skip to main content
US FlagAn official website of the United States government
dot gov icon
Official websites use .gov
A .gov website belongs to an official government organization in the United States.
https lock icon
Secure .gov websites use HTTPS
A lock ( lock ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.


Title: DNA metabarcoding captures different macroinvertebrate biodiversity than morphological identification approaches across a continental scale
Abstract DNA‐based aquatic biomonitoring methods show promise to provide rapid, standardized, and efficient biodiversity assessment to supplement and in some cases replace current morphology‐based approaches that are often less efficient and can produce inconsistent results. Despite this potential, broad‐scale adoption of DNA‐based approaches by end‐users remains limited, and studies on how these two approaches differ in detecting aquatic biodiversity across large spatial scales are lacking. Here, we present a comparison of DNA metabarcoding and morphological identification, leveraging national‐scale, open‐source, ecological datasets from the National Ecological Observatory Network (NEON). Across 24 wadeable streams in North America with 179 paired sample comparisons, we found that DNA metabarcoding detected twice as many unique taxa than morphological identification overall. The two approaches showed poor congruence in detecting the same taxa, averaging 59%, 35%, and 23% of shared taxa detected at the order, family, and genus levels, respectively. Importantly, the two approaches detected different proportions of indicator taxa like %EPT and %Chironomidae. DNA metabarcoding detected far fewer Chironomid and Trichopteran taxa than morphological identification, but more Ephemeropteran and Plecopteran taxa, a result likely due to primer choice. Overall, our results showed that DNA metabarcoding and morphological identification detected different benthic macroinvertebrate communities. Despite these differences, we found that the same environmental variables were correlated with invertebrate community structure, suggesting that both approaches can accurately detect biodiversity patterns across environmental gradients. Further refinement of DNA metabarcoding protocols, primers, and reference libraries–as well as more standardized, large‐scale comparative studies–may improve our understanding of the taxonomic agreement and data linkages between DNA metabarcoding and morphological approaches.  more » « less
Award ID(s):
2207680
PAR ID:
10442051
Author(s) / Creator(s):
 ;  ;  ;  ;  ;  ;  
Publisher / Repository:
Wiley Blackwell (John Wiley & Sons)
Date Published:
Journal Name:
Environmental DNA
Volume:
5
Issue:
6
ISSN:
2637-4943
Format(s):
Medium: X Size: p. 1307-1320
Size(s):
p. 1307-1320
Sponsoring Org:
National Science Foundation
More Like this
  1. Abstract Biodiversity is changing at an accelerating rate at both local and regional scales. Beta diversity, which quantifies species turnover between these two scales, is emerging as a key driver of ecosystem function that can inform spatial conservation. Yet measuring biodiversity remains a major challenge, especially in aquatic ecosystems. Decoding environmental DNA (eDNA) left behind by organisms offers the possibility of detecting species sans direct observation, a Rosetta Stone for biodiversity. While eDNA has proven useful to illuminate diversity in aquatic ecosystems, its utility for measuring beta diversity over spatial scales small enough to be relevant to conservation purposes is poorly known. Here we tested how eDNA performs relative to underwater visual census (UVC) to evaluate beta diversity of marine communities. We paired UVC with 12S eDNA metabarcoding and used a spatially structured hierarchical sampling design to assess key spatial metrics of fish communities on temperate rocky reefs in southern California. eDNA provided a more-detailed picture of the main sources of spatial variation in both taxonomic richness and community turnover, which primarily arose due to strong species filtering within and among rocky reefs. As expected, eDNA detected more taxa at the regional scale (69 vs. 38) which accumulated quickly with space and plateaued at only ~ 11 samples. Conversely, the discovery rate of new taxa was slower with no sign of saturation for UVC. Based on historical records in the region (2000–2018) we found that 6.9 times more UVC samples would be required to detect 50 taxa compared to eDNA. Our results show that eDNA metabarcoding can outperform diver counts to capture the spatial patterns in biodiversity at fine scales with less field effort and more power than traditional methods, supporting the notion that eDNA is a critical scientific tool for detecting biodiversity changes in aquatic ecosystems. 
    more » « less
  2. Microorganisms are ubiquitous in the biosphere, playing a crucial role in both biogeochemistry of the planet and human health. However, identifying these microorganisms and defining their function are challenging. Widely used approaches in comparative metagenomics, 16S amplicon sequencing and whole genome shotgun sequencing (WGS), have provided access to DNA sequencing analysis to identify microorganisms and evaluate diversity and abundance in various environments. However, advances in parallel high-throughput DNA sequencing in the past decade have introduced major hurdles, namely standardization of methods, data storage, reproducible interoperability of results, and data sharing. The National Ecological Observatory Network (NEON), established by the National Science Foundation, enables all researchers to address queries on a regional to continental scale around a variety of environmental challenges and provide high-quality, integrated, and standardized data from field sites across the U.S. As the amount of metagenomic data continues to grow, standardized procedures that allow results across projects to be assessed and compared is becoming increasingly important in the field of metagenomics. We demonstrate the feasibility of using publicly available NEON soil metagenomic sequencing datasets in combination with open access Metagenomics Rapid Annotation using the Subsystem Technology (MG-RAST) server to illustrate advantages of WGS compared to 16S amplicon sequencing. Four WGS and four 16S amplicon sequence datasets, from surface soil samples prepared by NEON investigators, were selected for comparison, using standardized protocols collected at the same locations in Colorado between April-July 2014. The dominant bacterial phyla detected across samples agreed between sequencing methodologies. However, WGS yielded greater microbial resolution, increased accuracy, and allowed identification of more genera of bacteria, archaea, viruses, and eukaryota, and putative functional genes that would have gone undetected using 16S amplicon sequencing. NEON open data will be useful for future studies characterizing and quantifying complex ecological processes associated with changing aquatic and terrestrial ecosystems. 
