As it happened, the world was not kept long in suspense. Almost before the proposition had taken shape in the minds of the other leaders, Pasteur had found a solution. Guided by the empirical success of Jenner, he, like many others, had long practised inoculation experiments, and on February 9, 1880, he announced to the French Academy of Sciences that he had found a method of so reducing the virulence of a disease germ that when introduced into the system of a susceptible animal it produced only a mild form of the disease, which, however, sufficed to protect against the usual virulent form exactly as vaccinia protects against small-pox. The particular disease experimented with was that infectious malady of poultry known familiarly as "chicken cholera." In October of the same year Pasteur announced the method by which this "attenuation of the virus," as he termed it, had been brought about - by cultivation of the disease germs in artificial media, exposed to the air, and he did not hesitate to assert his belief that the method would prove "susceptible of generalization" - that is to say, of application to other diseases than the particular one in question.

Within a few months he made good this prophecy, for in February, 1881, he announced to the Academy that with the aid, as before, of his associates MM. Chamberland and Roux, he had produced an attenuated virus of the anthrax microbe by the use of which, as he affirmed with great confidence, he could protect sheep, and presumably cattle, against that fatal malady. "In some recent publications," said Pasteur, "I announced the first case of the attenuation of a virus by experimental methods only. Formed of a special microbe of an extreme minuteness, this virus may be multiplied by artificial culture outside the animal body. These cultures, left alone without any possible external contamination, undergo, in the course of time, modifications of their virulency to a greater or less extent. The oxygen of the atmosphere is said to be the chief cause of these attenuations - that is, this lessening of the facilities of multiplication of the microbe; for it is evident that the difference of virulence is in some way associated with differences of development in the parasitic economy.

"There is no need to insist upon the interesting character of these results and the deductions to be made therefrom. To seek to lessen the virulence by rational means would be to establish, upon an experimental basis, the hope of preparing from an active virus, easily cultivated either in the human or animal body, a vaccine-virus of restrained development capable of preventing the fatal effects of the former. Therefore, we have applied all our energies to investigate the possible generalizing action of atmospheric oxygen in the attenuation of virus.

"The anthrax virus, being one that has been most carefully studied, seemed to be the first that should attract our attention. Every time, however, we encountered a difficulty. Between the microbe of chicken cholera and the microbe of anthrax there exists an essential difference which does not allow the new experiment to be verified by the old. The microbes of chicken cholera do not, in effect, seem to resolve themselves, in their culture, into veritable germs. The latter are merely cells, or articulations always ready to multiply by division, except when the particular conditions in which they become true germs are known.

"The yeast of beer is a striking example of these cellular productions, being able to multiply themselves indefinitely without the apparition of their original spores. There exist many mucedines (Mucedinae?) of tubular mushrooms, which in certain conditions of culture produce a chain of more or less spherical cells called Conidae. The latter, detached from their branches, are able to reproduce themselves in the form of cells, without the appearance, at least with a change in the conditions of culture, of the spores of their respective mucedines. These vegetable organisms can be compared to plants which are cultivated by slipping, and to produce which it is not necessary to have the fruits or the seeds of the mother plant.

The anthrax bacterium, in its artificial cultivation, behaves very differently. Its mycelian filaments, if one may so describe them, have been produced scarcely for twenty-four or forty-eight hours when they are seen to transform themselves, those especially which are in free contact with the air, into very refringent corpuscles, capable of gradually isolating themselves into true germs of slight organization. Moreover, observation shows that these germs, formed so quickly in the culture, do not undergo, after exposure for a time to atmospheric air, any change either in their vitality or their virulence. I was able to present to the Academy a tube containing some spores of anthrax bacteria produced four years ago, on March 21, 1887. Each year the germination of these little corpuscles has been tried, and each year the germination has been accomplished with the same facility and the same rapidity as at first. Each year also the virulence of the new cultures has been tested, and they have not shown any visible falling off. Therefore, how can we experiment with the action of the air upon the anthrax virus with any expectation of making it less virulent?

"The crucial difficulty lies perhaps entirely in this rapid reproduction of the bacteria germs which we have just related. In its form of a filament, and in its multiplication by division, is not this organism at all points comparable with the microbe of the chicken cholera?

"That a germ, properly so called, that a seed, does not suffer any modification on account of the air is easily conceived; but it is conceivable not less easily that if there should be any change it would occur by preference in the case of a mycelian fragment. It is thus that a slip which may have been abandoned in the soil in contact with the air does not take long to lose all vitality, while under similar conditions a seed is preserved in readiness to reproduce the plant. If these views have any foundation, we are led to think that in order to prove the action of the air upon the anthrax bacteria it will be indispensable to submit to this action the mycelian development of the minute organism under conditions where there cannot be the least admixture of corpuscular germs. Hence the problem of submitting the bacteria to the action of oxygen comes back to the question of presenting entirely the formation of spores. The question being put in this way, we are beginning to recognize that it is capable of being solved.

"We can, in fact, prevent the appearance of spores in the artificial cultures of the anthrax parasite by various artifices. At the lowest temperature at which this parasite can be cultivated - that is to say, about +16 degrees Centigrade - the bacterium does not produce germs - at any rate, for a very long time. The shapes of the minute microbe at this lowest limit of its development are irregular, in the form of balls and pears - in a word, they are monstrosities - but they are without spores. In the last regard also it is the same at the highest temperatures at which the parasite can be cultivated, temperatures which vary slightly according to the means employed. In neutral chicken bouillon the bacteria cannot be cultivated above 45 degrees. Culture, however, is easy and abundant at 42 to 43 degrees, but equally without any formation of spores. Consequently a culture of mycelian bacteria can be kept entirely free from germs while in contact with the open air at a temperature of from 42 to 43 degrees Centigrade. Now appear the three remarkable results. After about one month of waiting the culture dies - that is to say, if put into a fresh bouillon it becomes absolutely sterile.

