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Drop it all: extraction-free detection of targeted marine species through optimized direct droplet digital PCR

Authors :
Michelle Scriver
Ulla von Ammon
Cody Youngbull
Xavier Pochon
Jo-Ann L. Stanton
Neil J. Gemmell
Anastasija Zaiko
Source :
PeerJ, Vol 12, p e16969 (2024)
Publication Year :
2024
Publisher :
PeerJ Inc., 2024.

Abstract

Molecular biomonitoring programs increasingly use environmental DNA (eDNA) for detecting targeted species such as marine non-indigenous species (NIS) or endangered species. However, the current molecular detection workflow is cumbersome and time-demanding, and thereby can hinder management efforts and restrict the “opportunity window” for rapid management responses. Here, we describe a direct droplet digital PCR (direct-ddPCR) approach 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 samples. This first proof-of-concept aquarium study was conducted with three distinct marine species: 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 marine water samples using an optimized species-specific ddPCR assay. The results demonstrated the consistent detection of S. spallanzanii and B. neritina free-eDNA when these organisms were present in high abundance. Once organisms were removed, the free-eDNA signal exponentially declined, noting that free-eDNA persisted between 24–72 h. Results indicate that organism biomass, specimen characteristics (e.g., stress and viability), and species-specific biological differences may influence free-eDNA detectability. This study represents the first step in assessing the feasibility of direct-ddPCR technology for the detection of marine species. Our results provide information that could aid in the development of new technology, such as a field development of ddPCR systems, which could allow for automated continuous monitoring of targeted marine species, enabling point-of-need detection and rapid management responses.

Details

Language :
English
ISSN :
21678359
Volume :
12
Database :
Directory of Open Access Journals
Journal :
PeerJ
Publication Type :
Academic Journal
Accession number :
edsdoj.5bb4e056f97a45ad93e8893fd0fef289
Document Type :
article
Full Text :
https://doi.org/10.7717/peerj.16969