Drop it all: Extraction-free detection of non-indigenous marine species through optimized direct-droplet digital PCR

Molecular biosecurity surveillance programs increasingly use environmental DNA (eDNA) for detecting marine non-indigenous species (NIS). However, the current molecular detection workflow is cumbersome, prone to errors and delays, and thereby can hinder management efforts and restrict the “opportunity window” for a rapid response to new marine NIS incursions.
Here, we describe the first method to use direct digital droplet PCR (direct-ddPCR) to detect species-specific free-floating extra-cellular eDNA (free-eDNA) signals, i.e. detection of species-specific eDNA without the need for filtration or DNA extraction, with seawater sample. This first proof-of-concept aquarium study was conducted with three distinct marine NIS: the Mediterranean fanworm Sabella spallanzanii, the ascidian clubbed tunicate Styela clava, and the brown bryozoan Bugula neritina to evaluate the detectability of free-eDNA in seawater. The detectability of targeted free-eDNA was assessed by directly analysing aquarium m...