Toxinology in the proteomics era: a review on arachnid venom proteomics

The word venomics was coined to acknowledge the studies that use omics to investigate venom proteins and peptides. Venomics has evolved considerably over the last 20 years. The first works on scorpion or spider venomics were published in the early 2000's. Such studies relied on peptide mass fingerprinting (PMF) to characterize venom complexity. After the introduction of new mass spectrometers with higher resolution, sensitivity and mass accuracy, and the next-generation nucleotide sequencing, the complexity of data reported in research on scorpion and spider venomics increased exponentially, which allowed more comprehensive studies. In the present review article, we covered key publications on scorpion venomics and spider venomics, presenting historical grounds and implemented technologies over the last years. The literature presented in this review was selected after searching the PubMed database using the terms "(scorpion venom) AND (proteome)" for scorpion venomics, and "(spider venom) AND (proteome)" for publications on spider venomics. We presented the key aspects related to proteomics in the covered papers including, but not restricted to, the employed proteomic strategy (i.e., PMF, two-dimensional gel electrophoresis, shotgun/bottom-up and/or top-down/peptidome), and the type of mass spectrometer used. Some conclusions can be drawn from the present study. For example, the scorpion genus Tityus is the most studied concerning venomics, followed by Centruroides; whereas for spiders the studied genera were found more equally distributed. Another interesting conclusion is the lack of high throughput studies on post-translational modifications (PTMs) of scorpion and spider proteins. In our opinion, PTMs should be more studied as they can modulate the activity of scorpion and spider toxins.(AU)

Recombinant expression and antigenicity of two peptide families of neurotoxins from Androctonus sp

Background: Scorpion neurotoxins such as those that modify the mammalian voltagegated sodium ion channels (Nav) are the main responsible for scorpion envenomation. Their neutralization is crucial in the production of antivenoms against scorpion stings. Methods: In the present study, two in silico designed genes ­ one that codes for a native neurotoxin from the venom of the Anatolian scorpion Androctonus crassicauda, named Acra 4 ­ and another non-native toxin ­ named consensus scorpion toxin (SccTx) obtained from the alignment of the primary structures of the most toxic neurotoxins from the Middle Eastern and North African scorpions ­ were recombinantly expressed in E. coli Origami. Results: Following bacterial expression, the two expressed neurotoxins, hereafter named HisrAcra4 and HisrSccTx, were obtained from inclusion bodies. Both recombinant neurotoxins were obtained in multiple Cys-Cys isoforms. After refolding, the active protein fractions were identified with molecular masses of 8,947.6 and 9,989.1 Da for HisrAcra4 and HisrSccTx, respectively, which agreed with their expected theoretical masses. HisrAcra4 and HisrSccTx were used as antigens to immunize two groups of rabbits, to produce either anti-HisrAcra4 or anti-HisrSccTx serum antibodies, which in turn could recognize and neutralize neurotoxins from venoms of scorpion species from the Middle East and North Africa. The antibodies obtained from rabbits neutralized the 3LD50 of Androctonus australis, Leiurus quinquestriatus hebraeus and Buthus occitanus venoms, but they did not neutralize A. crassicauda and A. mauritanicus venoms. In addition, the anti-HisrAcra4 antibodies did not neutralize any of the five scorpion venoms tested. However, an antibody blend of anti-HisrAcra4 and anti-HisrSccTx was able to neutralize A. crassicauda and A. mauritanicus venoms. Conclusions: Two recombinant Nav neurotoxins, from different peptide families, were used as antigens to generate IgGs for neutralizing scorpion venoms of species from the Middle East and North Africa.(AU)

Voracity, reaction to stings, and survival of domestic hens when feeding on the yellow scorpion (Tityus serrulatus)

