Deciphering environmental factors influencing phytoplankton community structure in a polluted urban river.

Tuojiang River Basin is a first-class tributary of the upper reaches of the Yangtze River-which is the longest river in China. As phytoplankton are sensitive indicators of trophic changes in water bodies, characterizing phytoplankton communities and their growth influencing factors in polluted urban rivers can provide new ideas for pollution control. Here, we used direct microscopic count and environmental DNA (eDNA) metabarcoding methods to investigate phytoplankton community structure in Tuojiang River Basin (Chengdu, Sichuan Province, China). The association between phytoplankton community structure and water environmental factors was evaluated by Mantel analysis. Additional environmental monitoring data were used to pinpoint major factors that influenced phytoplankton growth based on structural equation modeling. At the phylum level, the dominant phytoplankton taxa identified by the conventional microscopic method mainly belonged to Bacillariophyta, Chlorophyta, and Cyanophyta, in contrast with Chlorophyta, Dinophyceae, and Bacillariophyta identified by eDNA metabarcoding. In α-diversity analysis, eDNA metabarcoding detected greater species diversity and achieved higher precision than the microscopic method. Phytoplankton growth was largely limited by phosphorus based on the nitrogen-to-phosphorus ratios > 16:1 in all water samples. Redundancy analysis and structural equation modeling also confirmed that the nitrogen-to-phosphorus ratio was the principal factor influencing phytoplankton growth. The results could be useful for implementing comprehensive management of the river basin environment. It is recommended to control the discharge of point- and surface-source pollutants and the concentration of dissolved oxygen in areas with excessive nutrients (e.g., Jianyang-Ziyang). Algae monitoring techniques and removal strategies should be improved in 201 Hospital, Hongrihe Bridge and Colmar Town areas.

Faecal source apportionment using molecular methods: A proof of concept using the FEAST algorithm.

Faecal contamination of freshwater and marine environments represents a significant risk for public health, recreational activity and food safety, and tools for evaluating complex multi-source contamination remain largely in the development phase. We evaluated the efficacy of the Fast Expectation Maximization (FEAST) microbial source tracking (MST) algorithm to apportion sources of faecal contamination among four mammalian species of interest in coastal waters in New Zealand. Using 16S ribosomal DNA metabarcoding of faecal samples from cows, fur seals, and sheep, as well as human wastewater, we aimed to differentiate and quantify the contribution of these sources in mixed faecal samples. Multivariate analysis confirmed significant differences in the microbial communities associated with each mammalian source, with specific bacterial classes indicative of different sources. The FEAST algorithm was tested using mixed DNA and mixed faecal samples, and we found that the algorithm correctly assigned the dominant source from all samples, but underestimated the dominant source's proportional contribution. This underestimation suggests the need for further refinement and validation to ensure accurate source apportionment in environmental samples where the faecal signal is likely to be a minor component. Despite these limitations, the findings of our study, in combination with the evidence from others who have tested the FEAST algorithm in environmental settings, indicates that it represents an advance on existing tools for microbial source tracking and may become a useful addition to the toolbox for environmental management.

Environmental DNA metabarcoding reveals the influence of environmental heterogeneity on microeukaryotic plankton in the offshore waters of East China Sea.

Microeukaryotic plankton are essential to marine food webs and biogeochemical cycles, with coastal seas playing a critical role in aquatic ecosystems. Understanding the diversity of microeukaryotic plankton, deciphering their community structure and succession patterns, and identifying the key factors influencing these dynamics remain central challenges in coastal ecology. In this study, we examine patterns of biodiversity, community structure, and co-occurrence using environmental DNA (eDNA)-based methods. Our results show a linear correlation between α-diversity and distance from the shore, with nutrient-related factors, especially inorganic nitrogen, being the primary determinants of the spatial distribution of plankton communities. Alternation of coastal habitat have shifted the succession patterns of coastal eukaryotic plankton communities from stochastic to deterministic processes. Additionally, our observations indicate that the topology and structure of eukaryotic plankton symbiotic patterns and networks are significantly influenced by environmental heterogeneity such as nutrients, which increase the vulnerability and decrease the stability of offshore ecological networks. Overall, our study demonstrates that the distribution of microeukaryotic plankton communities is influenced by factors related to environmental heterogeneity.

