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The results of this experiment were that of four checks three died, two in four and five days after the inoculation with the germ, the third in seventeen days, and the fourth check recovered.
Of the three pigs treated with the ptomaine one recovered, two died in five and six days after the inoculation.
Of the two pigs treated with the synthetical compound one died in thirty-nine days after the inoculation and fourteen days after the last check ;the other one recovered entirely. Though this experiment is not conclusive, it certainly indicates that the pigs which had been treated offered considerable resistance to the disease, and that the synthetical compound is more effective than the ptomaine obtained from the culture liquids.
Had a somewhat larger quantity of the ptomaines been used for treatment, and the injections been made in smaller quantities, extending over a longer period of time, it is probable that all the treated animals would have recovered. At any rate we are sufficiently encouraged to continue the experiments. I may take occasion here to mention the valuable assistance rendered me by Dr. V. A. Moore in connection with the bacteriological work, autopsies, etc., and by Dr. Theobald Smith in allowing the use of the facilities of the bacteriological laboratory in his charge, and also the uniform kindly encouragement of Dr. Salmon, Chief of the Bureau.
While awaiting the results of further experiments upon hog cholera it was thought well to begin a study of the swine plague cultures, with the object of obtaining from them albuminoid and alkaloid poisons. The swine plague germ grows but slightly in the ordinary beef infusion culture. Dr. Moore, however, found that if instead of making a simple beef infusion a beef broth was prepared by boiling the meat the growth of the swine plague germ in this liquid was much more abundant. Alkaline media
of this description were therefore used, 1,000 cubic centimeters in Erlenmeyer flasks being inoculated and kept in the incubator for two days at a temperature of 37° C. The growth of the germ was by this time very perceptible. The contents of the flasks proved to be uncontaminated. When opened a disagreeable, pungent odor was noticed.
After filtration about eight times its volume of absolute alcohol was added to the solution, and a considerable amount of a white flocculent precipitate was obtained. This, after settling, was filtered off, redissolved with water and again precipitated with absolute alcohol. The precipitate was thoroughly washed with absolute alcohol and dried over sulphuric acid in vacuo. A white translucent mass was thus obtained, with difficulty soluble in water and having properties of an albuminoid or proteid body. The filtrate from the albumose was neutralized with hydrocholoric acid evaporated to dryness, and the residue extracted with absolute alcohol. This alcohol extract gave alkaloidal reactions with mercuric chloride, phosphomolybdic acid, platinum chloride, etc., showing the presence of a ptomaine. The double platinum salt of this body I have prepared, but have not at this writing been able to make a satisfactory analysis of it. I have demonstrated, however, the existence in the culture liquids of the swine plague germ of a ptomaine and albumose. The name suplagatoxin may be given to the ptomaire (from the Greek evs, a hog, iri, plague, and tozzzóv, poison), and suplagoalbumin to the al
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bumose. This would be distinctive from the hog cholera ptomaines. While purifying a larger quantity of these substances in order to make a closer study of them chemically, I thought it advisable to use the material at hand for making some experiments in the production of immunity in guinea pigs from swine plague by preventive treatment. Previous to this Dr. Moore had made a number of inoculations of guinea pigs with swine plague, which showed that one one-thousandth of a cubic centimeter of beef infusion peptone culture of swine plague one day old, was sufficient to kill a guinea pig in from twenty-four to forty-eight hours. Further, in order to see if the treatment which proved satisfactory for producing immunity against hog cholera might have any effect in retarding the disease of swine plague, two guinea pigs that had been submitted to the preventive treatment for hog cholera, but never exposed by inoculation, were inoculated with one one-thousandth of a cubic centimeter of beef infusion swine plague culture one day old. Both animals succumbed in forty-eight hours to the disease of swine plague. Two guinea pigs that had been subjected to the preventive treatment, then inoculated with hog cholera and recovered and were perfectly well, were inoculated with one one-thousandth of a cubic centimeter each of beef infusion peptone swine plague culture one day old. Both died, as was expected, in forty-eight hours. These experiments serve further to demonstrate, if proof is necessary, that the diseases of hog cholera and swine plague are distinct, and that an animal that has had the hog cholera and recovered is still susceptible as ever to the swine plague.
Experiment XII.-Two guinea pigs were selected, and on three successive days .0030 gram of swine plague albumose was injected subcutaneously in the thigh. About .0010 gram of substance was given at each injection. There was a slight swelling at the point of injection, which disappeared in four or five days and the animal appeared well.
Two checks were now taken and the four inoculated with one one-thousandth of a cubic centimeter swine plague culture. The checks died, one in forty-eight hours and the other in thirty-six hours. The treated pigs appeared a little stupid for a day or two and then recovered entirely.
