Exposure of marine mammals to genotoxic environmental contaminants: Application of the P-32-postlabeling assay for measuring DNA-xenobiotic adducts

There is little information on exposure of marine mammals to genotoxic envi ronmental contaminants. The P-32-postlabeling assay has been successfully u sed to assess exposure to genotoxic polycyclic aromatic compounds in fish a nd humans. In the present study, a preliminary investigation showed that po lycyclic aromatic compound-like DNA adducts were present in hepatic tissues of harbor seals (Phoca vitulina richardsi) exposed to petroleum following the Exxon Valdez oil spill. However, for marine mammals, effects from chang es in tissue condition on DNA recovery and quality is of concern, because t issue samples are often collected from animals that have been dead for unkn own periods of time. To assess the effects of postmortem thermal history on DNA recovery from tissue and on DNA adduct quantitation, samples of harbor porpoise (Phocoena phocoena) hepatic tissue were incubated for up to 10 d at 4 and 30 degrees C. Only traces (<4 mu g) of hepatic DNA were recovered from 200 mg of tissue after incubation at 30 degrees C for 36 h. At 4 degre es C, DNA (50-130 mu g) was recovered from tissue incubated for up to 6 d; whereas DNA recovery at 10 d was minimal. Chromatograms of P-32-labeled DNA digests of liver tissue held at 4 and 30 degrees C and salmon sperm DNA he ld at 30 degrees C for 2 d had comparable profiles, suggesting that alterat ion of DNA bases had occurred during incubation of porpoise liver tissue. M oreover, the chromatograms of DNA extracted from liver tissues of harbor po rpoises caught incidentally in a northwest Atlantic fishery, packed in ice and sampled several days later also exhibited similar altered DNA structure s. Although, altered DNA structures that can interfere with the DNA adduct quantitation were present in autolyzed tissue, changes in the P-32-postlabe ling chromatography conditions can decrease the interference. Moreover, in a study with tissues taken from California sea lions (Zalophus californianu s) immediately postmortem and stored at -80 degrees C until processing, DNA structures associated with tissue breakdown were not observed. The DNA fro m sea lions, however, had putative age-dependent hepatic DNA modifications, which have a distinctive profile, and must be considered when evaluating e xposure of marine mammals to polycyclic aromatic compounds. Overall, the fi ndings showed that with attention to the postmortem thermal history of the tissue samples hepatic DNA adducts, as measured by P-32-postlabeling, have the potential to serve as a biological indicator of exposure of marine mamm als to environmental genotoxic compounds.