Snake venoms

Several colubrid snakes produce venomous oral secretions. In this work, the venom collected from Venezuelan opisthoglyphous (rear-fanged) Philodryas olfersii snake was studied. Different proteins were present in its venom and they were characterized by 20% SDS-PAGE protein electrophoresis. The secretion exhibited proteolytic (gelatinase) activity, which was partially purified on a chromatography ionic exchange mono Q2 column. Additionally, the haemorrhagic activity of Philodryas olfersii venom on chicken embryos, mouse skin and peritoneum was demonstrated. Neurotoxic symptoms were demonstrated in mice inoculated with Philodryas olfersii venom. In conclusion, Philodryas olfersii venom showed proteolytic, haemorrhagic, and neurotoxic activities, thus increasing the interest in the high toxic action of Philodryas venom. Key-words: Colubridae. Haemorrhage. Neurotoxic. Philodryas olfersii. Proteolytic activity. Venom.

Snake bite is a result of an unfortunate accidental interaction between a snake and a human victim. It is the single most important toxin-related injury, causing substantial mortality in many parts of the Africa, Asia, and the Americas. A World Health Organization-funded study estimated about 8,000 cases of snakebite with over 20% mortality in Bangladesh annually. A medically important land snake in Bangladesh includes the krait, cobras, and vipers. Medically important snake species are the ones that fall into one of the three categories: bite usually causes death or disability. Snake venom is highly modified saliva (Reptile Venom Research) that is produced by special glands) of certain species of snakes. The glands which secrete the zootoxin are a modification of the parotid salivary gland of other vertebrates, and are usually situated on each side of the head below and behind the eye, encapsulated in a muscular sheath. The glands have large alveoli in which venom is stored before ...

G. ROJAS, J. M. JIM~NEZ and J. M. GUTII~RREZ. Caprylic acid fractionation of hyperimmune horse plasma: description of a simple procedure for antivenom production. Toxicon 32, 351-363, 1994.--A simple methodology for hyperimmune horse plasma fractionation, based on caprylic acid precipitation, is described. Optimal conditions for fractionation were studied; the method gives best results when concentrated caprylic acid was added to plasma, whose pH had been adjusted to 5.8, until a final caprylic acid concentration of 5% was reached. The mixture was vigorously stirred during caprylic acid addition and then for 60 min; afterwards the mixture was filtered. Non-immunoglobulin proteins precipitated in these conditions, whereas a highly enriched immunoglobulin preparation was obtained in the filtrate, which was then dialysed to remove caprylic acid before the addition of NaC1 and phenol. Thus, antivenom was produced after a single precipitation step followed by dialysis. In order to compare this methodology with that based on ammonium sulfate fractionation, a sample of hyperimmune plasma was divided into two aliquots which were fractionated in parallel by both methods. It was found that caprylic acid-fractionated antivenom was superior in terms of yield, production time, albumin/globulin ratio, turbidity, protein aggregates, electrophoretic pattern and neutralizing potency against several activities of Bothrops asper venom. Owing to its efficacy and simplicity, this method could be of great value in antivenom and antitoxin production laboratories.

Bone marrow derived hematopoietic stem cells can function as endothelial progenitor cells. They are recruited to malignant tumors and differentiate into endothelial cells. This mechanism of neovascularization termed vasculogenesis is distinct from proliferation of preexisting vessels. To better understand vasculogenesis we developed a cell culture model with expansion and subsequent endothelial differentiation of human CD133 + progenitor cells in vitro. a v b 3-integrins are expressed by endothelial cells and play a role in the attachment of endothelial cells to the extracellular matrix. We investigated the effect of Cilengitide, a peptide-like, high affinity inhibitor of a v b 3-and a v b 5-integrins in our in vitro system. We could show expression of a v b 3-integrin on 60 ± 9% of non-adherent endothelial progenitors and on 91 ± 7% of differentiated endothelial cells. a v b 3-integrin was absent on CD133 + hematopoietic stem cells. Cilengitide inhibited proliferation of CD133 + cells in a dose-dependent manner. The development of adherent endothelial cells from expanded CD133 + cells was reduced even stronger by Cilengitide underlining its effect on integrin mediated cell adhesion. Expression of endothelial antigens CD144 and von Willebrand factor on differentiating endothelial precursors was decreased by Cilengitide. In summary, Cilengitide inhibits proliferation and differentiation of human endothelial precursor cells underlining its anti-angiogenic effects.

