Snake Venom Composition Research Articles

Venoms, venomics, antivenomics

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.

Transcriptomic basis for an antiserum against Micrurus corallinus (coral snake) venom

BackgroundMicrurus corallinus (coral snake) is a tropical forest snake belonging to the family Elapidae. Its venom shows a high neurotoxicity associated with pre- and post-synaptic toxins, causing diaphragm paralysis, which may result in death. In spite of a relatively small incidence of accidents, serum therapy is crucial for those bitten. However, the adequate production of antiserum is hampered by the difficulty in obtaining sufficient amounts of venom from a small snake with demanding breeding conditions. In order to elucidate the molecular basis of this venom and to uncover possible immunogens for an antiserum, we generated expressed sequences tags (ESTs) from its venom glands and analyzed the transcriptomic profile. In addition, their immunogenicity was tested using DNA immunization.ResultsA total of 1438 ESTs were generated and grouped into 611 clusters. Toxin transcripts represented 46% of the total ESTs. The two main toxin classes consisted of three-finger toxins (3FTx) (24%) and phospholipases A2 (PLA2s) (15%). However, 8 other classes of toxins were present, including C-type lectins, natriuretic peptide precursors and even high-molecular mass components such as metalloproteases and L-amino acid oxidases. Each class included an assortment of isoforms, some showing evidence of alternative splicing and domain deletions. Five antigenic candidates were selected (four 3FTx and one PLA2) and used for a preliminary study of DNA immunization. The immunological response showed that the sera from the immunized animals were able to recognize the recombinant antigens.ConclusionBesides an improvement in our knowledge of the composition of coral snake venoms, which are very poorly known when compared to Old World elapids, the expression profile suggests abundant and diversified components that may be used in future antiserum formulation. As recombinant production of venom antigens frequently fails due to complex disulfide arrangements, DNA immunization may be a viable alternative. In fact, the selected candidates provided an initial evidence of the feasibility of this approach, which is less costly and not dependent on the availability of the venom.

Comparative venom gland transcriptome surveys of the saw-scaled vipers (Viperidae: Echis) reveal substantial intra-family gene diversity and novel venom transcripts

BackgroundVenom 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.ResultsVenom gland transcriptomes were created for E. pyramidum leakeyi, E. coloratus and E. carinatus sochureki by sequencing ~1000 expressed sequence tags from venom gland cDNA libraries. A standardised methodology allowed a comprehensive intra-genus comparison of the venom gland profiles to be undertaken, including the previously described E. ocellatus transcriptome. Blast annotation revealed the presence of snake venom metalloproteinases, C-type lectins, group II phopholipases A2, serine proteases, L-amino oxidases and growth factors in all transcriptomes throughout the genus. Transcripts encoding disintegrins, cysteine-rich secretory proteins and hyaluronidases were obtained from at least one, but not all, species. A representative group of novel venom transcripts exhibiting similarity to lysosomal acid lipase were identified from the E. coloratus transcriptome, whilst novel metallopeptidases exhibiting similarity to neprilysin and dipeptidyl peptidase III were identified from E. p. leakeyi and E. coloratus respectively.ConclusionThe comparison of Echis venom gland transcriptomes revealed substantial intrageneric venom variation in representations and cluster numbers of the most abundant venom toxin families. The expression profiles of established toxin groups exhibit little obvious association with venom-related adaptations to diet described from this genus. We suggest therefore that alterations in isoform diversity or transcript expression levels within the major venom protein families are likely to be responsible for prey specificity, rather than differences in the representation of entire toxin families or the recruitment of novel toxin families, although the recruitment of lysosomal acid lipase as a response to vertebrate feeding cannot be excluded. Evidence of marked intrageneric venom variation within the medically important genus Echis strongly advocates further investigations into the medical significance of venom variation in this genus and its impact upon antivenom therapy.

