Part 15

"The best method of procedure for the purpose of vaccinating large animals destined to produce anti-venomous serum consists in injecting them from the outset with gradually increasing quantities of the venom of the cobra mixed with diminishing quantities of a one-in-sixty solution of hypochlorite of lime. The condition and the variations in the weights of the animals are carefully followed, in order that the injections may be made less frequently if the animals do not thrive well. Quantities of stronger and stronger venom are in turn injected, first considerably diluted, and then more concentrated; and when the animals have already acquired a sufficiently perfect immunity, the venoms derived from as large a number of different species of snakes as possible are injected. The duration of the treatment is of considerable length--at least fifteen months--before the serum is sufficiently active to be used for the purposes of treatment."

More recently the snake venom employed by Dr. Calmette for the immunisation of his horses consists of a mixture of colubrine and viperine poisons, the former making up about 80 per cent. of the mixture. A solution of this mixture is heated at about 73° C. for half an hour and then filtered, and injected into horses.

An immense number of animals have been vaccinated by this method at the Pasteur Institute at Lille, where Dr. Calmette is now director; and in one of his memoirs we are told that they have horses there which have yielded during a period of eighteen months serum extremely active against venom. These horses receive in a single inoculation, without suffering the least inconvenience, doses of venom sufficient to kill fifty horses fresh to the treatment.

Large quantities of this serum have been forwarded from the Lille Institute to various parts of the world where venomous serpents are most frequently met with, and already important evidence has been collected as to its efficacy in cases of human beings bitten by dangerous reptiles. So impressed with its importance are Indian medical authorities, that its preparation has been included in the work which the new great bacteriological institute at Agra is carrying on.

The importance of the production in situ of this anti-venomous serum has been recently demonstrated by the experiments which have been conducted in the Plague Research Laboratory, Bombay, by Mr. Lamb and his colleagues, on the keeping properties of such serums in India. From the careful investigations which have been made on this subject, these gentlemen state that anti-venomous serum undergoes a progressive and fairly rapid deterioration when stored in hot climates, and that this deterioration is greater and more rapid the higher the mean temperature to which it is subjected.

The protective potency of this horse-serum may be gathered from the fact that it suffices to inject a rabbit, for example, with a quantity amounting to about one two-hundred-thousandth of its weight to ensure the latter acquiring complete immunity from a dose of venom capable of otherwise killing it in twelve hours.

The rapidity with which it acts is also extremely remarkable. Thus, if a rabbit receive two cubic centimetres (about fifty drops) of anti-venomous serum in the marginal vein of one of its ears, it will suffer with absolute impunity an injection of venom into the marginal vein of the other ear capable of killing it under ordinary circumstances in a quarter of an hour. Its curative powers are not less remarkable, for it is possible to inject venom sufficient to kill an animal in two hours, and to let one hour and three-quarters elapse before administering the antidote, and yet at this late stage to save the victim's life, although it is necessary where such a long interval has occurred between the respective venom and serum injections to employ the latter in larger quantities than is usually required. Dr. Calmette believes that the anti-toxin may be applied at an even more advanced stage of the disease if it is employed in yet larger doses. Another novel and important feature about this anti-venomous serum is the fact that it not only protects animals from one species of very active venom, such as that of the cobra and other poisonous snakes, but it also affords protection from the dreaded venom of scorpions. This is a very remarkable and significant discovery, for hitherto the opinion has been stubbornly held that each toxin requires its specific anti-toxin for its correction. Dr. Calmette has, however, frequently indicated by his researches that this view cannot be considered so completely proven as is claimed by its supporters, and his latest investigations support the theory that particular toxins may be counteracted by several anti-toxins of different origin. Thus it has been shown by Calmette and Roux that rabbits hyper-vaccinated against rabies acquire the power of resisting venom-poison, and that the serum of horses vaccinated against tetanus or lock-jaw also nullifies the action of serpent venom.

The practical bearing of this discovery is obvious, and the hope is justified that the at present cumbrous appliances required for the elaboration of anti-toxins of such varied origin will ultimately give way to simpler and less costly methods, which will admit of these new antidotes being more widely circulated and applied.

