eDNA Surveys Research Articles

The downside of eDNA as a survey tool in water bodies

Summary There is a growing literature on the detection of species‐specific DNA fragments in water to reveal animals' presence in aquatic environments. Analysis of Rees et al. (Journal of Applied Ecology, 2014, 51, 1450) and additional publications highlight the downside of the method and suggest that some conclusions have to be toned down. Non‐detection of species‐specific DNA fragments in a water sample does not automatically imply the absence of the target species. A positive signal does not necessarily mean the species is present because the environmental DNA (eDNA) could have been transported or preserved after an animal's death. Detection of aquatic invertebrates is weakly documented. The effect of species abundance on detection efficiency is not always established. Synthesis and applications. Before successfully becoming an efficient tool to assist proper management policies for monitoring of aquatic species, eDNA surveys must implement sampling strategies that allow estimating detection rates and scaling up information from water samples to larger spatial areas. The effects of discharge variations and dendritic organization in running waters require particular consideration.

Environmental DNA particle size distribution from Brook Trout (Salvelinus fontinalis)

Environmental DNA (eDNA) sampling has become a widespread approach for detecting aquatic animals with high potential for improving conservation biology. However, little research has been done to determine the size of particles targeted by eDNA surveys. In this study, we conduct particle distribution analysis of eDNA from a captive Brook Trout (Salvelinus fontinalis) in a flow-through mesocosm. Our data suggest that 1.2–10-μm particles are the most common size of eDNA from Brook Trout, which is consistent with our hypotheses that eDNA in the environment is comprised of loosely aggregated smaller particles, resulting in high inter-sample heterogeneity. These findings are similar to those of a study on Common Carp (Cyprinus carpio) in lentic systems.

How will the ‘molecular revolution’ contribute to biological recording?

Soaring throughput, plummeting costs, and increased sensitivity for assaying degraded or low-concentration DNA are driving a revolution in the way that we monitor biodiversity. Arguably the biggest ‘game-changer’ is environmental DNA (eDNA), which refers to free-floating DNA released by organisms into their environment. Rare or elusive species can be detected with greater sensitivity and accuracy using eDNA than by most conventional methods, and we have the capability to screen and describe whole communities, as well as perform targeted monitoring of single species. This paper discusses the basic approaches for molecular monitoring of biodiversity, provides case studies to demonstrate the effectiveness of the techniques, and considers any challenges and limitations that could impact molecular biological recording. It is argued that eDNA surveys offer exciting new opportunities to engage the public in biological recording and that molecular approaches will complement conventional surveys, enabling unprecedented insights into species distributions. Finally, with the number of eDNA studies increasing at a rapid pace, it is argued that there is a need to rapidly establish ways of managing molecular records. Integrating molecular records into existing biological records databases would enhance our understanding of species distributions and may be something that the Biological Records Centre should be considering to mark its landmark anniversary. © 2015 The Linnean Society of London, Biological Journal of the Linnean Society, 2015, ●●, ●●–●●.

Monitoring the near-extinct European weather loach in Denmark based on environmental DNA from water samples

The European weather loach (Misgurnus fossilis) represents one of many European freshwater fishes in decline. Efficient monitoring is essential if conservation efforts are to be successful, but due to the species’ cryptic biology, traditional monitoring methods currently in use are inefficient, time consuming and likely prone to non-detection error. Here, we investigate the usefulness of environmental DNA (eDNA) monitoring as an alternative or supplementary method for surveying the Danish weather loach population, which is presumed to consist primarily of a single group of no more than 50 individuals. In 2008, the majority of historical Danish localities were surveyed, using traditional fishing techniques. We then applied eDNA methods to a number of these, as well as other potential localities. We successfully detected the weather loach at multiple sites in the single known remaining locality; a result that was later confirmed when local managers caught eight live specimens. Furthermore, the eDNA method indicated presence of the weather loach in another historical locality, where the species has not been observed since 1995. At the remaining localities, weather loach eDNA was not detected, providing further evidence for its absence. Importantly, the eDNA survey required less effort in person-hours and lower costs than the traditional fishing survey. This study confirms that eDNA monitoring is a valid supplement to traditional monitoring methods currently applied to monitor rare freshwater fishes. We propose that by providing reliable distribution data at lower cost and limited effort, the eDNA method can allow for increased management efficiency of endangered freshwater species such as the European weather loach.

