Proteomic analysis and Lethality of the venom of Aegaeobuthus nigrocinctus, a scorpion of medical significance in the Middle East.

The scorpion Aegaeobuthus nigrocinctus inhabits areas in Turkey and the Levant region of the Middle East where severe/lethal envenomings have been reported. Previous research indicated its extreme venom lethality to vertebrates and distinct envenomation syndrome. We report on the composition of A. nigrocinctus venom from Lebanese specimens using nESI-MS/MS, MALDI-TOF MS, SDS-PAGE and RP-HPLC. Venom lethality in mice was also assessed (LD50 = 1.05 (0.19-1.91) mg/kg, i.p), confirming A. nigrocinctus venom toxicity from Levantine populations. Forty-seven peaks were resolved using RP-HPLC, 25 of which eluted between 20-40% acetonitrile. In reducing SDS-PAGE, most predominant components were <10 kDa, with minor components at higher molecular masses of 19.6, 26.1, 46.3 and 57.7 kDa. MALDI-TOF venom fingerprinting detected 20 components within the 1,000-12,000 m/z range. Whole venom 'shotgun' bottom-up nLC-MS/MS approach, combined with in-gel tryptic digestion of SDS-PAGE bands, identified at least 67 different components belonging to 15 venom families, with ion channel-active components (K+ toxins (23); Na+ toxins (20); Cl- toxins (2)) being predominant. The sequence of a peptide (named α-KTx9.13) ortholog to Leiurus hebraeus putative α-KTx9.3 toxin was fully determined, which exhibited 81-96% identity to other members of the α-KTx9 subfamily targeting Kv1.x and Ca2+-activated K+ channels. Chlorotoxin-like peptides were also identified. Our study underscores the medical significance of A. nigrocinctus in the region and reveals the potential value of its venom components as lead templates for biomedical applications. Future work should address whether available antivenoms in the Middle East are effective against A. nigrocinctus envenoming in the Levant area.

Divergence in toxin antigenicity and venom enzymes in Tityus melici, a medically important scorpion, despite transcriptomic and phylogenetic affinities with problematic Brazilian species.

The Brazilian scorpion Tityus melici, native to Minas Gerais and Bahia, is morphologically related to Tityus serrulatus, the most medically significant species in Brazil. Despite inhabiting scorpion-envenomation endemic regions, T. melici venom remains unexplored. This work evaluates T. melici venom composition and function using transcriptomics, enzymatic activities, and in vivo and in vitro immunological analyses. Next-Generation Sequencing unveiled 86 components putatively involved in venom toxicity: 39 toxins, 28 metalloproteases, seven disulfide isomerases, six hyaluronidases, three phospholipases and three amidating enzymes. T. serrulatus showed the highest number of toxin matches with 80-100 % sequence similarity. T. melici is of medical importance as it has a venom LD50 of 0.85 mg/kg in mice. We demonstrated venom phospholipase A2 activity, and elevated hyaluronidase and metalloprotease activities compared to T. serrulatus, paralleling our transcriptomic findings. Comparison of transcriptional levels for T. serrulatus and T. melici venom metalloenzymes suggests species-specific expression patterns in Tityus. Despite close phylogenetic association with T. serrulatus inferred from COI sequences and toxin similarities, partial neutralization of T. melici venom toxicity was achieved when using the anti-T. serrulatus antivenom, implying antigenic divergence among their toxins. We suggest that the Brazilian therapeutic scorpion antivenom could be improved to effectively neutralize T. melici venom.

Venomics of Leiurus abdullahbayrami, the most lethal scorpion in the Levant region of the Middle East.

