- My Adaptive Biotoxicology Lab specialises in reconstructing the evolutionary history of venomous animals, both of the... moreMy Adaptive Biotoxicology Lab specialises in reconstructing the evolutionary history of venomous animals, both of the toxins themselves as well as the glands. This not only sheds light on the evolutionary processes that shape venom evolution but also provides an integrated search strategy for biodiscovery.edit
While elucidating the evolutionary trajectory of green anacondas, we previously documented the existence of two distinct species, Eunectes akayima sp. nov. and Eunectes murinus (Linnaeus, 1758), that separated approximately 10 million... more
While elucidating the evolutionary trajectory of green anacondas, we previously documented the existence of two distinct species, Eunectes akayima sp. nov. and Eunectes murinus (Linnaeus, 1758), that separated approximately 10 million years ago. Our research integrates a novelmolecular clock approach, focuses on tectonic plate movements with fossil records as minimal chronological markers, and offers a refined understanding of speciation events in relation to major biogeographical occurrences in South
America. Mitochondrial DNA analysis demonstrates a significant genetic divergence between the species,bwhich is supported by a notable difference in sexual size dimorphism (SSD) intensity between the two species, along with other morphological differences. This paper also rectifies earlier oversights in the description of the new species and clarifies taxonomic ambiguities in compliance with the International Code of Zoological Nomenclature (henceforth ICZN). In addition, we designate a neotype for E. murinus to stabilize the group. In an effort to honor Indigenous nations, E. akayima sp. nov. derives its name from the Carib language, advocating for the inclusion of traditional names in scientific discourse. Our paper not
only contributes to the taxonomic stability of anacondas but also advocates for the usage of Indigenous names in zoological nomenclature by adopting amore inclusive and flexible approach to the ICZN and eliminating unintended exclusionary practices that we have inherited in science as in other disciplines.
America. Mitochondrial DNA analysis demonstrates a significant genetic divergence between the species,bwhich is supported by a notable difference in sexual size dimorphism (SSD) intensity between the two species, along with other morphological differences. This paper also rectifies earlier oversights in the description of the new species and clarifies taxonomic ambiguities in compliance with the International Code of Zoological Nomenclature (henceforth ICZN). In addition, we designate a neotype for E. murinus to stabilize the group. In an effort to honor Indigenous nations, E. akayima sp. nov. derives its name from the Carib language, advocating for the inclusion of traditional names in scientific discourse. Our paper not
only contributes to the taxonomic stability of anacondas but also advocates for the usage of Indigenous names in zoological nomenclature by adopting amore inclusive and flexible approach to the ICZN and eliminating unintended exclusionary practices that we have inherited in science as in other disciplines.
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Despite their evolutionary novelty, lizard venoms are much less studied in comparison to the intense research on snake venoms. While the venoms of helodermatid lizards have long been assumed to be for defensive purposes, there is... more
Despite their evolutionary novelty, lizard venoms are much less studied in comparison to the intense research on snake venoms. While the venoms of helodermatid lizards have long been assumed to be for defensive purposes, there is increasing evidence of toxic activities more useful for predation than defence (such as paralytic neurotoxicity). This study aimed to ascertain the effects of Heloderma, Lanthanotus, and Varanus lizard venoms on the coagulation and cardiovascular systems. Anticoagulant toxicity was demonstrated for the Varanus species studied, with the venoms prolonging clotting times in human and bird plasma due to the destructive cleavage of fibrinogen. In contrast, thromboelastographic analyses on human and bird plasmas in this study demonstrated a procoagulant bioactivity for Heloderma venoms. A previous study on Heloderma venom using factor-depleted plasmas as a proxy model suggested a procoagulant factor was present that activated either Factor XI or Factor XII, but could not ascertain the precise target. Our activation studies using purified zymogens confirmed FXII activation. Comparisons of neonate and adult H. exasperatum, revealed the neonates to be more potent in the ability to activate FXII, being more similar to the venom of the smaller species H. suspectum than the adult H. exasperatum. This suggests potent FXII activation a basal trait in the genus, present in the small bodied last common ancestor. This also indicates an ontogenetic difference in prey preferences in the larger Heloderma species paralleing the change in venom biochemistry. In addition, as birds lack Factor XII, the ability to clot avian plasma suggested an additional procoagulant site of action, which was revealed to be the activation of Factor VII, with H. horridum being the most potent. This study also examined the effects upon the cardiovascular system, including the liberation of kinins from kininogen, which contributes to hypotension induction. This form of toxicity was previously described for Heloderma venoms, and was revealed in this study was to also be a pathophysiological effect of Lanthanotus and Varanus venoms. This suggests that this toxic activity was present in the venom of the last common ancestor of the anguimorph lizards, which is consistent with kallikrein enzymes being a shared toxin trait. This study therefore uncovered novel actions of anguimorph lizard venoms, not only contributing to the evolutionary biology body of knowledge but also revealing novel activities to mine for drug design lead compounds.
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The study investigated the intraspecific and interspecific variability in venom effects of Agkistro-don viperid snake species and subspecies (eleven venoms total) on plasma clotting times, fibrino-gen levels, and fibrin clot strength.... more
The study investigated the intraspecific and interspecific variability in venom effects of Agkistro-don viperid snake species and subspecies (eleven venoms total) on plasma clotting times, fibrino-gen levels, and fibrin clot strength. Significant delays in plasma clotting time were observed for A. conanti, A. contortrix mokasen, A. contortrix phaeogaster, A. howardgloydi, A. piscivorus leu-costoma, and A. piscivorus piscivorus. Notably the phylogenetically disjunct lineages A. conanti, A. contortrix mokasen and A. howardgloydi exhibited the most potent anticoagulant effects, indicating independent amplification of a basal trait. Inhibition assays with activated clotting enzymes (Factor XIa, IXa, Xa, and thrombin) revealed that FXa inhibition is another basal trait amplified independently on multiple occasions within the genus but with A. howardgloydi notably more po-tent than all others. Phospholipid degradation and zymogen destruction were identified as mechanisms underlying the variability in venom effects observed experimentally and previous clinical reports. Thromboelastography demonstrated that venoms did not clot fibrinogen directly but affected fibrin clot strength by damaging fibrinogen, that thrombin was subsequently only able to cleave into weak, unstable clots. The ability to activate Protein C, an endogenous anticoag-ulant enzyme, varied across species, with some venoms exceeding that of A. contortrix contortrix venom, which previously yielded the protein diagnostic agent Protac®. Phylogenetic analysis suggested that both fibrinogen degradation and Protein C activation were each amplified multi-ple times within the genus, albeit with negative correlation between these two modes of action. This study highlights the evolutionary, clinical, and biodiscovery implications of venom varia-bility in Agkistrodon species, underscoring their dynamic evolution, emphasizing the need for tailored clinical approaches, and highlighting the potential for novel diagnostic and therapeutic developments inspired by the unique properties of snake venoms.
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Snakes in the family Elapidae largely produce venoms rich in three-finger toxins (3FTx) that bind to the α 1 subunit of nicotinic acetylcholine receptors (nAChRs), impeding ion channel activity. These neurotoxins immobilize the prey by... more
Snakes in the family Elapidae largely produce venoms rich in three-finger toxins (3FTx) that bind to the α 1 subunit of nicotinic acetylcholine receptors (nAChRs), impeding ion channel activity. These neurotoxins immobilize the prey by disrupting muscle contraction. Coral snakes of the genus Micrurus are specialist predators who produce many 3FTx, making them an interesting system for examining the coevolution of these toxins and their targets in prey animals. We used a bio-layer interferometry technique to measure the binding interaction between 15 Micrurus venoms and 12 taxon-specific mimotopes designed to resemble the orthosteric binding region of the muscular nAChR subunit. We found that Micrurus venoms vary greatly in their potency on this assay and that this variation follows phylogenetic patterns rather than previously reported patterns of venom composition. The long-tailed Micrurus tend to have greater binding to nAChR orthosteric sites than their short-tailed relatives and we conclude this is the likely ancestral state. The repeated loss of this activity may be due to the evolution of 3FTx that bind to other regions of the nAChR. We also observed variations in the potency of the venoms depending on the taxon of the target mimotope. Rather than a pattern of prey-specificity, we found that mimotopes modeled after snake nAChRs are less susceptible to Micrurus venoms and that this resistance is partly due to a characteristic tryptophan→serine mutation within the orthosteric site in all snake mimotopes. This resistance may be part of a Red Queen arms race between coral snakes and their prey.
