Project description:A large data set from the Eastern Mediterranean was analyzed to explore the relationship between seawater column variables and benthic community status. Our results showed a strong quantitative link between the seawater column variables (Chlorophyll a and Eutrophication Index) and various indicators describing benthic diversity and community composition. The percentage of benthic opportunistic species increased significantly in the stations with high trophic status of the seawater column and so did the strength of the coupling between values of seawater column and benthic indicators. The Eutrophication Index threshold level of 0.85, separating the "Bad and Poor" from "Moderate to High" conditions could serve as an acceptable critical value above which there is a readily observable change in benthic community composition.
Project description:Environmental DNA (eDNA) is the DNA suspended in the environment (e.g., water column), which includes cells, gametes, and other material derived from but not limited to shedding of tissue, scales, mucus, and fecal matter. Amplifying and sequencing marker genes (i.e., metabarcoding) from eDNA can reveal the wide range of taxa present in an ecosystem through analysis of a single water sample. Metabarcoding of eDNA provides higher resolution data than visual surveys, aiding in assessments of ecosystem health. This study conducted eDNA metabarcoding of two molecular markers (cytochrome c oxidase I (COI) and 18S ribosomal RNA (rRNA) genes) to survey eukaryotic diversity across multiple trophic levels in surface water samples collected at three sites along the coral reef tract within the Florida Keys National Marine Sanctuary (FKNMS) during four research cruises in 2015. The 18S rRNA gene sequences recovered 785 genera while the COI gene sequences recovered 115 genera, with only 33 genera shared between the two datasets, emphasizing the complementarity of these marker genes. Community composition for both genetic markers clustered by month of sample collection, suggesting that temporal variation has a larger effect on biodiversity than spatial variability in the FKNMS surface waters. Sequences from both marker genes were dominated by copepods, but each marker recovered distinct phytoplankton groups, with 18S rRNA gene sequences dominated by dinoflagellates and COI sequences dominated by coccolithophores. Although eDNA samples were collected from surface waters, many benthic species such as sponges, crustaceans, and corals were identified. These results show the utility of eDNA metabarcoding for cataloging biodiversity to establish an ecosystem baseline against which future samples can be compared in order to monitor community changes.
Project description:BackgroundCharacterizing biodiversity in a habitat or in targeted taxonomically or socioeconomically important groups remains a challenge. Standard DNA-based biodiversity identification tools such as DNA barcoding coupled with high-throughput Next-Generation Sequencing (NGS) technologies are rapidly changing the landscape of biodiversity analysis by targeting various habitats and a wide array of organisms. However, effective use of these technological advances requires optimized protocols and benchmarking against traditional tools. Here we investigate the use of commonly used preservative ethanol as a non-destructive and inexpensive source of DNA for NGS biodiversity analysis of benthic macroinvertebrates. We used the preservative ethanol added to field collected organisms (live sorted bulk benthic samples) as a source of community DNA for NGS environmental barcoding. We directly compare this approach with a DNA barcode library generated using Sanger sequencing of all individuals separated from abenthic sample as well as with NGS environmental barcoding of DNA extracted from mixed/homogenized tissue specimens of the same benthic sample. We also evaluate a multiplex PCR strategy, as compared to commonly used single amplicon workflow, using three newly designed primer sets targeting a wide array of benthic macroinvertebrate taxa.ResultsOur results indicate the effectiveness of ethanol-based DNA in providing sequence information from 87% of taxa identified individually from mixture as compared to 89% in conventional tissue extracted DNA. Missing taxa in both DNA sources were from species with the lowest abundance (e.g. 1 individual) in the benthic mixture. Interestingly, we achieved 100% detection for taxa represented with more than 1% individuals in the mixture in both sources of DNA. Our multiplex amplification regime increased the detection as compared to any single primer set indicating the usefulness of using multiple primer sets in initial amplification of target genes.ConclusionsAlthough NGS approaches have significantly increased the potential of using DNA information in biodiversity analysis, robust methods are needed to provide reliable data and alleviate sample-processing bottlenecks. Here we coupled non-destructive DNA access and a multiplex PCR approach in NGS environmental barcoding for effective data generation from benthic live-sorted samples collected in bulk and preserved in ethanol. Our study provides a possible solution to sampling and vouchering challenges in using benthic samples through next-generation environmental barcoding and facilitates wider utility of DNA information, especially species-specific DNA barcodes, in ecological and environmental studies and real-world applications such as biomonitoring programs.
