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In this study, we investigated the action of varespladib (VPL) alone or in combination with a coral snake antivenom (CAV) on the local and systemic effects induced by Micrurus corallinus venom in rats. Adult male Wistar rats were exposed to venom (1.5 mg/kg – i.m.) and immediately treated with CAV (antivenom:venom ratio 1:1.5 ‘v/w’ – i.p.), VPL (0.5 mg/kg – i.p.), or both of these treatments. The animals were monitored for 120 min and then anesthetized to collect blood samples used for haematological and serum biochemical analysis; after euthanasia, skeletal muscle, renal and hepatic tissue samples were collected for histopathological analysis. M. corallinus venom caused local oedema without subcutaneous haemorrhage or apparent necrosis formation, although there was accentuated muscle morphological damage; none of the treatments prevented oedema formation but the combination of CAV and VPL reduced venom-induced myonecrosis. Venom caused neuromuscular paralysis and respiratory impairment in approximately 60 min following envenomation; CAV alone did not prevent the neurotoxic action, whereas VPL alone prevented neurotoxic symptoms developing as did the combination of CAV and VPL. Venom induced significant increase of serum CK and AST release, mostly due to local and systemic myotoxicity, which was partially prevented by the combination of CAV and VPL. The release of hepatotoxic serum biomarkers (LDH and ALP) induced by M. corallinus venom was not prevented by CAV and VPL when individually administered; their combination effectively prevented ALP release. The venom-induced nephrotoxicity (increase in serum creatinine concentration) was prevented by all the treatments. VPL alone or in combination with CAV significantly prevented the venom-induced lymphocytosis. In conclusion, VPL shows to be effective at preventing the neurotoxic, nephrotoxic, and inflammatory activities of M. corallinus venom. In addition, VPL acts synergistically with antivenom to prevent a number of systemic effects caused by M. corallinus venom. Fundação de Amparo à Pesquisa do Estado de São Paulo/[No.2020/04287-6]/FAPESP/Brasil Fundação de Amparo à Pesquisa do Estado de São Paulo/[No.2020/14191-6]/FAPESP/Brasil Conselho Nacional de Desenvolvimento Científico e Tecnológico/[No.309320/2016-0]/CNPq/Brasil University of the West of São Paulo/[No.2020/04287-6]/UNOESTE/Brasil UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto Clodomiro Picado (ICP) UCR::Vicerrectoría de Docencia::Salud::Facultad de Microbiología

Major variations in venom composition can occur between juvenile and adult venomous snakes. However, due to logistical constraints, antivenoms are produced using adult venoms in immunising mixtures, possibly resulting in limited neutralisation of juvenile snake venoms. Daboia russelii is one of the leading causes of snakebite death across South Asia. Its venom is potently procoagulant, causing stroke in prey animals but causing in humans consumptive coagulopathy—a net anticoagulant state—and sometimes death resulting from hemorrhage. In this in vitro study, we compared the venom activity of—and antivenom efficacy against—six 2-week-old D. russelii relative to that of their parents. Using a coagulation analyser, we quantified the relative coagulotoxicity of these venoms in human, avian, and amphibian plasma. The overall potency on human plasma was similar across all adult and neonate venoms, and SII (Serum Institute of India) antivenom was equipotent in neutralising these coagulotoxic effects. In addition, all venoms were also similar in their action upon avian plasma. In contrast, the neonate venoms were more potent on amphibian plasma, suggesting amphibians make up a larger proportion of neonate diet than adult diet. A similar venom potency in human and avian plasmas but varying selectivity for amphibian plasma suggests ontogenetic differences in toxin isoforms within the factor X or factor V activating classes, thereby providing a testable hypothesis for future transcriptomics work. By providing insights into the functional venom differences between adult and neonate D. russelii venoms, we hope to inform clinical treatment of patients envenomated by this deadly species and to shed new light on the natural history of these extremely medically important snakes.

The aim of this study was to evaluate the effects of different doses of alternagin-C, a disintegrin-like protein from Rhinocerophis alternatus venom, on myocardial contractility of the freshwater fish Hoplias malabaricus, an alternative model to contractile function studies. Alternagin-C treatment exhibited a hormetic-like dose-response curve with a strong positive inotropism and enhanced cardiac pumping capacity at low dose, whereas a modest inotropism and a left shift in the force-frequency relationship was registered at high dose.

The scorpions of the Buthidae family exhibit diverse toxins with proven pharmacological activities and yet underexplored. The Hottentotta rugiscutis is a commonly found south-Indian buthid scorpion, whose venom proteomic profile is unknown. In this study, the venom was biochemically and immunologically characterized by SDS-PAGE, MALDI-TOF MS, western blot and ELISA. The regional and seasonal variation in the venom composition from the same species was also assessed at the molecular mass level. The venom was further studied in albino mice to understand its impact on various blood parameters. The venom has varied MW proteins between 6-275 kDa, four of them were found to be major immunodominant proteins. The mass spectra have revealed that some proteins are predominantly present in the venom of 3–4.5 kDa or 6.5–8.0 kDa, which could be the K+ or Na+ channel blockers respectively whose ratio varied by season. The obtained venom-mass spectra could also be used as H. rugiscutis specific finger-print in identifying the region-specific species. The venom was found to elicit a stress-induced innate immune response in mice, giving rise to a strong Th2 mediated humoral immune response. Overall, this study has provided a glimpse of the venom composition and its immunogenicity.

