Transient Hematotoxicity After Emerald Horned Pitviper (Ophryacus smaragdinus) Envenomation: A Case Report.

BACKGROUND: A minority of snake envenomations in the United States involve non-native snakes. In this report, we describe what we believe is the first documented human envenoming from an emerald horned pitviper, Ophryacus smaragdinus. CASE REPORT: A previously healthy 36-year-old woman was bitten on her left index finger by a captive emerald horned pitviper she was medicating at work. Swelling to the entire hand was present on emergency department arrival. She had no systemic symptoms and her initial laboratory studies were unremarkable. The affected limb was elevated. We administered five vials of Antivipmyn TRIⓇ (Bioclon), which specifically lists Ophryacus among the envenomations for which it is indicated. She developed pruritus and was treated with IV diphenhydramine and famotidine. Her swelling improved, but her repeat laboratory studies were notable for a platelet count of 102 K/µL and a fibrinogen level of 116 mg/dL. She declined additional antivenom because of the previous allergic reaction. She was admitted for further monitoring and pain control. Subsequent laboratory tests were better, but a small hemorrhagic bleb developed at the bite site. She was discharged the next day and followed up as an outpatient. Her swelling had resolved, her bleb had healed, and her laboratory studies continued to improve. WHY SHOULD AN EMERGENCY PHYSICIAN BE AWARE OF THIS?: Emergency physicians may be required to treat bites from non-native snakes. Many of these bites will warrant treatment with non-U.S. Food and Drug Administration-approved antivenoms. Consultation with a regional poison center or medical toxicologist may be necessary to procure the proper antivenom.

Acute ischaemic stroke and deep vein thrombosis following snakebite.

Snakebite envenomation remains a neglected tropical public health issue claiming thousands of lives every year. It is a common medical emergency and a threat to the impoverished populations of low-income and middle-income countries including India. A combination of ischaemic stroke and deep vein thrombosis is a devastating duo complication of snake bite, with no literature report to date. Here, the authors report an unusual case of a young woman developing ischaemic stroke and deep vein thrombosis following snakebite even after the use of antivenom. MRI brain showed right thalamic infarct with haemorrhagic transformation and, ultrasound Doppler revealed right lower limb deep vein thrombosis. The pathophysiology of deep vein thrombosis and ischaemic stroke is complex. It is believed that the activation of the coagulation cascade, complement system together with endothelial injury and immune activation leads to inflammation, thrombosis and occlusion of smaller and even larger vessels.

Improving in vivo assays in snake venom and antivenom research: A community discussion.

On the 26 th January 2023, a free to attend, 'improving in vivo snake venom research: a community discussion' meeting was held virtually. This webinar brought together researchers from around the world to discuss current neutralisation of venom lethality mouse assays that are used globally to assess the efficacy of therapies for snakebite envenoming. The assay's strengths and weaknesses were highlighted, and we discussed what improvements could be made to refine and reduce animal testing, whilst supporting preclinical antivenom and drug discovery for snakebite envenoming. This report summarises the issues highlighted, the discussions held, with additional commentary on key perspectives provided by the authors.

Mass spectrometric analysis of Odonthobuthus Doriae scorpion venom and its non-neutralized fractions after interaction with commercial antivenom.

It is believed that antivenoms play a crucial role in neutralizing venoms. However, uncontrolled clinical effects appear in patients stung by scorpions after the injection of antivenom. In this research, non-neutralized components of the venom of the Iranian scorpion Odonthobuthus doriae were analyzed after interacting with the commercial antivenom available in the market. The venom and antivenom interaction was performed, then centrifuged, and the supernatant was analyzed by high-performance liquid chromatography (HPLC). Two peaks of Odonthobuthus doriae venom were observed in the chromatogram of the supernatant. Two components were isolated by HPLC and analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) instruments. Peptide sequencing was done by Liquid Chromatography Quadrupole Time-of-Flight Tandem Mass Spectrometry (LC-Q-TOF MS/MS). Results indicate that the components of scorpion venom mainly have a molecular weight below 10 kDa, consisting of toxic peptides that disrupt the function of sodium and potassium channels. The MALDI-TOF MS results show that two toxic peptides with molecular masses of 6941 Da and 6396 Da were not neutralized by the antivenom. According to the MS/MS sequencing data, the components have been related to peptides A0A5P8U2Q6_MESEU and A0A0U4FP89_ODODO, which belong to the sodium and potassium channels toxins family, respectively.

