Fig. 84.—Platurus laticaudatus(syn.P. fischeri).(After Sir Joseph Fayrer.)
Fig. 84.—Platurus laticaudatus(syn.P. fischeri).(After Sir Joseph Fayrer.)
Two large poison-fangs, and only one or two small solid teeth near the posterior extremity of the maxillary. Head shields large;nostrils lateral, the nasal shields separated by the internasals. Body greatly elongate; scales smooth and imbricate; ventrals and subcaudals large.
Four species, distributed in the eastern parts of the Indian Ocean and in the Western Pacific.
P. laticaudatus(syn.P. fischeri;fig. 84).—Olive above, yellowish on the belly, with 29-48 black annuli.
Total length: 970 millimetres; tail 90.
Habitat: From the Bay of Bengal to the China Sea and the Western South Pacific Ocean.
P. colubrinus(fig. 83).—Olive above, yellowish on the belly, with 28-54 black annuli, some or all of which may be interrupted below.
Total length, 1,270 millimetres; tail 125.
Habitat: From the Bay of Bengal to the China Sea and the Western South Pacific Ocean.
P. muelleri.—62 black annuli, some of which are interrupted on the belly.
Habitat: Only found in the South Pacific Ocean (subtropical zone), as far as the New Hebrides and the shores of Tasmania.
P. schistorhynchus.—Coloration and size as inP. colubrinus: body with 25-45 annuli.
Habitat: Western Tropical Pacific.
Non-poisonousas well aspoisonoussnakes possessparotidandupper labialglands capable of secreting venom. In the former the organs of inoculation are wanting, but we shall see later on that the toxic secretion of their glands is just as indispensable to them as to the snakes of the second category for the purpose of enabling them to digest their prey.
For the morphological, histological, and physiological demonstration of the existence of these glands in harmless reptiles we are indebted to Leydig (1873), whose discovery has since been confirmed and extended by the researches of Phisalix and Bertrand, Alcock, L. Rogers, and L. Lannoy.
The parotids of Grass Snakes are mixed glands of the sero-mucous type. The serous tubes are situate almost exclusively in the posterior portion of the gland. As we proceed towards the anterior portion, we find that these serous tubes are interspersed with others which are exclusively mucous or sero-mucous, and they become entangled with those of the upper labial gland, properly so-called. The substance of the gland is divided into several lobes by bands of connective tissue; the tubes are separated byseptaof the same tissue, in extremely delicate layers (Lannoy).
In poisonous snakes these glands are much more developed, especially in their hinder portions, which sometimes assumeenormous dimensions. They may attain the size of a large almond (Crotalus,Naja), and they then occupy the spacious chamber already described (Chap. I., p. 10), which is situated behind the eye on each side of the skull.
Each gland is surrounded by a thick capsule of fibrous tissue, two prolongations of which, the one anterior, the other posterior, keep it in its place beneath themassetermuscle. A portion of the latter is inserted in the capsule itself, in such a way that when the snake closes its jaws to bite, the gland is forcibly compressed and the contained liquid is squeezed into its excretory duct.
Between the muscle and the envelope of the gland there is a serous pouch, which enables the one to slide over the other.
The excretory duct runs along the outer side of the upper jaw, and opens by a slit at the base of the poison-fang, with which it inosculates at right angles in a little muscular mass forming asphincter.
In the normal position of repose, the poison-fang is always concealed by a gingival fold of mucous membrane, in the substance of which are buried a few fibres of the tendon of the internal pterygoid muscle. When the latter contracts, the tooth is almost completely exposed, and the efferent duct of the gland then assumes an oblique position, which allows of the direct discharge of the venom through the canal which runs along the greater portion of the length of the tooth.
When the poison-fangs are folded back in their sheath, the poisonous secretion can escape freely into the buccal cavity by the slit situated at the base of the fangs.
At the moment when the animal is about to bite, when it throws back its head and opens its jaws, directing its fangs forwards, the muscles that come into action (masseters,temporals, andpterygoids) compress the glands on each side, and cause the venom to be expelled in a sudden jet, as if by a sort of ejaculatory process. In the case of certain species the venom may be projected to a distance of more than a yard.
The quantity of venom secreted by the glands varies greatly, according to the length of time which has elapsed since the animal took its last meal, and in accordance with a number of other conditions not very easy to determine.
The Common Viper of Europe yields scarcely 10 centigrammes of poison, while an adult Indian Cobra may excrete more than 1 gramme.
Freshly collected venom is a syrupy liquid, citron-yellow or slightly opalescent white in colour.
When dried rapidlyin vacuoor in a desiccator over calcium chloride, it concretes in cracked translucent lamellæ like albumin or gum arabic, and thus assumes a crystalloid aspect. In this condition it may be kept indefinitely, if protected from light, air, and moisture. It dissolves again in water just as readily as albumin or dried serums.
I regularly weighed the dry residue from eleven bites made on a watch-glass by twoNaja haje, received at my laboratory from Egypt at the same time, and placed in the same case. Both snakes were approximately of equal length, 1,070 millimetres. Throughout the entire course of the experiment, which lastedone hundred and two days, neither of them took any food, but they drank water and frequently bathed.
The results that I obtained are shown in the table on next page.