Most traditional research on snake venoms has focused on front-fanged snake families (Viperidae, Elapidae, and Atractaspididae). However, venom is now generally accepted as being a much more broadly possessed trait within snakes, including species traditionally considered harmless. Unfortunately, due to historical inertia and methodological challenges, the toxin repertoires of non-front-fanged snake families (e.g., Colubridae, Dipsadidae, and Natricidae) have been heavily neglected despite the knowledge of numerous species capable of inflicting medically relevant envenomations. Integrating proteomic data for validation, we perform a de novo assembly and analysis of the Duvernoy's venom gland transcriptome of the Central American Road Guarder (Dipsadidae: Xenodontinae: Conophis lineatus), a species known for its potent bite. We identified 28 putative toxin transcripts from 13 toxin families in the Duvernoy's venom gland transcriptome, comprising 63.7% of total transcriptome expression. In addition to ubiquitous snake toxin families, we proteomically confirmed several atypical venom components. The most highly expressed toxins (55.6% of total toxin expression) were recently described snake venom matrix metalloproteases (svMMPs), with 48.0% of svMMP expression contributable to a novel svMMP isoform. We investigate the evolution of the new svMMP isoform in the context of rear-fanged snakes using phylogenetics. Finally, we examine the morphology of the venom apparatus using μCT and explore how the venom relates to autecology and the highly hemorrhagic effects seen in human envenomations. Importantly, we provide the most complete venom characterization of this medically relevant snake species to date, producing insights into the effects and evolution of its venom, and point to future research directions to better understand the venoms of 'harmless' non-front-fanged snakes.

Hydractinia symbiolongicarpus is a colonial hydrozoan that displays a division of labor through morphologically distinct and functionally specialized polyp types. As with all cnidarians, their venoms are housed in nematocysts, which are scattered across an individual. Here, we investigate the spatial distribution of a specific protein family, jellyfish toxins, in which multiple paralogs are differentially expressed across the functionally specialized polyps. Jellyfish toxins (JFTs) are known pore-forming toxins in the venoms of medically relevant species such as box jellyfish (class Cubozoa), but their role in other medusozoan venoms is less clear. Utilizing a publicly available single-cell dataset, we confirmed that four distinct H. symbiolongicarpus JFT paralogs are expressed in nematocyst-associated clusters, supporting these as true venom components in H. symbiolongicarpus. In situ hybridization and immunohistochemistry were used to localize the expression of these JFTs across the colony. These expression patterns, in conjunction with known nematocyst type distributions, suggest that two of these JFTs, HsymJFT1c-I and HsymJFT1c-II, are localized to specific types of nematocysts. We further interpret JFT expression patterns in the context of known regions of nematogenesis and differential rates of nematocyst turnover. Overall, we show that JFT expression patterns in H. symbiolongicarpus are consistent with the subfunctionalization of JFT paralogs across a partitioned venom system within the colony, such that each JFT is expressed within a specific set of functionally distinct polyp types and, in some cases, specific nematocyst types.

The genus Conus includes over 900 species of marine invertebrates known as cone snails, whose venoms are among the most powerful described so far. This potency is mainly due to the concerted action of hundreds of small bioactive peptides named conopeptides, which target different ion channels and membrane receptors and thus interfere with crucial physiological processes. By swiftly harpooning and injecting their prey and predators with such deadly cocktails, the slow-moving cone snails guarantee their survival in the harsh, competitive marine environment. Each cone snail species produces a unique venom, as the mature sequences of conopeptides from the venoms of different species share very little identity. This biochemical diversity, added to the numerous species and conopeptides contained in their venoms, results in an immense biotechnological and therapeutic potential, still largely unexplored. That is especially true regarding the bioprospection of the venoms of cone snail species found off the Brazilian coast - a region widely known for its biodiversity. Of the 31 species described in this region so far, only four - Conus cancellatus, Conus regius, Conus villepinii, and Conus ermineus - have had their venoms partially characterized, and, although many bioactive molecules have been identified, only a few have been actually isolated and studied. In addition to providing an overview on all the cone snail species found off the Brazilian coast to date, this review compiles the information on the structural and pharmacological features of conopeptides and other molecules identified in the venoms of the four aforementioned species, paving the way for future studies.

The Crotalus intermedius group is a clade of rattlesnakes consisting of several species adapted to a high elevation habitat, primarily in México. Crotalus tancitarensis was previously classified as C. intermedius, until individuals occurring on Cerro Tancítaro in Michoacán, México, were reevaluated and classified as a new species (C. tancitarensis) based on scale pattern and geographic location. This study aimed to characterize the venom of C. tancitarensis and compare the venom profile to those of other species within the Crotalus intermedius group using gel electrophoresis, biochemical assays, reverse-phase high performance liquid chromatography, mass spectrometry, and lethal toxicity (LD50) assays. Results show that the venom profiles of species within the Crotalus intermedius group are similar, but with distinct differences in phospholipase A2 (PLA2), metalloproteinase PI (SVMP PI), and kallikrein-like serine proteinase (SVSP) activity and relative abundance. Proteomic analysis indicated that the highland forms produce venoms with 50–60 protein isoforms and a composition typical of type I rattlesnake venoms (abundant SVMPs, lack of presynaptic PLA2-based neurotoxins), as well as a diversity of typical Crotalus venom components such as serine proteinases, PLA2s, C-type lectins, and less abundant toxins (LAAOs, CRiSPs, etc.). The overall venom profile of C. tancitarensis appears most similar to C. transversus, which is consistent with a previous mitochondrial DNA analysis of the Crotalus intermedius group. These rattlesnakes of the Mexican highlands represent a radiation of high elevation specialists, and in spite of divergence of species in these Sky Island habitats, venom composition of species analyzed here has remained relatively conserved. The majority of protein family isoforms are conserved in all members of the clade, and as seen in other more broadly distributed rattlesnake species, differences in their venoms are largely due to relative concentrations of specific components.

