Venom-Induced Blood Disturbances by Palearctic Viperid Snakes, and Their Relative Neutralization by Antivenoms and Enzyme-Inhibitors

Palearctic vipers are medically significant snakes in the genera Daboia, Macrovipera, Montivipera, and Vipera which occur throughout Europe, Central Asia, Near and Middle East. While the ancestral condition is that of a small-bodied, lowland species, extensive diversification has occurred in body size, and niche specialization. Using 27 venom samples and a panel of in vitro coagulation assays, we evaluated the relative coagulotoxic potency of Palearctic viper venoms and compared their neutralization by three antivenoms (Insoserp Europe, VIPERFAV and ViperaTAb) and two metalloprotease inhibitors (prinomastat and DMPS). We show that variation in morphology parallels variation in the Factor X activating procoagulant toxicity, with the three convergent evolutions of larger body sizes (Daboia genus, Macrovipera genus, and Vipera ammodytes uniquely within the Vipera genus) were each accompanied by a significant increase in procoagulant potency. In contrast, the two convergent evolutions of high altitude specialization (the Montivipera genus and Vipera latastei uniquely within the Vipera genus) were each accompanied by a shift away from procoagulant action, with the Montivipera species being particularly potently anticoagulant. Inoserp Europe and VIPERFAV antivenoms were both effective against a broad range of Vipera species, with Inoserp able to neutralize additional species relative to VIPERFAV, reflective of its more complex antivenom immunization mixture. In contrast, ViperaTAb was extremely potent in neutralizing V. berus but, reflective of this being a monovalent antivenom, it was not effective against other Vipera species. The enzyme inhibitor prinomastat efficiently neutralized the metalloprotease-driven Factor X activation of the procoagulant venoms. In contrast, DMPS (2,3-dimercapto-1-propanesulfonic acid), which as been suggested as another potential treatment option in the absence of antivenom, DMPS failed against all venoms tested. Overall, our results highlight the evolutionary variations within Palearctic vipers and help to inform clinical management of viper envenomation.

Intravenous Vipera berus Venom-Specific Fab Fragments and Intramuscular Vipera ammodytes Venom-Specific F(ab’)2 Fragments in Vipera ammodytes-Envenomed Patients

Vipera ammodytes (V. ammodytes) is the most venomous European viper. The aim of this study was to compare the clinical efficacy and pharmacokinetic values of intravenous Vipera berus venom-specific (paraspecific) Fab fragments (ViperaTAb) and intramuscular V. ammodytes venom-specific F(ab’)2 fragments (European viper venom antiserum, also called “Zagreb” antivenom) in V.ammodytes-envenomed patients. This was a prospective study of V.ammodytes-envenomed patients that were treated intravenously with ViperaTAb or intramuscularly with European viper venom antiserum that was feasible only due to the unique situation of an antivenom shortage. The highest venom concentration, survival, length of hospital stay and adverse reactions did not differ between the groups. Patients treated with intravenous Fab fragments were sicker, with significantly more rhabdomyolysis and neurotoxicity. The kinetics of Fab fragments after one or more intravenous applications matched better with the venom concentration in the early phase of envenomation compared to F(ab’)2 fragments that were given intramuscularly only on admission. F(ab’)2 fragments given intramuscularly had 25-fold longer apparent total body clearance and 14-fold longer elimination half-time compared to Fab fragments given intravenously (2 weeks vs. 24 h, respectively). In V.ammodytes-envenomed patients, the intramuscular use of specific F(ab’)2 fragments resulted in a slow rise of antivenom serum concentration that demanded their early administration but without the need for additional doses for complete resolution of all clinical signs of envenomation. Intravenous use of paraspecific Fab fragments resulted in the immediate rise of antivenom serum concentration that enabled their use according to the clinical progress, but multiple doses might be needed for efficient therapy of thrombocytopenia due to venom recurrence, while the progression of rhabdomyolysis and neurotoxic effects of the venom could not be prevented.