Combined snake venomics and venom gland transcriptomic analysis of the ocellated carpet viper, Echis ocellatus

Snakebite in Africa causes thousands of deaths annually and considerable permanent physical disability. The saw-scaled viper, Echis ocellatus, represents the single most medically important snake species in West Africa. To provide a detailed compositional analysis of the venom of E. ocellatus for designing novel toxin-specific immunotherapy and to delineate sequence structure-function relationships of individual toxins, we characterised the venom proteome and the venom gland transcriptome. Whole E. ocellatus venom was fractionated by reverse-phase HPLC, followed by analysis of each chromatographic fraction using a combination of SDS-PAGE, N-terminal sequencing, MALDI-TOF mass fingerprinting, and CID-MS/MS of tryptic peptides. This analysis identified around 35 distinct proteins of molecular masses in the range of 5.5-110 kDa belonging to 8 different toxin families (disintegrin, DC-fragment, phospholipase A(2), cysteine-rich secretory protein, serine proteinase, C-type lectin, l-amino acid oxidase, and Zn(2+)-dependent metalloprotease). Comparison of the toxin composition of E. ocellatus venom determined using a proteomic approach, with the predicted proteome derived from assembly of 1000 EST sequences from a E. ocellatus venom gland cDNA library, shows some differences. Most notably, peptides derived from 26% of the venom proteins could not be ascribed an exact match in the transcriptome. Similarly, 64 (67%) out of the 95 putative toxin clusters reported in the transcriptome did not match to peptides detected in the venom proteome. These data suggest that the final composition of venom is influenced by transcriptional and post-translational mechanisms that may be more complex than previously appreciated. This, in turn, emphasises the value of combining proteomic and transcriptomic approaches to acquire a more complete understanding of the precise composition of snake venom, than would be gleaned from using one analysis alone. From a clinical perspective, the large amount of SVMPs (66.5% of the total venom proteins) is consistent with the haemorrhagic pathology associated with E. ocellatus envenoming. More significantly, whilst the proteomic analysis confirms the majority of these metalloproteinases (58%) belong to the SVMP PIII class, MS/MS derived peptide sequencing also demonstrates a major constituent (32%) of E. ocellatus venom is a PIV-SVMP with a quaternary structure comprising a 48 kDa (Q2UXQ4 or Q2UXQ5) PIII-SVMP subunit, and two 14-16 kDa C-type lectin-like domains [EOC_00087 and EOC_00124] which display similarity to echicetin alpha [P81017] and beta [P81996] subunits.

Complementary DNA sequencing and identification of mRNAs from the venomous gland of Agkistrodon piscivorus leucostoma

To advance our knowledge on the snake venom composition and transcripts expressed in venom gland at the molecular level, we constructed a cDNA library from the venom gland of Agkistrodon piscivorus leucostoma for the generation of expressed sequence tags (ESTs) database. From the randomly sequenced 2112 independent clones, we have obtained ESTs for 1309 (62%) cDNAs, which showed significant deduced amino acid sequence similarity (scores >80) to previously characterized proteins in National Center for Biotechnology Information (NCBI) database. Ribosomal proteins make up 47 clones (2%) and the remaining 756 (36%) cDNAs represent either unknown identity or show BLASTX sequence identity scores of <80 with known GenBank accessions. The most highly expressed gene encoding phospholipase A 2 (PLA 2) accounting for 35% of A. p. leucostoma venom gland cDNAs was identified and further confirmed by crude venom applied to sodium dodecyl sulfate/polyacrylamide gel electrophoresis (SDS-PAGE) electrophoresis and protein sequencing. A total of 180 representative genes were obtained from the sequence assemblies and deposited to EST database. Clones showing sequence identity to disintegrins, thrombin-like enzymes, hemorrhagic toxins, fibrinogen clotting inhibitors and plasminogen activators were also identified in our EST database. These data can be used to develop a research program that will help us identify genes encoding proteins that are of medical importance or proteins involved in the mechanisms of the toxin venom.