This work is aimed to bring insights on the potential sexual dimorphism differences on the venom composition of Bothrops asper and Crotalus simus to expand the knowledge of the venom variability that might improve the antivenom design. Biological characterization of venoms of each sex in both species did not show significant qualitative differences. Considerations on the sexual venom variations in these species are not relevant for choosing the snake donors for venom production.

Understanding the mechanisms that produce cellular cytotoxicity is fundamental in the field of toxicology. Cytotoxic stimuli can include organic toxins such as hemorrhagic snake venom, which can lead to secondary complications such as the development of necrotic tissue and profuse scarring. These clinical manifestations mimic cytotoxic responses induce by other organic compounds such as organic acids. We used hemorrhagic snake venom and human embryonic kidney cells (HEK 293T) as a model system to better understand the cellular responses involved in venom induced cytotoxicity. Cells stimulated with Crotalus atrox (CA) (western diamondback) venom for 4 or 10 h demonstrated significant cytotoxicity. Results from 2',7'-Dichlorodihydrofluorescein diacetate (H2 DCF-DA) assays determine CA venom stimulation induces a robust production of reactive oxygen species (ROS) over a 3-h time course. In contrast, pretreatment with polyethylene glycol (PEG)-catalase or N-acetyl cysteine (NAC) prior to CA venom stimulation significantly blunted H2 DCFDA fluorescence fold changes and showed greater cytoprotective effects than cells stimulated with CA venom alone. Pre- incubating HEK293T cells with the NADPH oxidase (NOX) pan-inhibitor VAS2870 prior venom stimulation significantly minimized the venom-induced oxidative burst at early timepoints (≤2 h). Collectively, our experiments show that pre-application of antioxidants reduces CA venom induce cellular toxicity. This result highlights the importance of ROS in the early stages of cytotoxicity and suggests muting ROS production in noxious injuries may increase positive clinical outcomes.

The Trimeresurus complex consists of diverse medically important venomous pit vipers that cause snakebite envenomation. Antivenoms, however, are in limited supply, and are specific to only two out of the many species across Asia. This study thus investigated the immunoreactivities of regional pit viper antivenoms toward selected Trimeresurus pit viper venoms, and examined the neutralization of their hemotoxic activities. Trimeresurus albolabris Monovalent Antivenom (TaMAV, Thailand) exhibited a higher immunoreactivity than Hemato Bivalent Antivenom (HBAV, raised against Trimeresurus stejnegeri and Protobothrops mucrosquamatus, Taiwan) and Gloydius brevicaudus Monovalent Antivenom (GbMAV, China), attributed to its monovalent nature and conserved antigens in the Trimeresurus pit viper venoms. The venoms showed moderate-to-strong in vitro procoagulant and in vivo hemorrhagic effects consistent with hemotoxic envenomation, except for the Sri Lankan Trimeresurus trigonocephalus venom which lacked hemorrhagic activity. TaMAV was able to differentially neutralize both in vitro and in vivo hemotoxic effects of the venoms, with the lowest efficacy shown against the procoagulant effect of T. trigonocephalus venom. The findings suggest that TaMAV is a potentially useful treatment for envenomation caused by hetero-specific Trimeresurus pit vipers, in particular those in Southeast Asia and East Asia. Clinical study is warranted to establish its spectrum of para-specific effectiveness, and dosages need be tailored to the different species in respective regions.

Inefficacy and associated risks of current antivenom has raised the need for alternative approaches of snakebite management. Aptamers are one such alternative which is being pursued for therapeutic interventions as well as for design of diagnostic kits due to its high specificity. Present study focussed on designing and validating nucleic acid aptamers against snake venom PLA2, a hydrolytic enzyme present in all venomous snakes. The aptamers were designed by adding nucleic acid chain on the surface of Daboxin P, a major PLA2 enzyme of Daboia russelii venom. Binding characteristics of the aptamers were confirmed by docking to Daboxin P as well as acidic and basic PLA2s from different snake species using in silico docking. The aptamers folded into different tertiary structures and bound to the active and Ca2+ binding site of PLA2 enzymes. Molecular dynamics simulation analysis of Daboxin P-aptamer complexes showed that the complexes were stable in an aqueous environment. The electrophoretic mobility shift assay further confirmed the binding of the synthetic aptamers to Daboxin P and other snake venom PLA2 enzymes. The aptamers inhibited the sPLA2 activity with an IC50 value ranging between 0.52 μM and 0.77 μM as well as the anticoagulant activity of Daboxin P. The aptamers could also inhibit the PLA2 activity of Echis carinatus crude venom and anti-coagulant activity of Bungarus caeruleus crude venom, members of big four snakes. However, the aptamers didn't inhibit fibrinogenolytic or proteolytic activity of big four venom as well as the coagulation and hemolytic activities. Thus, aptamers can be rationally designed to inhibit the biochemical and biological activities of snake venom proteins.

Bungarus multicinctus is one of the top ten venomous snakes in China, and its bite causes acute and severe diseases, but its pathophysiology remains poorly elucidated. Thus, an animal model of Bungarus multicinctus bite was established by intramuscular injection of 30μg/kg of Bungarus multicinctus venom, and then the serum metabolites were subsequently screened, identified and validated by ultra-performance liquid chromatography-quadrupole-time of flight-mass spectrometry (UPLC-Q-TOF-MS) methods to explore the potential biomakers and possible metabolic pathways. Untargeted metabolomics analysis showed that 36 and 38 endogenous metabolites levels changed in ESI+ and ESI-, respectively, KEGG pathway analysis showed that 5 metabolic pathways, including mineral absorption, central carbon metabolism in cancer, protein digestion and absorption, aminoacyl-tRNA biosynthesis and ABC transporters might be closely related to Bungarus multicinctus bite. Targeted metabolomics analysis showed that there were significant differences in serum D-proline, L-leucine and L-glutamine after Bungarus multicinctus bite (P < 0.05). In addition, receiver operating characteristic (ROC) analysis showed that the diagnostic efficiency of L-Glutamine was superior to other potential biomarkers and the AUC value was 0.944. Moreover, we found evidence for differences in the pathophysiology of glutamine between Bungarus multicinctus bite group and normal group, specifically with the content of glutamine synthetase (GS) and glutaminase (GLS). Taken together, the current study has successfully established an animal model of Bungarus multicinctus bite, and further identified the links between the metabolic perturbations and the pathophysiology and the potential diagnostic biomakers of Bungarus multicinctus bite, which provided valuable insights for studying the mechanism of Bungarus multicinctus bite.