Snake venom hemorrhagic metalloproteinases (SVMPs) of the PI, PII and PIII classes were compared in terms of tissue localization and their ability to hydrolyze basement membrane components in vivo, as well as by a proteomics analysis of exudates collected in tissue injected with these enzymes. Immunohistochemical analyses of co-localization of these SVMPs with type IV collagen revealed that PII and PIII enzymes co-localized with type IV collagen in capillaries, arterioles and post-capillary venules to a higher extent than PI SVMP, which showed a more widespread distribution in the tissue. The patterns of hydrolysis by these three SVMPs of laminin, type VI collagen and nidogen in vivo greatly differ, whereas the three enzymes showed a similar pattern of degradation of type IV collagen, supporting the concept that hydrolysis of this component is critical for the destabilization of microvessel structure leading to hemorrhage. Proteomic analysis of wound exudate revealed similarities and differences between the action of the three SVMPs. Higher extent of proteolysis was observed for the PI enzyme regarding several extracellular matrix components and fibrinogen, whereas exudates from mice injected with PII and PIII SVMPs had higher amounts of some intracellular proteins. Our results provide novel clues for understanding the mechanisms by which SVMPs induce damage to the microvasculature and generate hemorrhage.

The extract of Marsypianthes chamaedrys, a plant used against snakebites, in the present study was shown to inhibit fibrinoclotting induced by several Brazilian snake venoms or thrombin. These data indicate that this extract affected thrombinlike enzymes. In this first report we determine some features of the components present in the extract regarding the antifibrinoclotting action. Our results show that active components responsible for those effects are thermo-resistant and are concentrated in the methanolic fraction.

Protein glycosylation represents one of the most important post-translational events, and is a mean of diversifying a protein without recourse to the genome. The venoms produced by snakes contain an abundance of glycoproteins with N-linked carbohydrates. N-linked glycosylation can ensure the correct folding of important functional domains. Characterization of carbohydrates structures aids in development of human therapeutics by snake venom toxins.

Aqueous extracts, prepared from dried or fresh roots, stems or leaves of Mikania glomerata, a plant found in Mata Atlântica in Southeastern Brazil, were able to efficiently neutralize different toxic, pharmacological, and enzymatic effects induced by venoms from Bothrops and Crotalus snakes. Phospholipase A2 activity and the edema induced by Crotalus durissus terrificus venom were inhibited around 100 and ∼40%, respectively, although this inhibition was only partial for Bothrops venoms. The hemorrhagic activity of Bothrops venoms (Bothrops altenatus, Bothrops moojeni, Bothrops neuwiedi, and Bothrops jararacussu) was significantly inhibited by this vegetal species, while the clotting activity of Crotalus durissus terrificus, Bothrops jararacussu, and Bothrops neuwiedi venoms was totally inhibited. Although, the mechanism of action of Mikania glomerata extract is still unknown, the finding that no visible change was detected in the electrophoretic pattern of snake venom after incubation with the extract excludes proteolytic degradation as a potential mechanism. Since the extract of Mikania glomerata significantly inhibited the studied snake venoms, it may be used as an alternative treatment to serumtherapy and, in addition, as a rich source of potential inhibitors of PLA2s, metalloproteases and serineproteases, enzymes involved in several physiopathological human and animal diseases.

We investigated the putative toxins of Philodryas olfersii (Colubridae), a representative of a family of snakes neglected in venom studies despite their growing medical importance. Transcriptomic data of the venom gland complemented by proteomic analysis of the gland secretion revealed the presence of major toxin classes from the Viperidae family, including serine proteases, metalloproteases, C-type lectins, Crisps, and a C-type natriuretic peptide (CNP). Interestingly, the phylogenetic analysis of the CNP precursor showed it as a linker between two related precursors found in Viperidae and Elapidae snakes. We suggest that these precursors constitute a monophyletic group derived from the vertebrate CNPs.

