Monitoring bloom-forming Aphanizomenon using environmental DNA metabarcoding: Method development, validation, and field application.

Harmful algal blooms (HABs) have emerged as a critical global environmental and ecological concern. Timely and accurate monitoring of the prevalent bloom-forming genera is crucial for HAB management. Conventional microscope-based methods are time-consuming, labor-intensive, and specialized expertise-dependent, often making them impractical for large-scale surveillance. Molecular methods, such as metabarcoding, provide efficient technical solutions; however, the lack of competent PCR primers and further field validation present obstacles to their wide use. Here, we successfully developed Aphanizomenon-specific primers and validated the application of environmental DNA (eDNA) metabarcoding for field-based monitoring of Aphanizomenon in 37 sites across lentic and lotic freshwater ecosystems in Beijing. The sensitivity and specificity tests of newly developed primers demonstrated high performance - comprehensive recovery of biodiversity in Aphanizomenon communities and high ratios (>95%) of Aphanizomenon sequences in datasets. We observed significant correlations between the sequence abundance derived from eDNA metabarcoding and the total cell density determined through microscopic identification across all the sampling sites, both in the spring (r = 0.8086, p < 0.0001) and summer (r = 0.7902, p < 0.0001), thus validating the utility of eDNA metabarcoding based on the newly developed primers for monitoring in the field. Further, we identified key environmental variables that were primary drivers responsible for the spatiotemporal distribution of Aphanizomenon abundance. These variables included temperature, total nitrogen, and dissolved oxygen in lentic ecosystems, and total phosphorus in lotic ecosystems. The method developed and validated here offers an accurate, efficient, and high-throughput tool for the monitoring of Aphanizomenon blooms in freshwater ecosystems.

GoEnrich: creating high quality genomic DNA resources from limited voucher specimen tissues or museum specimens of at-risk species for conservation-friendly use in the validation of environmental DNA assays.

OBJECTIVE: Environmental DNA (eDNA) methods are crucial for monitoring populations, particularly rare and cryptic species. For confident eDNA application, rigorous assay validation is required including specificity testing with genomic DNA (gDNA). However, this critical step is often difficult to achieve as obtaining fresh tissue samples from at-risk species can be difficult, highly limited, or impossible. Natural history museum collections could serve as a valuable and ethical voucher specimen resource for eDNA assay validation. The present study demonstrates the effectiveness of whole genome amplification (WGA) in providing enough gDNA to assemble high quality mitogenomes from which robust targeted eDNA assays can be designed. RESULTS: Using fresh and historical museum tissue samples from six species spanning fish, birds, and mammals, we successfully developed a WGA method with an average yield of 380 to 1,268 ng gDNA per 20 µL reaction. This gDNA was used for whole genome shotgun sequencing and subsequent assembly of high quality mitogenomes using mtGrasp. These mitogenomes were then used to develop six new robust, targeted quantitative real time polymerase chain reaction-based eDNA assays and 200 ng WGA-enriched yielded satisfactory Cq values and near 100% detection frequencies for all assays tested. This approach offers a cost-effective and non-invasive alternative, streamlining eDNA research processes and aiding in conservation efforts.

Unmanaged grasslands are a reservoir of Alternaria and other important fungal species with differing emission patterns.

Alternaria is a ubiquitous fungal genus with many allergenic and pathogenic species inhabiting grasslands. We hypothesise that grasslands (natural/man-made) host a diversity of fungal species whose spores have varying emission patterns. Therefore, the purpose of this study was to examine the potential of grasslands for emission, diversity and composition of Alternaria and other fungal species. To test the hypothesis, Hirst-type and multi-vial Cyclone samplers collected air samples from two grassland sites (unmanaged and managed) and a non-grassland site at Lakeside campus of the University of Worcester, United Kingdom for the period May to September 2019. The unmanaged grassland was originally planted with grasses and left uncut for three years. The managed grassland was a roadside verge that was cut once every year, typically after most grasses have flowered. We used optical microscopy and Illumina MiSeq sequencing to investigate the emission, abundance, diversity and composition of the fungal spores from each site alongside meteorological variables. Kruskal-Wallis and Wilcoxon tests examined differences in the bi-hourly Alternaria concentrations between the sites. Shannon's and Simpson's Index determined the diversity of the fungal spores between the unmanaged and non-grassland sites. The results showed that grasslands are a strong source of Alternaria spores with considerably higher numbers of clinically important days compared with the non-grassland site. The managed grassland varied in Alternaria spore emission pattern from the unmanaged, probably due to differences in environmental variables and cutting frequency. The unmanaged grassland and non-grassland sites showed a high diversity of fungi including Alternaria, Cladosporium, Ascochyta, Botrytis and Aureobasidium. Overall, the study shows that grasslands are a strong source of fungal spores with allergenic and pathogenic potential and have varying emission patterns, compared with nearby urban areas where monitoring stations are located. This information is useful for atmospheric modelling of airborne fungal spore sources and has implications for allergy sufferers in particular.

