Alkaline single-cell gel (comet) assay and genotoxicity monitoring using two species of tadpoles

✍ Scribed by Steven Ralph; Michael Petras; Raj Pandrangi; Maria Vrzoc

• Publisher: John Wiley and Sons
• Year: 1996
• Tongue: English
• Weight: 923 KB
• Volume: 28
• Category: Article
• ISSN: 0893-6692
• DOI: 10.1002/(sici)1098-2280(1996)28:2<112::aid-em7>3.0.co;2-h

✦ Synopsis

Small bodies of water (e.g., creeks, ponds, and drainage ditches) have received very little attention in genotoxicity studies, yet these areas are important because they are often the first to be affected by industrial effluents, sewage contaminants, accidental spills, internal combustion engine emissions, landfill runoffs, and pesticide uses. To address this deficiency, we examined erythrocytes in two species of tadpoles, Rana clamitans and Bufo americanus, using the alkaline singlecell gel (SCG) ("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. Exposure of R. clamitans tadpoles to a range of concentrations of methyl methanesulfonate (MMS) prcduced a linear increase in DNA length to DNA core width ratios. This is consistent with findings in a number of other species. Timedose experiments using MMS suggest that the peak level of DNA damage in R. clamitans tadpoles occurred 42 hr after exposure. B. americanus tadpoles exposed to 6.25 mg/l of MMS for 12 hours had a significant increase in DNA damage over that seen in the controls. Freshly caught R. clamitans tadpoles from Highgate and B. americanus tadpoles from Duart, both on the north shore of lake Erie, gave ratios of 2.78 and 2.07, respectively. This region of Ontario is a prime agricultural area and pesticide use is extensive. Tadpoles from Highgate and Duart, maintained in the laboratory for 4 months and 6 weeks, respectively, gave ratios of 1.29 and 1.44. The results of the SCG procedure in tadpoles indicate that this assay is extremely sensitive and suitable for detecting genotoxicity in the environment.