Se reporta por primera vez la presencia de Melanomys caliginosus Ratón arrocero moreno (Rodentia: Cricetidae) en la dieta de la serpiente Bothrops asper para el Ecuador.  Se encontró un individuo hembra adulto de la serpiente equis o terciopelo recientemente muerta por los trabajadores del sector durante una exploración al bosque de la Hacienda El Mirador en el Sector de Chitoa, Santo Domingo de los Tsáchilas. En su estómago se registró dos individuos en buen estado de Melanomys caliginosus, un macho juvenil y una hembra adulta gestante.

Leishmania parasites of several species cause cutaneous and visceral disease to millions of people worldwide, and treatment for this vector-borne protozoan parasite typically involves administration of highly toxic antimonial drugs. Snake venoms are one of the most concentrated enzyme sources in nature, displaying a broad range of biological effects, and several drugs now used in humans were derived from venoms. In this study, we compared the effects of the venoms of the South American rear-fanged snakes Philodryas baroni (PbV), Philodryas olfersii olfersii (PooV) and Philodryas patagoniensis (PpV), and the North American rear-fanged snakes Hypsiglena torquata texana (HttV) and Trimorphodon biscutatus lambda (TblV), on the growth of Leishmania major, a causative agent of cutaneous leishmaniasis. Different concentrations of each venom were incubated with the log-phase promastigote stage of L. major. TblV showed significant anti-leishmanial activity (IC 50 of 108.6 mg/mL) at its highest concentrations; however, it induced parasite proliferation at intermediate concentrations. PpV was not very active in decreasing the parasitic growth, and a high final concentration (1.7 mg/mL) was necessary to inhibit proliferation by only 51.5% AE 3.6%. PbV, PooV and HttV, at final concentrations of 562, 524 and 438 mg/mL respectively, had no significant effect on L. major growth. The phospholipase A 2 of TblV (trimorphin) was isolated and assayed as for crude venom, and it also exhibited dose-dependent biphasic effects on the parasite culture, with potent cytotoxicity at higher concentrations (IC 50 of 0.25 mM; 3.6 mg/mL) and stimulation of proliferation at very low concentrations. Anti-leishmanial activity of TblV appears to be solely due to the action of trimorphin. This is the first report of anti-leishmanial activity of rear-fanged snake venoms, and these results suggest novel possibilities for discovering new protein-based drugs that might be used as possible agents against leishmaniasis as well as tools to study the biology of Leishmania parasites.

Purity is a characteristic that, together with effectiveness and safety, must be tested to determine the quality of biopharmaceutical products. In therapeutic immunoglobulins, such as human intravenous immunoglobulin (IVIG), purity is evaluated on the basis of physicochemical properties, and is usually assessed by chromatography and electrophoresis. However, in the case of antivenoms these methods fail to discriminate between antibodies towards venom antigens, which constitute the active substance, and antibodies towards non-venom antigens, which are the major impurities in most of the current formulations. The assessment of this aspect of purity requires the use of the immunochemical methods. In this study, it was demonstrated that antivenoms showing physicochemical purity higher than 90% might present immunochemical purity lower than 40%. It is proposed that a comprehensive analysis of antivenom purity should combine physicochemical and immunochemical parameters. In addition, these results are crucial to decide the more appropriate strategies to improve antivenom purity. Taking into account that the current methods of antivenom purification remove most non-antibodies proteins, we propose that efforts must be primarily directed to the improvement of immunization protocols to enhance the antibody response towards venom components in hyperimmunized animals, and secondarily, in the realm of immunoglobulin purification technology.