Evaluating the quantitative performance of environmental DNA metabarcoding for freshwater zooplankton community: a case study in Lake Biwa, Japan.

Zooplankton monitoring is important for understanding their population dynamics and life history, ecosystem health, and environmental changes. Compared with traditional morphological identification, environmental DNA (eDNA) analysis allows for more sensitive and efficient monitoring of zooplankton diversity. Previous eDNA studies have primarily used metabarcoding approaches to reveal their richness and composition, whereas its performance in predicting zooplankton abundance remains understudied. We conducted water and bulk sampling in Lake Biwa, Japan, showing that the number of sequence reads by metabarcoding moderately correlated with eDNA concentrations estimated by quantitative real-time PCR (qPCR). In addition, the eDNA read number was significantly related to cladoceran and copepod abundance estimated by microscopy sorting, although there remained too much uncertainty in the read-abundance relationship. Moreover, there was a significant difference in species composition between eDNA metabarcoding and sorting. Although our results indicated the potential applicability of eDNA metabarcoding for quantifying multiple zooplankton abundance, several methodological validations in eDNA metabarcoding would also be required to optimize its performance in the future.

Comparative analysis of size-fractional eukaryotic microbes in subtropical riverine systems inferred from 18S rRNA gene V4 and V9 regions.

Eukaryotic microbes play key ecological roles in riverine ecosystems. Amplicon sequencing has greatly facilitated the identification and characterization of eukaryotic microbial communities. Currently, 18S rRNA gene V4 and V9 hypervariable regions are widely used for sequencing eukaryotic microbes. Identifying optimal regions for the profiling of size-fractional eukaryotic microbial communities is critical for microbial ecological studies. In this study, we spanned three rivers with typical natural-human influenced transition gradients to evaluate the performance of the 18S rRNA gene V4 and V9 hypervariable regions for sequencing size-fractional eukaryotic microbes (>180 µm, 20-180 µm, 5-20 µm, 3-5 µm, 0.8-3 µm). Our comparative analysis revealed that amplicon results depend on the specific species and microbial size. The V9 region was most effective for detecting a broad taxonomic range of species. The V4 region was superior to the V9 region for the identification of microbes in the minor 3 µm and at the family and genus levels, especially for specific microbial groups, such as Labyrinthulomycetes. However, the V9 region was more effective for studies of diverse eukaryotic groups, including Archamoebae, Heterolobosea, and Microsporidia, and various algae, such as Haptophyta, Florideophycidae, and Bangiales. Our results highlight the importance of accounting for potential misclassifications when employing both V4 and V9 regions for the identification of microbial sequences. The use of optimal regions for amplification could enhance the utility of amplicon sequencing in environmental studies. The insights gained from this work will aid future studies that employ amplicon-based identification approaches for the characterization of eukaryotic microbial communities and contribute to our understanding of microbial ecology within aquatic systems.

Metagenomic approaches and opportunities in arid soil research.

Arid soils present unique challenges and opportunities for studying microbial diversity and bioactive potential due to the extreme environmental conditions they bear. This review article investigates soil metagenomics as an emerging tool to explore complex microbial dynamics and unexplored bioactive potential in harsh environments. Utilizing advanced metagenomic techniques, diverse microbial populations that grow under extreme conditions such as high temperatures, salinity, high pH levels, and exposure to metals and radiation can be studied. The use of extremophiles to discover novel natural products and biocatalysts emphasizes the role of functional metagenomics in identifying enzymes and secondary metabolites for industrial and pharmaceutical purposes. Metagenomic sequencing uncovers a complex network of microbial diversity, offering significant potential for discovering new bioactive compounds. Functional metagenomics, connecting taxonomic diversity to genetic capabilities, provides a pathway to identify microbes' mechanisms to synthesize valuable secondary metabolites and other bioactive substances. Contrary to the common perception of desert soil as barren land, the metagenomic analysis reveals a rich diversity of life forms adept at extreme survival. It provides valuable findings into their resilience and potential applications in biotechnology. Moreover, the challenges associated with metagenomics in arid soils, such as low microbial biomass, high DNA degradation rates, and DNA extraction inhibitors and strategies to overcome these issues, outline the latest advancements in extraction methods, high-throughput sequencing, and bioinformatics. The importance of metagenomics for investigating diverse environments opens the way for future research to develop sustainable solutions in agriculture, industry, and medicine. Extensive studies are necessary to utilize the full potential of these powerful microbial communities. This research will significantly improve our understanding of microbial ecology and biotechnology in arid environments.

