Development of a novel TaqMan qPCR assay for rapid detection and quantification of Gymnodinium catenatum for application to harmful algal bloom monitoring in coastal areas of Tunisia

Abstract

Gymnodinium catenatum is a dinoflagellate known to cause paralytic shellfish poisoning (PSP), commonly associated with human muscular paralysis, neurological symptoms, and, in extreme cases, death. In the present work, we developed a real-time PCR-based assay for the rapid detection of the toxic microalgal species, G. catenatum, in environmental bivalve mollusc samples as well as seawater samples. G. catenatum-specific primers and probe were designed on the ITS1-5.8S-ITS2 rDNA region. Hydrolysis probe qPCR assay was optimized. ITS1-5.8S-ITS2 rDNA region copy numbers per G. catenatum cell genome were estimated to be 122.73 ± 5.54 copies/cell, allowing cell quantification. The application of the optimized qPCR assay for G. catenatum detection and quantification in field samples has been conducted, revealing high sensitivity (detection of around 1.310⁵ cells/L of seawater samples. Thus, the designed hydrolysis probe qPCR assay could be considered an efficient tool for phytoplankton monitoring whilst ensuring accuracy and sensitivity and providing cost and time savings.

Keywords: Environmental samples; Gymnodinium catenatum; Harmful algae; Hydrolysis probe; ITS1-5.8S-ITS2 region; Quantification; qPCR assay.