Trapped DNA fragments in marine sponge specimens unveil North Atlantic deep-sea fish diversity.

Sponges pump water to filter feed and for diffusive oxygen uptake. In doing so, trace DNA fragments from a multitude of organisms living around them are trapped in their tissues. Here we show that the environmental DNA retrieved from archived marine sponge specimens can reconstruct the fish communities at the place of sampling and discriminate North Atlantic assemblages according to biogeographic region (from Western Greenland to Svalbard), depth habitat (80-1600 m), and even the level of protection in place. Given the cost associated with ocean biodiversity surveys, we argue that targeted and opportunistic sponge samples - as well as the specimens already stored in museums and other research collections - represent an invaluable trove of biodiversity information that can significantly extend the reach of ocean monitoring.

Environmental DNA persistence and fish detection in captive sponges.

Large and hyperdiverse marine ecosystems pose significant challenges to biodiversity monitoring. While environmental DNA (eDNA) promises to meet many of these challenges, recent studies suggested that sponges, as "natural samplers" of eDNA, could further streamline the workflow for detecting marine vertebrates. However, beyond pilot studies demonstrating the ability of sponges to capture eDNA, little is known about the dynamics of eDNA particles in sponge tissue, and the effectiveness of the latter compared to water samples. Here, we present the results of a controlled aquarium experiment to examine the persistence and detectability of eDNA captured by three encrusting sponge species and compare the sponge's eDNA capturing ability with established water filtration techniques. Our results indicate that sponges and water samples have highly similar detectability for fish of different sizes and abundances, but different sponge species exhibit considerable variance in performance. Interestingly, one sponge appeared to mirror the eDNA degradation profile of water samples, while another sponge retained eDNA throughout the experiment. A third sponge yielded virtually no DNA sequences at all. Overall, our study suggests that some sponges will be suitable as natural samplers, while others will introduce significant problems for laboratory processing. We suggest that an initial optimization phase will be required in any future studies aiming to employ sponges for biodiversity assessment. With time, factoring in technical and natural accessibility, it is expected that specific sponge taxa may become the "chosen" natural samplers in certain habitats and regions.

The Multiple States of Environmental DNA and What Is Known about Their Persistence in Aquatic Environments.

Increased use of environmental DNA (eDNA) analysis for indirect species detection has spurred the need to understand eDNA persistence in the environment. Understanding the persistence of eDNA is complex because it exists in a mixture of different states (e.g., dissolved, particle adsorbed, intracellular, and intraorganellar), and each state is expected to have a specific decay rate that depends on environmental parameters. Thus, improving knowledge about eDNA conversion rates between states and the reactions that degrade eDNA in different states is needed. Here, we focus on eukaryotic extraorganismal eDNA, outline how water chemistry and suspended mineral particles likely affect conversion among each eDNA state, and indicate how environmental parameters affect persistence of states in the water column. On the basis of deducing these controlling parameters, we synthesized the eDNA literature to assess whether we could already derive a general understanding of eDNA states persisting in the environment. However, we found that these parameters are often not being measured or reported when measured, and in many cases very few experimental data exist from which to draw conclusions. Therefore, further study of how environmental parameters affect eDNA state conversion and eDNA decay in aquatic environments is needed. We recommend analytic controls that can be used during the processing of water to assess potential losses of different eDNA states if all were present in a water sample, and we outline future experimental work that would help determine the dominant eDNA states in water.

Mapping biodiversity hotspots of fish communities in subtropical streams through environmental DNA.

Large tropical and subtropical rivers are among the most biodiverse ecosystems worldwide, but also suffer from high anthropogenic pressures. These rivers are hitherto subject to little or no routine biomonitoring, which would be essential for identification of conservation areas of high importance. Here, we use a single environmental DNA multi-site sampling campaign across the 200,000 km2 Chao Phraya river basin, Thailand, to provide key information on fish diversity. We found a total of 108 fish taxa and identified key biodiversity patterns within the river network. By using hierarchical clustering, we grouped the fish communities of all sites across the catchment into distinct clusters. The clusters not only accurately matched the topology of the river network, but also revealed distinct groups of sites enabling informed conservation measures. Our study reveals novel opportunities of large-scale monitoring via eDNA to identify relevant areas within whole river catchments for conservation and habitat protection.

Assessing the impact of the threatened crucian carp (Carassius carassius) on pond invertebrate diversity: A comparison of conventional and molecular tools.