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In the ongoing evolutionary arms race between predators and prey, adaptive innovations often trigger a reciprocal response. For instance, the emergence of α-neurotoxins in snake venom has driven prey species targeted by these snakes to... more
In the ongoing evolutionary arms race between predators and prey, adaptive innovations often trigger a reciprocal response. For instance, the emergence of α-neurotoxins in snake venom has driven prey species targeted by these snakes to evolve sophisticated defense mechanisms. This study zeroes in on the particular motifs within the orthosteric sites of post-synaptic nicotinic acetylcholine receptors (nAChR) that confer resistance to α-neurotoxins, often through structural alterations of nAChR. This research examined Australian agamid lizards, a primary prey group for Australian elapid snakes, which are subject to predatory selection pressures. We previously showed that Pogona vitticeps (Central bearded dragon) was resistant to α-neurotoxic snake venoms through a steric hindrance form resistance evolving within the nAChR orthosteric, specifically through the 187–189NVT motif resulting in the presence of N-glycosylation, with the branching carbohydrate chains impeding the binding by the neurotoxins. This adaptive trait is thought to be a compensatory mechanism for the lizard's limited escape capabilities. Despite the significance of this novel adaptation, the prevalence and evolutionary roots of such venom resistance in Australian agamids have not been thoroughly investigated. To fill this knowledge gap, we undertook a comprehensive sequencing analysis of the nAChR ligand-binding domain across the full taxonomical diversity of Australian agamid species. Our findings reveal that the N-glycosylation resistance mechanism is a trait unique to the Pogona genus and absent in other Australian agamids. This aligns with Pogona's distinctive morphology, which likely increases vulnerability to neurotoxic elapid snakes, thereby increasing selective pressures for resistance. In contrast, biolayer interferometry experiments with death adder (Acanthophis species) venoms did not indicate any resistance-related binding patterns in other agamids, suggesting a lack of similar resistance adaptations, consistent with these lineages either being fast-moving, covered with large defensive spines, or being arboreal. This research not only uncovers a novel α-neurotoxin resistance mechanism in Australian agamids but also highlights the complex dynamics of the predator-prey chemical arms race. It provides a deeper understanding of how evolutionary pressures shape the interactions between venomous snakes and their prey.
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The evolutionary interplay between predator and prey has significantly shaped the de-velopment of snake venom, a critical adaptation for subduing prey. This arms race has spurred the diversification of the components of venom and the... more
The evolutionary interplay between predator and prey has significantly shaped the de-velopment of snake venom, a critical adaptation for subduing prey. This arms race has spurred the diversification of the components of venom and the corresponding emergence of resistance mecha-nisms in the prey and predators of venomous snakes. Our study investigates the molecular basis of venom resistance in pythons, focusing on electrostatic charge repulsion as a defense against α-neurotoxins binding to the alpha-1 subunit of the postsynaptic nicotinic acetylcholine receptor. Through phylogenetic and bioactivity analyses of orthosteric site sequences from various python species, we explore the prevalence and evolution of amino acid substitutions that confer resistance by electrostatic repulsion, which initially evolved in response to predatory pressure by Naja (co-bra) species (which occurs across Africa and Asia). The small African species Python regius re-tains the two resistance-conferring lysines (positions 189 and 191) of the ancestral Python genus, conferring resistance to sympatric Naja venoms. This differed from the giant African species Py-thon sebae, which has secondarily lost one of these lysines, potentially due to its rapid growth out of the prey size range of sympatric Naja species. In contrast, the two Asian species Python brong-ersmai (small) and Python bivittatus (giant) share an identical orthosteric site, which exhibits the highest degree of resistance, attributed to three lysine residues in the orthosteric sites. One of these lysines (at orthosteric position 195) evolved in the last common ancestor of these two species, which may reflect an adaptive response to increased predation pressures from the sympatric α-neurotoxic snake-eating genus Ophiophagus (King Cobras) in Asia. All these terrestrial Python species, however, were less neurotoxin-susceptible than pythons in other genera which have evolved under different predatory pressure as: the Asian species Malayopython reticulatus which is arboreal as neonates and juveniles before rapidly reaching sizes as terrestrial adults too large for sympatric Ophiophagus species to consider as prey; and the terrestrial Australian species Aspidites melanocephalus which occupies a niche, devoid of selection pressure from α-neurotoxic predatory snakes. Our findings underline the importance of positive selection in the evolution of venom re-sistance and suggest a complex evolutionary history involving both conserved traits and second-ary evolution. This study enhances our understanding of the molecular adaptations that enable pythons to survive in environments laden with venomous threats and offers insights into the on-going co-evolution between venomous snakes and their prey.
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The interplay between predator and prey has catalyzed the evolution of venom systems, with predators honing their venoms in response to the evolving resistance of prey. A previous study showed that the African varanid species Varanus... more
The interplay between predator and prey has catalyzed the evolution of venom systems, with predators honing their venoms in response to the evolving resistance of prey. A previous study showed that the African varanid species Varanus exanthematicus has heightened resistance to snake venoms compared to the Australian species V. giganteus, V. komodoensis, and V. mertensi, likely due to increased predation by sympatric venomous snakes on V. exanthematicus. To understand venom resistance among varanid lizards, we analyzed the receptor site targeted by venoms in 27 varanid lizards, including 25 Australian varanids. The results indicate an active evolutionary arms race between Australian varanid lizards and sympatric neurotoxic elapid snakes. Large species preying on venomous snakes exhibit inherited neurotoxin resistance, a trait potentially linked to their predatory habits. Consistent with the ‘use it or lose it’ aspect of venom resistance, this trait was secondarily reduced in two lineages that had convergently evolved gigantism (V. giganteus and the V. komodoensis/V. varius clade), suggestive of increased predatory success accompanying extreme size and also increased mechanical protection against envenomation due to larger scale osteoderms. Resistance was completely lost in the mangrove monitor V. indicus, consistent with venomous snakes not being common in their arboreal and aquatic niche. Conversely, dwarf varanids demonstrate a secondary loss at the base of the clade, with resistance subsequently re-evolving in the burrowing V. acanthurus/V. storri clade, suggesting an ongoing battle with neurotoxic predators. Intriguingly, within the V. acanthurus/V. storri clade, resistance was lost again in V. kingorum, which is morphologically and ecologically distinct from other members of this clade. Resistance was also re-evolved in V. glebopalma which is terrestrial in contrast to the arboreal/cliff dwelling niches occupied by the other members of its clade (V. glebopalma, V. mitchelli, V. scalaris, V. tristis). This ‘Russian doll’ pattern of venom resistance underscores the dynamic interaction between dwarf varanids and Australian neurotoxic elapid snakes. Our research, which included testing Acanthophis (death adder) venoms against varanid receptors as models for alpha-neurotoxic interactions, uncovered a fascinating instance of the Red Queen Hypothesis: some death adders have developed more potent toxins specifically targeting resistant varanids, a clear sign of the relentless predator–prey arms race. These results offer new insight into the complex dynamics of venom resistance and highlight the intricate ecological interactions that shape the natural world.
