Invasive species often encounter environmental conditions well outside those found in their native geographic ranges, and thus provide ideal model systems with which to explore responses to novel abiotic challenges. Within Australia, the invasive cane toad Rhinella marina has colonized areas that are considerably more arid than those found within its native range. Has the colonization of these novel environments been accompanied by shifts in physiology and/or locomotor performance? We measured rates of evaporative water loss, water gain, and effects of desiccation on locomotor performance of cane toads from two invasion fronts: one mesic (the wet-dry tropics) and one arid (the semi-desert). The two populations diverged substantially. Contrary to intuition (but consistent with intra-specific comparisons between other toad populations from mesic vs arid areas), rates of evaporative water loss were lower (not higher) in toads from the mesic population. However, arid-zone toads gained water more rapidly through their ventral surfaces, and rates of water loss and gain were highly correlated within individual toads from the arid-zone population. Rates of water exchange in laboratory-acclimated toads from the semi-arid zone did not differ from those of free-ranging conspecifics from the same population, suggesting that divergences between mesic and semi-arid toads reflect genetic changes that have occurred during the species’ Australian invasion. Mesic and semi-arid toads showed similar locomotor performance (endurance, distance per hop) when fully hydrated, but locomotor performance declined much more rapidly with desiccation in the mesic toads. Thus, within the short (decades-long) timespan of the cane toad's Australian invasion, there has been substantial population divergence in the ability to withstand desiccating conditions. If we are to accurately predict the distributions (and hence impacts) of invading organisms, we will need to include adaptation potential in risk assessment schemes.