It will be seen that in one hundred and two days, an adultNaja hajeis capable of producing on an average 0·632 gramme of liquid venom, equal to a mean weight of 0·188 gramme of dry extract; and we may conclude that 1 gramme ofliquidgives 0·336 gramme ofdryvenom.
In Australia it has been found by MacGarvie Smith, of Sydney, thatPseudechis porphyriacusyields at each bite a quantity of venom varying from 0·100 gramme to 0·160 gramme (equal to 0·024 gramme to 0·046 gramme of dry venom), and that aHoplocephalus curtus(Tiger Snake) yields 0·065 gramme to 0·150 gramme ofliquid venom, with 0·017 gramme to 0·055 gramme of dry residue. In all the experiments of this physiologist, the proportion of dry residue varied from 9 to 38 per cent. of the liquid venom excreted by the reptile.
ALachesis lanceolatus(Fer-de-lance) from Martinique, of medium size, when both of its glands were squeezed, furnished me with 0·320 gramme of liquid venom, and 0·127 gramme of dry extract.
Two largeCerastesvipers, from Egypt, yielded me, one 0·123 gramme, the other 0·085 gramme of liquid venom, which, after desiccation, left respectively 0·027 gramme and 0·019 gramme of dry residue.
Under the same conditions, a magnificentCrotalus confluentus(Mottled Rattle-Snake), for which I was indebted to the kindness of Mr. Retlie, of New York, yielded, two months after reaching my laboratory, 0·370 gramme of liquid venom and 0·105 gramme of dry extract ina single bite.
The total quantity of liquid venom that I found contained in the two glands of the same reptile, when extirpated after death, and after the snake had been in the laboratory for five months, amounted to 1·136 gramme, which gave 0·480 gramme of dry extract.
We see, therefore, that the proportion of dry residue, including albumin, salts, thedébrisof leucocytes, and the toxic substance, oscillates between 20 and 38 per cent. Its strength varies with the length of time that has elapsed since the snake’s last bite or last meal.
From thehistological standpoint, the process of the secretion of venom, in the cells of the glands, may be divided into two stages:—
(a) A stage of nuclear elaboration.
(b) A stage of cytoplasmic elaboration.
These two stages are superposed and successive.
In addition to the passive exchanges between the nucleus and the cytoplasm, the nuclear mass actively participates in the secretion. This participation is rendered evident:—
(1) By the difference of chromaticity in the granules of chromatin.
(2) By the emission of formed granules into the cytoplasm, granules which are spherical and of equal bulk, with the chromatic reactions of differentiated intranuclear chromatin.
(3) By the exosmosis of the dissolved nuclear substance, accessorily formed in an ergastoplasmic shape.
These formations constitute, on the one hand, the granules ofvenogen; on the other, the ergastoplasmic venogen. In the poison-cell ofVipera aspis, and in the serous cell of the parotid glands ofTropidonotus natrix(Grass Snake) the venogen is elaborated chiefly in granular form.
On entering the perinuclear cytoplasm, the granule of venogen and the ergastoplasmic venogen may either disappear immediately,as happens in periods of cellular stimulation, or else continue to exist for some time within the cell, indicating a period of saturation by the elaborated material.
During cytoplasmic activity the granule of venogen and the ergastoplasmic venogen disappear.
Nuclear elaboration and cytoplasmic elaboration constitute two different cycles of secretion. The effect of the nuclear cycle is to furnish the cytoplasm with the elements necessary for the work of secretion properly so-called. Cytoplasmic elaboration is not confined to the basal protoplasm, but takes place throughout the entire cell: it is especially active in the perinuclear cytoplasm.
The granule of venogen is distinguished from the granule of elaborated venom by its affinity for Unna’s blue, safranin, and fuchsin. The granule of venom has an affinity for eosin; it is never excreted in granular form, but after intracellular dissolution.
Venogen is never met with in the lumen of the gland-tube.6
Venom can be extracted from the poison-glands of either freshly killed or living snakes.
In cases in which the venom of dead snakes has to be collected, the best method of extraction consists in fixing the head of the animal to a sheet of cork and carefully dissecting out the gland on each side. The reptile being placed on its back, the lower jaw is removed with a pair of scissors; two strong pins or two tacks are thrust through the skull, in the median line, in order to keep the head from moving. The poison-fangs are next drawn out of their sheaths, and, without injuring them, the two poison-ducts, which open at their bases, are isolated and tied with a thread in order to prevent the poison from running out.
The dissection of the glands is then very easy; they are liftedout and placed in a saucer. The end of the duct is cut between the gland and the ligature, and with a pair of fenestrated or polypus forceps the whole of the glandular mass is gently squeezed from behind forwards, the liquid which flows out being received in a large watch-glass.
If pressed for time, a more simple method of operating is to hold the head of the snake in the left hand, with the mouth open and the lower jaw directed downwards. A watch-glass, capsule, or receptacle of some sort, such as a cup or plate, is then introduced by an assistant between the jaws, and, with the index finger and thumb of the right hand, the whole of the region occupied by the glands on each side of the upper jaw is forcibly compressed from behind forwards; the poison flows out by the fangs.