The scorpion Leiurus abdullahbayrami has been associated with severe/lethal envenomings throughout the Levant region of the Middle East, encompassing Turkey, Syria, and Lebanon, and only scarce information is available on its venom composition, activity, and antigenicity. We report on the composition of L. abdullahbayrami venom collected from Lebanese specimens using nESI-MS/MS, MALDI-TOF MS, SDS-PAGE and RP-HPLC. Venom lethality, through LD50 determination in mice (intraperitoneal), was also assessed (0.75 (0.16-1.09) mg/kg), confirming L.abdullahbayrami venom vertebrate toxicity. Fifty-four peaks were detected using RP-HPLC, half of which eluted in the gradient region between 20 and 40% acetonitrile. In reducing SDS-PAGE, most predominant components were <10 kDa, with minor components at higher molecular masses of 24.4, 43.1, and 48.9 kDa. Venom mass fingerprint by MALDI-TOF detected 21 components within the 1000-12,000 m/z range. Whole venom 'shotgun' bottom-up nLC-MS/MS approach, combined with in-gel tryptic digestion of SDS-PAGE bands, identified at least 113 different components belonging to 15 venom families and uncharacterized proteins, with ion channel-active components (K+ channel toxins (28); Na+ channel toxins (42); Cl- channel toxins (4); Ca+2 toxins (2)) being predominant. A single match for a L. adbullahbayrami NaTx was found in the UniProt database with other congeneric species, toxin h3.1 from Leiurus hebraeus, suggesting this might be an indication of venom divergence within Leiurus, eventhough this warrants further investigation involving venom proteomics and transcriptomics of relevant species. Considering such potential interspecific venom variation, future work should address whether preparation of a specific anti-L. abdullahbayrami antivenom is justified.

Scorpion Venom as a Source of Antimicrobial Peptides: Overview of Biomolecule Separation, Analysis and Characterization Methods.

Scorpion venoms have long captivated scientific researchers, primarily due to the potency and specificity of the mechanism of action of their derived components. Among other molecules, these venoms contain highly active compounds, including antimicrobial peptides (AMPs) and ion channel-specific components that selectively target biological receptors with remarkable affinity. Some of these receptors have emerged as prime therapeutic targets for addressing various human pathologies, including cancer and infectious diseases, and have served as models for designing novel drugs. Consequently, extensive biochemical and proteomic investigations have focused on characterizing scorpion venoms. This review provides a comprehensive overview of the key methodologies used in the extraction, purification, analysis, and characterization of AMPs and other bioactive molecules present in scorpion venoms. Noteworthy techniques such as gel electrophoresis, reverse-phase high-performance liquid chromatography, size exclusion chromatography, and "omics" approaches are explored, along with various combinations of methods that enable bioassay-guided venom fractionation. Furthermore, this review presents four adapted proteomic workflows that lead to the comprehensive dissection of the scorpion venom proteome, with an emphasis on AMPs. These workflows differ based on whether the venom is pre-fractionated using separation techniques or is proteolytically digested directly before further proteomic analyses. Since the composition and functionality of scorpion venoms are species-specific, the selection and sequence of the techniques for venom analyses, including these workflows, should be tailored to the specific parameters of the study.

Luring Triatomines (Hemiptera: Reduviidae) into a Trap: Aliphatic and Aromatic Aldehydes as Attractants of Triatoma infestans.

To assess the attracting capacity of aliphatic and aromatic aldehydes to Triatoma infestans, the Chagas disease vector, laboratory tests were conducted using individual compounds and mixtures to evaluate their potential use in baited traps for intradomicile population dynamics analysis. Commercial samples of hexanal, nonanal, and benzaldehyde were used at 95% purity. The experiments were performed at 25°C and 65% relative humidity using two procedures: a glass arena with filter papers impregnated with 1, 5, and 10 µL of the tested compounds and a double-choice olfactometer. Attraction was scored positively if the insect remained more than 30 seconds on one of the surfaces. The results of the study showed that hexanal was attractive to females at higher concentrations (5-10 µL; P < 0.0001), and IV instar nymphs were only attracted at the highest concentration (10 µL; P < 0.01). Nonanal was attractive to IV instar nymphs at 1 and 5 µL (P < 0.0001), whereas males and females were more attracted at 1 µL (P < 0.01 and P < 0.05, respectively). Benzaldehyde showed significant differences with respect to controls, attracting females at low concentrations (1 µL; P < 0.0001) and IV instar nymphs at 5 and 10 µL (P < 0.0001 and P < 0.001, respectively). In the olfactometer, the 60:40 hexanal/nonanal mixture was the most effective. In conclusion, this study demonstrated that the aliphatic and aromatic aldehydes studied here, both individually and in mixtures, could be used as effective attractants for T. infestans in intradomicile-baited traps. These results suggest that mixtures of these compounds could be implemented in field trials for Chagas disease surveillance.

