Snake venom
Snake venom is a highly modified saliva containing zootoxins that facilitate the immobilization and digestion of prey, and defense against threats. It is injected by unique fangs during a bite, and some species are also able to spit their venom.
The glands that secrete the zootoxins are a modification of the parotid salivary glands found in other vertebrates, and are usually situated on each side of the head, below and behind the eye, and encapsulated in a muscular sheath. The glands have large alveoli in which the synthesized venom is stored before being conveyed by a duct to the base of channeled or tubular fangs through which it is ejected.
Venoms contain more than 20 different compounds, mostly proteins and polypeptides. A complex mixture of proteins, enzymes, and various other substances with toxic and lethal properties serves to immobilize the prey animal, enzymes play an important role in the digestion of prey, and various other substances are responsible for important but non-lethal biological effects. Some of the proteins in snake venom have very specific effects on various biological functions including blood coagulation, blood pressure regulation, and transmission of the nervous or muscular impulses, and have been developed for use as pharmacological or diagnostic tools, and even useful drugs.
Chemistry
, the son of Lucien Bonaparte, younger brother of Napoleon Bonaparte, was the first to establish the proteinaceous nature of snake venom in 1843.Proteins constitute 90-95% of venom's dry weight and they are responsible for almost all of its biological effects. Among hundreds, even thousands of proteins found in venom, there are toxins, neurotoxins in particular, as well as nontoxic proteins, and many enzymes, especially hydrolytic ones. Enzymes make up 80-90% of viperid and 25-70% of elapid venoms, including digestive hydrolases, L-amino acid oxidase, phospholipases, thrombin-like pro-coagulant, and kallikrein-like serine proteases and metalloproteinases, which damage vascular endothelium. Polypeptide toxins include cytotoxins, cardiotoxins, and postsynaptic neurotoxins, which bind to acetylcholine receptors at neuromuscular junctions. Compounds with low molecular weight include metals, peptides, lipids, nucleosides, carbohydrates, amines, and oligopeptides, which inhibit angiotensin converting enzyme and potentiate bradykinin. Inter- and intra-species variation in venom chemical composition is geographical and ontogenic. Phosphodiesterases interfere with the prey's cardiac system, mainly to lower the blood pressure. Phospholipase A2 causes hemolysis by lysing the phospholipid cell membranes of red blood cells. Amino acid oxidases and proteases are used for digestion. Amino acid oxidase also triggers some other enzymes and is responsible for the yellow colour of the venom of some species. Hyaluronidase increases tissue permeability to accelerate absorption of other enzymes into tissues. Some snake venoms carry fasciculins, like the mambas, which inhibit cholinesterase to make the prey lose muscle control.
| Type | Name | Origin |
| Oxydoreductases | dehydrogenase lactate | Elapidae |
| L-amino-acid oxidase | All species | |
| Catalase | All species | |
| Transferases | Alanine amino transferase | |
| Hydrolases | Phospholipase A2 | All species |
| Lysophospholipase | Elapidae, Viperidae | |
| Acetylcholinesterase | Elapidae | |
| Alkaline phosphatase | Bothrops atrox | |
| Acid phosphatase | Deinagkistrodon acutus | |
| 5'-Nucleotidase | All species | |
| Phosphodiesterase | All species | |
| Deoxyribonuclease | All species | |
| Ribonuclease 1 | All species | |
| Adenosine triphosphatase | All species | |
| Amylase | All species | |
| Hyaluronidase | All species | |
| NAD-Nucleotidase | All species | |
| Kininogenase | Viperidae | |
| Factor-X activator | Viperidae, Crotalinae | |
| Heparinase | Crotalinae | |
| α-Fibrinogenase | Viperidae, Crotalinae | |
| β-Fibrinogenase | Viperidae, Crotalinae | |
| α-β-Fibrinogenase | Bitis gabonica | |
| Fibrinolytic enzyme | Crotalinae | |
| Prothrombin activator | Crotalinae | |
| Collagenase | Viperidae | |
| Elastase | Viperidae | |
| Lyases | Glucosamine ammonium lyase |
Snake toxins vary greatly in their functions. The two broad classes of toxins found in snake venoms are neurotoxins and hemotoxins. However, exceptions occur — the venom of the black-necked spitting cobra, an elapid, consists mainly of cytotoxins, while that of the Mojave rattlesnake, a viperid, is primarily neurotoxic. Both elapids and viperids may carry numerous other types of toxins.