Fishes stocked for recreation and angling can damage freshwater habitats and negatively impact biodiversity. The pond-associated crucian carp (Carassius carassius) is rare across Europe and is stocked for conservation management in England, but its impacts on pond biota are understudied. Freshwater invertebrates contribute substantially to aquatic biodiversity, encompassing many rare and endemic species, but their small size and high abundance complicate their assessment. Practitioners have employed sweep-netting and kick-sampling with microscopy (morphotaxonomy), but specimen size/quality and experience can bias identification. DNA and environmental DNA (eDNA) metabarcoding offer alternative means of invertebrate assessment. We compared invertebrate diversity in ponds (N = 18) with and without crucian carp using morphotaxonomic identification, DNA metabarcoding and eDNA metabarcoding. Five 2 L water samples and 3 min sweep-net samples were collected at each pond. Inventories produced by morphotaxonomic identification of netted samples, DNA metabarcoding of bulk tissue samples and eDNA metabarcoding of water samples were compared. Alpha diversity was greatest with DNA or eDNA metabarcoding, depending on whether standard or unbiased methods were considered. DNA metabarcoding reflected morphotaxonomic identification, whereas eDNA metabarcoding produced markedly different communities. These complementary tools should be combined for comprehensive invertebrate assessment. Crucian carp presence minimally reduced alpha diversity in ponds, but positively influenced beta diversity through taxon turnover (i.e., ponds with crucian carp contained different invertebrates to fishless ponds). Crucian carp presence contributes to landscape-scale invertebrate diversity, supporting continued conservation management in England. Our results show that molecular tools can enhance freshwater invertebrate assessment and facilitate development of more accurate and ecologically effective pond management strategies.

Targeted and passive environmental DNA approaches outperform established methods for detection of quagga mussels, Dreissena rostriformis bugensis in flowing water.

The early detection of invasive non-native species (INNS) is important for informing management actions. Established monitoring methods require the collection or observation of specimens, which is unlikely at the beginning of an invasion when densities are likely to be low. Environmental DNA (eDNA) analysis is a highly promising technique for the detection of INNS-particularly during the early stages of an invasion.Here, we compared the use of traditional kick-net sampling with two eDNA approaches (targeted detection using both conventional and quantitative PCR and passive detection via metabarcoding with conserved primers) for detection of quagga mussel, Dreissena rostriformis bugensis, a high priority INNS, along a density gradient on the River Wraysbury, UK.All three molecular tools outperformed traditional sampling in terms of detection. Conventional PCR and qPCR both had 100% detection rate in all samples and outperformed metabarcoding when the target species was at low densities. Additionally, quagga mussel DNA copy number (qPCR) and relative read count (metabarcoding) were significantly influenced by both mussel density and distance from source population, with distance being the most significant predictor. Synthesis and application. All three molecular approaches were more sensitive than traditional kick-net sampling for the detection of the quagga mussel in flowing water, and both qPCR and metabarcoding enabled estimates of relative abundance. Targeted approaches were more sensitive than metabarcoding, but metabarcoding has the advantage of providing information on the wider community and consequently the impacts of INNS.

Needle in a haystack? A comparison of eDNA metabarcoding and targeted qPCR for detection of the great crested newt (Triturus cristatus).

Environmental DNA (eDNA) analysis is a rapid, cost-effective, non-invasive biodiversity monitoring tool which utilises DNA left behind in the environment by organisms for species detection. The method is used as a species-specific survey tool for rare or invasive species across a broad range of ecosystems. Recently, eDNA and "metabarcoding" have been combined to describe whole communities rather than focusing on single target species. However, whether metabarcoding is as sensitive as targeted approaches for rare species detection remains to be evaluated. The great crested newt Triturus cristatus is a flagship pond species of international conservation concern and the first UK species to be routinely monitored using eDNA. We evaluate whether eDNA metabarcoding has comparable sensitivity to targeted real-time quantitative PCR (qPCR) for T. cristatus detection. Extracted eDNA samples (N = 532) were screened for T. cristatus by qPCR and analysed for all vertebrate species using high-throughput sequencing technology. With qPCR and a detection threshold of 1 of 12 positive qPCR replicates, newts were detected in 50% of ponds. Detection decreased to 32% when the threshold was increased to 4 of 12 positive qPCR replicates. With metabarcoding, newts were detected in 34% of ponds without a detection threshold, and in 28% of ponds when a threshold (0.028%) was applied. Therefore, qPCR provided greater detection than metabarcoding but metabarcoding detection with no threshold was equivalent to qPCR with a stringent detection threshold. The proportion of T. cristatus sequences in each sample was positively associated with the number of positive qPCR replicates (qPCR score) suggesting eDNA metabarcoding may be indicative of eDNA concentration. eDNA metabarcoding holds enormous potential for holistic biodiversity assessment and routine freshwater monitoring. We advocate this community approach to freshwater monitoring to guide management and conservation, whereby entire communities can be initially surveyed to best inform use of funding and time for species-specific surveys.