Mock samples resolve biases in diversity estimates and quantitative interpretation of zooplankton metabarcoding data

Metabarcoding is a rapidly developing tool in marine zooplankton ecology, although most zooplankton surveys continue to rely on visual identification for monitoring purposes. We attempted to resolve some of the biases associated with metabarcoding by sequencing a 313-b.p. fragment of the COI gene in 34 “mock” samples from the North Sea which were pre-sorted to species level, with biomass and abundance estimates obtained for each species and taxonomic group. The samples were preserved either in 97% ethanol or dehydrated for 24 h in a drying oven at 65 °C (the routine way of preserving samples for dry weight measurements). The visual identification yielded a total of 59 unique holoplanktonic and 16 meroplanktonic species/taxa. Metabarcoding identified 86 holoplanktonic and 124 meroplanktonic species/taxa, which included all but 3 of the species identified visually as well as numerous species of hard-to-identify crustaceans, hydrozoan jellyfish, and larvae of benthic animals. On a sample-to-sample basis, typically 90–95% of visually registered species were recovered, but the number of false positives was also high. We demonstrate robust correlations of relative sequence abundances to relative biomass for most taxonomic groups and develop conversion factors for different taxa to account for sequencing biases. We then combine the adjusted sequencing data with a single bulk biomass measurement for the entire sample to produce a quantitative parameter akin to species biomass. When examined with multivariate statistics, this parameter, which we call BWSR (biomass-weighed sequence reads) showed very similar trends to species biomass and comparable patterns to species abundance, highlighting the potential of metabarcoding not only for biodiversity estimation and mapping of presence/absence of species but also for quantitative assessment of zooplankton communities.publishedVersio

How Does Nursing Staff Perceive the Use of Electronic Handover Reports? A Questionnaire-Based Study

Following the implementation of electronic nursing records in a psychogeriatric ward, we examined nursing staff's attitudes and perceptions to the implementation of an electronic handover routine. A web-based anonymous and secure questionnaire was distributed by e-mail to all nursing staff at a psychogeriatric ward at a university hospital. Most respondents were satisfied with the electronic handover, and they believed they managed to keep informed by the new routine. The simultaneous introduction of a morning meeting, to ensure a forum for oral professional discussion, was a success. A minority of staff did not fully trust the information conveyed in the electronic handover, and a significant proportion expressed a need for guidance in using the system. Staff that had a high level of trust in written reports believed these saved time, had little trouble finding time and a place to read the reports, and were more positive to the new handover routine

The role of morphological evolution and prey specialization in adaptive radiations: The spider genus Dysdera in the Canary Islands

Non peer reviewe

To denoise or to cluster? That is not the question. Optimizing pipelines for COI metabarcoding and metaphylogeography

Background: The recent blooming of metabarcoding applications to biodiversity studies comes with some relevant methodological debates. One such issue concerns the treatment of reads by denoising or by clustering methods, which have been wrongly presented as alternatives. It has also been suggested that denoised sequence variants should replace clusters as the basic unit of metabarcoding analyses, missing the fact that sequence clusters are a proxy for species-level entities, the basic unit in biodiversity studies. We argue here that methods developed and tested for ribosomal markers have been uncritically applied to highly variable markers such as cytochrome oxidase I (COI) without conceptual or operational (e.g., parameter setting) adjustment. COI has a naturally high intraspecies variability that should be assessed and reported, as it is a source of highly valuable information. We contend that denoising and clustering are not alternatives. Rather, they are complementary and both should be used together in COI metabarcoding pipelines. Results: Using a COI dataset from benthic marine communities, we compared two denoising procedures (based on the UNOISE3 and the DADA2 algorithms), set suitable parameters for denoising and clustering, and applied these steps in diferent orders. Our results indicated that the UNOISE3 algorithm preserved a higher intra-cluster variability. We introduce the program DnoisE to implement the UNOISE3 algorithm taking into account the natural variability (measured as entropy) of each codon position in protein-coding genes. This correction increased the number of sequences retained by 88%. The order of the steps (denoising and clustering) had little infuence on the fnal outcome. Conclusions: We highlight the need for combining denoising and clustering, with adequate choice of stringency parameters, in COI metabarcoding. We present a program that uses the coding properties of this marker to improve the denoising step. We recommend researchers to report their results in terms of both denoised sequences (a proxy for haplotypes) and clusters formed (a proxy for species), and to avoid collapsing the sequences of the latter into a single representative. This will allow studies at the cluster (ideally equating species-level diversity) and at the intra-cluster level, and will ease additivity and comparability between studies