Why a randomized double blind controlled study is missing ? Why we don’t have impartial independent truly scientific studies ? How an InVitro , claim of the manufacturer will constitute sufficient proof of effectiveness . As always , world play is substituted for actual effectiveness. What studies confirmed , the variations in nature & quality of venoms of snakes of the same species , but existing at geographically / regionally different locations ? 
How come everyone involved at the top are high ranking freemasons , british agents , and many non scientists.   
A major  reason  for  death  from a snakebite  is  fear.
When they say polyvalent antivenoms are only 25% effective ( while that too is actually not proven ) , how come its being administered extensively & promoted as if it is well proven ??
Detoxified ( how a detoxified venom is useful in creating immunity / resistence creating antivenom , since we are trying to create immunity / resistence against toxicity? what processes make it detoxified ? ) venom collected from 10’s of 1000’s of snakes of the same or different species ( for mono / poly valent ) injected into horse or certain other animals to make them immune / resistant  to  those venoms , then their blood is processed to make antivenom – the theory says .  How it is confirmed , these animals develop immunity / resistence to venoms – which study proves that ? How can polyvalent antivenom be effective at all , since the quantity of each will be much lesser than the quantity in monovalent? What about cross interactions between venoms , when put together – nullifying or even creating altogether different substance which could be dangerous or useless?  Is it proven scientifically ?  If at all , the particular animal becomes immune / resistant , what proof do we have to claim that the blood represents the immunity / resistence?  If at all the blood represents immunity / resistence , how do we isolate purified , immunity / resistence carrying material from that blood ? How do we identify / know that , a particular factor is the one carrying immunity / resistence ? What about the person taking the so called antivenom getting infested with the diseases of that animal? .  What about the health issues because of foreign bodies injected into human bodies? . What is the meaning of purification ? How it is made certain , that the so called purified antivenom is really safe , since we don’t know almost anything about what constitute blood? what processes make purified antivenom? 

Even when there is , supposedly,  randomized double blind control studies , proving the effectiveness & safety of medicines , we see them creating havocs and the products being recalled / banned for their previously unforeseen dangers and uselessness .  Since we don’t have any such studies in the cases of antivenom , what confirms the usefulness & safety ?
If death occurs after antivenom vaccine , how to confirm the cause of death –caused from venom or antivenom ??
Antivenom comes from the same people / gang that brought us , unproven virus & preventive vaccines & who claimed elephentiasis is caused by certain kinds of mosqitos & bcg is effective treatment for tuberculosis – All Fraudulent  , Baseless.   

What data do we have on snakes , its actions ?  when the bad effects of antivenom is confirmed / certain , grave , cab even cause death in many cases , do we have any credible truly scientific record proving the usefulness ? when death / complications occur , how do we identify the causes ? fear , effects of antivenom ? how the interpretation / confirmation by the medical cartel /mafia to be trusted ? what data do they have to substantiate their claims ?

More than 95% of the snakes are Non venomous . Of the venomous  ones , they will not be venomous all the time. Even when their fangs are full , mostly they give DRY bites , meaning they bite without injecting venom , seems like they don’t want to waste their venoms unnecessarily . 

Are we talking about truly scientific treatment , in the sense we perceive what science to be or it is modern day witchcraft ??  How much of science is scientific ?

What authority do the medical doctors , ascertaining / confirming these treatment to be truly scientific / useful  , really have . Do they study the concepts in deapth or they are just executives ? Why do we see even the very young doctors with just the basic medical degrees being so assertive on everything medical ( many a times even on non medical ) ? are they knowledgeable at all ?
Indian medical association is a member of world medical association , which has the top most pharma giants run by rothschilds , as their corporate partners . how genuine their intentions will be . bill gates & his wife run the whole show for queen of England & rothschilds .  They are Commanders of British Empire & Padma Vibhushan .
Bill Gates & his father – prominent in Planned Parenthood ,which kills millions of unborn children all over the world & recommending post birth abortion, up to one year after birth ( one of the prominent agent of which is Vegan Peter Singer ). Planned Parenthood, a branch of British Eugenics Society , the utmost racist ,depopulation promoting outfit.

Australia is the stronghold of the front-fanged venomous snake family Elapidae. The Australasian elapid snake radiation, which includes approximately 100 terrestrial species in Australia, as well as Melanesian species and all the world's true sea snakes, may be less than 12 million years old. The incredible phenotypic and ecological diversity of the clade is matched by considerable diversity in venom composition. The clade's evolutionary youth and dynamic evolution should make it of particular interest to toxinologists, however, the majority of species, which are small, typically inoffensive, and seldom encountered by non-herpetologists, have been almost completely neglected by researchers. The present study investigates the venom composition of 28 species proteomically, revealing several interesting trends in venom composition, and reports, for the first time in elapid snakes, the existence of an ontogenetic shift in the venom composition and activity of brown snakes (Pseudonaja sp.). Trends in venom composition are compared to the snakes' feeding ecology and the paper concludes with an extended discussion of the selection pressures shaping the evolution of snake venom.