The application of eDNA for monitoring of the Great Crested Newt in the UK.

Current ecological surveys for great crested newts are time-consuming and expensive and can only be carried out within a short survey window. Additional survey methods which would facilitate the detection of rare or protected species such as the great crested newt (Triturus cristatus) would be extremely advantageous. Environmental DNA (eDNA) analysis has been utilized for the detection of great crested newts in Denmark. Here, the same methodology has been applied to water samples taken from UK ponds concurrently with conventional field surveying techniques. Our eDNA analysis exhibited an 84% success rate with a kappa coefficient of agreement between field and eDNA surveys of 0.86. One pond determined to be negative for great crested newt by field survey was positive by eDNA analysis, revealing the potential for improved detection rates using this methodology. Analysis of water samples collected in late summer indicates that eDNA analysis could be used to detect great crested newt after the optimal survey window for current field techniques had passed. Consequently, eDNA analysis could augment currently stipulated techniques for great crested newt surveying as a relatively quick and inexpensive tool for collecting great crested newt presence and distribution data within the UK instead of or prior to full field surveys.

Detection of invasive freshwater fish species using environmental DNA survey

Invasive species are one of the most significant problem in freshwater ecosystems. Most common non-native freshwater species in Turkish freshwater fish fauna are Prussian Carp (Carassius gibelio), North African Catfish (Clarias gariepinus), Nile Tilapia (Oreochromis niloticus) and Topmouth Gudgeon (Pseudorasbora parva).Recent studies showed that environmental DNA could be used to detect target species inhabiting the ecosystem with higher precision and less effort compared to traditional field surveys. In this study, eDNA approach was used to investigate non-native freshwater fish species from fifteen different locations of Upper Sakarya Basin. eDNA was successfully extracted from the water samples of locations where the species were visually observed. Mean amplification rate of eDNA was calculated as 77.03%.This study is the first environmental DNA study used in detection of four of the most common invasive freshwater fish species. Results clearly indicating that eDNA surveys could be used as an important molecular tool to monitor invasive fish species in freshwater ecosystems.

Site occupancy models in the analysis of environmental DNA presence/absence surveys: a case study of an emerging amphibian pathogen

Summary The use of environmental DNA (eDNA) to detect species in aquatic environments such as ponds and streams is a powerful new technique with many benefits. However, species detection in eDNA‐based surveys is likely to be imperfect, which can lead to underestimation of the distribution of a species. Site occupancy models account for imperfect detection and can be used to estimate the proportion of sites where a species occurs from presence/absence survey data, making them ideal for the analysis of eDNA‐based surveys. Imperfect detection can result from failure to detect the species during field work (e.g. by water samples) or during laboratory analysis (e.g. by PCR). To demonstrate the utility of site occupancy models for eDNA surveys, we reanalysed a data set estimating the occurrence of the amphibian chytrid fungus Batrachochytrium dendrobatidis using eDNA. Our reanalysis showed that the previous estimation of species occurrence was low by 5–10%. Detection probability was best explained by an index of the number of hosts (frogs) in ponds. Per‐visit availability probability in water samples was estimated at 0·45 (95% CRI 0·32, 0·58) and per‐PCR detection probability at 0·85 (95% CRI 0·74, 0·94), and six water samples from a pond were necessary for a cumulative detection probability >95%. A simulation study showed that when using site occupancy analysis, researchers need many fewer samples to reliably estimate presence and absence of species than without use of site occupancy modelling. Our analyses demonstrate the benefits of site occupancy models as a simple and powerful tool to estimate detection and site occupancy (species prevalence) probabilities despite imperfect detection. As species detection from eDNA becomes more common, adoption of appropriate statistical methods, such as site occupancy models, will become crucial to ensure that reliable inferences are made from eDNA‐based surveys.

Environmental DNA as a new method for early detection of New Zealand mudsnails (Potamopyrgus antipodarum)

Early detection of aquatic invasive species is a critical task for management of aquatic ecosystems. This task is hindered by the difficulty and cost of surveying aquatic systems thoroughly. The New Zealand mudsnail (Potamopyrgus antipodarum) is a small, invasive parthenogenic mollusk that can reach very high population densities and severely affects ecosystem functioning. To assist in the early detection of this invasive species, we developed and validated a highly sensitive environmental deoxyribonucleic acid (eDNA) assay. We used a dose–response laboratory experiment to investigate the relationship between New Zealand mudsnail density and eDNA detected through time. We documented that as few as 1 individual in 1.5 L of water for 2 d could be detected with this method, and that eDNA from this species may remain detectable for 21 to 44 d after mudsnail removal. We used the eDNA method to confirm the presence of New Zealand mudsnail eDNA at densities as low as 11 to 144 snails/m2 in a eutrophic 5th-order river. Combined, these results demonstrate the high potential for eDNA surveys to assist with early detection of a widely distributed invasive aquatic invertebrate.