Metabarcoding reveals high-resolution biogeographical and metaphylogeographical patterns through marine barriers

Aim: It has been predicted that there should be concordance between biogeographical and phylogeographical processes structuring multi-species regional assemblages. We hypothesise that oceanographic barriers in the marine environment affect concomitantly the distribution and the connectivity of the marine biota, thus producing congruent biogeographical and phylogeographical structures. We also predict that macro- and meio-eukaryotes will be differentially affected by hydrological features. Location: The Atlanto-Mediterranean transition along the E Iberian coast marked by the Almeria-Oran Front (AOF) and the Ibiza Channel hydrological discontinuities. Taxon: Eukaryotes. Methods: A new analytical framework based in the metabarcoding of community DNA and a hypervariable marker is presented. This framework allows the simultaneous detection of multispecies biogeographical and phylogeographical structures. Shallow hard-bottom communities were sampled at 12 sites over the littoral zone and community-DNA metabarcoding was performed using the cytochrome c oxidase I marker. The resulting dataset was analysed at several levels: beta diversity of Molecular Operational Taxonomic Units (MOTUs) as surrogate for species, and Exact Sequence Variants as surrogate for haplotypes. We also assessed genetic differentiation within MOTUs (metaphylogeography). Analyses were performed for the combined dataset and separately for macro- and meio-eukaryotes. Results: Both hydrological discontinuities had a detectable effect, more marked at all levels for the AOF than for the Ibiza Channel. The MOTU dataset provided more clear-cut patterns than the ESVs. The metaphylogeographical approach provided the highest resolution in terms of differentiating localities and identifying geographical barriers. The separate analyses of macro- and meio-eukaryotes showed a higher differentiation of the latter, both in terms of beta diversity and genetic differentiation. Main Conclusions: Metabarcoding coupled with metaphylogeography allowed the characterisation of the heterogeneity in community composition and population genetic structure along the Atlanto-Mediterranean transition, coherent with known hydrological discontinuities. This methodology unlocks a vast amount of information on the geographical distribution of different components of biodiversity for basic and applied research

Using DNA Metabarcoding to Characterize the Prey Spectrum of Two Co-Occurring Themisto Amphipods in the Rapidly Changing Atlantic-Arctic Gateway Fram Strait

The two congeneric hyperiids Themisto libellula and T. abyssorum provide an important trophic link between lower and higher trophic levels in the rapidly changing Arctic marine ecosystem. These amphipods are characterized by distinct hydrographic affinities and are hence anticipated to be impacted differently by environmental changes, with major consequences for the Arctic food web. In this study, we applied DNA metabarcoding to the stomach contents of these Themisto species, to comprehensively reveal their prey spectra at an unprecedented-high-taxonomic-resolution and assess the regional variation in their diet across the Fram Strait. Both species feed on a wide variety of prey but their diet strongly differed in the investigated summer season, showing overlap for only a few prey taxa, such as calanoid copepods. The spatially structured prey field of T. libellula clearly differentiated it from T. abyssorum, of which the diet was mainly dominated by chaetognaths. Our approach also allowed the detection of previously overlooked prey in the diet of T. libellula, such as fish species and gelatinous zooplankton. We discuss the reasons for the differences in prey spectra and which consequences these may have in the light of ongoing environmental changes

DNA metabarcoding reveals the importance of gelatinous zooplankton in the diet of Pandalus borealis, a keystone species in the Arctic

