The European continent is inhabited by medically important venomous Viperinae snakes. Vipera ammodytes, Vipera berus, and Vipera aspis cause the greatest public health problems in Europe, but there are other equally significant snakes in specific regions of the continent. Immunotherapy is indicated for patients with systemic envenoming, of which there are approximately 4000 annual cases in Europe, and was suggested as an indication for young children and pregnant women, even if they do not have systemic symptoms. In the present study, the safety and venom-neutralizing efficacy of Inoserp Europe—a new F(ab’)₂ polyvalent antivenom, designed to treat envenoming by snakes in the Eurasian region—were evaluated. In accordance with World Health Organization recommendations, several quality control parameters were applied to evaluate the safety of this antivenom. The venom-neutralizing efficacy of the antivenom was evaluated in mice and the results showed it had appropriate neutralizing potency against the venoms of several species of Vipera, Montivipera, and Macrovipera. Paraspecificity of the antivenom was demonstrated as well, since it neutralized venoms of species not included in the immunization schemes and contains satisfactory levels of total proteins and F(ab’)₂ fragment concentration. Therefore, this new polyvalent antivenom could be effective in the treatment of snake envenoming in Europe, including Western Russia and Turkey.

The African viperid snake genera Atheris, Cerastes, and Proatheris are closely related, similar in size, but occupy extremely divergent ecological niches (arboreal in tropical rainforests, fossorial in deserts, and swamp-dwelling, respectively). Their venoms have not previously been subjected to comparative analyses for their action upon the coagulation of blood, most notably with significant data deficiencies from Atheris and Proatheris. In contrast, the closely related genus Echis is well-documented as capable of producing potent procoagulant effects. In light of this, we set out to compare the coagulotoxic actions of Atheris ceratophora, A. chlorechis, A. desaixi, A. nitschei, A. squamigera, C. cerastes, C. cerastes gasperettii, C. vipera, and Proatheris superciliaris and explore potential pharmacological interventions to reestablish normal blood coagulation. All venoms displayed extremely potent procoagulant effects, over twice as fast as the most potent Echis reported to date. Although Cerastes is used in the immunising mixture of two different regionally available antivenoms (Inoserp-MENA with C. cerastes, C. cerastes gasperettii, C. vipera and Saudi Arabian polyvalent with C. cerastes), none of the other species in this study are included in the immunising mixture of any antivenom. Notably, all the Cerastes species were only neutralised by the Inoserp-MENA antivenom. C. cerastes venom was not neutralised well by the Saudi Arabian antivenom, with the low levels of recognition for any of the Cerastes venoms suggesting a strong regional variation in the venom of this species, as the C. cerastes venom tested was of African (Tunisian) origin versus Saudi locality used in that antivenom’s production. The other antivenoms (Micropharm EchiTAbG, ICP EchiTAb-Plus-ICP, Inosan Inoserp Pan-Africa, Premium Serums PANAF Sub-Sahara Africa, South African Vaccine Producers Echis, South African Vaccine Producers Polyvalent) all displayed trivial-to-no ability to neutralise the procoagulant toxicity of any of the Atheris, Cerastes, or Proatheris venoms. Comparative testing of the enzyme inhibitors DMPS, marimastat, and prinomastat, revealed a very potent neutralising capacity of marimastat, with prinomastat showing lower but still significant potency at the same molar concentration, while a 5× molar concentration of DMPS had no apparent effect on procoagulant venom effects normalized by the other inhibitors. These results and methods contribute to the body of knowledge of potential clinical effects and data necessary for evidence-based advancement of clinical management strategies.

The Eastern Long-Nosed Viper (Vipera ammodytes meridionalis) is considered one of the most venomous snakes in Europe. However, it is unknown whether ontogenetic variation in venom effects occurs in this subspecies and how this may impact antivenom efficacy. In this study, we compared the procoagulant activities of V. a. meridionalis venom on human plasma between neonate and adult venom phenotypes. We also examined the efficacy of three antivenoms—Viperfav, ViperaTAb, and Inoserp Europe—across our neonate and adult venom samples. While both neonate and adult V. a. meridionalis venoms produced procoagulant effects, the effects produced by neonate venom were more potent. Consistent with this, neonate venom was a stronger activator of blood-clotting zymogens, converting them into their active forms, with a rank order of Factor X >> Factor VII > Factor XII. Conversely, the less potent adult venom had a rank order of FXII marginally more activated than Factor VII, and both much more so than Factor X. This adds to the growing body of evidence that activation of factors besides FII (prothrombin) and FX are significant variables in reptile venom-induced coagulopathy. Although all three examined antivenoms displayed effective neutralization of both neonate and adult V. a. meridionalis venoms, they generally showed higher efficacy on adult venom than on neonate venom. The ranking of antivenom efficacy against neonate venom, from the most effective to the least effective, were Viperfav, Inoserp Europe, ViperaTAb; for adult venom, the ranking was Inoserp Europe, Viperfav, ViperaTAb. Our data reveal ontogenetic variation in V. a meridionalis, but this difference may not be of clinical concern as antivenom was effective at neutralizing both adult and neonate venom phenotypes. Regardless, our results highlight a previously undocumented ontogenetic shift, likely driven by the documented difference in prey preference observed for this species across age classes

