Taking the sting out of scorpions: Electrophysiological investigation of the relative efficacy of three antivenoms against medically significant Centruroides species.

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 (hNaV) channel subtypes: hNaV1.4 and hNaV1.5, 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, hNaV1.4. 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.

Tiny but Mighty: Vipera ammodytes meridionalis (Eastern Long-Nosed Viper) Ontogenetic Venom Variations in Procoagulant Potency and the Impact on Antivenom Efficacies.

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.

Extreme Procoagulant Potency in Human Plasma of Venoms from the African Viperid Genera Atheris, Cerastes, and Proatheris and the Relative Efficacy of Antivenoms and Synthetic Enzyme-Inhibitors.

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.

Effect of the bio-inspired modification of low-cost membranes with TiO2:ZnO as microbial fuel cell membranes.

Microbial fuel cells (MFCs) are a novel technique for converting biodegradable materials into electricity. In this study, the efficiency of mixed crystal (TiO2:ZnO) as a membrane modifier of a low-cost, antifouling and self-cleaning cation exchange membrane for MFCs was studied. The modification was prepared using polydopamine (PDA) as the bio-inspired glue, followed by gravity deposition of a mixture of catalyst nanoparticles (TiO2:ZnO 0.03%, 1:1 ratio) as anti-biofouling agents. The effects of the membrane modification were evaluated in terms of power density, open circuit potential, coulombic efficiency, anti-biofouling properties and also color and COD removal efficiency. The results showed that the use of the PDA-modified membrane and a mixture of catalysts facilitated the transfer of cations released during the oxidation process in the anodic compartment of the MFC, which increased the power generation in the MFC by 2.5 times and 5.7 times the current compared to pristine and PDA pristine membranes, decreased the MFC operating cycle time from 5 to 3 days, doubled the lifetime of the membranes and demonstrated higher COD removal efficiency and color removal. Finally, SEM and AFM analysis showed that the modification significantly minimized surface fouling. The modified membranes in this study proved to be a potential alternative to the expensive membranes currently used in MFCs, furthermore, this modification could be an interesting alternative modification for other potential membranes for use in MFCs, due to the fact that the catalyst activation was only performed with visible light (artificial and solar), which could decrease operating costs.

Clinical implications of differential procoagulant toxicity of the palearctic viperid genus Macrovipera, and the relative neutralization efficacy of antivenoms and enzyme inhibitors.

Species within the viperid genus Macrovipera are some of the most dangerous snakes in the Eurasian region, injecting copious amounts of potent venom. Despite their medical importance, the pathophysiological actions of their venoms have been neglected. Particularly poorly known are the coagulotoxic effects and thus the underlying mechanisms of lethal coagulopathy. In order to fill this knowledge gap, we ascertained the effects of venom upon human plasma for Macrovipera lebetina cernovi, M. l. lebetina, M. l. obtusa, M. l. turanica, and M. schweizeri using diverse coagulation analysing protocols. All five were extremely potent in their ability to promote clotting but varied in their relative activation of Factor X, being equipotent in this study to the venom of the better studied, and lethal, species Daboia russelii. The Insoserp European viper antivenom was shown to be highly effective against all the Macrovipera venoms, but performed poorly against the D. russelii venom. Reciprocally, while Daboia antivenoms performed well against D. russelii venom, they failed against Macrovipera venom. Thus despite the two genera sharing a venom phenotype (Factor X activation) driven by the same toxin type (P-IIId snake venom metalloproteases), the surface biochemistries of the toxins differed significantly enough to impede antivenom cross- neutralization. The differences in venom biochemistry were reflected in coagulation co-factor dependence. While both genera were absolutely dependent upon calcium for the activation of Factor X, dependence upon phospholipid varied. The Macrovipera venoms had low levels of dependence upon phospholipid while the Daboia venom was three times more dependent upon phospholipid for the activation of Factor X. This suggests that the sites on the molecular surface responsible for phospholipid dependence, are the same differential sites that prevent inter-genera antivenom cross- neutralization. Due to cold-chain requirements, antivenoms may not be stocked in rural settings where the need is at the greatest. Thus we tested the efficacy of enzyme inhibitor Prinomastat as a field-deployable treatment to stabilise patients while being transported to antivenom stocks, and showed that it was extremely effective in blocking the Factor X activating pathophysiological actions. Marimastat however was less effective. These results thus not only shed light on the coagulopathic mechanisms of Macrovipera venoms, but also provide data critical for evidence-based design of snakebite management strategies.

