Caged amphibian tadpoles and in situ genotoxicity monitoring of aquatic environments with the alkaline single cell gel electrophoresis (comet) assay

Abstract

In previous studies we demonstrated that indigenous amphibian tadpoles are suitable organisms for monitoring small bodies of water (e.g., creeks, ponds, and drainage ditches) using the alkaline single cell gel electrophoresis (SCG) or `comet' assay. This approach involves detection, under alkaline conditions, of cell DNA fragments which on electrophoresis migrate from the nuclear core, resulting in a `comet with tail' formation. However, although often plentiful, tadpoles are not present in all aquatic environments. Both larger bodies of water (e.g., lakes and rivers) and those impacted upon heavily by man (e.g., bodies of water near industrial sites, on landfills, and in urban areas) often do not support amphibian tadpole populations. An alternative approach to the collection of indigenous tadpoles is to place caged tadpoles at these sites for short term exposures to environmental contaminants. To determine the feasibility of such an approach, Rana clamitans (green frog) and Bufo americanus (American toad) tadpoles were housed in cages at 11 sites in southwestern Ontario (Canada). In a preliminary experiment, we found that tadpoles caged at a polluted reference site (Tallgrass Prairie Heritage Park in Windsor, Ontario) for either 7 or 14 days showed significant ( P<0.05) increases in DNA damage, relative to tadpoles caged in the laboratory in dechlorinated water. As a result we routinely used a 7 day exposure time. Significantly ( P<0.05) increased levels of DNA damage, relative to their controls, were observed in tadpoles caged at three sites along two creeks draining a large petrochemical installation south of Sarnia, Ontario; at two sites in the Tallgrass Prairie Heritage Park; and at a site along the Ecarte Channel which is part of the St. Clair River. The DNA damage levels of animals caged in Lake St. Clair, in the Trenton Channel of the Detroit River, at a landfill site, and in two creeks in the city of Windsor did not differ significantly ( P>0.05) from their controls. This study demonstrates that caged tadpoles are suitable for monitoring most bodies of fresh water, particularly those aquatic habitats mentioned above where indigenous tadpoles are not present. A combined approach of collecting indigenous tadpoles and using caged tadpoles should provide a sensitive system for aquatic genotoxicity monitoring.