Jair Mendes Lima Ferreira Junior^1,2, Lorena Vitória da Costa Sousa^1,2, Donat Alexander de Chapeaurouge¹, Bárbara Cibelle Soares Farias Quintela¹, ANNA CAROLINA MACHADO MARINHO¹

Introduction: Bothrops venoms comprise complex mixtures of proteins and peptides with hemotoxic, neurotoxic, and cytotoxic effects. Characterizing these components is essential for understanding venom mechanisms and improving antivenom efficacy.  Snakebite envenomation is a neglected tropical disease that causes significant morbidity and mortality in rural Brazil, where antivenom therapy remains the only definitive treatment. Regional variations in venom composition may reflect ecological adaptations, highlighting the need for geographically targeted proteomic studies to optimize snakebite therapies. Objective: Identify and compare protein profiles in Bothrops venoms (B. leucurus, B. atrox, B. jararacussu) from Northeast, North, and Southeast Brazil using advanced proteomics, elucidating regional variations and potential biomarkers. Materials and Methods: Lyophilized venoms were solubilized, reduced/alkylated, and digested with trypsin (37°C, 16 h) after RapiGest SF treatment. Peptides were separated by RP-LC (Q Exactive Plus Orbitrap MS) and analyzed via HCD fragmentation. Data were processed with Proteome Discoverer against UniProt’s snake venom database, considering fixed carbamidomethylation and variable modifications (e.g., oxidation). Results and Discussion: Conserved components: 138 proteins were shared across all three species, suggesting core biological functions. Unique signatures: Species-specific proteins included 69 (B. jararacussu), 191 (B. atrox), and 67 (B. leucurus), potentially serving as regional biomarkers. Toxin families identified: Phospholipases A₂ (PLA₂s), metalloproteinases (SVMPs), serine proteases, L-amino acid oxidases, myotoxins, C-type lectins, and thrombin-like enzymes dominated the profiles, consistent with known venom pathomechanisms. Conclusion: This study reveals significant regional variations in Bothrops venom composition, likely driven by ecological or genetic factors. These findings underscore the importance of region-specific antivenom development and provide a foundation for future research on venom evolution and clinical management of envenomation.

Agradecimentos: Os autores agradecem à Fundação Oswaldo Cruz Ceará (Fiocruz Ceará), ao Sistema Único de Saúde (SUS), à Escola Estadual de Ensino Profissional Eusébio de Queiroz, à Escola Politécnica Joaquim José Venâncio (FIOCRUZ) e ao Programa de Vocação Científica (PROVOC) pelo apoio institucional e pela contribuição para a realização deste trabalho.