Case Report: Fatal Scorpion Envenomation in a Shuar Child by Tityus cisandinus from Amazonian Ecuador: A Call for Specific Antivenom Availability in the Amazon Basin.

Envenomation by scorpions belonging to the genus Tityus can be life threatening in the Americas, particularly in the Amazon Basin. We report a 4-month-old Ecuadorean boy of Shuar origin stung by a scorpion identified as Tityus cisandinus in the Amazonian province of Morona Santiago, presenting with pulmonary edema and systemic inflammation. We administered immunotherapy using the scorpion antivenom available in Ecuador, of Mexican origin (anti-Centruroides). Catecholamine discharge-related events such as hyperglycemia and thrombocytosis were resolved after treatment but leukocytosis did not, suggesting that factors associated with the sting-admission delay and specificity of antivenom played a role in the envenomation outcome. Cardiorespiratory arrest determined a fatal outcome, despite specific maneuvers. The case severity and the limited supply of nonspecific scorpion antivenoms in problematic areas of Amazonian Ecuador and elsewhere in northwestern Amazonia are discussed in regard to the need for specific therapeutic immunoglobulins in the area and in the Amazon Basin as a whole.

An overview of Tityus cisandinus scorpion venom: Transcriptome and mass fingerprinting reveal conserved toxin homologs across the Amazon region and novel lipolytic components.

Tityus cisandinus, a neglected medically important scorpion in Ecuadorian and Peruvian Amazonia, belongs to a complex of species related to the eastern Amazon endemic Tityus obscurus, spanning a distribution of ca. 4000 km. Despite high morbidity and mortality rates, no effective scorpion antivenom is currently available in the Amazon region. Knowledge of the structural/functional relationships between T. cisandinus venom components and those from related Amazonian species is crucial for designing region-specific therapeutic antivenoms. In this work, we carried out the first venom gland transcriptomic study of an Amazonian scorpion outside Brazil, T. cisandinus. We also fingerprinted its total venom through MALDI-TOF MS, which supported our transcriptomic findings. We identified and calculated the expression level of 94 components: 60 toxins, 25 metalloproteases, five disulfide isomerases, three amidating enzymes, one hyaluronidase, and also uncovered transcripts encoding novel lipolytic beta subunits produced by New World buthid scorpions. This study demonstrates the high similarity between T. cisandinus and T. obscurus venoms, reinforcing the existence of a neglected complex of genetically and toxinologically related Amazonian scorpions of medical importance. Finally, we demonstrated the low recognition of currently available therapeutic sera against T. cisandinus and T. obscurus venoms, and concluded that these should be improved to protect against envenomation by Amazonian Tityus spp.

Amazonian scorpions and scorpionism: integrating toxinological, clinical, and phylogenetic data to combat a human health crisis in the world's most diverse rainfores.

Venom from Amazonian scorpions of the genus Tityus contains components capable of eliciting a distinct clinical, mostly neurological, syndrome. This contrasts with the mainly autonomic manifestations produced after envenomation by congeneric southern and northern South American species. Herein, we summarize Pan-Amazonian scorpionism by synthesizing available toxinological, clinical, and molecular data gathered from all affected areas in Amazonia, including Brazil, Ecuador, Colombia, Peru, Venezuela, and French Guiana. We searched multiple databases, as well as our own records, for reports of scorpion envenomations in Amazonia by confirmed Tityus spp., and compared the clinical manifestations. To help uncover clinical and venom relationships among problematic species, we explored phylogenetic relationships with a rate-calibrated analysis of mitochondrial COI data from available species. The possible existence of diversity gradients for venom toxic and immunogenic components despite the predicted strong phylogenetic association among species is underscored by discussed clinical and toxinological findings. A multicentric effort, involving all nations affected by this neglected disease, is urgently needed to offer alternatives for treating and understanding this pathology, including the preparation of neutralizing antibodies with a broad range of efficacy.

