Smith Kirsty F, de Salas Miguel, Adamson Janet, Rhodes Lesley L
Cawthron Institute, 98 Halifax Street East, Private Bag 2, Nelson 7042, New Zealand.
Tasmanian Herbarium, Tasmanian Museum and Art Gallery, Private Bag 4, Hobart, Tasmania 7001, Australia.
Mar Drugs. 2014 Mar 7;12(3):1361-76. doi: 10.3390/md12031361.
The identification of toxin-producing dinoflagellates for monitoring programmes and bio-compound discovery requires considerable taxonomic expertise. It can also be difficult to morphologically differentiate toxic and non-toxic species or strains. Various molecular methods have been used for dinoflagellate identification and detection, and this study describes the development of eight real-time polymerase chain reaction (PCR) assays targeting the large subunit ribosomal RNA (LSU rRNA) gene of species from the genera Gymnodinium, Karenia, Karlodinium, and Takayama. Assays proved to be highly specific and sensitive, and the assay for G. catenatum was further developed for quantification in response to a bloom in Manukau Harbour, New Zealand. The assay estimated cell densities from environmental samples as low as 0.07 cells per PCR reaction, which equated to three cells per litre. This assay not only enabled conclusive species identification but also detected the presence of cells below the limit of detection for light microscopy. This study demonstrates the usefulness of real-time PCR as a sensitive and rapid molecular technique for the detection and quantification of micro-algae from environmental samples.
为监测计划和生物化合物发现而鉴定产毒素的甲藻需要相当多的分类学专业知识。从形态上区分有毒和无毒的物种或菌株也可能很困难。各种分子方法已被用于甲藻的鉴定和检测，本研究描述了针对裸甲藻属、卡伦藻属、卡罗藻属和高山藻属物种的大亚基核糖体RNA（LSU rRNA）基因开发的八种实时聚合酶链反应（PCR）检测方法。检测方法被证明具有高度特异性和敏感性，并且针对链状裸甲藻的检测方法因应对新西兰马努考港的藻华而进一步开发用于定量分析。该检测方法估计环境样品中的细胞密度低至每个PCR反应0.07个细胞，相当于每升三个细胞。该检测方法不仅能够进行确定性的物种鉴定，还能检测到低于光学显微镜检测限的细胞的存在。本研究证明了实时PCR作为一种灵敏且快速的分子技术用于检测和定量环境样品中的微藻的实用性。
