Marine benthic metabarcoding:Anthropogenic effects on benthic diversity from shore to deep sea; assessed by metabarcoding and traditional taxonomy

Marien benthische biodiversiteit, de diversiteit aan soorten die in de zeebodem leven, is van grote waarde voor het mariene ecosysteem. Echter, deze benthische biodiversiteit wordt bedreigd door anthropogene invloeden zoals habitatsveranderingen, vervuiling en (over-)bevissing. Om te kunnen anticiperen op zulke invloeden is het belangrijk om verandering in de benthische biodiversiteit te kunnen meten. Traditionele methodes, gebaseerd op morphologische taxonomie zijn vaak tijd-rovend en vereist een uitgebreide taxonomische kennis. In het afgelopen decennium zijn moleculaire methodes, zoals DNA metabarcoding, ontwikkelend tot een alternatieve methode. In mijn proefschrift heb ik onderzoek gedaan naar de toepassing van zulke metabarcoding methodes om de mariene bentische biodiversiteit te analyseren en menselijke invloeden op dit ecosysteem te kunnen meten. De resultaten in mijn proefschrift laten zien dat de metabarcoding methode een waardevolle methode is om antropogene effecten op de benthische biodiversiteit te bepalen. Vergeleken met traditionele morfologische werden met de metabarcoding methode dezelfde conclusies getrokken omtrent het antropogene effect. Met de metabarcoding methode was het bovendien mogelijk om dit effect voor een grotere groep soorten te laten zien. Echter, de metabarcoding methode heeft ook een aantal nadelen ten opzichte van de traditionele morfologische methode. Zo is het bijvoorbeeld lastig om soortgroepen te kwantificeren op zowel abundantie als biomassa en kunnen door hiaten in de referentie databank niet altijd alle DNA-sequenties worden vertaald in een taxonomische groep. De methode ontwikkeld zich nog altijd erg snel, en deze snelle ontwikkelingen zullen hopelijk in de toekomst huidige limiteringen overkomen en de methode een nog waardevolle aanvulling in het werkveld maken

Quantification of marine benthic communities with metabarcoding

DNA metabarcoding methods have been implemented in studies aimed at detecting and quantifying marine benthic biodiversity. In such surveys, universal barcodes are amplified and sequenced from environmental DNA. To quantify biodiversity with DNA metabarcoding, a relation between the number of DNA sequences of a species and its biomass and/or the abundance is required. However, this relationship is complicated by many factors, and it is often unknown. In this study, we validate estimates of biomass and abundance from molecular approaches with those from the traditional morphological approach. Abundance and biomass were quantified from 126 samples of benthic intertidal mudflat using traditional morphological approaches and compared with frequency of occurrence and relative read abundance estimates from a molecular approach. A relationship between biomass and relative read abundance was found for two widely dispersed annelid taxa (Pygospio and Scoloplos). None of the other taxons, however, showed such a relationship. We discuss how quantification of abundance and biomass using molecular approaches are hampered by the ecology of DNA i.e. all the processes that determine the amount of DNA in the environment, including the ecology of the benthic species as well as the compositional nature of sequencing data

Diversity of Wadden Sea macrofauna and meiofauna communities highest in DNA from extractions preceded by cell lysis

Metabarcoding of genetic material in environmental samples has increasingly been employed as a means to assess biodiversity, also of marine benthic communities. Current protocols employed to extract DNA from benthic samples and subsequent bioinformatics pipelines differ considerably. The present study compares three commonly deployed metabarcoding approaches against a morphological approach to assess benthic biodiversity in an intertidal bay in the Dutch Wadden Sea. Environmental DNA was extracted using three different approaches; extraction of extracellular DNA, extraction preceded by cell lysis of a sieved fraction of the sediment, and extraction of DNA directly from small amounts of sediment. DNA extractions after lysis of sieved sediment fractions best recovered the macrofauna diversity whereas direct DNA extraction of small amounts of sediment best recovered the meiofauna diversity. Extractions of extracellular DNA yielded the lowest number of OTUs per sample and hence an incomplete view of benthic biodiversity. An assessment of different bioinformatic pipelines and parameters was conducted using a mock sample with a known species composition. The RDP classifier performed better than BLAST for taxonomic assignment of the samples in this study. Novel metabarcodes obtained from local specimens were added to the SILVA 18S rRNA database to improve taxonomic assignment. This study provides recommendations for a general metabarcoding protocol for marine benthic surveys in the Wadden Sea

Impact of an artificial structure on the benthic community composition in the southern North Sea:Assessed by a morphological and molecular approach

Man-made structures in the North Sea are known to act as artificial reefs by providing a habitat for sessile epifauna in a predominantly soft sediment environment. This epifauna is hypothesized to cast a so-called "shadow"over the soft sediment ecosystem by altering the nutrient composition in the overlying water column. In addition, the structure itself could alter currents and thereby influence the deposition and erosion of the sediments in the wake of the platform. This study aims to assess the long-Term effects of a gas platform in the southern North Sea on the surrounding benthic community by both morphological and molecular identification of benthic species. The species composition and a set of abiotic factors of the sediment around a gas platform were assessed along four transects. Differences for the abiotic factors were found in the closer vicinity of the platform in the direction corresponding to the predominant currents. The number of benthic fauna families found in the molecular approach were on average three times higher than for the morphological approach. Both approaches showed that small differences occurred primarily due to changes in sedimentary organic matter content. Differences in species composition were more pronounced between transects rather than between distances from the platform.</p

Erratum: Impact of an artificial structure on the benthic community composition in the southern North Sea: Assessed by a morphological and molecular approach

The following affiliation for Lise Klunder was not included in the earlier version of this article. This has now been added: Marine Evolution and Conservation, Groningen Institute of Evolutionary Life Sciences, University of Groningen, Nijenborgh 7, 9747 AG Groningen, The Netherlands.</p

A molecular approach to explore the background benthic fauna around a hydrothermal vent and their larvae:Implications for future mining of deep-sea SMS deposits

Seafloor massive sulfide (SMS) deposits are commonly found at hydrothermal vents and recently gained the special interest of mining industries. These deposits contain valuable metals and methods are currently developed to mine deep sea SMS deposits. However, excavation of SMS deposits potentially pose a threat to benthic life at the mining site itself, and also in the surrounding environment with plumes of suspended sediment and fine-grained SMS debris created during deep sea mining activities being highlighted as one of the major threats to deep-sea benthic fauna. The benthic communities surrounding the vents are, however, poorly known. As they are often exposed to natural plumes studying such communities could provide valuable information on their resilience toward mining related plumes. The Rainbow hydrothermal vent site at the Mid-Atlantic Ridge is a site characterized by one of the largest continuous natural plumes, which is found persisting over an extensive area. Sediment and water samples were taken both upstream and downstream of the Rainbow hydrothermal vent. Approximately 25 km away from the vent reference sites were samples as well. In addition to detecting the plume itself, concentrations of major and trace-metals in the sediments were used as tracers for long time sustained plume influence. At all sites, we assessed benthic species composition and detected larvae. Metabarcoding methods were used to determine species composition. Benthic species composition in the sediment was shown to differ between all locations and was highly influenced by the plume's fall out. Arthropoda were more dominant closer to the vent whereas Annelida and Nematoda were more dominant at the reference locations. Conservation and restoration of all these communities after a deep sea mining event will be difficult due to the spatial variation of these benthic communities