Various proteins are involved in fish venom toxicity, but limited information is available regarding their structure and mode of action. Here, we analyzed RNA transcripts in the dorsal spine of the devil stinger Inimicus japonicus using next-generation sequencing (NGS), and identified two putative protein toxins, a natterin-like protein (Ij-natterin) and a phospholipase A2 (Ij-PLA2), as well as a previously reported stonustoxin-like protein. The deduced amino acid sequence of Ij-natterin suggested that it acts as a pore-forming toxin through the cooperation of the N-terminal lectin-like domain and the C-terminal pore-forming domain. Ij-PLA2 showed significant homology with secreted Ca2+-dependent PLA2s from snake venom and mammals (sPLA2-I/II). The recombinant Ij-PLA2 protein exhibited PLA2 activity in the absence of Ca2+, in contrast to canonical sPLA2-I/II. Comparison of the amino acid sequences of Ij-PLA2 with the other sPLA2-I/II suggests that the C-terminal extended peptide region of Ij-PLA2 is involved in its Ca2+-independent activity.

Made available in DSpace on 2022-04-28T19:44:45Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-10-30 Medical Research Council Australian Research Council Endoparasitoid wasps use complex biochemical arsenals to suppress the normal humoral and cellular immune responses of their hosts in order to transform them into a suitable environment for development of their eggs and larvae. Venom injected during oviposition is a key component of this arsenal, but the functions of individual venom toxins are still poorly understood. Furthermore, there has been little investigation of the potential biotechnological use of these venom toxins, for example for control of agricultural pests. The endoparasitoid Cotesia flavipes (Hymenoptera: Braconidae) is a biocontrol agent reared in biofactories and released extensively in Brazil to control the sugarcane borer Diatraea saccharalis (Lepidoptera: Crambidae). The objectives of this work were to reveal venom components produced by C. flavipes and explore the function of a major venom peptide, Cf4. Using a combined proteomic/transcriptomic approach, we identified 38 putative venom toxins including both linear and disulfide-rich peptides, hydrolases, protease inhibitors, apolipophorins, lipid-binding proteins, and proteins of the odorant binding families. Because of its high abundance in the venom, we selected Cf4, a 33-residue peptide with three disulfide bonds, for synthesis and further characterization. We found that synthetic Cf4 reduced the capacity of D. saccharalis hemocytes to encapsulate foreign bodies without any effect on phenoloxidase activity, consistent with a role in disruption of the cellular host immune response. Feeding leaves coated with Cf4 to neonate D. saccharalis resulted in increased mortality and significantly reduced feeding compared to caterpillars fed untreated leaves, indicating that Cf4 is a potential candidate for insect pest control through ingestion. This study adds to our knowledge of endoparasitoid wasp venoms composition, host regulation mechanisms and their biotechnological potential for pest management. School of Agricultural and Veterinarian Sciences São Paulo State University (UNESP) Institute for Molecular Bioscience The University of Queensland Australian Research Council Centre of Excellence for Innovations in Peptide and Protein Science The University of Queensland School of Agricultural and Veterinarian Sciences São Paulo State University (UNESP) Australian Research Council: CE200100012

The present study was made to determine the mouse in vivo micronuclei induction capacity of the venom obtained from three Mexican endemic snakes: Ophryacus undulatus, Ophryacus smaragdinus, and Oph...

Graphical abstract, Highlights • A comprehensive review about number of toxins with pharmacological importances. • Toxicological evaluations of several toxins with its targeted protein. • Benchmark of venom peptides with mechanism of action., Venom peptides have been evolving complex therapeutic interventions that potently and selectively modulate a range of targets such as ion channels, receptors, and signaling pathways of physiological processes making it potential therapeutic. Several venom peptides were deduced in vivo for clinical development targeting pain management, diabetes, cardiovascular diseases, antimicrobial activity. Several contributions have been detailed for a clear perspective for a better understanding of venomous animals, their venom, and their pharmacological effects. Here we unravel and summarize the recent advances in wide venom peptides across varieties of species for their therapeutics prospects.

18 páginas, 8 figuras, 1 tabla, Snake venoms are a complex biological mixture of proteins with or without enzymatic activity, peptides, and nucleotides, among other components. It is produced in specialized secretory glands located in the maxillary region, being the result of millions of years of evolution and whose biological functions are defense, immobilization, and digestion of prey. Venoms present intraspecific (i.e., individual, ontogenetic, geographical) and interspecific (i.e., between sympatric and allopatric species) variation, and the study of this variability has become the focus of toxinological research. Bothrops asper is responsible for highest incidence, morbimortality and severe cases of envenoming in Mesoamerica and northern South America. Given its clinical importance, its venom has been characterized and compared qualitatively and quantitatively across the species range. More than 50 years of research show that B. asper venom is endowed with an interesting intraspecific variability. Knowing this variation has allowed advances in the elucidation of the biological role of the venom, a better understanding of the clinical signs and symptoms in patients envenomed by B asper, the immunological implications in the context of antivenoms production, and the generation of new ideas that could be useful to solve different biological and evolutionary questions of one of the venomous snakes with the greatest distribution and strongest public health impact in Latin America.