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Anacondas, genus Eunectes, are a group of aquatic snakes with a wide distribution in South America. The taxonomic status of several species has been uncertain and/or controversial. Using genetic data from four recognized anaconda species... more
Anacondas, genus Eunectes, are a group of aquatic snakes with a wide distribution in South America. The taxonomic status of several species has been uncertain and/or controversial. Using genetic data from four recognized anaconda species across nine countries, this study inves-tigates the phylogenetic relationships within the genus Eunectes. A key finding was the identifica-tion of two distinct clades within Eunectes murinus, revealing two species as cryptic yet genetical-ly deeply divergent. This has led to the recognition of the Northern Green Anaconda as a separate species (Eunectes akayima sp. nov), distinct from its southern counterpart (E. murinus), the Southern Green Anaconda. Additionally, our data challenge the current understanding of Yellow Anaconda species by proposing the unification of Eunectes deschauenseei and Eunectes beniensis into a single species with Eunectes notaeus. This reclassification is based on comprehensive genetic and phylo-geographic analyses, suggesting closer relationships than previously recognized and the realiza-tion that our understanding of their geographic ranges is insufficient to justify its use as a separa-tion criterion. We also present a phylogeographic hypothesis that traces the Miocene diversifica-tion of anacondas in western South America. Beyond its academic significance, this study has vital implications for the conservation of these iconic reptile species, highlighting our lack of knowledge about the diversity of the South American fauna and the need for revised strategies to conserve the newly identified and reclassified species.
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Bites from helodermatid lizards can cause pain, paresthesia, paralysis, and tachycardia, as well as other symptoms consistent with neurotoxicity. Furthermore, in vitro studies have shown that Heloderma horridum venom inhibits ion flux and... more
Bites from helodermatid lizards can cause pain, paresthesia, paralysis, and tachycardia, as well as other symptoms consistent with neurotoxicity. Furthermore, in vitro studies have shown that Heloderma horridum venom inhibits ion flux and blocks the electrical stimulation of skeletal muscles. Helodermatids have long been considered the only venomous lizards, but a large body of robust evidence has demonstrated venom to be a basal trait of Anguimorpha. This clade includes varanid lizards, whose bites have been reported to cause anticoagulation, pain, and occasionally paralysis and tachycardia. Despite the evolutionary novelty of these lizard venoms, their neuromuscular targets have yet to be identified, even for the iconic helodermatid lizards. Therefore, to fill this knowledge gap, the venoms of three Heloderma species (H. exasperatum, H. horridum and H. suspectum) and two Varanus species (V. salvadorii and V. varius) were investigated using Gallus gallus chick biventer cervicis ner...
Research Interests: Biology, Medicine, Reptiles, Venom Evolution, Venom, and 5 moreToxins, Venomous Reptiles, Sodium channel, Varanus, and Heloderma
The evolution of an aquatic lifestyle from land dwelling venomous elapids is a radical ecological modification, bringing about many evolutionary changes from morphology to diet. Diet is an important ecological facet which can play a key... more
The evolution of an aquatic lifestyle from land dwelling venomous elapids is a radical ecological modification, bringing about many evolutionary changes from morphology to diet. Diet is an important ecological facet which can play a key role in regulating functional traits such as venom composition and prey-specific targeting of venom. In addition to predating upon novel prey (e.g., fish, fish eggs and invertebrates), the venoms of aquatic elapids also face the challenge of increased prey-escape potential in the aquatic environment. Thus, despite the independent radiation into an aquatic niche on four separate occasions, the venoms of aquatic elapids are evolving under convergent selection pressures. Utilising a biolayer interferometry binding assay, this study set out to elucidate whether crude venoms from representative aquatic elapids were target-specific to the orthosteric site of postsynaptic nicotinic acetylcholine receptor mimotopes of fish compared to other terrestrial prey ...
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The binding of compounds to nicotinic acetylcholine receptors is of great interest in biomedical research. However, progress in this area is hampered by the lack of a high-throughput, cost-effective, and taxonomically flexible platform.... more
The binding of compounds to nicotinic acetylcholine receptors is of great interest in biomedical research. However, progress in this area is hampered by the lack of a high-throughput, cost-effective, and taxonomically flexible platform. Current methods are low-throughput, consume large quantities of sample, or are taxonomically limited in which targets can be tested. We describe a novel assay which utilizes a label-free bio-layer interferometry technology, in combination with adapted mimotope peptides, in order to measure ligand binding to the orthosteric site of nicotinic acetylcholine receptor alpha-subunits of diverse organisms. We validated the method by testing the evolutionary patterns of a generalist feeding species (Acanthophis antarcticus), a fish specialist species (Aipysurus laevis), and a snake specialist species (Ophiophagus hannah) for comparative binding to the orthosteric site of fish, amphibian, lizard, snake, bird, marsupial, and rodent alpha-1 nicotinic acetylchol...
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Research Interests: Evolutionary Biology, Toxicology, Molecular Evolution, Biology, Evolution, and 15 moreEvolutionary Ecology, Medicine, Snake venoms, Animal venoms and toxins, Coagulopathy, Molecular Phylogenetics and Evolution, Blood Coagulation, General medicine, Antivenom, Snake Venom, Coagulation, Snake Venom Antidote, Coagulation Factors, ENVIRONMENTAL SCIENCE AND MANAGEMENT, and Pharmacology and pharmaceutical sciences
The genus Bitis comprises 17 snake species that inhabit Africa and the Arabian Peninsula. They are responsible for a significant proportion of snakebites in the region. The venoms of the two independent lineages of giant Bitis (B.... more
The genus Bitis comprises 17 snake species that inhabit Africa and the Arabian Peninsula. They are responsible for a significant proportion of snakebites in the region. The venoms of the two independent lineages of giant Bitis (B. arietans and again in the common ancestor of the clade consisting of B. gabonica, B. nasicornis, B. parviocula and B. rhinoceros) induce an array of debilitating effects including anticoagulation, hemorrhagic shock and cytotoxicity, whilst the dwarf species B. atropos is known to have strong neurotoxic effects. However, the venom effects of the other species within the genus have not been explored in detail. A series of coagulation assays were implemented to assess the coagulotoxic venom effects of fourteen species within the genus. This study identified procoagulant venom as the ancestral condition, retained only by the basal dwarf species B. worthingtoni, suggesting anticoagulant venom is a derived trait within the Bitis genus and has been secondarily am...
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The functional activities of Anguimorpha lizard venoms have received less attention compared to serpent lineages. Bite victims of varanid lizards often report persistent bleeding exceeding that expected for the mechanical damage of the... more
The functional activities of Anguimorpha lizard venoms have received less attention compared to serpent lineages. Bite victims of varanid lizards often report persistent bleeding exceeding that expected for the mechanical damage of the bite. Research to date has identified the blockage of platelet aggregation as one bleeding-inducing activity, and destructive cleavage of fibrinogen as another. However, the ability of the venoms to prevent clot formation has not been directly investigated. Using a thromboelastograph (TEG5000), clot strength was measured after incubating human fibrinogen with Heloderma and Varanus lizard venoms. Clot strengths were found to be highly variable, with the most potent effects produced by incubation with Varanus venoms from the Odatria and Euprepriosaurus clades. The most fibrinogenolytically active venoms belonged to arboreal species and therefore prey escape potential is likely a strong evolutionary selection pressure. The results are also consistent wit...
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Research Interests: Herpetology, Toxicology, Biology, Toxinology, Medicine, and 14 moreSnake venoms, Pergamon, Animal venoms and toxins, Coagulopathy, Venom, Antivenom, Envenomation, Elapidae, In Vitro Toxicology, Disseminated Intravascular Coagulation, Snake Venom Antidote, Pseudonaja, Oxyuranus, and Pharmacology and pharmaceutical sciences
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Lancehead pit-vipers ( genus) are an extremely diverse and medically important group responsible for the greatest number of snakebite envenomations and deaths in South America. (common lancehead), responsible for majority of snakebites... more
Lancehead pit-vipers ( genus) are an extremely diverse and medically important group responsible for the greatest number of snakebite envenomations and deaths in South America. (common lancehead), responsible for majority of snakebites and related deaths within the Brazilian Amazon, is a highly adaptable and widely distributed species, whose venom variability has been related to several factors, including geographical distribution and habitat type. This study examined venoms from four populations (Belterra and Santarém, PA; Pres. Figueiredo, AM and São Bento, MA) and two additional species ( and ) from Southeastern region for their coagulotoxic effects upon different plasmas (human, amphibian, and avian). The results revealed inter⁻ and intraspecific variations in coagulotoxicity, including distinct activities between the three plasmas, with variations in the latter two linked to ecological niche occupied by the snakes. Also examined were the correlated biochemical mechanisms of ven...