The extraction of the venom from living snakes is effected in the same manner. The animal being firmly held by the neck, as close as possible to the head, so that it cannot turn and bite; it can be made to eject the greater portion of the liquid contained in its two glands by compressing the latter with force from behind forwards, as one would squeeze out the juice from a quarter of an orange (fig. 85).
It is necessary to take care that the reptile cannot coil itself round furniture or other objects in the vicinity of the operator, for if this should happen there would be the greatest difficulty in making it let go, especially if dealing with a strong animal such as a Cobra, Rattle-Snake, or Fer-de-lance.
Snakes of the last-mentioned kind are especially difficult to manage. In order to avoid the risk of being bitten, it is always wise to begin by pinning down the head of the animal in a corner of its cage by means of a stick, and to seize it with a pair of long fenestrated tongs shaped like forceps. The operator then easily draws the reptile towards him and grasps it firmly by the neck with his left hand, always as close to the head as possible, at the same time raising the body quickly in order to prevent it from takinghold of anything. In this way the most powerful snake is perfectly under control.
Fig. 85.—Collecting Venom from aLachesisat the Serotherapeutic Institute at São Paulo(Brazil).
Fig. 85.—Collecting Venom from aLachesisat the Serotherapeutic Institute at São Paulo(Brazil).
Fig. 86.—Chloroforming a Cobra in order to Collect Venom, at the French Settlement of Pondicherry, in India(Stage I.).
Fig. 86.—Chloroforming a Cobra in order to Collect Venom, at the French Settlement of Pondicherry, in India(Stage I.).
At Pondicherry, where is collected the greater portion of the venom ofNaja tripudiansused by me for the vaccination of the horses that produce antivenomous serum, it is customary to chloroform the snakes in order to render them easier to manipulate.
The reptile is placed in a large covered jar, containing a pad of absorbent wool impregnated with chloroform (figs. 86, 87), and in a few minutes it is stupefied. It is then grasped by the neck with the hands, and the edge of a plate is slipped between its jaws. On compressing the two poison-glands with the fingers, the venom dribbles out on to the plate.
A detailed description of this technique will be found in a note kindly drawn up for me by my friend Dr. Gouzien, late head of the Medical Staff of the French Settlements in India, and reproduced further on in the section of this book devoted to documents. The note in question was accompanied by figs. 17, 18, 19, 86, 87, and 88, which are reproduced from photographs, for which I am indebted to the kindness of M. Geracki, Engineer of the Savanna Spinning Mill at Pondicherry, Dr. Lhomme, and M. Serph, Assistant Surgeon-Dispenser.
The collection of the venom having been completed, the snake is put back into its cage again, the tail and the body being introduced first, and then the head. The lid or trap-door is half closed with the left hand, and, with a quick forward thrust, the right hand releases its grasp of the reptile and is immediately withdrawn; at the same time the left hand completes the closure of the cage. The snake is temporarily dazed, as though stunned, and it is only after the lapse of a moment that it thinks of darting open-mouthed at the walls of its prison.
When it is desired to procure large quantities of venom, as is indispensable in laboratories where antivenomous serum is prepared, the endeavour must be made to keep the snakes alive for the longest possible time. It then becomes necessary to resortto artificial feeding in the manner previously described (see p. 17), for they very often refuse to feed themselves.
Fig. 87.—Chloroforming a Cobra in order to Collect Venom, at the French Settlement of Pondicherry, in India(Stage II.).
Fig. 87.—Chloroforming a Cobra in order to Collect Venom, at the French Settlement of Pondicherry, in India(Stage II.).
Except when a snake ismoulting, the venom can be extracted from its glands about every fortnight; and it is better that the extraction be not performed concurrently with artificial feeding, since, owing to the fact that the venom serves the animal as digestive juice, the reptile will soon perish if deprived of the means of digesting the food that it is obliged to receive. It is best, therefore, to select one day of the week for artificial feeding, and the corresponding day of the following week for the extraction of the venom.
When the venom has been collected, it must immediately beplaced in a desiccator over calcium chloride or sulphuric acid, in order to dry it rapidly. In hot countries, and where no laboratory specially equipped for the purpose exists, it will suffice to dry the venom in a current of air, or even in the sun. It then concretes in scales of a citrin colour, more or less dark, according to the concentration of the liquid. In this dry condition, placed in well-corked bottles, protected from damp air, it may be kept almost indefinitely without losing anything of its original toxic power. On the contrary, if the desiccation be imperfect it undergoes a somewhat rapid change, and assumes a disagreeable odour of meat peptone. I have kept samples of various venoms, dried as described, forfifteenyears without any sensible diminution of their activity.
Fig. 88.—Collecting Cobra Venom at Pondicherry(Stage III.).
Fig. 88.—Collecting Cobra Venom at Pondicherry(Stage III.).
In the condition in which they are received on issuing from the glands, venoms always present the appearance of a thick saliva, of an oily consistency and more or less tinged with yellow, according to the species of snake by which the poison has been produced. They are entirely soluble in water, the addition of which renders them opalescent. Tested with litmus they exhibit a slightly acid reaction; this acidity, which is due to the presence of a very small quantity of an indeterminate volatile acid, disappears on desiccation, so that solutions of dried venom are neutral. The taste of venoms is very bitter. Their density, which is slightly greater than that of water, varies from 1030 to 1050.
