Recombinase polymerase amplification combined with lateral flow dipstick for the rapid detection of Chattonella marina

Chattonella marina (Raphidophyceae) is a common microalgal species that has frequently caused harmful algal blooms (HABs). Here, recombinase polymerase amplification (RPA) was combined with a lateral flow dipstick (RPA-LFD) to establish a rapid detection method for this alga, aimed at providing an effective early warning of C. marina-forming HABs. A pair of specific RPA primers and an LFD probe was designed to target the internal transcribed spacer (ITS) sequence of C. marina. The specificity test with 20 control microalgal species indicated that the RPA-LFD was highly specific. Sensitivity test showed that the established RPA-LFD was 10 - 100 times more sensitive than conventional PCR. The detection limit of RPA-LFD was as low as 9.5 x 10(-1) ng mu L-1 for genomic DNA, 1.17 x 10(3) copies mu L-1 for recombinant plasmid containing the inserted ITS sequence, and 10 cells mL(-1) for the crude DNA extract of C. marina. In addition, the RPA-LFD was proved to be more accurate and sensitive than conventional PCR in the detection of target cells from natural samples, exhibiting its ability to be applied in the actual in situ detection scenarios. In conclusion, the developed RPA-LFD is rapid, specific, sensitive, and promising for being used in the point-of-care testing.