Snakebites are common in tropical countries like Bangladesh where most snakebite victims dwell in rural areas. Among the
management options after snakebite in Bangladesh, snake charmers (Ozha in Bengali language) are the first contact following
a snakebite for more than 80% of the victims and they are treated mostly with the help of some medicinal plants. Our aim of the
study is to compile plants used for the treatment of snakebite occurrence in Bangladesh.The field survey was carried out in a period
of almost 3 years. Fieldwork was undertaken in Chittagong Hill Tracts, Bangladesh, including Chittagong, Rangamati, Bandarban,
and Khagrachari. Open-ended and semistructured questionnaire was used to interview a total of 110 people including traditional
healers and local people. A total of 116 plant species of 48 families were listed. Leaves were the most cited plant part used against
snake venom. Most of the reported species were herb in nature and paste mostly used externally is the mode of preparation. The
survey represents the preliminary information of certain medicinal plants having neutralizing effects against snake venoms, though
further phytochemical investigation, validation, and clinical trials should be conducted before using these plants as an alternative
to popular antivenom.

This communication revises the state of the art concerning antivenoms against snakes, spiders and scorpions. An overview of the historical facts that preceded the therapeutic use of antibodies is mentioned. A brief list of the major protein components of these venomous animals is revised with a short discussion of what is known on the proteomic analysis of their venoms, but the emphasis is placed on the type of antivenoms available commercially, including pertinent literature and addresses of the companies that prepare these antivenoms. The final section revises and discusses current research on the field and new potential applications that are being developed geared at obtaining new therapeutic antibodies or fragments of antibodies for neutralization of toxic components of venomous animals.

Before the introduction of the first Australian antivenom was the era of the self-styled 'snakemen' and their diverse snakebite remedies. Many received multiple bites from highly dangerous snakes, some of which were deliberately taken to either prove a certain treatment or liven up their show. The mortality rate among these handlers and showmen was high. Production of the first effective Australian antivenom, the tiger snake antivenom, in 1930, began the scientific approach to treating snakebite and opened new frontiers for professional and amateur snake people. Collecting venoms in the development and early production of antivenoms was carried out by a number of professional herpetologists often with little or no reward and in some instances at the ultimate cost of their lives. This paper reviews the most important of those late nineteenth and twentieth century snakemen and their contributions to venom research, antivenom production and current toxinological knowledge. r

The aim of this work was to investigate the involvement of caspases in apoptosis induced by L-amino acid oxidase isolated from Bothrops atrox snake venom. The isolation of LAAO involved three chromatographic steps: molecular exclusion on a G-75 column; ion exchange column by HPLC and affinity chromatography on a Lentil Lectin column. SDS-PAGE was used to confirm the expected high purity level of BatroxLAAO. It is a glycoprotein with 12% sugar and an acidic character, as confirmed by its amino acid composition, rich in "Asp and Glu" residues. It displays high specificity toward hydrophobic L-amino acids. The N-terminal amino acid sequence and internal peptide sequences showed close structural homology to other snake venom LAAOs. This enzyme induces in vitro platelet aggregation, which may be due to H 2 O 2 production by LAAOs, since the addition of catalase completely inhibited the aggregation effect. It also showed cytotoxicity towards several cancer cell lines: HL60, Jurkat, B16F10 and PC12. The cytotoxicity activity was abolished by catalase. A fluorescence microscopy evaluation revealed a significant increase in the apoptotic index of these cells after BatroxLAAO treatment. This observation was confirmed by phosphatidyl serine exposure and activation of caspases. BatroxLAAO is a protein with various biological functions that can be involved in envenomation. Further investigations of its function will contribute to toxicology advances.

