Lancehead pit-vipers (Bothrops genus) are an extremely diverse and medically important group responsible for the greatest number of snakebite envenomations and deaths in South America. Bothrops atrox (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 B. atrox populations (Belterra and Santarém, PA; Pres. Figueiredo, AM and São Bento, MA), and two additional Bothrops species (B. jararaca and B. neuwiedi) 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 venom action. Significant variation in the relative reliance upon the cofactors calcium and phospholipid were revealed, and the relative dependency did not significantly correlate with potency. Relative levels of Factor X or prothrombin activating toxins correlated with prey type and prey escape potential. The antivenom was shown to perform better in neutralising prothrombin activation activity than neutralising Factor X activation activity. Thus, the data reveal new information regarding the evolutionary selection pressures shaping snake venom evolution, while also having significant implications for the treatment of the envenomed patient. These results are, therefore, an intersection between evolutionary biology and clinical medicine.

The genus Bitis comprises 18 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 amplified on at least four occasions. A wide range of anticoagulant mechanisms were identified, such as pseudo-procoagulant and destructive activities upon fibrinogen in both giant and dwarf Bitis and the action of inhibiting the prothrombinase complex, which is present in a clade of dwarf Bitis. Antivenom studies revealed that while the procoagulant effects of B. worthingtoni were poorly neutralized, and thus a cause for concern, the differential mechanisms of anticoagulation in other species were all well neutralized. Thus, this study concludes there is a wide range of coagulotoxic mechanisms which have evolved within the Bitis genus and that clinical management strategies are limited for the procoagulant effects of B. worthingtoni, but that anticoagulant effects of other species are readily treated by the South African polyvalent antivenom. These results therefore have direct, real-work implications for the treatment of envenomed patients.

Snakes of the genera Pseudocerastes and Eristicophis (Viperidae: Viperinae) are known as the desert vipers due to their association with the arid environments of the Middle East. These species have received limited research attention and little is known about their venom or ecology. In this study, a comprehensive analysis of desert viper venoms was conducted by visualising the venom proteomes via gel electrophoresis and assessing the crude venoms for their cytotoxic, haemotoxic, and neurotoxic properties. Plasmas sourced from human, toad, and chicken were used as models to assess possible prey-linked venom activity. The venoms demonstrated substantial divergence in composition and bioactivity across all experiments. Pseudocerastes urarachnoides venom activated human coagulation factors X and prothrombin and demonstrated potent procoagulant activity in human, toad, and chicken plasmas, in stark contrast to the potent neurotoxic venom of P. fieldi. The venom of E. macmahonii also induced coagulation, though this did not appear to be via the activation of factor X or prothrombin. The coagulant properties of P. fieldi and P. persicus venoms varied among plasmas, demonstrating strong anticoagulant activity in the amphibian and human plasmas but no significant effect in that of bird. This is conjectured to reflect prey-specific toxin activity, though further ecological studies are required to confirm any dietary associations. This study reinforces the notion that phylogenetic relatedness of snakes cannot readily predict venom protein composition or function. The significant venom variation between these species raises serious concerns regarding antivenom paraspecificity. Future assessment of antivenom is crucial.

The snake genus Daboia (Viperidae: Viperinae; Oppel, 1811) contains five species: D. deserti, D. mauritanica, and D. palaestinae, found in Afro-Arabia, and the Russell’s vipers D. russelii and D. siamensis, found in Asia. Russell’s vipers are responsible for a major proportion of the medically important snakebites that occur in the regions they inhabit, and their venoms are notorious for their coagulopathic effects. While widely documented, the extent of venom variation within the Russell’s vipers is poorly characterised, as is the venom activity of other species within the genus. In this study we investigated variation in the haemotoxic activity of Daboia using twelve venoms from all five species, including multiple variants of D. russelii, D. siamensis, and D. palaestinae. We tested the venoms on human plasma using thromboelastography, dose-response coagulometry analyses, and calibrated automated thrombography, and on human fibrinogen by thromboelastography and fibrinogen gels. We assessed activation of blood factors X and prothrombin by the venoms using fluorometry. Variation in venom activity was evident in all experiments. The Asian species D. russelii and D. siamensis and the African species D. mauritanica possessed procoagulant venom, while D. deserti and D. palaestinae were net-anticoagulant. Of the Russell’s vipers, the venom of D. siamensis from Myanmar was most toxic and D. russelli of Sri Lanka the least. Activation of both factor X and prothrombin was evident by all venoms, though at differential levels. Fibrinogenolytic activity varied extensively throughout the genus and followed no phylogenetic trends. This venom variability underpins one of the many challenges facing treatment of Daboia snakebite envenoming. Comprehensive analyses of available antivenoms in neutralising these variable venom activities are therefore of utmost importance.

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.