Scorpionism is a worldwide problem that has already made thousands of victims, and multi-disciplinary approaches for controlling their populations are to be more successful. Hens are often mentioned as tools for controlling scorpions; however, systematic/experimental behavioral studies are not available. Moreover, there is no systematic information on the effect of scorpion venoms on hens. Using the venomous yellow scorpion Tityus serrulatus, the present study aimed to clarify the following aspects: (1) voracity of hens, (2) how hens react when stung, (3) the effect of scorpion stings on hen behavior during attacks, and (4) hen survivorship after feeding on scorpions. Methods: We attracted hens with corn powder, offered them scorpions and then recorded the hen-scorpion interaction. To test the effects of the sting we manually removed the scorpion's telson. Results: We found that some hens ate up to six scorpions within minutes. By means of an ethogram and drawings, we showed that they exhibited several aversive behaviors when capturing scorpions. Removal of the scorpion telson stopped the aversive reactions, which was not observed in the control group. Finally, hens did not exhibit atypical behaviors after 1, 7 and 30 days and were all alive after 30 days. Conclusion: This is the first empirical and video recorded study providing evidence that hens are clearly affected by scorpion venom but do not die. Therefore, they may have potential to be used in biological control of these arthropods.(AU)

Proteinograma de ovelhas imunizadas com veneno detoxificado do escorpião Tityus serrulatus / Proteinogram of immunized sheep with detoxified Tityus serrulatus scorpion venom

Background: Scorpionism is a worldwide medical issue, especially relevant in the tropical and subtropical countries. Tityusserrulatus is the species responsible for most cases in Brazil. Antivenom administration to victims is the sole specific therapyobtained from donor animals. Most of these donors suffer with symptoms of the poisoning, debilitating their health andreducing their life expectancy. The aim of the present research was to evaluate whether the immunogens prepared fromthe crude and detoxified venom of T. serrulatus promoted different changes in fractionated sheep plasma proteins, duringa scorpion antivenom serum production.Materials, Methods & Results: Twelve sheep, healthy, mean weight of 30 kg, were distributed into 3 groups (n = 4): G1(control), G2 (crude venom) and G3 (detoxified venom). The adopted immunization protocol (first cycle) had 6 doses, 3using Freund’s adjuvant, with a 21-day interval between each one (day 0, 22 and 43), and 3 doses with no adjuvant (booster)and 0.2 mg of antigen (reinforcement), spaced 3 days between each other (day 50, 53 and 56). Group control (G1) received6 immunizations with phosphate buffered saline (PBS) associated with Freund’s adjuvant (1:1), while the other 2 groupsreceived 0.5 mg of venom (G2) and detoxified venom (G3), respectively, diluted in PBS, associated with the Freund adjuvant. The boosters were 1/3 of the initial dose, diluted only PBS. At baseline (T0) and at 24 and 48 h after immunization,all animals underwent clinical examinations. Blood samples were collected at day 0, 22, 43, 53 and 56 for proteinogramanalysis. Total protein, albumin and globulins fractions were measured. Plasma albumin concentration at T0 ranged from3.41-4.86 g/dL, with a mean value of 4.12 g/dL. There was no statistical difference between...

ß-micrustoxin (Mlx-9), a PLA2 from Micrurus lemniscatus snake venom: biochemical characterization and anti-proliferative effect mediated by p53

Background: Endogenous phospholipases A2 (PLA2 ) play a fundamental role in inflammation, neurodegenerative diseases, apoptosis and cellular senescence. Neurotoxins with PLA2 activity are found in snake venoms from the Elapidae and Viperidae families. The mechanism of action of these neurotoxins have been studied using hippocampal and cerebellar neuronal cultures showing [Ca2+]i increase, mitochondrial depolarization and cell death. Astrocytes are rarely used as a model, despite being modulators at the synapses and responsible for homeostasis and defense in the central nervous system. Preserving the cell division ability, they can be utilized to study the cell proliferation process. In the present work cultured astrocytes and glioblastoma cells were employed to characterize the action of ß-micrustoxin (previously named Mlx-9), a PLA2 isolated from Micrurus lemniscatus snake venom. The ß-micrustoxin structure was determined and the cell proliferation, cell cycle phases and the regulatory proteins p53, p21 and p27 were investigated. Methods: ß-micrustoxin was characterized biochemically by a proteomic approach. Astrocytes were obtained by dissociation of pineal glands from Wistar rats; glioblastoma tumor cells were purchased from ATCC and Sigma and cultured in DMEM médium. Cell viability was evaluated by MTT assay; cell proliferation and cell cycle phases were analyzed by flow cytometry; p53, p21 and p27 proteins were studied by western blotting and immunocytochemistry. Results: Proteomic analysis revealed fragments on ß-micrustoxin that aligned with a PLA2 from Micrurus lemniscatus lemniscatus previously identified as transcript ID DN112835_C3_g9_i1/m.9019. ß-micrustoxin impaired the viability of astrocytes and glioblastoma tumor cells. There was a reduction in cell proliferation, an increase in G2/M phase and activation of p53, p21 and p27 proteins in astrocytes. Conclusion: These findings indicate that ß-micrustoxin from Micrurus lemniscatus venom could inhibit cell proliferation through p53, p21 and p27 activation thus imposing cell cycle arrest at the checkpoint G2/M.(AU)

