Edward A. Meyer

Minimising exposure of amphibians to pathogens during field studies

Many of the recent global amphibian mass mortalities, declines and extinctions have been attributed to the emerging infectious disease chytridiomycosis. There have been mass mortalities due to ranaviral disease but no major declines or extinctions. Controlling the transmission and spread of disease is of utmost importance, especially where there is the potential for human involvement. We have reviewed current hygiene guidelines for working with wild frogs, identified potential flaws and recommended those most suitable and effective for the field environment. Our within-site hygiene measures aim to reduce the risk of transmission among individuals. These measures encompass the capture, handling and holding of amphibians, skin disinfection before and after invasive procedures, marking frogs, sealing open wounds and treatment of accessory equipment. Our between-site hygiene measures aim to mitigate the risk of pathogen spread among populations. We have designed a risk calculator to help simplify and standardise the decision-making process for determining the level of risk and appropriate risk mitigation strategies to reduce the risk of increasing pathogen spread above background levels. Calculation of an overall risk score for pathogen spread takes into account the prior activity of field workers, the proposed activity, remoteness of the site, presence of known pathogens and the consequences of increased pathogen spread for amphibians in a given area.

Overview of the conservation status of Australian frogs

A review of the current conservation status of Australian amphibians was recently completed as part of a World Conservation Union (IUCN) sponsored Global Amphibian Assessment (GAA). Fifty of 216 amphibian species (23%) in Australia are now recognized as threatened or extinct in accord with IUCN Red List Categories and Criteria. Here we report on the categories and criteria under which individual species qualified for listing and provide a summary of supporting information pertaining to population and distribution declines. Major threatening processes contributing to listing of species are also reviewed.

Changes in cutaneous microbial abundance with sloughing: possible implications for infection and disease in amphibians

The emergence of disease as a significant global threat to amphibian diversity has generated considerable interest in amphibian defenses against cutaneous microbial infection and disease. To date, however, the influence of sloughing on the susceptibility of amphibians to infection and disease has been largely overlooked. To investigate the potential for sloughing to regulate topical microbial loads, the abundance of cultivable cutaneous bacteria and fungi in the cane toad Rhinella marina were compared before and after sloughing. Toads were also exposed to fluctuating thermal regimes (10-20 and 20-30°C) and variable photoperiods to investigate possible effects of season and climate on sloughing periodicity. Sloughing substantially reduced the abundance of cultivable cutaneous bacteria and fungi by up to 100%. The intermoult interval of toads maintained at 10-20°C was twice that of animals at 20-30°C and did not appear to thermally acclimate. Photoperiod had no discernable influence on sloughing periodicity. Results of this study suggest that normal sloughing cycles could play a significant role in controlling the persistence and build-up of cutaneous microbes, including pathogens. The loss of non-pathogenic commensal and protective skin microbiota after sloughing may also influence host susceptibility to cutaneous pathogens. We suggest that the spatio-temporal dynamics of chytridiomycosis, the widespread and often fatal disease caused by the fungal pathogen Batrachochytrium dendrobatidis, are related to temperature not only because of its effect on the growth of the fungus, but also because of its effect on the frequency of host sloughing.

First line of defence: the role of sloughing in the regulation of cutaneous microbes in frogs

The skin is the first line of defence in preventing the establishment of pathogens and associated infections in frogs. Regular sloughing of the outer layer can reduce the abundance of cultivable cutaneous microbes in green tree frogs which has ramifications for our understanding of cutaneous pathogens like the amphibian chytrid fungus.

Functional and morphological plasticity of crocodile (Crocodylus porosus) salt glands.

SUMMARY The estuarine crocodile, Crocodylus porosus, inhabits both freshwater and hypersaline waterways and maintains ionic homeostasis by excreting excess sodium and chloride ions via lingual salt glands. In the present study, we sought to investigate the phenotypic plasticity, both morphological and functional, in the lingual salt glands of the estuarine crocodile associated with chronic exposure to freshwater (FW) and saltwater (SW) environments. Examination of haematological parameters indicated that there were no long-term disruptions to ionic homeostasis with prolonged exposure to SW. Maximal secretory rates from the salt glands of SW-acclimated animals (100.8±14.7 μmol 100 g–0.7 body mass h–1) were almost three times greater than those of FW-acclimated animals (31.6±6.2 μmol 100 g–0.7 body mass h–1). There were no differences in the mass-specific metabolic rate of salt gland tissue slices from FW- and SW-acclimated animals (558.9±49.6 and 527.3±142.8 μl O2 g–1 h–1, respectively). Stimulation of the tissue slices from SW-acclimated animals by methacholine resulted in a 33% increase in oxygen consumption rate. There was no significant increase in the metabolic rate of tissues from FW-acclimated animals in response to methacholine. Morphologically, the secretory cells from the salt glands of SW-acclimated animals were larger than those of FW-acclimated animals. In addition, there were significantly more mitochondria per unit volume in secretory tissue from SW-acclimated animals. The results from this study demonstrate that the salt glands of C. porosus are phenotypically plastic, both morphologically and functionally and acclimate to changes in environmental salinity.