Barcoding of Italian mosquitoes (BITMO): generation and validation of DNA barcoding reference libraries for native and alien species of Culicidae.

BACKGROUND: Mosquitoes (Culicidae), as disease vectors, represent a risk for human health worldwide. Repeated introductions of alien mosquito species and the spread of invasive species have been recorded in different countries. Traditionally, identification of mosquitoes relies on morphological observation. However, morphology-based identification is associated with a number of potential disadvantages, such as the high level of specialisation of the operator and its limited applicability to damaged samples. In these cases, species identification is achieved through molecular methods based on DNA amplification. Molecular-based taxonomy has also enabled the development of techniques for the study of environmental DNA (eDNA). Previous studies indicated the 16S mitochondrial ribosomal RNA (rRNA) gene as a promising target for this application; however, 16S rRNA sequences are available for only a limited number of mosquito species. In addition, although primers for the 16S rRNA gene were designed years ago, they are based on limited numbers of mosquito sequences. Thus, the aims of this study were to: (i) design pan-mosquito 16S rRNA gene primers; (ii) using these primers, generate a 16S rRNA gene mosquito reference library (with a focus on mosquitoes present in Italy); and (iii) compare the discriminatory power of the 16S rRNA gene with two widely used molecular markers, cytochrome c oxidase subunit 1 mitochondrial gene (COI) and internal transcribed spacer 2 (ITS2). METHODS: A total of six mosquito genera (28 mosquito species) were included in this study: Aedes (n = 16 species), Anopheles (5 species), Coquillettidia (1 species), Culex (3 species), Culiseta (2 species) and Uranotaenia (1 species). DNA was extracted from the whole mosquito body, and more than one specimen for each species was included in the analysis. Sanger sequencing was used to generate DNA sequences that were then analysed through the Barcode of Life Data Systems (BOLD). Phylogenetic analyses were also performed. RESULTS: Novel 16S rDNA gene, COI and ITS2 sequences were generated. The 16S rRNA gene was shown to possess sufficient informativeness for the identification of mosquito species, with a discriminatory power equivalent to that of COI. CONCLUSIONS: This study contributes to the generation of DNA barcode libraries, focussed on Italian mosquitoes, with a significant increase in the number of 16S rRNA gene sequences. We hope that these novel sequences will provide a resource for studies on the biodiversity, monitoring and metabarcoding of mosquitoes, including eDNA-based approaches.

Pollinator behaviour and prevalence of the anther smut Antherospora vindobonensis in its host, the Hungarian two-leaf squill (Scilla vindobonensis).

Anther smuts are fungal diseases that have detrimental effects on the biology of their host plants. We investigated the transmission of Anther smut Antherospora vindobonensis to its host, Scilla vindobonensis under natural and laboratory conditions. Plants flowering early in spring were less vulnerable to disease (∼ 20-30% of plants which appeared early in the season were diseased) than flowers that bloom at the end of the season (∼ 60% of plants which appeared late in the season were diseased) which contradicts the patterns observed for other anther smuts species. Diseased plants showed thicker stems, lower nectar production and smaller flower radius than healthy plants. The disease prevalence in S. vindobonensis was not influenced by plant density. The production of spores in diseased flowers was 125 times higher than the production of pollen in healthy flowers. The number of pollen grains attached to pollinator bodies from healthy plants under controlled conditions was similar to the number of spores attached from diseased plants. Experienced honeybees in the field showed almost identical preferences for healthy plants (65%) over diseased ones, as did naïve bumblebees under controlled laboratory conditions (69%). eDNA metabarcoding revealed that healthy flowers attracted a significantly higher number of arthropod species (M = 6.23, SE = 1.48) than diseased flowers (M = 2.32, SE = 0.31). Strong seasonal differences in the risk of infection suggest that S. vindobonensis solves the trade-off between low pollinator availability and low risk of infection at the beginning of the season and high risk of infection and high pollinator availability at the end of season. By reproducing early in the season, plants can reduce the risk of disease.

Methodological assessment for efficient collection of Schistosoma mansoni environmental DNA and improved schistosomiasis surveillance in tropical wetlands.

