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Freshwater habitats are under stress from agricultural land use, most notably the influx of neonicotinoid pesticides and increased nutrient pressure from fertilizer. Traditional studies investigating the effects of stressors on freshwater systems are often limited to a narrow range of taxa, depending heavily on morphological expertise. Additionally, disentanglement of multiple simultaneous stressors can be difficult in field studies, whereas controlled laboratory conditions do not accurately reflect natural conditions and food webs. To overcome these drawbacks, we investigated the impacts of two agricultural stressors (the neonicotinoid insecticide thiacloprid and fertilizer) in full‐factorial design in a semi‐natural research site, using environmental DNA sampling to study three different taxonomic groups representing three trophic levels: bacteria (decomposers), phytoplankton (primary producers), and chironomids (consumers). The results show considerable impact of both stressors across trophic levels, with an additive effect of fertilizer and thiacloprid on community composition at all levels. These findings suggest that agricultural stressors affect the entire food web, either directly or through cascade reactions. They are also consistent with morphological assessments that were performed in the same study site, even at a lower number of replicates. The study presented shows that the use of multimarker environmental DNA provides a more comprehensive assessment of stressor impacts across multiple trophic levels, at a higher taxonomic resolution than traditional surveys. Additionally, many putative novel bioindicators for both agricultural stressors were discovered. We encourage further investigations into stressors impacts at different trophic levels, which will lead to more effective monitoring and management of freshwater systems. [ABSTRACT FROM AUTHOR]

The hydrozoan species Nemalecium lighti (Hargitt, 1924) is widely distributed in tropical marine waters around the world. Here we report the complete linear mitochondrial genome of N. lighti from Sint Eustatius (Lesser Antilles). The mitochondrial genome with a length of 14,320 bp encodes for 13 protein-coding genes, two tRNA genes, and two rRNA genes. Gene arrangement differs from that found in other species of the same taxonomic order and a phylogenetic analysis shows that based on mitochondrial genes, N. lighti clusters outside of the Leptothecata, rendering the order paraphyletic. [ABSTRACT FROM AUTHOR]

DNA-based identification through the use of metabarcoding has been proposed as the next step in the monitoring of biological communities, such as those assessed under the Water Framework Directive (WFD). Advances have been made in the field of metabarcoding, but challenges remain when using complex samples. Uneven biomass distributions, preferential amplification and reference database deficiencies can all lead to discrepancies between morphological and DNA-based taxa lists. The effects of different taxonomic groups on these issues remain understudied. By metabarcoding WFD monitoring samples, we analyzed six different taxonomic groups of freshwater organisms, both separately and combined. Identifications based on metabarcoding data were compared directly to morphological assessments performed under the WFD. The diversity of taxa for both morphological and DNA-based assessments was similar, although large differences were observed in some samples. The overlap between the two taxon lists was 56.8% on average across all taxa, and was highest for Crustacea, Heteroptera, and Coleoptera, and lowest for Annelida and Mollusca. Taxonomic sorting in six basic groups before DNA extraction and amplification improved taxon recovery by 46.5%. The impact on ecological quality ratio (EQR) scoring was considerable when replacing morphology with DNA-based identifications, but there was a high correlation when only replacing a single taxonomic group with molecular data. Different taxonomic groups provide their own challenges and benefits. Some groups might benefit from a more consistent and robust method of identification. Others present difficulties in molecular processing, due to uneven biomass distributions, large genetic diversity or shortcomings of the reference database. Sorting samples into basic taxonomic groups that require little taxonomic knowledge greatly improves the recovery of taxa with metabarcoding. Current standards for EQR monitoring may not be easily replaced completely with molecular strategies, but the effectiveness of molecular methods opens up the way for a paradigm shift in biomonitoring. [ABSTRACT FROM AUTHOR]

