test promises a new field of research.

IV.—Cultivation, &c., of Microscopic Fungi.
By R. L. MADDOX, M.D.

PLATE XXXVIII.

THE cultivation of Microscopic Fungi offering such a wide and tempting field for research, and the few experiments conducted in reference to "Mucor Mucedo," as stated in a previous article, p. 140, vol. ii. of this Journal, being incomplete, others were set forward without delay; but unfortunately, from unavoidable absence from home shortly after, they must for the present be entirely passed

EXPLANATION OF PLATE.

FIG. 1.-a. a. Spores or conidia of Oidium Tuckerii.
b. The same, germinating.

c. Pedicels with fruit.

d. Older stalks, rough on the outside.
e. Small nucleated spores and bacteria.

2-f. Spores or conidia from brand on orange leaf.
g, h. The same, germinating.

i. Stalk with spores-one-rough on the outside.

j. Small spores nucleated.

k. Part of an old mycelial thread, with minute bodies
enclosed between the septa.

3.-1. Spore or conidia from brand on leaf of a climbing plant.
m. The same germinating and producing irregular cells.
n. Terminal cell supposed to have discharged the minute
bodies.

o. Moniliform rows of Penicillium glaucum.

p. Mycelium from the brand on the leaf.

4.-Figures of the bodies now found (Dec. 13) in the two large drops

of juice alluded to in pp. 144, 145, vol. ii., of this Journal, representing the true spores, the small schizonematous bodies and minute granules or molecules, and which show them to appear at least as fungoidal elements.

5. Cultivating slide.

over, being too imperfect for publication, and others only slightly sketched.

As the subject is likely to occupy considerable attention at home and abroad, and different observers employ different plans, I venture to shortly notice the method adopted by myself, which proved useful and inexpensive, and allude more particularly to that employed by Drs. Billings and Curtis, as stated in their portion of the late "Report on the Cattle Plague in America."

The great difficulty, if not almost absolute impossibility, of conducting any series of experiments, which, however free to legitimate deduction, shall not be open to useless controversy, determined me to use only such means as might be called simply precautionary, of easy arrangement and manipulation, one object being to determine some useful form of cultivating-slide with the ordinary 3-inch microscopic object-slide. Several plans were tried, but the one now described selected. It was made as follows, to be used with thin ths of an inch square covering-glass, such as is usually employed for a th-objective.

A piece of tin-foil, of the stoutness of ordinary note-paper, was cut into squares of one inch diameter, a portion was then removed from a square by cutting it from one side almost to the opposite, then at right angles and again at right angles, thus leaving a strip of the shape of the letter with a flat base, and limbs of equal length and width; from the piece removed a narrow slip was cut from three sides, and then a portion removed from it, as in the first piece, thus forming a smaller letter ; the largest was. cemented to the surface of a well-cleaned slide with easily-melting marine glue, or Bell's cement thickened with gum-shellac; the smaller one was then similarly fixed within the larger one, the limbs being turned to opposite sides of the slide, as seen in the figure, thus affording a central space for the material under examination and a narrow channel each side for the admission of air.

The fungi or spores selected were placed on the cover, under the dissecting or erecting microscope, with a droplet of the medium to be employed or cultivating solution. This was examined under the microscope with a power of 150 or 200 diameters, and if satisfactory, placed at once over the central space of the slide (which, after cleaning with liquor potassæ water and alcohol, if cemented with marine glue, or diluted alcohol if with Bell's cement, was kept turned upside down, one edge resting on a piece of glass until wanted), and the edges of the thin cover resting on the slip of tinfoil, were then closed up with wax softened by oil or with Bell's thin cement, except at the spots corresponding to the two narrow passages marked ** in the figure. If thought necessary, a duplicate slide was similarly prepared, one being left in diffused light, the

other in darkness in the moist chamber.

Journal,

The medium should not touch the inner edges of the tin-foil at any part.

To procure a satisfactory moist chamber occupying but little space, the slides were set in small porous battery cells, previously thoroughly cleansed and moistened with freshly-boiled water, and set in a basin of the same to the depth of half an inch, the basin, with levelled edges, being covered by a plate of clean glass. Those slides intended for diffused light were placed in cleaned damp white porous cells set in water as the others, and covered with glass, the face or cover side of the slide leaning towards the inner surface of the battery-cell; a small cell would thus hold four slides occupying little space. The external temperature, if not sufficiently high, must be raised artificially by a water-bath, or the basin set in a warm place.

The slides when removed are bedewed with moisture; but if rested on end, in a clean tumbler (warm if necessary), and covered with a bell-glass, this soon disappears, and permits examination with any power up to ath.

Although the slide and cover were generally fogged with moisture, I sometimes found the droplet had considerably diminished, and the slide presented surfaces barely moist, especially if cover and slide had not been most carefully cleaned. To meet this difficulty, which was somewhat serious, especially if the slide had to be watched many days, it occurred to me that if an artificial cultivating fluid was made, which would be hygrometric, besides containing the elements of nourishment considered useful for the colour and growth of the fungi, much of this difficulty might be overcome; hence, after several trials, the following was selected and successfully used:

Dextrine, 2 grains; phosphate soda and ammonia, 2 grains; saturated solution of acetate potash, 12 drops; grape sugar, 16 grains; freshly-distilled water, 1 ounce; boiled in a clean glass vessel (thin beaker or large test-tube) for 15 minutes, covered whilst boiling and cooling; when settled poured into perfectly clean two-drachm stoppered bottles, and set aside for use. Sometimes with the cultivating fluid other media were added on the slide.