Schistosomiasis, caused by trematodes of genus Schistosoma, is among the most seriously neglected tropical diseases. Although rapid surveillance of risk areas for Schistosoma transmission is vital to control schistosomiasis, the habitat and infection status of this parasite are difficult to assess. Environmental DNA (eDNA) analysis, involving the detection of extra-organismal DNA in water samples, facilitates cost-efficient and sensitive biomonitoring of aquatic environments and is a promising tool to identify Schistosoma habitat and infection risk areas. However, in tropical wetlands, highly turbid water causes filter clogging, thereby decreasing the filtration volume and increasing the risk of false negatives. Therefore, in this study, we aimed to conduct laboratory experiments and field surveys in Lake Victoria, Mbita, to determine the appropriate filter pore size for S. mansoni eDNA collection in terms of particle size and filtration volume. In the laboratory experiment, aquarium water was sequentially filtered using different pore size filters. Targeting >3 µm size fraction was found to be sufficient to capture S. mansoni eDNA particles, regardless of their life cycle stage (egg, miracidia, and cercaria). In the field surveys, GF/D (2.7 µm nominal pore size) filter yielded 2.5-times the filtration volume obtained with a smaller pore size filter and pre-filtration methods under the same time constraints. Moreover, a site-occupancy model was applied to the field detection results to estimate S. mansoni eDNA occurrence and detection probabilities and assess the number of water samples and PCR replicates necessary for efficient eDNA detection. Overall, this study reveals an effective method for S. mansoni eDNA detection in turbid water, facilitating the rapid and sensitive monitoring of its distribution and cost-effective identification of schistosomiasis transmission risk areas.

Storms facilitate airborne DNA from leaf fragments outside the main tree pollen season.

Bioaerosols are useful indicators of plant phenology and can demonstrate the impacts of climate change on both local and regional scales (e.g. pollen monitoring/flowering phenology). Analysing bioaerosols with eDNA approaches are becoming more popular to quantify the diversity of airborne plant environmental DNA (eDNA) and flowering season of plants and trees. Leaf abscission from broadleaved trees and other perennial species can also indicate the status of plant health in response to climate. This happens primarily during autumn in response to seasonal growth conditions and environmental factors, such as changing photoperiod and reduced temperatures. During this period biological material is released in larger quantities to the environment. Here, rural bioaerosol composition during late summer and autumn was captured by MiSEQ sequencing of the rRNA internal transcribed spacer 2 (ITS2) region, a common marker for taxonomic variation. Meteorological parameters were recorded from a proximal weather station. The composition of atmospheric taxa demonstrated that deciduous tree DNA forms part of the bioaerosol community during autumn and, for several common broadleaved tree species, atmospheric DNA abundance correlated to high wind events. This suggests that both flowering and autumn storms cause bioaerosols from deciduous trees that can be detected with eDNA approaches. This is an aspect that must be considered when eDNA methods are used to analyse either pollen or other fragments from trees. Supplementary Information: The online version contains supplementary material available at 10.1007/s10453-024-09826-w.

Fungi of Great Salt Lake, Utah, USA: a spatial survey.

The natural system at Great Salt Lake, Utah, USA was augmented by the construction of a rock-filled railroad causeway in 1960, creating two lakes at one site. The north arm is sequestered from the mountain snowmelt inputs and thus became saturated with salts (250-340 g/L). The south arm is a flourishing ecosystem with moderate salinity (90-190 g/L) and a significant body of water for ten million birds on the avian flyways of the western US who engorge themselves on the large biomass of brine flies and shrimp. The sediments around the lake shores include calcium carbonate oolitic sand and clay, and further away from the saltwater margins, a zone with less saline soil. Here a small number of plants can thrive, including Salicornia and Sueda species. At the north arm at Rozel Point, halite crystals precipitate in the salt-saturated lake water, calcium sulfate precipitates to form gypsum crystals embedded in the clay, and high molecular weight asphalt seeps from the ground. It is an ecosystem with gradients and extremes, and fungi are up to the challenge. We have collected data on Great Salt Lake fungi from a variety of studies and present them here in a spatial survey. Combining knowledge of cultivation studies as well as environmental DNA work, we discuss the genera prevalent in and around this unique ecosystem. A wide diversity of taxa were found in multiple microniches of the lake, suggesting significant roles for these genera: Acremonium, Alternaria, Aspergillus, Cladosporium, Clydae, Coniochaeta, Cryptococcus, Malassezia, Nectria, Penicillium, Powellomyces, Rhizophlyctis, and Wallemia. Considering the species present and the features of Great Salt Lake as a terminal basin, we discuss of the possible roles of the fungi. These include not only nutrient cycling, toxin mediation, and predation for the ecosystem, but also roles that would enable other life to thrive in the water and on the shore. Many genera that we discovered may help other organisms in alleviating salinity stress, promoting growth, or affording protection from dehydration. The diverse taxa of Great Salt Lake fungi provide important benefits for the ecosystem.