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Atractaspis snake species are enigmatic in their natural history, and venom effects are correspondingly poorly described. Bite reports are scarce but bites have been described as causing severe hypertension, profound local tissue damage... more
Atractaspis snake species are enigmatic in their natural history, and venom effects are correspondingly poorly described. Bite reports are scarce but bites have been described as causing severe hypertension, profound local tissue damage leading to amputation, and deaths are on record. Clinical descriptions have largely concentrated upon tissue effects, and research efforts have focused upon the blood-pressure affecting sarafotoxins. However, coagulation disturbances suggestive of procoagulant functions have been reported in some clinical cases, yet this aspect has been uninvestigated. We used a suite of assays to investigate the coagulotoxic effects of venoms from six different Atractaspis specimens from central Africa. The procoagulant function of factor X activation was revealed, as was the pseudo-procoagulant function of direct cleavage of fibrinogen into weak clots. The relative neutralization efficacy of South African Antivenom Producer's antivenoms on Atractaspis venoms we...
Research Interests: Immunology, Central Africa, Medicine, Fibrinogen, Humans, and 15 moreAnimal venoms and toxins, Animals, Coagulopathy, Blood Coagulation, Antivenom, Snake Venom, Bothrops, Coagulation, Neutralization, Factor Xa, ENVIRONMENTAL SCIENCE AND MANAGEMENT, Blood coagulation disorders, Antivenins, Pharmacology and pharmaceutical sciences, and In Vitro Techniques
Pseudechis (black snakes) is an Australasian elapid snake genus that inhabits much of mainland Australia, with two representatives confined to Papua New Guinea. The present study is the first to analyse the venom of all 9 described... more
Pseudechis (black snakes) is an Australasian elapid snake genus that inhabits much of mainland Australia, with two representatives confined to Papua New Guinea. The present study is the first to analyse the venom of all 9 described Pseudechis species (plus one undescribed species) to investigate the evolution of venom composition and functional activity. Proteomic results demonstrated that the typical Pseudechis venom profile is dominated by phospholipase A2 toxins. Strong cytotoxicity was the dominant function for most species. P. porphyriacus, the most basal member of the genus, also exhibited the most divergent venom composition, being the only species with appreciable amounts of procoagulant toxins. The relatively high presence of factor Xa recovered in P. porphyriacus venom may be related to a predominantly amphibian diet. Results of this study provide important insights to guide future ecological and toxinological investigations.
Research Interests: Molecular Evolution, Australia, Biology, Proteomics, Cytotoxicity, and 15 moreMedicine, Snake venoms, Biological Sciences, Phylogeny, Environmental Sciences, Animal venoms and toxins, Animals, Antivenom, New Guinea, Envenomation, Elapidae, Hydrophiidae, Molecular Conformation, Coagulants, and Medical and Health Sciences
While some US populations of the Mohave rattlesnake (Crotalus scutulatus scutulatus) are infamous for being potently neurotoxic, the Mexican subspecies C. s. salvini (Huamantlan rattlesnake) has been largely unstudied beyond crude... more
While some US populations of the Mohave rattlesnake (Crotalus scutulatus scutulatus) are infamous for being potently neurotoxic, the Mexican subspecies C. s. salvini (Huamantlan rattlesnake) has been largely unstudied beyond crude lethality testing upon mice. In this study we show that at least some populations of this wide-ranging snake are as potently neurotoxic as its northern cousin. Testing of the Mexican antivenom Antivipmyn showed a complete lack of neutralisation for the neurotoxic effects of C. s. salvini venom, while the neurotoxic effects of the US subspecies C. s. scutulatus were time-delayed but ultimately not eliminated. These results document unrecognised potent neurological effects of a Mexican snake and highlight the medical importance of this subspecies, a finding augmented by the ineffectiveness of the Antivipmyn antivenom. These results also influence our understanding of the venom evolution of Crotalus scutulatus, suggesting that neurotoxicity is the ancestral f...
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A paradigm of venom research is adaptive evolution of toxins as part of a predator-prey chemical arms race. This examined differential co-factor dependence, variations relative to dietary preference, and the impact upon relative... more
A paradigm of venom research is adaptive evolution of toxins as part of a predator-prey chemical arms race. This examined differential co-factor dependence, variations relative to dietary preference, and the impact upon relative neutralisation by antivenom of the procoagulant toxins in the venoms of a clade of Australian snakes. All genera were characterised by venoms rich in factor Xa which act upon endogenous prothrombin. Examination of toxin sequences revealed an extraordinary level of conservation, which indicates that adaptive evolution is not a feature of this toxin type. Consistent with this, the venoms did not display differences on the plasma of different taxa. Examination of the prothrombin target revealed endogenous blood proteins are under extreme negative selection pressure for diversification, this in turn puts a strong negative selection pressure upon the toxins as sequence diversification could result in a drift away from the target. Thus this study reveals that adap...
Research Interests: Evolutionary Biology, Australia, Biology, Ecology, Evolution, and 15 moreMedicine, Biological Sciences, Phylogeny, Environmental Sciences, Humans, Animal venoms and toxins, Animals, Coagulopathy, Blood Coagulation, Antivenom, Elapidae, Disseminated Intravascular Coagulation, Coagulation, Antivenins, and Medical and Health Sciences
While snake venoms have been the subject of intense study, comparatively little work has been done on lizard venoms. In this study, we have examined the structural and functional diversification of anguimorph lizard venoms and associated... more
While snake venoms have been the subject of intense study, comparatively little work has been done on lizard venoms. In this study, we have examined the structural and functional diversification of anguimorph lizard venoms and associated toxins, and related these results to dentition and predatory ecology. Venom composition was shown to be highly variable across the 20 species of Heloderma, Lanthanotus, and Varanus included in our study. While kallikrein enzymes were ubiquitous, they were also a particularly multifunctional toxin type, with differential activities on enzyme substrates and also ability to degrade alpha or beta chains of fibrinogen that reflects structural variability. Examination of other toxin types also revealed similar variability in their presence and activity levels. The high level of venom chemistry variation in varanid lizards compared to that of helodermatid lizards suggests that venom may be subject to different selection pressures in these two families. The...
Research Interests: Evolutionary Biology, Phylogenetics, Biology, Evolution, Evolutionary Ecology, and 15 moreMedicine, Fibrinogen, Animal venoms and toxins, Lizards, Animals, Male, Monitor Lizard, Molecular Phylogenetics and Evolution, Lizard, Monitor lizards, Muscle contraction, Coagulation, Heloderma, Ileum, and In Vitro Techniques
Coral snake envenomations are well characterized to be lethally neurotoxic. Despite this, few multispecies, neurotoxicity and antivenom efficacy comparisons have been undertaken and only for the Micrurus genus; Micruroides has remained... more
Coral snake envenomations are well characterized to be lethally neurotoxic. Despite this, few multispecies, neurotoxicity and antivenom efficacy comparisons have been undertaken and only for the Micrurus genus; Micruroides has remained entirely uninvestigated. As the USA's supplier of antivenom has currently stopped production, alternative sources need to be explored. The Mexican manufacturer Bioclon uses species genetically related to USA species, thus we investigated the efficacy against Micrurus fulvius (eastern coral snake), the main species responsible for lethal envenomations in the USA as well as additional species from the Americas. The use of Coralmyn® coral snake antivenom was effective in neutralizing the neurotoxic effects exhibited by the venom of M. fulvius but was ineffective against the venoms of Micrurus tener, Micrurus spixii, Micrurus pyrrhocryptus, and Micruroides euryxanthus. Our results suggest that the Mexican antivenom may be clinically useful for the tre...
