ABSTRACT The Eurasian otter ( Lutra lutra ) is a semiaquatic, freshwater top predator of conservation concern due to its significant global population decline in the last century. Following its recovery and recolonization in parts of Europe, the population faces ongoing threats, including habitat loss and fragmentation due to human activities, climate change, prey depletion, and exposure to emerging contaminants. Accurately identifying its diet is crucial for understanding ecological interactions and proposing effective management strategies. DNA metabarcoding based on high‐throughput sequencing is a reliable diet analysis tool with better resolution than traditional methods. This study, for the first time, applied two 16S rRNA primers targeting different prey classes on the dietary spectrum of two Eurasian otters populations returning to Northern Germany (Schleswig‐Holstein and Lower Saxony). DNA fragments of prey items from the gut contents of 156 otters collected between 2005 and 2022 were sequenced. Fifty‐five operational taxonomic units (OTUs) were identified in the dietary remains, including freshwater and marine fish, anurans, caudates, and water birds. Otters in the study area are opportunistic predators with wide dietary plasticity, feeding on the most abundant species. Nine‐spined sticklebacks, common roach, and common frogs were important prey items, and marine fish species were detected in the diet of some of the investigated otters. The otters' diet differed between Lower Saxony and Schleswig‐Holstein, between sexes, and a slight difference was observed between adults and subadults. This study provides new insights into the diet composition of otters and their ability to exploit a variety of resources in their habitat. This information is vital for understanding their chances of survival in an environment that is continuously changing due to human impacts and for developing effective management tools.
ABSTRACT Effective management of fecal contamination in coastal areas requires identifying the biological sources of contamination to protect human and ecosystem health. Current microbial source tracking (MST) methods experience limitations in source identification and/or reliability. Therefore, a need exists for robust next generation source tracking methods to improve understanding of fecal sources. Environmental DNA (eDNA) metagenomics is an approach that enables sampling of organisms without physically coming into contact with or observing species and can also provide valuable ecological context to field sites. Despite its advantages, metagenomic MST has not yet been fully implemented and remains uncommon in routine monitoring programs. The eDNA Source Tracking Toolkit was created to serve as a framework that enables community partners and stakeholders to implement this technology on local, need‐based scales, increasing impact and benefiting communities. The Toolkit contains recommendations, protocols, and best practices for sample processing, sequencing, bioinformatics, and data visualization, as well as a comprehensive list of supplies needed for implementing each step of the eDNA Source Tracking Toolkit workflow. An eDNA Source Tracking Toolkit Workshop was hosted at the University of South Alabama in Mobile, AL to introduce local researchers and community stakeholders to the Toolkit and increase use of this new MST tool. Workshop participants were surveyed before, immediately after, and 6 months after the workshop to gauge utility of the Workshop and knowledge retention. Overall, the Workshop and Toolkit were well received, with the majority of participants reporting learning new skills, gaining new information, and stimulating new ideas or perspectives. This study exemplifies how structured resources and training frameworks can lower barriers to adopting next generation eDNA approaches in water quality monitoring.
ABSTRACT Kelp farming in the North Atlantic is emerging as a new sustainable source of food, feed, nutraceuticals and biostimulants. Kelp biofouling by epibiont taxa, including bryozoans, hydrozoans, amphipods, bivalves, and gastropods, is a key constraint in kelp yield and quality. The planktonic stages of these species belonging to meroplankton precede visible settlement on seaweed fronds. Thus, the precise taxonomic detection of the planktonic stage and the identification of the temporal lag with the biofouling stage could help form an early warning signal helping farmers plan harvesting times. To achieve this, monthly samples of plankton and kelp fronds were collected from a Scottish kelp farm during a kelp production period (September 2021–July 2022). The taxonomic identity of kelp biofouling epibionts was established by microscopy and DNA‐barcoding. The taxa were then traced in the plankton samples using both microscopy counts and eDNA‐metabarcoding. COI barcoding identified 14 kelp epibiont species, sharpening the morphological classification which only reached the class or genus level. Plankton microscopy confirmed two of these at the genus level while plankton eDNA metabarcoding recovered 9 of the 14 species and detected an additional 14 taxa. Microscopy counts of hydromedusae, bryozoan cyphonautes, and bivalve larvae peaked in the plankton 2 weeks before the maximum coverage on Saccharina latissima fronds. Metabarcoding and microscopy counts frequently identified different peak months for the same taxa. Depth‐stratified frond surveys revealed that kelp segments located deeper within the water column supported the highest levels of hydrozoan colonization, indicating either depth‐related colonization processes, kelp age‐related exposure effects, or UV avoidance. The results indicate that the combination of molecular diagnostics and conventional microscopy improves the sensitivity and specificity of biofouling detection, facilitating earlier and more informed farm management decisions.
ABSTRACT Analyses of environmental DNA (eDNA), that is, DNA obtained from environmental samples, has emerged as a highly promising tool for monitoring fish communities. While the ability to obtain quantitative estimates from eDNA metabarcoding surveys remains debatable; recent efforts have aimed to enhance the quantitative nature. Comparative studies assessing the impact of sampling strategies, refinements to molecular and data analyses protocols are, however, lacking. We compared two common sampling strategies (i.e., spatially integrated sampling vs. point sampling) in two natural lakes and six approaches designed to improve the quantitative interpretation of eDNA metabarcoding data. Metabarcoding estimates of eDNA concentrations were correlated against standard gillnet fishing estimates of species' abundance and biomass, absolute eDNA quantifications for five target species obtained through digital droplet PCR analyses, and the allometrically scaled abundance (i.e., correcting for the non‐linear relationship between eDNA production and body mass). Results showed that a spatially integrated sampling strategy and the use of internal standards to convert read counts to eDNA concentrations increases the probability of detecting positive correlations between eDNA concentrations and species' abundance and biomass estimates and increases model predictive performance. To our knowledge, we provide the first comparative study to determine the optimal strategies to move toward a quantitative interpretation of fish eDNA metabarcoding surveys. These findings are essential to pave the way for routine implementation of eDNA metabarcoding surveys.
ABSTRACT Airborne environmental DNA (eDNA) is an emerging tool for biodiversity assessment, enabling the detection of organisms from biological particles suspended in the air. Recent studies have demonstrated its potential for detecting diverse taxonomic groups, with applications in conservation, biosecurity, and ecological monitoring. However, given the early stage of development and the diversity of collection approaches, it remains unclear how key sampling design parameters influence airborne eDNA collection, detection, and ecological inference. Here, we use a simple, low‐cost, 3D‐printable passive collection device to systematically evaluate the effects of filter surface area, sampling duration, and post‐collection storage on target‐species detection and community recovery. Using species‐specific qPCR targeting European rabbit ( Oryctolagus cuniculus ), shingleback lizard ( Tiliqua rugosa ), and tiger ( Panthera tigris ), alongside general vertebrate 12SV5 and mammal‐specific Mam1/2 metabarcoding, we found that smaller filters generally improved detection sensitivity, recovered signal increased rapidly over short deployment periods before showing diminishing gains, and dry storage for 28 days had only modest effects on detection outcomes. Zoo deployments recovered ecologically plausible signals, although zoo‐held taxa were detected inconsistently across replicate filters. Together, these findings demonstrate that sampling design can substantially influence detection within passive airborne eDNA workflows and provide practical guidance for developing scalable and reproducible monitoring strategies.