Surto de botulismo tipo C em aves de subsistência em Santa Catarina, Brasil / Outbreak of type C botulism in backyard poultry in Santa Catarina, Brazil

Background: Botulism is a disease caused by the ingestion of neurotoxin produced by Clostridium botulinum, characterizedby flaccid paralysis, which can lead to high mortality. They have seven types of neurotoxins (A, B, C, D, E, F, and G) and,in birds, most cases are attributed to type C. They are considered sources of botulinum toxins where the decomposition oforganic matter occurs, like stagnant water and rotting food. The main feature of the disease in birds is ascending symmetricflaccid paralysis. The present study aims to describe an outbreak of type C botulism in backyard poultry in the state ofSanta Catarina, Southern Brazil.Case: A visit was made to the property with 160 backyard poultry with a history of high mortality in the municipality ofAgrolândia, Santa Catarina. Clinical signs were characterized by paralysis of the pelvic limbs, neck and pendular wings,which progressed to death within 48 h. There was a mortality rate of 37.5% (60/160) between March and May 2019. Thesebirds were kept in an overcrowded environment, with different species (chickens, ducks, teals, and turkeys) fed irregularly.The water supplied was provided from kitchen exhaust, accumulating in puddles on the floor that contained organic matterresidues such as animal feces, food waste and bone fragments. The disposal of the carcasses of birds that died was in thesame enclosure, buried superficially, facilitating the access of other birds to dig them up and consume them. Necropsywas performed on 2 chickens and one duck, no macroscopic or histopathological lesions were observed. Blood, liver, andgastrointestinal content samples were sent for research and identification of botulinum toxin through the serum neutralization test in mice. The presence of type C botulinum toxin was confirmed in the liver chicken of one sampled animals.Discussion: The identification of type C botulism toxin enabled the characterization of the outbreak, which is...

Surto de botulismo tipo C em aves de subsistência em Santa Catarina, Brasil / Outbreak of type C botulism in backyard poultry in Santa Catarina, Brazil

Background: Botulism is a disease caused by the ingestion of neurotoxin produced by Clostridium botulinum, characterizedby flaccid paralysis, which can lead to high mortality. They have seven types of neurotoxins (A, B, C, D, E, F, and G) and,in birds, most cases are attributed to type C. They are considered sources of botulinum toxins where the decomposition oforganic matter occurs, like stagnant water and rotting food. The main feature of the disease in birds is ascending symmetricflaccid paralysis. The present study aims to describe an outbreak of type C botulism in backyard poultry in the state ofSanta Catarina, Southern Brazil.Case: A visit was made to the property with 160 backyard poultry with a history of high mortality in the municipality ofAgrolândia, Santa Catarina. Clinical signs were characterized by paralysis of the pelvic limbs, neck and pendular wings,which progressed to death within 48 h. There was a mortality rate of 37.5% (60/160) between March and May 2019. Thesebirds were kept in an overcrowded environment, with different species (chickens, ducks, teals, and turkeys) fed irregularly.The water supplied was provided from kitchen exhaust, accumulating in puddles on the floor that contained organic matterresidues such as animal feces, food waste and bone fragments. The disposal of the carcasses of birds that died was in thesame enclosure, buried superficially, facilitating the access of other birds to dig them up and consume them. Necropsywas performed on 2 chickens and one duck, no macroscopic or histopathological lesions were observed. Blood, liver, andgastrointestinal content samples were sent for research and identification of botulinum toxin through the serum neutralization test in mice. The presence of type C botulinum toxin was confirmed in the liver chicken of one sampled animals.Discussion: The identification of type C botulism toxin enabled the characterization of the outbreak, which is...(AU)

