Characterization of serum complement immune activity in the prairie rattlesnake (Crotalus viridis)

Abstract

Background: Reptile populations face a growing number of threats including global climate change and emerging pathogens. Unfortunately, research investigating the reptile immune system lags behind other taxa groups, hindering our ability to predict or mitigate species' response to threats. Baseline studies are critical for our understanding of reptile immune response and overall health of wild populations. Results: Treatment of unsensitized sheep red blood cells (SRBCs) with plasma collected from captive prairie rattlesnakes (Crotalus viridis) resulted in volume-dependent hemolysis, with a CH50 of 0.23 [plus or minus] 0.01 mL. Kinetic analyses at different temperatures revealed that the hemolysis was relatively rapid, with 50% of hemolytic activity occurring within 15 min (25 [degree]C), 18 min (35 [degree]C), or 23 min (15 [degree]C), and near maximum hemolysis within 60-90 min at all three temperatures. A comparative thermal profile revealed that complement activity was low at 5 [degree]C, but increased sharply at 10 and 15 [degree]C, and was maximal at 20-30 [degree]C. A steep decline in activity was noted at temperatures > 30 [degree]C. Mild heat treatment of the snake plasma (56 [degree]C, 30 min) or treatment with proteases completely abolished the hemolytic activity. In addition, inclusion of 5 mM EDTA inhibited 90% of the hemolysis, but the activity could be reconstituted with the addition of 15 mM Ca2+ or Mg2+, but not Ba2+, Cu2+, or Fe2+. Furthermore, the hemolysis was unaffected in the presence of 20 mM methylamine, indicating that the alternative mechanism of complement activation is responsible for the observed activities. Conclusions: Rattlesnakes show a relatively robust innate immune response as measured by hemolysis of SRBCs. However, hemolytic activity is reduced at high temperatures, indicating that rising global temperatures may have immune consequences for snake species, making them more vulnerable to known and emerging pathogens.