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Venoms can deleteriously affect any physiological system reachable by the bloodstream, including directly interfering with the coagulation cascade. Such coagulopathic toxins may be anticoagulants or procoagulants. Snake venoms are unique... more
Venoms can deleteriously affect any physiological system reachable by the bloodstream, including directly interfering with the coagulation cascade. Such coagulopathic toxins may be anticoagulants or procoagulants. Snake venoms are unique in their use of procoagulant toxins for predatory purposes. The boomslang (Dispholidus typus) and the twig snakes (Thelotornis species) are iconic African snakes belonging to the family Colubridae. Both species produce strikingly similar lethal procoagulant pathologies. Despite these similarities, antivenom is only produced for treating bites by D. typus, and the mechanisms of action of both venoms have been understudied. In this study, we investigated the venom of D. typus and T. mossambicanus utilising a range of proteomic and bioactivity approaches, including determining the procoagulant properties of both venoms in relation to the human coagulation pathways. In doing so, we developed a novel procoagulant assay, utilising a Stago STA-R Max analys...
Research Interests: Biology, Evolution, Medicine, Humans, Blood Coagulation Factors, and 15 moreAnimals, Blood Coagulation, Biological evolution, Colubridae, Colubrid Snake Venom, Antivenom, Disseminated Intravascular Coagulation, Enzyme activity, Biodiscovery, Coagulation Factors, Metalloproteases, Exocrine glands, Antivenins, Procoagulant, and Pharmacology and pharmaceutical sciences
Venom is a key evolutionary trait, as evidenced by its widespread convergent evolution across the animal kingdom. In an escalating prey-predator arms race, venoms evolve rapidly to guarantee predatory or defensive success. Variation in... more
Venom is a key evolutionary trait, as evidenced by its widespread convergent evolution across the animal kingdom. In an escalating prey-predator arms race, venoms evolve rapidly to guarantee predatory or defensive success. Variation in venom composition is ubiquitous among snakes. Here, we tested variation in venom activity on substrates relevant to blood coagulation among Pseudonaja (brown snake) species, Australian elapids responsible for the majority of medically important human envenomations in Australia. A functional approach was employed to elucidate interspecific variation in venom activity in all nine currently recognised species of Pseudonaja. Fluorometric enzymatic activity assays were performed to test variation in whole venom procoagulant activity among species. Analyses confirmed the previously documented ontogenetic shift from non-coagulopathic venom in juveniles to coagulopathic venom as adults, except for the case of P. modesta, which retains non-coagulopathic venom ...
Research Interests: Biochemistry, Evolutionary Biology, Australia, Biology, Diet, and 15 moreCell Biology, Evolutionary Ecology, Biological Sciences, Environmental Sciences, Animal venoms and toxins, Animals, Coagulopathy, Antivenom, Envenomation, Elapidae, Disseminated Intravascular Coagulation, Coagulation, Enzymatic Activity, Factor Xa, and Coagulants
Venom systems have evolved on multiple occasions across the animal kingdom, and they can act as key adaptations to protect animals from predators [1]. Consequently, venomous animals serve as models for a rich source of mimicry types, as... more
Venom systems have evolved on multiple occasions across the animal kingdom, and they can act as key adaptations to protect animals from predators [1]. Consequently, venomous animals serve as models for a rich source of mimicry types, as non-venomous species benefit from reductions in predation risk by mimicking the coloration, body shape, and/or movement of toxic counterparts [2-5]. The frequent evolution of such deceitful imitations provides notable examples of phenotypic convergence and are often invoked as classic exemplars of evolution by natural selection. Here, we investigate the evolution of fangs, venom, and mimetic relationships in reef fishes from the tribe Nemophini (fangblennies). Comparative morphological analyses reveal that enlarged canine teeth (fangs) originated at the base of the Nemophini radiation and have enabled a micropredatory feeding strategy in non-venomous Plagiotremus spp. Subsequently, the evolution of deep anterior grooves and their coupling to venom se...
Research Interests: Evolutionary Biology, Pigmentation, Biology, Evolution, Medicine, and 15 moreBiological Sciences, Animal venoms and toxins, Predation, Animals, Fishes, Mimicry, Biological evolution, Blennies, Phenotype, Jellyfish, Aposematism, Confusion, Convergent Evolution, Batesian Mimicry, and Medical and Health Sciences
Research Interests: Evolutionary Biology, Pigmentation, Biology, Evolution, Evolutionary Ecology, and 15 moreCytotoxicity, Medicine, Cell line, Humans, Animal venoms and toxins, Animals, Effects of Proteins Extracted from Snake Venom on Cancerous Cells, Population Ecology of Venomous Snakes, Biological evolution, Neurotoxins, Chickens, Aposematism, Elapidae, Neuromuscular Junction, and Cell Survival
Fish venoms are often poorly studied, in part due to the difficulty in obtaining, extracting, and storing them. In this study, we characterize the cardiovascular and neurotoxic effects of the venoms from the following six species of fish:... more
Fish venoms are often poorly studied, in part due to the difficulty in obtaining, extracting, and storing them. In this study, we characterize the cardiovascular and neurotoxic effects of the venoms from the following six species of fish: the cartilaginous stingrays Neotrygon kuhlii and Himantura toshi, and the bony fish Platycephalus fucus, Girella tricuspidata, Mugil cephalus, and Dentex tumifrons. All venoms (10-100 μg/kg, i.v.), except G. tricuspidata and P. fuscus, induced a biphasic response on mean arterial pressure (MAP) in the anesthetised rat. P. fucus venom exhibited a hypotensive response, while venom from G. tricuspidata displayed a single depressor response. All venoms induced cardiovascular collapse at 200 μg/kg, i.v. The in vitro neurotoxic effects of venom were examined using the chick biventer cervicis nerve-muscle (CBCNM) preparation. N. kuhlii, H. toshi, and P. fucus venoms caused concentration-dependent inhibition of indirect twitches in the CBCNM preparation. T...
