Identification of protective B-cell epitopes of Atroxlysin-I: A metalloproteinase from Bothrops atrox snake venom.

Atroxlysin-I (Atr-I) is a hemorrhagic snake venom metalloproteinase (SVMP) from Bothrops atrox venom, the snake responsible for the majority of bites in the north region of South America. SVMPs like Atr-I produce toxic effects in victims including hemorrhage, inflammation, necrosis and blood coagulation deficiency. Mapping of B-cell epitopes in SVMPs might result in the identification of non-toxic molecules capable of inducing neutralizing antibodies and improving the anti-venom therapy. Here, using the SPOT-synthesis technique we identified two epitopes located in the N-ter region of Atr-I (AtrEp1-(22)YNGNSDKIRRRIHQM(36); and AtrEp2-(55)GVEIWSNKDLINVQ(68)). Based on the sequence of AtrEp1 and AtrEp2 a third peptide named Atr-I biepitope (AtrBiEp) was designed and synthesized ((23)NGNSDKIRRRIH(34)GG(55)GVEIWSNKDLINVQ(68)). AtrBiEp was used to immunize BALB/c mice. Anti-AtrBiEp serum cross-reacted against Atr-I in western blot and was able to fully neutralize the hemorrhagic activity of Atr-I. Our results provide a rational basis for the identification of neutralizing epitopes on Atr-I snake venom toxin and show that the use of synthetic peptides could improve the generation of immuno-therapeutics.

Partial in vitro analysis of toxic and antigenic activities of eleven Peruvian pitviper snake venoms.

This work used eleven Peruvian snake venoms (Bothrops andianus, Bothrops atrox, Bothrops barnetti, Bothrops castelnaudi, Bothriopsis chloromelas, Bothrocophias microphthalmus, Bothrops neuwiedi, Bothriopsis oligolepis, Bothriopsis peruviana, Bothrops pictus and Bothriopsis taeniata) to perform in vitro experimentation and determine its main characteristics. Hyaluronidase (HYAL), phospholipase A2 (PLA2), snake venom metalloproteinase (SVMP), snake venom serine protease (SVSP) and L-amino acid oxidase (LAAO) activities; toxicity by cell viability assays using MGSO3, VERO and HeLa cell lineages; and crossed immunoreactivity with Peruvian (PAV) and Brazilian (BAV) antibothropic polyvalent antivenoms, through ELISA and Western Blotting assays, were determined. Results show that the activities tested in this study were not similar amongst the venoms and each species present their own peculiarities, highlighting the diversity within Bothrops complex. All venoms were capable of reducing cell viability of all tested lineages. It was also demonstrated the crossed recognition of all tested venoms by both antivenoms.

Serological, biochemical and enzymatic alterations in rodents after experimental envenomation with Hadruroides lunatus scorpion venom.

Toxic effects of Peruvian Hadruroides lunatus scorpion venom on different biochemical and enzymatic parameters in blood serum of Wistar rats and Swiss mice were determined after experimental envenomation. An increase in enzymatic activities of Aspartate Aminotransferase (AST), Lactate Dehydrogenase (LDH) and levels of serum protein and albumin were observed while a decrease in creatinine level in serum was perceived after 30 min of envenomation. No alterations in urea levels and in kidney histology were detected in the envenomed rats. The global leukocytes count was diminished, with decrease in lymphocytes, eosinophils and neutrophils levels in the bloodstream, while no alterations were found in hematological parameters of red series in rats injected with H. lunatus venom. IL-2, IL-4, IL-6, INF-γ, TNF, IL-17A and IL-10 levels were evaluated 0.5, 3 and 6 h after experimental envenomation of mice with H. lunatus venom. From all the analyzed cytokines, only IL-6 showed an increase in serum levels. Taken together, these results point out that envenomation by H. lunatus can impair hematological and immunological parameters and therefore might be monitored in accidents involving this species.