Mortality caused by venomous animals in Venezuela (2000-2009): A new epidemiological pattern / Mortalidad causada por animales venenosos en Venezuela (2000-2009): nuevo patrón epidemiológico.

Introduction: Injuries by venomous animals frequently occur in impoverished communities with limited access to health services. They are considered neglected diseases that stand out as important causes of morbidity and mortality in various countries, including Venezuela. Objective: To assess mortalities resulting from contact with venomous animals in Venezuela from 2000 to 2009 (X20-X29 series). Materials and Methods: The data were obtained from the annual mortality records of the Venezuelan Ministry of Health. Results: From 2000-2009, 759 fatalities were recorded with the greatest number taking place in 2009. Snakebites (n=323; 42.6%) accounted for the largest percentage of envenomation-related deaths in that period, followed by hymenopteran stings (n=170; 22.4%), centipede bites (n=106; 14.0%), and scorpion stings (n=76; 10.0%). The median value of envenomation-related deaths per 100,000 inhabitants (period 2000-2009) was 0.285: 0.120 corresponded to snakebites, 0.065 to hymenopteran stings, 0.035 to centipede bites, and 0.025 to scorpion stings. Conclusions: Taking into account previous records of animal envenomations in Venezuela, we provided evidence for a shift in the pattern of mortality. Deaths due to centipede bites have increased, making it the third leading cause of envenomation-related mortality in Venezuela. Scorpionism, on the other hand, has declined to the fourth most common cause of fatal envenomations in the country.

Introducción. Los accidentes causados por animales venenosos ocurren con mucha frecuencia en comunidades pobres con acceso limitado a los servicios de salud. Se les consideran enfermedades desatendidas y son una de las causas importantes de morbimortalidad en varias naciones del mundo, incluida Venezuela. Objetivo. Evaluar la mortalidad por contacto traumático con animales venenosos (serie X20-X29) en Venezuela en el periodo de 2000 a 2009. Materiales y métodos. Los datos se obtuvieron de los anuarios de mortalidad del Ministerio de Salud. Resultados. Se registraron 759 decesos, la mayoría de ellos en el 2009. La primera causa fue la mordedura de serpientes (n=323; 42,6 %), seguida por la picadura de himenópteros (n=170; 22,4 %), la mordedura de centípedos (n=106; 14,0 %) y la picadura de escorpiones (n=76; 10,0 %). La mediana de la tasa de mortalidad general para el periodo fue de 0,285 fallecidos por 100.000 habitantes, en tanto que, por grupo específico, fue de 0,120 para ofidios, de 0,065 para himenópteros, de 0,035 para centípedos y de 0,025 para escorpiones. Conclusión. Al comparar estos datos con los antecedentes históricos, se evidenció la modificación del patrón de mortalidad en el país caracterizada por un aumento significativo de los decesos por centípedos, tercera causa de muerte, lo que reubica la picadura de escorpiones como la cuarta causa de mortalidad.

Mortalidad causada por animales venenosos en Venezuela (2000-2009): nuevo patrón epidemiológico / Mortality caused by venomous animals in Venezuela (2000-2009): A new epidemiological pattern

Resumen | Introducción. Los accidentes causados por animales venenosos ocurren con mucha frecuencia en comunidades pobres con acceso limitado a los servicios de salud. Se les consideran enfermedades desatendidas y son una de las causas importantes de morbimortalidad en varias naciones del mundo, incluida Venezuela. Objetivo. Evaluar la mortalidad por contacto traumático con animales venenosos (serie X20-X29) en Venezuela en el periodo de 2000 a 2009. Materiales y métodos. Los datos se obtuvieron de los anuarios de mortalidad del Ministerio de Salud. Resultados. Se registraron 759 decesos, la mayoría de ellos en el 2009. La primera causa fue la mordedura de serpientes (n=323; 42,6 %), seguida por la picadura de himenópteros (n=170; 22,4 %), la mordedura de centípedos (n=106; 14,0 %) y la picadura de escorpiones (n=76; 10,0 %). La mediana de la tasa de mortalidad general para el periodo fue de 0,285 fallecidos por 100.000 habitantes, en tanto que, por grupo específico, fue de 0,120 para ofidios, de 0,065 para himenópteros, de 0,035 para centípedos y de 0,025 para escorpiones. Conclusión. Al comparar estos datos con los antecedentes históricos, se evidenció la modificación del patrón de mortalidad en el país caracterizada por un aumento significativo de los decesos por centípedos, tercera causa de muerte, lo que reubica la picadura de escorpiones como la cuarta causa de mortalidad.