Venoms are composed of a mixture, in variable proportions, of proteid substances, mucus and epithelialdébris, fatty matters and salts (chlorides and phosphates of lime, ammonia and magnesia), with from 65 to 80 per cent. of water.
The elementary analysis of Cobra-venom made by H. Armstrong7gave the following results:—
Not much is to be learnt from these figures; it would be of far greater importance to know the exact constitution of theproteid substances to which venom owes its physiological properties. Unfortunately, our knowledge of the chemistry of the albuminoid matters is still too imperfect for it to be possible for us to determine their nature.
As early as 1843 it was pointed out by Lucien Bonaparte that in the venom ofVipera berusthe most important principle is a proteid substance to which he gave the name ofviperinorechidnin, and which he compared to the digestive ferments. Later on Weir Mitchell and Reichert, and subsequently Norris Wolfenden, Pedlar, Wall, Kanthack, C. J. Martin, and MacGarvie Smith, showed that venoms, like diastases, exhibit a great complexity in composition; that all their characteristic toxic constituents are precipitable by absolute alcohol, and that the precipitate, when redissolved in water, recovers the properties possessed by the venom before precipitation.
According to Armand Gautier,8venoms contain alkaloids. The latter may be obtained, in very small amounts, however, by finely pulverizing dried venom with carbonate of soda, and systematically exhausting the mixture with alcoholic ether at a temperature of 50° C. These alkaloids have yielded crystallized chloraurates and chloroplatinates, and slightly deliquescent crystallized chlorhydrates. The latter produce Prussian blue when treated with very dilute ferric salts, and mixed with a little red prussiate. They therefore represent reductive bodies analogous to ptomaines.
Norris Wolfenden did not succeed in extracting these alkaloids from Cobra-venom, whence they had nevertheless been isolated by Armand Gautier. Wolcott Gibbs, and afterwards Weir Mitchell and Reichert, likewise failed to find them inCrotalus-venom. The toxicity of these bases is, moreover, but very slight, for the totality of the alkaloids extracted by A. Gautier from 0·3 gramme of Cobra-venom did not kill a small bird.
It is therefore to thetoxalbuminsthat the toxic properties of venoms are essentially due.
All venoms are not equally affected by heat. The venoms ofColubridæ(Naja,Bungarus,Hoplocephalus,Pseudechis) and those of theHydrophiidæare entirely uninjured by temperatures approaching 100° C., and even boiling for a short time. When the boiling is prolonged, or when venoms are heated beyond 100° C., their toxic power at first diminishes, and then disappears altogether. At 120° C. it is always destroyed.
The venoms ofViperidæ(Lachesis,Crotalus,Vipera) are much less resistant. By heating to the coagulating point of albumin,i.e., to about 70° C., their toxic properties become attenuated, and they are entirely suppressed between 80° and 85° C.Lachesis-venoms are the most sensitive; their toxicity is lost if they be heated beyond 65° C.
On separating the coagulable albumins of the venoms ofColubridæ, by heating to 72° C., followed by filtration, we obtain a perfectly limpid liquid, which is no longer injured by boiling, and in which the toxic substance remains wholly in solution. The albuminous precipitate, when separately collected and washed, is no longer toxic. The clear liquid, after being filtered, is again precipitated by absolute alcohol, and the precipitate, redissolved in an equal quantity of water, is just as toxic as the original filtered liquid.
The venoms ofViperidæ, when coagulated, by heating them to a temperature of 72° C., and filtered, are almost always inert. The albuminous coagula, if washed, redissolved in water, and injected into the most sensitive animals, produce no harmful effect whatever.
The results of dialysis likewise differ when we experiment with the venoms ofColubridæandViperidæ. The former pass slowly through vegetable membranes, and with greater difficulty through animal parchment. The latter do not dialyse.
Filtration through porcelain (Chamberland candle F) does not sensibly modify the toxicity of the venoms ofColubridæ;on the contrary, it diminishes that of the venom ofViperidæby nearly one-half.
By using a special filter at a pressure of 50 atmospheres, C. J. Martin has succeeded in separating from the venom of an AustralianPseudechistwo substances: a non-diffusiblealbuminoid, coagulable at 82° C., and a diffusible, non-coagulablealbumose. The former produces hæmorrhages; the second attacks the nerve-cell of the respiratory centres.
All venoms exhibit most of the chemical reactions characteristic of the proteids:—
Millon’s reaction.
Xantho-proteic reaction(heating with nitric acid and subsequent addition of ammonia = orange coloration).
Biuret reaction(caustic potash and traces of sulphate of copper).
Precipitation by picric acid, disappearing on being heated, reappearing when cooled.
Precipitation bysaturation withchloride of sodium.
Precipitation bysaturation withsulphate of magnesium.
Precipitation bysaturation withammonium sulphate.
Precipitation by a 5 per cent. solution of sulphate of copper.
Precipitation by alcohol.
