Background: Toxicity Test Methods

Toxicity tests are used to determine whether the concentration of contaminants in water or sediments is biologically available and likely to produce harmful biological effects. Organisms are exposed to a test sample under controlled conditions and examined for lethal (mortality) and sublethal effects. Sublethal effects include reduced growth, development or reproduction. Ideally a toxicity test should use a local species, be relatively simple to perform and capable of providing answers quickly, and have sensitive endpoints that are easily interpreted and ecologically relevant.

Figure 1. Toxicity tests are usually conducted in beakers supplied with aeration and maintained at a constant temperature by using a water bath (pictured) or environmental chamber.

Sea Urchin Fertilization Test

The sea urchin fertilization test provides a rapid assessment of toxicity in aqueous samples. This test is conducted using the purple sea urchin (Stronglocentrotus purpuratus; Figure 2). Sea urchin sperm are exposed to the sample, and the ability of the sperm to fertilize the egg (Figure 3) is evaluated. The particular merits of this test are its short duration of exposure (40 minutes total), small sample size requirements (10 ml per replicate) and easily identifiable endpoint (presence of a fertilization membrane visible around the egg in Figure 3). The test is based on methods in the EPA’s Short-Term Methods for Estimating the Chronic Toxicity of Effluents and Receiving Waters to West Coast Marine and Estuarine Organisms (EPA/600/R-95/136).

Figure 2. Stronglocentrotus purpuratus, the purple urchin.

Figure 3. Fertilized Stronglocentrotus purpuratus egg.

Sea Urchin Development Test

This method measures toxicity of aqueous samples by the determining the ability of sea urchin embryos to develop normally within 72 hours (Figure 4). Like the fertilization test, the embryo development test can be conducted on small a volume which makes it useful for monitoring programs where the test sample often has to be shared among laboratories. Sea urchin embryo development has been extensively studied and the transparent embryo enables the researcher to detect abnormalities in the skeletal and digestive system. The test is based on methods in the EPA’s Short-Term Methods for Estimating the Chronic Toxicity of Effluents and Receiving Waters to West Coast Marine and Estuarine Organisms (EPA/600/R-95/136).

Figure 4. Normal Stronglocentrotus purpuratus embryo.

Mussel Embryo Development Test

This test measures the effect of aqueous samples on the ability of mussel (Mytilus galloprovincialis) embryos (Figure 5) to develop normally with in 48 hours. The test endpoint of normal embryo development is easily identifiable as a “D” shape (Figure 5). Mussels in spawning condition can be collected from the field or purchased from aquaculture companies, which allows this test to be conducted at any time of the year. The test is based on methods in the EPA’s Short-Term Methods for Estimating the Chronic Toxicity of Effluents and Receiving Waters to West Coast Marine and Estuarine Organisms (EPA/600/R-95/136).

Figure 5. Normal (left) and abnormal Mytilus galloprovincialis embryos (right).

Polychaete Growth Test

The polychaete growth test is a sublethal test that measures growth and survival in the marine worm Neanthes arenaceodentata (Figure 6). The test utilizes juvenile worms (≤7 days old) which are exposed to sediment for 28 days. This is one of the few standardized marine tests available to measure long-term toxicity. The 28 day N. arenaceodentata test was developed by the US Army Engineer Research and Development Center (Bridges and Farrar 1997, Bridges, Farrar et al. 1997).

Figure 6. Marine worm Neanthes arenaceodentata.

Amphipod Survival Test

Amphipods like Eohaustorius estuarius, (Figure 7) are the most common organisms used to measure sediment toxicity. In these tests, the mortality of animals exposed to test sediments for 10 days is compared with the mortality of animals exposed to control sediment. Increased mortality indicates toxicity. The amphipod survival test is the most widely used method used to assess toxicity in regional monitoring and dredged material characterization programs. Longer term exposures with some amphipod species can be conducted to measure growth and reproductive impairment. Ten-day survival tests with Eohaustorius estuarius are commonly conducted using USEPA testing procedures (1994).

Figure 7. Eohaustorius estuarius.

Sediment Bioaccumulation Tests

Bioaccumulation tests measure the accumulation of contaminants from sediment into benthic infauna. The data from these tests can also indicate the potential for contaminants to bioconcentrate in species occupying higher levels in the food chain. These tests tend to be the longest in duration; exposures are typically a minimum of 28 days. At the end of the exposure, the tissues from the animals are removed and analyzed for metals and trace organic compounds. Bioaccumulation tests usually do not provide a sensitive measure of sediment toxicity because the species used are selected to be resistant to contaminant exposure so that the survivors can be used for chemical analysis. The marine worm, Nereis virens (Figure 8), and the marine clam, Macoma nasuta (Figure 9), are commonly utilized.

Figure 8. Nereis virens.

Figure 9. Macoma nasuta.

References

Bridges, T. S. and J. D. Farrar (1997). "The influence of worm age, duration of exposure and endpoint selection on bioassay sensitivity for Neanthes arenaceodentata (Annelida: Polychaeta)." Environmental Toxicology and Chemistry 16(8): 1650-1658.

Bridges, T. S., J. D. Farrar, et al. (1997). "The influence of food ration on sediment toxicity in Neanthes arenaceodentata (Annelida: Polychaeta)." Environmental Toxicology and Chemistry 16(8): 1659-1665.

USEPA (1994). Methods for assessing the toxicity of sediment-associated contaminants with estuarine and marine amphipods. Narragansett, RI, Office of Research and Development, U.S. Environmental Protection Agency: 140.

USEPA (1995). Short-term methods for estimating the chronic toxicity of effluents and receiving waters to west coast marine and estuarine organisms. G. A. Chapman, D. L. Denton and J. M. Lazorchak. Cincinnati, OH, Office of Research and Development: 661.

For more information on Background: Toxicity Test Methods, contact Steve Bay at steveb@sccwrp.org (714) 755-3204.

This page was last updated on: 1/21/2010