Abstract | Introduction: Injuries by venomous animals frequently occur in impoverished communities with limited access to health services. They are considered neglected diseases that stand out as important causes of morbidity and mortality in various countries, including Venezuela. Objective: To assess mortalities resulting from contact with venomous animals in Venezuela from 2000 to 2009 (X20-X29 series). Materials and Methods: The data were obtained from the annual mortality records of the Venezuelan Ministry of Health. Results: From 2000-2009, 759 fatalities were recorded with the greatest number taking place in 2009. Snakebites (n=323; 42.6%) accounted for the largest percentage of envenomation-related deaths in that period, followed by hymenopteran stings (n=170; 22.4%), centipede bites (n=106; 14.0%), and scorpion stings (n=76; 10.0%). The median value of envenomation-related deaths per 100,000 inhabitants (period 2000-2009) was 0.285: 0.120 corresponded to snakebites, 0.065 to hymenopteran stings, 0.035 to centipede bites, and 0.025 to scorpion stings. Conclusions: Taking into account previous records of animal envenomations in Venezuela, we provided evidence for a shift in the pattern of mortality. Deaths due to centipede bites have increased, making it the third leading cause of envenomation-related mortality in Venezuela. Scorpionism, on the other hand, has declined to the fourth most common cause of fatal envenomations in the country.

Novel components of Tityus serrulatus venom: A transcriptomic approach.

Several research groups have studied the components produced by the venom gland of the scorpion Tityus serrulatus, which has one of the most lethal venoms in the world. Various methodologies have been employed to clarify the complex mechanisms of action of these components, especially neurotoxins and enzymes. Transcriptomes and proteomes have provided important information for pharmacological, biochemical, and immunological research. Next-generation sequencing (NGS) has allowed the description of new transcripts and completion of partial sequence descriptions for peptides, especially those with low expression levels. In the present work, after NGS sequencing, we searched for new putative venom components. We present a total of nine new transcripts with neurotoxic potential (Ts33-41) and describe the sequences of one hyaluronidase (TsHyal_4); three enzymes involved in amidation (peptidyl-glycine alpha-amidating monooxygenase A, peptidyl-alpha-hydroxyglycine alpha-amidating lyase, and peptidylglycine alpha-hydroxylating monooxygenase), which increases the lethal potential of neurotoxins; and also the enzyme Ts_Chitinase1, which may be involved in the venom's digestive action. In addition, we determined the level of transcription of five groups: toxins, metalloproteases, hyaluronidases, chitinases and amidation enzymes, including new components found in this study. Toxins are the predominant group with an expression level of 91.945%, followed by metalloproteases with only 7.790% and other groups representing 0.265%.

Amazonian scorpions and scorpionism: integrating toxinological, clinical, and phylogenetic data to combat a human health crisis in the worlds most diverse rainfores

Abstract Venom from Amazonian scorpions of the genus Tityus contains components capable of eliciting a distinct clinical, mostly neurological, syndrome. This contrasts with the mainly autonomic manifestations produced after envenomation by congeneric southern and northern South American species. Herein, we summarize Pan-Amazonian scorpionism by synthesizing available toxinological, clinical, and molecular data gathered from all affected areas in Amazonia, including Brazil, Ecuador, Colombia, Peru, Venezuela, and French Guiana. We searched multiple databases, as well as our own records, for reports of scorpion envenomations in Amazonia by confirmed Tityus spp., and compared the clinical manifestations. To help uncover clinical and venom relationships among problematic species, we explored phylogenetic relationships with a rate-calibrated analysis of mitochondrial COI data from available species. The possible existence of diversity gradients for venom toxic and immunogenic components despite the predicted strong phylogenetic association among species is underscored by discussed clinical and toxinological findings. A multicentric effort, involving all nations affected by this neglected disease, is urgently needed to offer alternatives for treating and understanding this pathology, including the preparation of neutralizing antibodies with a broad range of efficacy.

