# Innate Immunity in the Cottonmouth Watersnake (Agkistrodon piscivorus)

## TL;DR

This study explores the strong innate immune system of the cottonmouth snake, revealing its powerful ability to fight infections and potential applications for medical research.

## Contribution

The first characterization of innate immunity in the cottonmouth snake (Agkistrodon piscivorus), identifying potent antibacterial and hemolytic activities.

## Key findings

- Cottonmouth plasma shows strong antibacterial activity against both Gram-negative and Gram-positive bacteria.
- Plasma exhibits potent hemolytic activity, inhibited by heat, proteases, and EDTA, suggesting complement protein involvement.
- A 36 kDa protein was identified, indicating the lectin pathway of complement activation may be active in this species.

## Abstract

Reptiles play an important role in nature, but unlike mammals, their immune systems are not well understood. This study examined the cottonmouth snake, a venomous species found in the southeastern United States, to better understand how it defends itself against infection. We found that the snake’s blood has strong natural defenses, able to kill harmful bacteria and break down foreign cells very effectively. In fact, its defensive power was far stronger than that found in other ectothermic animals. This study also identified specific proteins in the blood that may be responsible for these protective effects. These findings are valuable because they help fill a major gap in knowledge about reptile health and survival. Understanding how reptiles fight infection could also inspire new medical research, potentially leading to improved treatments for human and animal diseases.

Despite their ecological importance and unique evolutionary history, reptiles remain underrepresented in immunological research. The innate immunity of the cottonmouth (Agkistrodon piscivorus), a semi-aquatic pit viper native to the southeastern United States, was characterized to provide insight into the molecular and cellular mechanisms underlying its first line of defense against pathogens. Plasma collected from wild A. piscivorus exhibited strong antibacterial activities against both Gram-negative and Gram-positive bacteria. In addition, plasma from A. piscivorus showed potent hemolytic activities in unsensitized sheep red blood cell (SRBC) hemolysis assays. This activity was concentration-, time-, and temperature-dependent. In addition, the hemolytic activity was inhibited by mild heat treatment (56 °C, 30 min) of plasma and proteases and also by EDTA, suggesting that the hemolytic activity was due to the presence of serum complement proteins. SDS-PAGE analysis of plasma proteins isolated from a mannan-agarose affinity column revealed the presence of a protein with a mass of 36 kDa, raising the strong possibility that the lectin pathway of complement activation is active. The EC50 for hemolysis of SRBCs by plasma from A. piscivorus was approximately 10–100× lower than that of any other reptilian species described. This is the first study to characterize innate immunity in A. piscivorus.