Characterization of the first two toxins isolated from the venom of the ancient scorpion Tityus (Archaeotityus) mattogrossensis (Borelli, 1901)

Background: Almost all Tityus characterized toxins are from subgenera Atreus and Tityus, there are only a few data about toxins produced by Archaeotityus, an ancient group in Tityus genus. Methods: Tityus (Archaeotityus) mattogrossensis crude venom was fractionated by high performance liquid chromatography, the major fractions were tested in a frog sciatic nerve single sucrose-gap technique. Two fractions (Tm1 and Tm2) were isolated, partially sequenced by MALDI-TOF/MS and electrophysiological assayed on HEK293 Nav 1.3, HEK293 Nav 1.6, DUM and DRG cells. Results: The sucrose-gap technique showed neurotoxicity in four fractions. One fraction caused a delay of action potential repolarization and other three caused a reduction in amplitude. An electrophysiological assay showed that Tm1 is active on HEK293 Nav 1.3, HEK293 Nav 1.6, DUM and DRG cells, and Tm2 on HEK293 Nav 1.3 and DRG cells, but not in HEK293 Nav 1.6. In addition, Tm1 and Tm2 did promote a shift to more negative potentials strongly suggesting that both are α-NaScTx. Conclusion: Although Tityus (Archaeotityus) mattogrossensis is considered an ancient group in Tityus genus, the primary structure of Tm1 and Tm2 is more related to Tityus subgenus. The patch clamp electrophysiological tests suggest that Tm1 and Tm2 are NaScTx, and also promoted no shift to more negative potentials, strongly suggesting that both are α-NaScTx. This paper aimed to explore and characterize for the first time toxins from the ancient scorpion Tityus (Archaeotityus) mattogrossensis.(AU)

MS/MS analysis of four scorpion venoms from Colombia: a descriptive approach

Background: Scorpions are widely known for the neurotoxic effects of their venoms, which contain peptides affecting ionic channels. Although Colombia is recognized for its scorpion diversity, only a few studies are available describing the venom content. Methods: In this descriptive study, we analyzed the MS/MS sequence, electrophoretic and chromatographic profile linked to a bioinformatics analysis of the scorpions Chactas reticulatus (Chactidae), Opisthacanthus elatus (Hormuridae), Centruroides edwardsii (Buthidae) and Tityus asthenes (Buthidae) from Colombia. Results: Each scorpion showed a specific electrophoretic and chromatographic profile. The electrophoretic profiles indicate the presence of high molecular mass compounds in all venoms, with a predominance of low molecular mass compounds in the Buthidae species. Chromatographic profiles showed a similar pattern as the electrophoretic profiles. From the MS/MS analysis of the chromatographic collected fractions, we obtained internal peptide sequences corresponding to proteins reported in scorpions from the respective family of the analyzed samples. Some of these proteins correspond to neurotoxins affecting ionic channels, antimicrobial peptides and metalloproteinase-like fragments. In the venom of Tityus asthenes, the MSn analysis allowed the detection of two toxins affecting sodium channels covering 50% and 84% of the sequence respectively, showing 100% sequence similarity. Two sequences from Tityus asthenes showed sequence similarity with a phospholipase from Opisthacanthus cayaporum indicating the presence of this type of toxin in this species for the first time. One sequence matching a hypothetical secreted protein from Hottentotta judaicus was found in three of the studied venoms. We found that this protein is common in the Buthidae family whereas it has been reported in other families - such as Scorpionidae - and may be part of the evolutionary puzzle of venoms in these arachnids. Conclusion: Buthidae venoms from Colombia can be considered an important source of peptides similar to toxins affecting ionic channels. An interesting predicted antimicrobial peptide was detected in three of the analyzed venoms.(AU)