Amazonian scorpions and scorpionism: integrating toxinological, clinical, and phylogenetic data to combat a human health crisis in the world's most diverse rainfores

Venom from Amazonian scorpions of the genus Tityus contains components capable of eliciting a distinct clinical, mostly neurological, syndrome. This contrasts with the mainly autonomic manifestations produced after envenomation by congeneric southern and northern South American species. Herein, we summarize Pan-Amazonian scorpionism by synthesizing available toxinological, clinical, and molecular data gathered from all affected areas in Amazonia, including Brazil, Ecuador, Colombia, Peru, Venezuela, and French Guiana. We searched multiple databases, as well as our own records, for reports of scorpion envenomations in Amazonia by confirmed Tityus spp., and compared the clinical manifestations. To help uncover clinical and venom relationships among problematic species, we explored phylogenetic relationships with a rate-calibrated analysis of mitochondrial COI data from available species. The possible existence of diversity gradients for venom toxic and immunogenic components despite the predicted strong phylogenetic association among species is underscored by discussed clinical and toxinological findings. A multicentric effort, involving all nations affected by this neglected disease, is urgently needed to offer alternatives for treating and understanding this pathology, including the preparation of neutralizing antibodies with a broad range of efficacy.(AU)

Amazonian scorpions and scorpionism: integrating toxinological, clinical, and phylogenetic data to combat a human health crisis in the world’s most diverse rainfores

Venom from Amazonian scorpions of the genus Tityus contains components capable of eliciting a distinct clinical, mostly neurological, syndrome. This contrasts with the mainly autonomic manifestations produced after envenomation by congeneric southern and northern South American species. Herein, we summarize Pan-Amazonian scorpionism by synthesizing available toxinological, clinical, and molecular data gathered from all affected areas in Amazonia, including Brazil, Ecuador, Colombia, Peru, Venezuela, and French Guiana. We searched multiple databases, as well as our own records, for reports of scorpion envenomations in Amazonia by confirmed Tityus spp., and compared the clinical manifestations. To help uncover clinical and venom relationships among problematic species, we explored phylogenetic relationships with a rate-calibrated analysis of mitochondrial COI data from available species. The possible existence of diversity gradients for venom toxic and immunogenic components despite the predicted strong phylogenetic association among species is underscored by discussed clinical and toxinological findings. A multicentric effort, involving all nations affected by this neglected disease, is urgently needed to offer alternatives for treating and understanding this pathology, including the preparation of neutralizing antibodies with a broad range of efficacy.

Genetic and toxinological divergence among populations of Tityus trivittatus Kraepelin, 1898 (Scorpiones: Buthidae) inhabiting Paraguay and Argentina.

Envenoming by scorpions in genus Tityus is a public health problem in Tropical America. One of the most medically significant species is Tityus trivittatus, which is known to occur from southwest Brazil to central-northern and eastern Argentina. In this work, we studied the lethality, composition, antigenicity, and enzymatic activity of venom from a T. trivittatus population found further north in urban areas of eastern Paraguay, where it has caused serious envenomation of children. Our results indicate that the population is of medical importance as it produces a potently toxic venom with an LD50 around 1.19 mg/kg. Venom neutralization in preliminary mouse bioassays was complete when using Brazilian anti-T. serrulatus antivenom but only partial when using Argentinean anti-T. trivittatus antivenom. Venom competitive solid-phase enzyme immunoassays and immunoblotting from Argentinean and Paraguayan T. trivittatus populations indicated that antigenic differences exist across the species range. SDS-PAGE showed variations in type and relative amounts of venom proteins between T. trivitattus samples from Argentina and Paraguay. MALDI-TOF mass spectrometry indicated that while some sodium channel toxins are shared, including ß-toxin Tt1g, others are population-specific. Proteolytic activity by zymography and peptide identification through nESI-MS/MS also point out that population-specific proteases may exist in T. trivitattus, which are postulated to be involved in the envenoming process. A time-calibrated molecular phylogeny of mitochondrial COI sequences revealed a significant (8.14%) genetic differentiation between the Argentinean and Paraguayan populations, which appeared to have diverged between the mid Miocene and early Pliocene. Altogether, toxinological and genetic evidence indicate that T. trivitattus populations from Paraguay and Argentina correspond to distinct, unique cryptic species, and suggest that further venom and taxonomic diversity exists in synanthropic southern South American Tityus than previously thought.