The widely used small subunit 18S rDNA molecule greatly underestimates true diversity in biodiversity surveys of the meiofauna

Molecular tools have revolutionized the exploration of biodiversity, especially in organisms for which traditional taxonomy is difficult, such as for microscopic animals (meiofauna). Environmental (eDNA) metabarcode surveys of DNA extracted from sediment samples are increasingly popular for surveying biodiversity. Most eDNA surveys use the nuclear gene-encoding small-subunit rDNA gene (18S) as a marker; however, different markers and metrics used for delimiting species have not yet been evaluated against each other or against morphologically defined species (morphospecies). We assessed more than 12,000 meiofaunal sequences of 18S and of the main alternatively used marker [Cytochrome c oxidase subunit I (COI) mtDNA] belonging to 55 datasets covering three taxonomic ranks. Our results show that 18S reduced diversity estimates by a factor of 0.4 relative to morphospecies, whereas COI increased diversity estimates by a factor of 7.6. Moreover, estimates of species richness using COI were robust among three of four commonly used delimitation metrics, whereas estimates using 18S varied widely with the different metrics. We show that meiofaunal diversity has been greatly underestimated by 18S eDNA surveys and that the use of COI provides a better estimate of diversity. The suitability of COI is supported by cross-mating experiments in the literature and evolutionary analyses of discreteness in patterns of genetic variation. Furthermore its splitting of morphospecies is expected from documented levels of cryptic taxa in exemplar meiofauna. We recommend against using 18S as a marker for biodiversity surveys and suggest that use of COI for eDNA surveys could provide more accurate estimates of species richness in the future.

Departmental status desired, EDNA survey shows

Departmental status desired, EDNA survey shows

Chronic nephritis due to ingestion of lead-contaminated flour.

<h3>Abstract</h3> Short-term target species for mitigation should be realistically and effectively selected for restoration within a few years. Current short-term target species selection is based on spatial distribution data of past and present biota in the neighborhood based on capture surveys. However, current methods have the following problems: If historical spatial distribution data are used, the risk of including large areas, where target species cannot be dispersed, exists, as well as the risk of including extinct species. If recent spatial distribution data are used, the survey area may be narrowed due to time constraints, thereby increasing the possibility of missing species that should be selected as the target species. We examined the possibility of using the environmental DNA (eDNA) metabarcoding method, which can obtain spatial distribution data quickly, sensitively, and extensively, as a new short-term target species selection method. The study site was a restoration mitigation area (Kaihotsu–Kasumi) in the Shigenobu River system, Ehime Prefecture, Japan, where freshwater fish target species were selected based on capture surveys and past literature information. Furthermore, eDNA surveys were conducted at 17 sites in the Kaihotsu–Kasumi and its inflow and outflow rivers in 2020, and metabarcoding analysis using MiFish primers was conducted to determine the current freshwater fish fauna at each site. The visualization of the fish community composition obtained from the eDNA survey using NMDS showed that the Kaihotsu–Kasumi and surrounding watersheds were significantly divided into three clusters: upper reaches, middle and lower reaches, and estuarine reaches. Kaihotsu–Kasumi was included in Cluster 2, comprising the middle and lower reaches of the inflow and outflow rivers. Twenty-six species (including five non-native species) were detected in Cluster 2. Twenty-one fish species, excluding non-native species, were considered suitable as short-term target species as taxa with high potential for dispersal into Kaihotsu–Kasumi. The selection of short-term target species using the eDNA metabarcoding method covered all short- and medium-term target species (14 species) selected based on the capture surveys, and seven additional species were also proposed. The study also showed that intraspecific lineages of <i>Misgurnus anguillicaudatus</i> (Clades A and B), which were missed by the capture surveys, can be identified. These results indicate that the eDNA metabarcoding method can provide more comprehensive and realistic short-term target species selection than capture surveys, providing higher resolution monitoring through intraspecific lineage detection.