Prorocentrum lima (P. lima) is a widely spread dinoflagellate in the Mediterranean Sea and it has become increasingly involved in harmful algal blooms. The purpose of this study is to develop a probe-based real-time polymerase chain reaction (PCR) targeting the ITS1-5.8S-ITS2 region for the detection and absolute quantification of P. lima based on linear and circular DNA standards. The results have shown that the quantitative PCR (q-PCR), using circular plasmid as a template, gave a threshold cycle number 1.79–5.6 greater than equimolar linear standards. When microalgae, commonly found in aquatic samples were tested, no cross-amplification was observed. The q-PCR brought about a good intra and inter-run reproducibility and a detection limit of 5 copies of linear plasmid per reaction. A quantitative relationship between the cell numbers and their corresponding plasmid copy numbers was attained. Afterwards, the effectiveness of the developed protocol was tested with 130 aquatic samples taken from 19 Tunisian sampling sites. The developed q-PCR had a detection sensitivity of up to 1 cell. All the positive samples were taken from three sampling sites of Medenine Governorate with cell abundances that ranged from 22 to 156,000 cells L⁻¹ of seawater. The q-PCR assay revealed a high sensitivity in monitoring the aquatic samples in which the low concentrations of P. lima were not accurately detected by light microscopy. Indeed, this approach is at the same time rapid, specific and sensitive than the traditional microscopy techniques and it represents a great potential for the monitoring of P. lima blooms.

利马原螯虾（P. lima）是地中海中广泛分布的甲鞭毛藻，它已越来越多地参与有害的藻华。这项研究的目的是开发针对ITS1-5.8S-ITS2区域的基于探针的实时聚合酶链反应（PCR），用于检测和绝对定量利马毕赤酵母。基于线性和圆形DNA标准。结果表明，以环状质粒为模板的定量PCR（q-PCR）的阈值循环数比等摩尔线性标准物大1.79-5.6。对水生样品中常见的微藻进行测试时，未观察到交叉扩增。q-PCR具有良好的内部和内部运行重现性，每个反应的线性质粒检测限为5个拷贝。获得了细胞数目与其相应质粒拷贝数之间的定量关系。之后，对从19个突尼斯采样点采集的130个水生样品进行了测试，验证了开发方案的有效性。开发的q-PCR的检测灵敏度高达1个细胞。^-1的海水。q-PCR分析显示，在监测水生样品中具有很高的灵敏度，在这些样品中，光学显微镜无法准确检测出低浓度的利马疟原虫。的确，与传统的显微镜技术相比，该方法同时具有快速，特异性和灵敏性，在监测利马假单胞菌花期方面具有巨大的潜力。