Metabarcoding free-living marine nematodes using curated 18S and CO1 reference sequence databases for species-level taxonomic assignments

Ecology and Evolution
Lara MacheriotouSofie Derycke

Abstract

High-throughput sequencing has the potential to describe biological communities with high efficiency yet comprehensive assessment of diversity with species-level resolution remains one of the most challenging aspects of metabarcoding studies. We investigated the utility of curated ribosomal and mitochondrial nematode reference sequence databases for determining phylum-specific species-level clustering thresholds. We compiled 438 ribosomal and 290 mitochondrial sequences which identified 99% and 94% as the species delineation clustering threshold, respectively. These thresholds were evaluated in HTS data from mock communities containing 39 nematode species as well as environmental samples from Vietnam. We compared the taxonomic description of the mocks generated by two read-merging and two clustering algorithms and the cluster-free Dada2 pipeline. Taxonomic assignment with the RDP classifier was assessed under different training sets. Our results showed that 36/39 mock nematode species were identified across the molecular markers (18S: 32, JB2: 19, JB3: 21) in UClust_ref OTUs at their respective clustering thresholds, outperforming UParse_denovo and the commonly used 97% similarity. Dada2 generated the most realistic number of A...Continue Reading

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Citations

Jun 12, 2020·Genome Génome / Conseil National De Recherches Canada·Thomas O PowersKirsten S Powers
Jul 16, 2020·Revista Do Instituto De Medicina Tropical De São Paulo·Gessica Baptista de MeloRonaldo Cesar Borges Gryschek
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Jan 19, 2021·Genome Génome / Conseil National De Recherches Canada·Leonardo Tresoldi GonçalvesCláudia Calegaro-Marques
Apr 28, 2021·Molecular Ecology Resources·Jose S HleapMelania E Cristescu
Jul 9, 2021·Molecular Ecology Resources·Zachary GoldPaul H Barber
Aug 23, 2021·Parasites & Vectors·Marie Louise DaveyFrode Fossøy

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Methods Mentioned

BETA
Assay
PCR
electrophoresis
density gradient centrifugation

Software Mentioned

ExCaliBar
Mega7
UParse
RDP
Geneious®
GraphPad Prism
Qiime2
denovo
MegaX
GraphPad

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