To preserve some of the perfect forms of fungi found on plants, or produced by cultivation, &c., a saturated solution of acetate of potash was employed with success. There is, however, a little difficulty often from the repellent nature of the heads when beset with spores, and retaining air in their interstices, in using this medium for mounting; but flooding the specimen momentarily with alcohol diluted to the point when it readily touches or wets all the surfaces, and draining it from the slide before applying the mounting solution, readily overcomes this little trouble, and with scarcely any appreciable change in the appearance of the spores or mycelium; at least such is my experience. I prefer it to glycerine or any other solution

used. The edges of the cover, if wetted by the acetate solution, attract moisture readily; so it is best, if the specimen be worth closing up, to dry the edges by a sable pencil and twist of tissue-paper before applying Bell's cement, at first thin, then thickened, as aforementioned.

On the 1st September, a few of the spores (conidia or sporanges) were removed from the surface of a grape, covered with Oidium Tuckerii, and sown as described in the cultivating solution, placed in the porous cell in the moist chamber, and left in diffused light. On the 4th there was an abundant mycelium from several of the spores or conidia (Fig. 1b), afterwards becoming of a light-brown colour, divided by septa, enclosing generally two oil globules or minute spores. Later, on the 15th, from the mycelium several stalks had sprouted, and fruited into the air-space beyond the edge of the liquid; the fruit resembling Penicillium, but the spores in rows on the pedicels were slightly oval. Some of the older stalks had a character which I had not noticed before, and which I have endeavoured to show in Fig. 1 d. They appeared rough outside, or covered with most minute bodies. I am of opinion that this was not an accidental character, but from absence was unable to trace this point further, or whether such stalks might not break up into other bodies.

Lying about amongst the threads were numerous minute bodies (?)bacteria, also small oval bodies with a central spot or nucleus, mostly collected into groups (Fig. 1e). It is possible these may have been drawn in from some slight shrinking of the fluid, and have originated, as supposed, from the heads that fruited in the airspace; but whence they were derived I am uncertain.

On August 30th, a small speck from a brand on an orangeleaf (from the same greenhouse), which for trial had been set in fresh gooseberry juice, was removed, as it had not altered, and set in the cultivating solution, &c., in the light. The spores or conidia commenced sprouting very shortly after, and soon became filled with oil globules or spores, as in Fig. 2g, h. On the 5th many of the mycelium threads had formed heads outside the fluid, the spores being very slightly oval (Fig. 2).

In the fluid lying by the side of some of the original conidia or spores which had sprouted were minute oval bodies with a central nucleus in each (Fig. 2). On the 15th the stems of the older fruited threads appeared roughened on the exterior, with minute bodies, as in the former case. The young threads were growing beautifully, and filled with closely-packed oil globules or spores, no septa being distinguishable, while in the older mycelial threads, which were of a brownish colour, septa existed; each division inclosing about six small bodies (Fig. 2).

Another slide was set from a black brand (found on a leaf of a climbing plant, * in the same greenhouse) with the culti

VOL. III.

C

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vating solution on the 1st September; on the 5th some of the conidia or sporanges showed in their interior four distinct spores (Fig. 31); some of the loose spores gradually sprouted, forming threads consisting of irregularly-shaped cells, many of them containing from one to four, either oil globules or spores (Fig. 3 m); these, like the former, were not tested with ether, &c., as it was desired to preserve the specimen intact. On the 15th the threads were full of these, and along their edges thousands of bacteria-like bodies were present. I am doubtful if these were not ejected from the sporiferous (?) ends of some of the offshoots, as in Fig. 3n. Though I had not witnessed their ejection, their position and the appearance of the terminal cells in some parts led to this supposition. The heads with spores in the air-space resembled Penicillium glaucum with its moniliform chain of round spores, and were larger than in the other specimens (Fig. 3 o). The mycelium from the same brand is seen in Fig. 3p. At this interesting point of inquiry they were obliged to be neglected, and are only alluded to here in this imperfect sketch, rather to show the utility of the cultivating slide and solution than otherwise; for in the two last experiments it would be difficult to prove more than that the fruit sprang from some particular spore of those found in the brand, which possibly might be of a mixed character.

In the very valuable and suggestive report of experiments by Drs. Billings and Curtis, of the U.S. army, appended to the comprehensive and extended researches furnished by Prof. John Gamgee, M.D., on the cattle plague in the United States, under the various titles of "lung disease, or pleuro-pneumonia," "the ill effects of smutty corn on cattle," and "the Texan, periodic or splenic fever," undertaken by the authority of the commissioners of agriculture, we find the microscopic examination and the cultivating experiments handled in a very careful, instructive, and trustworthy manner. The omission of any experiments carried out in darkness in reference to the cultivation of fungi from the healthy or pathological tissues or fluids, more especially those relating to the most serious and difficult question of the cause of Texan or splenic fever, which destroys life so largely, is to be regretted, as it leaves this point open for future investigation.

Experiments performed outside the body, if darkness be omitted as one of the conditions, however carefully temperature and moisture may have been regarded, may not sufficiently closely imitate the natural relations, and determine a difference as regards the results obtained. This is not to say that such differences do exist, nor is it to be expected that all the circumstances can be rendered similar, hence there may remain always some degree of questioning, but this condition adopted would tend to narrow the circle. My object is not to enter into a review of this most interesting and valuable