# First characterization of the venom of the endemic coral snake Micrurus camilae (Serpentes: Elapidae) from Colombia: Proteome, toxic activities, immunorecognition, and neutralization by antivenoms

## TL;DR

This study is the first to characterize the venom of the Colombian endemic coral snake Micrurus camilae, revealing its protein composition and toxic effects, and testing the effectiveness of antivenoms.

## Contribution

The first proteomic, toxicological, and antivenomic characterization of Micrurus camilae venom, including its phylogenetic placement.

## Key findings

- The venom proteome is rich in phospholipase A2 and includes 17 protein families.
- The venom exhibits lethal, myotoxic, edematogenic, and hemorrhagic activities.
- Commercial antivenoms failed to neutralize lethality, but an experimental rabbit antivenom prolonged mouse survival.

## Abstract

Thirty-one species of Micrurus (coral snakes) are distributed in Colombia. However, functional and proteomic analyses of their venoms have only been performed on six of them. Micrurus camilae is endemic to Colombia, and no information exists regarding its venom. The proteome of M. camilae venom, its biochemical and toxic activities, immunorecognition, and neutralization by commercial equine antivenoms and an experimental one prepared in rabbits are here reported. In addition, the phylogenetic position of M. camilae within the genus was explored. The venom was characterized by RP-HPLC, SDS-PAGE, and nESI-MS/MS, and functional analyses were performed using in vitro (proteolytic, coagulant, phospholipase A2, and L-amino acid oxidase activity) and in vivo (myotoxic, edematogenic, hemorrhagic) assays. Immunorecognition and neutralization were evaluated using ELISA and mouse lethality, respectively. To determine phylogenetic relationships, sequences of the mitochondrial ND4 gene from 48 Micrurus species were analyzed. The venom proteome revealed a PLA2-rich phenotype and identified 17 protein families, the four most abundant being PLA2, LAO, 3FTx, and MP. The myotoxic and hemorrhagic activities observed in mice correlated with the relative abundance of PLA2s and MPs, respectively. Furthermore, the i.p. lethal effect in mice was associated with only one fraction, a 3FTx. Two commercial equine antivenoms (INS-anticoral and ICP-anticoral) immunologically recognized both the whole venom and the chromatographic fractions by ELISA. However, they did not neutralize venom lethality in mice in a preincubation assay. On the other hand, the experimental rabbit antivenom was shown to recognize the whole venom and its fractions and, although it did not completely neutralize lethality, it prolonged mouse survival by several hours compared to the venom-only control. Our phylogenetic hypothesis showed M. camilae within the mipartitus group as a sister species of M. mipartitus.

Snakebite envenomation is a global public health problem. Snakes of the genus Micrurus are distributed throughout the Americas, and due to the composition of their venoms, they induce a neurotoxic syndrome that can be fatal if antivenom is not administered. Colombia harbors 31 Micrurus species, among which M. camilae is endemic, and its venom has not been previously studied. In this work, we present the first characterization of the composition and biological activities of M. camilae venom. In addition, we evaluated whether commercial antivenom and an experimental antivenom could neutralize its toxicity.The results revealed that the venom is composed of 17 protein families, with phospholipases A₂ being the most abundant, followed by L-amino acid oxidases, three-finger toxins, and metalloproteinases. The venom exhibited lethal activity, which was attributable to a single fraction corresponding to a three-finger toxin. Moreover, the venom showed myotoxic, edematogenic, and hemorrhagic effects, the latter being uncommon among venoms of this genus. The commercial antivenom did not neutralize the lethal effect of the venom; however, a species-specific experimental antivenom was able to prolong survival compared with the control. These findings highlight the urgent need to advance venomic and antivenomic studies on clinically relevant Micrurus species in Colombia.