Variability in antivenom neutralization of Mexican viperid snake venoms.

BACKGROUND: Each year, 3,800 cases of snakebite envenomation are reported in Mexico, resulting in 35 fatalities. The only scientifically validated treatment for snakebites in Mexico is the use of antivenoms. Currently, two antivenoms are available in the market, with one in the developmental phase. These antivenoms, produced in horses, consist of F(ab')2 fragments generated using venoms from various species as immunogens. While previous studies primarily focused on neutralizing the venom of the Crotalus species, our study aims to assess the neutralization capacity of different antivenom batches against pit vipers from various genera in Mexico. METHODOLOGY: We conducted various biological and biochemical tests to characterize the venoms. Additionally, we performed neutralization tests using all three antivenoms to evaluate their effectiveness against lethal activity and their ability to neutralize proteolytic and fibrinogenolytic activities. RESULTS: Our results reveal significant differences in protein content and neutralizing capacity among different antivenoms and even between different batches of the same product. Notably, the venom of Crotalus atrox is poorly neutralized by all evaluated batches despite being the primary cause of envenomation in the country's northern region. Furthermore, even at the highest tested concentrations, no antivenom could neutralize the lethality of Metlapilcoatlus nummifer and Porthidium yucatanicum venoms. These findings highlight crucial areas for improving existing antivenoms and developing new products. CONCLUSION: Our research reveals variations in protein content and neutralizing potency among antivenoms, emphasizing the need for consistency in venom characteristics as immunogens. While Birmex neutralizes more LD50 per vial, Antivipmyn excels in specific neutralization. The inability of antivenoms to neutralize certain venoms, especially M. nummifer and P. yucatanicum, highlights crucial improvement opportunities, given the medical significance of these species.

Thrombotic microangiopathy following a white-lipped pit viper envenomation.

Certain snake envenomation patients with consumptive coagulopathy, termed venom-induced consumption coagulopathy, develop thrombotic microangiopathy (TMA). Due to predominant renal involvement, TMA is said to resemble haemolytic uraemic syndrome and is treated with haemodialysis. We present a case of a young male who presented to the emergency department after being bitten by a white-lipped pit viper (Trimeresurus albolabris). He developed heart failure in addition to acute kidney injury secondary to TMA. He was treated with 30 vials of anti-snake venom according to national guidelines and underwent haemodialysis. Despite haemodialysis, the patient's ventilatory parameters continued to worsen, necessitating invasive mechanical ventilation. Thus, he was initiated on plasma exchange therapy, to which the patient responded well. TMA has not been reported in Trimeresurus envenomations yet, to the best of our knowledge. Additionally, plasma exchange therapy can be considered an adjunctive therapy for snakebite patients who develop TMA.

An indigenously developed tele-ICU model to treat severe cases of envenomation in remote regions in India.

In cases of severe envenomation due to snakebites, patients require antivenom, intensive care management, including respiratory support, haemodynamic monitoring and renal replacement therapy. Early recognition and treatment of complications such as acute kidney injury, rhabdomyolysis and coagulopathy are important to improve outcomes.Tele-ICU models can play a critical role in providing access to critical care expertise and nuanced support to remote healthcare facilities that may not have the necessary resources or expertise to manage complex cases of envenomation. With the help of telemedicine technology, remote intensivists can provide timely guidance on diagnosis and ongoing management, improving the quality of care and outcomes for patients. We discuss two patients in resource-constrained regions of India with severe envenomation who were managed with tele-ICU support.

Development and Efficacy of the Antivenom Specific to Severe Envenomations in Morocco and North Africa: Advancements in Scorpion Envenomation Management.

