A field-ready molecular workflow for sample-to-result detection of the harmful dinoflagellate species Prorocentrum cordatum in coastal waters

Abstract

The globally distributed dinoflagellate Prorocentrum cordatum contributes to harmful algal blooms, posing serious risks to marine ecosystems and coastal economies. Conventional monitoring of toxic phytoplankton is time-consuming, requires large sample volumes, and depends on expert taxonomic identification, highlighting the need for faster, more sensitive and widely accessible methods. This study presents a sample-to-result molecular assay for detection P. cordatum using polymerase chain reaction (PCR) and loop-mediated isothermal amplification (LAMP) targeting the ITS and psbA genes. Analyses were performed on small volumes of water (5-50 mL), concentrated with syringe filters and membranes. Sensitivity was evaluated through 10-fold serial dilutions of P. cordatum DNA, and specificity tested against three other potentially harmful dinoflagellates, P. lima, P. hoffmannianum, and Takayama acrotrocha. Detection limits were 5.3 x 102 cells L-1 for ITS-PCR and 5.3 x 103 cells L-1 for psbA-PCR; and 5.3 x 10 and 5.3 x 10 cells L-1 for ITS-LAMP and psbA-LAMP, respectively. In field tests (n = 48, all < 106 cells L-1), P. cordatum was detected in 32 samples by ITS-PCR and 29 by psbA-PCR. By contrast, ITS-LAMP detected the species in only three samples, and psbA-LAMP in none. Overall, PCR proved highly sensitive and specific for detecting low cell densities, while LAMP, although less sensitive, offers a rapid and portable option for field monitoring during moderate-to-high bloom conditions. Highlights* P. cordatum was isolated for molecular assays development. * Biomass concentration was achieved using small volumes. * PCR and LAMP assays targeting ITS and psbA were validated.