Centre for Integrative Ecology

The question about whether evolution is unpredictable and stochastic or intermittently constrained along predictable pathways is the subject of a fundamental debate in biology, in which understanding convergent evolution plays a central role. At the molecular level, documented examples of convergence are rare and limited to occurring within specific taxonomic groups. Here we provide evidence of constrained convergent molecular evolution across the metazoan tree of life. We show that resistance to toxic cardiac glycosides produced by plants and bufonid toads is mediated by similar molecular changes to the sodium-potassium-pump (Na + /K + -ATPase) in insects, amphibians, reptiles, and mammals. In toadfeeding reptiles, resistance is conferred by two point mutations that have evolved convergently on four occasions, whereas evidence of a molecular reversal back to the susceptible state in varanid lizards migrating to toad-free areas suggests that toxin resistance is maladaptive in the absence of selection. Importantly, resistance in all taxa is mediated by replacements of 2 of the 12 amino acids comprising the Na + /K + -ATPase H1-H2 extracellular domain that constitutes a core part of the cardiac glycoside binding site. We provide mechanistic insight into the basis of resistance by showing that these alterations perturb the interaction between the cardiac glycoside bufalin and the Na + /K + -ATPase. Thus, similar selection pressures have resulted in convergent evolution of the same molecular solution across the breadth of the animal kingdom, demonstrating how a scarcity of possible solutions to a selective challenge can lead to highly predictable evolutionary responses. constraint | parallelism | genotype phenotype | ion transporters | bufotoxin cardenolide

Invasive species can induce shifts in habitat use by native taxa: either by modifying habitat availability,
or by repelling or attracting native species to the vicinity of the invader. The ongoing invasion of cane toads (Rhinella
marina) through tropical Australia might affect native frogs by affecting refuge-site availability, because both frogs
and toads frequently shelter by day in burrows. Our laboratory and field studies in the wet-dry tropics show that
native frogs of at least three species (Litoria tornieri, Litoria nasuta and Litoria dahlii) preferentially aggregate with
conspecifics, and with (some) other species of native frogs. However, the frogs rarely aggregated with cane toads
either in outdoor arenas or in standardized experimental burrows that we monitored in the field. The native frogs
that we tested either avoided burrows containing cane toads (or cane toad scent) or else ignored the stimulus (i.e.
treated such a burrow in the same way as they did an empty burrow). Native frogs selected a highly non-random
suite of burrows as diurnal retreat sites, whereas cane toads were less selective. Hence, even in the absence of toads,
frogs do not use many of the burrows that are suitable for toads. The invasion of cane toads through tropical
Australia is unlikely to have had a major impact on retreat-site availability for native frogs.

The question about whether evolution is unpredictable and stochastic or intermittently constrained along predictable pathways is the subject of a fundamental debate in biology, in which understanding convergent evolution plays a central role. At the molecular level, documented examples of convergence are rare and limited to occurring within specific taxonomic groups. Here we provide evidence of constrained convergent molecular evolution across the metazoan tree of life. We show that resistance to toxic cardiac glycosides produced by plants and bufonid toads is mediated by similar molecular changes to the sodium-potassium-pump (Na+/K+-ATPase) in insects, amphibians, reptiles, and mammals. In toadfeeding reptiles, resistance is conferred by two point mutations that have evolved convergently on four occasions, whereas evidence of a molecular reversal back to the susceptible state in varanid lizards migrating to toad-free areas suggests that toxin resistance is maladaptive in the absence of selection. Importantly, resistance in all taxa is mediated by replacements of 2 of the 12 amino acids comprising the Na+/K+-ATPase H1–H2 extracellular domain that constitutes a core part of the cardiac glycoside binding site. We provide mechanistic insight into the basis of resistance by showing that these alterations perturb the interaction between the cardiac glycoside bufalin and the Na+/K+-ATPase. Thus, similar selection pressures have resulted in convergent evolution of the same  molecular solution across the breadth of the animal kingdom, demonstrating how a scarcity of possible solutions to a selective challenge can lead to highly predictable evolutionary responses.

Invasive species can induce shifts in habitat use by native taxa: either by modifying habitat availability, or by repelling or attracting native species to the vicinity of the invader. The ongoing invasion of cane toads (Rhinella marina) through tropical Australia might affect native frogs by affecting refuge-site availability, because both frogs and toads frequently shelter by day in burrows. Our laboratory and field studies in the wet-dry tropics show that native frogs of at least three species (Litoria tornieri, Litoria nasuta and Litoria dahlii) preferentially aggregate with conspecifics, and with (some) other species of native frogs. However, the frogs rarely aggregated with cane toads either in outdoor arenas or in standardized experimental burrows that we monitored in the field. The native frogs that we tested either avoided burrows containing cane toads (or cane toad scent) or else ignored the stimulus (i.e. treated such a burrow in the same way as they did an empty burrow). Native frogs selected a highly non-random suite of burrows as diurnal retreat sites, whereas cane toads were less selective. Hence, even in the absence of toads, frogs do not use many of the burrows that are suitable for toads. The invasion of cane toads through tropical Australia is unlikely to have had a major impact on retreat-site availability for native frogs.