Environmental DNA最新文献

{"title":"Sedimentary DNA Metabarcoding and Indigenous Knowledge Reconstruct Natural and Anthropogenic Disturbances to a Freshwater Lake in the Oil Sands Region of Alberta, Canada","authors":"Mark Louie D. Lopez,&nbsp;Ave Dersch,&nbsp;Paul Drevnick,&nbsp;Rute Clemente-Carvalho,&nbsp;Evan Morien,&nbsp;Christopher F. G. Hebda,&nbsp;Erin Ussery,&nbsp;Mark E. McMaster,&nbsp;Matthew A. Lemay,&nbsp;Caren C. Helbing","doi":"10.1002/edn3.70169","DOIUrl":"https://doi.org/10.1002/edn3.70169","url":null,"abstract":"<p>Sedimentary DNA (sedDNA), a form of environmental DNA (eDNA) shed by aquatic organisms and preserved in sediment, is crucial for reconstructing historical community compositions in aquatic ecosystems. In Cowpar Lake (Dene name: Doghostú), Alberta, a significant landslide event in the early 1940s CE impacted the lake's geochemistry and fish populations, as documented by Indigenous Knowledge from the Chipewyan Prairie First Nation and corroborated by targeted fish sedDNA analyses. The present study used 18S rRNA and cytochrome oxidase I (COI) genes for DNA metabarcoding of a sediment core from Cowpar Lake to assess the effect of the documented landslide and to reconstruct the historical community composition of eukaryotic functional trophic groups, including photoautotrophs, mixotrophs, parasites, and consumers. Between 1948 and 1956 CE, a notable shift in community composition occurred, with a decline in the alpha diversity of eukaryotic amplicon sequence variants. The increased primary productivity and terrestrial organic input post-1950 is correlated with an increased diversity of phototrophs and mixotrophs, suggesting potential algal blooms. While parasite diversity remained stable, consumer diversity declined, likely due to increased microbial respiration of organic matter, reducing oxygen levels and making the lake less hospitable for consumers like whitefish, which eventually disappeared in the lake. The reconstructed eukaryotic community profiles from sedDNA were consistent with Indigenous Knowledge of natural changes around the lake. The present study highlights the potential of braiding sedDNA data with Indigenous Knowledge to reconstruct long-term changes in aquatic communities, offering high-resolution baseline data for environmental monitoring and a deeper understanding of how freshwater systems respond to natural and human-induced impacts.</p>","PeriodicalId":52828,"journal":{"name":"Environmental DNA","volume":"7 4","pages":""},"PeriodicalIF":6.2,"publicationDate":"2025-08-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/edn3.70169","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144758553","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"How Fragmented Is eDNA? A Case Study on Shark DNA in Tropical Reef Seawater","authors":"Katrina M. West,&nbsp;Bruce Deagle","doi":"10.1002/edn3.70165","DOIUrl":"https://doi.org/10.1002/edn3.70165","url":null,"abstract":"<p>Environmental DNA (eDNA) is broadly assumed to be highly fragmented (&lt; 600 bp) in seawater. However, several marine eDNA studies that have successfully amplified longer fragments (from 600 up to 16,000 bp) are challenging this notion. We hypothesized that a small, yet amplifiable, proportion of eDNA templates contain fragment lengths exceeding 600 bp. To test this, we designed primers to target a series of mitochondrial fragment lengths between 119 and 15,727 bp for the tiger shark (<i>Galeocerdo cuvier</i>) and performed qPCR on seawater eDNA samples collected from the offshore, tropical Kimberley and Roebuck Marine Parks in Western Australia. We observed a steep decrease in eDNA copy number with increasing fragment size between 119 and 1518 bp, beyond which amplification was not successful. Importantly, we demonstrate that fragment sizes larger than conventionally targeted (e.g., 636, 840, and 1518 bp) can still be successfully amplified from seawater eDNA samples. Estimated mean nucleotide damage in seawater eDNA samples was found to be 3.9 breaks per 1000 bp; this equates to a mean undamaged fragment size of 256 bp and is less than damage observed in modern fecal DNA and ancient DNA. Characterizing the extent of eDNA fragmentation in various environmental samples will improve understanding of the genetic material available and enable practitioners to target standard length barcodes and longer hypervariable gene regions. Through the recovery of more informative data, eDNA applications will extend to finer-scale taxonomic resolution, including complex species and sub-species discrimination, as well as population analyses.