Environmental DNA最新文献

Estuarine habitats serve as critical feeding and nursery grounds for many aquatic species and support fisheries. However, monitoring these complex ecosystems using conventional trawling methods is destructive, costly, and labor-intensive. This study compared trawling and a multi-marker environmental DNA (eDNA) metabarcoding approach to monitor marine vertebrate and crustacean communities in an estuarine environment in subtropical Hong Kong. We analyzed 16 bottom trawl samples and the eDNA from 32 two-liter water samples using primer sets specific to fishes and mammals (MiFish-U, 12S-V5, and Berry-Fish) and decapod crustaceans (MiDeca). We found that the eDNA approach detected more pelagic and demersal fishes (237 vs. 106 in trawling) and elasmobranchs (6 vs. 3) than trawling. The eDNA approach was also more effective than trawling in detecting threatened vertebrates (16 vs. 4), including the Indo-Pacific Finless Porpoise and the critically endangered Large Yellow Croaker. Among the detected fish at species level, 70 species were detected by both approaches, 32 species were detected by trawling only, and 142 species were detected by the eDNA approach only. Regarding crustaceans, the eDNA approach detected slightly fewer decapods (61 vs. 77) and stomatopods (5 vs. 8) than trawl surveys. However, the eDNA approach could be enhanced through the development of suitable decapod-specific primers and the expansion of the local reference database. In addition, multivariate analyses of the eDNA data revealed spatial patterns of fish and crustacean assemblages that might be associated with sediment loading, oxygen, and nutrient levels. Furthermore, there was a positive correlation between eDNA read counts and trawl catch, but their correlation coefficient was low. We conclude that eDNA metabarcoding can provide high-resolution detection of species, composition, and unravel community–environment relationships in estuarine ecosystems. Overall, integrating the non-destructive eDNA approach can complement the conventional trawling method for better-informed sustainable fishery management and conservation.

Environmental DNA (eDNA) has significant potential to improve the efficiency of biological sampling and detect species that pose challenges for traditional sampling methods. However, a key obstacle in utilizing eDNA data for ecosystem management is uncertainty surrounding the ability to estimate abundance or biomass of multiple species simultaneously. In this study, we use experimental trials with known biomasses of multiple species to explore the feasibility of (1) estimating species proportions from eDNA metabarcoding data and (2) estimating absolute eDNA concentrations of multiple species by scaling metabarcoding proportions with eDNA concentrations of a single species obtained from qPCR. The focal species for this study were three gadid fishes that are key components of marine ecosystems in Alaska and vary in their distribution and habitat use: Walleye pollock (Gadus chalcogrammus), Pacific cod (Gadus macrocephalus), and Arctic cod (Boreogadus saida). After designing gadid-specific metabarcoding primers and accounting for PCR biases in the metabarcoding data, we found corrected read proportions closely approximated the true biomass proportions of species. Furthermore, we found strong positive relationships between absolute eDNA concentration and absolute biomass for Arctic cod and Pacific cod using quantitative metabarcoding data combined with estimates of Walleye pollock eDNA concentration derived from qPCR. These findings suggest that it is possible to accurately quantify species compositions and estimate metrics of biomass for gadids in real-world scenarios. Furthermore, this work provides a framework for developing primers and analytical approaches that can be applied to other species to improve the quantitative utility of eDNA.

Marine pest introductions continue to occur and increase at accelerated rates, threatening the marine environment and blue economy. Environmental DNA (eDNA) is a tool for determining the presence of both indigenous and nonindigenous species, via the detection of genetic material that is shed into the local environment. Although eDNA approaches have gained widespread adoption in the last decade, fundamental knowledge gaps remain around factors that can influence the probability of detection, and how to optimize eDNA sampling in aquatic environments. Here, we partition eDNA research into four major research themes: eDNA concentration (shedding and decay), transport (advection and mixing), sampling design strategies, and the modeling of these dynamics. We review current developments and challenges in each theme with a focus on field sampling strategies and the use of biophysical models for understanding the movement of modeling eDNA in complex aquatic environments. We then introduce three modeling case studies from a large embayment where we (1) quantify the spatial and temporal variability of eDNA dispersion, (2) use biophysical models to inform a field sampling strategy, and (3) demonstrate a backtracking modeling technique to identify upstream DNA sources to an existing sample (monitoring) site. We conclude by identifying specific recommendations to help improve future eDNA studies. This work highlights how biophysical models can be applied to improve early detection and informing response and management decisions.

