Ecosystem biomonitoring with eDNA: metabarcoding across the tree of life in a tropical marine environment

Abstract

Effective marine management requires comprehensive data on the status of marine biodiversity. However, efficient methods that can document biodiversity in our oceans are currently lacking. Environmental DNA (eDNA) sourced from seawater offers a new avenue for investigating the biota in marine ecosystems. Here, we investigated the potential of eDNA to inform on the breadth of biodiversity present in a tropical marine environment. Directly sequencing eDNA from seawater using a shotgun approach resulted in only 0.34% of 22.3 million reads assigning to eukaryotes, highlighting the inefficiency of this method for assessing eukaryotic diversity. In contrast, using ‘tree of life’ (ToL) metabarcoding and 20-fold fewer sequencing reads, we could detect 287 families across the major divisions of eukaryotes. Our data also show that the best performing ‘universal’ PCR assay recovered only 44% of the eukaryotes identified across all assays, highlighting the need for multiple metabarcoding assays to catalogue biodiversity. Lastly, focusing on the fish genus Lethrinus, we recovered intra- and inter-specific haplotypes from seawater samples, illustrating that eDNA can be used to explore diversity beyond taxon identifications. Given the sensitivity and low cost of eDNA metabarcoding we advocate this approach be rapidly integrated into biomonitoring programs.

Non‐specific amplification compromises environmental DNA metabarcoding with COI

“…Particularly important for regulatory applications, or where researchers wish to compare results over time or between studies, some degree of standardization is desirable (Hering et al, ). Our results – and those of previous studies using similar primer sets (Bakker et al, ; Jeunen et al, ; Lim et al, ; Macher et al, ; Singer et al, ; Stat et al, ; Yang et al, ) – show that environmental metabarcoding for restricted taxonomic groups using degenerate COI primers results in excessive volumes of ‘wasted’ sequencing effort. This co‐amplification of prokaryotic and non‐target eukaryotic DNAs and subsequent lack of specificity is due to the nature of mutation patterns in COI (Siddall et al, ).…”

“…That the highly degenerate Leray‐XT primers produced a low proportion of fish reads is unsurprising given that previous studies on environmental samples using degenerate COI primers have demonstrated that they can amplify widely beyond their target taxa, and can produce large proportions of unassigned reads (Lim et al., ; Macher et al, ; Singer et al, ; Stat et al, ). The proportion of bacterial reads are generally lower when metabarcoding bulk organismal samples, however, with most reads belonging to metazoans (Leray & Knowlton, ; Macher et al, ; Wangensteen et al, ).…”

Environmental DNA (eDNA) metabarcoding reveals strong discrimination among diverse marine habitats connected by water movement

“…Library preparation followed the protocol described in (Berry et al, ; Stat et al, ). Briefly, samples were analyzed using three metabarcoding assays targeting two fragments of the 16S rRNA gene region and one fragment of the cytochrome c oxidase subunit I (COI) gene region (Supporting information Table S2).…”

Counting with DNA in metabarcoding studies: How should we convert sequence reads to dietary data?

“…Fortunately, we are starting to move towards a point where markers used in different applications are better understood and alternative less‐biased approaches are being explored. This includes the use of multiple target markers (Stat et al., ) and PCR‐free approaches (Srivathsan, Ang, Vogler, & Meier, ) that can be combined with prey DNA enrichment (Krehenwinkel, Kennedy, Pekár, & Gillespie, ). Inclusion of control materials in sequencing runs can also ensure consistency between experiments (Hardwick, Deveson, & Mercer, ).…”