We have seen that although most animals fall an easy prey to serpent venom, yet there are a few notable exceptions, amongst which may be mentioned hedgehogs, swine, and the mongoose. Now the very natural question arises why, if these animals are already in such a high degree immune from this poison, should not they be employed to furnish forth protective serum, instead of laboriously training up susceptible animals to become artificially immune and supply this venom anti-toxin?

This brings us face to face with one of the many problems connected with the subject of immunity which so far have successfully eluded all attempts made to solve them. Experience has shown repeatedly that although artificially acquired immunity from a particular poison can be handed on by means of an animal's serum, yet the natural immunity from a given poison enjoyed by one species of animal cannot be similarly transferred to less-favoured varieties.

This fact has long been recognised in the case of poisons of bacterial origin. Thus, white rats are absolutely immune from diphtheria, but Wassermann showed some years ago that the serum of these animals has no power whatever to counteract the action of diphtheria-toxin in other animals. Guinea-pigs were inoculated with fatal doses of diphtheria toxin along with white-rat serum; but although other guinea-pigs treated with the same toxin mixed with the ordinary artificially elaborated anti-diphtheritic serum survived, those which received the rat serum died in every case.

Now very similar results have been obtained by Calmette in respect to the serum of animals naturally immune from serpent venom. The serum of the refractory little mongoose, as well as that of the hedgehog, is wholly unable to save other animals from the lethal effect of venom poison, and similar results have been noted in respect to swine serum. But a very curious fact has also been discovered by Calmette--i.e. that these so-called naturally immune animals very frequently are quite incapable of being artificially trained to elaborate a serum possessing protective powers which can be transferred to another animal.

How splendid a domain for beneficent research lies before the scientific investigator is apparent to all, and the important work already accomplished is but an augury of yet greater discoveries awaiting the labours of such leaders as Calmette. It is not surprising, therefore, that the scientific interest in toxins and anti-toxins shows no signs of abatement. On the contrary, the competition for obtaining and working the new "claims" which pioneer research enthusiasts are constantly engaged in "pegging out" remains as keen as ever.

Despite, however, the extraordinary interest which this subject has aroused in scientific circles all over the world, nearly ten years elapsed before any notice was taken of the curious discovery made by two brothers that the blood of eels contained a highly poisonous principle, and the memoir containing this remarkable announcement remained until comparatively recently buried in the Italian journals where it was first published.

Calmette was, we believe, the first to call attention to this discovery of the brothers Mosso and give it the prominence it deserves, and both he and other investigators have not only fully confirmed it, but have greatly added to our knowledge concerning the character of the poison contained in eel serum.

Now the venerable Izaak Walton, in one of his quaint and most fascinating discourses, which although written more than two centuries ago have a freshness as if penned but yesterday, waxes enthusiastic over the eel, and supplies an elaborate recipe for its preparation for the table, telling us "it is agreed by most men that the eel is a most dainty fish; the Romans have esteemed her the Helena of their feasts, and some the queen of palate-pleasure." The announcement that the blood of eels is poisonous will hardly, despite its scientific interest, form a comfortable subject for reflection to the modern votaries of this novel Helena. Indeed, in the present timid temper of the public, this article of diet would not improbably share the ill-odour which befell the unfortunate oyster and be practically banished from our tables; but although the oyster is perhaps justifiably at present ostracised from our menus, taking the majority of its breeding-grounds into consideration, it would be the height of injustice to measure out similar drastic treatment to the eel.

That the oyster bred in sewage-contaminated beds may revenge itself upon its consumer by infecting him with the germs of typhoid has been repeatedly contended, but that the eel, although its unsavoury surroundings are proverbial, can be held responsible for poisoning those who eat it has never, we believe, been seriously maintained, although there is an old Italian saying which bids us "give eels and no wine to our enemies."