Cross-cultural folk beliefs about milk-suckling or milk-drinking amphibians and reptiles have long been noted by scholars. European dialectal folklore, for instance, has countless instances of cow-suckling and milk-stealing animals including butterflies, reptiles, batrachians, hares, hedgehogs and nocturnal birds. These creatures are regularly said to sneak into the domestic space at night to suck life-giving milk or blood from cattle and women. The early documentary evidence for this set of ideas, which relies, in great part, on the motif of breasts or udders suckled by a snake or similar animals such as toads or lizards, has not yet received the study it so richly deserves. Ideally, a comparative study of the milk-suckling reptile (both animal-human and animal-animal) would be carried out across the full gamut of relevant disciplines including ethnology, linguistics, philology, folklore and historical-religious studies: this would naturally include pre-modern written references and an analysis of their transmission. This paper aims to open up new avenues for research on the traditional fondness of snakes for milk, a truly ‘impossible biology’. It is built around several known and little known pre-modern literary and iconographic sources – examples can be found from much of Eurasia – and adopts an interdisciplinary and retrospective comparative method

Snakebite is a serious medical and socio-economic problem affecting the rural and agricultural laborers of tropical and sub-tropical region across the world leading to high morbidity and mortality. In most of the snakebite incidences, victims usually end up with permanent tissue damage and sequelae with high socioeconomic and psychological impacts. Although, mortality has been reduced markedly due to anti-venom regimen, it is associated with several limitations. Snake venom metalloprotease, hyaluronidase and myotoxic phospholipase A2 are the kingpins of tissue necrosis and extracellular matrix degradation. Thus, inhibition of these enzymes is considered to be the rate limiting step in the management of snakebite. Unfortunately, tissue necrosis and extracellular matrix degradation persists even after the administration of anti-venom. At present, inhibitors from snake serum and plasma, several synthetic compounds and their analogs have been demonstrated to possess anti-snake venom activities, but the use of plant metabolites for this purpose has an added advantage of traditional knowledge and will make the treatment cheaper and more accessible to the affected population. Therefore, the clinical and research forums are highly oriented towards plant metabolites and interestingly, certain phytochemicals are implicated as the antibody elicitors against venom toxicity that can be exploited in designing effective anti-venoms. Based on these facts, we have made an effort to enlist plant based secondary metabolites with antiophidian abilities and their mechanism of action against locally acting enzymes/toxins in particular. The review also describes their functional groups responsible for therapeutic beneficial and certainly oblige in designing potent inhibitors against venom toxins.

Many issues and complications in treating snakebite are a result of poor human social, economic and clinical intervention and management. As such, there is scope for significant improvements for reducing incidence and increasing patient outcomes. Snakes do not target humans as prey, but as our dwellings and farms expand ever farther and climate change increases snake activity periods, accidental encounters with snakes seeking water and prey increase drastically. Despite its long history, the snakebite crisis is neglected, ignored, underestimated and fundamentally misunderstood. Tens of thousands of lives are lost to snakebites each year and hundreds of thousands of people will survive with some form of permanent damage and reduced work capacity. These numbers are well recognized as being gross underestimations due to poor to non-existent record keeping in some of the most affected areas. These underestimations complicate achieving the proper recognition of snakebite's socioeconomic impact and thus securing foreign aid to help alleviate this global crisis. Antivenoms are expensive and hospitals are few and far between, leaving people to seek help from traditional healers or use other forms of ineffective treatment. In some cases, cheaper, inappropriately manufactured antivenom from other regions is used despite no evidence for their efficacy, with often robust data demonstrating they are woefully ineffective in neutralizing many venoms for which they are marketed for. Inappropriate first-aid and treatments include cutting the wound, tourniquets, electrical shock, immersion in ice water, and use of ineffective herbal remedies by traditional healers. Even in the developed world, there are fundamental controversies including fasciotomy, pressure bandages, antivenom dosage, premedication such as adrenalin, and lack of antivenom for exotic snakebites in the pet trade. This review explores the myriad of human-origin factors that influence the trajectory of global snakebite causes and treatment failures and illustrate that snakebite is as much a sociological and economic problem as it is a medical one. Reducing the incidence and frequency of such controllable factors are therefore realistic targets to help alleviate the global snakebite burden as incremental improvements across several areas will have a strong cumulative effect.

Vipera ammodytes is the most venomous European snake, whose venom has been used as antigen for immunization of antivenom-producing animals. Same as venom of any other snake, it is a complex mixture of proteins, peptides and other compounds which biochemical and pharmacological variability has been demonstrated at interspecies and intraspecies level. In this work we demonstrated intraspecific variability between 8 venom production batches using both the conventional and the new methodology. Moreover, in contrast to the literature on different venoms' variability, for the first time we were able to select those biochemical differences that are related to and give information on the venom's toxicity and immunogenicity. We have shown that methods quantifying ammodytoxin (the most toxic compound identified so far in the Vipera ammodytes ammodytes venom) content of the venom clearly distinguish between high and low immunogenic venoms.

