(235) The Thread Method. In the thread method capillary glass rods are made by drawing out ordinary glass tubing in the blow-pipe flame, these are tipped with sealing-wax, and to the wax a little bit of sterilized cotton wool is made to adhere. The end of the rod thus prepared is infected with the bacterium by a short immersion in a pure cultivation and then placed in the disinfectant for a definite time. The rod on removal is soaked for a little time in sterilized water until all trace of the disinfectant has been removed. The rod thus charged and purified is next inserted into a mass of solid nutrient gelatin in a test-tube, and put on one side at the ordinary temperature of the atmosphere, protected of course from external contamination by a suitable plug of sterilized wool. Whether the process used is the "drop" or the "thread," in each case "control" experiments are made with threads infected with the bacterium, but which have not been submitted to disinfection. Enumeration of Colonies.-This is an entirely different method of procedure. The number of colonies in a gram of sewage or other suitable liquid are carefully determined by a modification of known methods. The same sewage is then treated by substances the disinfectant properties of which form the subject of inquiry, and the number of colonies capable of growing in a nutrient soil, representing the micro-organisms which have escaped destruction, again enumerated. The only special apparatus used requiring description is the "drop-bottle" and the "rings and plates." The Drop-bottle.-The figure represents its shape and size, the capacity is about 25 c.c. The stopper is hollow and terminates in a pipette; it has a pinhole at a, which can be closed by the finger. a FIG. 47. The Glass Plates and Rings.-The glass plates are 4 by 2 inches square, the rings 4 inches in diameter, inch thick, and inch high. The plates have a ground surface the size of the ring thickness; the rings are cemented to the plates in the following manner. After heating the rings and plates in a hot air oven for many hours a little peptone gelatin is run on to the ribbon of ground surface, the ring adjusted, and the whole allowed to cool in a glass chamber formed by a small dish covered by a slightly larger one; at the bottom of the dish is some filter-paper moistened with a solution of mercuric chloride. The plates are not used until the gelatin cement has perfectly set. The plates are ruled by means of a diamond into squares for the purpose of easy enumeration. Solid substances, such as ferrous sulphate, may be weighed and dissolved in definite quantities of the sewage; in other cases solutions of known strength are mixed with the sewage. The method of cultivation is as follows:-A small quantity, whether of diluted or disinfected sewage, is transferred to the previously cleansed and sterilized drop-bottle, the bottle and its contents carefully weighed, then by means of the pipette stopper one or two drops spotted on to the surface of the glass cell formed by the plate and ring already described; the weight of the drops is ascertained by reweighing the drop-bottle. Ordinary nutrient gelatin liquefied at a gentle heat is run from a Lister flask into the glass cell, and mixed equally with the drops by inclining the plate in different directions. During these several operations dust is excluded as far as possible by covering the glass cell by a second glass plate, merely shifting the plate sufficiently on one side to allow the insertion of the nozzle of the Lister flask or the point of the pipette. The cells thus charged are placed in the moist chamber; the gelatin rapidly sets, and at the end of from three to five days the colonies of growth are counted in the usual way and their general nature determined. The weight of the drop or drops taken may vary from 20 to 100 mgrs., the gelatin in which the drop is cultivated from 15 to 20 grams, so that the minute quantity of disinfectant contained in the drop itself is diluted from 200 to 1000 times. This amount of dilution with the comparatively weak percentages of most disinfectants save those approaching corrosive sublimate in power reduces the action of the disinfectant on the gelatin, the cultivating soil-to a minimum, so that practically as soon as the microorganisms still surviving are floated into the nutrient gelatin they are removed from the sphere of disinfectant influence, the following is an example of this method : Two quantities of sewage were respectively treated with phenol and cresol, so that the mixtures were equivalent to 1.9 per cent. and allowed to act for twenty-four hours; the mean of two strictly concordant experiments gave the following as the number of colonies which at the end of four days could be enumerated No. of colonies per gram of the sewage taken. Phenol The control 33,333 33,410 1,490,000 (236) Experiments of Parsons and Klein. VALUABLE experiments have been made in this country by Drs. Parsons and Klein, as to the details of heat disinfection; the following is condensed from Dr. Parsons's report on the subject. (Supplement, 14th Annual Report, Local Government Board.) The infective materials employed were: (1) Blood of guinea pig dead of anthrax, containing spore free bacillus anthracis. (2) Pure cultivation of b. anthracis in rabbit broth, without spores. (3) Spore holding b. anthracis in gelatin cultivation. (4) Cultivation of bacillus of swine fever (infectious pneumo-enteritis of the pig) in pork broth. (5) Tubercular pus, from an abscess in a guinea pig which had been inoculated with tubercle. Strips of clean flannel were steeped in the respective infective fluids, dried in the air, wrapped separately and loosely in a single layer of thin blotting paper, and suspended in the centre of the apparatus in company with a thermometer, so placed that its bulb was close to the packets of infected material. The results of the disinfection were tested by inoculation of animals. Control inoculations with unheated portions of the same materials were also in all cases made. (237) Experiments with Dry Heat. They found that spores of anthrax lost their vitality after exposure for four hours to a temperature a little over that of boiling water (212°-216° F.); or for one hour at a temperature of 245° F. Non-spore-bearing anthrax, as well as the bacilli of swine fever, were rendered inert by exposure for an hour to a temperature of 212-218°, and even five minutes' exposure to this temperature sufficed to destroy the vitality of the former and impair that of the latter. (238) Experiments with Moist Heat. (a) Boiling Water.-Dr. Klein found that spore holding bacillus anthracis was killed by boiling for so short a space as one minute. (b) Steam at 212°.-The experiments were conclusive that it destroyed all contagia; in one instance only, was there room for doubt, this was in the case of highly resisting anthrax spores exposed to steam for five minutes only. (239) On the Penetration of Heat into Articles Submitted for Disinfection. Experiments were made to ascertain how far the enclosing of infective objects in blotting paper or test-tubes plugged with cotton wool hindered the full access of heat to them. Two similar registering thermometers were taken; the bulb of one was tied up in a single layer of thin white blotting paper, that of the other was placed in a test-tube inch wide, in such a manner as not to touch the sides, and a plug of white cotton wool 1 inch deep was pushed into the tube around the stem of the thermometer, but not as far as the bulb. Both the paper and cotton wool were previously dried. The two thermometers were then suspended in a proper apparatus which need not be here described. It took two and a half hours before the thermometers thus covered attained the same heat as the naked thermometer. In a second experiment in which a thermometer was covered with a single layer of blanket; the blanket-covered thermometer marked 4° lower than the control at the end of two and a half hours. The following tables give results of similar experiments on pillows, and show conclusively how difficult it is to secure the penetration of a dry heat sufficient for disinfection into the interior of such an article as a pillow. It can only be effected by |