Reappraisal of Vipera aspis Venom Neurotoxicity

BackgroundThe variation of venom composition with geography is an important aspect of intraspecific variability in the Vipera genus, although causes of this variability remain unclear. The diversity of snake venom is important both for our understanding of venomous snake evolution and for the preparation of relevant antivenoms to treat envenomations. A geographic intraspecific variation in snake venom composition was recently reported for Vipera aspis aspis venom in France. Since 1992, cases of human envenomation after Vipera aspis aspis bites in south-east France involving unexpected neurological signs were regularly reported. The presence of genes encoding PLA2 neurotoxins in the Vaa snake genome led us to investigate any neurological symptom associated with snake bites in other regions of France and in neighboring countries. In parallel, we used several approaches to characterize the venom PLA2 composition of the snakes captured in the same areas.Methodology/Principal FindingsWe conducted an epidemiological survey of snake bites in various regions of France. In parallel, we carried out the analysis of the genes and the transcripts encoding venom PLA2s. We used SELDI technology to study the diversity of PLA2 in various venom samples. Neurological signs (mainly cranial nerve disturbances) were reported after snake bites in three regions of France: Languedoc-Roussillon, Midi-Pyrénées and Provence-Alpes-Côte d'Azur. Genomes of Vipera aspis snakes from south-east France were shown to contain ammodytoxin isoforms never described in the genome of Vipera aspis from other French regions. Surprisingly, transcripts encoding venom neurotoxic PLA2s were found in snakes of Massif Central region. Accordingly, SELDI analysis of PLA2 venom composition confirmed the existence of population of neurotoxic Vipera aspis snakes in the west part of the Massif Central mountains.Conclusions/SignificanceThe association of epidemiological studies to genetic, biochemical and immunochemical analyses of snake venoms allowed a good evaluation of the potential neurotoxicity of snake bites. A correlation was found between the expression of neurological symptoms in humans and the intensity of the cross-reaction of venoms with anti-ammodytoxin antibodies, which is correlated with the level of neurotoxin (vaspin and/or ammodytoxin) expression in the venom. The origin of the two recently identified neurotoxic snake populations is discussed according to venom PLA2 genome and transcriptome data.

Eggs-Only Diet: Its Implications for the Toxin Profile Changes and Ecology of the Marbled Sea Snake (Aipysurus eydouxii)

Studies so far have correlated the variation in the composition of snake venoms with the target prey population and snake's diet. Here we present the first example of an alternative evolutionary link between venom composition and dietary adaptation of snakes. We describe a dinucleotide deletion in the only three finger toxin gene expressed in the sea snake Aipysurus eydouxii (Marbled Sea Snake) venom and how it may have been the result of a significant change in dietary habits. The deletion leads to a frame shift and truncation with an accompanying loss of neurotoxicity. Due to the remarkable streamlining of sea snake venoms, a mutation of a single toxin can have dramatic effects on the whole venom, in this case likely explaining the 50- to 100-fold decrease in venom toxicity in comparison to that of other species in the same genus. This is a secondary result of the adaptation of A. eydouxii to a new dietary habit--feeding exclusively on fish eggs and, thus, the snake no longer using its venom for prey capture. This was parallel to greatly atrophied venom glands and loss of effective fangs. It is interesting to note that a potent venom was not maintained for use in defense, thus reinforcing that the primary use of snake venom is for prey capture.

사독 관련논문의 유형분석(TOXICON 1990-2000)

The study was carried out to investigate the researches of Snake which was published papers in the TOXlCON(1990-2000), one of the most famous Journal of toxicology. And the results were as follows : 1. The number related with Snake is 195papers. 2. There were great papers related wih Cobra, and next is Tigris, Viper, etc. 3. There were great papers related wih protein in the composition of snake venom. 4. There were great papers related wih neurotoxin in the research of poisonous character. 5. There were great papers related wih Viper according to the anticoagulation. 6 Eight papers were published to study the immune response of snake venom. 7. The papers of molecular study of snake venom were seven. 8. The papers of anti-snake venom study were three.

