Biological and proteolytic variation in the venom of Crotalus scutulatus scutulatus from Mexico

Rattlesnake venoms may be classified according to the presence/absence and relative abundance of the neurotoxic phospholipases A2s (PLA2s), such as Mojave toxin, and snake venom metalloproteinases (SVMPs). In Mexico, studies to determine venom variation in Mojave Rattlesnakes (Crotalus scutulatus scutulatus) are limited and little is known about the biological and proteolytic activities in this species. Tissue (34) and venom (29) samples were obtained from C. s. scutulatus from different locations within their distribution in Mexico. Mojave toxin detection was carried out at the genomic (by PCR) and protein (by ELISA) levels for all tissue and venom samples. Biological activity was tested on representative venoms by measuring LD50 and hemorrhagic activity. To determine the approximate amount of SVMPs, 15 venoms were separated by RP-HPLC and variation in protein profile and proteolytic activity was evaluated by SDS-PAGE (n = 28) and Hide Powder Azure proteolytic analysis (n = 27). Three types of venom were identified in Mexico which is comparable to the intraspecific venom diversity observed in the Sonoran Desert of Arizona, USA: Venom Type A ( Type II), with Mojave toxin, highly toxic, lacking hemorrhagic activity, and with scarce proteolytic activity; Type B ( Type I), without Mojave toxin, less toxic than Type A, highly hemorrhagic and proteolytic; and Type A + B, containing Mojave toxin, as toxic as venom Type A, variable in hemorrhagic activity and with intermediate proteolytic activity. We also detected a positive correlation between SVMP abundance and hemorrhagic and proteolytic activities. Although more sampling is necessary, our results suggest that venoms containing Mojave toxin and venom lacking this toxin are distributed in the northwest and southeast portions of the distribution in Mexico, respectively, while an intergradation in the middle of both zones is presentConsejo Nacional de Ciencia y Tecnologia/[221343]/CONACYT/MéxicoUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto Clodomiro Picado (ICP)UCR::Vicerrectoría de Docencia::Salud::Facultad de Microbiologí

Conocimiento etnoherpetológico de dos comunidades aledañas a la reserva estatal sierra de montenegro, Morelos, México

La fauna silvestre aporta a las comunidades campesinas de México satisfactores como carne, huevos o estructuras anatómicas para la elaboración de artesanías y servicios ambientales que garantizan su bienestar social. El objetivo del presente trabajo fue analizar el conocimiento etnoherpetológico en dos comunidades aledañas a la Reserva Estatal Sierra de Montenegro, Morelos. Se utilizaron técnicas etnozoológicas: entrevistas abiertas y se aplicaron 105 cuestionarios semi-estructurados a hombres y mujeres. Para la identificación de las especies se llevaron a cabo recorridos guiados con el apoyo de expertos naturalistas locales. Los informantes reconocen 32 especies herpetofaunísticas, de las cuales 15 (48%) presentan valor cultural y, de éstas, 9 (29%) tienen lossiguientes valores de uso: alimento, medicinal, ornamental y mascota. Las especies de anfibios más conocidas fueron el sapo (Rhinella marina) y la rana (Lithobates zweifeli), utilizados como medicina y alimento. Ya para los reptiles, la iguana negra (Ctenosaura pectinata) como alimento, medicina y mascota; la víbora de cascabel (Crotalus culminatus) como alimento, medicina, ornamento y mascota; y el tilcuate (Drymarchon melanurus) en relatos. Los habitantes poseen conocimiento acerca de anfibios y reptiles, reflejado en las formas de manejo y uso y en la manera en que los perciben, así como creencias, cuentos y relatos. Los habitantes identifican hábitos de las diferentes especies, así como aquellas venenosas y no venenosas. Los estudios etnoherpetológicos aportanelementos factibles de ser integrados a planes de manejo y conservación de la herpetofauna

Intraspecies variation in the venom of the rattlesnake Crotalus simus from Mexico: Different expression of crotoxin results in highly variable toxicity in the venoms of three subspecies

The composition and toxicological profile of the venom of the rattlesnake Crotalus simus in Mexico was analyzed at the subspecies and individual levels. Venoms of the subspecies C. s. simus, C. s. culminatus and C. s. tzabcan greatly differ in the expression of the heterodimeric neurotoxin complex ‘crotoxin’, with highest concentrations in C. s. simus, followed by C. s. tzabcan, whereas the venom of C. s. culminatus is almost devoid of this neurotoxic PLA2. This explains the large variation in lethality (highest in C. s. simus, which also exerts higher myotoxicity). Coagulant activity on plasma and fibrinogen occurs with the venoms of C. s. simus and C. s. tzabcan, being absent in C. s. culminatus which, in turn, presents higher crotamine-like activity. Proteomic analysis closely correlates with toxicological profiles, since the venom of C. s. simus has high amounts of crotoxin and of serine proteinases, whereas the venomof C. s. culminatus presents higher amounts of metalloproteinases and crotamine. This complex pattern of intraspecies venom variation provides valuable information for the diagnosis and clinicalmanagement of envenoming by this species in Mexico, as well as for the preparation of venom pools for the production and quality control of antivenoms. Biological significance This study describes the variation in venomcomposition and activities of the three subspecies of Crotalus simus fromMexico. Results demonstrate that there is a notorious difference in these venoms, particularly regarding the content of the potent neurotoxic phospholipase A2 complex ‘crotoxin’. In addition, other differences were observed regarding myotoxic and coagulant activities, and expression of the myotoxin ‘crotamine’. These findings have implications in, at least, three levels: (a) the adaptive role of variations in venom composition; (b) the possible differences in the clinical manifestations of envenomings by these subspecies in Mexico; and (c) the design of venom mixtures for the preparation of antivenoms effective in the neutralization of the venoms of the three subspecies.Consejo Nacional de Rectores//CONARE/Costa RicaUniversidad de Costa Rica/[741-B2-652]/UCR/Costa RicaConsejo Nacional de Ciencia y Tecnología//CONACyT/MéxicoUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto Clodomiro Picado (ICP