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Research Interests: Toxicology, Molecular Evolution, Biology, Proteomics, Medicine, and 15 morePhylogeny, Sequence alignment, Lizards, Animals, Tandem Mass Spectrometry, Venom, High Performance Liquid Chromatography, Lizard, Toxins, Adaptive evolution, Toxin, Amino Acid Sequence, Heloderma, Two dimensional Gel Electrophoresis, and Molecular Sequence Data
Research Interests: Biochemistry, Analytical Chemistry, Molecular Biology, Molecular Evolution, Biology, and 15 moreMedicine, Multidisciplinary, Snake venoms, Transcriptome, Phylogeny, Sequence alignment, Maximum Likelihood, Lizards, Animals, Snakes, Venom, Positive Selection, Adaptive evolution, Amino Acid Sequence, and Molecular Sequence Data
Research Interests: Bioinformatics, Molecular Evolution, Biology, Life Sciences, Medicine, and 15 moreMultidisciplinary, Phylogeny, Humans, Biomedical Research, Animal venoms and toxins, Mutation, Animals, Nicotinic Acetylcholine Receptors, Peptides, Metalloproteinase, Amino Acid Sequence, Disintegrin, Metalloproteases, Molecular Sequence Data, and binding sites
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Research Interests: Bioinformatics, Evolutionary Biology, Genetics, Molecular Biology, Computational Biology, and 15 moreMolecular Evolution, Biology, Evolution, Medicine, Phylogeny, Animal venoms and toxins, Reptiles, Lizards, Animals, Natriuretic Peptides, Lizard, Amino Acid Sequence, Arboreal Locomotion, Biochemistry and cell biology, and Molecular Sequence Data
Background: The explosive radiation and diversification of the advanced snakes (superfamily Colubroidea) was associated with changes in all aspects of the shared venom system. Morphological changes included the partitioning of the mixed... more
Background: The explosive radiation and diversification of the advanced snakes (superfamily Colubroidea) was associated with changes in all aspects of the shared venom system. Morphological changes included the partitioning of the mixed ancestral glands into two discrete glands devoted for production of venom or mucous respectively, as well as changes in the location, size and structural elements of the venom-delivering teeth. Evidence also exists for homology among venom gland toxins expressed across the advanced snakes. However, despite the evolutionary novelty of snake venoms, in-depth toxin molecular evolutionary history reconstructions have been mostly limited to those types present in only two front-fanged snake families, Elapidae and Viperidae. To have a broader understanding of toxins shared among extant snakes, here we first sequenced the transcriptomes of eight taxonomically diverse rear-fanged species and four key viperid species and analysed major toxin types shared across the advanced snakes. Results: Transcriptomes were constructed for the following families and species: Colubridae-Helicops leopardinus, Heterodon nasicus, Rhabdophis subminiatus; Homalopsidae-Homalopsis buccata; Lamprophiidae-Malpolon monspessulanus, Psammophis schokari, Psammophis subtaeniatus, Rhamphiophis oxyrhynchus; and Viperidae-Bitis atropos, Pseudocerastes urarachnoides, Tropidolaeumus subannulatus, Vipera transcaucasiana. These sequences were combined with those from available databases of other species in order to facilitate a robust reconstruction of the molecular evolutionary history of the key toxin classes present in the venom of the last common ancestor of the advanced snakes, and thus present across the full diversity of colubroid snake venoms. In addition to differential rates of evolution in toxin classes between the snake lineages, these analyses revealed multiple instances of
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The evolution of venom resistance through coevolutionary chemical arms races has arisen multiple times throughout animalia. Prior documentation of resistance to snake venom α-neurotoxins consists of the N-glycosylation motif or the... more
The evolution of venom resistance through coevolutionary chemical arms races has arisen multiple times throughout animalia. Prior documentation of resistance to snake venom α-neurotoxins consists of the N-glycosylation motif or the hypothesized introduction of arginine at positions 187 at the α-1 nicotinic acetylcholine receptor orthosteric site. However, no further studies have investigated the possibility of other potential forms of resistance. Using a biolayer interferometry assay, we first confirm that the previously hypothesized resistance conferred by arginine at position 187 in the honey badger does reduce binding to α-neurotoxins, which has never been functionally tested. We further discovered a novel form of α-neurotoxin resistance conferred by charge reversal mutations, whereby a negatively charged amino acid is replaced by the positively charged amino acid lysine. As venom α-neurotoxins have evolved strong positive charges on their surface to facilitate binding to the negatively charged α-1 orthosteric site, these mutations result in a positive charge/positive charge interaction electrostati-cally repelling the α-neurotoxins. Such a novel mechanism for resistance has gone completely undiscovered, yet this form of resistance has convergently evolved at least 10 times within snakes. These coevolutionary innovations seem to have arisen through convergent phenotypes to ultimately evolve a similar biophysical mechanism of resistance across snakes.
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Envenomations are complex medical emergencies that can have a range of symptoms and sequelae. The only specific, scientifically-validated treatment for envenomation is antivenom administration, which is designed to alleviate venom... more
Envenomations are complex medical emergencies that can have a range of symptoms and sequelae. The only specific, scientifically-validated treatment for envenomation is antivenom administration, which is designed to alleviate venom effects. A paucity of efficacy testing exists for numerous antivenoms worldwide, and understanding venom effects and venom potency can help identify antivenom improvement options. Some spider venoms can produce debilitating injuries or even death, yet have been largely neglected in venom and antivenom studies because of the low venom yields. Coagulation disturbances have been particularly under studied due to difficulties in working with blood and the coagulation cascade. These circumstances have resulted in suboptimal spider bite treatment for medically significant spider genera such as Loxosceles and Sicarius. This study identifies and quantifies the anticoagulant effects produced by venoms of three Loxoscles species (L. reclusa, L. boneti, and L. laeta) and that of Sicarius terrosus. We showed that the venoms of all studied species are able to cleave the fibrinogen Aα-chain with varying degrees of potency, with L. reclusa and S. terrosus venom cleaving the Aα-chain most rapidly. Thromboelastography analysis revealed that only L. reclusa venom is able to reduce clot strength, thereby presumably causing anticoagulant effects in the patient. Using the same thromboelastography assays, antivenom efficacy tests revealed that the commonly used Loxoscles-specific SMase D recombinant based antivenom failed to neutralize the anticoagulant effects produced by Loxosceles venom. This study demonstrates the fibrinogenolytic activity of Loxosceles and Sicarius venom and the neutralization failure of Loxosceles antivenom, thus providing impetus for antivenom improvement. Key Contribution: Loxosceles and Sicarius venom was shown to be fibrinogenolytic, with antivenom unable to neutralize these effects.
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Slow lorises are enigmatic animal that represent the only venomous primate lineage. Their defensive secretions have received little attention. In this study we determined the full length sequence of the protein secreted by their unique... more
Slow lorises are enigmatic animal that represent the only venomous primate lineage. Their defensive secretions have received little attention. In this study we determined the full length sequence of the protein secreted by their unique brachial glands. The full length sequences displayed homology to the main allergenic protein present in cat dander. We thus compared the molecular features of the slow loris brachial gland protein and the cat dander allergen protein, showing remarkable similarities between them. Thus we postulate that allergenic proteins play a role in the slow loris defensive arsenal. These results shed light on these neglected, novel animals. Key Contribution: We show the remarkable similarities between the proteins of cat dander allergen and slow loris brachial gland secretions. These proteins play a role in the defensive arsenal of both mammals.
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Dispholidus typus and Thelotornis mossambicanus are closely related rear-fanged colubrid snakes that both possess strongly procoagulant venoms. However, despite similarities in overall venom biochemistry and resulting clinical... more
Dispholidus typus and Thelotornis mossambicanus are closely related rear-fanged colubrid snakes that both possess strongly procoagulant venoms. However, despite similarities in overall venom biochemistry and resulting clinical manifestations, the underlying venom composition differs significantly between the two species. As a result, the only available antivenom-which is a monovalent antivenom for D. typus-has minimal cross reactivity with T. mossambicanus and is not a clinically viable option. It was hypothesised that this lack of cross reactivity is due to the additional large metalloprotease protein within T. mossambicanus venom, which may also be responsible for faster coagulation times. In this study, we found that T. mossambicanus venom is a more powerful activator of prothrombin than that of D. typus and that the SVMP transcripts from T. mossambicanus form a clade with those from D. typus. The sequences from D. typus and T. mossambicanus were highly similar in length, with the calculated molecular weights of the T. mossambicanus transcripts being significantly less than the molecular weights of some isoforms on the 1D SDS-PAGE gels. Analyses utilising degylcosylating enzymes revealed that T. mossambicanus SVMPs are glycosylated during post-translational modification, but that this does not lead to the different molecular weight bands observed in 1D SDS-PAGE gels. However, differences in gly-cosylation patterns may still explain some of the difference between the enzymatic activities and neutralization by antivenom that have been observed in these venoms. The results of this study provide new information regarding the treatment options for patients envenomated by T. mossambicanus as well as the evolution of these dangerous snakes.