According to C. J. Martin and MacGarvie Smith, the albumoses of the venoms ofColubridæarehetero-albumoses,proto-albumoses, and perhapsdeutero-albumosesin small quantities. They can be separated in the following manner:—
The solution of venom is heated to 90° C., and filtered in order to separate the albumins coagulable by heat. The filtrate, saturated with sulphate of magnesium, is shaken for twelve hours. By this means there is obtained a flocculent precipitate, which is placed upon a filter and washed with a saturated solution of sulphate of magnesium. The filtrate is dialysed for twenty-four hours in a stream of distilled water, and then concentrated, likewise by dialysis, in absolute alcohol. Thus we obtain a few cubic centimetresof liquid, which contains a small quantity ofproteidsin solution. Theseproteidscan be nothing but a mixture ofproto-anddeutero-albumoses with peptones. That there is actually no trace of the latter can easily be ascertained.
Neumeister9has shown that it is impossible to precipitate all theproto-albumosesof a solution by saturation with neutral salts, and, since the filtrate becomes slightly turbid when a few drops of a 5 per cent. solution of sulphate of copper are added to it, we must conclude that it contains a small proportion of theseproto-albumoses.
The deposit retained upon the filter after washing with sulphate of magnesium is redissolved in distilled water, and dialysed for three days. An abundant precipitate then becomes collected in the dialyser. This is centrifuged. The clear liquid is decanted with a pipette, then concentrated by dialysis in absolute alcohol, and finally evaporated at 40° C. until completely desiccated. The solid residue is washed and centrifuged several times in distilled water, after which it is dried on chloride of sodium.
This method enables us to separate two albumoses, both precipitable by saturation with sulphate of magnesium, and belonging to the class ofprimary albumoses: one of these,proto-albumose, is soluble in distilled water, the other,hetero-albumose, is insoluble; but the latter can be dissolved in dilute solutions of neutral salts. These bodies are respectively identical with those obtained by the pepsic digestion of proteids.10
In order to study separately the local and general effects of these different albumoses, C. J. Martin and MacGarvie Smith performed the following experiment:—
They introduced beneath the skin of the belly of a guinea-pig, previously shaved and rendered aseptic, two small pieces of sterilized sponge, about 2 c.mm., one of which was impregnatedwith the solution of proteid, while the other served as control. The two small incisions, one on either side of the median line, were then sutured and covered with collodion. In this way the maximum of local effect and the minimum of general effects was obtained. The solutions of albumoses introduced by this method into the organism produced an enormous œdema, which, in from six to eight hours, extended along the whole side of the abdomen containing the sponge charged with poison.
To test the general toxic effects, the solutions were injected into a vein or into the peritoneal cavity. It was thus found that theproto-andhetero-albumoseskilled the animals in a few hours.
It must therefore be concluded from these facts that the active principles of venom areproto-andhetero-albumoses, the albumins that it contains being devoid of all toxic power.
Many chemical substances modify or destroy venoms, and we shall see in another chapter that several of them, by reason of their properties, may be very usefully employed for the destruction, in the actual wound resulting from a venomous bite, of the venom that has not yet been absorbed in the circulation.
Among these substances the most important are:—
A 1 per cent. solution ofpermanganate of potash(Lacerda).
A 1 per cent. solution ofchloride of gold(Calmette).
Chloride of limeor evenhypochloride of calcium(Calmette), in a solution of 1 in 12, which is augmented, at the moment of use, by 5 to 6 volumes of distilled water, so as to bring it to the standard strength of about 850 cubic centimetres of activechlorineper litre of solution.
A 1 per cent. solution ofchromic acid(Kaufmann).
Saturatedbromized water(Calmette).
A 1 per cent. solution oftrichloride of iodine(Calmette).
All these chemical bodies also modify or destroy the diastases and the microbic toxins. The venoms, although more resistant to the influence of heat, behave, therefore, like these latter, and exhibit the closest affinity with them. Moreover, like all thenormal glandular juices, they possess very manifest zymotic properties, which singularly complicate their physiological action, and upon which we shall dwell later on.
Electricity, employed in the form of continuous electrolytic currents passing through a solution of venom, destroys the toxicity of the latter, because under these conditions there is always formed, at the expense of the salts accompanying the venom, a sufficient quantity of chlorinated products (hypochlorites, chlorates, &c.), and a small amount of ozone, the oxidizing action of which is extremely powerful.
With alternating currents of high frequency, Phisalix, repeating the experiments that Arsonval and Charrin had performed upon diphtheria toxin, thought that he had succeeded in attenuating venom to the point of transforming it into vaccine.11But it has been shown by Marmier that this attenuation was simply the result of thermic actions. When, by means of a suitable arrangement, any rise of temperature was carefully avoided, no modification of toxicity was obtained.12
The influence oflight, which has no effect upon venom preserved in a dry state, is, on the contrary, very marked upon venom in solution. Solutions of venom that are destined for physiological experiments should therefore not be employed without controls, if they be several days old. Apart from the fact that, if care be not taken to render them aseptic, they very soon become contaminated with the germs of all kinds of microbes, it is found that they gradually lose a large part of their activity, especially when they remain in contact with the air. By filtering them through a Chamberland candle and keeping them in the dark, in a refrigerator, in perfectly closed phials, they may be kept unimpaired for several months.
The addition ofglycerinein equal parts to a concentrated solution of venom is also an excellent means of preservation.
Phisalix has shown that the emanations fromradiumattenuate and then destroy the virulence of Cobra- and also of Viper-venom.