Scorpion envenomation poses a global public health issue, with an estimated 1,500,000 cases worldwide annually resulting in 2600 deaths. North Africa, particularly Morocco, experiences severe envenomations, mainly attributed to Androctonus mauretanicus and Buthus occitanus in Morocco, and Buthus occitanus and Androctonus australis hector in Algeria and Tunisia, with case numbers often underestimated. Current treatment relies mainly on symptomatic approaches, except in Morocco, where management is limited to symptomatic treatment due to controversies regarding specific treatment. In Morocco, between 30,000 and 50,000 scorpion envenomation cases are reported annually, leading to hundreds of deaths, mainly among children. Controversies among clinicians persist regarding the appropriate course of action, often limiting treatments to symptomatic measures. The absence of a specific antivenom for the venoms of the most lethal scorpions further exacerbates the situation. This study aims to address this gap by developing a monovalent antivenom against the endemic and most dangerous scorpion, Androctonus mauretanicus. The antivenom was produced by immunizing albino rabbits with a mixture of Androctonus mauretanicus venom collected from high-risk areas in Morocco. Immunizations were performed by subcutaneous injections at multiple sites near the lymphatic system, following an immunization schedule. Production control of neutralizing antibody titers was conducted through immunodiffusion. Once a sufficient antibody titer was achieved, blood collection was performed, and the recovered plasma underwent affinity chromatography. The efficacy of purified IgG was evaluated by determining the ED50 in mice, complemented by histological and immunohistochemical studies on its ability to neutralize venom-induced tissue alterations and the neutralization of toxins bound to receptors in the studied organs. The monovalent antivenom demonstrated specificity against Androctonus mauretanicus venom and effective cross-protection against the venom of the scorpions Buthus occitanus and Androctonus australis hector, highly implicated in lethal envenomations in the Maghreb. This study shows that the developed monovalent antivenom exhibits notable efficacy against local scorpions and a surprising ability to neutralize the most lethal envenomations in North Africa. These results pave the way for a new, more specific, and promising therapeutic approach to countering severe scorpion envenomations, especially in Morocco, where specific treatment is lacking.

Pulmonary Thromboembolism following Russell's Viper Bites.

Snakebite envenoming and its resulting complications are serious threats to the health of vulnerable people living in rural areas of developing countries. The knowledge of the heterogeneity of symptoms associated with snakebite envenoming and their management strategies is vital to treat such life-threatening complications to save lives. Russell's viper envenomation induces a diverse range of clinical manifestations from commonly recognised haemotoxic and local effects to several rare conditions that are often not reported. The lack of awareness about these unusual manifestations can affect prompt diagnosis, appropriate therapeutic approaches, and positive outcomes for patients. Here, we report pulmonary thromboembolism that developed in three patients following Russell's viper envenomation and demonstrate their common clinical features and diagnostic and therapeutic approaches used. All patients showed clinical signs of local (oedema) and systemic (blood coagulation disturbances) envenomation, which were treated using polyvalent antivenom. They exhibited elevated heart rates, breathlessness, and reduced oxygen saturation, which are non-specific but core parameters in the diagnosis of pulmonary embolism. The recognition of pulmonary embolism was also achieved by an electrocardiogram, which showed sinus tachycardia and computed tomography and echocardiogram scans further confirmed this condition. Anti-coagulant treatment using low-molecular-weight heparin offered clinical benefits in these patients. In summary, this report reinforces the broad spectrum of previously unreported consequences of Russell's viper envenomation. The constant updating of healthcare professionals and the dissemination of major lessons learned in the clinical management of snakebite envenoming through scientific documentation and educational programs are necessary to mitigate the adverse impacts of venomous snakebites in vulnerable communities.

In vivo neutralization of coral snake venoms with an oligoclonal nanobody mixture in a murine challenge model.

Oligoclonal mixtures of broadly-neutralizing antibodies can neutralize complex compositions of similar and dissimilar antigens, making them versatile tools for the treatment of e.g., infectious diseases and animal envenomations. However, these biotherapeutics are complicated to develop due to their complex nature. In this work, we describe the application of various strategies for the discovery of cross-neutralizing nanobodies against key toxins in coral snake venoms using phage display technology. We prepare two oligoclonal mixtures of nanobodies and demonstrate their ability to neutralize the lethality induced by two North American coral snake venoms in mice, while individual nanobodies fail to do so. We thus show that an oligoclonal mixture of nanobodies can neutralize the lethality of venoms where the clinical syndrome is caused by more than one toxin family in a murine challenge model. The approaches described may find utility for the development of advanced biotherapeutics against snakebite envenomation and other pathologies where multi-epitope targeting is beneficial.