사독(蛇毒)에 대한 문헌적(文獻的) 고찰(考察)

This study was carried out to invastigate the researches of Snake Venom and snakes which used in treatment 1. The fist literature that used the snake for treatment is Shin Nong Ben Cao Jing 2. Composition of Snake Venom is consist of Enzymatic proteins ; Phospholipase A(A1-2), Protease, L-amino acid oxidase etc, and Non-enzymatic proteins ; Crotamine(Cytolysin), Proteolytic factor(Hematoxin), Crotoxin(Neurotoxin) etc. 3. Main toxins in Snake Venom are Hematoxin, Cytolysin, Neurotoxin and Cardiotoxin. Lethal dose 50 value of Agkistrodon brevicaudus is /18g, Agkistrodon saxatilis is /18g, Agkistrodon ussuriensis is /18g, therefore Agkistrodon ussuriensis has strongist Snake Venom of all in Korea. 4. Pharmacological actions of Snake Venom are anticoagulation, thrombolytic function, hypotensor etc. 5. Systemic syndromes and signs after snakebite are Dizziness(25.7%), Vomitting(23.1%), Fever(22%), Visual disturbance(18%), Headache(17.7%) and Dyspnea(17.6%), etc. 6. Local syndrome and sign after snakebite is Discoloration(54.2%), Bleeding(20.2%), Bullae(10.7%), Skinulcer(10.8%), etc. 7. Pathological syndromes after snakebite are WBC increase, Urine protein, Urine sugar, Haematuria and elevation of S-GDT, S-GPT etc. These syndromes are leaded by Hematoxin and Cytolysin. 8. Complication signs after snakebite are Cellulitis, Gastritis, Lympoma, Abscess etc. 9. Common function of Viperidae(Agkistrodon acutus or Zaocys dhumnades etc) is expelling the wind(祛風), removing obstruction in the channels(通絡), antipastic function(止痙). And it is used in order to cure hemiparesis, hemiplegia, facial palsy and CVA disease, etc. 10. Using way of snake for medical treatment is various like Herbal alchol therapy, pill, powder and injection etc. The Study on the Snake Venom should be carried out continuously for using of medical treatment.

Bothrops jararaca snakes produce several bothrojaracin isoforms following an individual pattern

More than one isoform of bothrojaracin (BJC), a potent and specific thrombin inhibitor isolated from Bothrops jararaca venom, has been found in individual venoms collected from adult snakes. Variations in snake venom composition have previously been associated with factors such as age, sex, geographic origin, season of the year and diet. In order to obtain further information concerning individual patterns of expression of BJC isoforms, we have analyzed five individual Bothrops jararaca snake venoms collected at the same time from adult female snakes from the same geographic region. As expected, crude venoms showed a similar migration pattern on SDS-PAGE. BJC was purified using a procedure which includes an affinity chromatography step (PPACK-thrombin Sepharose). A slight variation in the amount of BJC obtained from individual venom samples was noticed. Inhibition of thrombin-induced platelet aggregation as well as migration pattern on SDS-PAGE (under reducing and non-reducing conditions) and isoelectric focusing varied considerably among BJC samples from the five snakes. The amino-terminal sequences (residues 1–34) of individual BJC samples were compared with the sequence deduced from isolated cDNAs encoding α and β chains of BJC. A high degree of homology was detected, although some residues differed from one sample to other. Altogether, data confirmed the heterogeneity found for BJC purified from individual snakes. Thus, the results indicate that: (1) individual specimens of Bothrops jararaca have different patterns of BJC isoform expression; and (2) it seems that genetic factors, at least in part, determine the variability found in BJC production.

Venom yields from several species of colubrid snakes and differential effects of ketamine

The composition of rear-fanged colubrid snake venoms is largely unknown due primarily to the difficulty involved in venom collection. Several different methods have been used to maximize the yield of Duvernoy's secretions. The method proposed by Rosenberg in 1992, which includes the use of ketamine hydrochloride anesthetic and pilocarpine to induce Duvernoy's glands secretion, was used in the present study to collect venom from eight species of colubrids. Protein concentrations, using a dye-binding microassay technique, were determined for the venoms collected. Average protein concentrations ranged from 49.8 to 96.4%. Most yields (dry weight/snake) obtained from specimens in this study were significantly greater than yields previously reported. There was a wide range of effects that occurred due to the ketamine injections; however, all snakes recovered from the effects of the ketamine hydrochloride/pilocarpine with no apparent ill effects. Recommended doses of ketamine hydrochloride have thus been adjusted, depending on previous reactions to the drug. The use of ketamine/pilocarpine in the collection of Duvernoy's secretion has proven to be highly effective in increasing yields. Some caution should be observed when administering ketamine to various species of colubrids, as effects do not necessarily scale to body mass.