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The binding of compounds to nicotinic acetylcholine receptors is of great interest in biomedical research. However, progress in this area is hampered by the lack of a high-throughput, cost-effective, and taxonomically flexible platform.... more
The binding of compounds to nicotinic acetylcholine receptors is of great interest in biomedical research. However, progress in this area is hampered by the lack of a high-throughput, cost-effective, and taxonomically flexible platform. Current methods are low-throughput, consume large quantities of sample, or are taxonomically limited in which targets can be tested. We describe a novel assay which utilizes a label-free bio-layer interferometry technology, in combination with adapted mimotope peptides, in order to measure ligand binding to the orthosteric site of nicotinic acetylcholine receptor alpha-subunits of diverse organisms. We validated the method by testing the evolutionary patterns of a generalist feeding species (Acanthophis antarcticus), a fish specialist species (Aipysurus laevis), and a snake specialist species (Ophiophagus hannah) for comparative binding to the orthosteric site of fish, amphibian, lizard, snake, bird, marsupial, and rodent alpha-1 nicotinic acetylcholine receptors. Binding patterns corresponded with diet, with the Acanthophis antarcticus not showing bias towards any particular lineage, while Aipysurus laevis showed selectivity for fish, and Ophiophagus hannah a selectivity for snake. To validate the biodiscovery potential of this method, we screened Acanthophis antarcticus and Tropidolaemus wagleri venom for binding to human alpha-1, alpha-2, alpha-3, alpha-4, alpha-5, alpha-6, alpha-7, alpha-9, and alpha-10. While A. antarcticus was broadly potent, T. wagleri showed very strong but selective binding, specifically to the alpha-1 target which would be evolutionarily selected for, as well as the alpha-5 target which is of major interest for drug design and development. Thus, we have shown that our novel method is broadly applicable for studies including evolutionary patterns of venom diversification, predicting potential neurotoxic effects in human envenomed patients, and searches for novel ligands of interest for laboratory tools and in drug design and development.
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Australian elapid snakes are some of the most venomous snakes in the world and are unique among venomous snakes in having mutated forms of the blood clotting factor X in an activated form (FXa) as a key venom component. In human bite... more
Australian elapid snakes are some of the most venomous snakes in the world and are unique among venomous snakes in having mutated forms of the blood clotting factor X in an activated form (FXa) as a key venom component. In human bite victims, an overdose of this activated clotting enzyme results in the systemic consumption of fibrinogen due to the large amounts of endogenous thrombin generated by the conversion of pro-thrombin to thrombin by venom FXa. Within Australian elapids, such procoagulant venom is currently known from the tiger snake clade (Hoplocephalus, Notechis, Paroplocephalus, and Tropidechis species), brown/taipan (Oxyuranus and Pseudonaja species) clade, and the red-bellied black snake Pseudechis porphyriacus. We used a STAR Max coagulation analyser and TEG5000 thromboelastographers to test 47 Australian elapid venoms from 19 genera against human plasma in vitro. In addition to activity being confirmed in the two clades above, FXa-driven potent procoagulant activity was found in four additional genera (Cryptophis, Demansia, Hemiaspis, and Suta). Ontogenetic changes in procoagulant function was also identified as a feature of Suta punctata venom. Phylogenetic analysis of FX sequences confirmed that snake venom FXa toxins evolved only once, that the potency of these toxins against human plasma has increased in a stepwise fashion, and that multiple convergent amplifications of procoagulant activity within Australian elapid snakes have occurred. Cofactor dependence tests revealed all procoagulant venoms in our study, except those of the tiger snake clade, to be highly calcium-dependent, whereas phospholipid dependence was less of a feature but still displayed significant variation between venoms. Antivenom testing using CSL Tiger Snake Antivenom showed broad but differential cross-reactivity against procoagulant venoms, with P. porphyriacus and S. punctata extremely well neutralised but with Cryptophis, Demansia, and Hemiaspis less well-neutralised. The relative variation was not in accordance to genetic relatedness of the species used in antivenom production (Notechis scutatus), which underscores a fundamental principle that the rapid evolution characteristic of venoms results in organismal phylogeny being a poor pre-dictor of antivenom efficacy. Our results have direct and immediate implications for the design of clinical management plans in the event of snakebite by such lesser known Australian elapid snake species that have been revealed in this study to be as potent as the better studied, and proven lethal, species.
Research Interests: Evolutionary Biology, Molecular Evolution, Evolution, Evolutionary Ecology, Snake venoms, and 10 moreAnimal venoms and toxins, Venom Evolution, Coagulopathy, Molecular Phylogenetics and Evolution, Blood Coagulation, Antivenom, Venomous snakes, Coagulation, Snake Venom Antidote, and Coagulation Factors
Snake envenomation globally is attributed to an ever-increasing human population encroaching into snake territories. Responsible for many bites in Asia is the widespread genus Trimeresurus. While bites lead to haemorrhage , only a few... more
Snake envenomation globally is attributed to an ever-increasing human population encroaching into snake territories. Responsible for many bites in Asia is the widespread genus Trimeresurus. While bites lead to haemorrhage , only a few species have had their venoms examined in detail. We found that Trimeresurus venom causes haemorrhaging by cleaving fibrinogen in a pseudo-procoagulation manner to produce weak, unstable, short-lived fibrin clots ultimately resulting in an overall anticoagulant effect due to fibrinogen depletion. The monovalent antivenom 'Thai Red Cross Green Pit Viper antivenin', varied in efficacy ranging from excellent neutralisation of T. albolabris venom through to T. gumprechti and T. mcgregori being poorly neutralised and T. hageni being unrecognised by the antivenom. While the results showing excellent neutralisation of some non-T. albolabris venoms (such as T. flavomaculaturs, T. fucatus, and T. macrops) needs to be confirmed with in vivo tests, conversely the antivenom failure T. hageni, and the very poor results against T. gumprechti and T. mcgregori, despite being conducted in the ideal scenario of preincubation of antivenom:venom, indicates that the likelihood of clinically relevant cross-reactivity for these species is low (T. gumprechti and T. mcgregori) to non-existent (T. hageni). These same latter three species were also not inhibited by the serine protease inhibitor AEBSF, suggesting that the toxins leading to a coagulotoxic effect in these species are non-serine proteases while in contrast T. albolabris coagulotoxicity was completely impeded by AEBSF, and thus driven by kallikrein-type serine proteases. There was a conspicuous lack of phylogenetic pattern in venom variation, with the most potent venoms (T. albolabris and T. hageni) being distant to each other on the organismal tree, and with the three most divergent and poorly neutralised venoms (T. gumprechti, T. hageni, and T. mcgregori) were also not each others closest relatives. This reinforces the paradigm that the fundamental dynamic evolution of venom results in organismal phylogeny being a poor predictor of venom potency or antivenom efficacy. This study provides a robust investigation on the differential venom effects from a wide range of Trimeresurus species on coagulation, highlighting differential fibrinogenolytic effects, while also investigating the relative antivenom neutralisation capabilities of the widely available Thai Red Cross Green Pit Viper antivenom. These results therefore have immediate, real-world implications for patients envenomed by Trimeresurus species.
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Snake bite is currently one of the most neglected tropical diseases affecting much of the developing world. Asian pit vipers are responsible for a considerable amount of envenomations annually and bites can cause a multitude of clinical... more
Snake bite is currently one of the most neglected tropical diseases affecting much of the developing world. Asian pit vipers are responsible for a considerable amount of envenomations annually and bites can cause a multitude of clinical complications resulting from coagulopathic and neuropathic effects. While intense research has been undertaken for some species of Asian pit viper, functional coagulopathic effects have been neglected for others. We investigated their effects upon the human clotting cascade using venoms of four species of Gloydius and Ovophis okinavensis, a species closely to Gloydius. All species of included within this investigation displayed varying fibrinogenolytic effects, resulting in a net anticoagulant outcome. Gloydius saxatilis and Gloydius ussuriensis displayed the most variable effects from differing localities, sampled from Russia and Korea. As this Gloydius investigation includes some geographical variation, notable results indicate key variations of these species that point to possible limitations in antivenom cross-reactivities, which may have implications for the clinical care of victims envenomed by these snakes.
