As environmental DNA (eDNA) studies have grown in popularity for use in ecological applications, it has become clear that their results differ in significant ways from those of traditional, non-PCR-based surveys. In general, eDNA studies that rely on amplicon sequencing may detect hundreds of species present in a sampled environment, but the resulting species composition can be idiosyncratic, reflecting species’ true biomass abundances poorly or not at all. Here, we use a set of simulations to develop a mechanistic understanding of the processes leading to the kinds of results common in mixed-template PCR-based (metabarcoding) studies. In particular, we focus on the effects of PCR cycle number and primer amplification efficiency on the results of diversity metrics in sequencing studies. We then show that proportional indices of amplicon reads capture trends in taxon biomass with high accuracy, particularly where amplification efficiency is high (median correlation up to 0.97). Our results explain much of the observed behavior of PCR-based studies, and lead to recommendations for best practices in the field.

We can recover genetic information from organisms of all kinds using environmental sampling. In recent years, sequencing this environmental DNA (eDNA) has become a tractable means of surveying many species using water, air, or soil samples. The technique is beginning to become a core tool for ecologists, environmental scientists, and biologists of many kinds, but the temporal resolution of eDNA sampling is often unclear, limiting the ecological interpretations of the resulting datasets. Here, in a temporally and spatially replicated field study using ca. 313 bp of eukaryotic COI mtDNA as a marker, we find that nearshore organismal communities are largely consistent across tides. Our findings suggest that nearshore eDNA from both benthic and planktonic taxa tends to be endogenous to the site and water mass sampled, rather than changing with each tidal cycle. However, where physiochemical water mass characteristics change, we find that the relative contributions of a broad range of organisms to eDNA communities shift in concert.

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Amplicon-sequence data from environmental DNA (eDNA) and microbiome studies provide important information for ecology, conservation, management, and health. At present, amplicon-sequencing studies-known also as metabarcoding studies, in which the primary data consist of targeted, amplified fragments of DNA sequenced from many taxa in a mixture-struggle to link genetic observations to the underlying biology in a quantitative way, but many applications require quantitative information about the taxa or systems under scrutiny. As metabarcoding studies proliferate in ecology, it becomes more important to develop ways to make them quantitative to ensure that their conclusions are adequately supported. Here we link previously disparate sets of techniques for making such data quantitative, showing that the underlying polymerase chain reaction mechanism explains the observed patterns of amplicon data in a general way. By modeling the process through which amplicon-sequence data arise, rather than transforming the data post hoc, we show how to estimate the starting DNA proportions from a mixture of many taxa. We illustrate how to calibrate the model using mock communities and apply the approach to simulated data and a series of empirical examples. Our approach opens the door to improve the use of metabarcoding data in a wide range of applications in ecology, public health, and related fields.

To understand the ecosystem dynamics that underpin the year-round presence of a large generalist consumer, the Bryde’s whale (Balaenoptera edeni brydei), we use a DNA metabarcoding approach and systematic zooplankton surveys to investigate seasonal and regional changes in zooplankton communities and if whale diet reflects such changes. Twenty-four zooplankton community samples were collected from three regions throughout the Hauraki Gulf, New Zealand, over two temperature regimes (warm and cool seasons), as well as 20 samples of opportunistically collected Bryde’s whale scat. Multi-locus DNA barcode libraries were constructed from 18S and COI gene fragments, representing a trade-off between identification and resolution of metazoan taxa. Zooplankton community OTU occurrence and relative read abundance showed regional and seasonal differences based on permutational analyses of variance in both DNA barcodes, with significant changes in biodiversity indices linked to season in COI only. In contrast, we did not find evidence that Bryde’s whale diet shows seasonal or regional trends, but instead indicated clear prey preferences for krill-like crustaceans, copepods, salps and ray-finned fishes independent of prey availability. The year-round presence of Bryde’s whales in the Hauraki Gulf is likely associated with the patterns of distribution and abundance of these key prey items.

Organisms of all kinds leave genetic traces in their environments, and in recent years, sequencing this environmental DNA (eDNA) has become a tractable means of surveying many species using water, air, or soil samples. The technique is beginning to become a core tool for ecologists, environmental scientists, and biologists of many kinds, but the temporal resolution of eDNA sampling is often unclear, limiting the ecological interpretations of the resulting datasets. Here, in a temporally and spatially replicated field study using ca. 330bp of eukaryotic COI mtDNA as a marker, we find that nearshore organismal communities are largely consistent across tides. Our findings suggest that nearshore eDNA tends to be endogenous to the site and water mass sampled, rather changing systematically as waters change over during the tidal cycle. However, where water-mass characteristics change, we find that the eDNA communities change in concert, again suggesting a close association between the habitat sampled and the eDNA community recovered.

Amplicon-sequence data from environmental DNA (eDNA) and microbiome studies provides important information for ecology, conservation, management, and health. At present, amplicon-sequencing studies – known also as metabarcoding studies, in which the primary data consist of targeted, amplified fragments of DNA sequenced from many taxa in a mixture – struggle to link genetic observations to underlying biology in a quantitative way, but many applications require quantitative information about the taxa or systems under scrutiny. As metabarcoding studies proliferate in ecology following decades of microbial and microbiome work using similar techniques, it becomes more important to develop ways ot make them quantitative to ensure that their conclusions are adequately supported. Here we link previously disparate sets of techniques for making such data quantitative, showing that the underlying PCR mechanism explains observed patterns of amplicon data in a general way. By modeling the process through which amplicon-sequence data arises, rather than transforming the data post-hoc, we show how to estimate the starting DNA proportions from a mixture of many taxa. We illustrate how to calibrate the model using mock communities and apply the approach to simulated data and a series of empirical examples. Our approach opens the door to improve the use of metabarcoding data in a wide range of applications in ecology, public health, and related fields.

Meroplankton community studies in the Antarctic have primarily focused on the coastal waters of both the Antarctic Peninsula and the Ross Sea. The New Zealand International Polar Year -Census of Antarctic Marine Life (IPY-CAML) voyage to the Ross Sea during the late summer (February-March) 2008 provided the first meroplankton samples from three regions in the deep, oceanic waters of the Ross Sea (shelf, slope and adjacent offshore Antarctic waters of Admiralty Seamount and Scott Island). We used a combined morphological and molecular approach to identify 36 larval operational taxonomic units based on sequences from three loci (16S, 18S, COI), and exclude early developmental stages of holoplankton. Overall, larval abundance was lower than previous Antarctic studies (5.19 specimens per 100 m 3 ), with larvae most abundant in the first 200 m of the water column and most diverse in the shelf region. Multivariate analysis revealed significant differences in the meroplankton community between regions and depth ranges, but with low similarity within these groupings; differences between water masses were undetectable due to the confounding effect with both region and depth. The influence of nearby benthic populations (e.g. the acorn barnacle Bathylasma corolliforme) and/or locally abundant taxa (e.g. the nudibranch Tergipes antarcticus) was evident in the meroplankton community.