This study investigates the concerted hepatoprotective effects for three doses of bradykinin potentiating factor (BPF) and/or followed by exposure to a low dose of γ-radiation (LDR) against Naja haje envenoming in rats. Male rats were injected with three consecutive doses of BPF (1 μg/g i.p. for 3 days), followed by exposure to a low dose of gamma radiation (0.5 Gy), and then rats were injected with a dose of Naja haje venom (250 μg/kg i.p.). Results showed that Naja haje causes liver damage, significant elevation of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), cytochrome c, Nitric oxide (NO), malondialdehyde (MDA) and significant depletion in glutathione peroxidase (GPx) contents. In addition, significant depletion in B-cell lymphoma 2 (Bcl-2) and significant elevation in BcL-2 associated X (Bax protein), nuclear factor kappa B (NF-κB), interleukin-1β (IL-1β) in hepatocytes. Bradykinin potentiating factor and/or low dose of γ-radiation caused improvement in liver damage caused by Naja haje venom by a significant decrease in ALT, AST, ALP levels, Bax, cytochrome c, NF-κB, IL-1β, NO and MDA contents, BPF alone or combined with low dose radiation caused a significant increase in Bcl2 and GPx contents. In conclusion, the concerted impact of BPF and LDR may provide an effective venom detoxification tool that helps to reduce hepatic toxicity and extends the lifespan.

The reef stonefish (Synanceia verrucosa) is a venomous fish which causes excruciatingly painful envenomations. While some research on the pathophysiology and functions of the venom have been conducted, there are still some gaps in the understanding of the venom effects due to the extreme lability of fish venom toxins and the lack of available testing platforms. Here we set out to assess new functions of the venom whilst also attempting to address some unclear pathophysiological effects from previous literature. Utilising a biolayer interferometry assay, our results highlight that the venom binds to the orthosteric site of the α-1 nicotinic acetylcholine receptor as well as the domain IV of voltage-gated Ca2+ (CaV1.2) channel mimotopes. Both these results add some clarity to the previously ambiguous literature. We further assessed the coagulotoxic effects of the venom using thromboelastography and Stago STA-R Max coagulation analyser assays. We reveal that the venom produced anticoagulant activity and significantly delayed time until clot formation of recalcified human plasma which is likely through the degradation of phospholipids. There was a difference between fresh and lyophilised venom activity toward the nicotinic acetylcholine receptor mimotopes and coagulation assays, whilst no difference was observed in the activity toward the domain IV of CaV1.2 mimotopes. This research adds further insights into the neglected area of fish venom whilst also highlighting the extreme labile nature of fish venom toxins.

The black mamba (Dendroaspis polylepis) is, due to its extremely toxic venom, one of the most dangerous snake species in Sub-Saharan Africa. A D. polylepis bite is a medical emergency and requires adequate action to prevent severe complications. However, there are no comprehensive reviews available based on clinical cases, and no readily accessible guidelines for standardized treatment. Therefore, we aim to provide an overview regarding the currently available clinical literature on D. polylepis envenomations; in order to promote knowledge on symptomatology and treatment options. We searched for cases reporting humans bitten by D. polylepis in PubMed, Embase, Scopus, and Sabinet. We searched the reference lists of all eligible articles for additional articles. After quality assessment, 29 cases were included in this review. We used descriptive analysis to create an overview of the collected parameters. Among the included case reports and case series, D. polylepis envenomations most frequently resulted in decreased respiratory function, sweating and paralysis. The onset of symptoms usually occurred within 60 minutes. Neurological symptoms occurred more often than symptoms of autonomic dysfunction. In the reported cases most patients (26/29) received antivenom and most survived (25/29). We recommend the reporting of additional structured case reports to improve future analyses on the clinical course of envenomations, in order to improve public health response to D. polylepis envenomations.

The metalloproteinases and phospholipase A2 are the main enzymes in the venom of Macrovipera lebetina obtusa that play a decisive role in the destructive and toxic effects on the organism of the prey. Metalloproteinases cause hemorrhagic damage, destroy the basement membrane of the blood vessel and disrupt the connections between endothelial cells. Phospholipase A2 causes hemolysis of erythrocytes, destroy the cell membranes, and inhibits the adhesion of platelets and so on. The state of the capillaries of the rat brain and microglia under the action of the venom with separately inhibited enzymes was investigated and compared to the action of the crude venom. Also, the toxicity LD50 of the venom of Macrovipera lebetina obtusa with the inhibited enzymatic activity was determined. The histochemical study showed that the inhibition of phospholipase A2 enzymatic activity did not significantly change the vasodestructive effect of the venoms. In case of action of a venom with inhibited enzymatic activity of metalloproteinases, low activity of microglia and less damaged capillaries were observed. The toxicity of the venom with inhibited phospholipase A2 and with inhibited metalloproteinases was respectively 1.8 and 3.7 times weaker than that of the crude venom. We can claim that both the toxicity of the venom of Macrovipera lebetina obtusa, the damaged brain vessels and the increased activity of CNS microglia are determined mainly by the action of metalloproteinases.

Photobiomodulation using light-emitting diode (LED) treatment has analgesic and anti-inflammatory effects which can be an effective therapeutic associated with serum therapy for local treatment of snakebites. Here we explored the effects of LED treatment on isolated macrophage under Bothrops jararacussu venom. Results showed that LED induced IL-6 and TNF-α genes down-regulation and, TGF and ARG1 genes up-regulation which indicates a polarization of macrophages to an M2 phenotype contributing to both tissue repair and resolution of inflammation.