The use of rotational thromboelastometry to guide management following Bitis nasicornis envenoming.

A man in his thirties presented following Bitis nasicornis envenoming. His coagulation was assessed using rotational thromboelastometry (ROTEM). It identified a subtle abnormality, not detected using standard laboratory assessments of coagulation, and influenced ongoing management. The abnormality resolved following treatment with antivenom. There are few documented cases of using ROTEM to assess patients following haemotoxic envenoming. This case highlights some of the potential benefits and limitations of doing so.

Protobothrops jerdonii (Jerdon's pit viper) and Protobothrops himalayanus (Himalayan lance-headed pit viper) bites: Clinical report on envenomings from North-East India, managed through remote consultation by a national-level Poison control center.

Members of the genus Protobothrops are amongst the more than twenty-eight range-restricted Indian pit viper species. Their bites and envenomings are rarely documented from India. Pit viper envenomings can be challenging to treat in the Indian setting, since available antivenoms do not satisfactorily neutralize their venoms. Herein, we present the first Indian reports on bites and envenoming by Protobothrops jerdonii and Protobothrops himalayanus resulting in local effects, coagulopathy and acute kidney injury in the case of the former and possible mild, isolated coagulopathy in the case of the latter; and discuss management-related challenges in the context of absent specific antivenoms.

Snakebite envenoming: A systematic review and meta-analysis of global morbidity and mortality.

BACKGROUND: Snakebite envenoming represents a significant and often neglected public health challenge, particularly in rural communities across tropical and subtropical regions. An estimated 1.2-5.5 million people are envenomed by snakebites annually. More than 125,000 of these bites are fatal, and 3-4 times as many results in disability/disfigurement. Despite its prevalence, collecting accurate epidemiological data on snakebite is challenging. This systematic review and meta-analysis collates global epidemiology data on snakebite morbidity and mortality. METHODS: Medline, Embase, Cochrane and CINAHL Plus databases were searched for articles published between 2001-2022. Pooled incidence and mortality were obtained using random effects modelling, heterogeneity (I2) was tested, and sensitivity analyses performed. Newcastle-Ottawa Scale assessed study quality. RESULTS: Out of the four databases, 5,312 articles were found. After removing duplicates, 3,953 articles were screened by title and abstract and 65 articles containing information on snakebite epidemiology, encompassing 663,460 snakebites, were selected for analysis. The people most at risk for snakebite were men (59%), engaged in agricultural labour (27.5%), and residing in rural areas (66.7%). More than half (57%) of the reported bites resulted in envenoming. Incidents occurred frequently in the summer season (38.5%), during daytime (56.7%), and bites were most often to the lower limb (56.4%). Envenoming severity was frequently mild (46.7%), treated in hospital (68.3%), and was treated with anti-venom (64.7%). The pooled global incidence and mortality was 69.4 /100,000 population (95%CI: 36.8 to 101.9) and 0.33/100,000 population (95%CI, 0.14 to 0.52) per year, respectively. Stratified by continents, Asia had the highest incidence of 130.7/100,000 population (95%CI: 48.3 to 213.1) while Europe has the lowest with 0.7/100,000 population (95%CI: -0.2 to 1.5). The highest mortality was reported in Asia at 0.96/100,000 population (95% CI: 0.22 to 1.70), and Africa 0.44/100,000 population (95%CI: -0.03 to 0.84). Incidence was highest among inhabitants of lower-middle-income countries 132.7/100,000 population (95%CI: 55.4 to 209.9) while mortality was highest in low-income countries at 0.85/100,000 population (95% CI: -0.06 to 2.31). CONCLUSION: Incidence and mortality rates noted here highlight the global impact of snakebite and underscore the critical need to address the burden of snakebite envenoming. It also reveals that while reported snakebite incidence was higher in lower-middle-income countries, the burden of mortality was greatest among inhabitants of low-income countries, again emphasising the need for greater efforts to tackle this neglected tropical disease.

Analysis of intra-specific variations in the venom of individual snakes based on Raman spectroscopy.