MS/MS analysis of four scorpion venoms from Colombia: a descriptive approach

Background: Scorpions are widely known for the neurotoxic effects of their venoms, which contain peptides affecting ionic channels. Although Colombia is recognized for its scorpion diversity, only a few studies are available describing the venom content. Methods: In this descriptive study, we analyzed the MS/MS sequence, electrophoretic and chromatographic profile linked to a bioinformatics analysis of the scorpions Chactas reticulatus (Chactidae), Opisthacanthus elatus (Hormuridae), Centruroides edwardsii (Buthidae) and Tityus asthenes (Buthidae) from Colombia. Results: Each scorpion showed a specific electrophoretic and chromatographic profile. The electrophoretic profiles indicate the presence of high molecular mass compounds in all venoms, with a predominance of low molecular mass compounds in the Buthidae species. Chromatographic profiles showed a similar pattern as the electrophoretic profiles. From the MS/MS analysis of the chromatographic collected fractions, we obtained internal peptide sequences corresponding to proteins reported in scorpions from the respective family of the analyzed samples. Some of these proteins correspond to neurotoxins affecting ionic channels, antimicrobial peptides and metalloproteinase-like fragments. In the venom of Tityus asthenes, the MSn analysis allowed the detection of two toxins affecting sodium channels covering 50% and 84% of the sequence respectively, showing 100% sequence similarity. Two sequences from Tityus asthenes showed sequence similarity with a phospholipase from Opisthacanthus cayaporum indicating the presence of this type of toxin in this species for the first time. One sequence matching a hypothetical secreted protein from Hottentotta judaicus was found in three of the studied venoms. We found that this protein is common in the Buthidae family whereas it has been reported in other families - such as Scorpionidae - and may be part of the evolutionary puzzle of venoms in these arachnids. Conclusion: Buthidae venoms from Colombia can be considered an important source of peptides similar to toxins affecting ionic channels. An interesting predicted antimicrobial peptide was detected in three of the analyzed venoms.(AU)

Exploring the five-paced viper (Deinagkistrodon acutus) venom proteome by integrating a combinatorial peptide ligand library approach with shotgun LC-MS/MS

Snake venoms are complex mixtures of toxic proteins or peptides encoded by various gene families that function synergistically to incapacitate prey. In the present study, in order to unravel the proteomic repertoire of Deinagkistrodon acutus venom, some trace abundance components were analyzed. Methods Shotgun proteomic approach combined with shotgun nano-LC-ESI-MS/MS were employed to characterize the medically important D. acutus venom, after collected samples were enriched with the combinatorial peptide ligand library (CPLL). Results This avenue helped us find some trace components, undetected before, in D. acutus venom. The results indicated that D. acutus venom comprised 84 distinct proteins from 10 toxin families and 12 other proteins. These results are more than twice the number of venom components obtained from previous studies, which were only 29 distinct proteins obtained through RP-HPLC for the venom of the same species. The present results indicated that in D. acutus venom, the most abundant components (66.9%) included metalloproteinases, serine proteinases, and C-type lectin proteins; the medium abundant components (13%) comprised phospholipases A2 (PLA2) and 5'-nucleotidases and nucleases; whereas least abundant components (6%) were aminopeptidases, L-amino acid oxidases (LAAO), neurotoxins and disintegrins; and the trace components. The last were undetected before the use of conventional shotgun proteomics combined with shotgun nano-LC-ESI-MS/MS, such as cysteine-rich secretory proteins Da-CRPa, phospholipases B-like 1, phospholipases B (PLB), nerve growth factors (NGF), glutaminyl-peptide cyclortransferases (QC), and vascular non-inflammatory molecules 2 (VNN2). Conclusion These findings demonstrated that the CPLL enrichment method worked well in finding the trace toxin proteins in D. acutus venom, in contrast with the previous venomic characterization of D. acutus by conventional LC-MS/MS. In conclusion, this approach combined with the CPLL enrichment was effective for allowing us to explore the hidden D. acutus venomic profile and extended the list of potential venom toxins.(AU)

