Abstract Background Saxitoxin (SXT) can cause the paralytic shellfish poisoning (PSP) with severe impacts on both human health and marine ecosystems. Dinoflagellates are the major marine producers of STX and other variety of secondary toxic metabolites that can gather in molluscs becoming a risk for consumers’ poisoning. The aim of this work was to investigate the presence of the main algal species producing SXT in an area intended for molluscs breeding. Methods During 2020-2023, 40 seawater samples were collected from two macro-areas in the Adriatic Sea (North-Western) at different distances from the coast. After water filtration, DNA was extracted (DNeasy PLANT Mini Kit) and analysed with Real Time PCR for Alexandrium minutum and Gymnodinium catenatum detection. The possible ability to produce SXT was confirmed by sxt4A gene detection. Molecular results were compared with the microscopy gold-standard method. Results A. minutum was detected in 62.5% (CI 95% 0.47-0.75) of the samples, with 84% (CI 95% 0.65-0.94) of them possibly producing SXT. G. catenatum was found in 95% (CI 95% 0.83-0.99) of samples, with 73.7% (CI 95% 0.58-0.85) of them presenting STX gene. By microscopy, only Alexandrium spp. were identified without successful species identification. No other saxitoxin-producing genera or species were observed. Conclusions Our results underlined that A. minutum and G. catenatum can play an important role in the production of SXT, confirming their harmful presence in the areas intended for molluscs breeding. Microscopic identification can lead to an underestimation of the presence of potentially toxic algal species, stressing the relevance of improving molecular tools also in this field. These findings highlighted the importance of reporting and monitoring toxic algae presence, given the saxitoxin impact for food safety, and its significance for public health. Key messages • Saxitoxin-producing algae detection through molecular methods in molluscs breeding areas may represent an early warning useful to improve food safety and human health. • Harmful algal monitoring can improve water management in a One Health perspective.