Dammed coastal waterways are less diverse, more homogenous, and dominated by non-native species: Comprehensive insights from quantitative analysis of environmental DNA.

Small dams are commonplace worldwide and impact local and regional aquatic diversity by altering habitats and disrupting dispersal networks. Quantifying the local and regional impacts of dams requires nearly comprehensive species occurrence data. We used environmental DNA (eDNA) metabarcoding to test theoretical predictions about the impacts of dams on local and regional bony fish diversity within the Chesapeake Bay Watershed, USA. We analyzed eDNA from 465 sampling points within 34 waterbodies documenting the distributions of 61 species. On average, dammed waterbodies had approximately half (48 %) as many species per site as undammed (lower alpha diversity) and more homogenous species composition (lower beta diversity). Native migratory species were less than one tenth (0.08) as likely to be detected at dammed sites than undammed sites, native resident (non-migratory) species were one third (0.34) as likely, whereas introduced species were 2.6 times more likely to be detected. Our sampling and bioinformatics methods were validated by a diverse mock community control. Our results suggest that dams in coastal waterways homogenize fish metacommunities, reduce local biodiversity through dispersal limitation and habitat alteration, and favor the dominance of lentic-adapted introduced species while potentially restricting the spread of introduced catfish. Decisions to construct or decommission dams should consider local and regional impacts on biodiversity.

eProbe: Sampling of Environmental DNA within Tree Canopies with Drones.

Environmental DNA (eDNA) analysis is a powerful tool for studying biodiversity in forests and tree canopies. However, collecting representative eDNA samples from these high and complex environments remains challenging. Traditional methods, such as surface swabbing or tree rolling, are labor-intensive and require significant effort to achieve adequate coverage. This study proposes a novel approach for unmanned aerial vehicles (UAVs) to collect eDNA within tree canopies by using a surface swabbing technique. The method involves lowering a probe from a hovering UAV into the canopy and collecting eDNA as it descends and ascends through branches and leaves. To achieve this, a custom-designed robotic system was developed featuring a winch and a probe for eDNA collection. The design of the probe was optimized, and a control logic for the winch was developed to reduce the risk of entanglement while ensuring sufficient interaction force to facilitate transfer of eDNA onto the probe. The effectiveness of this method was demonstrated during the XPRIZE Rainforest Semi-Finals as 10 eDNA samples were collected from the rainforest canopy, and a total of 152 molecular operational taxonomic units (MOTUs) were identified using eDNA metabarcoding. We further investigate how the number of probe interactions with vegetation, the penetration depth, and the sampling duration influence the DNA concentration and community composition of the samples.

Using grass inflorescence as source material for biomonitoring through environmental DNA metabarcoding.

BACKGROUND: Over the last decade, increasing attention has been directed to using different substrates as sources of environmental DNA (eDNA) in ecological research. Reports on the use of environmental DNA located on the surface of plant leaves and flowers have highlighted the utility of this DNA source in studies including, but not limited to, biodiversity, invasive species, and pollination ecology. The current study assesses grass inflorescence as a source of eDNA for detecting invertebrate taxa. METHODS AND RESULTS: Inflorescences from four common grass species in a central South African grassland were collected for high-throughput sequencing analysis. Universal COI primers were utilised to detect Metazoan diversity. The sequencing results allowed for the detection of three Arthropoda orders, with most OTUs assigned to fungal taxa (Ascomycota and Basidiomycota). Some biases were detected while observing the relative read abundance (RRA) results. DISCUSSION: The observed biases could be explained by the accidental inclusion of invertebrate specimens during sample collection and DNA extraction. Primer biases towards the amplified taxa could be another reason for the observed RRA results. This study provided insight into the invertebrate community associated with the four sampled grass species. It should be noted that with the lack of negative field controls, it is impossible to rule out the influence of airborne eDNA on the observed diversity associated with each grass species. The lack of the inclusion of PCR and extraction blanks in the sequencing step, as well as the inclusion of negative field controls, including other areas for refinement were highlighted, and suggestions were provided to improve the outcomes of future studies.

A comparative analysis of eDNA metabarcoding and field surveys: Exploring freshwater plant communities in rivers.