Application of Synchronous Evaluation-Diagnosis Model with Quantitative Stressor-Response Analysis (SED-QSR) to Urban Lake Ecological Status: A Proposed Multiple-Level System.

Ecological integrity assessment and degradation diagnosis are used globally to evaluate the health of water bodies and pinpoint critical stressors. However, current studies mainly focus on separate evaluation or diagnosis, leading to an inadequate exploration of the relationship between stressors and responses. Here, based on multiple data sets in an urban lake system, a synchronous evaluation-diagnosis model with quantitative stressor-response analysis was advanced, aiming to improve the accuracy of evaluation and diagnosis. The weights for key physicochemical stressors were quantitatively determined in the sequence of NDAVIadj > CODMn > TP > NH4+-N by the combination of generalized additive model and structural equation modeling, clarifying the most significant effects of aquatic vegetation on the degradation of fish assemblages. Then, sensitive biological metrics were screened by considering the distinct contributions of four key stressors to alleviate the possible deviation caused by common methods. Finally, ecological integrity was evaluated by summing the key physicochemical stressors and sensitive biological metrics according to the model-deduced weights instead of empirical weights. Our system's diagnosis and evaluation results achieved an accuracy of over 80% when predicting anthropogenic stress and biological status, which highlights the great potential of our multiple-level system for ecosystem management.

Stability of environmental DNA methylation and its utility in tracing spawning in fish.

The use of environmental DNA (eDNA) is becoming prevalent as a novel method of ecological monitoring. Although eDNA can provide critical information on the distribution and biomass of particular taxa, the DNA sequences of an organism remain unaltered throughout its existence, which complicates the accurate identification of crucial events, including spawning. Therefore, we examined DNA methylation as a novel source of information from eDNA, considering that the methylation patterns in eggs and sperm released during spawning differ from those of somatic tissues. Despite its potential applications, little is known about eDNA methylation, including its stability and methods for detection and quantification. Therefore, we conducted tank experiments and performed methylation analysis targeting 18S rDNA through bisulphite amplicon sequencing. In the target region, eDNA methylation was not affected by degradation and was equivalent to the methylation rate of genomic DNA from somatic tissues. Unmethylated DNA, abundant in the ovaries, was detected in the eDNA released during fish spawning. These results indicate that eDNA methylation is a stable signal reflecting targeted gene methylation and further demonstrate that germ cell-specific methylation patterns can be used as markers for detecting fish spawning.

Scanning amplicons with CRISPR-Dx detects endangered amphibians in environmental DNA.

More efficient methods for extensive biodiversity monitoring are required to support rapid measures to address the biodiversity crisis. While environmental DNA (eDNA) metabarcoding and quantitative PCR (qPCR) methods offer advantages over traditional monitoring approaches, their large-scale application is limited by the time and labour required for developing assays and/or for analysis. CRISPR (clustered regularly interspaced short palindromic repeats) diagnostic technologies (Dx) may overcome some of these limitations, but they have been used solely with species-specific primers, restricting their versatility for biodiversity monitoring. Here, we demonstrate the feasibility of designing species-specific CRISPR-Dx assays in silico within a short metabarcoding fragment using a general primer set, a methodology we term 'ampliscanning', for 18 of the 22 amphibian species in Switzerland. We sub-selected nine species, including three classified as regionally endangered, to test the methodology using eDNA sampled from ponds at nine sites. We compared the ampliscanning detections to data from traditional monitoring at these sites. Ampliscanning was successful at detecting target species with different prevalences across the landscape. With only one visit, we detected more species per site than three traditional monitoring visits (visual and acoustic detections by trained experts), in particular more elusive species and previously undocumented but expected populations. Ampliscanning detected 25 species/site combinations compared to 12 with traditional monitoring. Sensitivity analyses showed that larger numbers of field visits and PCR replicates are more important for reliable detection than many technical replicates at the CRISPR-Dx assay level. Given the reduced sampling and analysis effort, our results highlight the benefits of eDNA and CRISPR-Dx combined with universal primers for large-scale monitoring of multiple endangered species across landscapes to inform conservation measures.