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The functional activities of Anguimorpha lizard venoms have received less attention compared to serpent lineages. Bite victims of varanid lizards often report persistent bleeding exceeding that expected for the mechanical damage of the... more
The functional activities of Anguimorpha lizard venoms have received less attention compared to serpent lineages. Bite victims of varanid lizards often report persistent bleeding exceeding that expected for the mechanical damage of the bite. Research to date has identified the blockage of platelet aggregation as one bleeding-inducing activity, and destructive cleavage of fibrinogen as another. However, the ability of the venoms to prevent clot formation has not been directly investigated. Using a thromboelastograph (TEG5000), clot strength was measured after incubating human fibrinogen with Heloderma and Varanus lizard venoms. Clot strengths were found to be highly variable, with the most potent effects produced by incubation with Varanus venoms from the Odatria and Euprepriosaurus clades. The most fibrinogenolytically active venoms belonged to arboreal species and therefore prey escape potential is likely a strong evolutionary selection pressure. The results are also consistent with reports of profusive bleeding from bites from other notably fibrinogenolytic species, such as V. giganteus. Our results provide evidence in favour of the predatory role of venom in varanid lizards, thus shedding light on the evolution of venom in reptiles and revealing potential new sources of bioactive molecules useful as lead compounds in drug design and development.
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Venom can affect any part of the body reachable via the bloodstream. Toxins which specifically act upon the coagulation cascade do so either by anticoagulant or procoagulant mechanisms. Here we investigated the coagulotoxic effects of six... more
Venom can affect any part of the body reachable via the bloodstream. Toxins which specifically act upon the coagulation cascade do so either by anticoagulant or procoagulant mechanisms. Here we investigated the coagulotoxic effects of six species within the medically important pit viper genus Protobothrops (Habu) from the Chinese mainland and Japanese islands, a genus known to produce hemorrhagic shock in envenomed patients. Differential coagulotoxicity was revealed: P. jerdonii and P. mangshanensis produced an overall net anticoagulant effect through the pseudo-procoagulant clotting of fibrinogen; P. flavoviridis and P. tokarensis exhibit a strong anticoagulant activity through the destructive cleavage of fibrinogen; and while P. elegans and P. mucrosquamatus both cleaved the A-alpha and B-beta chains of fibrinogen they did not exhibit strong anticoagulant activity. These variations in coagulant properties were congruent with phylogeny, with the closest relatives exhibiting similar venom effects in their action upon fibrinogen. Ancestral state reconstruction indicated that anticoagulation mediated by pseudo-procoagulant cleavage of fibrinogen is the basal state, while anticoagulation produced by destructive cleavage of fibrinogen is the derived state within this genus. This is the first in depth study of its kind highlighting extreme enzymatic variability, functional diversification and clotting diversification within one genus surrounding one target site, governed by variability in co-factor dependency. The documentation that the same net overall function, anticoagulation, is mediated by differential underlying mechanics suggests limited antivenom cross-reactivity, although this must be tested in future work. These results add to the body of knowledge necessary to inform clinical management of the envenomed patient.
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Abstract: Cnidarians are the oldest extant lineage of venomous animals. Despite their simple anatomy, they are capable of subduing or repelling prey and predator species that are far more complex and recently evolved. Utilizing... more
Abstract: Cnidarians are the oldest extant lineage of venomous animals. Despite their
simple anatomy, they are capable of subduing or repelling prey and predator species that are
far more complex and recently evolved. Utilizing specialized penetrating nematocysts,
cnidarians inject the nematocyst content or “venom” that initiates toxic and immunological
reactions in the envenomated organism. These venoms contain enzymes, potent pore
forming toxins, and neurotoxins. Enzymes include lipolytic and proteolytic proteins that
catabolize prey tissues. Cnidarian pore forming toxins self-assemble to form robust
membrane pores that can cause cell death via osmotic lysis. Neurotoxins exhibit rapid ion
channel specific activities. In addition, certain cnidarian venoms contain or induce the
release of host vasodilatory biogenic amines such as serotonin, histamine, bunodosine and
caissarone accelerating the pathogenic effects of other venom enzymes and porins. The
cnidarian attacking/defending mechanism is fast and efficient, and massive envenomation
of humans may result in death, in some cases within a few minutes to an hour after sting.
The complexity of venom components represents a unique therapeutic challenge and probably
reflects the ancient evolutionary history of the cnidarian venom system. Thus, they are
invaluable as a therapeutic target for sting treatment or as lead compounds for drug design.
simple anatomy, they are capable of subduing or repelling prey and predator species that are
far more complex and recently evolved. Utilizing specialized penetrating nematocysts,
cnidarians inject the nematocyst content or “venom” that initiates toxic and immunological
reactions in the envenomated organism. These venoms contain enzymes, potent pore
forming toxins, and neurotoxins. Enzymes include lipolytic and proteolytic proteins that
catabolize prey tissues. Cnidarian pore forming toxins self-assemble to form robust
membrane pores that can cause cell death via osmotic lysis. Neurotoxins exhibit rapid ion
channel specific activities. In addition, certain cnidarian venoms contain or induce the
release of host vasodilatory biogenic amines such as serotonin, histamine, bunodosine and
caissarone accelerating the pathogenic effects of other venom enzymes and porins. The
cnidarian attacking/defending mechanism is fast and efficient, and massive envenomation
of humans may result in death, in some cases within a few minutes to an hour after sting.
The complexity of venom components represents a unique therapeutic challenge and probably
reflects the ancient evolutionary history of the cnidarian venom system. Thus, they are
invaluable as a therapeutic target for sting treatment or as lead compounds for drug design.
Research Interests:
Abstract: Cnidarian venom research has lagged behind other toxinological fields due to technical difficulties in recovery of the complex venom from the microscopic nematocysts. Here we report a newly developed rapid, repeatable and cost... more
Abstract: Cnidarian venom research has lagged behind other toxinological fields due to
technical difficulties in recovery of the complex venom from the microscopic nematocysts.
Here we report a newly developed rapid, repeatable and cost effective technique of venom
preparation, using ethanol to induce nematocyst discharge and to recover venom contents
in one step. Our model species was the Australian box jellyfish (Chironex fleckeri), which
has a notable impact on public health. By utilizing scanning electron microscopy and
light microscopy, we examined nematocyst external morphology before and after ethanol
treatment and verified nematocyst discharge. Further, to investigate nematocyst content or
“venom” recovery, we utilized both top-down and bottom-up transcriptomics–proteomics
approaches and compared the proteome profile of this new ethanol recovery based method
to a previously reported high activity and recovery protocol, based upon density purified
intact cnidae and pressure induced disruption. In addition to recovering previously
characterized box jellyfish toxins, including CfTX-A/B and CfTX-1, we recovered putative
metalloproteases and novel expression of a small serine protease inhibitor. This study not
only reveals a much more complex toxin profile of Australian box jellyfish venom but also
suggests that ethanol extraction method could augment future cnidarian venom proteomics
research efforts.
technical difficulties in recovery of the complex venom from the microscopic nematocysts.
Here we report a newly developed rapid, repeatable and cost effective technique of venom
preparation, using ethanol to induce nematocyst discharge and to recover venom contents
in one step. Our model species was the Australian box jellyfish (Chironex fleckeri), which
has a notable impact on public health. By utilizing scanning electron microscopy and
light microscopy, we examined nematocyst external morphology before and after ethanol
treatment and verified nematocyst discharge. Further, to investigate nematocyst content or
“venom” recovery, we utilized both top-down and bottom-up transcriptomics–proteomics
approaches and compared the proteome profile of this new ethanol recovery based method
to a previously reported high activity and recovery protocol, based upon density purified
intact cnidae and pressure induced disruption. In addition to recovering previously
characterized box jellyfish toxins, including CfTX-A/B and CfTX-1, we recovered putative
metalloproteases and novel expression of a small serine protease inhibitor. This study not
only reveals a much more complex toxin profile of Australian box jellyfish venom but also
suggests that ethanol extraction method could augment future cnidarian venom proteomics
research efforts.