While environmental DNA (eDNA) metabarcoding holds promise as a holistic approach to assess vegetation changes and community composition across diverse spatial and temporal scales, systematic investigations of its efficacy compared to conventional field surveys remain scarce in the literature. The present study explores the differences in plant diversity recovered from field surveys and captured with a multi-marker eDNA metabarcoding approach (two nrDNA ITS1 and ITS2, and two cpDNA rbcL and trnL) from river water samples. The eDNA metabarcoding approach retrieved 46 aquatic plants (hydrophytes and helophytes) and 245 terrestrial plants, compared to 24 and 127 species identified from field surveys. On average, eDNA samples collected immediately downstream of the survey sites recovered 43 % and 39 % of the aquatic and terrestrial species observed, respectively. Discrepancies were explained by differences in taxonomic resolution, the stochasticity of the retrieval of rare and elusive species, and the presence of reference sequences. We found a significant positive correlation between spatial and community distances at scales ranging from 2 to 9 km and identified turnover as the driving force of these differences. Metabarcoding demonstrated sensitivity to community changes and both approaches converge on a similar community structure. Interestingly, eDNA samples collected immediately upstream of the survey sites exhibited significant species overlap with the downstream samples (c. 100 m apart). Overall, our results demonstrate that within-site species mismatches between the methods are nonnegligible, and they question the use of eDNA for generating complete species lists at scales comparable to our field surveys (< 100-m transects). However, with adequate sampling and a multi-marker metabarcoding approach, eDNA has the potential to approximate catchment gamma diversity with less sampling effort than conventional surveys.

Environmental DNA-Based Identification of Non-Native Fish in Beijing: Diversity, Geographical Distribution, and Interactions with Native Taxa.

Rapid urbanization and its associated human activities have facilitated the colonization and spread of non-native species, rendering urban ecosystems, particularly in megacities such as Beijing, highly susceptible to biological invasions. This study employed environmental DNA (eDNA) metabarcoding to evaluate the biodiversity and geographical distribution of non-native fish, as well as their interactions with native fish species, across three river basins in Beijing pertaining to the Daqing River, the North Canal, and the Ji Canal. Across all the 67 sampling sites, we identified 60 fish taxa, representing 11 orders, 23 families, and 40 genera, with an average of 33.0 taxa per site. Of these, 40 taxa were native, accounting for only 47.1% of the historically recorded native fish species. Additionally, we detected 20 non-native fish taxa, spanning 11 orders, 13 families, and 17 genera. Native fish exhibited geographical homogenization across the basins, while non-native taxa displayed varied geographical distributions. Non-metric multidimensional scaling (NMDS) and analysis of similarities (ANOSIM) revealed no significant variation in the non-native communities across the river basins. Although most of the non-native taxa were widespread, some were restricted to specific sites or basins. The North Canal exhibited significantly lower non-native biodiversity compared with the Ji Canal across all alpha diversity indices. Simple linear regression analyses indicated positive correlations between the number of taxa and species richness for both native and non-native taxa. Interestingly, species co-occurrence analyses revealed predominantly positive interactions among both native and non-native species pairs, with only two negative relationships involving one native and two non-native taxa. This study provides insights into the biodiversity and geographical distribution of non-native fish in Beijing and establishes a baseline for future biomonitoring and conservation efforts. The findings underscore the need for further investigation into the mechanisms and dynamics of biological invasions within urban environments in Beijing.

Effect of uneven tolerance to human disturbance on dominance interactions of top predators.

Anthropogenic activities may alter felid assemblage structure, facilitating the persistence of tolerant species (commonly mesopredators), excluding ecologically demanding ones (top predators) and, consequently, changing coexistence rules. We aimed to determine how human activities influence intraguild relationships among top predators and their cascading effects on mesopredators, which remain poorly understood despite evidence of top carnivore decline. We used structural equation modeling at a continental scale to investigate how habitat quality and quantity, livestock density, and other human pressures modified the intraguild relations of the 3 species that are at the top of the food chain in the Neotropics: jaguars (Panthera onca), pumas (Puma concolor), and ocelots (Leopardus pardalis). We included presence-absence data derived from systematic studies compiled in Neocarnivores data set for these felid species at 0.0833° resolution. Human disturbance reduced the probability of jaguar occurrence by -0.35 standard deviations. Unexpectedly, the presence of sheep (Ovis aries) or goats (Capra aegagrus hircus) and jaguars was positively related to the presence of pumas, whereas puma presence was negatively related to the presence of ocelots. Extent of forest cover had more of an effect on jaguar (ß = 0.23) and ocelot (ß = 0.12) occurrences than the extent of protected area, which did not have a significant effect. The lack of effect of human activities on puma presence and the positive effect of small livestock supports the notion that pumas are more adaptable to habitat disturbance than jaguars. Our findings suggest that human disturbance has the potential to reverse the hierarchical competition dominance among large felids, leading to an unbalanced community structure. This shift disadvantages jaguars and elevates the position of pumas in the assemblage hierarchy, resulting in the exclusion of ocelots, despite their relatively lower susceptibility to anthropogenic disturbance. Our results suggest that conservation efforts should extend beyond protected areas to encompass the surrounding landscape, where complexities and potential conflicts are more pronounced.