In this study, we examined the potential use of N-acetyl-L-cysteine (NAC) in association with a polyvalent antivenom and as stand-alone therapy to reduce the acute local and systemic effects induced by Lachesis muta muta venom in rats. Male Wistar rats (300–350 g) were exposed to L. m. muta venom (1.5 mg/kg – i.m.) and subsequently treated with anti-Bothrops/Lachesis serum (antivenom:venom ratio 1:3 ‘v/w’ – i.p.) and NAC (150 mg/kg – i.p.) separately or in association; the animals were monitored for 120 min to assess changes in temperature, locomotor activity, local oedema formation and the prevalence of haemorrhaging. After this time, animals were anesthetized in order to collect blood samples through intracardiac puncture and then euthanized for collecting tissue samples; the hematological-biochemical and histopathological analyses were performed through conventional methods. L. m. muta venom produced pronounced local oedema, subcutaneous haemorrhage and myonecrosis, with both antivenom and NAC successfully reducing the extent of the myonecrotic lesion when individually administered; their association also prevented the occurrence of subcutaneous haemorrhage. Venom-induced creatine kinase (CK) release was significantly prevented by NAC alone or in combination with antivenom; NAC alone failed to reduce the release of hepatotoxic (alanine aminotransferase) and nephrotoxic (creatinine) serum biomarkers induced by L. m. muta venom. Venom induced significant increase of leucocytes which was also associated with an increase of neutrophils, eosinophils and monocytes; antivenom and NAC partially reduced these alterations, with NAC alone significantly preventing the increase of eosinophils whereas neither NAC or antivenom prevented the increase in monocytes. Venom did not induce changes in the erythrogram parameters. In the absence of a suitable antivenom, NAC has the potential to reduce a number of local and systemic effects caused by L. m. muta venom.

The peptide, denominated Ct1a, is a β-toxin of 66 amino acids, isolated from venom of the scorpion, Centruroides tecomanus, collected in Colima, Mexico. This toxin was purified using size exclusion, cationic exchange, and reverse phase chromatography. It is the most abundant toxin, representing 1.7% of the soluble venom. Its molecular mass of 7588.9 Da was determined by mass spectrometry. The amino acid sequence was determined by Edman degradation and confirmed by transcriptomic analysis. Since neurons of the suprachiasmatic nucleus (SCN) maintain a spontaneous firing rate (SFR), we evaluated the physiological effects of toxin Ct1a on these neurons. The SFR exhibited a bimodal concentration-dependent response: 100 nM of Ct1a increased the SFR by 223%, whereas 500 nM and 1000 nM reduced it to 42% and 7%, respectively. Control experiments, consisting of recordings of the SFR during a time similar to that used in Ct1a testing, showed stability throughout the trials. Experiments carried out with denatured Ct1a toxin (500 nM) caused no variation in SFR recordings. Action potentials of SCN neurons, before and after Ct1a (100 nM) showed changes in the time constants of depolarization and repolarization phases, amplitude, and half-time. Finally, recordings of hNav1.6 sodium currents indicated that Ct1a shifts the channel activation to a more negative potential and reduces the amplitude of the peak current. These results all demonstrate that toxin Ct1a affects the SFR of SCN neurons by acting upon sodium channels of sub-type 1.6, implicating them in regulation of the SFR of SCN neurons.

Calloselasma rhodostoma (Malayan pit viper) is a medically important snake species that is widely distributed across Southeast Asia. Systemic coagulopathy causing severe haemorrhage and local tissue injury is commonly observed following C. rhodostoma envenoming. However, nephrotoxicity and congestive heart failure were previously reported in a patient who had a long length of hospital stay. In this study, we determined the effect of C. rhodostoma envenoming on cardiovascular disturbances and the associated morphological changes in the liver, heart and kidneys using animal models. We also evaluated the efficacy of Hemato polyvalent antivenom (HPAV; Queen Saovabha Memorial Institute (QSMI) of the Thai Red Cross Society, Thailand) in neutralising the histopathological effects of C. rhodostoma venom. The intravenous (i.v.) administration of C. rhodostoma venom (1000 µg/kg) caused a rapid decrease in mean arterial pressure (MAP) followed by complete cardiac collapse in anaesthetized rats. Moreover, the intraperitoneal (i.p.) administration of C. rhodostoma venom (11.1 mg/kg; 3 × LD50) for 24 h caused cellular lesions in the liver and heart tissues. C. rhodostoma venom also induced nephrotoxicity, as indicated by the presence of tubular injury, interstitial vascular congestion and inflammatory infiltration in the whole area of the kidney. The administration of HPAV, at manufacturer-recommended doses, 15 min prior to or after the addition of C. rhodostoma venom reduced the extent of the morphological changes in the liver, heart and kidneys. This study found that experimental C. rhodostoma envenoming induced cardiovascular disturbances, hepatotoxicity and nephrotoxicity. We also highlighted the potential broad utility of HPAV to neutralise the histopathological effects of C. rhodostoma venom. The early delivery of antivenom appears capable of preventing envenoming outcomes.

