Integration of genotoxicity and population genetic analyses in kangaroo rats (Dipodomys merriami) exposed to radionuclide contamination at the Nevada Test Site, USA.

We examined effects of radionuclide exposure at two atomic blast sites on kangaroo rats (Dipodomys merriami) at the Nevada Test Site, Nevada, USA, using genotoxicity and population genetic analyses. We assessed chromosome damage by micronucleus and flow cytometric assays and genetic variation by randomly amplified polymorphic DNA (RAPD) and mitochondrial DNA (mtDNA) analyses. The RAPD analysis showed no population structure, but mtDNA exhibited differentiation among and within populations. Genotoxicity effects were not observed when all individuals were analyzed. However, individuals with mtDNA haplotypes unique to the contaminated sites had greater chromosomal damage than contaminated-site individuals with haplotypes shared with reference sites. When interpopulation comparisons used individuals with unique haplotypes, one contaminated site had greater levels of chromosome damage than one or both of the reference sites. We hypothesize that shared-haplotype individuals are potential migrants and that unique-haplotye individuals are potential long-term residents. A parsimony approach was used to estimate the minimum number of migration events necessary to explain the haplotype distributions on a phylogenetic tree. The observed predominance of migration events into the contaminated sites supported our migration hypothesis. We conclude the atomic blast sites are ecological sinks and that immigration masks the genotoxic effects of radiation on the resident populations.

The slider turtle as an environmental sentinel: multiple tissue assays using flow cytometric analysis.

We used flow cytometry (FCM) to conduct a multiple-tissue assay on slider turtles (Trachemys scripta) inhabiting radioactive seepage basins. Duplicate samples of blood, heart, spleen and kidney were analysed on two different cytometers (Leitz MPV and Coulter Profile II), each employing distinct staining protocols (DAPI and PI, respectively). Both DAPI and PI assays of spleen cells demonstrated significantly greater variation in DNA content for the basin turtles than for 'control' animals from nearby, uncontaminated sites. Basin turtles also exhibited significant cell-cycle effects for blood and spleen, again revealed by both assays. These corroborative findings demonstrate the consistency and repeatability of FCM assays in environmental monitoring and identify the particularly sensitive nature of turtle blood and spleen to mutagenic agents. Our survey complements previous FCM studies on sliders from contaminated sites and thereby underscores the species' potential as a sentinel for biomarker assays.

Flow cytometry for monitoring contaminant exposure in black-crowned night-herons.

The flow cytometry methods (FCM) was employed to determine cellular DNA content of black-crowned night-heron (Nycticorax nycticorax) embryos and 10-day-old chicks collected at sites differing in types of chemical contamination. The coefficient of variation of DNA content (CV) in blood collected from embryos suggested cytogenetic damage at a side in Louisiana known to be contaminated with petroleum. Blood CV from chicks suggested genetic damage at a site in Texas also known to be contaminated with petroleum. Spleen CVs in chicks were significantly lower than respective means from the reference site. The CVs of chick blood and liver and spleen negatively correlated, suggesting recovery of spleen and liver cells after exposure to a clastogenic compound. Thus, the lower CVs may also have been indicative of genetic damage. Based on the findings of this study, FCM is a potential indicator of certain environmental contaminants in black-crowned night-herons.

The application of bioassays in risk assessment of environmental pollution.

Increased contamination of the environment by toxic chemicals has resulted in the need for sensitive assays to be used in risk assessment of polluted sites. Traditional tests are useful to detect and measure concentrations of chemicals in the environment and in tissues. However, physicochemical assays possess deficiencies that impair their use in evaluating complex environmental contamination. We have developed cytogenetic procedures, including chromosomal, micronucleus, and flow cytometric assays, to assess the mutagenic damage of petrochemicals and low-level radioactivity on indigenous terrestrial and aquatic wildlife populations. These procedures are sensitive to the perturbation of DNA that results from exposure to mutagenic contaminants in both field and laboratory studies. The use of natural populations of animals in biomonitoring, combined with traditional chemical assays, will ultimately provide sufficient information to estimate the risk to human health and environmental quality from anthropogenic pollution.

Sequential expression of biomarkers in Bluegill Sunfish exposed to contaminated sediment.

The temporal expression of various biological rsponses was determined in Bluegill SunfishLepomis macrochirus exposed under controlled laboratory conditions to sediment containing high concentrations of polynuclear aromatic hydrocarbons, polychlorinated biphenyls and heavy metals. Liver, gill, blood, kidney, brain, spleen and intestine were removed from Sunfish sampled at 1, 2, 4, 8, 16, and 40 weeks post-exposure. Biomarker data were recorded for specific proteins, enzymatic activities, DNA integrity, and histopathology. Biomarkers in the laboratory exposed fish were similar to those of indigenous Sunfish sampled from the site of origin of the contaminated sediment. Several patterns of development of biomarkers over time were also evident. For example, the responses of certain biomarkers are not time-dependent (i.e., intestine and gill ATPase activities) while that of others, such as brain ATPase activity, liver cytochrome P450 and NADPH content, stress proteins, chromatin proteins and DNA strand breaks, fluctuate over time. Still other biomarkers, such as EROD activity, zinc protoporphyrin content of the blood, and DNA adducts, showed marked increases over time. Such patterns need to be considered when comparing laboratory and field results and deciding which biomarkers to use for biomonitoring programs. Implications for natural selection and population/community level responses are also discussed.