Efectos de la tolerancia dispareja a las perturbaciones humanas sobre las interacciones de dominancia de los depredadores mayores Resumen Las actividades antropogénicas pueden alterar la estructura de las poblaciones de félidos, lo que facilita la persistencia de especies tolerantes (normalmente mesodepredadores), excluye a otras especies con exigencias ecológicas (depredadores mayores) y, en consecuencia, modifica las reglas de coexistencia. Buscamos determinar cómo influyen las actividades humanas sobre las relaciones entre gremios de los depredadores superiores y sus efectos en cascada sobre los mesodepredadores, que aún son poco conocidos a pesar de las pruebas del declive de los carnívoros superiores. Utilizamos modelos de ecuaciones estructurales a escala continental para investigar cómo la calidad y cantidad del hábitat, la densidad ganadera y otras presiones humanas modifican las relaciones entre gremios de las tres especies que se encuentran en la cima de la cadena trófica en el Neotrópico: jaguares (Panthera onca), pumas (Puma concolor) y ocelotes (Leopardus pardalis). Incluimos datos de presencia­ausencia derivados de estudios sistemáticos recopilados en el conjunto de datos Neocarnivores para estas especies de félidos con una resolución de 0.0833°. Las perturbaciones humanas redujeron la probabilidad de aparición del jaguar en ­0.35 desviaciones estándar. De forma inesperada, la presencia de ovejas (Ovis aries) o cabras (Capra aegargrus hircus) y jaguares tuvo una relación positiva con la presencia de pumas, mientras que la presencia de pumas tuvo una relación negativa con la presencia de ocelotes. La extensión de la cubierta forestal tuvo más efecto sobre la presencia de jaguares (ß = 0.23) y ocelotes (ß = 0.12) que la extensión del área protegida, que no tuvo un efecto significativo. La falta de efectos de las actividades humanas sobre la presencia del puma y el efecto positivo del ganado menor apoyan la idea de que los pumas son más adaptables a las perturbaciones del hábitat que los jaguares. Nuestros hallazgos sugieren que las perturbaciones humanas tienen el potencial de invertir la dominancia jerárquica de competencia entre los grandes félidos, lo que lleva a una estructura comunitaria desequilibrada. Este cambio perjudica a los jaguares y eleva la posición de los pumas en la jerarquía del ensamblaje, lo que excluye a los ocelotes, a pesar de su relativamente menor susceptibilidad a las perturbaciones antropogénicas. Nuestros resultados sugieren que los esfuerzos de conservación deberían extenderse más allá de las áreas protegidas para abarcar el paisaje circundante, donde las complejidades y los conflictos potenciales son más pronunciados.

Three-dimensional conservation planning of fish biodiversity metrics to achieve the deep-sea 30×30 conservation target.

Accelerating rate of human impact and environmental change severely affects marine biodiversity and increases the urgency to implement the Convention on Biological Diversity (CBD) 30×30 plan for conserving 30% of sea areas by 2030. However, area-based conservation targets are complex to identify in a 3-dimensional (3D) ocean where deep-sea features such as seamounts have been seldom studied mostly due to challenging methodologies to implement at great depths. Yet, the use of emerging technologies, such as environmental DNA combined with modern modeling frameworks, could help address the problem. We collected environmental DNA, echosounder acoustic, and video data at 15 seamounts and deep island slopes across the Coral Sea. We modeled 7 fish community metrics and the abundances of 45 individual species and molecular operational taxonomic units (MOTUs) in benthic and pelagic waters (down to 600-m deep) with boosted regression trees and generalized joint attribute models to describe biodiversity on seamounts and deep slopes and identify 3D protection solutions for achieving the CBD area target in New Caledonia (1.4 million km2). We prioritized the identified conservation units in a 3D space, based on various biodiversity targets, to meet the goal of protecting at least 30% of the spatial domain, with a focus on areas with high biodiversity. The relationship between biodiversity protection targets and the spatial area protected by the solution was linear. The scenario protecting 30% of each biodiversity metric preserved almost 30% of the considered spatial domain and accounted for the 3D distribution of biodiversity. Our study paves the way for the use of combined data collection methodologies to improve biodiversity estimates in 3D structured marine environments for the selection of conservation areas and for the use of biodiversity targets to achieve area-based international targets.