“Dry Viper-venom, dissolved inaqua chloroformiin the proportion of 1 in 1,000, is put up in four tubes, three of which are irradiated, the first for six hours, the second for twenty hours, and the third for thirty-six hours. Three guinea-pigs, of equal weight, are inoculated with equal quantities of the irradiated venom; a control receives the non-irradiated venom. The latter dies in ten hours; the animal inoculated from the first tube dies in twelve hours; the one inoculated from the second tube in twenty hours, and the third proves resistant without any symptom of poisoning. A second inoculation produces a transitory lowering of the animal’s temperature by half a degree. At the end of four days it dies after inoculation with a lethal dose.”
The nature of the solvent exerts a great influence upon the action of the emanations from radium: if the same experiment be performed with venom dissolved in a 50 per cent. mixture of glycerine and water, the attenuation is merely relative after six hours.
Auguste Lumière and Joseph Nicolas, of Lyons, conceived the idea of studying the effect upon venom of the prolonged action of the intensecoldproduced by the evaporation of liquid air.13The Cobra-venom employed by these investigators was in solution at a strength of 1 in 1,000. It was submitted to the action of liquid air, partly for twenty-four hours and partly for nine days at -191° C. Its toxicity was in no way diminished.
Lastly, I must mention the recent researches of HideyoNoguchi,14with reference to the photodynamic action ofeosinanderythrosinupon the venoms of the Cobra,Vipera russellii, andCrotalus. It was found by the scientist in question that the toxicity of these various venoms is more or less diminished in the presence of these aniline colours, when the mixtures are insolated. Cobra-venom is the most resistant, just as it is in regard to the other physical or chemical agents. That ofCrotalus, on the contrary, is the least stable.
The bites of poisonous snakes produce very different effects according to the species of snake, the species to which the animal bitten belongs, and according to the situation of the bite. It is therefore necessary to take these various factors into account, in describing the symptoms of poisoning in different animals.
When the quantity of venom introduced into the tissues by the bite of the reptile is sufficient to produce fatal results—which is happily not always the case—the venom manifests its toxic action in two series of phenomena: the first of these is local and affects only the seat and surroundings of the bite; the second, or general series, is seen in the effects produced upon the circulation and nervous system.
It is remarkable to find how great is the importance of the local disorders when the venomous reptile belongs to theSolenoglyphagroup (Viperidæ), while it is almostnilin the case of theProteroglypha(ColubridæandHydrophiidæ).
The effects of general intoxication, on the contrary, are much more intense and more rapid with the venom ofProteroglypha, than with that ofSolenoglypha.
In considering the usual phenomena of snake-poisoning in man, we must therefore take this essential difference into account, anddraw up separately a clinical description of the symptoms observed after a bite from aCobra(Colubridæ), for instance, and another list of those that accompany a bite fromLachesisorVipera berus(Viperidæ).
The bite of aCobra, even of large size, is not very painful; it is characterized especially by numbness, that supervenes in the bitten part, rapidly extends throughout the body, and produces attacks of syncope and fainting. The patient soon experiences a kind of lassitude and irresistible desire to sleep; his legs scarcely support him; he breathes with difficulty and his respiration becomes of the diaphragmatic type.
By degrees the drowsiness and the difficulty of breathing become greater; the pulse, which at first is more rapid, becomes slower and gradually weaker, the mouth contracts, and there is profuse salivation, the tongue appears swollen, the eyelids remain drooping, and, after a few hiccoughs frequently accompanied by vomiting and involuntary emissions of urine or fæcal matter, the unfortunate victim falls into the most profound coma and dies. The pupils react to luminous impressions up to the last moment, and the heart continues to beat sometimes for two hours after respiration has ceased.
All this takes but a few hours, most frequently from two to six or seven, rarely more.
When the reptile by which the bite is inflicted is one of theSolenoglypha, such as aLachesisfor example, the seat of the bite immediately becomes very painful and red, then purple. The surrounding tissues are soon infiltrated with sanguinolent serosity. Sharp pains, accompanied by attacks of cramp, extend towards the base of the limb. The patient complains of intense thirst, and extreme dryness of the mouth and throat; the mucous membranes of the eyes, mouth, and genitalia become congested.
These phenomena often continue for a very long period, even for more than twenty-four hours, and are sometimes accompanied by hæmorrhages from the eyes, mouth, stomach, intestines, or bladder, and by more or less violent delirium.
If the quantity of venom absorbed be sufficient to cause death, the patient exhibits, a few hours after being bitten, stupor, insensibility, and then somnolence, with increasing difficulty of respiration, which ends by becoming stertorous. Loss of consciousness seems complete a good while before coma appears. Asphyxia then ensues, and the heart continues to beat for nearly a quarter of an hour after respiratory movements have entirely ceased.
In certain exceptional cases death is very rapid; it may supervene suddenly in a few minutes, even before the local phenomena have had time to manifest themselves; in this case the venom, having penetrated directly into a vein, has produced almost immediate coagulation of the blood, thus causing the formation of a generalized embolism.
If the venom be introduced in a highly vascular region, or directly into a vein, the result is almost invariably fatal. On the contrary, if the derm be scarcely broken, or if the clothing has acted as a protection, scarcely any absorption will take place. We are here confronted with the same factors of gravity as in the case of bites inflicted upon human beings by animals suffering fromrabies.
