COI metabarcoding primer choice affects richness and recovery of indicator taxa in freshwater systems

Abstract

Mixed community or environmental DNA marker gene sequencing has become a commonly used technique for biodiversity analyses in freshwater systems. Many cytochrome c oxidase subunit I (COI) primer sets are now available for such work. The purpose of this study is to test whether COI primer choice affects the recovery of arthropod richness, beta diversity, and recovery of target assemblages in the benthos kick-net samples typically used in freshwater biomonitoring. We examine six commonly used COI primer sets on samples collected from six freshwater sites. Biodiversity analyses show that richness is sensitive to primer choice and the combined use of multiple COI amplicons recovers higher richness. Thus, to recover maximum richness, multiple primer sets should be used with COI metabarcoding. In ordination analyses based on community dissimilarity, samples consistently cluster by site regardless of amplicon choice or PCR replicate. Thus, for broadscale community analyses, overall beta diversity patterns are robust to COI marker choice. Recovery of traditional freshwater bioindicator assemblages such as Ephemeroptera, Trichoptera, Plectoptera, and Chironomidae as well as Arthropoda site indicators were differentially detected by each amplicon tested. This work will help future biodiversity and biomonitoring studies develop not just standardized, but optimized workflows that either maximize taxon-detection or the selection of amplicons for water quality or Arthropoda site indicators.

Zooplankton biogeographic boundaries in the California Current System as determined from metabarcoding

“…Banzai in general annotated taxa more specifically than USEARCH and generated an order of magnitude higher number of OTUs, discussed below). Similarly, choice of metabarcoding marker and primer set biases results towards different sets of taxa [24]. Here, results from two different genetic markers, 18S and COI, revealed the same overall trends but with varying sensitivities to the taxa detected and the specificity of taxonomic annotations permitted by the genetic marker for different groups.…”

Calibrating Environmental DNA Metabarcoding to Conventional Surveys for Measuring Fish Species Richness

“…However, multi-marker approaches do not guarantee better performance if reference databases are insufficiently populated or if primers impart amplification bias. The impact of these two factors on biodiversity inferences from eDNA metabarcoding has been explored (Hajibabaei et al, 2019;Kelly et al, 2019) but is inconsistently reported in field studies. Lecaudey et al (2019) detected only 23 of 43 fish species (53%) known to occur in their study area despite using three gene markers (cytb, 12S, 16S) in their analysis of Volga River eDNA samples.…”

Improved freshwater macroinvertebrate detection from eDNA through minimized non-target amplification

“…Thus, especially for indicator taxa, species or OTUs diversity was low. Li et al (2018) In the study by Hajibabaei et al (2019), which coined the term 'watered-down biodiversity', number of reads assigned to benthic invertebrates per eDNA sample was two orders of magnitude lower and diversity, measured as the number of exact sequence variants (ESVs) per sample, one order of magnitude lower compared to bulk samples. Pereira-da-Conceicoa et al (2019) showed that water samples from South African streams contained less than 10 % reads assigned to targeted macroinvertebrate taxa.…”