"So much for the life and nutrition of this organism. In respect to its virulence, it is an extraordinary fact that it disappears entirely after eight days' culture at 42 to 43 degrees Centigrade, or, at any rate, the cultures are innocuous for the guinea-pig, the rabbit, and the sheep, the three kinds of animals most apt to contract anthrax. We are thus able to obtain, not only the attenuation of the virulence, but also its complete suppression by a simple method of cultivation. Moreover, we see also the possibility of preserving and cultivating the terrible microbe in an inoffensive state. What is it that happens in these eight days at 43 degrees that suffices to take away the virulence of the bacteria? Let us remember that the microbe of chicken cholera dies in contact with the air, in a period somewhat protracted, it is true, but after successive attenuations. Are we justified in thinking that it ought to be the same in regard to the microbe of anthrax? This hypothesis is confirmed by experiment. Before the disappearance of its virulence the anthrax microbe passes through various degrees of attenuation, and, moreover, as is also the case with the microbe of chicken cholera, each of these attenuated states of virulence can be obtained by cultivation. Moreover, since, according to one of our recent Communications, anthrax is not recurrent, each of our attenuated anthrax microbes is, for the better-developed microbe, a vaccine - that is to say, a virus producing a less-malignant malady. What, therefore, is easier than to find in these a virus that will infect with anthrax sheep, cows, and horses, without killing them, and ultimately capable of warding off the mortal malady? We have practised this experiment with great success upon sheep, and when the season comes for the assembling of the flocks at Beauce we shall try the experiment on a larger scale.

"Already M. Toussaint has announced that sheep can be saved by preventive inoculations; but when this able observer shall have published his results; on the subject of which we have made such exhaustive studies, as yet unpublished, we shall be able to see the whole difference which exists between the two methods - the uncertainty of the one and the certainty of the other. That which we announce has, moreover, the very great advantage of resting upon the existence of a poison vaccine cultivable at will, and which can be increased indefinitely in the space of a few hours without having recourse to infected blood."[8]

This announcement was immediately challenged in a way that brought it to the attention of the entire world. The president of an agricultural society, realizing the enormous importance of the subject, proposed to Pasteur that his alleged discovery should be submitted to a decisive public test. He proposed to furnish a drove of fifty sheep half of which were to be inoculated with the attenuated virus of Pasteur. Subsequently all the sheep were to be inoculated with virulent virus, all being kept together in one pen under precisely the same conditions. The "protected" sheep were to remain healthy; the unprotected ones to die of anthrax; so read the terms of the proposition. Pasteur accepted the challenge; he even permitted a change in the programme by which two goats were substituted for two of the sheep, and ten cattle added, stipulating, however, that since his experiments had not yet been extended to cattle these should not be regarded as falling rigidly within the terms of the test.

It was a test to try the soul of any man, for all the world looked on askance, prepared to deride the maker of so preposterous a claim as soon as his claim should be proved baseless. Not even the fame of Pasteur could make the public at large, lay or scientific, believe in the possibility of what he proposed to accomplish. There was time for all the world to be informed of the procedure, for the first "preventive" inoculation - or vaccination, as Pasteur termed it - was made on May 5th, the second on May 17th, and another interval of two weeks must elapse before the final inoculations with the unattenuated virus. Twenty-four sheep, one goat, and five cattle were submitted to the preliminary vaccinations. Then, on May 31 st, all sixty of the animals were inoculated, a protected and unprotected one alternately, with an extremely virulent culture of anthrax microbes that had been in Pasteur's laboratory since 1877. This accomplished, the animals were left together in one enclosure to await the issue.

Two days later, June 2d, at the appointed hour of rendezvous, a vast crowd, composed of veterinary surgeons, newspaper correspondents, and farmers from far and near, gathered to witness the closing scenes of this scientific tourney. What they saw was one of the most dramatic scenes in the history of peaceful science - a scene which, as Pasteur declared afterwards, "amazed the assembly." Scattered about the enclosure, dead, dying, or manifestly sick unto death, lay the unprotected animals, one and all, while each and every "protected" animal stalked unconcernedly about with every appearance of perfect health. Twenty of the sheep and the one goat were already dead; two other sheep expired under the eyes of the spectators; the remaining victims lingered but a few hours longer. Thus in a manner theatrical enough, not to say tragic, was proclaimed the unequivocal victory of science. Naturally enough, the unbelievers struck their colors and surrendered without terms; the principle of protective vaccination, with a virus experimentally prepared in the laboratory, was established beyond the reach of controversy.

That memorable scientific battle marked the beginning of a new era in medicine. It was a foregone conclusion that the principle thus established would be still further generalized; that it would be applied to human maladies; that in all probability it would grapple successfully, sooner or later, with many infectious diseases. That expectation has advanced rapidly towards realization. Pasteur himself made the application to the human subject in the disease hydrophobia in 1885, since which time that hitherto most fatal of maladies has largely lost its terrors. Thousands of persons bitten by mad dogs have been snatched from the fatal consequences of that mishap by this method at the Pasteur Institute in Paris, and at the similar institutes, built on the model of this parent one, that have been established all over the world in regions as widely separated as New York and Nha-Trang.