Venom of the neotropical rattlesnake, Crotalus culminatus: Intraspecific variation, neutralization by antivenoms, and immunogenicity in rabbits

Crotalus culminatus is a medically significant species of rattlesnake in Mexico [1]. While the proteomic composition of its venom has been previously reported for both juvenile and adult specimens, there has been limited research into its functional properties, with only a few studies, including one focusing on coagulotoxicity mechanisms. In this study, we aimed to compare the biochemical and biological activities of the venom of juvenile and adult snakes. Additionally, we assessed antibody production using the venoms of juveniles and adults as immunogens in rabbits. Our findings reveal lethality and proteolytic activity differences between the venoms of juveniles and adults. Notably, juvenile venoms exhibited high proportions of crotamine, while adult venoms displayed a reduction of this component. A commercially available antivenom demonstrated effective neutralization of lethality of both juvenile and adult venoms in mice. However, it failed to neutralize the paralytic activity induced by crotamine, which, in contrast, was successfully inhibited by antibodies obtained from hyperimmunized rabbits. These results suggest the potential inclusion of C. culminatus venom from juveniles in commercial antivenom immunization schemes to generate antibodies targeting this small myotoxin.UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto Clodomiro Picado (ICP)UCR::Vicerrectoría de Docencia::Salud::Facultad de Microbiologí

Functional, proteomic and transcriptomic characterization of the venom from Micrurus browni browni: identification of the first lethal multimeric neurotoxin in coral snake venom

Proteomic characterization of Micrurus browni browni venom showed approximately 41 components belonging to 9 protein families, mainly phospholipases A2 (PLA2s) and three-finger toxins (3FTxs). Venom gland transcriptome yielded 39 venom transcripts belonging to 10 protein families. Functional characterization identified a multimeric toxin, here designated Brownitoxin-1, which comprises at least one PLA2 and one 3FTx. Its components have no or very low lethality individually but become extremely lethal when combined; both were partially characterized. Other two lethal components were identified: A neurotoxic PLA2, and a postsynaptic α-neurotoxin. LD50s as well as PLA2 and nAChR-blocking activities were determined for whole venom and isolated components. Application of venom to murine neuromuscular preparations caused a progressive decrease of twitch force that was irreversible after washing. Inhibition of PLA2 activity with p-bromophenacyl bromide (pBPB) showed that approximately 90% of toxicity is dependent on this activity. Non-lethal components include diverse 3FTxs, at least three enzymatically active PLA2s and the nociceptive toxin MitTx. No evidence of specificity towards prey was observed. This work is one of the most complete characterizations of a coral snake venom so far and its findings highlight the relevance of protein complexes in venom function.UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto Clodomiro Picado (ICP

Proteomic and toxicological characterization of the venoms of the most enigmatic group of rattlesnakes: The long-tailed rattlesnakes

The most enigmatic group of rattlesnakes is the long-tailed rattlesnake group, consisting of three species: Crotalus ericsmithi, Crotalus lannomi and Crotalus stejnegeri. These species have been the least studied rattlesnakes in all aspects, and no study on the characterization of their venoms has been carried out to date. Our main objective was to investigate the proteomic composition, as well as some of the biochemical and toxic activities of these venoms, and their neutralization by commercial antivenom. The venom proteome of C. ericsmithi mainly contains metalloproteinases (SVMP; 49.3%), phospholipases A2 (PLA2; 26.2%), disintegrins (Dis; 12.6%), and snake venom serine proteases (SVSP; 6.8%), while C. lannomi venom mainly consists of SVMP (47.1%), PLA2 (19.3%), Dis (18.9%), SVSP (6%) and l-amino acid oxidase (LAAO; 2.6%). For these venoms high lethality was recorded in mice, the most potent being that of C. lannomi (LD50 of 0.99 μg/g body weight), followed by C. ericsmithi (1.30 μg/g) and finally C. stejnegeri (1.79 μg/g). The antivenoms Antivipmyn® from SILANES and Fabotherapic polyvalent antiviperin® from BIRMEX neutralized the lethal activity of the three venoms. Although this group of snakes is phylogenetically related to the C. viridis group, no neurotoxic components (crotoxin or crotoxin-like proteins) common in rattlesnakes were found in their venoms. This study expands current knowledge on the venoms of understudied snake species of the Mexican herpetofaunaPrograma de Apoyo a Proyectos de Investigación e Innovación Tecnológica/[IN- 211621]/PAPIIT/MéxicoConsejo Nacional de Ciencia y Tecnología/[264255]/CONACYT/MéxicoConsejo Nacional de Ciencia y Tecnología/[303045]/CONACYT/MéxicoUniversidad de Costa Rica/[]/UCR/Costa RicaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto Clodomiro Picado (ICP