Toxins
Neurotoxins
The beginning of a new neural impulse goes as follows:- An exchange of ions across the nerve cell membrane sends a depolarizing current towards the end of the nerve cell.
- When the depolarizing current arrives at the nerve cell terminus, the neurotransmitter acetylcholine, which is held in vesicles, is released into the space between the two nerves. It moves across the synapse to the postsynaptic receptors.
- ACh binds to the receptors and transfers the signal to the target cell, and after a short time, it is destroyed by acetylcholinesterase.
;Dendrotoxins
;α-neurotoxins
Cytotoxins
;Phospholipases;Cardiotoxins / Cytotoxins
; Hemotoxins
Determining venom toxicity (LD50)
The toxicity of snake venom is assessed by a toxicological test called the median lethal dose, LD50,, which determines the concentration of a toxin required to kill half the members of a tested population of animals. The potency of the venom of wild snakes varies considerably, even within any single species, because of assorted influences such as biophysical environment, physiological status, ecological variables, genetic variation, and various other molecular and ecological evolutionary factors. Such variation necessarily is smaller in captive populations in controlled laboratory settings, though it cannot be eliminated altogether. However, studies to determine snake venom lethality or potency need to be designed to minimize variability, so several techniques have been designed to this end. One approach that is considered particularly helpful is to use 0.1% bovine serum albumin as a diluent in determining LD50 values for various species. It results in far more accurate and consistent LD50 determinations than for example using 0.1% saline as a diluent. Fraction V produces about 95% purified albumin, which is the dried crude venom. Saline as a diluent consistently produces widely varying LD50 results for nearly all venomous snakes; it produces unpredictable variation in the purity of the precipitate. Fraction V is structurally stable because it has seventeen disulfide bonds; it is unique in that it has the highest solubility and lowest isoelectric point of all the major plasma proteins. This makes it the final fraction to be precipitated from its solution. Bovine serum albumin is located in fraction V. The precipitation of albumin is done by reducing the pH to 4.8, which is near the pI of the proteins, and maintaining the ethanol concentration at 40%, with a protein concentration of 1%. Thus, only 1% of the original plasma remains in the fifth fraction. When the ultimate goal of plasma processing is a purified plasma component for injection or transfusion, the plasma component must be highly pure. The first practical large-scale method of blood plasma fractionation was developed by Edwin J. Cohn during World War II. It is known as the Cohn process. This process is also known as cold ethanol fractionation, as it involves gradually increasing the concentration of ethanol in the solution at 5oC and 3oC. The Cohn Process exploits differences in properties of the various plasma proteins, specifically, the high solubility and low pI of albumin. As the ethanol concentration is increased in stages from 0 to 40%, the is lowered from neutral to about 4.8, which is near the pI of albumin. At each stage, certain proteins are precipitated out of the solution and removed. The final precipitate is purified albumin. Several variations to this process exist, including an adapted method by Nitschmann and Kistler that uses fewer steps, and replaces centrifugation and bulk freezing with filtration and diafiltration. Some newer methods of albumin purification add additional purification steps to the Cohn process and its variations. Chromatographic albumin processing as an alternative to the Cohn Process emerged in the early 1980s, however, it was not widely adopted until later due to the inadequate availability of large-scale chromatography equipment. Methods incorporating chromatography generally begin with cryo-depleted plasma undergoing buffer exchange via either diafiltration or buffer exchange chromatography, to prepare the plasma for following ion exchange chromatography steps. After ion exchange, generally further chromatographic purification steps and buffer exchange occur.However, chromatographic methods for separation started being adopted in the early 1980s. Developments were ongoing in the time between when Cohn fractionation started being used, in 1946, and when chromatography started being used, in 1983. In 1962, the Kistler and Nistchmann process was created which was a spin-off of the Cohn process. Chromatographic processes began to take shape in 1983. In the 1990s, the Zenalb and the CSL Albumex processes were created, which incorporated chromatography with a few variations. The general approach to using chromatography for plasma fractionation for albumin is: recovery of supernatant I, delipidation, anion exchange chromatography, cation exchange