Information about the dietary composition of a species is crucial to understanding their position and role in the food web. Increasingly, molecular approaches such as DNA metabarcoding are used in studying trophic relationships, not least because they may alleviate problems such as low taxonomic resolution or underestimation of digestible taxa in the diet. Here, we used DNA metabarcoding with universal primers for cytochrome c oxidase I (COI) to study the diet composition of the northern shrimp (Pandalus borealis), an Arctic keystone species with large socio-economic importance. Across locations, jellyfish and chaetognaths were the most important components in the diet of P. borealis, jointly accounting for 40%–60% of the total read abundance. This dietary importance of gelatinous zooplankton contrasts sharply with published results based on stomach content analysis. At the same time, diet composition differed between fjord and shelf locations, pointing to different food webs supporting P. borealis in these two systems. Our study underlines the potential of molecular approaches to provide new insights into the diet of marine invertebrates that are difficult to obtain with traditional methods, and calls for a revision of the role of gelatinous zooplankton in the diet of the key Arctic species P. borealis, and in extension, Arctic food webs

Spatio-temporal patterns of eukaryotic biodiversity in shallow hard-bottom communities from the West Antarctic Peninsula revealed by DNA metabarcoding

Aim: We studied molecular eukaryotic biodiversity patterns in shallow hard-bottom Antarctic benthic communities using community DNA metabarcoding. Polar ecosystems are extremely exposed to climate change, and benthic macroinvertebrate communities have demonstrated rapid response to a range of natural and anthropogenic pressures. However, these rich and diverse ecosystems are poorly studied, revealing how little is known about the biodiversity of the Antarctic benthos associated with hard-bottom habitats. Location: West Antarctic Peninsula and South Shetland Islands. Methods: Using data collected in seven localities along the western Antarctic Peninsula, we calculated spatial patterns of alpha and beta diversities. Furthermore, we analysed temporal changes in benthic composition in one location (Deception Island) over 3 years. We calculated the temporal alpha and beta diversities to reveal changes in this community over time. Results: We obtained a final list of 2057 molecular operational taxonomic units. We found significant differences in benthic community composition between localities and among years. Our dataset revealed a total of 10 different kingdom-level lineages and 34 different phyla in the samples. The most diverse phylum was Arthropoda, followed by Bacillariophyta, and Annelida, while the highest relative read abundances belonged to Annelida, Porifera and Echinodermata. Benthic community compositions changed between 2016 and 2018 in Deception Island, and decreasing species richness was the main component of temporal beta diversity. Main Conclusions: Direct sampling methods are required for monitoring these complex communities. Informative biodiversity patterns can be retrieved even though most of the benthic biodiversity found in Antarctic habitats is yet to be taxonomically described and barcoded. Hard-bottom assemblages exhibit high spatial variability and heterogeneity, not related to depth, which represent a huge challenge for largescale studies in the Southern Ocean. Local patchiness and structure within the communities are probably a consequence of a combination of several biotic and abiotic factors (i.e. ice disturbance, food supply and competition)

Persistence of environmental DNA in marine systems

As environmental DNA (eDNA) becomes an increasingly valuable resource for marine ecosystem monitoring, understanding variation in its persistence across contrasting environments is critical. Here, we quantify the breakdown of macrobial eDNA over a spatio-temporal axis of locally extreme conditions, varying from ocean-influenced offshore to urban-inshore, and between winter and summer. We report that eDNA degrades 1.6 times faster in the inshore environment than the offshore environment, but contrary to expectation we find no difference over season. Analysis of environmental covariables show a spatial gradient of salinity and a temporal gradient of pH, with salinity—or the biotic correlates thereof—most important. Based on our estimated inshore eDNA half-life and naturally occurring eDNA concentrations, we estimate that eDNA may be detected for around 48 h, offering potential to collect ecological community data of high local fidelity. We conclude by placing these results in the context of previously published eDNA decay rates