    more » « less
  3. Abstract Marine zooplankton are key players in pelagic food webs, central links in ecosystem function, useful indicators of water masses, and rapid responders to environmental variation and climate change. Characterization of biodiversity of the marine zooplankton assemblage is complicated by many factors, including systematic complexity of the assemblage, with numerous rare and cryptic species, and high local-to-global ratios of species diversity. The papers in this themed article set document important advances in molecular protocols and procedures, integration with morphological taxonomic identifications, and quantitative analyses (abundance and biomass). The studies highlight several overarching conclusions and recommendations. A primary issue is the continuing need for morphological taxonomic experts, who can identify species and provide voucher specimens for reference sequence databases, which are essential for biodiversity analyses based on molecular approaches. The power of metabarcoding using multi-gene markers, including both DNA (Deoxyribonucleic Acid) and RNA (Ribonucleic Acid)templates, is demonstrated. An essential goal is the accurate identification of species across all taxonomic groups of marine zooplankton, with particular concern for detection of rare, cryptic, and invasive species. Applications of molecular approaches include analysis of trophic relationships by metabarcoding of gut contents, as well as investigation of the underlying ecological and evolutionary forces driving zooplankton diversity and structure. 
    more » « less
  4. ABSTRACT DNA metabarcoding of zooplankton biodiversity is used increasingly for monitoring global ocean ecosystems, requiring comparable data from different research laboratories and ocean regions. The MetaZooGene Intercalibration Experiment (MZG‐ICE) was designed to examine1 and analyse patterns of variation of DNA sequence data resulting from multi‐gene metabarcoding of 10 zooplankton samples carried out by 10 research groups affiliated with the Scientific Committee for Ocean Research (SCOR). Aliquots of DNA extracted from the 10 zooplankton samples were distributed to MZG‐ICE groups for metabarcoding of four gene regions: V1‐V2, V4 and V9 of nuclear 18S rRNA and mitochondrial COI. Molecular protocols and procedures were recommended; substitutions were allowed as necessary. Resulting data were uploaded to a common repository for centralised statistics and bioinformatics. Based on proportional sequence numbers for abundant phyla, overall patterns of variation were consistent across many—but not all—MZG‐ICE groups. V9 showed highest similarity, followed (in order) by V4, V1‐V2, and COI. Outlier data were hypothesised to result from the use of different PCR protocols and sequencing platforms, and possible contamination. MZG‐ICE results indicated that DNA metabarcoding data from different laboratories and research groups can provide reliable, accurate and valid descriptions of biodiversity of zooplankton throughout the ocean. Recommendations included: pre‐screening QA/QC of raw data, detailed records for laboratory protocols, reagents, and instrumentation, and centralised bioinformatics and multivariate statistics. In the absence of universal agreement on standardised protocols or best practices, intercalibration is the best way forward toward validation of DNA metabarcoding of zooplankton diversity for global ocean monitoring. 
    more » « less
  5. Biodiversity monitoring based on DNA metabarcoding depends on primer performance. Here, we develop a new metabarcoding primer pair that targets a ~ 318 bp fragment of the 28S rRNA gene. We validate the primer pair in assessing sponges, a notoriously challenging group for coral reef metabarcoding studies, by using mock and natural complex reef communities to examine its performance in species detection, amplification efficiency, and quantitative potential. Mock community experiments revealed a high number of sponge species (n = 94) spanning a broad taxonomic scope (15 orders), limited taxon-specific primer biases (only a single species exceeded a two-fold deviation from the expected number of reads), and its suitability for quantitative metabarcoding – there was a significant relationship between read abundance and visual percent coverage of sponge taxa (R = 0.76). In the natural complex coral reef community experiments, commonly used COI metabarcoding primers detected only 30.9% of sponge species, while the new 28S primer increased detection to 79.4%. These new 28S primers detect a broader taxonomic array of species across phyla and classes within the complex cryptobiome of coral reef communities than the Leray-Geller COI primers. As biodiversity assessments using metabarcoding tools are increasingly being leveraged for environmental monitoring and guide policymaking, these new 28S rRNA primers can improve biodiversity assessments for complex ecological coral reef communities. 
    more » « less