Exploring the five-paced viper (Deinagkistrodon acutus) venom proteome by integrating a combinatorial peptide ligand library approach with shotgun LC-MS/MS

Snake venoms are complex mixtures of toxic proteins or peptides encoded by various gene families that function synergistically to incapacitate prey. In the present study, in order to unravel the proteomic repertoire of Deinagkistrodon acutus venom, some trace abundance components were analyzed. Methods Shotgun proteomic approach combined with shotgun nano-LC-ESI-MS/MS were employed to characterize the medically important D. acutus venom, after collected samples were enriched with the combinatorial peptide ligand library (CPLL). Results This avenue helped us find some trace components, undetected before, in D. acutus venom. The results indicated that D. acutus venom comprised 84 distinct proteins from 10 toxin families and 12 other proteins. These results are more than twice the number of venom components obtained from previous studies, which were only 29 distinct proteins obtained through RP-HPLC for the venom of the same species. The present results indicated that in D. acutus venom, the most abundant components (66.9%) included metalloproteinases, serine proteinases, and C-type lectin proteins; the medium abundant components (13%) comprised phospholipases A2 (PLA2) and 5'-nucleotidases and nucleases; whereas least abundant components (6%) were aminopeptidases, L-amino acid oxidases (LAAO), neurotoxins and disintegrins; and the trace components. The last were undetected before the use of conventional shotgun proteomics combined with shotgun nano-LC-ESI-MS/MS, such as cysteine-rich secretory proteins Da-CRPa, phospholipases B-like 1, phospholipases B (PLB), nerve growth factors (NGF), glutaminyl-peptide cyclortransferases (QC), and vascular non-inflammatory molecules 2 (VNN2). Conclusion These findings demonstrated that the CPLL enrichment method worked well in finding the trace toxin proteins in D. acutus venom, in contrast with the previous venomic characterization of D. acutus by conventional LC-MS/MS. In conclusion, this approach combined with the CPLL enrichment was effective for allowing us to explore the hidden D. acutus venomic profile and extended the list of potential venom toxins.(AU)

Escherichia coli vacuolating factor, involved in avian cellulitis, induces actin contraction and binds to cytoskeleton proteins in fibroblasts

Avian pathogenic Escherichia coli (APEC) isolated from avian cellulitis lesions produces a toxin, named Escherichia coli vacuolating factor (ECVF), that causes cell vacuolization and induces inflammatory response in broiler chicken. Methods We investigated the intracellular activities of ECVF in avian fibroblasts using fluorescence staining, electron microscopy, MTT and LDH measurements. As ECVF act specifically in avian cells, we performed blotting assay followed by mass spectrometry to better understand its initial intracellular protein recognition. Results ECVF induced actin contraction, mitochondrial damage and membrane permeability alterations. Ultrastructural analysis showed intracellular alterations, as nuclear lobulation and the presence of degraded structures inside the vacuoles. Moreover, ECVF induced cell death in fibroblasts. ECVF-biotin associates to at least two proteins only in avian cell lysates: alpha-actinin 4 and vinculin, both involved in cytoskeleton structure. Conclusion These findings demonstrated that ECVF plays an important role in avian cellulitis, markedly in initial steps of infection. Taken together, the results place this toxin as a target for drug and/or vaccine development, instead of the use of large amounts antibiotics.(AU)

Escherichia coli vacuolating factor, involved in avian cellulitis, induces actin contraction and binds to cytoskeleton proteins in fibroblasts

Avian pathogenic Escherichia coli (APEC) isolated from avian cellulitis lesions produces a toxin, named Escherichia coli vacuolating factor (ECVF), that causes cell vacuolization and induces inflammatory response in broiler chicken. Methods We investigated the intracellular activities of ECVF in avian fibroblasts using fluorescence staining, electron microscopy, MTT and LDH measurements. As ECVF act specifically in avian cells, we performed blotting assay followed by mass spectrometry to better understand its initial intracellular protein recognition. Results ECVF induced actin contraction, mitochondrial damage and membrane permeability alterations. Ultrastructural analysis showed intracellular alterations, as nuclear lobulation and the presence of degraded structures inside the vacuoles. Moreover, ECVF induced cell death in fibroblasts. ECVF-biotin associates to at least two proteins only in avian cell lysates: alpha-actinin 4 and vinculin, both involved in cytoskeleton structure. Conclusion These findings demonstrated that ECVF plays an important role in avian cellulitis, markedly in initial steps of infection. Taken together, the results place this toxin as a target for drug and/or vaccine development, instead of the use of large amounts antibiotics.(AU)