The knowledge of variations in the composition of venoms from different snakes is important from both theoretical and practical points of view, in particular, at developing and selecting an antivenom. Many studies on this topic are conducted with pooled venoms, while the existence and significance of variations in the composition of venoms between individual snakes of the same species are emphasized by many authors. It is important to study both inter- and intra-specific, including intra-population, venom variations, because intra-specific variations in the venom composition may affect the effectiveness of antivenoms as strongly as inter-specific. In this work, based on venom Raman spectroscopy with principal component analysis, we assessed the variations in venoms of individual snakes of the Vipera nikolskii species from two populations and compared these intra-specific variations with inter-specific variations (with regard to the other related species). We demonstrated intra-specific (inter- and intra-population) differences in venom compositions which are smaller than inter-specific variations. We also assessed the compositions of V. nikolskii venoms from two populations to explain inter-population differences. The method used is rapid and requires virtually no preparation of samples, used in extremely small quantities, allowing the venoms of individual snakes to be analyzed. In addition, the method is informative and capable of detecting fairly subtle differences in the composition of venoms.

Venom diversity in Naja mossambica: Insights from proteomic and immunochemical analyses reveal intraspecific differences.

BACKGROUND: Intraspecific variations in snake venom composition have been extensively documented, contributing to the diverse clinical effects observed in envenomed patients. Understanding these variations is essential for developing effective snakebite management strategies and targeted antivenom therapies. We aimed to comprehensively investigate venoms from three distinct populations of N. mossambica from Eswatini, Limpopo, and KwaZulu-Natal regions in Africa in terms of their protein composition and reactivity with three commercial antivenoms (SAIMR polyvalent, EchiTAb+ICP, and Antivipmyn Africa). METHODOLOGY/PRINCIPAL FINDINGS: Naja mossambica venoms from Eswatini region exhibited the highest content of neurotoxic proteins, constituting 20.70% of all venom proteins, compared to Limpopo (13.91%) and KwaZulu-Natal (12.80%), and was characterized by the highest diversity of neurotoxic proteins, including neurotoxic 3FTxs, Kunitz-type inhibitors, vespryns, and mamba intestinal toxin 1. KwaZulu-Natal population exhibited considerably lower cytotoxic 3FTx, higher PLA2 content, and significant diversity in low-abundant proteins. Conversely, Limpopo venoms demonstrated the least diversity as demonstrated by electrophoretic and mass spectrometry analyses. Immunochemical assessments unveiled differences in venom-antivenom reactivity, particularly concerning low-abundance proteins. EchiTAb+ICP antivenom demonstrated superior reactivity in serial dilution ELISA assays compared to SAIMR polyvalent. CONCLUSIONS/SIGNIFICANCE: Our findings reveal a substantial presence of neurotoxic proteins in N. mossambica venoms, challenging previous understandings of their composition. Additionally, the detection of numerous peptides aligning to uncharacterized proteins or proteins with unknown functions underscores a critical issue with existing venom protein databases, emphasizing the substantial gaps in our knowledge of snake venom protein components. This underscores the need for enhanced research in this domain. Moreover, our in vitro immunological assays suggest EchiTAb+ICP's potential as an alternative to SAIMR antivenom, requiring confirmation through prospective in vivo neutralization studies.

Standard Quality Characteristics and Efficacy of a New Third-Generation Antivenom Developed in Colombia Covering Micrurus spp. Venoms.

