Assessing biodiversity of remote deep-sea chemosynthetic environments: a case study comparing morphological and metabarcoding analysis of seep macrofauna from the U.S. Cascadia margin

Anthropogenic disturbances are drastically affecting marine biodiversity across diverse ecosystems, including the deep sea. However, the relative inaccessibility of the deep sea poses difficulties in assessing the biodiversity of vulnerable habitats such as chemosynthetic environments. Chemosynthetic cold seeps host distinct macrofaunal communities adapted to living in harsh environmental conditions and contribute significantly to productivity and nutrient cycling of the barren seafloor. Traditionally, biodiversity of deep-sea macrofauna has been assessed via morphological approaches, in which sediment samples are collected and animals sorted and identified—a time-consuming process that relies heavily on taxonomic expertise. Technological advancements in eDNA metabarcoding may now provide a rapid means for the analysis and monitoring of these remote habitats. However, few studies have compared metabarcoding methods to traditional approaches and baseline genetic data are needed prior to its use in biomonitoring. The vulnerability of dense macrofaunal communities in seeps make them an ideal habitat for evaluating metabarcoding’s effectiveness in estimating taxa abundance and diversity. Sediment macrofaunal communities were examined in core samples collected via ROV from cold seeps in Astoria Canyon, located on the Cascadia margin off the coasts of Oregon and Washington, USA. For traditional morphology-based analyses, cores were preserved, sieved, and sorted, and macrofauna were identified to the lowest possible taxonomic level. Additional sediment cores were collected, preserved, and processed for eDNA metabarcoding methods; this processing included isolating the fauna via Ludox density gradient separation, followed by DNA extraction, amplification of the 18S rRNA gene, and sequencing (Illumina NextSeq 2x300 bp). Here we compare the community metrics (e.g., abundance estimates, taxa richness, Shannon diversity index, community structure) resulting from both approaches, while also describing the macrofaunal communities across various depths and seep habitats of Astoria Canyon. The role of environmental variables (e.g., organic carbon, total nitrogen, C:N ratios, δ13C and δ15N stable isotopes, redox potential, and grain size) in shaping these communities was examined for both methodologies. These findings will inform future experimental designs and biomonitoring efforts for macrofaunal communities vulnerable to human-induced disturbances, while also broadening the knowledge of biodiversity in these deep-sea cold seeps.