Analgesic effects of Phα1ß toxin: a review of mechanisms of action involving pain pathways

Phα1ß is a neurotoxin purified from spider venom that acts as a high-voltage-activated (HVA) calcium channel blocker. This spider peptide has shown a high selectivity for N-type HVA calcium channels (NVACC) and an analgesic effect in several animal models of pain. Its activity was associated with a reduction in calcium transients, glutamate release, and reactive oxygen species production from the spinal cord tissue and dorsal ganglia root (DRG) in rats and mice. It has been reported that intrathecal (i.t.) administration of Phα1ß to treat chronic pain reverted opioid tolerance with a safer profile than ω-conotoxin MVIIA, a highly selective NVACC blocker. Following a recent development of recombinant Phα1ß (CTK 01512-2), a new molecular target, TRPA1, the structural arrangement of disulphide bridges, and an effect on glial plasticity have been identified. CTK 01512-2 reproduced the antinociceptive effects of the native toxin not only after the intrathecal but also after the intravenous administration. Herein, we review the Phα1ß antinociceptive activity in the most relevant pain models and its mechanisms of action, highlighting the impact of CTK 01512-2 synthesis and its potential for multimodal analgesia.

Analgesic effects of Phalfa1 beta toxin: a review of mechanisms of action involving pain pathways

Phα1ß is a neurotoxin purified from spider venom that acts as a high-voltage-activated (HVA) calcium channel blocker. This spider peptide has shown a high selectivity for N-type HVA calcium channels (NVACC) and an analgesic effect in several animal models of pain. Its activity was associated with a reduction in calcium transients, glutamate release, and reactive oxygen species production from the spinal cord tissue and dorsal ganglia root (DRG) in rats and mice. It has been reported that intrathecal (i.t.) administration of Phα1ß to treat chronic pain reverted opioid tolerance with a safer profile than ω-conotoxin MVIIA, a highly selective NVACC blocker. Following a recent development of recombinant Phα1ß (CTK 01512-2), a new molecular target, TRPA1, the structural arrangement of disulphide bridges, and an effect on glial plasticity have been identified. CTK 01512-2 reproduced the antinociceptive effects of the native toxin not only after the intrathecal but also after the intravenous administration. Herein, we review the Phα1ß antinociceptive activity in the most relevant pain models and its mechanisms of action, highlighting the impact of CTK 01512-2 synthesis and its potential for multimodal analgesia.

Why does the number of dangerous species of scorpions increase? The particular case of the genus Leiurus Ehrenberg (Buthidae) in Africa

The aim of this contribution is to bring some precise information on the reasons why the number of noxious scorpion species is constantly growing. This fact is directly associated with the zoological research on the domains generally defined as systematics and taxonomy. The classification of any zoological group is in most cases a source of problem for most biologists not directly involved with this almost confidential aspect of the zoological research. Much information has been gathered and published over two centuries on the classification but it is remains poorly accessible and too technical for non-experts. The exposed example could be taken from several groups of scorpions possessing infamous species, but the choice went to the genus Leiurus Ehrenberg, 1828 distributed from North Africa to the Middle East. Maybe this contribution will help to explain why so numerous cases of species misidentification are regularly present in the general literature devoted to scorpion venoms and incidents.(AU)

The coevolution between telson morphology and venom glands in scorpions (Arachnida)