chromatography, and gel filtration chromatography. The recovered purified material is formulated with combinations of sodium octanoate and sodium N-acetyl tryptophanate and then subjected to viral inactivation procedures, including pasteurization at 60°C. This is a more efficient alternative than the Cohn process for four main reasons: 1) smooth automation and a relatively inexpensive plant was needed, 2) easier to sterilize equipment and maintain a good manufacturing environment, 3) chromatographic processes are less damaging to the albumin protein, and 4) a more successful albumin end result can be achieved. Compared with the Cohn process, the albumin purity went up from about 95% to 98% using chromatography, and the yield increased from about 65% to 85%. Small percentage increases make a difference in regard to sensitive measurements such as purity. The one big drawback in using chromatography has to do with the economics of the process. Although the method was efficient from the processing aspect, acquiring the necessary equipment is a big task. Large machinery is necessary, and for a long time, the lack of equipment availability was not conducive to its widespread use. The components are more readily available now, but it is still a work in progress.
Evolution
Venom evolved just once among all Toxicofera about 170 million years ago, and then diversified into the huge venom diversity seen today. The original toxicoferan venom was a very simple set of proteins that were assembled in a pair of glands. Subsequently, this set of proteins evolved independently in the various lineages of toxicoferans, including Serpentes, Anguimorpha, and Iguania. Several snake lineages have since lost the ability to produce venom, often due to a change in diet or a change in predatory tactics. The evolution of venom is thought to be responsible for the enormous expansion of snakes across the globe.The mechanism of evolution in most cases has been gene duplication in tissues unrelated to the venom, followed by expression of the new protein in the venom gland. This was followed by natural selection for adaptive traits following the birth-and-death model, where duplication is followed by functional diversification, resulting in the creation of structurally related proteins that have slightly different functions. The study of venom evolution has been a high priority for scientists in terms of scientific research, due to the medical relevance of snake venom, in terms of making antivenom and cancer research. Knowing more about the composition of venom and the ways it can potentially evolve is very beneficial. Three main factors affect venom evolution that have been closely studied: predators of the snake that are resistant to snake venom, prey that are in an evolutionary arms race with snakes, and the specific diets that affect the intraspecific evolution of venom. Venoms continue to evolve as specific toxins and are modified to target a specific prey, and toxins are found to vary according to diet in some species.
Rapid venom evolution can also be explained by the arms race between venom-targeted molecules in resistant predators, such as the opossum, and the snake venom that targets the molecules. Scientists performed experiments on the opossums and found that multiple trials showed replacement to silent substitutions in the von Willebrand factor gene that encodes for a venom-targeted hemostatic blood protein. These substitutions are thought to weaken the connection between vWf and a toxic snake venom ligand, which changes the net charge and hydrophobicity. These results are significant to the venom evolution because it is the first citation of rapid evolution in a venom-targeted molecule. This shows that an evolutionary arms race may be occurring in terms of defensive purposes. Alternative hypotheses suggest that venom evolution is due to trophic adaption, whereas these scientists believe, in this case, that selection would occur on traits that help with prey survival in terms of venom evolution instead of predation success. Several other predators of the pit viper show the same type of relationship between snakes, which helps to support the hypothesis that venom has a very strong defensive role along with a trophic role. Which in turn supports the idea that predation on the snakes can be the arms race that produces snake venom evolution.
Some of the various adaptations produced by this process include venom more toxic to specific prey in several lineages, proteins that pre-digest prey, as well as a method to track down prey after a bite. The presence of digestive enzymes in snake venom was once believed to be an adaptation to assist digestion. However, studies of the western diamondback rattlesnake, a snake with highly proteolytic venom, show that venom has no impact on the time required for food to pass through the gut. These various adaptations of venom have also led to considerable debate about the definition of venom and venomous snakes.
