Snake Venomics of the Arboreal Talamancan Palm-Pitviper, <i>Bothriechis nubestris</i>, Provides Clues on the Origin of a Phenotypic Dichotomy between Type-I and Type-II Venoms

We report the biochemical and proteomic characterization of the venom of the arboreal Talamancan palm-pitviper, <i>Bothriechis nubestris</i>, a species endemic to north and central parts of the Costa Rican Cordillera de Talamanca, at 2100 to 3000 m above sea level. The Talamancan palm-pitviper venom arsenal comprised the translated products of 26 unique transcripts into 10 toxin classes, where metalloproteinases PIIa and PIII represent the dominant components. <i>In vitro</i>, the venom proteolyzed azocasein and gelatin but showed no phospholipase A₂ or human plasma coagulant activities. <i>In vivo</i>, <i>B. nubestris</i> venom exhibited an intravenous median lethal dose (LD₅₀) of 21.5 (95% confidence interval: 15.6-29.5) μg/mouse, and a minimum hemorrhagic dose (MHD) of 1.85 μg. PoliVal-ICP antivenom neutralized the venom's lethal activity with a potency of 9.7 mg of venom/g of antivenom and significantly reduced the hemorrhagic effect. Comparison of venom gland transcriptomes and venom proteomes of <i>B. nubestris</i> and its closest congeneric relative, <i>B. nigroviridis</i>, revealed that highly conserved venom gland transcriptomes are differentially processed by each species to produce divergent (type-I vs type-II, respectively) venoms. This phenomenon contributes to the remarkable venom phenotypic variability found across the palm-pitviper phylogeny. A possible mechanism for the occurrence of type-I/type-II venom phenotypic dichotomy is discussed.