Following the development of high-throughput DNA sequencers, environmental prokaryotic communities were usually described by metabarcoding on short markers of the 16S domain. Among third-generation sequencers, that offered the possibility to sequence the full 16S domain, the portable MinION from Oxford Nanopore was undervalued for metabarcoding because of its relatively higher error rate per read. Here we illustrate the limits and benefits of Nanopore sequencing devices by comparing the prokaryotic community structure in a mock community and 52 sediment samples from mangrove sites, inferred from full-length 16S long-reads (16S-FL, ca. 1.5 kbp) on a MinION device, with those inferred from partial 16S short-reads (16S-V4V5, ca. 0.4 kbp, 16S-V4V5) on Illumina MiSeq. 16S-V4V5 and 16S-FL retrieved all the bacterial species from the mock, but Nanopore long-reads overestimated their diversity (56 species vs. 15). Whether these supplementary OTUs were artefactual or not, they only accounted for ca. 10% of the reads. From the sediment samples, with a coverage-based rarefaction of reads and after singletons filtering, Mantel and Procrustean tests of co-inertia showed that bacterial community structures inferred from 16S-V4V5 and 16S-FL were significantly similar, showing both a comparable contrast between sites and a coherent sea-land orientation within sites. In our dataset, 84.7% and 98.8% of the 16S-V4V5 assigned reads were assigned strictly to the same species and genus, respectively, than those detected by 16S-FL. 16S-FL detected 92.2% of the 309 families and 87.7% of the 448 genera that were detected by the short 16S-V4V5. 16S-FL recorded 973 additional species and 392 genus not detected by 16S-V4V5 (31.5% and 10.4% of the 16S-FL reads, respectively, among which 67.8% and 79.3% were assigned), produced by both primer specificities and different error rates. Thus, our results concluded an overall similarity between 16S-V4V5 and 16S-FL sequencing strategies for this type of environmental samples.

Environmental DNA (eDNA) has evolved into a valuable asset of the ecologists' toolkit, enabling time- and cost-efficient biodiversity assessments in a wide variety of ecosystems. Since eDNA can be isolated from a broad range of environmental substrates, its persistence times in those media are of decisive importance for drawing inferences about species presence. For the first time, we characterize eDNA persistence in water and sediment samples of seminatural waterholes in a savanna system in South Africa to gain a better understanding of eDNA decay in these waterbodies. Using mesocosm experiments, we tracked eDNA decay in two different DNA states (extracellular and membrane bound), during two different seasons (wet and dry), and from two different substrates (surface water and sediment). Extracellular DNA degraded rapidly in a first-order exponential decay fashion and membrane-bound DNA exhibited a slower decline with more intricate patterns, involving initial reduction followed by a subsequent increase in measured DNA concentrations. The latter we attribute to cell disassociation and cell lysis at 24–48 h after introduction into the environment. Higher stochasticity of membrane-bound DNA capture in the dry season highlights the need for higher sampling efforts in natural systems in which eDNA is presumably more patchily distributed. Additionally, we observed longer eDNA persistence in sediments than in water samples, presumably due to better protection from nucleases. We could not reveal any effects of environmental parameters on eDNA decay, emphasizing that further research is needed to better understand eDNA dynamics in those waterbodies in order to exploit their full potential for eDNA-based bioassessments in those systems.

Oak wilt disease, caused by the fungus Bretziella fagacearum, can kill mature red oaks within months of infection, severely affecting biodiversity, landscapes, and industries. The disease, originally only present in the United States, was officially reported for the first time in Canada in June 2023. The aim of this study was to suggest a standardized assay and sample processing method to optimize oak wilt detection both in infection centers and ahead of the disease front. Two previously published molecular assays, a Nested PCR and a TaqMan qPCR, were compared to detect B. fagacearum in a variety of samples in a ring trial across five laboratories. Sample types investigated included eDNA from trapped insect vectors (sorted insects and bulk content from traps), infested and healthy oak wood chips, and B. fagacearum conidia dilutions. Results demonstrated that both Nested and TaqMan assays can be used for molecular confirmation of oak wilt, and results are reproducible across different labs. There is a general agreement between both detection assays when testing true-positive and true-negative samples. Both methods demonstrated overall good accuracy. The TaqMan assay was more sensitive and detected lower amounts of DNA target. Both tests were 100% specific to oak wood samples, which was the best sample type to use for detection. In general, samples with high Cts were more prompted to yield false negative Nested results. Detecting oak wilt from bulk insect samples was by far more rapid than sorted sap beetles, but resulted in lower detection signals, especially with the Nested assay. The time-period when the insect traps were set up also had considerable influence on detection results. We hope this study helps to formulate guidelines in oak wilt detection and biosurveillance management.

Recent research into the spatiotemporal dynamics of eDNA in marine environments indicates that eDNA signals are highly localized and may dissipate beyond detection levels within a few hours of production. This affects whether single-timepoint eDNA sampling, which generally occurs during daylight hours, or cyclic (day/night) interval eDNA sampling is necessary to detect both diurnal and nocturnal marine species. Our study investigated short-term variability in eDNA derived from fishes and macroinvertebrates across three temperate reef sites in eastern Tasmania, Australia. Simultaneous eDNA and underwater visual census (UVC) diver transect surveys were conducted every 6 h over a 24-h period to investigate whether eDNA was able to detect marine species outside of their UVC-observed diel activity. We report that single-timepoint eDNA sampling can detect both diurnal and nocturnal species on temperate reefscapes. A lack of eDNA compositional turnover between day and night suggests that eDNA persists beyond 12 h and/or is continuously produced by both diurnal and nocturnal reef taxa, irrespective of diel behavioral changes observed by UVC. Given high eDNA sample variability, however, we recommend a high replication level (> 10 × 1 L samples) to produce robust site community composition profiles. This study builds on emerging literature on short-term variability in eDNA, assisting in the design of future eDNA studies at sites with pronounced variation in faunal activity between day and night.