As in previous contributions to the JVATiTD, the aim of this note is to bring some general information on a particular aspect of the scorpion biology. An attempt is made to explain the possible coevolution of telson morphology and venom glands, which took place during several hundred million years and in particular since scorpions migrated from aquatic to terrestrial environments. Three components can be directly associated with predation and defensive behaviours: (1) morphology of the chelae and structure of the chelae fingers granulations; (2) morphology of the metasoma and in particular of the telson; (3) evolution of tegumentary glands in the telson toward different types of venom glands. Since a number of recent contributions already treated some of these aspects, I will limit my comments to the possible evolution of the telson in relation to the evolution of venom glands. As in previous contributions, the content of this article is basically addressed to non-specialists on scorpions whose research embraces scorpions in several fields such as venom toxins and public health.(AU)

Pediatric scorpionism in northern Amazonia: a 16-year study on epidemiological, environmental and clinical aspects

The Amazon basin is one of the seven major geographical areas where scorpionism is recorded. In French Guiana, 90 stings per 100,000 inhabitants are registered per year. As the severity of cases is higher in children, descriptive studies are needed to have a better understanding of this pathology. The aim of the present study is to describe pediatric scorpionism in French Guiana. Methods: We conducted a monocentric descriptive retrospective study on scorpion stings in all pediatric patients admitted to Cayenne General Hospital from January 1, 2002 to December 31, 2018. Results: In this survey, 132 patients were included. Of them, 63% were male. Patients with general signs of envenomation were younger and lighter (p = 0.04). The picture was "one sting" (95.3%) by a "big" (47.6%), "black" (60%) and "small pincer" (58%) scorpion on the extremity of the body (84%). Stings occurred mainly during the day, while patients changed clothes. There was no envenomation during night. The monthly evaluation highlights that the number of stings and percentage of general signs of envenomation were closely connected to a composite variable including the variation of the level of rivers (p = 0.005). Cardiac symptoms were recorded in 82% of cases with general signs of envenomation. The presence of pulmonary; ear, nose, and throat (ENT); or gastrointestinal symptoms are related to major envenomation (p = 0.001, p = 0.01, and p = 0.02 respectively). Leukocytosis and glycemia increased according to the envenomation grade whereas serum potassium and alkaline reserve decreased. Forty-six patients needed hospitalization and seven of them required intensive care. No patient died nor presented sequelae at discharge from the hospital. Conclusion: Pediatric scorpionism in French Guiana is closely associated with child activities and climatic conditions. Severe envenomation presented most of the time with cardiac, pulmonary, and gastrointestinal symptoms.(AU)

Analgesic and side effects of intravenous recombinant Ph alfa 1 beta

Background: Intrathecal injection of voltage-sensitive calcium channel blocker peptide toxins exerts analgesic effect in several animal models of pain. Upon intrathecal administration, recombinant Phα1β exerts the same analgesic effects as the those of the native toxin. However, from a clinical perspective, the intrathecal administration limits the use of anesthetic drugs in patients. Therefore, this study aimed to investigate the possible antinociceptive effect of intravenous recombinant Phα1β in rat models of neuropathic pain, as well as its side effects on motor, cardiac (heart rate and blood pressure), and biochemical parameters. Methods: Male Wistar rats and male Balb-C mice were used in this study. Giotto Biotech® synthesized the recombinant version of Phα1β using Escherichia coli expression. In rats, neuropathic pain was induced by chronic constriction of the sciatic nerve and paclitaxel-induced acute and chronic pain. Mechanical sensitivity was evaluated using von Frey filaments. A radiotelemeter transmitter (TA11PA-C10; Data Sciences, St. Paul, MN, USA) was placed on the left carotid of mice for investigation of cardiovascular side effects. Locomotor activity data were evaluated using the open-field paradigm, and serum CKMB, TGO, TGP, LDH, lactate, creatinine, and urea levels were examined. Results: Intravenous administration of recombinant Phα1β toxin induced analgesia for up to 4 h, with ED50 of 0.02 (0.01-0.03) mg/kg, and reached the maximal effect (Emax = 100% antinociception) at a dose of 0.2 mg/kg. No significant changes were observed in any of the evaluated motor, cardiac or biochemical parameters. Conclusion: Our data suggest that intravenous administration of recombinant Phα1β may be feasible for drug-induced analgesia, without causing any severe side effects.(AU)