In experiments we are able to eliminate all these factors, and to follow in an animal inoculated with a known quantity of venom the whole series of phenomena of poisoning, the intensity of which can be graduated. Let us see, then, how the various animals that it is possible to make use of in laboratories behave with regard to venoms of different origins.
In the monkey, the first apparent sign of the absorption ofCobra-venom, or of the venom of any other species ofColubridæ, is a sort of general lassitude; the eyelids next become half closed. The animal appears to be seeking a suitable spot in which to rest; it gets up again immediately, and walks with a jerky action; itslimbs have a difficulty in supporting it. It is soon attacked by nausea, vomiting and dyspnœa; it rests its head upon the ground, raises it, trying to get breath, and carries its hand to its mouth as if in order to pluck a foreign body from its throat. It totters upon its limbs, and lies down upon its side with its face against the ground. Ptosis increases, and complete asphyxia soon supervenes. The heart continues to beat for some time after respiration has ceased, and then stops in diastole.
Cadaveric rigidity very rapidly sets in, and persists for a long time, even after putrefaction has commenced. During the last moments of life the pupil remains very sensitive; the animal appears to retain unimpaired its sense of hearing and sensibility to pain. The electric excitability of the muscles of the face persists, but that of those of the limbs and body almost entirely disappears. The application of volta-faradic currents from the nape to the diaphragm produces no respiratory movement when asphyxia begins to manifest itself. The sphincters of the bladder and anus relax after a few spasms, which, in case of males, frequently provoke the ejaculation of semen; the urine and fæces immediately escape.
The autopsy reveals slight hæmorrhagic œdema at the point of inoculation, and hyperæmia of all the viscera, especially of the liver and spleen, with, very frequently, small hæmorrhagic patches on the surface of these organs, and on that of the intestine and kidneys. The serous membranes, especially the meninges, endocardium, pleuræ, and peritoneum, exhibit ecchymoses; the lungs are besprinkled with small infarcts, the more numerous the slower the intoxication. The blood remains fluid and laccate.
In poisoning by the venoms ofViperidæ, the hæmorrhagic phenomena appear at the outset, and are more intense. Death is always preceded by a period of asphyxia, indicating that the bulbar nuclei of the pneumogastric nerve have become affected. At the autopsy, however, the blood, instead of remaining fluid, is alwaysfound to be coagulated into a mass in all the vessels; it afterwards gradually becomes redissolved in six or eight hours, and then appears laccate, as after poisoning byCobra-venom, but darker.
All mammals exhibit the same symptoms after inoculation with lethal doses of venom. The same applies to birds; but in the latter the period of asphyxia is much longer, probably on account of the reserves of air accumulated in their air-sacs and pneumatic bones. They gape like pigeons that are being suffocated, rest the tip of the beak on the floor of the cage, and frequently have convulsive spasms of the pharynx, accompanied by flapping of the wings. Small birds and even pigeons are extremely sensitive to venom; fowls are more resistant.
Frogs, thanks to their cutaneous respiration, succumb very slowly. I have seen some survive for thirty hours after being inoculated with a quantity of venom which, when subcutaneously injected into a rabbit, causes death in ten minutes.
Lizards and chameleons succumb very rapidly. Grass Snakes and non-venomous snakes in general withstand doses of venom that in proportion to their weight are fairly large; nevertheless, as indeed we shall see in the sequel, they do not possess any real immunity. It is only poisonous snakes that are unaffected by enormous doses of their own venom, as has already been shown by Fontana, Weir Mitchell, and Viaud Grand Marais. They are, however, quite capable of being poisoned by snakes belonging to altogether different species; strong doses ofCrotalus- orLachesis-venom are fatal to Cobras or Kraits, and, when several poisonous snakes are shut up together in the same cage, they are not infrequently seen to kill each other as the result of repeated bites.
Fishes, which are particularly sensitive to the venom ofHydrophiidæ, readily succumb to inoculation with other venoms, such as that of the Cobra. At Saigon, in 1891, I made experiments upon the action of this latter venom on two specimens of thefighting fishes, that the natives of Annam rear in aquariums in order to witness their combats and make bets on them. The fishes died five hours after intramuscular inoculation with a dose which kills a pigeon in twenty minutes.
Many invertebrates, such as leeches, crayfish, and gastropod molluscs (snails), are killed by inoculation with very small quantities of venom.
It is very difficult to specify, even within broad limits, the dose of venom necessary to kill a human being. The quantity of poison introduced by the bite of a venomous snake depends, as has already been stated, upon a large number of factors, and, very fortunately, this quantity is not always sufficient to cause death. Thus in India, that is to say in the region in which snakes are most numerous and most dangerous, the mean mortality seems scarcely to exceed 35 to 40 per cent., so far as it is possible to judge from official statistics. But, by experimenting upon animals, and commencing with known doses of venom, which has first been dried and then dissolved again in always the same quantity of physiological saline solution or sterile distilled water, we can determine exactly,for each kind of venom, and for each species of animal, the minimum lethal doseper kilogramme of animal.