Two recently revised Azemiops snakes with apparent differences in their external appearances and skeletal morphologies but unclear genetic boundaries have been proposed. Some researchers have refrained from using the newly proposed taxonomy because these two “species” might be two clades corresponding to different geographical populations of Azemiops feae. To improve the understanding of the kinship of these two Burmese viper groups, more of their characteristics should be explored in depth. We performed a comparative analysis of the proteomic profiles and biochemical activities of snake venoms from these two groups (Sichuan A. feae and Zhejiang A. feae) and evaluated the immunorecognition capacity of commercial antivenoms toward them. Eight protein families were identified in venoms from these two groups, while phospholipase B was only detected in venom from Sichuan A. feae. These protein families displayed varying degrees of differences in relative abundance between venoms, and phospholipase A2 (Sichuan A. feae: 57.15%; Zhejiang A. feae: 65.94%) was the predominated component. Gloydius brevicaudus antivenom exhibited the strongest capacity to immunologically recognize these two venoms, but this was mainly limited to components with high molecular masses, some of which differed between venoms. Additionally, Zhejiang A. feae venom was more toxic than Sichuan A. feae venom, and the venoms expressed remarkable differences in enzymatic activities, probably resulting from the variation in the relative abundance of specific protein families. Our findings unveil differences between the two Burmese viper groups in terms of proteomic profiles, immunoreactivity, and the biochemical functions of their venoms. This information will facilitate the management of snakebites caused by these snakes.

Daboia siamensis (Russell’s viper) is a highly venomous and medically important snake in China, as well as much of Asia. There is minimal information on the pharmacological activity of the venom of the Chinese species, and currently no commercially available specific antivenom in China. This has led to the use of non-specific antivenoms to treat D. siamensis envenomation. In this study, the in vitro neurotoxicity and myotoxicity of D. siamensis venom was examined and the efficacy of four antivenoms was investigated, including the recently developed Chinese D. siamensis monovalent antivenom (C-DsMAV) and three commercially available antivenoms (Thai D. siamensis (Thai-DsMAV) monovalent antivenom, Deinagkistrodon acutus monovalent antivenom (DaAV), and Gloydius brevicaudus monovalent antivenom (GbAV). D. siamensis venom (10–30 µg/mL) caused the concentration-dependent inhibition of indirect twitches in the chick biventer cervicis nerve muscle preparation, without abolishing contractile responses to exogenous agonists ACh or CCh, indicating pre-synaptic neurotoxicity. Myotoxicity was also evident at these concentrations with inhibition of direct twitches, an increase in baseline tension, and the partial inhibition of ACh, CCh, and KCl responses. The prior addition of C-DsMAV or Thai-DsMAV prevented the neurotoxic and myotoxic activity of D. siamensis venom (10 µg/mL). The addition of non-specific antivenoms (GbAV and DaAV) partially prevented the neurotoxic activity of venom (10 µg/mL) but failed to neutralize the myotoxic effects. We have shown that D. siamensis venom exhibits in vitro weak presynaptic neurotoxicity and myotoxicity, which can be prevented by the pre-addition of the Chinese and Thai Russell’s viper antivenoms. Non-specific antivenoms were poorly efficacious. There should be further development of a monospecific antivenom against D. siamensis envenomation in China.

Small RNAs (sRNAs) and microRNAs (miRNAs) are small endogenous noncoding single-stranded RNAs that regulate gene expression in eukaryotes. Experiments in mice and humans have revealed that a typical small RNA can affect the expression of a wide range of genes, implying that small RNAs function as global regulators. Here, we used small RNA deep sequencing to investigate how jararhagin, a metalloproteinase toxin produced from the venom of Bothrops jararaca, affected mmu-miRNAs expression in mice 2 hours (Jar 2hrs) and 24 hours (Jar 24hrs) after injection compared to PBS control. The findings revealed that seven mmu-miRNAs were substantially differentially expressed (p value (p (Corr) cut-off 0.05, fold change ≥ 2) at 2 hrs after jararhagin exposure and that the majority of them were upregulated when compared to PBS. In contrast to these findings, a comparison of Jar 24hrs vs. PBS 24hrs demonstrated that the majority of identified mmu-miRNAs were downregulated. Furthermore, the studies demonstrated that mmu-miRNAs can target the expression of several genes involved in the MAPK signaling pathway. The steady antithetical regulation of mmu-miRNAs may correlate with the expression of genes that trigger apoptosis via MAPK in the early stages, and this effect intensifies with time. The findings expand our understanding of the effects of jararhagin on local tissue lesions at the molecular level.

Snake venom is an adaptive ecological trait that has evolved primarily as a form of prey subjugation. Thus, the selection pressure for toxin diversification is exerted by the prey’s physiological targets, with this pressure being particularly acute for specialist feeders, such as the King Cobra species, all of which are snake-prey specialists. However, while extensive research has been undertaken to elucidate key amino acids that guide toxin structure–activity relationships, reciprocal investigations into the specific sites guiding prey-lineage selective effects have been lacking. This has largely been due to the lack of assay systems amenable to systematic amino acid replacements of targeted proteins in the prey’s physiological pathways. To fill this knowledge gap, we used a recently described approach based upon mimotope peptides corresponding to the orthosteric site of nicotinic acetylcholine receptor alpha-1 subunits, a major binding site for snake venom neurotoxins that cause flaccid paralysis. We investigated the venoms of four different types of King Cobra (Cambodian, Javan, Malaysian, and Thai). This approach allowed for the determination of the key amino acid positions in King Cobra snake prey that are selectively bound by the toxins, whereby replacing these amino acids in the snake-prey orthosteric site with those from lizards or rats resulted in a significantly lower level of binding by the venoms, while conversely replacing the lizard or rat amino acids with those from the snake at that position increased the binding. By doing such, we identified three negatively charged amino acids in the snake orthosteric site that are strongly bound by the positively charged neurotoxic three-finger toxins found in King Cobra venom. This study, thus, sheds light on the selection pressures exerted by a specialist prey item for the evolution of lineage-selective toxins.

