Application of real-time PCR assay for detection and quantification of Alexandrium tamarense and Alexandrium catenella cysts from marine sediments
Application of real-time PCR assay for detection and quantification of Alexandrium tamarense and Alexandrium catenella cysts from marine sediments
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DOI：
10.1016/j.hal.2006.12.004
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发表时间：
2007-04-01
期刊：
影响因子：
6.6
通讯作者：
Sako, Yoshihiko
中科院分区：
文献类型：
--
作者：
Kamikawa, Ryoma;Nagai, Satoshi;Sako, Yoshihiko
甲藻纲（Dinophyceae）的塔玛亚历山大藻（Alexandrum tamarense（Lebor）Balech）和链状亚历山大藻（Alexandrum catenella（Whedon and Kofoid）Balech）是麻痹性贝毒（paralytic shellfish poisoning，PSP）的主要病原。有必要识别A。tamarense和A.由于孢囊是游动营养细胞引起水华的种子群体，因此，为了减少PSP对经济和食品质量的危害，需要对悬链孢囊进行研究，并监测其在沉积物中的分布。在这项研究中，我们开发了一种有效的DNA提取方法，从天然孢囊存在于海洋沉积物中，他们的大小分级后，浮游生物网（网目尺寸为20-150 μ m）。将10-3000个孢囊添加到从有明海收集的沉积物中，并且樱草碱染色方法没有发现任何孢囊。然后从每个样品中提取DNA，并将A. tamarense和A.从实时PCR的Ct值与孢囊初始密度的对数之间的相关性获得链状孢囊。我们监测了A。tamarense和A.环境样品中的链状孢囊密度。这种方法被证明是一个强有力的工具，用于识别，检测和定量的有毒甲藻孢囊。(C)2007 Elsevier B. V.保留所有权利。
The dinoflagellates Alexandrium tamarense (Lebor) Balech and Alexandrium catenella (Whedon and Kofoid) Balech (Dinophyceae) are believed to be the main species responsible for paralytic shellfish poisoning (PSP) all over the world. It is necessary to identify A. tamarense and A. catenella cysts and to monitor their distribution in sediment in order to minimize the damages caused by PSP to the economy and food quality because cysts are the seed population for blooms caused by motile vegetative cells. In this study, we developed an efficient DNA extraction method from the natural cysts present in marine sediments after they were size fractionated with a plankton net (mesh size of 20-150 mu m). The 10-3000 cysts were added to the sediments collected from the Ariake Sea, and for which the primuline-staining method did not reveal any cysts. DNA was then extracted from each sample, and linear standard curves for A. tamarense and A. catenella cysts were obtained from the correlation between the Ct values by real-time PCR and the log of the initial densities of cysts. We monitored the A. tamarense and A. catenella cyst densities in the environmental samples. This assay was demonstrated to be a powerful tool for the identification, detection, and quantification of the cysts of the toxic dinoflagellates. (C) 2007 Elsevier B.V. All rights reserved.