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.

Preclinical Assessment of a New Polyvalent Antivenom (Inoserp Europe) against Several Species of the Subfamily Viperinae.

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.

Application of a direct current circuit to pick up and to store bioelectricity produced by microbial fuel cells / Aplicación de un circuito de corriente continua para la colección y almacenamiento de bioelectricidad producida por celdas de combustible microbianas / Aplicação de um circuito de corrente contínua para coleta e armazenamento de bioeletricidade produzida por células a combustível microbianas

Abstract Every year the demand for energy worldwide is increasing. There are some alternatives to reduce these problems, such as clean energy or renewable energy. A particular alternative is the microbial fuel cells. These cells are biochemical reactors that convert chemical energy into electricity. The present research evaluated the dairy serum to produce bioelectricity from micro fuel cells (MFC) that were constructed with low-cost materials and with isolated bacteria in anaerobic sediments, located in Ecuadorian national territory, producing maximum voltages of 0.830 V in the circuit and a maximum power density of 30 mW / m2. This low voltage was worked with 50 mL MFCs and with an output voltage of 300 mV. Under these conditions, a FLYBACK lift circuit isolated by the transformer was designed. This new circuit could increase the voltage from 30 mV to enough voltage to light a 2.5 V LED. Therefore, the energy produced by the MFC can be directly used to light a LED and to charge capacitors. This study shows that these MFCs, together with the designed circuit, could be used potentially to generate clean energy.

Resumen Cada año la demanda de energía, en todo el mundo, va en aumento. Existen algunas alternativas para reducir estos problemas, tales como las energías limpias y renovables. Una alternativa muy específica es el uso de celdas de combustible microbianas. Dichas celdas son reactores bioquímicos que convierten la energía química en electricidad. La presente investigación evaluó el suero lácteo para la producción de bioelectricidad en celdas de combustible microbianas (MFC). Estas fueron construidas con materiales de bajo costo y con bacterias aisladas en sedimentos anaeróbicos, ubicados en territorio nacional ecuatoriano, produciendo voltajes máximos de 0,830 V en el circuito y una densidad de potencia máxima de 30 mW / m2. Este bajo voltaje se trabajó con MFC de 50 mL y con un voltaje de salida de 300 mV. Bajo estas condiciones, se diseñó un circuito de elevación FLYBACK aislado por transformador. Este nuevo circuito aumentará el voltaje de 30 mV a un voltaje suficiente para encender un LED de 2.5 V. Por lo tanto, la energía producida por las MFC puede ser directamente utilizable para encender un LED y cargar los condensadores. Este estudio muestra que dichas celdas MFC, junto con el circuito diseñado, podrían utilizarse, potencialmente, para generar energía limpia.

Resumo Todos os anos a demanda por energia, em todo o mundo, está aumentando. Existem algumas alternativas para reduzir esses problemas, como energias limpas e renováveis. Uma alternativa muito específica é o uso de células combustíveis microbianas. Essas células são reatores bioquímicos que convertem energia química em eletricidade. O presente trabalho avaliou o soro lácteo para a produção de bioeletricidade em células a combustível microbianas (CCM), Estes foram construídos com materiais de baixo custo e bactérias isoladas em sedimentos anaeróbios, localizados no território nacional equatoriano, produzindo tensões máximas de 0,830 V no circuito e uma densidade de potência máxima de 30 mW / m2. Esta baixa voltagem trabalhamos com CCM de 50 mL e com uma voltagem de saída de 300 mV. Sob essas condições, um circuito de elevação FLYBACK isolado por transformador foi projetado. Este novo circuito aumentará a tensão de 30 mV para uma tensão suficiente para ligar um LED de 2,5 V. Portanto, a energia produzida pelo MFC pode ser diretamente utilizável para ligar um LED e carregar os capacitores. Este estudo mostra que essas células CCM, juntamente com o circuito projetado, poderiam ser usadas para gerar energia limpa.