To celebrate our 10th Anniversary, we are highlighting a key article from each of our volumes. For Volume 7 we selected The ecologist’s field guide to sequence‐based identification of biodiversity’ by Creer et al. (2016).

In this post, two of our Associate Editors with expertise in eDNA, Lynsey Harper and Chloe Robinson, select their favourite MEE papers in this field.

Lynsey Harper Liverpool John Moores University, UK

Molecular biodiversity assessment is booming, with DNA being sequenced from a broad range of source material, including water, soil, air, faeces, gut contents and whole organisms. Molecular tools can enable targeted detection of rare, cryptic and elusive species and invasive non-native species, diversity estimation and structure of entire communities along biotic or abiotic gradients, dietary profiling and trophic network construction, and analysis of gene expression patterns. Creer et al. (2016), provided a beginner’s guide for ecologists venturing into this world, from sampling to sequencing. Since then, the field has continued to grow, with new applications, challenges and solutions discovered.

Ecologists trying to capture eDNA must decide what filter to use and how to preserve it. Water sampling and filtration can also generate excessive plastic waste as sterile equipment often equates to single-use, particularly when filtering on-site. Consequently, eDNA analysis can be a double-edged sword, both helping and damaging the environment. Thomas et al. (2019) offered a solution in the form of a self-preserving, partially biodegradable filter that facilitates on-site filtration, removes the need for a freezer or preservation buffer, and minimises plastic waste.

Ecologists using metabarcoding cannot underestimate the importance of marker gene(s) selection as performance will vary across sample types and taxonomic groups. The wrong choice could result in wasted sequencing effort, biased diversity and relative abundance estimates, and erroneous regulatory decisions. Collins et al. (2019) provided evidence for this in their marker gene comparison for freshwater and marine fishes from the British Isles. Despite extensive reference database representation, COI primers performed poorly in comparison to 12S primers due to non-specific amplification of non-target DNA (e.g. microbes, plankton) inherent to aquatic eDNA samples.

However, COI primers can perform extremely well for bulk tissue samples as demonstrated by Elbrecht et al. (2017). They showed that metabarcoding of macroinvertebrate specimens could dramatically enhance routine freshwater biomonitoring by identifying taxa missed by and taxa at higher resolution than morphological identification. Metabarcoding can also improve understanding of pollutants entering food chains. For example, Nelms et al. (2019) combined faecal DNA metabarcoding with a microplastic isolation method to associate prey items with higher or lower microplastic exposure in a marine top predator, the grey seal (Halichoerus grypus).

DNA in environmental, faecal and gut content samples is often assumed to be highly degraded, leading metabarcoding studies to target short fragments which may not always allow species-level identification. Deiner et al. (2017) showed that not all DNA may be degraded by amplifying mitochondrial genomes from aquatic eDNA samples using long-range PCR with high-throughput sequencing. This approach could enable environmental samples to be used for studying population genetics, systematics and phylogeography. Such studies will become more likely with continued development of long-read sequencers, such as the Oxford Nanopore MinION.

The Oxford Nanopore MinION is also an example of a portable sequencer, which could change molecular biodiversity assessment in remote and inaccessible locations. Truelove et al. (2019) showed that it was possible to detect charismatic marine megafauna, such as the great white shark (Carcharodon  carcharias), while onboard an oceanographic vessel with this technology.

Molecular tools have opened up new avenues for us to monitor, understand and protect biodiversity. Let’s continue to boldly go where no ecologist has gone before!

Chloe Robinson University of Guelph, Canada

Environmental DNA (eDNA) research, particularly in aquatic systems has been growing since the early 2000s. Methods in Ecology and Evolution has a plethora of published eDNA studies, ranging from development of field methods, species/taxa case study applications and R packages and tools.  For me, a few studies stand out as pivotal for influencing the future of eDNA research. One such paper, though not directly eDNA-based itself, helped lead the way for diagnostic detections of species from eDNA samples. Winder et al. (2010), evaluated the application of DNA melting analysis as a tool for species identification, to determine whether this approach enabled rapid, accurate and reliable identification of organisms. This premise formed the basis of analysis for single-species qPCR eDNA high resolution melt (HRM) assays, which have been widely applied for a range of species including crayfish (and associated pathogens), green sea turtles and invasive brown trout.

Two of the main limitations of eDNA research are the ability to detect DNA within large volumes of water, such as the ocean, and the (often) slow turnaround of sample processing. The paper by Truelove et al. (2019), overcame both of these issues through using a portable Oxford Nanopore MinIon sequencer to sequence eDNA samples for white shark (Carcharodon carcharias). This study fascinated me, regarding their ~48 hr time frame from sample collection to presenting of results and their robust study design has opened a world of opportunities for detecting elusive marine megafauna using eDNA.

As more and more eDNA studies are published, it is important to reflect on the current practices, in terms of field, lab and bioinformatic analysis, to best inform future directions. The review paper by Goldberg et al. (2016), presented a synthesis of knowledge for aquatic-based eDNA studies and established considerations for implementing eDNA methods to detect aquatic macroorganisms. This review was incredibly informative and provided supportive guidelines for all steps of undertaking eDNA research.

To find out more about the article chosen for Volume 7, read the authors’ reflections:

 10th Anniversary Volume 7: The ecologist’s field guide to sequence‐based identification of biodiversity

Find out about the Methods in Ecology and Evolution articles selected to celebrate the other volumes and our editors’ favourite papers in this collection of MEE blog posts.