Injection
Vipers
In the vipers, which have the most highly developed venom-delivery apparatus, the venom gland is very large and is surrounded by the masseter or temporal muscle, which consists of two bands, the superior arising from behind the eye, the inferior extending from the gland to the mandible. A duct carries venom from the gland to the fang. In vipers and elapids, this groove is completely closed, forming a hypodermic needle-like tube. In other species, the grooves are not covered, or only partially covered. From the anterior extremity of the gland, the duct passes below the eye and above the maxillary bone, to the basal orifice of the venom fang, which is ensheathed in a thick fold of mucous membrane. By means of the movable maxillary bone hinged to the prefrontal bone and connected with the transverse bone, which is pushed forward by muscles set in action by the opening of the mouth, the fang is erected and the venom discharged through the distal orifice. When the snake bites, the jaws close and the muscles surrounding the gland contract, causing venom to be ejected via the fangs.Elapids
In the proteroglyphous elapids, the fangs are tubular, but are short and do not possess the mobility seen in vipers.Colubrids
s have enlarged, grooved teeth situated at the posterior extremity of the maxilla, where a small posterior portion of the upper labial or salivary gland produces venom.Mechanics of biting
Several genera, including Asian coral snakes, burrowing asps, and night adders, are remarkable for having exceptionally long venom glands, extending along each side of the body, in some cases extending posterially as far as the heart. Instead of the muscles of the temporal region serving to press out the venom into the duct, this action is performed by those of the side of the body.Considerable variability in biting behavior is seen among snakes. When biting, viperid snakes often strike quickly, discharging venom as the fangs penetrate the skin, and then immediately release. Alternatively, as in the case of a feeding response, some viperids bite and hold. A proteroglyph or opisthoglyph may close its jaws and bite or chew firmly for a considerable time.
Differences in fang length between the various venomous snakes are likely due to the evolution of different striking strategies.
Mechanics of spitting
of the genera Naja and Hemachatus, when irritated or threatened, may eject streams or a spray of venom a distance of 4 to 8 ft. These snakes' fangs have been modified for the purposes of spitting; inside the fangs, the channel makes a 90° bend to the lower front of the fang. Spitters may spit repeatedly and still be able to deliver a fatal bite.Spitting is a defensive reaction only. The snakes tend to aim for the eyes of a perceived threat. A direct hit can cause temporary shock and blindness through severe inflammation of the cornea and conjunctiva. Although usually no serious symptoms result if the venom is washed away immediately with plenty of water, blindness can become permanent if left untreated. Brief contact with the skin is not immediately dangerous, but open wounds may be vectors for envenomation.
Physiological effects
The four distinct types of venom act on the body differently:- Proteolytic venom dismantles the molecular surroundings, including the bite.
- Hemotoxic venom act on the heart and cardiovascular system.
- Neurotoxic venom acts on the nervous system and brain.
- Cytotoxic venom has a localized action at the site of the bite.
Proteroglyphous snakes
Vipers
Viper venom acts more on the vascular system, bringing about coagulation of the blood and clotting of the pulmonary arteries; its action on the nervous system is not great, no individual group of nerve-cells appears to be picked out, and the effect upon respiration is not so direct; the influence upon the circulation explains the great depression, which is a symptom of viperine envenomation. The pain of the wound is severe and is rapidly followed by swelling and discoloration. The symptoms produced by the bite of the European vipers are thus described by Martin and Lamb:The bite is immediately followed by the local pain of a burning character; the limb soon swells and becomes discolored, and within one to three hours great prostration, accompanied by vomiting, and often diarrhea, sets in. Cold, clammy perspiration is usual. The pulse becomes extremely feeble, and slight dyspnoea and restlessness may be seen. In severe cases, which occur mostly in children, the pulse may become imperceptible and the extremities cold; the patient may pass into coma. In from twelve to twenty-four hours these severe constitutional symptoms usually pass off; but in the meantime, the swelling and discoloration have spread enormously. The limb becomes phlegmonous and occasionally suppurates. Within a few days recovery usually occurs somewhat suddenly, but death may result from the severe depression or from the secondary effects of suppuration. That cases of death, in adults as well as in children, are not infrequent in some parts of the Continent is mentioned in the last chapter of this Introduction.