Public confidence, however, in the eel as an article of food need not be shaken, for it is satisfactory to learn that researches which, on the one hand, condemn eels as living generators of a highly poisonous substance, on the other hand allay any alarm which they may have reasonably raised by showing that this toxic principle is entirely destroyed in the processes of digestion, and that, therefore, taken through the mouth it is rendered harmless, and only when introduced into the system by inoculation beneath the skin or injected into the peritoneum can it assert its dangerous properties. That the blood of eels is, however, justifiably to be in future classed amongst the toxins, the number of which has of late been so increased, is at once apparent when we learn that about a dozen drops inoculated into a dog weighing about fourteen pounds will destroy the latter in less than ten minutes, whilst pigeons, rabbits, and guinea-pigs similarly treated, only with smaller quantities, also invariably succumb to its lethal action.

Quite recently an endeavour has been made to determine precisely the degree of toxicity possessed by eel's blood, or, in other words, to standardise the poisonous principle contained in it, so as to afford a guide to those experimenting on the subject; and it has been asserted that one cubic centimetre, or about twenty drops, injected into the veins of a rabbit weighing four pounds, may be regarded as a fatal dose for such an animal. But many difficulties surround such an attempt to exactly define the degree of toxic action possessed by such a substance, for, in the first place, the blood varies in respect to this property in different eels, whilst it also differs widely in character at different stages of the life of the fish. This seasonable variation in toxic character has been noticed in the case of viper venom, which it will be remembered was shown to be far more lethal in action when collected from snakes in the spring of the year than in the winter months.

The toxic substance contained in eel serum was originally called by its discoverers, the Mosso brothers, ittio-tossina; and they record the fact that the blood of rabbits and frogs, which animals had succumbed to its action, did not coagulate after death, whilst, curiously, in the case of dogs this abnormal phenomenon was not observed.

There are various means which may be resorted to for destroying the poisonous principle contained in eel blood, and from a dietetic point of view it is satisfactory to know that heat-exposure for a quarter of an hour to a temperature of from 57·7° to 77·7° Cent. entirely removes it, whilst its virulence is greatly modified by submitting it for a longer period, twenty-four hours, to a much lower temperature, i.e. 37° Cent. It also gradually loses its toxic properties eight days after it has been collected, even when carefully shielded from light, a feature which contrasts favourably with viper venom, which can be kept for more than a year and remains as active as when first derived from the snake. We have seen also that its toxic properties invariably succumb to the processes of digestion, so that even if fashion or fad or advertising speculators, backed by scientific names, were to decree that a wealth of nourishment and support was contained in raw eel "juice," and the edict went out that it was a desirable and highly important article of invalid diet, the general public may, according to its wont, innocently accept the edict and in this case suffer no evil consequences.

But another and very remarkable method of mitigating the virulence of eel blood, and one which so far has received no explanation, is mentioned by Dr. Wehrmann, of Moscow, who has been lately studying the character of this fish's blood in Dr. Calmette's laboratory at the Pasteur Institute at Lille. Dr. Wehrmann found that if blood serum be taken from animals previously rendered artificially immune to the action of serpent venom, and if some of this so-called anti-venomous serum be injected under the skin of eels some hours before they are killed, the lethal properties of their blood after death are considerably reduced. Thus, an eel weighing about six ounces received subcutaneous injections of five cubic centimetres of anti-venomous serum; after the lapse of four-and-twenty hours it was killed and bled, and its serum inoculated into animals in the usual way. But whereas two cubic centimetres of normal eel blood sufficed to kill a guinea-pig, this eel's blood had to be administered in twice that quantity to produce a fatal result, so that its toxic character had been reduced to a very appreciable extent. The readiness with which eel serum parts with its lethal properties, and the restricted conditions under which they can operate, sufficiently assure us that in the present state of our knowledge there is no danger to be apprehended from this fish, and in the absence of any experiments to show what is the effect on human beings of subcutaneous inoculations of such blood, there is no call for this substance to be scheduled under the Poisons Act. We have, however, by no means exhausted the extremely curious properties which characterise this material, and these properties are brought to light in a remarkable manner in connection with the investigations which have been carried out to artificially protect animals from its lethal influence, and also in some interesting experiments which have been made to compare the toxicity of eel blood with that of vipers.