The entire series of data collected by investigators who have devoted themselves to this study may be summed up as follows:—
Minimal doses lethal in twenty-four hours for aguinea-pigweighing from 600 to 700 grammes:—
Cobra-venom. Dose lethal in twenty-four hours for different animals:—
Venom ofBungarus cæruleus(Common Krait), according to Elliot, Sillar, and Carmichael.15Minimal lethal doses for:—
Venom ofEnhydrina valakadien(according to Elliot and Fraser).16Minimal lethal doses per kilogramme:—
Venom ofEnhydris curtus:—
Rat 0·0005 to 0·0006 gramme per kilogramme
Venom ofNotechis scutatus(syn.Hoplocephalus curtus; the Tiger Snake of Australia):—
Venom ofVipera russellii(Daboia):—
Venom ofLachesis gramineus(Green Pit-Viper, India):—
Venom ofCrotalus adamanteus(Californian Rattle-Snake):—
It will have been seen from the foregoing figures, that the respective sensitiveness of the dog, cat, rabbit, guinea-pig, rat, mouse, and frog, with regard to the same venom, is in no way proportional to the weight of these animals.
The species mentioned are, per unit of weight, more or less resistant to intoxication; and, on experimenting with other animals, as for instance the monkey, pig, ass, and horse, we find that the monkey is much more susceptible to intoxication than the dog, and that the ass is extremely sensitive (0·010 gramme of Cobra-venom is sufficient to kill it), while the horse is less so, and the pig is by far the most resistant.
The same weight of dry Cobra-venom, let us say 1 gramme to be precise, will enable us to kill 1,250 kilogrammes of dog, 2,000 kilogrammes of rabbit, 2,500 kilogrammes of guinea-pig, 1,430 kilogrammes of rat, or 8,333 kilogrammes of mouse.
The lethal dose for a horse being, as I have ascertained by my own experiments, about 0·025 gramme, 1 gramme of dry Cobra-venom will therefore suffice to kill 20,000 kilogrammes of horse.
Assuming that man, in proportion to his weight, possesses a resistance intermediate between that of the dog and that of the horse, we may consider that the lethal dose for a human being is about 0·015 gramme. It follows, therefore, that 1 gramme of venom would kill 10,000 kilogrammes of man, or, let us say, 165 persons of an average weight of 60 kilogrammes.
Another extremely important fact, which must not be lost sight of, is that differences of toxicity, which are often considerable, are exhibited by the venoms of different specimens of the same species of snake, or by the venom of the same snake collected at different times. I have found, for instance, in the case of the specimens ofNajaandLachesisreared in my laboratory, that, according to the length of time that the animals had been without food, and to the nearness or otherwise of the moulting period, the venom was more or less active, and that on evaporation it left behind a more or less considerable quantity of dry extract. In certain cases, immediately after the moult and after a prolonged fast, the venom wasten timesmore active than after a plentiful meal or before the moult.
The figures given above must therefore not be regarded as determining the minimal lethal doses of the different venoms, except in a purely comparative way, and they must be considered only as data useful to know when it is desired to experiment upon animals with these substances.
Variations of this kind are observed in the case of all species of snakes. Thus Phisalix rightly insists upon the necessity of always noting, besides the species of snake, the place of origin and the season; for he has himself seen that, as regards French vipers, those of the Jura, for example, produce in the spring a venom almost devoid of local phlogogenic action; while vipers from the vicinity of Clermont-Ferrand, though less toxic, produce much more serious local effects.
On the other hand, it has been shown by Th. Madsen and H. Noguchi, in a very interesting study of venoms and anti-venoms,17that, when we examine the relation between dose and toxicity, we find that the interval separating the moment of inoculation from that of death diminishes only up to a certain point in proportion as the dose is increased. In the case of the guinea-pig, with 0·0005 gramme of Cobra-venom the interval is 3 hours 75 seconds; but after this, an increase in the dose produces only a relatively inconsiderable acceleration of death. There is therefore no strict ratio between the dose inoculated and the time that elapses until death supervenes.
When the quantity of venom introduced into the organism is insufficient to cause death, the phenomena that precede and accompany recovery differ very greatly according as the snake from which the venom was derived belongs to theColubridæorViperidæ.
After a non-lethal bite from a Cobra or Krait, for example, convalescence usually takes place very rapidly, and, apart from the local œdema of the subcutaneous tissue surrounding the wound, which in very many cases leads to the formation of a suppurating abscess, no lasting injury to health is observed. The venom is eliminated by the kidneys, without even causing albuminuria, and sensation gradually returns, in twenty-four or forty-eight hours, in the part affected by the original lesion.
If the bite has been inflicted by a Viperine snake, the local lesion, which is much more extensive, almost always results in the formation of a patch of gangrene. Hæmorrhages from the mucous membranes, and sanguineous suffusions into the serous cavities,such as the pleura or pericardium, may supervene more or less slowly. Pulmonary infarcts are sometimes produced, as well as desquamation and hæmorrhage from the kidneys, albuminuria, or hæmaturia. These lesions, which are more or less severe, last for several days, and then slowly disappear after a period of true convalescence. In many cases they leave behind them traces which last for months and even years, and they then more or less affect the health of the subjects according to the organs that were most seriously affected.
In certain cases, in domestic animals such as dogs, and more rarely in man, after recovery from the bite of a viper, total or partial loss of sight, smell, or hearing, has been observed. Such results, however, are fortunately exceptional.