Venom diversity in the Neotropical scorpion genus Tityus: Implications for antivenom design emerging from molecular and immunochemical analyses across endemic areas of scorpionism.

Scorpions of the Neotropical genus Tityus are responsible for most severe envenomations in the Caribbean, South America, and Lower Central America (LCA). Although Tityus is taxonomically complex, contains high toxin polymorphism, and produces variable clinical manifestations, treatment is limited to antivenoms produced against species with restricted distributions. In this study, we explored the compositional and antigenic diversity of Tityus venoms to provide improved guidelines for the use of available antivenoms at a broader geographic scale. We used immunoblotting, competitive ELISA, and in vivo studies to compare reactivity against commercial antivenoms from Brazil, Venezuela, and Mexico, as well as MALDI-TOF mass spectrometry, cDNA sequencing, and phylogenetic analyses to assess venom sodium channel-active toxin (NaTx) content from medically important Tityus populations inhabiting Brazil, Colombia, Costa Rica, Ecuador, Panama, Trinidad and Tobago, and Venezuela. Additionally, we raised rabbit antibodies against Tityus venoms from LCA to test for cross-reactivity with congeneric species. The results suggest that Tityus spp. possess high venom antigenic diversity, underlying the existence of four toxinological regions in Tropical America, based on venom composition and immunochemical criteria: LCA/Colombia/Amazonia (Region I), Venezuela (Region II), southeast South America (Region III), and a fourth region encompassing species related to toxinologically divergent Tityus cerroazul. Importantly, our molecular and cross-reactivity results highlight the need for new antivenoms against species inhabiting Region I, where scorpions may produce venoms that are not significantly reactive against available antivenoms.

Scorpion envenoming in Morona Santiago, Amazonian Ecuador: Molecular phylogenetics confirms involvement of the Tityus obscurus group.

Scorpion envenoming by species in the genus Tityus is hereby reported from rural locations in the Amazonian province of Morona Santiago, southeastern Ecuador. Twenty envenoming cases (18 patients under 15 years of age) including one death (a 4-year-old male) were recorded at the Macas General Hospital, Morona Santiago, between January 2015 and December 2016 from the counties of Taisha (n=17), Huamboyo (n=1), Palora (n=1), and Logroño (n=1). An additional fatality from 2014 (a 3-year-old female from Nayantza, Taisha county) is also reported. Leukocytosis and low serum potassium levels were detected in most patients. We observed a significant negative correlation between leukocytosis and hypokalemia. Scorpions involved in three accidents from Macuma, Taisha County, were identified as genetically related to Tityus obscurus from the Brazilian Amazonian region based on comparison of mitochondrial DNA sequences encoding cytochrome oxidase subunit I. These cases, along with previously reported envenoming from northern Manabí, reinforce the notion that scorpionism is a health hazard for children in Ecuador and emphasizes the need to supply effective antivenoms against local species, which are not currently available. The genetic affinity of the Ecuadorian specimens with T. obscurus may underlay toxinological, clinical, and venom antigenic relationships among Amazonian scorpions that deserves further exploration for designing therapeutic strategies to treat scorpionism in the region.

Scorpionism in Ecuador: First report of severe and fatal envenoming cases from northern Manabí by Tityus asthenes Pocock.

