Report from the 29th Meeting on Toxinology, "Toxins: From the Wild to the Lab", Organized by the French Society of Toxinology on 30 November-1 December 2023.

The French Society of Toxinology (SFET), which celebrated its 30th anniversary this year, organized its 29th annual Meeting (RT29), shared by 87 participants, on 30 November-1 December 2023. The RT29 main theme, "Toxins: From the Wild to the Lab", focused on research in the field of animal venoms and animal, bacterial, fungal, or plant toxins, from their discovery in nature to their study in the laboratory. The exploration of the functions of toxins, their structures, their molecular or cellular ligands, their mode of action, and their potential therapeutic applications were emphasized during oral communications and posters through three sessions, of which each was dedicated to a secondary theme. A fourth, "miscellaneous" session allowed participants to present recent out-of-theme works. The abstracts of nine invited and 15 selected lectures, those of 24 posters, and the names of the Best Oral Communication and Best Poster awardees, are presented in this report.

Peixes peçonhentos da bacia hidrográfica do Rio Paraná: aspectos ecológicos e toxinológicos de interesse em saúde

Aquatic ecosystems have a vast diversity of fauna and flora associated with them. Notably, rivers are of great social, economic, and environmental importance. The Paraná River basin comprises a river system used for energy production, recreation, production and consumption of fish resources. In addition, there is a record of a diverse fauna of venomous fish in this region which, due to their close proximity to the population, results in accidents known as ichthyism. Although these accidents are not lethal, they have a high morbidity rate and serious occupational consequences. Their main symptoms are intense pain, edema, and slow- healing lesions that can result in tissue necrosis. Albeit frequent, these accidents are underreported, making it difficult to map and treat them. Therefore, the aim of this study is to expand the knowledge about venomous fish by characterizing the venomous species found in the Paraná River with the potential to cause accidents. The venom and mucus samples were collected by the Taxonomy of Venomous Animals Laboratory - Toxicological Information and Assistance Center of Paraná and sent to the Immunoregulation Unit of the Applied Toxinology Laboratory of the Butantan Institute for analysis. The venoms were extracted by scraping the epithelium covering the spines and stingers and scraping the mucus covering the animal's body, and the supernatant was obtained by centrifugation. Proteins were quantified using the Bradford method and the protein profile was analyzed using polyacrylamide gel electrophoresis. A total of 133 individuals were collected, distributed among 19 species and 6 families (Auchenipteridae, Doradidae, Loricariidae, Pimelodidae, Cichlidae, and Myliobatiformes). The Pimelodidae family presented the highest species richness, with seven species in total. In addition, seven species collected occur outside their natural distribution area, with Pterodoras granulosus being the most abundant among all the samplings. The Auchenipteridae and Pimelodidae families have similar protein profiles, notably at 68, 38, 28, and 14 kDa in the stinger samples, although they vary for mucus. The Doradidae family, despite having two species from the same family (Pterodoras granulosus and Oxydoras kneri), did not show similarities in the protein pattern. The Cichlidae, with two families, showed only two similar bands (62 and 38 kDa). The expansion of venomous fish habitats can occur due to hydrological and limnological changes, such as river damming. Although it was expected that individuals from the same family would present similar protein profiles, this was not verified in the samples analyzed, suggesting greater diversity in the composition and types of venom. This work is the first endeavor towards notification and biochemical knowledge about fish species with the potential to cause accidents in the Paraná River basin and provides a better understanding of the ecological and toxicological characteristics of these animals.