Snakebite envenoming represents a neglected tropical disease that has a heavy public health impact, particularly in Asia, Africa and Latin America. A global initiative, aimed at increasing antivenom production and accessibility, is being promoted by the World Health Organization and others. This work discusses several aspects of antivenom manufacture and control in which the proteomic analysis of snake venoms, for which the term 'snake venomics' has been coined, might play a relevant supporting role. Snake venomics has already shown its usefulness for generating knowledge at different levels (ontogenetic, individual, and geographic) on inter-and intraspecies venom variability. This information has applications for the quality control of venom preparations used in antivenom manufacture. Moreover, the design of the best venom mixtures for immunization, aimed at increasing the effectiveness of antivenoms, may also be guided by venom proteome analysis, including molecular studies of the cross-reactivity of antivenoms and heterologous venoms through a recently developed methodological approach termed 'antivenomics'. Results generated by proteomic protocols should be complemented by preclinical testing of antivenom efficacy using functional neutralization assays. Snake venomics might be also helpful in designing alternative in vitro tests for the assessment of antivenom efficacy that would eventually substitute current in vivo tests.

Snakebite is a medically and socially significant issue in India, but the quality of treatment and reporting protocols need to be upgraded to international standards. There are currently seven pharmaceutical laboratories in India which produce antivenom against four medically important Indian snake species (cobra (Naja sp.), krait (Bungarus sp.), Russell's viper (Daboia russelii) and saw-scaled viper (Echis carinatus sp.), the 'big four'. Most venom for antivenom production is sourced from Chennai, South India. While the 'big four' are responsible for a majority of serious and fatal bites, the situation is actually much more complex. In this article, we review the production of venom and antivenom in India and suggest areas of improvement. We show that several factors complicate the treatment of snakebite in India. The first is geographic, intra-species variation in venoms of cobras and Russell's vipers. Secondly, there are four species of cobra, eight species of kraits, two distinct sub-species of saw-scaled viper. In view of these observations, it is felt that identifying , evaluating and implementing changes to venom and antivenom production protocols, public education, snakebite treatment and policy in India should be an immediate priority.

A renewed interest in the phenomenon of inter-and intra-species resistance towards the toxicity of snake venoms, coupled with the search for new strategies for treatment of snake envenomations, has prompted the discovery of proteins which neutralize the major toxic components of these venoms. Among these emerging groups of proteins are inhibitors of toxic phospholipases A 2 (PLA 2 s), many of which exhibit a wide range of toxic effects including muscle-tissue damage, neurotoxicity, and inflammation. These proteins have been isolated from both venomous and non-venomous snakes, mammals, and most recently from medicinal plant extracts. The snake blood-derived inhibitors have been grouped into three major classes, a, b, and g, based on common structural motifs found in other proteins with diverse physiological properties. In mammals, DM64, an anti-myotoxic protein isolated from opossum serum, belongs to the immunoglobulin super gene family and is homologous to human a 1 B-glycoprotein and DM43, a metalloproteinase inhibitor from the same organism. In plants, a short note is made of WSG, a newly described anti-toxic-PLA 2 glycoprotein isolated from Withania somnifera (Ashwaganda), a medicinal plant whose aqueous extracts neutralize the PLA 2 activity of the Naja naja venom. The implications of these new groups of PLA 2 toxin inhibitors in the context of our current understanding of snake biology as well as in the development of novel therapeutic reagents in the treatment of snake envenomations worldwide are discussed. q

Highest rates of snake envenoming and mortalities in South Asia, Southeast Asia, Sub-Share Africa with India reporting the most deaths. World Health Organization listed snakebite as a neglected disease. Clinical manifestations of snakebite include overwhelming fear, nausea vomiting diarrhea, vertigo, fainting, tachycardia, and cold, clammy skin. Most snakes cause neurotoxicity. Paresthesia throughout the body, as well as difficulty in speaking and breathing. Several other neurological features have been reported after snake envenomation which is likely to be direct neurotoxic effects. Mechanisms of many these acute manifestations are not clear. Myokymia has been reported mainly from United States following rattlesnake (Crotalusspp). Altered consciousness and deep comma has been reported in 64 % of patient after common krait envenomation. Delayed neurotoxicity also been reported. Anti-venom treatment should be given as soon possible, low dose anti-snake venom (ASV) is not inferior to high dose ASV. Improvement in neurotoxicity has been reported when anti-venom had been administered early. Prevention by avoiding areas heavily infested with snakes. Bite from a dead snake often contains large amount of venom. There is an urgent need for better treatment in neurotoxic envenoming.