Project description:Preserving biodiversity is a global challenge requiring data on species' distribution and abundance over large geographic and temporal scales. However, traditional methods to survey mobile species' distribution and abundance in marine environments are often inefficient, environmentally destructive, or resource-intensive. Metabarcoding of environmental DNA (eDNA) offers a new means to assess biodiversity and on much larger scales, but adoption of this approach for surveying whole animal communities in large, dynamic aquatic systems has been slowed by significant unknowns surrounding error rates of detection and relevant spatial resolution of eDNA surveys. Here, we report the results of a 2.5 km eDNA transect surveying the vertebrate fauna present along a gradation of diverse marine habitats associated with a kelp forest ecosystem. Using PCR primers that target the mitochondrial 12S rRNA gene of marine fishes and mammals, we generated eDNA sequence data and compared it to simultaneous visual dive surveys. We find spatial concordance between individual species' eDNA and visual survey trends, and that eDNA is able to distinguish vertebrate community assemblages from habitats separated by as little as ~60 m. eDNA reliably detected vertebrates with low false-negative error rates (1/12 taxa) when compared to the surveys, and revealed cryptic species known to occupy the habitats but overlooked by visual methods. This study also presents an explicit accounting of false negatives and positives in metabarcoding data, which illustrate the influence of gene marker selection, replication, contamination, biases impacting eDNA count data and ecology of target species on eDNA detection rates in an open ecosystem.
Project description:People value the existence of a variety of marine species and habitats, many of which are negatively impacted by human activities. The Convention on Biological Diversity and other international and national policy agreements have set broad goals for reducing the rate of biodiversity loss. However, efforts to conserve biodiversity cannot be effective without comprehensive metrics both to assess progress towards meeting conservation goals and to account for measures that reduce pressures so that positive actions are encouraged. We developed an index based on a global assessment of the condition of marine biodiversity using publically available data to estimate the condition of species and habitats within 151 coastal countries. Our assessment also included data on social and ecological pressures on biodiversity as well as variables that indicate whether good governance is in place to reduce them. Thus, our index is a social as well as ecological measure of the current and likely future status of biodiversity. As part of our analyses, we set explicit reference points or targets that provide benchmarks for success and allow for comparative assessment of current conditions. Overall country-level scores ranged from 43 to 95 on a scale of 1 to 100, but countries that scored high for species did not necessarily score high for habitats. Although most current status scores were relatively high, likely future status scores for biodiversity were much lower in most countries due to negative trends for both species and habitats. We also found a strong positive relationship between the Human Development Index and resilience measures that could promote greater sustainability by reducing pressures. This relationship suggests that many developing countries lack effective governance, further jeopardizing their ability to maintain species and habitats in the future.
Project description:The assessment of the actual impact of discharged wastewater on the whole ecosystem and, in turn, on human health requires the execution of bioassays. In effect, based on the chemical characterization alone, the synergistic/antagonistic effect of mixtures of pollutants is hardly estimable. The aim of this work was to evaluate the applicability of a battery of bioassays and to suggest a smart procedure for results representation. Two real wastewater treatment plants were submitted to analytical campaigns. Several baseline toxicity assays were conducted, together with tests for the determination of endocrine activity, genetic toxicity and carcinogenicity of wastewater. A "traffic light" model was adopted for an easy-to-understand visualization of the results. Although the legal prescriptions of chemical parameters are fully complied with, bioassays show that a certain biological activity still residues in the treated effluents. Moreover, influent and effluent responses are not always appreciably different. Some tests employing human cells were revealed to be only partially adequate for environmental applications. An interesting and helpful development of the present approach would consist in the estimation of biological equivalents of toxicity, as shown for the estrogenic compound 17-β-estradiol.
Project description:Decisions guiding environmental management need to be based on a broad and comprehensive understanding of the biodiversity and functional capability within ecosystems. Microbes are of particular importance since they drive biogeochemical cycles, being both producers and decomposers. Their quick and direct responses to changes in environmental conditions modulate the ecosystem accordingly, thus providing a sensitive readout. Here we have used direct sequencing of total DNA from water samples to compare the microbial communities of two distinct coastal regions exposed to different anthropogenic pressures: the highly polluted Port of Genoa and the protected area of Montecristo Island in the Mediterranean Sea. Analysis of the metagenomes revealed significant differences in both microbial diversity and abundance between the two areas, reflecting their distinct ecological habitats and anthropogenic stress conditions. Our results indicate that the combination of next generation sequencing (NGS) technologies and bioinformatics tools presents a new approach to monitor the diversity and the ecological status of aquatic ecosystems. Integration of metagenomics into environmental monitoring campaigns should enable the impact of the anthropogenic pressure on microbial biodiversity in various ecosystems to be better assessed and also predicted.
Project description:In a recent paper published in Cell Research, a cryo-EM structure reveals the interface between DNA-PKcs and the Ku70/80:DNA complex, together forming the DNA-dependent protein kinase holoenzyme in non-homologous DNA end joining. Insight from this structure suggests how an allosteric rearrangement of DNA-PKcs driven by Ku70/80:DNA binding regulates kinase activity in this largest member of a family of structurally homologous phosphoinositide 3-kinase-related protein kinases that includes mTOR, ATR, and ATM.