The presence in rural areas of western Ecuador of scorpions in the genus Tityus capable of producing pediatric mortality is hereby evidenced. The medical significance of scorpions in Ecuador has been underestimated partly because of the clinically unimportant stings delivered by Centruroides margaritatus and Teuthraustes atramentarius, which have venom with low toxicity to vertebrates. Five intra-domiciliary cases of scorpion envenoming in victims aged between 1.9 and 16 years old, including one fatality, are reported from rural settings in forest areas of Chone (n = 2) and Flavio Alfaro (n = 3) counties, northern Manabí province, western Ecuador. Three cases were graded as Class II (moderate) and two in Class III (severe) envenoming. Manifestations showed characteristic autonomic nervous system hyper-stimulation and the fatality (a 1.9-year-old boy from Flavio Alfaro) was due to cardio-respiratory failure. Marked leukocytosis in four of the cases (21,800-31,800 cells/mm(3)), with notable neutrophilia (58-82%), suggests induction of a venom-mediated systemic inflammatory response-like syndrome. Specimens responsible for cases in Flavio Alfaro County, including the fatality, were classified as Tityus asthenes Pocock, accountable for severe scorpionism in Colombia. These findings demand implementation of control and therapeutic measures in affected areas in Ecuador, including evaluation of available scorpion antivenoms.

A checklist of the scorpions of Ecuador (Arachnida: Scorpiones), with notes on the distribution and medical significance of some species.

Ecuador harbors one of the most diverse Neotropical scorpion faunas, hereby updated to 47 species contained within eight genera and five families, which inhabits the "Costa" (n = 17), "Sierra" (n = 34), "Oriente" (n = 16) and "Insular" (n = 2) biogeographical regions, corresponding to the western coastal, Andean, Amazonian, and the Galápagos archipelago regions, respectively. The genus Tityus Koch, in the family Buthidae, responsible for severe/fatal accidents elsewhere in northern South America and the Amazonia, is represented in Ecuador by 16 species, including T. asthenes, which has caused fatalities in Colombia and Panama, and now in the Ecuadorian provinces of Morona Santiago and Sucumbíos. Underestimation of the medical significance of scorpion envenoming in Ecuador arises from the fact that Centruroides margaritatus (Gervais) (family Buthidae) and Teuthraustes atramentarius Simon (family Chactidae), whose venoms show low toxicity towards vertebrates, frequently envenom humans in the highly populated Guayas and Pichincha provinces. This work also updates the local scorpion faunal endemicity (74.5 %) and its geographical distribution, and reviews available medical/biochemical information on each species in the light of the increasing problem of scorpionism in the country. A proposal is hereby put forward to classify the Ecuadorian scorpions based on their potential medical importance.

A checklist of the scorpions of Ecuador (Arachnida: Scorpiones), with notes on the distribution and medical significance of some species

Ecuador harbors one of the most diverse Neotropical scorpion faunas, hereby updated to 47 species contained within eight genera and five families, which inhabits the Costa (n= 17), Sierra (n= 34), Oriente (n= 16) and Insular (n= 2) biogeographical regions, corresponding to the western coastal, Andean, Amazonian, and the Galápagos archipelago regions, respectively. The genusTityus Koch, in the family Buthidae, responsible for severe/fatal accidents elsewhere in northern South America and the Amazonia, is represented in Ecuador by 16 species, including T. asthenes, which has caused fatalities in Colombia and Panama, and now in the Ecuadorian provinces of Morona Santiago and Sucumbíos. Underestimation of the medical significance of scorpion envenoming in Ecuador arises from the fact thatCentruroides margaritatus (Gervais) (family Buthidae) andTeuthraustes atramentarius Simon (family Chactidae), whose venoms show low toxicity towards vertebrates, frequently envenom humans in the highly populated Guayas and Pichincha provinces. This work also updates the local scorpion faunal endemicity (74.5 %) and its geographical distribution, and reviews available medical/biochemical information on each species in the light of the increasing problem of scorpionism in the country. A proposal is hereby put forward to classify the Ecuadorian scorpions based on their potential medical importance.