Is it worthy to use environmental DNA instead of scientific trawling or video survey to monitor taxa in soft-bottom habitats?

Non-extractive techniques such as video analysis are increasingly used by scientists to study marine communities instead of extractive methods such as trawling. Currently, environmental DNA (eDNA) analysis is seen as a revolutionary tool to study taxonomic diversity. We aimed to determine which method is the most appropriate to describe fish and commercial invertebrate diversity comparing bottom trawl hauls, video transects and seawater eDNA. Our results reveal that video detected the lowest number of taxa and trawling the highest. eDNA analysis is powerful to describe marine bony fish communities, but some taxa of importance for the ecosystem such as elasmobranchs, crustaceans or molluscs are poorly detected. This may be due to several factors such as marker specificity, incomplete reference gene databases or low DNA release in the environment. For now, the various methods provide different information and none is exhaustive enough to be used alone for biodiversity characterisation.

Analyzing the Interaction between Tetrahymena pyriformis and Bacteria under Different Physicochemical Conditions When Infecting Guppy Using the eDNA Method.

In the aquaculture system of ornamental fish, the interaction between bacterial microbiota and ciliate protozoa can prevent or promote disease outbreaks, and different physicochemical conditions will affect the relationships between them. We investigated the interaction between bacterial microbiota and the parasite Tetrahymena pyriformis when infecting Poecilia reticulata (guppy) under different physicochemical conditions. The abundance of T. pyriformis in water, the relative abundance of bacterial species, and histopathological observation were studied or monitored using environmental DNA (eDNA) extraction technology, the qPCR method, and 16s rRNA sequencing, respectively. The morphological identification and phylogenetic analysis of T. pyriformis were carried out. The infected guppy tissue was also stained by the hematoxylin and eosin methods. The results showed: (1) the bacterial communities of water samples were mainly composed of species assigned to Proteobacteria and Bacteroidetes, and Tabrizicola and Puniceicoccaceae were positively correlated with fish mortality, T. pyriformis abundance, and temperature. (2) Arcicella and Methyloversatilis universalis with different correlations between ciliates appeared in different treatment groups, the result of which proved that environmental factors affected the interaction between bacteria and T. pyriformis. (3) Lower temperatures and a higher pH were more beneficial for preventing disease outbreaks.

Measuring air metagenomic diversity in an agricultural ecosystem.

All species shed DNA during life or in death, providing an opportunity to monitor biodiversity via environmental DNA (eDNA). In recent years, combining eDNA, high-throughput sequencing technologies, bioinformatics, and increasingly complete sequence databases has promised a non-invasive and non-destructive environmental monitoring tool. Modern agricultural systems are often large monocultures and so are highly vulnerable to disease outbreaks. Pest and pathogen monitoring in agricultural ecosystems is key for efficient and early disease prevention, lower pesticide use, and better food security. Although the air is rich in biodiversity, it has the lowest DNA concentration of all environmental media and yet is the route for windborne spread of many damaging crop pathogens. Our work suggests that ecosystems can be monitored efficiently using airborne nucleic acid information. Here, we show that the airborne DNA of microbes can be recovered, shotgun sequenced, and taxonomically classified, including down to the species level. We show that by monitoring a field growing key crops we can identify the presence of agriculturally significant pathogens and quantify their changing abundance over a period of 1.5 months, often correlating with weather variables. We add to the evidence that aerial eDNA can be used as a source for biomonitoring in terrestrial ecosystems, specifically highlighting agriculturally relevant species and how pathogen levels correlate with weather conditions. Our ability to detect dynamically changing levels of species and strains highlights the value of airborne eDNA in agriculture, monitoring biodiversity changes, and tracking taxa of interest.

Human activities strengthen the influence of deterministic processes in the mechanisms of fish community assembly in tropical rivers of Yunnan, China.

