Environmental DNA Quantitative PCR Assays for Marine Species Detection

<p>University College Dublin (UCD) was tasked with developing quantitative (q)PCR assays for marine species to assess the possibility to detect presence of target species in environmental water samples. Marine eenvironmental (e)DNA is released by organisms into the water in the form of mucus, gametes, faeces or dead tissue as cells or extracellular material. To access marine eDNA, water samples are collected by CTD or other means and subsequently filtered. DNA is then extracted from filter and interrogated for the presence of target species through species specific eDNA assays. UCD has previously developed a range of species specific eDNA assays using Minor Grove Binding (MGB) probes for freshwater and marine species. The target species selected in the ATLAS project range from hydrothermal endemic shrimp (Mirocaris fortunata), cold-water coral (Lophelia pertusa), deep-sea fish (orange roughy and blackbelly rosefish) and pelagic fish (bigeye tuna). To date assays have been developed and validated for the hydrothermal vent shrimp and the deep sea fishes. Further development is underway for the pelagic fish and the cold-water coral. These assays are currently being deployed for field validation on water samples from the North Atlantic. Development of eDNA assays for Lophelia and bigeye tuna has been hampered by lack of suitable mitochondrial (mt)DNA regions in the Barcode of Life Gene (mtDNA COI) for anchoring primers and probes. Further investigations are ongoing to identify suitable areas for eDNA assays in alternative mtDNA regions for these two species (sequence provided by IFREMER for Lophelia pertusa and other mtDNA regions for bigeye tuna) to enable assay development and future deployment.