Environmental DNA (eDNA) metabarcoding surveys can support the acquisition of extensive biodiversity data to support ecosystem monitoring and conservation actions. However, the optimization of eDNA metabarcoding project design is essential to capture spatio-temporal heterogeneity of eDNA signals and maximize diversity detection. In this study, we developed a system-specific approach to detect fish communities in kelp forests, by analyzing fine-scale spatio-temporal patterns in eDNA signals at two sites along the South African coastline, as well as testing the effect of biological replication and pooling of replicates on species detection. At each site, samples were collected at two stations along the shoreline at two depth zones, and this was repeated at two time points (24 h apart). We detected 113 operational taxonomic units (OTUs) across 32 families, but fewer than 20% of OTUs could be assigned to species, indicating that barcode reference libraries need to be drastically improved. We detected significant differences in communities across small spatial scales (< 600 m) and time points, suggesting that to best capture a site's diversity patterns, samples should be collected at multiple points and times within at least 24 h. To detect ~80% of the fish community, including some low abundance species, a minimum of four samples appear sufficient. In addition, a higher number of OTUs (76 vs. 65) were found in individual replicates than in any of the pools. However, pooling samples prior to sequencing can still detect valuable broad-scale biodiversity patterns for monitoring and can offset the decrease in data resolution with the benefit of accumulating comprehensive data from increased sampling efforts over time. As a pilot investigation into how best to maximize kelp forest-associated fish communities, this study provides a basis for optimizing sampling design for coastal eDNA-based surveys in southern Africa and strengthens the development of long-term eDNA monitoring programs to better support conservation and management actions.

Environmental DNA (eDNA) is frequently used for detecting species and describing biodiversity through metabarcoding techniques. More recently, there has been emerging evidence that eDNA can be used to investigate intraspecific variability, providing novel pathways to explore population genetics questions. However, it can be difficult to distinguish between true intraspecific variation and PCR/sequence error, and the presence of DNA from multiple individuals makes using traditional frequency-based approaches challenging. Here, we explore how eDNA can be used to investigate population structure of Hector's dolphin (Cephalorhynchus hectori), an endemic, endangered, and culturally important (taonga) species. In doing so, we present a simple and effective method to filter out noise due to PCR/sequence error and show how treating haplotype detections equally can provide similar results to frequency-based approaches from traditional sampling methods. Over the 2022/23 Austral summer, we collected 85 water samples close to Hector's dolphins, and three negative controls, across three areas on the east coast of Aotearoa New Zealand's South Island: Banks Peninsula (n = 41), Timaru (n = 33), and Dunedin (n = 14). We targeted a 348 bp region of the cetacean D-loop in the mitochondrial DNA (mtDNA) and obtained positive detections in 68 (77%) water samples, confidently identifying seven haplotypes across the study area. The occurrence of specific haplotypes and the overall frequencies in Banks Peninsula and Timaru matched well with previous tissue-based studies and were similar to other East Coast South Island (ECSI) subpopulations. In Dunedin, however, our results indicate a closer relationship to South Coast populations, suggesting that the membership within the ECSI population be reconsidered, which has implications for how this subpopulation is managed. We show that eDNA sampling can be used to elucidate matrilineal population structure for Hector's dolphin and provide a simple method that could be applied to other eDNA-based studies of any taxa.

Environmental DNA (eDNA) analysis is a promising tool for monitoring wild animal populations and, more recently, their genetic variability. In this study, we used the mitochondrial Cytochrome B gene to develop and apply new eDNA metabarcoding assays targeting amphibian families and genera in order to estimate both inter- and intraspecific genetic diversity. We designed and tested seven new primer pairs (a) in silico against an amphibian reference database based on the target genera; (b) in vitro on tissue samples of the target species; and (c) in situ on water samples from 38 wetlands in the Province of Trento (Italy). Overall, most target species were amplified successfully, although some markers also amplified non-target amphibian species. In addition, to complete the workflow, we compared the performance of three different bioinformatic pipelines (namely, MICCA with VSEARCH, and OBITools using ecotag or metabinkit), in retrieving reads and exact sequence variants from the metabarcoding datasets. Overall, the MICCA based pipeline retrieved more reads, but less putative haplotypes of amphibians. After comparing these sequences with previously known haplotypes from tissue-based studies, when the aim is to both decrease the probability of detecting false haplotypes and retrieve the highest number of reads, we suggest using MICCA+VSEARCH, unless a direct comparison with tissue-based genetic data is possible.