</p>","PeriodicalId":52828,"journal":{"name":"Environmental DNA","volume":"7 4","pages":""},"PeriodicalIF":6.2,"publicationDate":"2025-08-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/edn3.70165","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144758554","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Using eDNA to Assess Freshwater Bacterial Diversity Along a Forest–Non-Forest Gradient in the Afrotropics","authors":"Zihui Wang,&nbsp;Vincent Munguleni,&nbsp;Innocent Kasekendi,&nbsp;Lauren J. Chapman,&nbsp;Marjorie Couton,&nbsp;Sadadi Ojoatre,&nbsp;Nick Atkinson,&nbsp;Florian Altermatt,&nbsp;T. Jonathan Davies","doi":"10.1002/edn3.70136","DOIUrl":"https://doi.org/10.1002/edn3.70136","url":null,"abstract":"<p>Healthy ecosystems are critical for maintaining ecosystem services and water security; yet many freshwater ecosystems have been subject to environmental degradation. Impacts are often greatest in water-scarce and developing regions, including across much of Sub-Saharan Africa, where many people lack access to basic drinking water. However, environmental monitoring programmes to track ecosystem health are generally lacking across this region due to limited resources and funding. Recent advances in environmental DNA (eDNA) methods offer an increasingly cost-effective and information-rich solution. Here, we explore the potential of eDNA as a tool for ecological monitoring of freshwater ecosystems in Uganda, East Africa. We sampled eDNA to quantify the bacterial diversity of rivers, streams, and swamps across a gradient of human disturbance in and around Kibale National Park, using off-the-shelf sampling methods that require minimal pre-existing infrastructure. We found distinct bacterial communities between intact and degraded habitats, but the bacterial community in rivers converged when flowing through intact forest. We identified several taxa with differential abundances that might serve as potential bioindicators of degraded ecosystems, and showed that a machine learning tool trained on eDNA can accurately differentiate between intact and degraded habitats. Our proof-of-concept study demonstrates the potential of eDNA as a practical and cost-effective biomonitoring tool for freshwater ecosystems in resource-limited regions, including Sub-Saharan Africa. We also highlight the potential benefits of protected forest in modulating bacterial composition in freshwater ecosystems.</p>","PeriodicalId":52828,"journal":{"name":"Environmental DNA","volume":"7 4","pages":""},"PeriodicalIF":6.2,"publicationDate":"2025-07-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/edn3.70136","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144740343","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Innovative Airborne DNA Approach for Monitoring Honey Bee Foraging and Health","authors":"Mateus Pepinelli,&nbsp;Alejandro José Biganzoli-Rangel,&nbsp;Katherine Lunn,&nbsp;Patrick Arteaga,&nbsp;Daniel Borges,&nbsp;Amro Zayed,&nbsp;Elizabeth L. Clare","doi":"10.1002/edn3.70150","DOIUrl":"https://doi.org/10.1002/edn3.70150","url":null,"abstract":"<p>Environmental DNA (eDNA) refers to genetic material collected from the environment and not directly from an organism. eDNA is best known as a tool in aquatic ecology but has been found associated with almost every substrate examined including soils, surfaces, and riding around on other animals. The collection of eDNA from air is one of the most recent advances and has been used to monitor a variety of organisms, including plants, animals, and microorganisms. Current evidence suggests a high turnover rate providing a recent signal for the presence of DNA associated with an organism. Here, we test whether material carried in air can be collected from honey bee hives to evaluate recent foraging behavior and colony health. We sampled air using purpose built “bee safe” air filters operating for 5–6 h at each colony. We successfully recovered plant, fungal and microbial DNA from the air within hives over a 3-week pilot period. From these data we identified the core honey bee microbiome and plant interaction data representing foraging behavior. We calculated beta diversity to estimate the effects of apiary sites and sampling date on data recovery. We observed that variance in ITS data was influenced by sampling date. Given that honey bees are generalist pollinators our ability to detect temporal signals in associated plant sequence data suggest this method opens new avenues into the ecological analysis of short term foraging behavior at the colony level. In comparison variance in microbial 16S sequencing data was more influenced by sampling location. As the assessment of colony health needs to be localized, spatial variance in these data indicate this may be an important tool in detecting infection. This pilot study demonstrates that colony air filtration has strong potential for the rapid screening of honey bee health and for the study of bee behavior.