Estudo de venenos de serpentes do complexo Bothrops, uma comparação da filogenia e da reatividade com antivenenos

Abstract

Frequently the venom composition has been considered as a reflex of snake phylogeny, with important implications for the production of antivenoms and treatment of envenomings. Snakes from Bothrops complex are responsible for most snake bites registered in Brazil, and their phylogenetic relationships are not yet completely clarified. Another important discussion is related to the efficacy of Bothrops Antivenom (SAB) in the treatment of envenoming by snakes whose venoms are not present in the immunization pool. Thus, the aim of this study was to analyze the composition of venoms from 6 species of snakes from Bothrops complex, and their correlation with snake phylogeny and reactivity with o SAB. In this way, venoms from Bothropoides jararaca, Bothropoides neuwiedi, Bothrops atrox, Bothrops jararacussu, Rhinocerophis cotiara and Rhinocerophis alternatus were analyzed by SDS-PAGE and reverse phase chromatography on HPLC C18 column. The reactivity of SAB with venoms and their fractions was performed by ELISA and western blot, followed by assessment of neutralization of lethality and hemorrhagic activity of the venoms of B. jararaca (present in immunization pool of SAB) and B. atrox (absent from the pool). The venoms showed distinct electrophoretic and chromatographic profiles, without apparent correlation with the snake phylogeny. Only venoms of the genus Rhinocerophis (R. alternatus and R. cotiara) showed relatively similar profiles. SAB recognized the venoms of different species with the same antibody titer of 640,000. Most of the fractions eluted from C18 column were recognized by SAB, especially those corresponding to snake venom metalloproteinases (SVMP) of class P-III. By SDS-PAGE, the venom of B. jararacussu presented the more distinct electrophoretic profile with predominance of phospholipase A2 (PLA2), while in the other venoms, the SVMPs were the predominant component. Two microlitters of antivenom were sufficient to protect 100 % of mice challenged with 3 LD50 of B. jararaca venom (105 µg), although for B. atrox venom (225 µg) the same effect was only obtained when doubling the antivenom amount. However, the SAB neutralized completely the hemorrhage induced by 10 µg of the both venoms, with the same volume of serum (4 µL). Thus, we conclude that SAB reacts similarly with 16 the same family of toxins from distinct venoms, regardless of the phylogeny of the snake or the presence of venom in the immunization pool employed in SAB production. This suggests that a Bothrops antivenom with greater efficiency and broad spectrum can be developed, if one considers the reactivity of antivenom to the different classes of toxins and not just the phylogeny of snakes.