Coral snakes of the Americas: biology, identification, and venoms

Part 1 General Information: Overview. Part 2 Form and Distinguishing Features: External Features Internal Features. Part 3 Biology and Evolution: Ecology Feeding and Food Reproduction Enemies and Defense Mimicry - Imitation Fellowships Biogeography, Origin and Evolution. Part 4 Venoms and Snakebite: Venoms in Biology and Their Value Studies of Coral Snake Venom Chemical and Biological Characteristics of Venom Coral Snake Bite Accidents. Part 5 Identification, Description and Distribution By Countries: Identification Description of Species and Subspecies with Keys to the Subspecies Distribution by Countries. Appendices: Scientific and Common Names of Coral Snakes Summary of the Composition of Coral Snake Venom.

Analysis of snake venoms by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and two-dimensional electrophoresis

The protein composition of snake venom has been analysed by sodium dodecyl sulfate—polyacrylamide gel electrophoresis (SDS-PAGE) and high-resolution two-dimensional electrophoresis (2-DE). SDS-PAGE suggests differences between species and similarities between related species. As expected, silver staining greatly enhanced detection sensitivity whilst 2-DE dramatically improved separation to reveal multiple strings of spots indicative of molecular heterogeneity. It is recommended that similarities (or differences) in the SDS-PAGE patterns of venoms from the same (or different) species of snake should be further characterized by 2-DE.

Comparative enzymatic composition of brazilian coral snake ( Micrurus) venoms

1. 1. Venoms of 11 coral snake taxa, including Micrurus albicinctus, M. corallinus, M. frontalis altirostris, M. f. brasiliensis, M. f. frontalis, M. fulvius fulvius, M. ibiboboca, M. lemniscatus ssp., M. rondonianus, M. spixii spixii, and M. surinamensis surinamensis, were examined for 13 enzymatic activities. 2. 2. These were compared with venoms of three outgroup taxa: Naja naja kaouthia, Bungarus multicinctus, and Bothrops moojeni. 3. 3. Enzyme activity levels in Micrurus venoms were highly variable from species to species. 4. 4. All venoms possessed phospholipase activity. 5. 5. Protease activity against synthetic or dyed natural substrates was generally negligible in all elapid venoms examined. By contrast, most Micrurus venoms displayed ample l-leucine aminopeptidase activity. 6. 6. Venom of M.s. surinamensis was significantly different from those of its congeners in most assays.

Immunological (Ouchterlony method) identification of intrasubspecies qualitative differences in snake venom composition

The Ouchterlony method of gel double-diffusion was employed to study the individual venoms of 298 snakes of six Bothrops neuwiedi subspecies. This species is widely distributed and its subdivision permits a more detailed comparative study between the morphology of these animals and the composition of their venoms. The venoms were compared to five standards: three venom pools and two sera. The results indicate that the venom of B. neuwiedi has five basic immunological variants with twenty different well-defined venoms. A single variant can be present in several subspecies; or several variants may be found in a single subspecies. These findings show that a very close relationship between morphology and venom composition does not exist in this species. No attempt was made to see if these immunological variations were correlated with specific biological activities. The geographical distribution of some of the snakes with these variations supports the existence of the variations, and seems to indicate that these variants are probably determined genetically. Staining and photographic improvements for immunoprecipitation are described.

The Antigenic Composition of Ammodytes Viper Venom

Abstract A considerable amount of information on the antigenic composition of snake venoms has been gained from cross-neutralizing effects between different venoms (or partially purified fractions) and their antisera. Grasset (1, 2) concludes, from a large number of cross experiments, that many venoms have an antigenic nucleus in common, but the group specificity does not always correspond to the zoological classification of snakes. Within a group, the venoms are supposed to be individually differentiated by secondary antigenic fractions in different proportions. The antiserum of one venom partially neutralizes all the venoms of the same group, but the effect is more complete for venoms containing more of the secondary components in common with the antigen used for immunization. Césari and Boquet (3) and other authors reach similar conclusions. The validity of these assumptions has not been confirmed directly; nevertheless they have proved very valuable for the preparation of polyvalent antisera.