</p>","PeriodicalId":52828,"journal":{"name":"Environmental DNA","volume":"7 4","pages":""},"PeriodicalIF":6.2,"publicationDate":"2025-07-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/edn3.70150","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144725617","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The Government eDNA Working Group 7th eDNA Technical Exchange Workshop","authors":"Carol A. Stepien,&nbsp;Richard F. Lance","doi":"10.1002/edn3.70163","DOIUrl":"https://doi.org/10.1002/edn3.70163","url":null,"abstract":"<p>The 7th Government eDNA Working Group (GEDWG) Workshop featured an in-person and virtual technical exchange conference coordinated by the GEDWG on September 17–19, 2024, at the Columbus Zoo and Aquarium in Columbus, Ohio, USA. GEDWG is a no-cost consortium that brings together stakeholders associated with federal, state, provincial, municipal, and other government agencies, universities, and various nongovernmental entities interested in environmental DNA (eDNA) and related fields. GEDWG shares technical expertise and experience during monthly online discussion meetings and annual workshops. Approximately 80 participants attended the 7th Workshop in person, and over 130 additional participants attended virtually. The Workshop featured seven keynote plenary speakers, five presentation sessions, 26 platform talks, 11 posters, and an overall discussion session. Attendees included research scientists, natural resource managers, conservation policy experts, industry professionals, and representatives of trade organizations and nongovernmental organizations. Key takeaways from the Workshop included moving the application of eDNA into resource management and developing ways to improve policy uptake for nationwide and worldwide biodiversity monitoring, including eDNA standard practices, eDNA networks, and national strategies. Suggested research directions that merit further growth include comprehensive studies of eDNA fate and transport in different environments, autonomous sampling/sample processing, and reference library curation. Additionally, the codesign of studies and improved engagement and communication among scientists, resource managers, and industry are needed to ensure clear expectations and outcomes and move forward with biodiversity assessments.</p>","PeriodicalId":52828,"journal":{"name":"Environmental DNA","volume":"7 4","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/edn3.70163","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144716691","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Isolation and Genetic Identification of Metazoan Cells From Seawater","authors":"Haylea Power,&nbsp;Mitchell J. O'Brien,&nbsp;Miwa Takahashi,&nbsp;Henry Hui,&nbsp;Kathy Fuller,&nbsp;Darren Korbie,&nbsp;Oliver Berry,&nbsp;Simon Jarman","doi":"10.1002/edn3.70154","DOIUrl":"https://doi.org/10.1002/edn3.70154","url":null,"abstract":"<p>Environmental DNA (eDNA) metabarcoding typically relies on collecting and characterising a pool of mixed, fragmented DNA from environmental samples for species identification. Here, we introduce environmental metazoan cells (emCells), representing whole individual cells shed by macro-organisms into aquatic ecosystems, and report on a method to successfully isolate and amplifying short amplicons to determine species identity. Using a custom fish probe and a novel multi-factor fluorescence-activated cell sorting (FACS) protocol on mesocosm water samples, we successfully enriched for target emCells, as confirmed by shifts in population density using FACS and imaging flow cytometry. Imaging flow cytometry demonstrated dual nuclear and mitochondrial staining of whole single cells, while multiplexed PCR assays (targeting both mitochondrial and nuclear DNA) confirmed the effective enrichment of fish emCells, with one-quarter of sorted cells identified as fish. Sequences obtained from isolated emCells matched known species in the mesocosm, validating our approach. Despite efforts to exclude non-target cells, diverse single-cell eukaryotes were also recovered, highlighting the need for additional strategies to enrich for target emCells given the abundance and diversity of off-target particles present in aquatic environments, which will be especially important for real-world environments. Isolation and analysis of emCells could provide a versatile complementary approach to current eDNA methodologies by providing genomic information that normally requires direct sampling from live organisms.