We investigated the toxicity of copper, zinc and cadmium to the following taxa: copepods Tigriopus angulatus (Lang) and Harpacticus sp. (Order Harpacticoida, Family Harpacticidae); flatworm Obrimoposthia ohlini (Bergendal) (Order Seriata, Family Procerodidae); bivalve Gaimardia trapesina (Lamarck) (Order Veneroida, Family Gaimardiidae); sea cucumber Pseudopsolus macquariensis (Dendy) (Order Dendrochirotida, Family Cucumriidae); sea star Anasterias directa (Koeler) (Order Forcipulatida, Family Asteriidae).
Sites chosen for the collection of invertebrates for this study were free of obvious signs of metal contamination, as verified by the analysis of seawater samples from collection sites by inductively coupled plasma optical emission spectrometry (ICP-OES).
Six invertebrate species were selected for toxicity tests to represent a range of taxa and ecological niches. Individuals of the copepod Tigriopus angulatus were collected using fine mesh dip nets from rock pools high on the intertidal zone. Individuals of the flatworm Obrimoposthia ohlini were collected from the undersides of boulders, high in the intertidal zone. The copepod Harpacticus sp. and bivalve Gaimardia trapesina were collected from several macroalgae species at high energy locations in the intertidal zone. Individuals of the sea cucumber Pseudopsolus macquariensis were collected from rocks from high energy locations from the intertidal to subtidal zones. Juveniles of the sea star Anasterias directa were collected from rocks in deep pools, low in the intertidal zone.
All experimental tests using O. ohlini, T. angulatus, P. macquariensis and A. directa were conducted at the AAD Kingston laboratories, while some tests with Harpacticus sp. and all tests with G. trapesina were conducted in the laboratory facilities on Macquarie Island. Adult life-stages were tested for all species except for P. macquairensis and A. directa in which juvenile stages were tested. Psedopsolus macquariensis released eggs in the aquarium which developed into juveniles prior to being used in tests, and juvenile A. directa were collected from the field.
Each test involved exposure to copper, zinc or cadmium solution under a static non-renewal test regime over 14 days. Five metal concentrations plus a control were used for each test, with 3-5 replicates of each concentration. Where possible, tests were replicated. Concentrations used in replicate tests sometimes varied, as species sensitivity information accrued in tests was used to optimise subsequent tests. Metal test solutions in seawater were prepared 24 hours prior to the addition of animals, using 500 micrograms/L CuSO4, 500 micrograms/L ZnCl2 and 500 micrograms/L Cd SO4 MilliQ stock solutions. Seawater was filtered to 0.45 microns and water quality parameters were measured using a TPS 90-FL multimeter at the start and end of tests. Dissolved oxygen (DO) was greater than 80% saturation, salinity 35 ppt plus or minus 0.5, and pH was ~8.1-8.3 at the start of tests. All experimental vials and glassware were acid washed with 10% nitric acid and rinsed with MilliQ three times before use. Metal concentrations were determined using ICP-OES; samples of test solutions were taken at the start (day 0) and end of tests (day 14), filtered through a 0.45 microns syringe filter and acidified with 1% ultra-pure nitric acid. Measured concentrations at the start of tests were within 96% of nominal concentrations. In order to estimate exposure concentrations, the measured concentrations at days 0 and 14 were averaged.
Tests were conducted in lidded plastic vials of varying sizes, depending on the size and number of individuals in the test. For both copepod species, there were 10 individuals per 50 mL in 70 mL vials; for P. macquariensis there were 8 individuals per 50 mL in 70 mL vials; and for O. ohlini, A. directa and G. trapesina, 10 individuals per 100 mL in 120 mL vials. Tests were conducted under a light-dark regime (at 2360 lux) of 18:6h light:dark in summer, 12:12 for tests for the rest of the year. Tests were kept in controlled temperature cabinets set at 6 degrees C, and temperatures within cabinets were monitored throughout the test using data loggers.
Vials were checked daily and survival recorded on days 1, 2, 4, 7, 10 and 14. Individuals were considered dead, and removed from test vials, when for G. trapesina adductor muscles no longer closed shell; O. ohlini were inactive and covered in mucous; P. macquariensis and A. directa tube feet were no longer moving; T. angulatus and Harpacticus sp. urosomes were perpendicular to prosomes.
Data are provided in a series of excel workbooks; one workbook per test species.