Ecossistemas aquáticos possuem vasta diversidade associada, apresentando alto grau de importância social, econômica e ambiental, e os ambientes fluviais contemplam diversos animais e espécies vegetais que usufruem e dependem de suas propriedades para a sobrevivência, apresentando assim alto grau de endemismo. A bacia do Rio Paraná compreende um grande sistema fluvial utilizado para recreação, produção e consumo de recursos pesqueiros e produção de energia por uma das principais hidrelétricas do país, Itaipu Binacional. Devido às suas particularidades, a bacia do Rio Paraná possui registros de uma grande diversidade da fauna de peixes peçonhentos que, por estar em áreas próximas à população, resulta em acidentes denominados de ictismo. Essas ocorrências, apesar de apresentarem baixa letalidade, têm alta morbidade e sérias consequências ocupacionais, cujos principais sintomas são dor intensa, edema e lesões de lenta cicatrização que podem resultar em necrose tecidual da região afetada. Apesar de frequentes, tais acidentes são subnotificados, dificultando seu mapeamento e tratamento. Assim, o objetivo deste trabalho foi expandir o conhecimento sobre peixes peçonhentos, através da caracterização das espécies peçonhentas encontradas no Rio Paraná com potencial de causarem acidentes. As amostras de peçonha e muco foram coletadas pelo Laboratório de Taxonomia de Animais Peçonhentos - Centro de Informações e Assistência Toxicológica do Paraná e encaminhadas à Unidade de Imunorregulação do Laboratório de Toxinologia Aplicada do Instituto Butantan para análise. A extração das peçonhas ocorreu por raspado do epitélio que recobre espinhos e ferrões e raspado do muco que recobre o corpo do animal, obtendo-se sobrenadante através de centrifugação. A quantificação de proteínas se deu pelo método de Bradford e o perfil eletroforético foi analisado através de eletroforese em gel de poliacrilamida. Ao todo foram coletados 133 indivíduos, distribuídos em 19 espécies e 6 famílias (Auchenipteridae, Doradidae, Loricariidae, Pimelodidae, Cichlidae e Myliobatiformes). A família Pimelodidae apresentou maior riqueza com sete espécies ao todo. Além disso, sete espécies capturadas ocorrem fora de sua área de distribuição natural, sendo Pterodoras granulosus a mais abundante entre a soma de todas as coletas. As famílias Auchenipteridae e Pimelodidae possuem semelhanças no perfil de proteínas, notadamente em 68, 38, 28 e 14 kDa, nas amostras de espinhos, embora variando para muco. A família Doradidae, apesar de possuir duas espécies de mesma família (Pterodoras granulosus e Oxydoras kneri), não apresentou semelhanças no padrão de proteínas. Os Cichlidae, com duas famílias, apresentaram apenas duas bandas semelhantes (62 e 38 kDa). A expansão de habitats dos peixes peçonhentos pode ocorrer por alterações hidrológicas e limnológicas, como o represamento do rio. Apesar de esperado que indivíduos de mesma família apresentassem perfis proteicos semelhantes, isso não foi verificado, sugerindo maior diversidade de quantidades e tipos de peçonha. Este trabalho é o primeiro passo para a notificação e conhecimento bioquímico sobre as espécies de peixes com potencial de causarem acidentes e possibilita o maior entendimento acerca das características ecológicas e toxicológicas desses animais.

Pulmonary involvement from animal toxins: the cellular mechanisms.

Venomous animals and their venom have always been of human interest because, despite species differences, coevolution has made them capable of targeting key physiological components of our bodies. Respiratory failure from lung injury is one of the serious consequences of envenomation, and the underlying mechanisms are rarely discussed. This review aims to demonstrate how toxins affect the pulmonary system through various biological pathways. Herein, we propose the common underlying cellular mechanisms of toxin-induced lung injury: interference with normal cell function and integrity, disruption of normal vascular function, and provocation of excessive inflammation. Viperid snakebites are the leading cause of envenomation-induced lung injury, followed by other terrestrial venomous animals such as scorpions, spiders, and centipedes. Marine species, particularly jellyfish, can also inflict such injury. Common pulmonary manifestations include pulmonary edema, pulmonary hemorrhage, and exudative infiltration. Severe envenomation can result in acute respiratory distress syndrome. Pulmonary involvement suggests severe envenomation, thus recognizing these mechanisms and manifestations can aid physicians in providing appropriate treatment.

Diversity of transgenes in sustainable management of insect pests.

Insecticidal transgenes, when incorporated and expressed in plants, confer resistance against insects by producing several products having insecticidal properties. Protease inhibitors, lectins, amylase inhibitors, and chitinase genes are associated with the natural defenses developed by plants to counter insect attacks. Several toxin genes are also derived from spiders and scorpions for protection against insects. Bacillus thuringiensis Berliner is a microbial source of insecticidal toxins. Several methods have facilitated the large-scale production of transgenic plants. Bt-derived cry, cyt, vip, and sip genes, plant-derived genes such as lectins, protease inhibitors, and alpha-amylase inhibitors, insect cell wall-degrading enzymes like chitinase and some proteins like arcelins, plant defensins, and ribosome-inactivating proteins have been successfully utilized to impart resistance to insects. Besides, transgenic plants expressing double-stranded RNA have been developed with enhanced resistance. However, the long-term effects of transgenes on insect resistance, the environment, and human health must be thoroughly investigated before they are made available for commercial planting. In this chapter, the present status, prospects, and future scope of transgenes for insect pest management have been summarized and discussed.

Animal Toxins: A Historical Outlook at the Institut Pasteur of Paris.

