Experiments on Animals

by Stephen Paget

Before bacteriology, the cause of tetanus (lock-jaw) was unknown, and men were free to imagine that it was due to inflammation travelling up an injured nerve to the central nervous system. This false and mischievous theory was abolished by the experimental work of Sternberg (1880), Carle and Rattone (1884), and Nicolaier (1884), who proved, once and for all, that the disease is an infection by a specific flagellate organism. Their work was of the utmost difficulty, for many reasons. First, because tetanus, in some tropical countries, is so common that it may fairly be called endemic; and many of these tropical cases, there being no record of any external infection, had been taken as evidence that the disease can occur "of itself." Of this frequency of tetanus in tropical countries, Sir Patrick Manson, in his book on Tropical Diseases (1898), says:—

"Tetanus is an exceedingly common disease in some tropical countries. In Western Africa, for example, a large proportion of wounds, no matter how trifling as wounds they may be, if they are fouled by earth or dirt, result in tetanus. The French in Senegambia have found this to their cost. A gentleman who had travelled much in Congoland told me that certain tribes poison their arrows by simply dipping the tips in a particular kind of mud. A wound from these arrows is nearly sure to cause tetanus. In many countries, so general and so extensive is the distribution of the tetanus-bacillus that trismus neonatorum (tetanus of newly-born infants) is a principal cause of the excessive infant mortality."

Next, because the tetanus-bacillus has its natural abode in the superficial layers of the soil: here it is associated with a vast number of other organisms, so that its identification and isolation were a work of immeasurable complexity. What mixed company it keeps, is shown by Houston's estimate of the number of microbes per gramme in twenty-one samples of different soils. This number ranged from 8326 in virgin sand, and 475,282 in virgin peat, to 115,014,492 in the soil from the trench of a sewage-farm. In all rich and well-manured soil the tetanus-bacillus may possibly be present; but it was the work of years to dissociate it from the myriads of organisms outnumbering it.

Next, because it cannot be got to grow in cultures exposed to the air: its proper place is below the surface of the soil, away from the air; it is "strictly anaërobic," and the attempts to cultivate it by ordinary methods failed again and again. It had to be cultivated below the surface of certain nutrient media, or in a special atmosphere of nitrogen or hydrogen.

These and other difficulties for many years delayed the final proof of the true pathology of tetanus. The success of the work was mainly due to Nicolaier. He started from the well-known fact that tetanus mostly comes of wounds or scratches contaminated with particles of earth—such mischances as the grinding of dirt or gravel into the skin, or the tearing of it by a splinter of wood or a rusty nail; as Dr. Poore says, in his Milroy Lectures (1899), "Every child who falls on the ground and gets an abrasion of the skin, all tillers of the soil who get accidental wounds in the course of duty, and every horse which 'breaks its knees' by falling in the London streets, runs potentially a risk of inoculation with tetanus." Nicolaier therefore studied the various microbes of the soil, and made inoculations of garden-mould under the skin of rabbits. He was able, by these inoculations, to produce tetanus in them; and the discharge from the points of inoculation, put under the skin of other rabbits, produced the disease again. He also identified the bacillus, and cultivated it; but in these cultures it was mixed with other organisms, and he failed to isolate it from them. Carle and Rattone, and Rosenbach, were able to produce tetanus in animals by inoculating them with discharge from the wounds of patients attacked by the disease. Finally, Kitasato, in 1889, found a way of obtaining pure cultures of the bacillus. Beginning with impure cultures such as Nicolaier had made, he kept these at a temperature of 36° C. till the bacillus had spored; then, by repeated exposures of the cultures to a temperature of 80° C. for three-quarters of an hour at a time, he killed-off all organisms except the spores of the tetanus-bacillus; then he kept these in an atmosphere of hydrogen, at a temperature of 20° C., and thus got pure cultures.

Brieger, Fränkel, Cohen, Sidney Martin, Kanthack, and others, have studied the chemical products of the disease, have obtained them from cultures and from infected tissues, and have been able with these toxins to produce the disease in animals. As with the other infective diseases, so with tetanus, there have been two main lines of researches; the one, toward a fuller knowledge of the chemical changes in the blood and in the central nervous system; the other, toward a fuller knowledge of the nature and ways of the bacillus, and its method of invasion. Before any study of immunity or immunisation, or of neutralisation of the toxins in man by an antitoxin, came the study of the toxins and of the bacillus. It was proved, by an immense quantity of hard work, that the bacillus does not tend to invade the blood, or to pass beyond the lymphatic glands in the immediate neighbourhood of the site of inoculation; that it stays in and about the wound, and there multiplies, and from this site pours into the blood the chemical products which cause the disease; and that these chemical substances have a selective action on certain nerve-cells in the brain and the spinal cord. This is the bare outline of the facts; and no account can be given here of the intricate problems of bacteriology and animal chemistry that have been answered, or are still waiting an answer. At least, it is evident that the whole pathology of tetanus was found, proved, and interpreted by the help of experiments on animals; and that these alone did away with the old false doctrine that the disease was due to rapid extension of inflammation up a nerve to the brain.

In 1894 came the use of an antitoxin in cases of the disease, and, in 1895, 42 cases were reported, with 27 recoveries. It cannot be said that any one of the diverse preparations of tetanus-antitoxin, up to this present time, has triumphed over the disease. Tetanus is of all diseases the hardest to reckon with: the first sign of it is the last stage of it; there is no warning, nothing, it may be, but a healed scratch, till the central nervous system is affected with sudden and rapidly advancing degeneration of certain cells. These and other difficulties have stood in the way of an antitoxin treatment; and there is no less difficulty in estimating the efficacy of that treatment. The recovery, under antitoxin, of a "chronic" case cannot always or altogether be attributed to the treatment; and in a very acute case, antitoxin, like everything else, has but small chance of success. Various reports on the antitoxin treatment, published during 1897-1899, give the following figures:—

26 cases,	with 12 recoveries.
98"	57"
36"	25"
22"	11"
51"	36"
10"	7"

Probably the paper by Dr. Lambert of New York, in the Medical News, July 1900, gives fairly the general opinion of the treatment, so far as the subcutaneous administration of antitoxin is concerned:—

"The following cases of tetanus, treated with antitoxin, comprise published and unpublished cases. We have a total of 279 cases, with a mortality of 44.08 per cent.: but of these we must rule out 17 cases—4 deaths from intercurrent diseases, 8 deaths in cases in which the antitoxin was given but a few hours before death, and 5 recoveries in which antitoxin was not given until after the twelfth day (as they probably would have recovered without it). We have left 262 cases, with 151 recoveries, and 111 deaths, a mortality of 42.36 per cent. Dividing the cases into acute and chronic, we have 124 acute cases, with 35 recoveries and 89 deaths, a mortality of 71.77 per cent., and 138 chronic cases, with 116 recoveries and 22 deaths, a mortality of 15.94 per cent. In interpreting critically these statistics, we see that in acute cases the mortality is but slightly reduced, being but 72 per cent. instead of 88 per cent. But, in the less acute cases, there is a decided improvement, from 40 per cent. to 16 per cent. Taking the statistics as a whole, there is a distinct improvement in the mortality of tetanus since the introduction of antitoxin."