Detection of the Toxic Marine Dinoflagellate Alexandrium pseudogonyaulax in Small-Volume Seawater Samples Using a ddPCR Assay for Environmental DNA at the Southern Baltic Sea Coast

Abstract

Alexandrium pseudogonyaulax is an increasingly recurrent harmful algal bloom species in Northern European waters, making reliable detection, including of low-abundance populations, important for monitoring. This study evaluated droplet digital PCR (ddPCR) for detecting A. pseudogonyaulax in a standardised 1-L seawater sample and compared molecular measurements with microscopic cell enumeration. Weekly surface-water samples were collected at Heiligendamm, southern Baltic Sea, from June to September 2021 (18 samples). Environmental DNA was extracted after filtration of 1 L of seawater, and A. pseudogonyaulax 28S rRNA gene copy numbers (GCN) were quantified by ddPCR. Microscopic enumeration was performed from a 25-mL subsample, corresponding to an approximate detection limit of 40 cells L−1. Molecular and microscopic measurements showed pronounced temporal co-variation. Across the time series, 28S GCN ranged from 3.8 × 103 to 9.93 × 106 GCN L−1. On five sampling dates, microscopy was below its detection limit, whereas A. pseudogonyaulax DNA was consistently detected at 3.8 × 103–3.19 × 105 GCN L−1. Among samples with microscopically detectable cells, GCN and cell abundance showed a positive but non-significant association (Spearman’s ρ = 0.58, p = 0.077, n = 10). Field-derived GCN-per-cell estimates were lower than culture-derived values, indicating that a fixed conversion factor should not be applied to environmental samples. Overall, the results support ddPCR as a sensitive complement to microscopy and indicate that a standardised 1-L sample can support species-specific detection also at low abundance. Further validation across sites, seasons, and molecular targets is required before routine implementation.