In the context of the sixth wave of extinction, reliable surveys of biodiversity are increasingly needed to infer the cause and consequences of species and community declines, identify early warning indicators of tipping points, and provide reliable impact assessments before engaging in activities with potential environmental hazards. DNA metabarcoding has emerged as having potential to provide speedy assessment of community structure from environmental samples. Here we tested the reliability of metabarcoding by comparing morphological and molecular inventories of invertebrate communities associated with seagrasses through estimates of alpha and beta diversity, as well as the identification of the most abundant taxa. Sediment samples were collected from six Zostera marina seagrass meadows across Brittany, France. Metabarcoding surveys were performed using both mitochondrial (Cytochrome Oxidase I) and nuclear (small subunit 18S ribosomal RNA) markers, and compared to morphological inventories compiled by a long-term benthic monitoring network. A sampling strategy was defined to enhance performance and accuracy of results by preventing the dominance of larger animals, boosting statistical support through replicates, and using two genes to compensate for taxonomic biases. Molecular barcodes proved powerful by revealing a remarkable level of diversity that vastly exceeded the morphological survey, while both surveys identified congruent differentiation of the meadows. However, despite the addition of individual barcodes of common species into taxonomic reference databases, the retrieval of only 36% of these species suggest that the remaining were either not present in the molecular samples or not detected by the molecular screening. This finding exemplifies the necessity of comprehensive and well-curated taxonomic reference libraries and multi-gene surveys. Overall, results offer methodological guidelines and support for metabarcoding as a powerful and repeatable method of characterizing communities, while also presenting suggestions for improvement, including implementation of pilot studies prior to performing full “blind” metabarcoding assessments to optimize sampling and amplification protocols.

The West Antarctic Peninsula (WAP) is the fastest warming region in Antarctica where climate impact on the cold-adapted marine ecosystem is already visible. To monitor faunal changes in remote vast bodies of Antarctic waters, efficient and informative tools are essential. High-throughput sequencing of environmental DNA (eDNA) has emerged as one such tool for monitoring biodiversity and ecosystems, as it increases detection sensitivity of taxa, and sampling is often simpler and less costly than traditional collection methods. We collected water samples from four WAP shallow (≤300m) shelf regions, recovered the eDNA therein, and performed metagenomic shotgun sequencing and analyses to determine the effectiveness of this method to assess marine benthic faunal diversity; this includes the detection of deep-water predatory king crabs whose potential shoreward expansion to warming shelves has sparked much concern. Using a customized bioinformatics pipeline, we identified abundant signatures of common benthic invertebrate fauna, endemic notothenioid fishes, as well as lithodid king crabs. We also uncovered species richness and diversity comparable to biological inventories compiled by the use of traditional survey methods, supporting the efficacy of the eDNA shotgun sequencing approach. As the rate of eDNA degradation affects faunal detection sensitivity, we also quantified mitochondrial ND2 gene copies in eDNA derived from a WAP icefish and found ND2 copies persisted to at least 20days in the cold WAP water, much longer than values reported for temperate environments. We propose that eDNA metagenomic sequencing complements traditional sampling, and combining both will enable more inclusive biodiversity detection and faunal change monitoring in the vast Southern Ocean.

Native crayfish species often face competition and displacement by non-indigenous invasive crayfishes. Management responses implemented to preserve imperiled crayfishes may include the construction of physical barriers to prevent the spread of invasive crayfishes, and movement of native populations to “ark” sites that have not yet been invaded. These strategies require ongoing monitoring to determine their effectiveness. We propose that environmental DNA (eDNA), genetic material identified from environmental samples, can be useful for assessing advancing invasions and imperiled freshwater species associated with management interventions. We monitored a series of management interventions intending to isolate the endangered Shasta crayfish Pacifastacus fortis (Faxon, 1914) from the invasive signal crayfish Pacifastacus leniusculus (Dana, 1852) in California, USA. We successfully detected P. fortis eDNA from two sites where it was known to occur, one site where its presence was uncertain, and one site (near an “ark” site) where it was believed absent. We also detected P. leniusculus eDNA from five sites it was known to occupy, but failed to detect its eDNA at two sites where it was believed to occur. We conclude with recommendations for improved eDNA monitoring of crayfish conservation and management interventions in the future.

Deep ocean hydrothermal vent ecosystems face physical disturbances from naturally occurring volcanic and tectonic activities and are at increasing risk of mineral resource exploitation, raising concerns about the resilience of endemic biological communities. Following destructive events, efficient and rapidly applicable surveys of organisms are required to monitor the state, evolution and a possible return of these ecosystems to their original baseline status. In this study, we explored the environmental DNA (eDNA) approach as a tool (1) to assess biodiversity of benthic communities associated with deep-sea hard substrata and (2) tracked the recolonization dynamics of benthic invertebrate communities living on the Montségur edifice within the Lucky Strike vent field (Mid-Atlantic Ridge), after an induced disturbance that consisted of faunal clearance within experimental quadrats. Hard substratum samples were collected prior to and one year after the disturbance, for eDNA metabarcoding using one marker of the mitochondrial Cytochrome Oxidase I (COI) gene and three markers of the nuclear 18S ribosomal RNA (18S) gene. We also generated a DNA barcoding inventory that consisted of taxa physically collected from Montségur and morphologically identified. This inventory contained amplified barcodes from COI, 18S and the nuclear large subunit ribosomal RNA (28S) gene. The resulting sequence information from the COI and 18S were used for eDNA taxonomic assignment. The eDNA datasets uncovered a high diversity of metazoan OTUs, which included macro- and meiofauna common to Lucky Strike. Baseline data collected at the start of the experiment identified higher OTU richness at sites peripheral to the active edifice, as well as at inactive sites. One year following the initial disturbance, analysis of recolonization data found no statistical difference in presence/absence from baseline communities. The eDNA protocols provide a reproducible strategy to quickly assess biodiversity associated with deep sea hard substratum, enabling comparisons across various habitats. To follow recolonization dynamics at small spatial scales, however, we recommend an approach that uses both molecular and morphological-based traditional methods. Finally, we present original data on the “unseen” diversity of the fauna inhabiting the poorly studied inactive sites, locations that are targeted by commercial mining. Continued monitoring of these sites is currently ongoing and will bring new insight on recovery potential over time, with the ultimate goal of informing conservation and management decisions in relation to the protection of hydrothermal vent ecosystems.