The effects of sediments contaminated with antifoulant paint on coral larvae, coral recuitment, coral recruits and adult corals from the Great Barrier Reef

On 2 November 2000, the 184 m cargo ship, Bunga Teratai Satu, ran aground on Sudbury Reef, on the Great Barrier Reef and remained grounded for 12 days. The ship was re-floated only 3 days prior to the November 2000 mass coral spawning. No cargo or fuel was lost but the impact resulted in significant contamination of the reef with antifoulant paint containing tributyltin (TBT), copper (Cu) and zinc (Zn). Sediment was collected from the grounding scar site and a control site 1 km from the grounding site in a similar habitat zone. Sediments were ground and sieved and the 1-4 mm fraction retained. Sediments from the control site were used to dilute contaminated sediment to the concentrations required for experimental treatments.Larvae were raised from gametes released by adult Acropora microphthalma, which had been collected from Pelorus Island and spawned in the outdoor aquaria at the Orpheus Island Research Station on November 17 2000. Live crustose coralline algae (Hydrolithon sp.) was collected from Pelorus Island, ground and extracted with methanol to produce a crude inducer stock, which was used to initiate attachment, settlement and metamorphosis of larvae. The influence of contaminated sediment on larval survival and competency was examined by firstly exposing groups of 200, 7 day old larvae for 24 hours to one of 4 contaminated sediment treatments (0, 5, 25 and 100%). Each 500 ml container contained 0.4 g of experimental sediment in 400 ml of filtered seawater. Larvae were then transferred to well cell culture plates (10-20 per well) containing inducer and filtered sea water. After 24 hours, counts of larvae and newly metamorphosed polyps were made using a dissecting microscope. The effect of contaminated sediment upon larval recruitment was examined by exposing previously unexposed 8 day old larvae to 0, 5, 25 and 100% contaminated sediment treatments. Approximately 660 larvae were placed in each 10 l aquaria containing a pre-conditioned terracotta tile and 6 g of experimental sediment in 6 l of filtered seawater. Tiles were raised 2cm above the sediment to allow larvae to settle on the top and bottom of the tiles. Recruitment was scored on each tile after 40 h using a dissecting microscope.The effect of contaminated sediments on previously unexposed coral recruits was examined by firstly placing 16 pre-conditioned tiles in a 500 l outdoor aquarium holding 8 day old larvae at a density of approximately 100 larvae/l. Tiles were elevated 2 cm from the bottom and left for 3 days after which the tiles were removed and the number of recruits counted. Each tile was then placed into individual 10 l aquaria containing 5 l of filtered seawater and one of four contaminated sediment treatments (0, 0.5, 5 and 25%). The number of coral recruits were counted after 38 and 72 hours.Six colonies of Acropora formosa were collected from Magnetic Island and transferred to the outdoor aquaria at the Orpheus Island Research Station. Branchlets, 4-6 cm long were cut from the colonies, mounted on cylindrical holders and acclimatised for 10 days prior to experiments. Five branchlets were randomly placed in each of twenty five 10 l aquaria. Five aquaria were exposed to each of 4 contaminated sediment treatments (0, 5, 25 and 100%) and 5 were used as controls (no sediment). Branchlets were visually inspected every 24 or 48 hours and categorised according to the degree of bleaching that had occurred. Branchlet mortality was also recorded at each census point.The maximum effective quantum yield (Fv/Fm') of each illuminated branchlet was determined using a DIVING-PAM fluorometer. Fv/Fm' ratios were recorded at 15 random locations over the surface of each coral branchlet at 3 mm distance to the coral surface. Zooxanthellae density counts were undertaken on a random subset of experimental branchlets (eight per treatment).The concentrations of TBT, DBT, MBT, Cu and Zn in sediment and water samples from the larval competency experiments and the coral recruit and branchlet toxicity experiments were measured. This study was undertaken to test the effects of sediments contaminated with anti-foulant paint on survival, competency and recruitment of coral larvae, coral recruits and adult corals.

