If a portion of the brain of an animal dead from street virus is taken and made up in a dilution of one to five hundred, and this is injected, we find that it does not produce death. But a dilution of one to three hundred will invariably kill. This is practically what very often happens when one is bitten through the clothing. The saliva may be filtered and held back so that a small amount is introduced; perhaps a dilution of one to five hundred of the virus may get into the wound, but this is usually not enough to cause the disease. There is no possible way of estimating the amount of the inoculation. In such cases one's chances of never contracting the disease are only decreased; that is all we can say.
The treating of individuals, bitten by rabid animals, in the Pasteur Institutes, is simply the practical application of results obtained by Pasteur from his original work on rabies virus. Pasteur was a French chemist living in Paris, and he began his search for the cause and cure of rabies in 1880. He hoped to find a sure method of preventing the development of the dread disease, even if he could not find a cure for it after it had developed. While he was pursuing this research Pasteur had access to the cases of rabies in the Paris hospitals, and these numbered sixty each year. He had practically an unlimited supply, for France could furnish him with twenty-five hundred more mad dogs, and a large number of other animals each year.
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Pasteur devoted the remainder of his life to the study of this subject. He collected some saliva from the mouth of a child, on December 11, 1880, who had died at the Hospital Trousseau four hours before. This saliva he diluted with distilled water, and this mixture he injected into rabbits, and they all died, and the saliva taken from these rabbits when injected into other rabbits caused their death with rabies. He found also that saliva from rabid dogs almost always caused the disease. The incubation period varied within wide limits, and very often the animals lived. He then used the blood of rabid dogs for inoculation, but these blood inoculations always failed to produce the disease. Pasteur was convinced after careful study of rabid animals during the many months necessary to complete his experiments, that rabies was a disease of the nervous system, and that the poison (virus) was transmitted from the wound to the brain by the way of the nerve trunks. Then to prove his theory Pasteur removed a portion of the brain of a dog that had died of rabies. A part of this was rubbed up in sterile water and used to inoculate other animals; and subcutaneous inoculations with this material almost always produced death.
After this Pasteur tried a new method and injected directly into the nervous system, either into the nerve trunk or directly into the brain, after trephining, and all such injections produced rabies in the injected animal and death. He also found that rabbits inoculated in the brain always died in the same length of time. When he injected into the nerve trunk the inoculation period was longer, depending upon the distance from the brain. Two problems now remained for Pasteur to solve, and these were, how could he obtain the definite virulence and how could he reduce the virulence regularly and gradually, so that it could be used by inoculation safely as a vaccine to produce immunity to rabies in healthy animals, and also to prevent the development of rabies in animals bitten by rabid animals. He first tried successive inoculations. These inoculations were made, after trephining, directly to the brain, and he used a portion of the brain as a virus each time. He inoculated rabbit number one with a portion of brain taken from a rabid dog, and this rabbit died on the fifteenth day. He then inoculated rabbit number two with a portion of the brain of rabbit number one; from the brain of rabbit number two the virus was supplied for inoculating rabbit number three, and thus the brain of each inoculated rabbit was taken, after its death, for material to inoculate the next rabbit in the series. This experimentation showed him that each rabbit in the series died a little sooner, showing that the virus was becoming more virulent, till no increase in activity of the poison was shown after the fiftieth successive inoculation. "Rabbits inoculated with a brain suspension of rabbit number fifty all died in seven days." This caused Pasteur to name the virus of number fifty "virus fixe," a virus of definite length. He now had obtained a virus of definite strength and the next question was, how could the virulence be gradually and definitely reduced.
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This he accomplished after many experiments. He proved that pieces of the "medulla oblongata" suspended in sterile tubes which contained fragments of caustic potash, steadily and gradually reduced their virulence as they dried, till the fourteenth day, when they were practically inert. New specimens were prepared each day and cords which had dried in one day Pasteur called "one-day virus;" cords which had dried in two days, "two day's virus," and so on up to the fourteenth day. With this graduated virus he now experimented on dogs, and the injection he used on the first day consisted of an emulsion of fourteen-day virus; for the second day, the thirteen-day virus, thus using a stronger virus each day, until on the fourteenth day he used the full strength virus. This treatment produced what is called immunity in the dog, and even the direct inoculation into the brain of the strong virus would not produce death.
After Pasteur had thoroughly satisfied himself by repeated trials, he announced his wonderful discovery, and it was in 1886 that Pasteur considered the preventive inoculation in human beings as resting upon a satisfactory experimental basis. During these five years this eminent man proved that it was possible to protect or immunize the lower animals, rabbits and dogs, against inoculation with the virulent virus.
The efficiency of this immunity was given trials by different methods of inoculation. It was found that sixty per cent of dogs inoculated under the "dura" (a membrane of the brain) were saved if treatment was given the second day. This test is more severe than is required to meet the ordinary infection of rabies. Pasteur, after a series of these final tests were so convincing, prescribed the preventive inoculations in human beings and on July 6th, 1886, the first human patient received the first treatment of his series of inoculations.
The method of obtaining the attenuated virus used in the treatment is as follows: A rabbit is inoculated by the brain method before described, each day, with suspension of the fresh, fixed virus. These rabbits die in six days after the inoculation. In this way a rabbit dies each day; the spinal cord is removed, divided into sections, and suspended in a flask containing potassium hydrate. The action of potassium hydrate is drying (desiccating). A series of these cords, which have been hung on fourteen successive days, are always kept in stock for the treatment of patients. The virus becomes less active with each successive day of exposure to drying (desiccation) and finally the virulence is altogether lost.