In Colombia, Micrurus snakebites are classified as severe according to the national clinical care guidelines and must be treated with specific antivenoms. Unfortunately, these types of antivenoms are scarce in certain areas of the country and are currently reported as an unavailable vital medicine. To address this issue, La Universidad de Antioquia, through its spin-off Tech Life Saving, is leading a project to develop third-generation polyvalent freeze-dried antivenom. The goal is to ensure access to this therapy, especially in rural and dispersed areas. This project aims to evaluate the physicochemical and preclinical parameters (standard quality characteristics) of a lab-scale anti-elapid antivenom batch. The antivenom is challenged against the venoms of several Micrurus species, including M. mipartitus, M. dumerilii, M. ancoralis, M. dissoleucus, M. lemniscatus, M. medemi, M. spixii, M. surinamensis, and M. isozonus, following the standard quality characteristics set by the World Health Organization (WHO). The antivenom demonstrates an appearance consistent with standards, 100% solubility within 4 min and 25 s, an extractable volume of 10.39 mL, a pH of 6.04, an albumin concentration of 0.377 mg/mL (equivalent to 1.22% of total protein), and a protein concentration of 30.97 mg/mL. Importantly, it maintains full integrity of its F(ab')2 fragments and exhibits purity over 98.5%. Furthermore, in mice toxicity evaluations, doses up to 15 mg/mouse show no toxic effects. The antivenom also demonstrates a significant recognition pattern against Micrurus venoms rich in phospholipase A2 (PLA2) content, as observed in M. dumerilii, M. dissoleucus, and M. isozonus. The effective dose 50 (ED50) indicates that a single vial (10 mL) can neutralize 2.33 mg of M. mipartitus venom and 3.99 mg of M. dumerilii venom. This new anti-elapid third-generation polyvalent and freeze-dried antivenom meets the physicochemical parameters set by the WHO and the regulators in Colombia. It demonstrates significant efficacy in neutralizing the venom of the most epidemiologically important Micrurus species in Colombia. Additionally, it recognizes seven other species of Micrurus venom with a higher affinity for venoms exhibiting PLA2 toxins. Fulfilling these parameters represents the first step toward proposing a new pharmacological alternative for treating snakebites in Colombia, particularly in dispersed rural areas, given that this antivenom is formulated as a freeze-dried product.

Harnessing the Cross-Neutralisation Potential of Existing Antivenoms for Mitigating the Outcomes of Snakebite in Sub-Saharan Africa.

Over 32,000 individuals succumb to snake envenoming in sub-Saharan Africa (sSA) annually. This results from several factors, including a lack of antivenom products capable of neutralising the venoms of diverse snake species in this region. Most manufacturers produce polyvalent antivenoms targeting 3 to 16 clinically important snake species in sSA. However, specific products are unavailable for many others, especially those with a restricted geographic distribution. While next-generation antivenoms, comprising a cocktail of broadly neutralising antibodies, may offer an effective solution to this problem, given the need for their clinical validation, recombinant antivenoms are far from being available to snakebite victims. One of the strategies that could immediately address this issue involves harnessing the cross-neutralisation potential of existing products. Therefore, we assessed the neutralisation potency of PANAF-Premium antivenom towards the venoms of 14 medically important snakes from 13 countries across sSA for which specific antivenom products are unavailable. Preclinical assays in a murine model of snake envenoming revealed that the venoms of most snake species under investigation were effectively neutralised by this antivenom. Thus, this finding highlights the potential use of PANAF-Premium antivenom in treating bites from diverse snakes across sSA and the utility of harnessing the cross-neutralisation potential of antivenoms.

Effect of Seaweed-Derived Fucoidans from Undaria pinnatifida and Fucus vesiculosus on Coagulant, Proteolytic, and Phospholipase A2 Activities of Snake Bothrops jararaca, B. jararacussu, and B. neuwiedi Venom.

BACKGROUND: Snakebite envenomation (SBE) causes diverse toxic effects in humans, including disability and death. Current antivenom therapies effectively prevent death but fail to block local tissue damage, leading to an increase in the severity of envenomation; thus, seeking alternative treatments is crucial. METHODS: This study analyzed the potential of two fucoidan sulfated polysaccharides extracted from brown seaweeds Fucus vesiculosus (FVF) and Undaria pinnatifida (UPF) against the fibrinogen or plasma coagulation, proteolytic, and phospholipase A2 (PLA2) activities of Bothrops jararaca, B. jararacussu, and B. neuwiedi venom. The toxicity of FVF and UPF was assessed by the hemocompatibility test. RESULTS: FVF and UPF did not lyse human red blood cells. FVF and UPF inhibited the proteolytic activity of Bothrops jararaca, B. jararacussu, and B. neuwiedi venom by approximately 25%, 50%, and 75%, respectively, while all venoms led to a 20% inhibition of PLA2 activity. UPF and FVF delayed plasma coagulation caused by the venoms of B. jararaca and B. neuwiedi but did not affect the activity of B. jararacussu venom. FVF and UPF blocked the coagulation of fibrinogen induced by all these Bothropic venoms. CONCLUSION: FVF and UPF may be of importance as adjuvants for SBE caused by species of Bothrops, which are the most medically relevant snakebite incidents in South America, especially Brazil.