The 'Wallum' of coastal east Australia is an area of sand plains and dunes where waters are both soft and acidic. Field and laboratory studies (chapter 2) show anuran amphibian species native to this area are highly acid-tolerant. Tolerance of Wallum-native anuran species to chronic acid stress is especially remarkable with larvae surviving to metamorphosis in waters pH 3.5 and less, waters lethal to larvae of most amphibian species and most fish.
In this PhD study, I investigated acid adaptation and mechanisms for softwater acid tolerance in larvae of anuran species native to the Wallum. To begin with, field and laboratory studies were undertaken to assess larval acid tolerance amongst a range of Wallum-native and non-Wallum-native species (chapter 2). These studies revealed a high level of acid tolerance amongst larvae of Wallum-native anuran species. Multiple independent comparisons between and within species suggest this
tolerance is the result of evolutionary adaptation. Next, I investigated histopathological changes to the gills and integument of anuran larvae acutely exposed to low pH (chapter 3). This investigation yielded significant insight into the toxicological effects of low pH on anuran larvae. Changes to the gills and integument of larvae with acid exposure included opening of cellular tight junctions of the integument and gills, epithelial necrosis, and sloughing of the integument. These changes in morphology were accompanied by disruption of Na+ balance due to both inhibition of Na+ uptake and increased passive ionic efflux of Na+.
To determine how larvae of Wallum-native species resist acid damage to gills and the integument and maintain ionic homeostasis at low pH, ion regulation and morphology of gills and the integument were compared in larvae of Wallum-native (Litoria
cooloolensis and Limnodynastes terraereginae) and acid-sensitive, non-Wallum-native anuran species (Litoriafallax and Limnodynastes peronii) (chapters 4 - 7). Ion regulation and morphology of gills and the integument were also compared in larvae of L. terraereginae from different pH (Wallum and non-Wallum) environs (chapters 6 and 7).
Studies of ion regulation (chapters 4 and 6) showed larvae of Wallum-native species were more resistant to Na+ loss than acid-sensitive L. fallax and L. peronii larvae in acid water. Wallum L. terraereginae larvae were also more resistant to increased ionic efflux than non-Wallum L. terraereginae at pH 3.5. In L. cooloolensis and L. terraereginae larvae (but not acid-sensitive L. fallax and
L. peronii larvae) the addition of Ca2+ to water reduced Na+ efflux at pH 3.5. As well as resisting increased Na+ efflux, acid-acclimated L. cooloolensis and L. terraereginae larvae were able to maintain Na+ uptake at pH 3.5. Analysis of Na+ transport kinetics suggests maintenance of Na+ uptake by these larvae is attributable to a low Km and high Vmax for Na+. Control of ionic efflux by acid-acclimated larvae may be attributed, in part, to depression of body Na+ content with the body Na+ content of acid-acclimated larvae 20% lower than larvae reared at pH 6.5.
Examination of the gills and integument via light, scanning and transmission electron microscopy
revealed significant differences between species and acid-naive and acid-acclimated larvae in mucous secretion and cell-cell junctional morphometry at both the gills and integument. These differences include closer apposition of epithelial cells of the gills and integument in L. cooloolensis larvae vs. L. fallax larvae, greater mucous production in L. terraereginae vs. L. peronii larvae and greater mucous production / secretion at the gills and integument in acid-acclimated L. cooloolensis and L. terraereginae larvae.
Based on these results, tolerance of Wallum-native species to acute acid stress may be attributed to greater mucous production and secretion at the integument and gills and a closer apposition of epithelial cells of the gills and integument. Tolerance of L. cooloolensis and L. terraereginae
larvae to chronic acid stress, meanwhile, may be attributed to a low Km and high Vmax for Na+ transport, depression of body Na+ content, and increased mucous production and secretion at the gills and integument.