2000年11月2日，总长184米的货船“Bunga Teratai Satu”在大堡礁（Great Barrier Reef）的萨德伯里礁（Sudbury Reef）搁浅，且持续搁浅12天之久。该船直至2000年11月大规模珊瑚产卵事件前3天才得以重新浮起。本次事故未造成货物或燃油泄漏，但却导致防污漆中的三丁基锡（tributyltin, TBT）、铜（copper, Cu）与锌（zinc, Zn）对该礁体造成了严重污染。

研究人员从搁浅疤痕位点以及距离搁浅点1公里、生境相似的对照位点采集了沉积物样本。将沉积物研磨过筛后，保留1-4 mm粒径的组分。随后使用对照位点的沉积物对污染沉积物进行稀释，以配置实验所需的各浓度处理组。

实验用珊瑚幼虫由采集自佩洛鲁斯岛（Pelorus Island）的成年微小鹿角珊瑚（Acropora microphthalma）释放的配子培育而来，这些成体珊瑚于2000年11月17日在奥费斯岛研究站（Orpheus Island Research Station）的室外水族箱中产卵。研究人员同时从佩洛鲁斯岛采集了活的壳状珊瑚藻（crustose coralline algae, Hydrolithon sp.），将其研磨后用甲醇萃取得到粗诱导剂储备液，用于诱导幼虫附着、沉降与变态发育。

为探究污染沉积物对幼虫存活与发育能力的影响，研究人员将200只7日龄幼虫分为若干组，每组暴露于4种不同浓度（0、5、25和100%）的污染沉积物中，暴露时长为24小时。每个500 ml容器中包含400 ml过滤海水与0.4 g实验用沉积物。随后将幼虫转移至含有诱导剂与过滤海水的多孔细胞培养板中（每孔10-20只幼虫）。24小时后，使用解剖显微镜对幼虫与新近变态发育的水螅体进行计数。

为探究污染沉积物对幼虫招募的影响，研究人员将未经过污染暴露的8日龄幼虫暴露于0、5、25和100%浓度的污染沉积物处理组中。每个10 L水族箱中放置一块预调理的陶土瓦片，以及6 g实验用沉积物与6 L过滤海水，每箱放置约660只幼虫。将瓦片抬升于沉积物上方2 cm处，以使幼虫可在瓦片的上下表面附着。40小时后，使用解剖显微镜对每块瓦片上的幼虫招募情况进行评分。

为探究污染沉积物对未经过暴露的珊瑚幼体的影响，研究人员首先将16块预调理的瓦片放入一个500 L的室外水族箱中，该水族箱中幼虫密度约为100只/L，幼虫为8日龄。将瓦片抬升于水族箱底部上方2 cm处，放置3天后取出瓦片并计数幼体数量。随后将每块瓦片单独放入10 L水族箱中，每个水族箱包含5 L过滤海水与4种污染沉积物处理组之一（0、0.5、5和25%）。分别在38小时与72小时后对珊瑚幼体数量进行计数。

研究人员从磁岛（Magnetic Island）采集了6株鹿角珊瑚（Acropora formosa），将其转移至奥费斯岛研究站的室外水族箱中。从各株珊瑚上剪下4-6 cm长的分枝，固定于圆柱形支架上，在实验前驯化10天。将5根分枝随机放置于25个10 L水族箱中，其中5个水族箱分别对应0、5、25和100%的污染沉积物处理组，另设5个无沉积物的对照组。每24或48小时对分枝进行目视检查，并根据其白化程度进行分类，同时记录每次普查时的分枝死亡率。

使用DIVING-PAM荧光计（DIVING-PAM fluorometer）测定每根受光照分枝的最大有效量子产率（Fv/Fm'）。在每根珊瑚分枝表面随机选取15个位点（距珊瑚表面3 mm处）记录Fv/Fm'比值。从各处理组的实验分枝中随机抽取子集（每组8根）进行虫黄藻密度计数。

对幼虫发育能力实验、珊瑚幼体与分枝毒性实验中采集的沉积物与水样中的三丁基锡（TBT）、二丁基锡（DBT）、一丁基锡（MBT）、铜（Cu）与锌（Zn）浓度进行了测定。

本研究旨在检验防污漆污染沉积物对珊瑚幼虫存活、发育能力与招募，以及珊瑚幼体和成体珊瑚的影响。