Planeación tridimensional de la conservación de las medidas de biodiversidad de peces para lograr el objetivo de conservación 30x30 de mar profundo Resumen El impacto antropogénico y el cambio ambiental acelerados afectan gravemente a la biodiversidad marina y aumentan la urgencia de aplicar el plan 30x30 del Convenio sobre la Diversidad Biológica (CDB) para conservar el 30% de las zonas marinas para el 2030. Sin embargo, la identificación de objetivos de conservación basados en zonas es compleja en un océano tridimensional (3D) en el que rara vez se han estudiado las características de las profundidades marinas, como los montes marinos, sobre todo por la dificultad de aplicar metodologías a grandes profundidades. No obstante, el uso de tecnologías emergentes, como el ADN ambiental combinado con marcos actuales de modelación, podría ayudar a resolver el problema. Recopilamos datos de ADN ambiental, acústica de ecosonda y video en 15 montes marinos y taludes de islas profundas del mar del Coral. Modelamos siete medidas de comunidades de peces y 45 abundancias de especies individuales y unidades taxonómicas moleculares (UTOM) en aguas bentónicas y pelágicas (hasta 600 m de profundidad) con árboles de regresión reforzada (ARR) y modelos de atributos conjuntos generalizados (MACJ) para describir la biodiversidad en los montes marinos y taludes profundos e identificar soluciones de protección en 3D para alcanzar el objetivo de área del CDB en Nueva Caledonia (1.4 millones de km2). Priorizamos las unidades de conservación identificadas en un espacio 3D con base en varios objetivos de biodiversidad para cumplir el objetivo de proteger al menos el 30% del dominio espacial con un enfoque en las zonas con una gran biodiversidad. La relación entre los objetivos de protección de la biodiversidad y el área espacial protegida por la solución fue lineal. El escenario que protegía el 30% de cada medida de biodiversidad preservó casi el 30% del dominio espacial considerado y consideró la distribución tridimensional de la biodiversidad. Nuestro estudio prepara el camino para el uso de metodologías combinadas de recopilación de datos con el fin de mejorar las estimaciones de biodiversidad en entornos marinos estructurados en 3D para la selección de áreas de conservación y para el uso de objetivos de biodiversidad con el fin de alcanzar objetivos internacionales basados en áreas.

Size-dependent release and temperature-dependent degradation of salmonid environmental DNA under laboratory conditions.

The degradation of environmental DNA (eDNA) and its release from fish were investigated in laboratory experiments for seven salmonid fishes. The eDNA concentration in experimental tanks without fish decreased exponentially, with a higher rate of decline observed under higher water temperature conditions. When a fish was introduced into a tank, the eDNA concentration was positively correlated with the length and weight of the fish.

Decomposing a San Francisco estuary microbiome using long-read metagenomics reveals species- and strain-level dominance from picoeukaryotes to viruses.

Although long-read sequencing has enabled obtaining high-quality and complete genomes from metagenomes, many challenges still remain to completely decompose a metagenome into its constituent prokaryotic and viral genomes. This study focuses on decomposing an estuarine metagenome to obtain a more accurate estimate of microbial diversity. To achieve this, we developed a new bead-based DNA extraction method, a novel bin refinement method, and obtained 150 Gbp of Nanopore sequencing. We estimate that there are ~500 bacterial and archaeal species in our sample and obtained 68 high-quality bins (>90% complete, <5% contamination, ≤5 contigs, contig length of >100 kbp, and all ribosomal and tRNA genes). We also obtained many contigs of picoeukaryotes, environmental DNA of larger eukaryotes such as mammals, and complete mitochondrial and chloroplast genomes and detected ~40,000 viral populations. Our analysis indicates that there are only a few strains that comprise most of the species abundances. IMPORTANCE: Ocean and estuarine microbiomes play critical roles in global element cycling and ecosystem function. Despite the importance of these microbial communities, many species still have not been cultured in the lab. Environmental sequencing is the primary way the function and population dynamics of these communities can be studied. Long-read sequencing provides an avenue to overcome limitations of short-read technologies to obtain complete microbial genomes but comes with its own technical challenges, such as needed sequencing depth and obtaining high-quality DNA. We present here new sampling and bioinformatics methods to attempt decomposing an estuarine microbiome into its constituent genomes. Our results suggest there are only a few strains that comprise most of the species abundances from viruses to picoeukaryotes, and to fully decompose a metagenome of this diversity requires 1 Tbp of long-read sequencing. We anticipate that as long-read sequencing technologies continue to improve, less sequencing will be needed.