Australian Institute of Marine Science, PMB no. 3, Townsville MC, Qld 4810, Australia Department of Marine Benthic Ecology and Evolution, Centre of Ecological and Evolutionary Studies, Biological Centre, Centre of Ecological and Evolutionary Studies, Biological Centre, University of Groningen, PO Box 14, 9750 AA Haren, The Netherlands.
Mol Ecol Resour. 2009 Jan;9(1):74-82. doi: 10.1111/j.1755-0998.2008.02222.x.
Understanding the flexibility of the endosymbioses between scleractinian corals and single-cell algae of the genus Symbiodinium will provide valuable insights into the future of coral reefs. Here, a real-time polymerase chain reaction (PCR) assay is presented to accurately determine the cell densities of Symbiodinium clades C and D in the scleractinian coral Acropora millepora, which can be extended to other coral-symbiont associations in the future. The assay targets single- to low-copy genes of the actin family of both the coral host and algal symbiont. Symbiont densities are expressed as the ratio of Symbiodinium cells to each host cell (S/H ratio, error within 30%), but can also be normalized to coral surface area. Greater accuracy in estimating ratios of associations involving multiple clades is achieved compared with previous real-time PCR assays based on high-copy ribosomal DNA loci (error within an order of magnitude). Healthy adult A. millepora containing ~1.4 × 10(6) zooxanthellae per cm(2) (as determined by haemocytometer counts) had S/H ratios of c. 0.15, i.e. ~15 symbiont cells per 100 host cells. In severely bleached colonies, this ratio decreased to less than 0.005. Because of its capacity to accurately determine both densities and ratios of multiple symbionts within one sample, the assay will open the door for novel research into the mechanisms of symbiont shuffling and switching.
理解硬珊瑚与单细胞藻类属共生体之间的灵活性Symbiodinium 将为珊瑚礁的未来提供有价值的见解。在这里，提出了一种实时聚合酶链反应 (PCR) 检测方法，可准确确定硬珊瑚 Acropora millepora 中共生体 clade C 和 D 的细胞密度，该方法将来可扩展到其他珊瑚共生体的关联中。该检测方法针对珊瑚宿主和藻类共生体的肌动蛋白家族的单拷贝到低拷贝基因。共生体密度表示共生体细胞与每个宿主细胞的比值（S/H 比值，误差在 30% 以内），但也可以归一化为珊瑚表面积。与以前基于高拷贝核糖体 DNA 基因座的实时 PCR 检测方法相比，该方法在估计涉及多个 clade 的关联的比值时具有更高的准确性（误差在数量级内）。健康的成年 A. millepora 每平方厘米含有约 1.4×10(6) 个虫黄藻（通过血球计数器计数确定），S/H 比值约为 0.15，即每 100 个宿主细胞中有 15 个共生体细胞。在严重白化的菌落中，该比值下降到低于 0.005。由于其能够准确地确定一个样品中多个共生体的密度和比值，该检测方法将为研究共生体 shuffling 和切换的机制开辟新的研究途径。