Background: The heterogeneous nature of environmental DNA (eDNA) and its effects on species detection and community composition estimates has been highlighted in several studies in the past decades. Mostly in the context of spatial distribution over large areas, in fewer occasions looking at spatial distribution within a single body of water. Temporal variation of eDNA, similarly, has mostly been studied as seasonality, observing changes over large periods of time, and often only for small groups of organisms such as fish and amphibians. Methods: We analyzed and compared small-scale spatial and temporal variation by sampling eDNA from two small, isolated dune lakes for 20 consecutive weeks. Metabarcoding was performed on the samples using generic COI primers. Molecular operational taxonomic unit (MOTUs) were used to assess dissimilarities between spatial and temporal replicates. Results: Our results show large differences between samples taken within one lake at one point in time, but also expose the large differences between temporal replicates, even those taken only 1 week apart. Furthermore, between-site dissimilarities showed a linear correlation with time frame, indicating that between-site differences will be inflated when samples are taken over a period of time. We also assessed the effects of PCR replicates and processing strategies on general patterns of dissimilarity between samples. While more inclusive PCR replicate strategies lead to higher richness estimations, dissimilarity patterns between samples did not significantly change. Conclusions: We conclude that the dissimilarity of temporal replicates at a 1 week interval is comparable to that of spatial replicate samples. It increases, however, for larger time intervals, which suggests that population turnover effects can be stronger than community heterogeneity. Spatial replicates alone may not be enough for optimal recovery of taxonomic diversity, and cross-comparisons of different locations are susceptible to inflated dissimilarities when performed over larger time intervals. Many of the observed MOTUs could be classified as either phyto- or zooplankton, two groups that have gained traction in recent years as potential novel bio-indicator species. Our results, however, indicate that these groups might be susceptible to large community shifts in relatively short periods of time, highlighting the need to take temporal variations into consideration when assessing their usability as water quality indicators. [ABSTRACT FROM AUTHOR]

Six species of dragonflies are recorded for the island of Sint Eustatius, Lesser Antilles. Breeding records are established for four out of the six species, despite the lack of natural fresh water sources. DNA barcoding was used to match the larvae with the adults. [ABSTRACT FROM AUTHOR]

The use of molecular tools for the detection and identification of invertebrate species enables the development of more easily standardisable inventories of biological elements for water quality assessments, as it circumvents human-based bias and errors in species identifications. Current Ecological Quality Ratio (EQR) assessments methods, however, often rely on abundance data. Translating metabarcoding sequence data into biomass or specimen abundances has proven difficult, as PCR amplification bias due to primer mismatching often provides skewed proportions of read abundances. While some potential solutions have been proposed in previous research, we instead looked at the necessity of abundance data in EQR assessments. In this study, we used historical monitoring data from natural (lakes, rivers and streams) and artificial (ditches and canals) water bodies to assess the impact of species abundances on the EQR scores for macroinvertebrates in the Water Framework Directive (WFD) monitoring programme of The Netherlands. By removing all the abundance data from the taxon observations, we simulated presence/absencebased monitoring, for which EQRs were calculated according to traditional methods. Our results showed a strong correlation between abundance-based and presence/absence-based EQRs. EQR scores were generally higher without abundances (75.8% of all samples), which resulted in 9.1% of samples being assigned to a higher quality class. The majority of the samples (89.7%) were assigned to the same quality class in both cases. These results are valuable for the incorporation of presence/absence metabarcoding data into water quality assessment methodology, potentially eliminating the need to translate metabarcoding data into biomass or absolute specimen counts for EQR assessments. [ABSTRACT FROM AUTHOR]

Assessing mosquito biodiversity is important for disease surveillance and ecosystem health assessments. Such studies are particularly needed in regions like the Caribbean, which have experienced a series of recent mosquito borne disease outbreaks but received little attention regarding its invertebrate biodiversity. Here, we report on results from a mosquito survey on the Dutch Leeward Islands (Sint Eustatius, Sint Maarten and Saba), carried out in April and October 2018, which is the first integrative survey since those conducted by Van der Kuyp (1947) and Wagenaar Hummelinck (1949). Moreover, we present a novel key for adults and fourth instar larvae of the mosquitoes of the Dutch Leeward Islands. Overall, eleven species were recorded, eight on Sint Maarten, five on Saba and two on Sint Eustatius. Two new potential disease vectors, Culex nigripalpus and Aedes taeniorhynchus, were recorded on Sint Maarten. One previously recorded species, Cx. habilitator, was not retrieved from any of the islands, which is further discussed in the paper. Species indicative of natural forest which previously occurred on all three islands were absent from Sint Eustatius and Sint Maarten but still present on Saba. In contrast, species indicative of human inhabitation, Ae. aegypti and Cx. quinquefasciatus, were highly abundant on Sint Maarten and Sint Eustatius and present in low numbers on Saba. Overall, the results of this study emphasize the importance of biodiversity surveys and indicate that frequent mosquito inventories may contribute to a better understanding of mosquito community composition and distribution of potential vector species. [ABSTRACT FROM AUTHOR]