Humans have faced poisonous animals since the most ancient times. It is recognized that certain animals, like specific plants, produce toxic substances that can be lethal, but that can also have therapeutic or psychoactive effects. The use of the term "venom", which initially designated a poison, remedy, or magic drug, is now confined to animal poisons delivered by biting. Following Louis Pasteur's work on pathogenic microorganisms, it was hypothesized that venoms could be related to bacterial toxins and that the process of pathogenicity attenuation could be applied to venoms for the prevention and treatment of envenomation. Cesaire Phisalix and Gabriel Bertrand from the National Museum of Natural History as well as Albert Calmette from the Institut Pasteur in Paris were pioneers in the development of antivenomous serotherapy. Gaston Ramon refined the process of venom attenuation for the immunization of horses using a formalin treatment method that was successful for diphtheria and tetanus toxins. This paved the way for the production of antivenomous sera at the Institut Pasteur, as well as for research on venom constituents and the characterization of their biological activities. The specific activities of certain venom components, such as those involved in blood coagulation or the regulation of chloride ion channels, raises the possibility of developing novel therapeutic drugs that could serve as anticoagulants or as a treatment for cystic fibrosis, for example. Scientists of the Institut Pasteur of Paris have significantly contributed to the study of snake venoms, a topic that is reported in this review.

Natural Peptide Toxins as an Option for Renewed Treatment of Type 2 Vasopressin Receptor-Related Diseases.

The type 2 vasopressin receptor (V2R) is expressed in the kidneys, and it is the keystone of water homeostasis. Under the control of the antidiuretic hormone vasopressin, the V2R ensures vital functions, and any disturbance has dramatic consequences. Despite decades of research to develop drugs capable of activating or blocking V2R function to meet real medical needs, only one agonist and one antagonist are virtually used today. These two drugs cover only a small portion of patients' needs, leaving millions of patients without treatment. Natural peptide toxins known to act selectively and at low doses on their receptor target could offer new therapeutic options.

Report from the 28th Meeting on Toxinology, "Toxins: What's up, Doc?", Organized by the French Society of Toxinology on 28-29 November 2022.

The French Society of Toxinology (SFET) organized its 28th annual meeting on 28-29 November 2022 (RT28). The central theme of this meeting was "Toxins: What's up, Doc?", emphasizing the latest findings on animal, bacterial, algal, plant and fungal toxins through sessions dedicated to deep learning, toxin tracking and toxinomic advances, shared by ca. 80 participants. The abstracts of the 10 invited and 11 selected lectures and 15 posters, along with the names of the Best Oral Communication and Best Poster awardees, are presented in this report.

Animal toxins: As an alternative therapeutic target following ischemic stroke condition.

Globally, Ischemic stroke (IS) has become the second leading cause of mortality and chronic disability. The process of IS has triggered by the blockages of blood vessels to form clots in the brain which initiates multiple interactions with the key signaling pathways, counting excitotoxicity, acidosis, ionic imbalance, inflammation, oxidative stress, and neuronal dysfunction of cells, and ultimately cells going under apoptosis. Currently, FDA has approved only tissue plasminogen activator therapy, which is effective against IS with few limitations. However, the mechanism of excitotoxicity and acidosis has spurred the investigation of a potential candidate for IS therapy. Acid-sensing ion channels (ASICs) and Voltage-gated Ca2+ channels (VDCCs) get activated and disturb the brain's normal physiology. Animal toxins are novel inhibitors of ASICs and VDCCs channels and have provided neuroprotective insights into the pathophysiology of IS. This review will discuss the potential directions of translational ASICs and VDCCs inhibitors research for clinical therapies.

Pulmonary involvement from animal toxins: the cellular mechanisms

Abstract Venomous animals and their venom have always been of human interest because, despite species differences, coevolution has made them capable of targeting key physiological components of our bodies. Respiratory failure from lung injury is one of the serious consequences of envenomation, and the underlying mechanisms are rarely discussed. This review aims to demonstrate how toxins affect the pulmonary system through various biological pathways. Herein, we propose the common underlying cellular mechanisms of toxin-induced lung injury: interference with normal cell function and integrity, disruption of normal vascular function, and provocation of excessive inflammation. Viperid snakebites are the leading cause of envenomation-induced lung injury, followed by other terrestrial venomous animals such as scorpions, spiders, and centipedes. Marine species, particularly jellyfish, can also inflict such injury. Common pulmonary manifestations include pulmonary edema, pulmonary hemorrhage, and exudative infiltration. Severe envenomation can result in acute respiratory distress syndrome. Pulmonary involvement suggests severe envenomation, thus recognizing these mechanisms and manifestations can aid physicians in providing appropriate treatment.

Report from the 27th (Virtual) Meeting on Toxinology, "Toxins: Mr Hyde or Dr Jekyll?", Organized by the French Society of Toxinology, 9-10 December 2021.