Human-induced global alterations have worsened the severe decrease in fish biodiversity in rivers. To successfully reduce the pace of reduction in fish diversity, it is crucial to prioritize the understanding of how human activities impact the processes that shape and maintain fish diversity. Traditional fish survey methods are based on catch collection and morphological identification, which is often time-consuming and ineffective. Hence, these methods are inadequate for conducting thorough and detailed large-scale surveys of fish ecology. The rapid progress in molecular biology techniques has transformed environmental DNA (eDNA) technique into a highly promising method for studying fish ecology. In this work, we conducted the first systematic study of fish diversity and its formation and maintenance mechanism in the Xishuangbanna section of the Lancang River using eDNA metabarcoding. The eDNA metabarcoding detected a total of 159 species of freshwater fishes spanning 13 orders, 34 families, and 99 genera. The fishes in the order cypriniformes were shown to be overwhelmingly dominant. At different intensities of anthropogenic activity, we found differences in fish community composition and assembly. The analysis of the Sloan's neutral community model fitting revealed that stochastic processes were the dominant factor in the shaping of fish communities in the Xishuangbanna section of the Lancang River. We have further confirmed this result by using the phylogenetic normalized stochasticity ratio. Furthermore, our findings indicate that as human activities get more intense, the influence of stochastic processes on the shaping of fish communities decreases, while the influence of deterministic processes eventually becomes more prominent. Finally, we discovered that salinity positively correlated with fish community changes in the high-intensity anthropogenic sample sites, but all environmental factors had little effect on fish community changes in the low-intensity and moderate-intensity anthropogenic sample sites. Our study not only validated the potential application of eDNA metabarcoding for monitoring fish diversity in tropical rivers, but also revealed how fish communities respond to human activities. This knowledge will serve as a solid foundation for the protection of fish resources in tropical rivers.

The Diatom Diversity and Ecological Status of a Tufa-Depositing River through eDNA Metabarcoding vs. a Morphological Approach-A Case Study of the Una River (Bosnia and Herzegovina).

Tufa deposits in karst rivers are unique habitats created by mutual interactions between specific environmental and biotope features and inhabited by diatoms as a highly abundant and diverse algal group. This pilot study aimed to investigate the diversity of diatom communities on tufa depositing habitats and assess the Una River's ecological status using a comparative molecular and morphological approach for diatom identification. The 312 base pairs of the rbcL gene were barcoded and analyzed using MiSeq reads and amplicon sequence variants (ASVs) obtained by the DADA2 pipeline. The reference database Diat.barcode v7 was used for taxonomic assignment. The morphological identification of the diatoms was carried out in parallel. In total, the combined dataset revealed 46 taxa identified at genus rank, 125 on the subgenus, and 145 on combined taxonomy rank. The metabarcoding approach mostly leads to a lower number of identified taxa at species rank (58 in molecular vs. 119 in optical inventory), resulting in higher values of beta diversity and heterogeneity in diatom assemblages in samples obtained by morphological approach. Despite the high percentage of taxonomically not assigned diatom ASVs to the species rank, high Shannon diversity index values and a similar number of taxa per locations compared to the morphological approach were obtained. Taxa Achnanthidium minutissimum (Kützing) Czarnecki, Achnanthidium pyrenaicum (Hustedt) H.Kobayasi, Amphora pediculus (Kützing) Grunow, Diatoma vulgaris Bory, Navicula cryptotenella Lange-Bertalot, and Navicula tripunctata (O.F.Müller) Bory were identified at all locations in both inventories. Although limited consistency in the diatom abundances between the two inventory datasets was found, a similar grouping of samples was observed connected to the river's longitudinal gradient. The data obtained using molecular approach in most sites indicated a mostly lower ecological status (good or moderate) compared to the data obtained from the morphological approach (high, good, and moderate). The potential of environmental DNA (eDNA) diatom metabarcoding for water monitoring and diversity studies is undeniable, but to fully realize the benefits of these methods in the future, it is essential to standardize protocols and expand the reference database for species found in specific habitats, such as tufa deposits.

Effects of season and water quality on community structure of planktonic eukaryotes in the Chaohu Lake Basin.