Snakebite envenomation is responsible for over 100,000 deaths and 400,000 cases of disability annually, most of which are preventable through access to safe and effective antivenoms. Snake venom toxins span a wide molecular weight range, influencing their absorption, distribution, and elimination within the body. In recent years, a range of scaffolds have been applied to antivenom development. These scaffolds similarly span a wide molecular weight range and subsequently display diverse pharmacokinetic behaviours. Computational simulations represent a powerful tool to explore the interplay between these varied antivenom scaffolds and venoms, to assess whether a pharmacokinetically optimal antivenom exists. The purpose of this study was to establish a computational model of systemic snake envenomation and treatment, for the quantitative assessment and comparison of conventional and next-generation antivenoms. A two-compartment mathematical model of envenomation and treatment was defined and the system was parameterised using existing data from rabbits. Elimination and biodistribution parameters were regressed against molecular weight to predict the dynamics of IgG, F(ab’)2, Fab, scFv, and nanobody antivenoms, spanning a size range of 15-150 kDa. As a case study, intramuscular envenomation by Naja sumatrana (equatorial spitting cobra) and its treatment using Fab, F(ab’)2, and IgG antivenoms was simulated. Variable venom dose tests were applied to visualise effective antivenom dose levels. Comparisons to existing antivenoms and experimental rescue studies highlight the large dose reductions that could result from recombinant antivenom use. This study represents the first comparative in silico model of snakebite envenomation and treatment.

The jellyfish venom stored in nematocysts contains highly toxic compounds comprising of polypeptides, enzymes and other proteins, which form their chemical defence armoury against predators. We have characterized the proteome of crude venom extract from three bloom-forming scyphozoan jellyfish along the south-west coast of India, Chrysaora caliparea, Cyanea nozakii and Lychnorhiza malayensis using a Quadrupole-Time of Flight (Q/TOF) mass spectrometry analysis. The most abundant toxin identified from Chrysaora caliparea and Lychnorhiza malayensis is similar to the pore-forming toxins and metalloproteinases. A protective antioxidant enzyme called peroxiredoxin was found abundantly in Cyanea nozakii. Metalloproteinase identified from the C. caliparea shows similarity with the venom of pit viper (Bothrops pauloensis), while that of L. malayensis was similar to the venom of snakes such as the Bothrops insularis and Bothrops asper. Kininogen-1 is a secreted protein, identified for the first time from the jellyfish L. malayensis. The proteome analysis of Cyanea nozakii, Chrysaora caliparea and Lychnorhiza malayensis contained 20, 12, 8 unique proteins, respectively. Our study characterized the proteome map of crude venom extract from L. malayensis and C. caliparea for the first time, and the venom profile is compared with published information elsewhere. Proteomic data from this study has been made available in the public domain.

Snakebite envenoming is responsible for as many as 138 000 deaths annually, making it the world's most lethal neglected tropical disease (NTD). There is an urgent need to improve snakebite treatment, which currently relies on outdated and poorly tolerated biologic antivenoms that are often weakly efficacious, must be given intravenously in a healthcare setting, and are expensive to those who need them the most. Herein we describe the challenges associated with the discovery and development of new snakebite treatments and detail the great potential of venom toxin-inhibiting small molecule drugs. We finish by highlighting successful enabling strategies applied to other NTDs that could be exploited to facilitate the development of next-generation small molecule-based snakebite treatments.

Understanding the time course of venom exposure in snakebite patients is important for the optimisation of treatment including antivenom dose and timing. We aimed to investigate the pharmacokinetics of red-bellied black snake (RBBS; Pseudechis porphyriacus) venom in envenomed patients. Timed venom concentration data were obtained from patients with RBBS envenomation recruited to the Australian Snakebite Project (ASP), including demographics and antivenom treatment. Venom concentrations were measured using an enzyme immunoassay. Data were modelled using NONMEM version 7.3. Uncertainty in venom ���dose��� was accounted for by arbitrarily fixing the average amount to 1 mg and incorporating between-subject variability on relative bioavailability. A scale parameter for venom clearance was implemented to account for the rapid venom clearance following antivenom dosing. A sensitivity analysis was performed to determine the magnitude of venom clearance amplification. There were 457 venom concentrations in 114 patients (median age 41, 2���90 y; 80 male). Antivenom was administered to 54 patients a median of 4.2 h post-bite (0.67 to 32 h). A one-compartment model with first-order absorption and elimination provided the best description of the data. The estimated clearance and volume of distribution were 5.21 L/h and 39.9 L, respectively. The calculated elimination half-life of P. porphyriacus venom from the final pharmacokinetic model was 5.35 �� 0.36 h. The variability in the relative dose of injected venom was 140%. Antivenom administration increased venom clearance by 40-fold. Ten patients showed evidence of a double peak in the absorption profile. The information on the exposure time of venom in the body following envenomation will help improve treatment and the timing of antivenom.

The venom color variation of Crotalus durissus terrificus (Cdt) is attributed to the presence of the toxin L-amino acid oxidase (LAAO). During the venom milking routine of Instituto Butantan, we have noticed that most venoms of captive Cdt specimens show a yellowish color, while most venoms of wild specimens are white. Here we describe a comparative analysis of long-term captive (LTC) and recently wild-caught (RWC) Cdt, focusing on LAAO variation. For the identification of LAAO in individual venoms, four different approaches were employed: evaluation of the enzymatic activity, SDS-PAGE, Western blotting, and ELISA. In addition, mass spectrometry analysis was performed using pooled samples. Although some variation among these methodologies was observed, it was possible to notice that the presence of LAAO was significantly higher in the venom of LTC individuals. LAAO was identified in 60–80% LTC specimens and in only 10–12% of RWC specimens. Furthermore, this enzyme accounts for 5.6% of total venom proteins of LTC Cdt pooled venom, while it corresponds to only 0.7% of RWC Cdt pooled venom. These findings strongly suggest that captive maintenance increases the expression of LAAO in Cdt venom.