It is far from an easy matter to secure for experimental purposes an adequate supply of eel serum, for even a big fish weighing nearly five pounds is not capable of yielding more than about twenty-five cubic centimetres of blood, and from this only from ten to twelve cubic centimetres of serum are obtainable. Calmette has shown that not only the venom glands of reptiles contain toxic substances, but that the blood of such snakes also possesses lethal properties, only in a far less degree. Curiously, the serum of eels is no less than three times as toxic as the serum of the most vicious viper, and, moreover, produces far more discomfort and pain to the animals into which it is introduced than accompanies the injection of viper blood. In the case of viper blood its introduction is followed by no symptoms of discomfort, the animal remains quite quiet, growing more and more somnolent, a condition which is followed by an abnormal fall of temperature, ultimately ending in complete collapse, symptoms which in a much more modified degree characterise the injection of heated eel serum into animals. This heated eel serum, which we have seen is deprived of the objectionable characteristics of ordinary eel serum, produces but very transitory symptoms in animals, occasioning some degree of somnolence, and now and again a reduction in temperature, a condition from which, however, the animals rapidly recover in from two to three hours. Animals, however, treated with this heated eel serum acquire a power of resisting the lethal action of unheated or ordinary eel serum, and this artificially induced condition of immunity continues for about three days after the completion of the treatment.

The protective properties of this heated serum are not restricted to animals subsequently inoculated with eel serum, but are extended also to animals which afterwards receive injections of viper serum; but of much greater interest and importance is the remarkable fact that heated eel serum, as well as weak doses of the latter not heated but diluted with water, are capable of protecting animals from the fatal consequences of the far more potent viper venom.

It is interesting to note that, although diluted eel serum can protect an animal from so deadly a poison as viper venom, the serum of vipers is quite unable to afford any such service in the case of animals inoculated with ordinary eel serum. The full complement of protective power obtainable from this treated eel serum is only able to slowly assert itself, for it is necessary for a period of as long as twenty-four hours to elapse after its introduction to ensure the animal's system being thoroughly impregnated with it and enable it to withstand a lethal dose of viper venom.

In this respect, what may be designated treated or protective eel serum differs very markedly from anti-venomous serum, which we have seen is serum derived from animals trained up to withstand fatal does of serpent venom, for anti-venomous serum acts immediately, and at once confers immunity on an animal from the lethal effects of such venom.

The rapidity with which it acts is indeed one of the most astonishing properties of this particular anti-toxin. Thus if two cubic centimetres of anti-venomous serum be inoculated into the marginal vein of a rabbit's ear, it at once confers upon the latter complete immunity from snake poison. Immediately after the injection of the serum, venom sufficient to destroy an ordinary rabbit in a quarter of an hour may be injected with impunity into the vein of the other ear. But not only are the protective powers of this serum so remarkable in their degree, but its curative powers, a much more difficult property to establish in a substance, are extraordinarily intense, as may be gathered from the following example. Four rabbits were inoculated with a quantity of venom calculated to destroy them in the space of two hours; one of these four animals was abandoned to its fate, but the other three received, practically at the eleventh hour, viz. just fifteen minutes before the expiration of the calculated two hours' respite, an intravenous injection of a small quantity of anti-venomous serum, only amounting to one four-hundredth part of the weight of each animal respectively. The rabbit which received only the venom died at the end of two hours, whilst the other three remained in perfect health.

But although eel serum can be persuaded to part with its poisonous character and even exercise protective powers over otherwise doomed victims, it is not able to stretch forth a healing hand to the afflicted, for, when once the poison has been introduced, whether it be eel or viper blood, or the venom of snakes, it is absolutely powerless to mitigate or stop in any way the deadly progress of the toxin. Thus whilst eel blood may acquire protective properties it cannot acquire curative properties, and, therefore, treated eel serum cannot be legitimately enrolled with the anti-toxins which have been elaborated, as, for example, anti-venomous serum, for, to be worthy of such rank, a substance must be capable of wielding both protective and curative powers.