The Viperidae differ much among themselves in the toxicity of their venoms. Some, such as the Indian Russell's viper and saw-scaled viper ; the American rattlesnakes, bushmasters, and lanceheads ; and the African adders, night adders, and horned vipers, cause fatal results unless a remedy is speedily applied. The bite of the larger European vipers may be very dangerous, and followed by fatal results, especially in children, at least in the hotter parts of the Continent; whilst the small meadow viper, which hardly ever bites unless roughly handled, does not seem to be possessed of a very virulent venom, and although very common in some parts of Austria and Hungary, is not known to have ever caused a serious accident.
Opisthoglyphous colubrids
Biologists had long known that some snakes had rear fangs, 'inferior' venom injection mechanisms that might immobilize prey; although a few fatalities were on record, until 1957, the possibility that such snakes were deadly to humans seemed at most remote. The deaths of two prominent herpetologists, Robert Mertens and Karl Schmidt, from African colubrid bites, changed that assessment, and recent events reveal that several other species of rear-fanged snakes have venoms that are potentially lethal to large vertebrates.Boomslang and twig snake venoms are toxic to blood cells and thin the blood. Early symptoms include headaches, nausea, diarrhea, lethargy, mental disorientation, bruising, and bleeding at the site and all body openings. Exsanguination is the main cause of death from such a bite.
The boomslang's venom is the most potent of all rear-fanged snakes in the world based on LD50. Although its venom may be more potent than some vipers and elapids, it causes fewer fatalities owing to various factors. Symptoms of a bite from these snakes include nausea and internal bleeding, and one could die from a brain hemorrhage and respiratory collapse.
Aglyphous snakes
Experiments made with the secretion of the parotid gland of Rhabdophis and Zamenis have shown that even aglyphous snakes are not entirely devoid of venom, and point to the conclusion that the physiological difference between so-called harmless and venomous snakes is only one of degree, just as various steps exist in the transformation of an ordinary parotid gland into a venom gland or of a solid tooth into a tubular or grooved fang.Use of snake venoms to treat disease
Given that snake venom contains many biologically active ingredients, some may be useful to treat disease.For instance, phospholipases type A2 from the Tunisian vipers Cerastes cerastes and Macrovipera lebetina have been found to have antitumor activity. Anticancer activity has been also reported for other compounds in snake venom. PLA2s hydrolyze phospholipids, thus could act on bacterial cell surfaces, providing novel antimicrobial activities.
The analgesic activity of many snake venom proteins has been long known. The main challenge, however, is to deliver protein to the nerve cells as proteins usually are not applicable as pills.
Immunity
Among snakes
The question whether individual snakes are immune to their own venom has not yet been definitively settled, though an example is known of a cobra that self-envenomated, resulting in a large abscess requiring surgical intervention, but showing none of the other effects that would have proven rapidly lethal in prey species or humans. Furthermore, certain harmless species, such as the North American common kingsnake and the Central and South American mussurana, are proof against the venom of the crotalines, which frequent the same districts, and which they are able to overpower and feed upon. The chicken snake is the enemy of the fer-de-lance in St. Lucia, and in their encounters, the chicken snake is invariably the victor. Repeated experiments have shown the European grass snake not to be affected by the bite of the European adder and the European asp, this being due to the presence, in the blood of the harmless snake, of toxic principles secreted by the parotid and labial glands, and analogous to those of the venom of these vipers. Several North American species of rat snakes, as well as king snakes, have proven to be immune or highly resistant to the venom of rattlesnake species. The king cobra, which does prey on cobras, is said to be immune to their venom.Among other animals
The hedgehog, the mongoose, the honey badger, the opossum, and a few other birds that feed on snakes, are known to be immune to a dose of snake venom. Recently, the honey badger and domestic pig were found to have convergently evolved amino-acid replacements in their nicotinic acetylcholine receptor, which are known to confer resistance to alpha-neurotoxins in hedgehogs. Whether the pig may be considered immune is still uncertain, though early studies show endogenous resistance in pigs tested against neurotoxins. Though the pig's subcutaneous layer of fat may protect it against snake venom, most venoms pass easily through vascular fat layers, making this unlikely to contribute to its ability to resist venoms. The garden dormouse has recently been added to the list of animals refractory to viper venom. Some populations of California ground squirrel are at least partially immune to rattlesnake venom as adults.Among humans