Introduction: Analyzing the correlation between planktonic eukaryotic communities (PECs) and aquatic physicochemical parameters (APPs) provides important references for predicting the impact of climate change and human activities on aquatic ecosystems. Methods: To assess the influence of seasons and APPs on PEC structures in lakes and rivers, we utilized high-throughput sequencing of the 18S rRNA gene to analyze PEC structures in a lake and seven rivers in the Chaohu Lake Basin and analyzed their correlations with APPs. Results: Our results revealed that PEC structure was significantly affected by season, with the highest α-diversity observed in summer. Furthermore, we identified several APPs, including water temperature, conductivity, dissolved oxygen, pH, phosphate, total phosphorus, trophic level index (TLI), nitrate, ammonia nitrogen, and total nitrogen, that significantly influenced PEC structures. Specifically, we found that Stephanodiscus hantzschii, Simocephalus serrulatus, Cryptomonas sp. CCAC_0109, Pedospumella encystans, Actinochloris sphaerica, Chlamydomonas angulosa, Gonyostomum semen, Skeletonema potamos, Chlamydomonas klinobasis, Pedospumella sp., and Neochlorosarcina negevensis were significantly correlated to TLI, while Limnoithona tetraspina, Theileria sp., and Pseudophyllomitus vesiculosus were significantly correlated to the water quality index (WQI). However, our random forest regression analysis using the top 100 species was unable to accurately predict the WQI and TLI. Discussion: These results provide valuable data for evaluating the impact of APPs on PEC and for protecting water resource in the Chaohu Lake Basin.

Phenological mismatches mitigate the ecological impact of a biological invader on amphibian communities.

Horizon scans have emerged as a valuable tool to anticipate the incoming invasive alien species (IAS) by judging species on their potential impacts. However, little research has been conducted on quantifying actual impacts and assessing causes of species-specific vulnerabilities to particular IAS due to persistent methodological challenges. The underlying interspecific mechanisms driving species-specific vulnerabilities therefore remain poorly understood, even though they can substantially improve the accuracy of risk assessments. Given that interspecific interactions underlying ecological impacts of IAS are often shaped by phenological synchrony, we tested the hypothesis that temporal mismatches in breeding phenology between native species and IAS can mitigate their ecological impacts. Focusing on the invasive American bullfrog (Lithobates catesbeianus), we combined an environmental DNA (eDNA) quantitative barcoding and metabarcoding survey in Belgium with a global meta-analysis, and integrated citizen-science data on breeding phenology. We examined whether the presence of native amphibian species was negatively related to the presence or abundance of invasive bullfrogs and whether this relationship was affected by their phenological mismatches. The field study revealed a significant negative effect of increasing bullfrog eDNA concentrations on native amphibian species richness and community structure. These observations were shaped by species-specific vulnerabilities to invasive bullfrogs, with late spring- and summer-breeding species being strongly affected, while winter-breeding species remained unaffected. This trend was confirmed by the global meta-analysis. A significant negative relationship was observed between phenological mismatch and the impact of bullfrogs. Specifically, native amphibian species with breeding phenology differing by 6 weeks or less from invasive bullfrogs were more likely to be absent in the presence of bullfrogs than species whose phenology differed by more than 6 weeks with that of bullfrogs. Taken together, we present a novel method based on the combination of aqueous eDNA quantitative barcoding and metabarcoding to quantify the ecological impacts of biological invaders at the community level. We show that phenological mismatches between native and invasive species can be a strong predictor of invasion impact regardless of ecological or methodological context. Therefore, we advocate for the integration of temporal alignment between native and IAS's phenologies into invasion impact frameworks.

Taxonomic and abundance biases affect the record of marine eukaryotic plankton communities in sediment DNA archives.

Environmental DNA (eDNA) preserved in marine sediments is increasingly being used to study past ecosystems. However, little is known about how accurately marine biodiversity is recorded in sediment eDNA archives, especially planktonic taxa. Here, we address this question by comparing eukaryotic diversity in 273 eDNA samples from three water depths and the surface sediments of 24 stations in the Nordic Seas. Analysis of 18S-V9 metabarcoding data reveals distinct eukaryotic assemblages between water and sediment eDNA. Only 40% of Amplicon Sequence Variants (ASVs) detected in water were also found in sediment eDNA. Remarkably, the ASVs shared between water and sediment accounted for 80% of total sequence reads suggesting that a large amount of plankton DNA is transported to the seafloor, predominantly from abundant phytoplankton taxa. However, not all plankton taxa were equally archived on the seafloor. The plankton DNA deposited in the sediments was dominated by diatoms and showed an underrepresentation of certain nano- and picoplankton taxa (Picozoa or Prymnesiophyceae). Our study offers the first insights into the patterns of plankton diversity recorded in sediment in relation to seasonality and spatial variability of environmental conditions in the Nordic Seas. Our results suggest that the genetic composition and structure of the plankton community vary considerably throughout the water column and differ from what accumulates in the sediment. Hence, the interpretation of sedimentary eDNA archives should take into account potential taxonomic and abundance biases when reconstructing past changes in marine biodiversity.