Ups and downs of intestinal function with prolonged fasting during aestivation in the burrowing frog, Cyclorana alboguttata

SUMMARY Although green striped burrowing frogs (Cyclorana alboguttata) experience large reductions in the mass and absorptive surface area of the small intestine (SI) during aestivation, little is known about how this may affect the functional capacity of the SI. We examined changes in the function (l-proline uptake rate and capacity) and metabolism of the SI (in vitro oxygen consumption, Na+/K+-ATPase activity and abundance) of C. alboguttata following 6 months of aestivation. l-Proline uptake rate was significantly higher in aestivating frogs, but overall uptake capacity was lower than in active frogs. Total SI oxygen consumption rate (VO2) was also lower in aestivating frogs, despite no difference in mass-specific V̇O2. The proportion of intestinal V̇O2 associated with Na+/K+-ATPase activity and protein synthesis was equivalent between active and aestivating frogs, suggesting these processes were unaffected by aestivation. Indeed, the activity of Na+/K+-ATPase transporters in the SI of aestivating frogs was not different from that of active animals. Aestivating frogs maintained Na+/K+-ATPase activity, despite experiencing a reduction in the density of Na+/K+-ATPase transporters, by increasing the molecular activity of the remaining pumps to 2–3 times that of active frogs. These results show that functionality of the SI is maintained at the cellular level, potentially facilitating the reclamation of nutrients from the intestinal lumen while in aestivation. Despite this, the functional capacity of the SI in aestivating C. alboguttata is significantly reduced due to a reduction in tissue mass, helping frogs to conserve energy while in aestivation.

Damage to the gills and integument of Litoria fallax larvae (Amphibia: Anura) associated with ionoregulatory disturbance at low pH

In fish, exposure to waters of low pH causes significant damage to the gill resulting in fatal iono- and osmoregulatory disturbance. In amphibians, exposure to acid waters also disrupts ionic homeostasis, however the extent and nature of injuries to amphibian larvae from acid exposure are poorly understood. Changes in gross morphology and ultrastructure of the gills and integument were examined, together with measures of Na(+) efflux/uptake, in larval Litoria fallax (Amphibia: Anura) following acute acid exposure. Examination of tissues revealed significant changes in morphology and ultrastructure of both gills and the integument following acutely lethal exposure to low pH water. Changes to the gills of acid-exposed L. fallax larvae included lifting of the branchial epithelium and opening of tight junctions between pavement cells (with a consequent reduction in tight junction length). Damage to epithelial cell-cell tight junctions was also apparent at the integument along with widespread oncosis and localised epithelial necrosis. Mucous secretory activity at the gills and body surface was largely unaffected by acid exposure, with little or no difference in density, cross-sectional area and number of epithelial mucous secretory vesicles in acid-exposed and control larvae. Changes in morphology and ultrastructure at low pH were accompanied by significant Na(+) loss (up to 50% of the total body Na(+) content) attributable in large part to increased paracellular ionic efflux across the gills as well as increased transcellular and paracellular efflux of ions across the integument.

Further observations of visual signalling in Australo-Papuan hylid frogs of the genus Litoria (Tschudi)

Intra-specific communication in anuran amphibians is primarily achieved with vocal signals (Duellman and Trueb 1994). However, some species also use visual displays for interand/ or intra-sexual communication (Hodl and Amezquita 2001). Such displays include movement of the fore and/ or hindlimbs (e.g. hand-waving, foot-flagging and/or legflicking), movement of the body or posturing, and exhibition of a brightly coloured vocal sac (Hodl and Amezquita 2001; Hirschmann and Hodl 2006). Visual signalling is hypothesised to be an alternative or complementary mode of communication in anurans used under certain environmental and/or social conditions (e.g. in noisy or visually complex environments) (Amezquita and Hodl 2004).

Long-range movement in the rare Cooloola sedgefrog Litoria cooloolensis

Understanding habitat usage is essential for the proper management of rare and threatened species in the wild. However, current knowledge of habitat usage by many rare and threatened Australian frog species is inadequate in this regard. Knowledge of non-breeding habitat usage in Australian amphibian species is particularly poor (Hines et al. 1999), with current understanding of the habitat requirements of many species based largely on habitat usage by calling animals during the breeding season, when fiogs are more readily detectable. However, like other fauna, the use of different habitats by amphibians can vary daily and seasonally as well as between different sexes and life stages (Law and Dickman 1998). Female frogs, for example, are likely to occupy habitat further away fiom a breeding water body while males tend to stay near the breeding site (Bartelt et al. 2004; Johnson et al. 2007; Rittenhouse and Semlitsch 2007). A majority of aquatic breeding amphibians will also undergo embryonic and larval development within the aquatic environment while spending their adult and juvenile lives in the terrestrial environment, unlike developing larvae, postmetamorphic adult and juvenile amphibians are not restricted to the aquatic body of their birth (Johnson et al 2007) and thus have the ability to disperse or migrate into adjacent, non-breeding, areas for the purposes of foraging, overwintering (Regosin et al 2003) or refuge use (Semlitsch and Bodie 2003).