</p>","PeriodicalId":52828,"journal":{"name":"Environmental DNA","volume":"7 4","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/edn3.70154","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144714876","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"DNA Metabarcoding as a Complementary Approach to Traditional Surveys for Monitoring the Plant Diversity in the Amazon canga","authors":"Vitória C. C. Martins,&nbsp;Gisele L. Nunes,&nbsp;Renato R. M. Oliveira,&nbsp;Markus Gastauer,&nbsp;Guilherme Oliveira,&nbsp;Santelmo Vasconcelos","doi":"10.1002/edn3.70155","DOIUrl":"https://doi.org/10.1002/edn3.70155","url":null,"abstract":"<p>The <i>canga</i> of the Serra dos Carajás in the Eastern Amazon (Pará, Brazil) has one of the largest iron ore deposits on the planet and is home to a community of endemic and rare plants. However, conservation and monitoring programs in megadiverse areas, as in the case of the region, are often hampered by the lack of knowledge of the species that inhabit these ecosystems. In this scenario, the comprehensive DNA barcoding effort directed to the complete flora of the <i>canga</i> in the Brazilian Amazon has enabled the implementation of DNA metabarcoding approaches for species monitoring. Here, we assessed the potential of implementing DNA metabarcoding with environmental DNA (eDNA) in future surveys of plant species of the ironstone outcrops of the Serra dos Carajás. After extracting eDNA from soil samples, the nuclear ITS2 region was amplified and sequenced using the Illumina MiSeq platform. With the metabarcoding analyses, we detected 95 species from 72 genera and 35 families, revealing a higher overall diversity than the morphology-based approach, including taxa that were not identified in a traditional floristic survey. The fact that DNA metabarcoding results mostly agreed with the data from the floristic survey indicates the robustness of the molecular approach to be used in monitoring studies of plant diversity in the region. Additionally, we discuss the relevance of our results to guide the development of broader applications of eDNA-based biodiversity monitoring in species-rich environments such as the Serra dos Carajás.</p>","PeriodicalId":52828,"journal":{"name":"Environmental DNA","volume":"7 4","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/edn3.70155","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144714875","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Dietary Differentiation Between Sympatric Ecotypes of Astatotilapia calliptera From Lake Masoko (Kisiba), Tanzania Revealed by Metabarcoding","authors":"Kirthana Pillay,&nbsp;Simon Creer,&nbsp;Alexandra M. Tyers,&nbsp;Enrica Gargiulo,&nbsp;William Perry,&nbsp;Georgina Brennan,&nbsp;Deborah A. Dawson,&nbsp;Kathryn H. Maher,&nbsp;Gavin J. Horsburgh,&nbsp;Msafiri Andrew,&nbsp;George Turner","doi":"10.1002/edn3.70146","DOIUrl":"https://doi.org/10.1002/edn3.70146","url":null,"abstract":"<p>Sympatric speciation is defined as the formation of new species in the absence of geographic barriers, but the genomic and life history strategy mechanisms underpinning sympatric speciation are still far from clear. It has recently been discovered that the cichlid fish <i>Astatotilapia calliptera</i> from crater Lake Masoko in Tanzania have diverged sympatrically into littoral (shallow water) and benthic (deep water) ecotypes, which differ in head and pharyngeal jaw morphology. Carbon stable isotope analysis has also broadly indicated trophic differentiation between ecotypes. Here, we explore trophic niche divergence on a finer scale, using metabarcoding of stomach contents. A combination of the mitochondrial COI region and 18S V4 region from the eukaryotic nuclear small subunit ribosomal DNA was used to target macroinvertebrate and broader eukaryotic taxonomic diversity, respectively, revealing dietary divergence between the ecotypes. Large proportions of Arthropoda (dipterans and copepod) were found in both ecotypes, indicating some food sources common to both microhabitats. However, gut contents of benthic <i>A. calliptera</i> individuals were characterized by an abundance of annelids and diatoms, while Lepidoptera, mayflies, fungi, freshwater mussels, and bivalves were common in littoral ecotypes. The variation observed in the dietary contents of the ecotypes indicates the presence of resource partitioning, facilitating adaptation to unique feeding strategies.