The French Society of Toxinology (SFET) organized its 27th annual meeting on 9-10 December 2021 as a virtual meeting (e-RT27). The central theme of this meeting was "Toxins: Mr Hyde or Dr Jekyll?", emphasizing the latest findings on plant, fungal, algal, animal and bacterial toxins during 10 lectures, 15 oral communications (shorter lectures) and 20 posters shared by ca. 80 participants. The abstracts of lectures and posters, as well as the winners of the best oral communication and poster awards, are presented in this report.

Design, Synthesis, and Antitumor Activities Study of Stapled A4K14-Citropin 1.1 Peptides.

A4K14-citropin 1.1 is a structurally optimized derivative derived from amphibians' skin secreta peptide Citropin, which exhibits broad biological activities. However, the application of A4K14-citropin 1.1 as a cancer therapeutic is restricted by its structural flexibility. In this study, a series of all-hydrocarbon stapled peptides derivatives of A4K14-citropin 1.1 were designed and synthesized, and their chemical and biological characteristics were also investigated. Among them, A4K14-citropin 1.1-Sp1 and A4K14-citropin 1.1-Sp4 displayed improved helicity levels, greater protease stability, and increased antitumor activity compared with the original peptide, which establishes them as promising lead compounds for novel cancer therapeutics development. These results revealed the important influence of all-hydrocarbon stapling side chain on the secondary structure, hydrolase stability, and biological activity of A4K14-citropin 1.1.

Cloning and identification of a new multifunctional Ascaris-type peptide from the hemolymph of Buthus martensii Karsch.

Only a few work have been done for peptides from non-venom gland tissues of venomous animals. Here, with the help of the whole body transcriptomic and the hemolymph proteomic data of the Chinese scorpion Buthus martensii Karsch, we identified the first Ascaris-type peptide BmHDP from scorpion hemolymph. The precursor of BmHDP has 80 residues, including a 16 residue signal peptide and a 64 residue mature peptide. The mature peptide has 10 conserved cysteines and adopts a conserved Ascaris-type fold. Using combined inclusion body refolding and biochemical identification strategies, recombinant BmHDP was obtained successfully. Protease inhibitory assays showed that BmHDP inhibited chymotrypsin apparently at a concentration of 8 nM. Patch-clamp experiments showed that BmHDP inhibited the Kv1.3 potassium channel apparently at a concentration of 1000 nM. Coagulation experiment assays showed that BmHDP inhibited intrinsic coagulation pathway apparently at a concentration of 500 nM. To the best of our knowledge, BmHDP is the first Ascaris-type peptide from scorpion hemolymph. Our work highlighted a functional link between scorpion non-venom gland peptides and venom gland toxin peptides, and suggested that scorpion hemolymph might be a new source of bioactive peptides.

Centipede Venom Peptides Acting on Ion Channels.

Centipedes are among the oldest venomous arthropods that use their venom to subdue the prey. The major components of centipede venom are a variety of low-molecular-weight peptide toxins that have evolved to target voltage-gated ion channels to interfere with the central system of prey and produce pain or paralysis for efficient hunting. Peptide toxins usually contain several intramolecular disulfide bonds, which confer chemical, thermal and biological stability. In addition, centipede peptides generally have novel structures and high potency and specificity and therefore hold great promise both as diagnostic tools and in the treatment of human disease. Here, we review the centipede peptide toxins with reported effects on ion channels, including Nav, Kv, Cav and the nonselective cation channel polymodal transient receptor potential vanilloid 1 (TRPV1).

Venom-Derived Peptide Modulators of Cation-Selective Channels: Friend, Foe or Frenemy.

Ion channels play a key role in our body to regulate homeostasis and conduct electrical signals. With the help of advances in structural biology, as well as the discovery of numerous channel modulators derived from animal toxins, we are moving toward a better understanding of the function and mode of action of ion channels. Their ubiquitous tissue distribution and the physiological relevancies of their opening and closing suggest that cation channels are particularly attractive drug targets, and years of research has revealed a variety of natural toxins that bind to these channels and alter their function. In this review, we provide an introductory overview of the major cation ion channels: potassium channels, sodium channels and calcium channels, describe their venom-derived peptide modulators, and how these peptides provide great research and therapeutic value to both basic and translational medical research.

