Although there is now a wealth of data on amphibian ecotoxicology, these data pertain to a relatively narrow range of taxa and ecological types. The atypical aquatic pipid Xenopus and temperate ranids and bufonids that have the common reproductive strategy of aquatic eggs and larvae account for a large percentage of the available information (Rowe et al., 2003). The Leptodactylidae, the largest family of anurans, have been neglected: McDiarmid and Mitchell (2000) list no leptodactylids among species that have been used in tests to evaluate water quality criteria chemicals, compared with 12 species of Bufo and 16 of Rana. Leptodactylids are also of interest in having diverse reproductive modes including aquatic and terrestrial foam nests and direct terrestrial development, and in being an important component of the fauna of the Neotropics, an area of particular conservation concern for amphibians (Young et al., 2004). Physalaemus pustulosus is the best-studied leptodactylid with classic work on many aspects of its ecology and behaviour (Ryan, 1985). Several characteristics of Physalaemus make it suitable for ecotoxicological work. It is widely distributed; abundant in savanna and modified savanna habitats, including agricultural and urban fringes; an explosive breeder during the rainy season, producing foam nests at the surface of shallow pools which are conspicuous and easily collected and contain a good sample size of a few hundred eggs. The eggs hatch in about 40 h so synchronous clutches from several pairs may be obtained easily from the field. The tadpoles are generalized benthic feeders (Kenny, 1969). Initial work showed that Physalaemus tadpoles may be reared at a density of 10/L in either still or flowing water systems, with better than 95% survival and 95% metamorphosis of survivors within 16 days of hatching. A soil substrate was the most important factor in successful rearing, achieving faster develeopment than even high temperature (Hailey et al., 2006). The soil substrate also mitigated a crowding effect, which otherwise reduced the growth performance of tadpoles reared at densities greater than 10/L; with soil there was little effect of density up to the highest values investigated (35 and 50/L) (Hailey et al., 2007). Nicole Sookoo paricipated in these experiments, peripheral to her own project, which also involved Asif Khan (who was registered for an M.Phil. project on amphibian diversity for a few months), and B.Sc. project students Devon Ramoo (2004) and Elena Hernandez (2005). Nicole's own research tested the susceptibility of Physalaemus eggs and tadpoles to three pesticides (Imidacloprid, Evisect, Fenobucarb) and three herbicides (Grammoxone, Roundup, Karmax) commonly used in Trinidad, particularly in sugar cane cultivation. Both acute toxicity at hatching and in older tadpoles, and developmental effects of chronic exposure in flowing water systems, were investigated. The second species studied was the cane toad Bufo marinus, which breeds in the dry season (January to April) and thus complements the wet-season breeder Physalaemus. The cane toad is also of interest in being more widely used in physiology and ecotoxicology, as are other Bufo species, and having a more typical reproductive strategy of aquatic eggs and larvae. The cane toad is also common in a variety of habitats, including agricultural and suburban, and is of sufficient body size to permit analysis of residues by tissue. Cane toad eggs are harder to collect from the wild, so adults were maintained in captivity and reproduction was induced by hormone injection (following the method of Reinier Mann and Mike Tyler in Adelaide). Tissue analysis for residues by GC-MS had to be abandoned due to problems with the equipment in the Department of Chemistry, an unfortunate circumstance as this was the part of the project of most interest to Nicole Sookoo to make use of her joint zoology/chemistry background.
Physalaemus pustulosus nests are collected from small tempoary pools, where the tadpoles are benthic feeders. Eggs hatch after two days, producing about 300 tadpoles per nest.
A soil substrate proved to be the most important factor in rearing Physalaemus successfully, either in still or flowing water systems. There was better than 95% survival and 95% metamorphosis of survivors within 16 days of hatching.
Acute toxicity is measured using nest fractions hatching directly into test solutions, and in older (Gosner stage 26-28) tadpoles. The latter are studied with (shown above) or without a soil substrate.
Cane toads are maintained in captivity and induced to spawn by subcutaneous injection of 10 μg LH-RH. Cane toad tadpoles will feed at the surface, unlike Physalaemus.
Hailey, A., Sookoo, N., Hernandez, E. & Ramoo, D. (2007). The influence of density and ration level on cultured Physalaemus pustulosus tadpoles, and the mitigation of a crowding effect by soil substrate. Appl. Herpetol. 4: 261-277.
Hailey, A., Sookoo, N., Mohammed, A. & Khan, A. (2006). Factors affecting tadpole growth: Development of a rearing system for the Neotropical leptodactylid Physalaemus pustulosus for ecotoxicological studies. Appl. Herpetol. 3: 111-128.
Kenny, J.S. (1969). The amphibia of Trinidad. Studies on the Fauna of Curacao and Other Caribbean Islands 29: 1-78.
McDiarmid, R.W., and Mitchell, J.C. (2000). Diversity and distribution of amphibians and reptiles. In: Ecotoxicology of Amphibians and Reptiles, p. 15-69. Sparling, D.W., Linder, G., and Bishop, C.A., Eds, Pensacoloa, FL, SETAC Press.
Rowe, C.L., Hopkins, W.A., and Bridges, C.M. (2003). Physiological ecology of amphibians in relation to susceptibility to natural and anthropogenic factors. In: Amphibian Decline: An Integrated Analysis of Multiple Stressor Effects, p. 9-57. Linder, G., Krest, S.K., and Sparling, D.W., Eds, Pensacoloa, FL, SETAC Press.
Ryan, M.J. (1985). The Túngara Frog: A Study in Sexual Selection and Communication. Chicago, University of Chicago Press.
Young, B.E., Stuart, S.N., Chanson, J.S., Cox, N.A., and Boucher, T.M. (2004): Disappearing Jewels: The Status of New World Amphibians. Arlington, VA, NatureServe.