Linear B-cell epitopes in BthTX-1, BthTX-II and BthA-1, phospholipase A2's from Bothrops jararacussu snake venom, recognized by therapeutically neutralizing commercial horse antivenom

Abstract

The benefits from treatment with antivenom sera are indubitable. However, the mechanism for toxin neutralization has not been completely elucidated. A mixture of anti-bothropic and anti-crotalic horse antivenom has been reported to be more effective in neutralizing the effects of Bothrops jararacussu snake venom than anti-bothropic antivenom alone. This study determined which regions in the three PLA₂s from B. jararacussu snake venom are bound by antibodies in tetravalent anti-bothropic and monovalent anti-crotalic commercial horse antivenom. Mapping experiments of BthTX-I, BthTX-II and BthA-I using two small libraries of 69 peptides each revealed six major IgG-binding epitopes that were recognized by both anti-bothropic and anti-crotalic horse antivenom. Two epitopes in BthTX-I were only recognized by the anti-bothropic horse antivenom, while anti-crotalic horse antivenom recognized four unique epitopes across the three PLA₂s. Our studies suggest that the harmful activities of the PLA₂s present in the venom of B. jararacussu are neutralized by the combinatorial treatment with both antivenom sera through their complementary binding sites, which provides a wide coverage on the PLA₂s. This is the first peptide microarray of PLA₂s from B. jararacussu snake venom to survey the performance of commercial horse antiophidic antivenom. Regions recognized by the protective antivenom sera are prime candidates for improved venom cocktails or a chimeric protein encoding the multiple epitopes to immunize animals as well as for designing future synthetic vaccines.

High-throughput immuno-profiling of mamba (Dendroaspis) venom toxin epitopes using high-density peptide microarrays

“…The results obtained clearly demonstrate the power of this high-throughput approach compared to similar methods based on SPOT synthesis6712 and mutation analysis11. Furthermore, toxin epitopes recognised by antivenom antibodies were unveiled for all but two of the manually curated toxins from every mamba and four cobra species endemic to sub-Saharan Africa.…”

“…During the immunisation process, a diverse pool of antibodies is raised, and these antibodies bind to and neutralise toxins and non-toxic components present in the venoms45. Despite the clinical importance of many snake venoms, little is known about the specific interactions between antivenom antibodies and snake venom toxins67.…”

“…Recently, studies involving immunoassay quantification of antivenom binding to immobilised synthetic peptides (SPOT synthesis), corresponding to individual segments of the amino acid sequence of a given toxin, have added valuable molecular insight by elucidating which sequences contain linear elements of epitopes recognised by given antivenoms67121314. These kind of meticulous epitope mapping experiments have also been performed on toxins from Tityus scorpions151617 and Loxosceles spiders18.…”

Cross-recognition of a pit viper (Crotalinae) polyspecific antivenom explored through high-density peptide microarray epitope mapping

“…simus D49 PLA 2 s in the immunization mixture. A previous study investigated the different recognition profiles of two antivenoms prepared by immunization with either four Bothrops species or a Crotalus snake [23]. The study involved peptides derived from three individual PLA 2 s (one from each of the non-crotoxin groups) from B .…”

“…To gain molecular level insight into para-specificity, antivenom cross-recognition of individual toxins can be assessed using synthetic peptides representing linear elements of B-cell epitopes on the toxins [23,24]. Despite the inability to evaluate discontinuous epitopes, a growing number of studies have proven the usefulness of linear epitope analyses in antivenom research.…”