This study examined the pathophysiological effects of venoms from neonate and adult specimens of the viperid snake Macrovipera lebetina obtusa, focusing on their ability to activate various blood clotting factors in human plasma. All venoms exhibited strong procoagulant properties. In concentration–response tests, the clotting potency of the neonate venoms fell within the range of their parents’ maximum clotting velocities and areas under the curve. Intriguingly, females were more potent than males within each age group, but this requires a larger sample size to confirm. Antivenom neutralization efficacy was equipotent across age groups. The venoms potently activated Factor X (FX) robustly, consistent with previous knowledge of this genus. For the first time, the ability to activate Factors VII (FVII) and XII (FXII) was identified in this genus, with FXII exhibiting particularly strong activation. The study found no significant ontogenetic variation in procoagulant venom potency on human plasma, convergent with the Daboia genus, the other large-bodied lineage within the Palearctic viperid clade. However, the activation of FXII and FVII reveals previously undocumented pathways in the procoagulant activity of these venoms, contributing to the broader understanding of venom evolution and its clinical impacts. These findings have implications for venom biodiscovery and the development of antivenoms, highlighting the complexity of clotting factor activation beyond traditional investigations that have myopically focused upon FX and prothrombin pathways, thereby underscoring the importance of exploring additional clotting factors.

Echis species (saw-scaled vipers) are WHO Category 1 medically significant venomous snakes with potent procoagulant venoms, which cause lethal venom-induced consumptive coagulopathy in human victims. Despite clinical presentations of bites varying significantly between individuals within the same species, the contribution of age-related changes in the venom biochemistry has not been investigated. This study investigated the ontogenetic changes in Echis pyramidum pyramidum venom and its impact on therapeutic efficacy. The efficacy of various antivenoms (Echitab, Echitab+ ICP, Inosan MENA, Inosan Pan African, and SAVP-Echis) was tested against both venom phenotypes. While both neonate and adult venoms were procoagulant, there were differences in the underlying biochemistry. Neonate venom was found to potently pathophysiologically activate Factor VII and Factor X, and to a lesser degree Factor XII. In contrast, adult venom was a slower clotter, less potent in activating FVII, equipotent with neonate venom on FXII, and inactive on FX. This is the first documentation of FVII and FXII activation for any Echis venom. The significant ontogenetic toxicological variations in Echis species were shown to impact antivenom efficacy. Among the tested antivenoms, SAVP-Echis was the most effective against both venom phenotypes, with adult venom being better neutralized. These findings suggest the need for a reconsideration of venom mixture selection in antivenom production through the inclusion of neonate venom. Additionally, the results indicate differential ontogenetic predatory ecology, providing a foundation for future natural history investigations.

In this study, we report the innovative application of whole-cell patch-clamp electrophysiology in assessing broad-spectrum neutralisation by three different antivenoms, of venoms from the medically significant scorpion genus Centruroides. Envenomations by as many as 21 species from the Centruroides genus result in up to 300,000 envenomations per year in Mexico, which poses significant and potentially life-threatening pathophysiology. We first evaluated the in vitro manifestation of envenomation against two human voltage-gated sodium (hNa_V) channel subtypes: hNa_V1.4 and hNa_V1.5, which are primarily expressed in skeletal muscles and cardiomyocytes, respectively. The neutralisation of venom activity was then characterised for three different antivenoms using a direct competition model against the more potent target, hNa_V1.4. While broad-spectrum neutralisation was identified, variation in neutralisation arose for Centruroides elegans, C. limpidus, C. noxius and C. suffusus venoms, despite the presence of a number of these venoms within the immunising mixture. This raises questions regarding the truly "broad" neutralisation capacity of the antivenoms. This study not only extends previous validation of the in vitro investigation of antivenom efficacy utilising the whole-cell patch-clamp technique but also underscores the potential of this animal-free model in exploring cross-reactivity, experimental scalability, and most importantly, informing clinical management practices regarding the administration of antivenom in Mexico.