</p>","PeriodicalId":52828,"journal":{"name":"Environmental DNA","volume":"7 4","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/edn3.70146","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144705152","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Tracking Ungulate Diet: Comparing Observational and DNA Metabarcoding Tools","authors":"Forman Roi,&nbsp;Berman Tali Sarah,&nbsp;Zhong Zhiwei,&nbsp;Dines Monica,&nbsp;Wang Deli,&nbsp;Inbar Moshe","doi":"10.1002/edn3.70157","DOIUrl":"https://doi.org/10.1002/edn3.70157","url":null,"abstract":"<p>Large mammalian herbivores (LMH) are abundant in grazing ecosystems and play a pivotal role in shaping vegetation characteristics. However, accurately determining their diets through traditional methods, such as direct observations, remains challenging, particularly in natural communities and mixed-species grazing systems. Recent studies have shown that DNA metabarcoding can effectively identify the plant composition in LMH diets as well as the plant-dwelling arthropods (PDA) incidentally ingested by LMH while grazing. Given the high specificity of herbivorous insects to their host plant, we hypothesize that DNA metabarcoding of arthropods ingested by LMH could offer valuable insights into their feeding preferences. The goal of this study is to evaluate the accuracy of plant and arthropod DNA metabarcoding methods in identifying the diets of sheep and cattle and to compare their performance with direct observations and known dietary patterns from the literature. To test this, we collected fecal samples from sheep and cattle grazing in the northeast Asian grasslands. We amplified arthropod DNA using COI mitochondrial markers and plant DNA using ITS1 markers, followed by Illumina sequencing. Additionally, we conducted field observations to identify plants grazed by sheep and cattle. The DNA metabarcoding methods provided a comprehensive view of the LMH diet. Both DNA metabarcoding methods successfully detected dietary differences between sheep and cattle, with sheep primarily consuming nutrient-rich forbs and cattle predominantly grazing on Poaceae, consistent with known foraging behaviors. While the constant presence of arthropods across multiple samples suggests that DNA of ingested arthropods could provide complementary information regarding LMH foraging behavior, we found such to be rather limited. However, our findings confirm that plant DNA metabarcoding is a reliable and accurate method for identifying LMH diets.</p>","PeriodicalId":52828,"journal":{"name":"Environmental DNA","volume":"7 4","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/edn3.70157","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144705151","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Evaluating eDNA Metabarcoding Techniques for Pollinator Community Assessment in a Field and Controlled Experiment","authors":"Stephanie S. Coster,&nbsp;Morgane Pommier,&nbsp;Nicholas J. Ruppel","doi":"10.1002/edn3.70162","DOIUrl":"https://doi.org/10.1002/edn3.70162","url":null,"abstract":"<p>Pollinators play a critical role in ensuring the stability of food systems, yet their populations are in decline. To better understand and promote pollinator biodiversity, this study explored the use of environmental DNA (eDNA) metabarcoding techniques to assess plant–pollinator interactions. We conducted two experiments to optimize eDNA metabarcoding strategies for detecting pollinators on flowering plants. In the first experiment, we compared visual observations and eDNA detection via Illumina sequencing to characterize pollinator visitation on two native plant species in public and private green spaces across the Richmond, Virginia metropolitan area. Our findings revealed notable differences between the two methods, with visual surveys more sensitive to Hymenoptera and eDNA more sensitive to plant pests and other organisms. We devised a second experiment in a controlled environment at the Lewis Ginter Botanical Garden butterfly exhibit. Here, we tested different sampling strategies, primer pairs, and DNA sequencing platform (using Oxford Nanopore Technology). Despite detecting two butterfly species present in the exhibit, the eDNA metabarcoding showed limited sensitivity to the expected Lepidoptera. Our results indicate that while eDNA metabarcoding can detect a broader range of eukaryotic organisms, it may not be as effective for monitoring specific pollinator taxa like Hymenoptera and Lepidoptera. Factors such as sample type, primer bias, sequencing platform, and bioinformatics pipeline may impact detection outcomes. This study underscores the need for combining traditional monitoring techniques with eDNA metabarcoding to gain a comprehensive understanding of plant–pollinator interactions and improve pollinator conservation efforts.</p>","PeriodicalId":52828,"journal":{"name":"Environmental DNA","volume":"7 4","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-07-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/edn3.70162","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144688154","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}