The voice of the little giants: Arcellinida testate amoebae in environmental DNA-based bioindication, from taxonomy free to haplotypic level.

Bioindication, evaluating biological responses to environmental disturbances, is crucial for assessing the ecological status of an ecosystem. While historical bioindication relied on macroscopic organisms, the introduction of environmental DNA (eDNA) techniques allows the application of protists without the necessity of morphological identification. In this study, we propose a novel bioindication methodology utilizing Arcellinida, a group of top predators among protists, as bioindicators of freshwater ecosystems. For that purpose, we first characterized the Arcellinida diversity over 1 year at three different points of Lake Sanabria, an ancient glacier lake known to be subjected to anthropogenic disturbances. We compared this diversity with an undisturbed control site. Second, we characterized the Arcellinida diversity in other ecosystems to generate the ecological background to test the connectivity between them. Results indicate limited connectivity between the different ecosystems and an edge effect between terrestrial and aquatic ecosystems. Disturbed freshwater ecosystems exhibited reduced Arcellinida diversity at both specific and infraspecific levels, providing valuable insight into recent disturbances. Arcellinida-based bioindication provides a sensitive, accurate and easy-to-interpret protocol for monitoring disturbances in freshwater ecosystems. It represents a valuable tool for environmental assessments and conservation strategies.

Lateral carbon export of macroalgae and seagrass from diverse habitats contributes particulate organic carbon in the deep sea of the Northern South China Sea.

Our study explored the lateral export of macroalgae and seagrass to the deep sea of the Northern South China Sea (NSCS). Particulate organic carbon (POC) collected from a depth of 500 m off southwestern Taiwan (station T) and Dongsha Atoll (station K) underwent environmental DNA (eDNA) and stable isotope assays. Metabarcoding using 18S V9 rDNA revealed lateral export of macrophyte detritus in NSCS. At station K, seagrass detritus predominated, while at station T, macroalgae-derived detritus was dominant. The consistency in the stable carbon isotope signature between POC and macrophytes indicates that stable carbon is an ideal bio-indicator for tracking macrophyte detritus destination and transformation after it has been laterally exported. Based on robust scientific methods, these findings provide valuable insights into the lateral export of macrophyte detritus to the deep sea in POC, influenced by habitat species, and shaped by distinct oceanographic physics around NSCS.

When does a parasite become a disease? eDNA unravels complex host-pathogen dynamics across environmental stress gradients in wild salmonid populations.

Infectious diseases stem from disrupted interactions among hosts, parasites, and the environment. Both abiotic and biotic factors can influence infection outcomes by shaping the abundance of a parasite's infective stages, as well as the host's ability to fight infection. However, disentangling these mechanisms within natural ecosystems remains challenging. Here, combining environmental DNA analysis and niche modelling at a regional scale, we uncovered the biotic and abiotic drivers of an infectious disease of salmonid fish, triggered by the parasite Tetracapsuloides bryosalmonae. We found that the occurrence and abundance of the parasite in the water-i.e., the propagule pressure- were mainly correlated to the abundances of its two primary hosts, the bryozoan Fredericella sultana and the fish Salmo trutta, but poorly to local abiotic environmental stressors. In contrast, the occurrence and abundance of parasites within fish hosts-i.e., proxies for disease emergence-were closely linked to environmental stressors (water temperature, agricultural activities, dams), and to a lesser extent to parasite propagule pressure. These results suggest that pathogen distribution alone cannot predict the risk of disease in wildlife, and that local anthropogenic stressors may play a pivotal role in disease emergence among wild host populations, likely by modulating the hosts' immune response. Our study sheds light on the intricate interplay between biotic and abiotic factors in shaping pathogen distribution and raises concerns about the effects of global change on pathogen emergence.

