Toxicity of copper, cadmium and zinc to Macquarie Island marine invertebrates 2012/15

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. 2018-09-24 - The original datasheet was reformatted to fit IPT Biodiversity.AQ standard. The new datasheet "toxicityTestMarineOrganisms.csv" provides the datasetID, occurrenceID, country, countryCode, decimalLatitude, decimaLongitude, island, localitionID, eventDate, year, month, day, identificationQualifier, occurrenceStatus, and basisOfRecord. The lowest taxonomical rank of the species identified that could be determined is provided, after matched in WoRMS (World Register of Marine Species).

本研究探究了铜、锌、镉对以下分类群的毒性：桡足类（copepods）角状猛水蚤（Tigriopus angulatus (Lang)）及猛水蚤属未定种（Harpacticus sp.，隶属于猛水蚤目Harpacticoida、猛水蚤科Harpacticidae）；扁形涡虫Obrimoposthia ohlini (Bergendal)（隶属于单肠目Seriata、原口涡虫科Procerodidae）；双壳类Gaimardia trapesina (Lamarck)（隶属于帘蛤目Veneroida、Gaimardiidae科）；海参Pseudopsolus macquariensis (Dendy)（隶属于枝手目Dendrochirotida、海参科Cucumriidae）；海星Anasterias directa (Koeler)（隶属于显带目Forcipulatida、海星科Asteriidae）。
本研究选取的无脊椎动物采集点位未出现明显金属污染迹象，该结论通过电感耦合等离子体发射光谱法（inductively coupled plasma optical emission spectrometry，ICP-OES）对采集点位的海水样本进行分析得以验证。本研究共选取6种无脊椎动物开展毒性试验，以覆盖多样的分类群与生态位。
角状猛水蚤（Tigriopus angulatus）个体采用细网抄网从潮间带高处的岩池采集；扁形涡虫Obrimoposthia ohlini个体采自潮间带高处的石块底面；猛水蚤属未定种（Harpacticus sp.）及双壳类Gaimardia trapesina采自潮间带高能区域的多种大型藻类之上；海参Pseudopsolus macquariensis个体采自潮间带至亚潮带高能区域的岩石表面；海星Anasterias directa幼体采自潮间带低处深池的石块表面。
针对扁形涡虫O. ohlini、角状猛水蚤T. angulatus、海参P. macquariensis及海星A. directa的所有试验均在AAD Kingston实验室开展；而猛水蚤属未定种Harpacticus sp.的部分试验及双壳类G. trapesina的全部试验则在麦夸里岛的实验室设施中进行。本试验对除P. macquariensis与A. directa外的所有物种采用成体阶段开展测试，后两者则使用幼体阶段。其中，海参P. macquariensis会在水族箱中产卵，卵发育为幼体后再用于试验；海星A. directa幼体直接采自野外。
每项试验均采用静态不更新试验体系，将受试生物暴露于铜、锌或镉溶液中，试验周期为14天。每项试验设置5个金属浓度梯度及1个对照组，每个浓度设置3~5个重复组；在条件允许的情况下，尽可能开展重复试验。由于试验过程中积累的物种敏感性数据可用于优化后续试验，因此重复试验中使用的浓度有时会有所调整。
金属受试溶液以海水为溶剂，在加入受试生物前24小时配制完成，使用的储备液分别为500 μg/L的硫酸铜（CuSO4）、500 μg/L的氯化锌（ZnCl2）及500 μg/L的硫酸镉（CdSO4），均以MilliQ水配制。海水经0.45 μm过滤后使用，试验开始与结束时均采用TPS 90-FL型多参数水质仪测定水质参数。试验初始阶段，溶解氧（dissolved oxygen，DO）饱和度大于80%，盐度为35±0.5 ppt，pH值介于8.1~8.3之间。
所有试验用瓶及玻璃器皿均经10%硝酸浸泡酸洗后，用MilliQ水冲洗3次方可使用。采用ICP-OES测定受试溶液中的金属浓度：分别在试验初始（第0天）与结束（第14天）采集受试溶液样本，经0.45 μm针式过滤器过滤后，加入1%超纯硝酸进行酸化。试验初始阶段的实测浓度与标称浓度的偏差不超过4%（即实测值为标称值的96%以上）。为估算实际暴露浓度，取第0天与第14天的实测浓度的平均值作为最终暴露浓度。
试验采用带盖塑料瓶进行，瓶具尺寸根据受试生物个体大小与试验个体数量进行选择。两种桡足类的试验体系为：70 mL试验瓶中加入50 mL溶液，每瓶放置10个个体；海参P. macquariensis的试验体系为：70 mL试验瓶中加入50 mL溶液，每瓶放置8个个体；扁形涡虫O. ohlini、海星A. directa及双壳类G. trapesina的试验体系为：120 mL试验瓶中加入100 mL溶液，每瓶放置10个个体。试验采用光暗循环培养：夏季为18小时光照:6小时黑暗，光照强度为2360 lux；其余季节为12小时光照:12小时黑暗。所有试验均置于设定为6℃的控温培养箱中进行，培养箱内的温度全程采用数据记录仪进行监测。
试验瓶每日进行检查，分别在第1、2、4、7、10和14天记录受试生物的存活情况。死亡个体的判定标准如下：双壳类G. trapesina的闭壳肌不再闭合贝壳；扁形涡虫O. ohlini失去活动能力且体表覆盖粘液；海参P. macquariensis与海星A. directa的管足停止运动；角状猛水蚤T. angulatus及猛水蚤属未定种Harpacticus sp.的尾体（urosomes）与头胸部（prosomes）呈垂直状态。
本数据集以一系列Excel工作簿文件提供，每个受试物种对应一个工作簿。2018-09-24：原始数据表已重新格式化，以符合IPT Biodiversity.AQ标准。新数据表"toxicityTestMarineOrganisms.csv"包含以下字段：datasetID、occurrenceID、country、countryCode、decimalLatitude、decimalLongitude、island、locationID、eventDate、year、month、day、identificationQualifier、occurrenceStatus及basisOfRecord。本数据集通过与世界海洋物种登记册（World Register of Marine Species，WoRMS）进行匹配后，提供可确定的最低分类阶元的物种鉴定信息。