Dataset Information

Divergence thresholds and divergent biodiversity estimates: can metabarcoding reliably describe zooplankton communities?

ABSTRACT: DNA metabarcoding is a promising method for describing communities and estimating biodiversity. This approach uses high-throughput sequencing of targeted markers to identify species in a complex sample. By convention, sequences are clustered at a predefined sequence divergence threshold (often 3%) into operational taxonomic units (OTUs) that serve as a proxy for species. However, variable levels of interspecific marker variation across taxonomic groups make clustering sequences from a phylogenetically diverse dataset into OTUs at a uniform threshold problematic. In this study, we use mock zooplankton communities to evaluate the accuracy of species richness estimates when following conventional protocols to cluster hypervariable sequences of the V4 region of the small subunit ribosomal RNA gene (18S) into OTUs. By including individually tagged single specimens and "populations" of various species in our communities, we examine the impact of intra- and interspecific diversity on OTU clustering. Communities consisting of single individuals per species generated a correspondence of 59-84% between OTU number and species richness at a 3% divergence threshold. However, when multiple individuals per species were included, the correspondence between OTU number and species richness dropped to 31-63%. Our results suggest that intraspecific variation in this marker can often exceed 3%, such that a single species does not always correspond to one OTU. We advocate the need to apply group-specific divergence thresholds when analyzing complex and taxonomically diverse communities, but also encourage the development of additional filtering steps that allow identification of artifactual rRNA gene sequences or pseudogenes that may generate spurious OTUs.

SUBMITTER: Brown EA

PROVIDER: S-EPMC4461424 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature

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Divergence thresholds and divergent biodiversity estimates: can metabarcoding reliably describe zooplankton communities?

Brown Emily A EA Chain Frédéric J J FJ Crease Teresa J TJ MacIsaac Hugh J HJ Cristescu Melania E ME

Ecology and evolution 20150513 11

DNA metabarcoding is a promising method for describing communities and estimating biodiversity. This approach uses high-throughput sequencing of targeted markers to identify species in a complex sample. By convention, sequences are clustered at a predefined sequence divergence threshold (often 3%) into operational taxonomic units (OTUs) that serve as a proxy for species. However, variable levels of interspecific marker variation across taxonomic groups make clustering sequences from a phylogenet ...[more]

PMID: 26078859

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Project description:For DNA metabarcoding to attain its potential as a community assessment tool, we need to better understand its performance versus traditional morphological identification and work to address any remaining performance gaps in incorporating DNA metabarcoding into community assessments. Using fragments of the 18S nuclear and 16S mitochondrial rRNA genes and two fragments of the mitochondrial COI marker, we examined the use of DNA metabarcoding and traditional morphological identification for understanding the diversity and composition of crustacean zooplankton at 42 sites across western Lake Superior. We identified 51 zooplankton taxa (genus or species, depending on the finest resolution of the taxon across all identification methods), of which 17 were identified using only morphological traits, 13 using only DNA and 21 using both methods. The taxa found using only DNA metabarcoding included four species and one genus-level identification not previously known to occur in Lake Superior, the presence of which still needs to be confirmed. A substantial portion of taxa that were identified to genus or species by morphological identification, but not identified using DNA metabarcoding, had zero ("no record") or ≤ 2 ("underrepresented records") reference barcodes in the BOLD or NCBI databases (63% for COI, 80% for 16S, 74% for 18S). The two COI marker fragments identified the most genus- and species-level taxa, whereas 18S was the only marker whose family-level percent sequence abundance patterns showed high correlation to composition patterns from morphological identification, based on a NMDS analysis of Bray-Curtis similarities. Multiple replicates were collected at a subset of sites and an occupancy analysis was performed, which indicated that rare taxa were more likely to be detected using DNA metabarcoding than traditional morphology. Our results support that DNA metabarcoding can augment morphological identification for estimating zooplankton diversity and composition of zooplankton over space and time, but may require use of multiple markers. Further addition of taxa to reference DNA databases will improve our ability to use DNA metabarcoding to identify zooplankton and other invertebrates in aquatic surveys.

Dataset Information

Divergence thresholds and divergent biodiversity estimates: can metabarcoding reliably describe zooplankton communities?

Publications

Divergence thresholds and divergent biodiversity estimates: can metabarcoding reliably describe zooplankton communities?

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