Comparison of Preclinical Properties of Several Available Antivenoms in the Search for Effective Treatment of Vipera ammodytes and Vipera berus Envenoming

Snakebites are a relatively rare medical emergency in Europe. In more than half of the annual cases caused by Vipera ammodytes, Vipera berus, and Vipera aspis, immunotherapy with animal-derived antivenom is indicated. Among eight products recently identified as available against European medically relevant species, only Zagreb antivenom, Viperfav, and ViperaTAb have been used almost exclusively for decades. Zagreb antivenom comprises V. ammodytes-specific F(ab′)2 fragments. Viperfav is a polyspecific preparation based on F(ab′)2 fragments against V. aspis, V. berus, and V. ammodytes venoms. ViperaTAb contains Fab fragments against the venom of V. berus. In 2014 the production of Zagreb antivenom was discontinued. Additionally, in the period of 2017 to 2018 a shortage of Viperfav occurred. Due to a lack of the product indicated for the treatment of V. ammodytes bites, other antivenoms were implemented into clinical practice without comparative assessment of their eligibility. The aim of our work was to identify a high-quality antivenom that might ensure the successful treatment of V. ammodytes and V. berus bites at the preclinical level. Differentiation between bites from these two species is difficult and unreliable in clinical practice, so the availability of a unique antivenom applicable in the treatment of envenoming caused by both species would be the most advantageous for Southeastern Europe. Zagreb antivenom, Viperfav, and ViperaTAb, as well as Viper venom antitoxin for V. berus envenoming and the in-development Inoserp Europe, which was designed to treat envenoming caused by all medically important European snakes, were comparatively tested for the first time. Emphasis was placed on their physicochemical properties, primarily purity and aggregate content, as well as their in vivo protective efficacies. As Zagreb antivenom is no longer available on the European market, Viperfav is the highest-quality product currently available and the only antivenom whose neutralisation potency against V. ammodytes and V. berus venoms was above regulatory requirements.

Analysis of the venomous snakebite patients treated in the Uzice general hospital (Western Serbia) between 2006 and 2018

Introduction/Objective. A countrywide survey of venomous snakebites was never conducted in Serbia: the making of a central register was initiated only in 2018. We collected and analyzed the available data regarding venomous snakebites in the Uzice region (Western Serbia). The previous analysis from this hospital was published in 1968. Methods. We retrospectively analyzed the data regarding the venomous snakebite patients treated in the Uzice General Hospital between 2006 and 2018 and compared these with the data from the 1960s, from one more hospital in Serbia, and from two ex- Yugoslav countries. Results. In 13 years, 249 persons were treated. Of all cases, 10.4% were with inconspicuous symptoms (mild pain at the place of bite), 68.7% were with mild to moderate symptoms, and 20.9% were more or less severe. No fatalities were recorded. Conclusion. Although usually not a life-threatening issue, venomous snakebites are quite common and can cause serious complications. With proper education, many can be avoided. Also, bearing in mind not only the biodiversity per se but also the importance of snakes? venoms for the making of various medically important products, we emphasize the need for proper protection of all three venomous snake species in Serbia, namely Vipera ammodytes (nose-horned viper), V. berus (European adder), and V. ursinii (meadow viper).

Crepuscular and nocturnal activity of the Nose-horned viper, Vipera ammodytes (Linnaeus, 1758) is more common than previously reported

Different snake species may be active at various times of the day. In Europe, most snake species are predominantly diurnal, but about a third are partially or principally crepuscular and/or nocturnal. Here, we report the first instance of multiple in situ observations of the crepuscular and nocturnal activity of V. ammodytes in Bulgaria. Overall, usually as a result of general herpetological surveys and chance observations between 2001–2020, we recorded crepuscular/nocturnal activity of twenty-seven individuals from both sexes and all age classes (juvenile to adult), observed from April to September, following warm (maximum daily Tair = 20–28 °C) and hot (Tair > 28 °C) days. These records represent less than 1.5% of all our personal viper observations. However, given that our observations occurred throughout the country, within five different climatic zones, we hypothesise that nocturnal activity of V. ammodytes is more common than previously reported.