The core of the matter – Importance of identification method and biological replication for benthic marine monitoring

Benthic macrofauna are important and widely used biological indicators of marine ecosystems as they have limited mobility and therefore integrate the effects of local environmental stressors over time. Recently, environmental DNA (eDNA) analysis has provided a potentially more resource-efficient approach for benthic biomonitoring than traditional morphology-based methods. Several studies have compared eDNA with morphology-based monitoring, but few have compared the two approaches using the exact same sediment cores. In addition, the meiofauna and pelagic organisms obtained as “bycatch” using eDNA have largely been disregarded from comparisons. Here, we address these shortcomings through comparative invertebrate analyses from six sediment sample replicates from each of four stations in Denmark, using eDNA metabarcoding and morphological identification. Our results revealed large variation between the six replicates for both methods and little overlap in taxon compositions between methods. While the morphological dataset was dominated by molluscs and annelids, the eDNA dataset was dominated by arthropods and annelids. Using community composition data, we found that sampling stations could be distinguished both with eDNA and with morphology. Finally, we evaluated expected total richness inferred from extrapolated accumulation curves of detected taxa from each method. This indicated that eDNA metabarcoding requires less replication than morphology for maximum coverage of diversity to be reached. However, both methods required high levels of replication, and our results on taxonomic composition add to the evidence that morphological and eDNA-based methods should preferably be used as complimentary tools for marine bioassessment.

Methods
This dataset represents environmental DNA metabarcoding data from sediment samples from Danish waters (collected in March/April 2019). Four sites were sampled (Samso, Hjelm, Ven and Karrebaeksminde (KBM)), and six samples were collected at each site (24 samples total, excluding controls). These eDNA samples were collected in connection with a morphological survey of benthic macrofauna carried out by the Danish Environmental Protection Agency. See connected publication for additional details.

DNA has been amplified using the forward primer mlCOIintF-XT (5′-GGWACWRGWTGRACWITITAYCCYCC-3′) and the reverse primer jgHCO2198 (5′-TAIACYTCIGGRTGICCRAARAAYCA-3′), which together amplify ~313 bp of the mitochondrial COI gene. The four libraries sequenced (SF1_5 - SF1_8) represent the four PCR replicates carried out in this study. The libraries were sequenced using paired-end NovaSeq 6000 sequencing (250 bp PE). Tags are consistent across libraries.