Antitumoral effects of Amblyomma sculptum Berlese saliva in neuroblastoma cell lines involve cytoskeletal deconstruction and cell cycle arrest / Efeito antitumoral da saliva do carrapato Amblyomma sculptum Berlese em células de neuroblastoma envolve desconstrução do citoesqueleto e parada do ciclo celular

Abstract The antitumor properties of ticks salivary gland extracts or recombinant proteins have been reported recently, but little is known about the antitumor properties of the secreted components of saliva. The goal of this study was to investigate the in vitro effect of the saliva of the hard tick Amblyomma sculptum on neuroblastoma cell lines. SK-N-SK, SH-SY5Y, Be(2)-M17, IMR-32, and CHLA-20 cells were susceptible to saliva, with 80% reduction in their viability compared to untreated controls, as demonstrated by the methylene blue assay. Further investigation using CHLA-20 revealed apoptosis, with approximately 30% of annexin-V positive cells, and G0/G1-phase accumulation (>60%) after treatment with saliva. Mitochondrial membrane potential (Δψm) was slightly, but significantly (p < 0.05), reduced and the actin cytoskeleton was disarranged, as indicated by fluorescent microscopy. The viability of human fibroblast (HFF-1 cells) used as a non-tumoral control decreased by approximately 40%. However, no alterations in cell cycle progression, morphology, and Δψm were observed in these cells. The present work provides new perspectives for the characterization of the molecules present in saliva and their antitumor properties.

Resumo As propriedades antitumorais de extratos de glândulas salivares de carrapatos ou proteínas recombinantes foram relatadas recentemente, mas pouco se sabe sobre as propriedades antitumorais dos componentes secretados da saliva. O objetivo deste estudo foi investigar o efeito in vitro da saliva bruta do carrapato duro Amblyomma sculptum sobre as linhagens celulares de neuroblastoma. Células SK-N-SK, SH-SY5Y, Be(2)-M17, IMR-32 e CHLA-20 foram suscetíveis à saliva, com redução de 80% na sua viabilidade em comparação com controles não tratados, como demonstrado pelo ensaio de Azul de Metileno. Investigações posteriores utilizando CHLA-20 revelaram apoptose, com aproximadamente 30% de células positivas para anexina-V, e G0/G1 (> 60%) após tratamento com saliva. O potencial de membrana mitocondrial (Δψm) foi reduzido significativamente (p <0,05), e o citoesqueleto de actina foi desestruturado, como indicado pela microscopia de fluorescência. A viabilidade do fibroblasto humano (células HFF-1), usado como controle não tumoral, diminuiu em aproximadamente 40%. No entanto, não foram observadas alterações na progressão do ciclo celular, morfologia e Δψm nestas células. O presente trabalho fornece novas perspectivas para a caracterização das moléculas presentes na saliva e suas propriedades antitumorais.

Defensins, a novel type of animal toxin-like potassium channel inhibitor.

The classical potassium channel inhibitors are toxin peptides from venomous animals, and whether there are peptide inhibitors from other species is an open question. Due to both the independent and interdependent relationships between the spear (peptide inhibitors) and the shield (potassium channels), human defensins were first identified by our group as endogenous potassium channel inhibitors. Encouraged by the discovery of human defensins as potassium channel inhibitors, defensins from invertebrates and fungi were successively found by our group to be potassium channel inhibitors. In addition, a plant defensin was reported to be a potassium channel inhibitor. Since defensins are widely produced by vertebrate, invertebrate, plant and fungi species, the recent work established a new research field on defensin-potassium channel interactions. Here, we review the current work on defensins from vertebrate, invertebrate, plant and fungi species as inhibitors of potassium channels and discuss future work in this research field.

Antitumoral effects of Amblyomma sculptum Berlese saliva in neuroblastoma cell lines involve cytoskeletal deconstruction and cell cycle arrest

The antitumor properties of ticks salivary gland extracts or recombinant proteins have been reported recently, but little is known about the antitumor properties of the secreted components of saliva. The goal of this study was to investigate the in vitro effect of the saliva of the hard tick Amblyomma sculptum on neuroblastoma cell lines. SK-N-SK, SH-SY5Y, Be(2)-M17, IMR-32, and CHLA-20 cells were susceptible to saliva, with 80% reduction in their viability compared to untreated controls, as demonstrated by the methylene blue assay. Further investigation using CHLA-20 revealed apoptosis, with approximately 30% of annexin-V positive cells, and G0/G1-phase accumulation (>60%) after treatment with saliva. Mitochondrial membrane potential (??m) was slightly, but significantly (p < 0.05), reduced and the actin cytoskeleton was disarranged, as indicated by fluorescent microscopy. The viability of human fibroblast (HFF-1 cells) used as a non-tumoral control decreased by approximately 40%. However, no alterations in cell cycle progression, morphology, and ??m were observed in these cells. The present work provides new perspectives for the characterization of the molecules present in saliva and their antitumor properties.