Monitoring offshore subtidal marine biodiversity in the Eastern Shore Islands Area of Interest using eDNA Metabarcoding

Environmental DNA metabarcoding is being applied as a biodiversity monitoring tool in marine conservation areas by the department of Fisheries and Oceans. Between 2021 to 2023, offshore surface and benthic eDNA samplers were collected in the Eastern Shore Islands Area of Interest in Nova Scotia to establish baseline fish and other eukaryote diversity data for monitoring potential changes in species richness and community structure over time. In each of the three years of this project, we collected water samples by a 6-bottle CTD rosette sampler deployed from the Canadian Coast Guard ship M. Perley along four transects with four stations in each transect (n=16 sites). These four transects extended from inshore to offshore and were spread across the AOI boundary to cover a longitudinal gradient. Water samples were filtered onto a 0.22um Sterivex filter (2021 and 2022) or a 1.2um Smith-Root filter (2023) to collect eDNA. eDNA was then extracted in a laboratory, and libraries were created to target fish (12S MiFishU marker in all 3 years, 16S in 2021) and general biodiversity (COI using Leray or Leray-XT primers) in 2022 and 2023. Samples from 2021 and 2022 were sequenced on an Illumina NovaSeq platform at the Centre for Environmental Genomics Applications (CEGA; St. John's, Newfoundland), while samples from 2023 were sequenced on an Illumina MiSeq at the Aquatic Biotechnology Lab (ABL) in Dartmouth, Nova Scotia. Samples were analyzed through the QIIME2 pipeline and denoised with DADA2. Amplicon sequence variant taxonomy was assigned using BLAST for 12S and 16S markers, while taxonomy for COI was assigned using an RDP Bayesian classifier. The overall objective for this study was to create baseline species richness and community data entirely from eDNA samples, and being a time series for detecting change or stability over time. Since our methods in the field and lab changed between 2021 to 2023, any temporal trends detected between years should be interpreted with caution.