The acquisition of human immunity against snake venom is ancient. Research into development of vaccines that will lead to immunity is ongoing. Bill Haast, owner and director of the Miami Serpentarium, injected himself with snake venom during most of his adult life, in an effort to build up an immunity to a broad array of venomous snakes, in a practice known as mithridatism. Haast lived to age 100, and survived a reported 172 snake bites. He donated his blood to be used in treating snake-bite victims when a suitable antivenom was not available. More than 20 so-treated individuals recovered. Amateur researcher Tim Friede also lets venomous snakes bite him in the hopes of a vaccine against snake venom being developed, and has survived over 160 bites from different species as of January 2016.Traditional treatments
The World Health Organization estimates that 80% of the world's population depends on traditional medicine for their primary health-care needs. Methods of traditional treatments of snakebites, although of questionable efficacy and perhaps even harmful, are nonetheless relevant.Plants used to treat snakebites in Trinidad and Tobago are made into tinctures with alcohol or olive oil and kept in rum flasks called snake bottles, which contain several different plants and/or insects. The plants used include the vine called monkey ladder, which is pounded and put on the bite. Alternatively, a tincture is made with a piece of the vine and kept in a snake bottle. Other plants used include mat root, cat's claw, tobacco, snake bush, obie seed, and wild gri gri root. Some snake bottles also contain the caterpillars that eat tree leaves. Emergency snake medicines are obtained by chewing a three-inch piece of the root of bois canôt and administering this chewed-root solution to the bitten subject. This is a common native plant of Latin America and the Caribbean, which makes it appropriate as an emergency remedy. Another native plant used is mardi gras , which are crushed together with the juice of wild cane and given to the bitten. Quick fixes have included applying chewed tobacco from cigarettes, cigars, or pipes. Making cuts around the puncture or sucking out the venom had been thought helpful in the past, but this course of treatment is now strongly discouraged, due to the risk of self-envenomation through knife cuts or cuts in the mouth.
Serotherapy
Especially noteworthy is progress regarding the defensive reaction by which the blood may be rendered proof against venom, by processes similar to vaccination — antipoisonous serotherapy. Studies have shown that the antitoxic sera do not act as chemical antidotes in destroying the venom, but as physiological antidotes; that, in addition to the venom glands, snakes possess other glands supplying their blood with substances antagonistic to the venom, such as also exist in various animals refractory to snake venom, the hedgehog and the mongoose for instance.Regional venom specificity
Unfortunately, the specificity of the different snake venoms is such that, even when the physiological action appears identical, serum injections or graduated direct inoculations confer immunity towards one species or a few allied species only.Thus, a European in Australia who had become immune to the venom of the deadly Australian tiger snake, manipulating these snakes with impunity, and was under the impression that his immunity extended also to other species, when bitten by a lowland copperhead, an allied elapine, died the following day.
In India, the serum prepared with the venom of monocled cobra Naja kaouthia has been found to be without effect on the venom of two species of kraits, Russell's viper, saw-scaled viper, and Pope's pit viper. Russell's viper serum is without effect on colubrine venoms, or those of Echis and Trimeresurus.
In Brazil, serum prepared with the venom of lanceheads is without action on rattlesnake venom.
Antivenom snakebite treatment must be matched as the type of envenomation that has occurred. In the Americas, polyvalent antivenoms are available that are effective against the bites of most pit vipers. Crofab is the antivenom developed to treat the bite of North American pit vipers. These are not effective against coral snake envenomation, which requires a specific antivenom to their neurotoxic venom. The situation is even more complex in countries such as India, with its rich mix of vipers and highly neurotoxic cobras and kraits of the Elapidae.
This article is based on the 1913 book The Snakes of Europe, by G. A. Boulenger, which is now in the public domain in the United States. Because of its age, the text in this article should not necessarily be viewed as reflecting the current knowledge of snake venom.