Experiment XIII.-Two guinea pigs were treated with a solution of the ptomaine extracted from the culture liquids. The injections were made subcutaneously in the inner side of thigh, the quantity of ptomaine used corresponding to about 15 cubic centimeters of the culture medium. There was a slight swelling and soreness at the point of injection, but otherwise the pigs appeared well. These, together with two checks, were inoculated with one one-thousandth of a cubic centimeter of swine plague culture. The checks died of swine plague. One of the treated animals died in thirty-six hours. The autopsy, however, showed but few marked characteristics of swine plague. At the point of inoculation there was a slight infiltration. Blood vessels in heart much injected ; liver slightly reddened. Bladder distended with urine. Otherwise the organs were norinal. Coverglass from spleen and liver showed no swine plague germs, but cultures from the liver showed that the swine plague germ was present. The other treated pig died five days after the checks, or eight days after the inoculation.
The ptomaine, therefore, produced resistance and a large dose would probably give immunity.
These few experiments, following the more extended ones upon hog cholera, prove conclusively that both these diseases can be prevented in guinea pigs by chemical inoculation. The experiments upon swine plague will be extended and a careful study made of the ptomaine and albumose produced by this germ, and their effect upon hogs.
Hankin holds that albumose is the one and principal factor in the production of immunity, and that the reason more results have not been secured in this direction is because the proper material has never been used. We think, however, that the albumose is only an intermediate product of the germs and the final and most fatal effect of the disease results from the ptomaines. At any rate the experiments upon hog cholera lead to the conclusion that while a mixture of the albumose and ptomaine seems to produce greater immunity than either substance alone, nevertheless when used separately they are of about equal value.
In addition to the study in connection with the disease just recorded the writer has given some little attention to the presence of tyrotoxicon in milk. In May a sample of milk from Maryland came into his hands which was supposed to have caused the sickness of a number of children. The symptoms as given indicated a possible tyrotoxicon poisoning. The milk was examined for the poison, but the latter could not be detected. Some months after this some cheese, which had produced sickness in this city, and two lots which had caused illness in Ohio, were received. In all three cases the questionable tyrotoxicon was blamed for the sickness. I could not, however, establish the presence of tyrotoxicon in any instance by the methods prescribed by Vaughan. This led me to repeat one of Vaughan's experiments, which should have given me considerable quantities of tyrotoxicon. Half a gallon of fresh normal milk was placed in a loosely stoppered glass jar and allowed to stand at the temperature of the room for three months during the summer. At the end of this time it was examined for tyrotoxicon, but the test failed to establish its presence. From this milk as well as from the samples of cheese Dr. Moore isolated several different germs, but other more important work has prevented a closer study of these and their products.
Our own experiments, supported by the negative results of a number of other chemists, force us to conclude that the toxic principles of poisonous cheese and milk have not been yet sufficiently studied, and that there is here a very important field for further investigations.
A number of other unimportant examinations and analyses have been made, but the facts established in regard to hog cholera and swine plague are the important results of our six months' work.
Tabulated experiments in producing immunity from hog cholera in guinea pigs.
EXPERIMENTS IN PRODUCING IMMUNITY FROM SWINE PLAGUE IN GUINEA PIGS.
2 and 3 Recovered; immunity. 2 and 3 One in 3 days, the other in
REPORT OF THE CHEMIST.
WASHINGTON, D. C., December 22, 1890. SIR: I have the honor to submit herewith a brief report of the work of my Division during the past year. I am, respectfully,
H. W. WILEY,
Chemist. Hon. J. M. RUSK,
STUDIES ON THE SEPARATION OF SUGAR FROM SORGHUM
For many years attempts have been made by the division to secure a more perfect separation of the sugar from the non-sugars in sorghum juices. Extensive practical experiments were made in this direction at Fort Scott in 1886, in the practical application of the process of carbonatation.
This process consists in the addition to the mill or diffusion juices of large quantities of lime. from 1.5 per cent to 3 per cent of the weight of the juice, according to the amount of impurities present. The lime is then precipitated by blowing through the liquid a current of carbonic acid derived from a limekiln or coke furnace, or even from the chimneys of the boiler furnaces. The result of this process was entirely successful in respect of the yield of sugar, but on account of the blackening of the molasses, which was at that time a valuable by-product, it met with no favor from sorghum sugar manufacturers, but on the contrary was condemned by them as being unsuitable for the purpose.
Subsequently extensive laboratory experiments were made looking to the precipitation of the crystallizable sugar in the juices as sucrates of lime. The process employed in the Steffen method of separating sugar from beet-root molasses was the one tried for this purpose. While these experiments were successful in separating the crystallizable sugars in the form of a precipitate, they were not wholly so in securing a separation from the non-sugars, the greater part of which were also thrown down as lime compounds or carried down mechanically with the precipitated sugar. This process was, therefore, abandoned as not being practical.
The destruction of the reducing sugars or glucoses present by boiling with excess of quicklime was next tried. This process was en. tirely successful in so far as destroying the glucoses was concerned.