The bioassessment of aquatic ecosystems is currently based on various biotic indices that use the occurrence and/or abundance of selected taxonomic groups to define ecological status. These conventional indices have some limitations, often related to difficulties in morphological identification of bioindicator taxa. Recent development of DNA barcoding and metabarcoding could potentially alleviate some of these limitations, by using DNA sequences instead of morphology to identify organisms and to characterize a given ecosystem. In this paper, we review the structure of conventional biotic indices, and we present the results of pilot metabarcoding studies using environmental DNA to infer biotic indices. We discuss the main advantages and pitfalls of metabarcoding approaches to assess parameters such as richness, abundance, taxonomic composition and species ecological values, to be used for calculation of biotic indices. We present some future developments to fully exploit the potential of metabarcoding data and improve the accuracy and precision of their analysis. We also propose some recommendations for the future integration of DNA metabarcoding to routine biomonitoring programs. [ABSTRACT FROM AUTHOR]

We calculated a Living Planet Index (LPI) for the Netherlands, based on 361 animal species from seven taxonomic groups occurring in terrestrial and freshwater habitats. Our assessment is basically similar to the global LPI, but the latter includes vertebrate species and trends in population abundance only. To achieve inferences on trends in biodiversity more generally, we added two insect groups (butterflies and dragonflies) and added occupancy trends for species for which we had no abundance trends available. According to the LPI, the state of biodiversity has slightly increased from 1990 to 2014. However, large differences exist between habitat types. We found a considerable increase in freshwater animal populations, probably because of improvement of chemical water quality and rehabilitation of marshland habitats. We found no trend in the LPI for woodland populations. In contrast, populations in farmland and open semi-natural habitats (coastal dunes, heathland and semi-natural grassland) declined, which we attribute to intensive agricultural practices and nitrogen deposition, respectively. The LPI shows that, even in a densely populated western European country, ongoing loss of animal biodiversity is not inevitable and may even be reversed if adequate measures are taken. Our approach enabled us to produce summary statistics beyond the level of species groups to monitor the state of biodiversity in a clear and consistent way. [ABSTRACT FROM AUTHOR]

Effective identification of species using short DNA fragments (DNA barcoding and DNA metabarcoding) requires reliable sequence reference libraries of known taxa. Both taxonomically comprehensive coverage and content quality are important for sufficient accuracy. For aquatic ecosystems in Europe, reliable barcode reference libraries are particularly important if molecular identification tools are to be implemented in biomonitoring and reports in the context of the EU Water Framework Directive (WFD) and the Marine Strategy Framework Directive (MSFD). We analysed gaps in the two most important reference databases, Barcode of Life Data Systems (BOLD) and NCBI GenBank, with a focus on the taxa most frequently used in WFD and MSFD. Our analyses show that coverage varies strongly among taxonomic groups, and among geographic regions. In general, groups that were actively targeted in barcode projects (e.g. fish, true bugs, caddisflies and vascular plants) are well represented in the barcode libraries, while others have fewer records (e.g. marine molluscs, ascidians, and freshwater diatoms). We also found that species monitored in several countries often are represented by barcodes in reference libraries, while species monitored in a single country frequently lack sequence records. A large proportion of species (up to 50%) in several taxonomic groups are only represented by private data in BOLD. Our results have implications for the future strategy to fill existing gaps in barcode libraries, especially if DNA metabarcoding is to be used in the monitoring of European aquatic biota under the WFD and MSFD. For example, missing species relevant to monitoring in multiple countries should be prioritized for future collaborative programs. We also discuss why a strategy for quality control and quality assurance of barcode reference libraries is needed and recommend future steps to ensure full utilisation of metabarcoding in aquatic biomonitoring., (Copyright © 2019 The Authors. Published by Elsevier B.V. All rights reserved.)