NGS-based biodiversity and community structure analysis of meiofaunal eukaryotes in shell sand from Hållö island, Smögen, and soft mud from Gullmarn Fjord, Sweden

Aim: The aim of this study was to assess the biodiversity and community structure of Swedish meiofaunal eukaryotes using metabarcoding. To validate the reliability of the metabarcoding approach, we compare the taxonomic resolution obtained using the mitochondrial cytochrome oxidase 1 (COI) 'mini-barcode' and nuclear 18S small ribosomal subunit (18S) V1-V2 region, with traditional morphology-based identification of Xenacoelomorpha and Nematoda.

Location: 30 samples were analysed from two ecologically distinct locations along the west coast of Sweden. 18 replicate samples of coarse shell sand were collected along the north-eastern side of Hållö island near Smögen, while 12 replicate samples of soft mud were collected in the Gullmarn Fjord near Lysekil.

Methods: Meiofauna was extracted using flotation and siphoning methods. Both COI and 18S regions were amplified from total DNA samples using Metazoan specific primers and subsequently sequenced using Illumina MiSeq, producing in total 24 132 875 paired-end reads of 300 bp in length, of which 15 883 274 COI reads and 8 249 601 18S reads. These were quality filtered resulting in 7 954 017 COI sequences and 890 370 18S sequences, clustered into 2805 and 1472 representative OTUs respectively, yielding 190 metazoan OTUs for COI and 121 metazoan OTUs for 18S using a 97% sequence similarity threshold.

Results: The Metazoan fraction represents 7% of the total dataset for COI (190 OTUs) and 8% of sequences for 18S (121 OTUs). Annelida (30% of COI metazoan OTUs and 23.97% of 18S metazoan OTUs) and Arthropoda (27.37% of COI metazoan OTUs and 11.57% of 18S metazoan OTUs), were the most OTU rich phyla identified in all samples combined. As well as Annelida and Arthropoda, other OTU rich phyla represented in our samples include Mollusca, Platyhelminthes and Nematoda. In total, 213 COI OTUs and 243 18S OTUs were identified to species using a 97% sequence similarity threshold, revealing some non-native species and highlighting the potential of metabarcoding for biological recording. Taxonomic community composition shows as expected clear differentiation between the two habitat types (soft mud versus coarse shell sand), and diversity observed varies according to choice of meiofaunal sampling method and primer pair used.