Snake venom proteomics and antivenomics of two Sundaic lance-headed pit vipers: Trimeresurus wiroti (Malaysia) and Trimeresurus puniceus (Indonesia)

Envenomation by two medically important Sundaic pit vipers, Trimeresurus wiroti (Malaysia) and Trimeresurus puniceus (Indonesia), causes hemotoxic syndrome with a potentially fatal outcome. Research on the compositions and antigenicity of these pit viper venoms is however lacking, limiting our understanding of the pathophysiology and treatment of envenomation. This study investigated the venom proteomes of both species through a protein decomplexation strategy, applying C₁₈ reverse-phase high-performance liquid chromatography followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and protein identification through nano-electrospray ionization liquid chromatography-tandem mass spectrometry (nano-ESI-LCMS/MS) of trypsin-digested peptides. The venom antigenicity was profiled against the Thai Green Pit Viper Antivenom (GPVAV, a hetero-specific antivenom), using indirect enzyme-linked immunosorbent assay (ELISA). The venom proteomes of T. wiroti and T. puniceus consisted of 10 and 12 toxin families, respectively. The major proteins were of diverse snake venom serine proteases (19–30% of total venom proteins), snake venom metalloproteinases (17–26%), disintegrins (9–16%), phospholipases A₂ (8–28%) and C-type lectins (~8%). These were putative snake toxins implicated in hemorrhage and coagulopathy, consistent with clinical hemotoxicity. GPVAV showed strong immunorecognition toward high and medium molecular weight proteins (e.g., SVMP and PLA₂) in both venoms, while a lower binding activity was observed toward small proteins such as disintegrins. Conserved antigenicity in the major hemotoxins supported toxicity cross-neutralization by GPVAV and indicated that the immunorecognition of low molecular weight toxins may be optimized for improved binding efficacy. Taken together, the study provides insights into the pathophysiology and antivenom treatment of envenomation caused by T. wiroti and T. puniceus in the region.