Interactive effects of competition and predator cues on immune responses of leopard frogs at metamorphosis

Recent hypotheses suggest that immunosuppression, resulting from altered environmental conditions, may contribute to the increased incidence of amphibian disease around the world. Antimicrobial peptides (AMPs) in amphibian skin are an important innate immune defense against fungal, viral and bacterial pathogens. Their release is tightly coupled with release of the stress hormone noradrenaline (norepinephrine). During metamorphosis, AMPs may constitute the primary immune response in the skin of some species because acquired immune functions are temporarily suppressed in order to prevent autoimmunity against new adult antigens. Suppression of AMPs during this transitional stage may impact disease rates. We exposed leopard frog tadpoles (Lithobates pipiens) to a factorial combination of competitor and caged-predator environments and measured their development, growth and production of hydrophobic skin peptides after metamorphosis. In the absence of predator cues, or if the exposure to predator cues was late in ontogeny, competition caused more than a 250% increase in mass-standardized hydrophobic skin peptides. Predator cues caused a decrease in mass-standardized hydrophobic skin peptides when the exposure was late in ontogeny under low competition, but otherwise had no effect. Liquid chromatography tandem mass spectrometry of the skin peptides showed that they include six AMPs in the brevinin and temporin families and at least three of these peptides are previously uncharacterized. Both of these peptide families have previously been shown to inhibit harmful microbes including Batrachochytrium dendrobatidis, the fungal pathogen associated with global amphibian declines. Our study shows that amphibians may be able to adjust their skin peptide defenses in response to stressors that are experienced early in ontogeny and that these effects extend through an important life-history transition.