G. ROJAS, J. M. JIM~NEZ and J. M. GUTII~RREZ. Caprylic acid fractionation of hyperimmune horse plasma: description of a simple procedure for antivenom production. Toxicon 32, 351-363, 1994.--A simple methodology for hyperimmune horse plasma fractionation, based on caprylic acid precipitation, is described. Optimal conditions for fractionation were studied; the method gives best results when concentrated caprylic acid was added to plasma, whose pH had been adjusted to 5.8, until a final caprylic acid concentration of 5% was reached. The mixture was vigorously stirred during caprylic acid addition and then for 60 min; afterwards the mixture was filtered. Non-immunoglobulin proteins precipitated in these conditions, whereas a highly enriched immunoglobulin preparation was obtained in the filtrate, which was then dialysed to remove caprylic acid before the addition of NaC1 and phenol. Thus, antivenom was produced after a single precipitation step followed by dialysis. In order to compare this methodology with that based on ammonium sulfate fractionation, a sample of hyperimmune plasma was divided into two aliquots which were fractionated in parallel by both methods. It was found that caprylic acid-fractionated antivenom was superior in terms of yield, production time, albumin/globulin ratio, turbidity, protein aggregates, electrophoretic pattern and neutralizing potency against several activities of Bothrops asper venom. Owing to its efficacy and simplicity, this method could be of great value in antivenom and antitoxin production laboratories.

Venom variation occurs at all taxonomical levels and can impact significantly upon the clinical manifestations and efficacy of antivenom therapy following snakebite. Variation in snake venom composition is thought to be subject to strong natural selection as a result of adaptation towards specific diets. Members of the medically important genus Echis exhibit considerable variation in venom composition, which has been demonstrated to co-evolve with evolutionary shifts in diet. We adopt a venom gland transcriptome approach in order to investigate the diversity of toxins in the genus and elucidate the mechanisms which result in prey-specific adaptations of venom composition.

Phospholipases A 2 (PLA 2 s) are commonly found in snake venoms from Viperidae, Hydrophidae and Elaphidae families and have been extensively studied due to their pharmacological and physiopathological effects in living organisms. This article reports a review on natural and artificial inhibitors of enzymatic, toxic and pharmacological effects induced by snake venom PLA 2 s. These inhibitors act on PLA 2 s through different mechanisms, most of them still not completely understood, including binding to specific domains, denaturation, modification of specific amino acid residues and others. Several substances have been evaluated regarding their effects against snake venoms and isolated toxins, including plant extracts and compounds from marine animals, mammals and snakes serum plasma, in addition to poly or monoclonal antibodies and several synthetic molecules. Research involving these inhibitors may be useful to understand the mechanism of action of PLA 2 s and their role in envenomations caused by snake bite. Furthermore, the biotechnological potential of PLA 2 inhibitors may provide therapeutic molecular models with antiophidian activity to supplement the conventional serum therapy against these multifunctional enzymes.

Venoms comprise mixtures of peptides and proteins tailored by Natural Selection to act on vital systems of the prey or victim. Here we review our proteomic protocols for uncoiling the composition, immunological profile, and evolution of snake venoms. Our long-term goal is to gain a deep insight of all viperid venom proteomes. Knowledge of the inter- and intraspecies ontogenetic, individual, and geographic venom variability has applied importance for the design of immunization protocols aimed at producing more effective polyspecific antivenoms. A practical consequence of assessing the cross-reactivity of heterologous antivenoms is the possibility of circumventing the restricted availability of species-specific antivenoms in some regions. Further, the high degree of target specificity makes toxins valuable scaffolds for drug development.