Streamlining large-scale oceanic biomonitoring using passive eDNA samplers integrated into vessel's continuous pump underway seawater systems.

Passive samplers are enabling the scaling of environmental DNA (eDNA) biomonitoring in our oceans, by circumventing the time-consuming process of water filtration. Designing a novel passive sampler that does not require extensive sample handling time and can be connected to ocean-going vessels without impeding normal underway activities has potential to rapidly upscale global biomonitoring efforts onboard the world's oceanic fleet. Here, we demonstrate the utility of an artificial sponge sampler connected to the continuous pump underway seawater system as a means to enable oceanic biomonitoring. We compared the performance of this passive sampling protocol with standard water filtration at six locations during a research voyage from New Zealand to Antarctica in early 2023. Eukaryote metabarcoding of the mitochondrial COI gene revealed no significant difference in phylogenetic α-diversity between sampling methods and both methods delineated a progressive reduction in number of Zero-Radius Operational Taxonomic Units (ZOTUs) with increased latitudes. While both sampling methods revealed comparable trends in geographical community compositions, distinct clusters were identified for passive samplers and water filtration at each location. Additionally, greater variability between replicates was observed for passive samplers, resulting in an increased estimated level of replication needed to recover 90 % of the biodiversity. Furthermore, traditional water filtration failed to detect three phyla observed by passive samplers and extrapolation analysis estimated passive samplers recover a larger number of ZOTUs compared to water filtration for all six locations. Our results demonstrate the potential of this passive eDNA sampler protocol and highlight areas where this emerging technology could be improved, thereby enabling large-scale offshore marine eDNA biomonitoring by leveraging the world's oceanic fleet without interfering with onboard activities.

An updated LSU database and pipeline for environmental DNA identification of arbuscular mycorrhizal fungi.

Recent work established a backbone reference tree and phylogenetic placement pipeline for identification of arbuscular mycorrhizal fungal (AMF) large subunit (LSU) rDNA environmental sequences. Our previously published pipeline allowed any environmental sequence to be identified as putative AMF or within one of the major families. Despite this contribution, difficulties in implementation of the pipeline remain. Here, we present an updated database and pipeline with (1) an expanded backbone tree to include four newly described genera and (2) several changes to improve ease and consistency of implementation. In particular, packages required for the pipeline are now installed as a single folder (conda environment) and the pipeline has been tested across three university computing clusters. This updated backbone tree and pipeline will enable broadened adoption by the community, advancing our understanding of these ubiquitous and ecologically important fungi.

Comparison of soil eDNA to camera traps for assessing mammal and bird community composition and site use.

Species detections often vary depending on the survey methods employed. Some species may go undetected when using only one approach in community-level inventory and monitoring programs, which has management and conservation implications. We conducted a comparative study of terrestrial mammal and bird detections in the spring and summer of 2021 by placing camera traps at 30 locations across a large military installation in northern Michigan, USA and testing replicate soil samples from these sites for environmental DNA (eDNA) using an established vertebrate metabarcoding assay. We detected a total of 48 taxa from both survey methods: 26 mammalian taxa (excluding humans, 24 to species and two to genus) and 22 avian taxa (21 to species and one to genus). We detected a relatively even distribution of mammalian taxa on cameras (17) and via eDNA analysis (15), with seven taxa detected from both methods. Most medium-to-large carnivores were detected only on cameras, whereas semi-fossorial small mammals were detected only via eDNA analysis. We detected higher bird diversity with camera traps (18 taxa) compared to eDNA analysis (eight taxa; four taxa were detected with both methods), but cameras alone were most effective at detecting smaller birds that frequently occupy arboreal environments. We also used Bayesian spatial occupancy models for two widely distributed game species (white-tailed deer, Odocoileus virginianus, and ruffed grouse, Bonasa umbellus) that were moderately detected with both survey methods and found species-specific site use (occupancy) estimates were similar between cameras and eDNA analysis. Concordant with similar studies, our findings suggest that a combination of camera trap and eDNA surveys could be most useful for assessing the composition of terrestrial mammal communities. Camera traps may be most efficient for assessing bird diversity but can be complemented with eDNA analysis, particularly for species that spend considerable time on the ground.