Effects of invasion history on physiological responses to immune system activation in invasive Australian cane toads

Abstract

The cane toad (Rhinella marina) has undergone rapid evolution during its invasion of tropical Australia. Toads from invasion front populations (in Western Australia) have been reported to exhibit a stronger baseline phagocytic immune response than do conspecifics from range core populations (in Queensland). To explore this difference, we injected wild-caught toads from both areas with the experimental antigen lipopolysaccharide (LPS, to mimic bacterial infection) and measured whole-blood phagocytosis. Because the hypothalamic-pituitary-adrenal axis is stimulated by infection (and may influence immune responses), we measured glucocorticoid response through urinary corticosterone levels. Relative to injection of a control (phosphate-buffered saline), LPS injection increased both phagocytosis and the proportion of neutrophils in the blood. However, responses were similar in toads from both populations. This null result may reflect the ubiquity of bacterial risks across the toad's invaded range; utilization of this immune pathway may not have altered during the process of invasion. LPS injection also induced a reduction in urinary corticosterone levels, perhaps as a result of chronic stress.

Keywords: Cane toad; Eco-immunology; Invasive species; Phagocytosis; Rhinella marina.

Figure 1. Map of cane toad distribution in Australia.

Current distribution of the cane toad throughout Australia. Toads were first introduced to Queensland (QLD) in 1935, and have since expanded their range into New South Wales, the Northern Territory (NT), and Western Australia (WA). Black diamonds indicate our toad collection sites: Cairns, QLD and Oombulgurri, WA.

Figure 2. Phagocytic responses of cane toads to lipopolysaccharide (LPS).

Phagocytosis curves of cane toads (Rhinella marina) from (A) Queensland (QLD) and (B) Western Australia (WA) both before and after injection with either lipopolysaccharide (LPS) or phosphate-buffered saline (PBS). The average of all samples within a treatment group at each time point (N = 5) was calculated to produce the points on the graph. (C) Difference in mean luminescence values between pre-injection and post-injection readings of each population and treatment group.

Figure 3. Neutrophil responses of cane toads to lipopolysaccharide (LPS).

(A) Changes in the percentages of neutrophils (as induced by injection of either lipopolysaccharide [LPS] or phosphate-buffered saline [PBS]) across two treatment groups of cane toads (Rhinella marina) from two populations (invasion front site in Western Australia [WA] and range core site in Queensland [QLD]). The percentage of neutrophils in the toad’s blood pre-injection was subtracted from the percentage of neutrophils in the same toad’s blood post-injection. The average of the difference between pre-injection and post-injection of all samples within a treatment group at each time point (N = 5) was calculated to produce the points on the graph. Error bars indicate standard error. (B) Positive correlation between the changes in neutrophil percentage and PC1 before vs after injection.

Figure 4. Urinary corticosterone responses of cane toads to lipopolysaccharide (LPS).

Changes in urinary corticosterone levels across two treatment groups of cane toads (Rhinella marina) from two populations (invasion front site in Western Australia [WA] and range core site in Queensland [QLD]). Toads were injected with either lipopolysaccharide (LPS) or phosphate-buffered saline (PBS). Error bars indicate standard error, and lines are fitted by linear regression to data from each treatment group.