Pest Alert Tool-a web-based application for flagging species of concern in metabarcoding datasets.

Advances in high-throughput sequencing (HTS) technologies and their increasing affordability have fueled environmental DNA (eDNA) metabarcoding data generation from freshwater, marine and terrestrial ecosystems. Research institutions worldwide progressively employ HTS for biodiversity assessments, new species discovery and ecological trend monitoring. Moreover, even non-scientists can now collect an eDNA sample, send it to a specialized laboratory for analysis and receive in-depth biodiversity record from a sampling site. This offers unprecedented opportunities for biodiversity assessments across wide temporal and spatial scales. The large volume of data produced by metabarcoding also enables incidental detection of species of concern, including non-indigenous and pathogenic organisms. We introduce an online app-Pest Alert Tool-for screening nuclear small subunit 18S ribosomal RNA and mitochondrial cytochrome oxidase subunit I datasets for marine non-indigenous species as well as unwanted and notifiable marine organisms in New Zealand. The output can be filtered by minimum length of the query sequence and identity match. For putative matches, a phylogenetic tree can be generated through the National Center for Biotechnology Information's BLAST Tree View tool, allowing for additional verification of the species of concern detection. The Pest Alert Tool is publicly available at https://pest-alert-tool-prod.azurewebsites.net/.

Fatty acid production and associated gene pathways are altered by increased salinity and dimethyl sulfoxide treatments during cryopreservation of Symbiodinium pilosum (Symbiodiniaceae).

The Symbiodinium genus is ancestral among other Symbiodiniaceae lineages with species that are both symbiotic and free living. Changes in marine ecosystems threaten their existence and crucial ecological roles. Cryopreservation offers an avenue for their long-term storage for future habitat restoration after coral bleaching. In our previous study we demonstrated that high salinity treatments of Symbiodiniaceae isolates led to changes in their fatty acid (FA) profiles and higher cell viabilities after cryopreservation. In this study, we investigated the role of increased salinity on FA production and the genes involved in FA biosynthesis and degradation pathways during the cryopreservation of Symbiodinium pilosum. Overall, there was a twofold increase in mass of FAs produced by S. pilosum after being cultured in medium with increased salinity (54 parts per thousand; ppt). Dimethyl sulfoxide (Me2SO) led to a ninefold increase of FAs in standard salinity (SS) treatment, compared to a fivefold increase in increased salinity (IS) treatments. The mass of the FA classes returned to baseline during recovery. Transcriptomic analyses showed an acyl carrier protein gene was significantly upregulated after Me2SO treatment in the SS cultures. Cytochrome P450 reductase genes were significantly down regulated after Me2SO addition in SS treatment preventing FA degradation. These changes in the expression of FA biosynthesis and degradation genes contributed to more FAs in SS treated isolates. Understanding how increased salinity changes FA production and the roles of specific genes in regulating FA pathways will help improve current freezing protocols for Symbiodiniaceae and other marine microalgae.

Hurdles and opportunities in implementing marine biosecurity systems in data-poor regions.

Managing marine nonindigenous species (mNIS) is challenging, because marine environments are highly connected, allowing the dispersal of species across large spatial scales, including geopolitical borders. Cross-border inconsistencies in biosecurity management can promote the spread of mNIS across geopolitical borders, and incursions often go unnoticed or unreported. Collaborative surveillance programs can enhance the early detection of mNIS, when response may still be possible, and can foster capacity building around a common threat. Regional or international databases curated for mNIS can inform local monitoring programs and can foster real-time information exchange on mNIS of concern. When combined, local species reference libraries, publicly available mNIS databases, and predictive modeling can facilitate the development of biosecurity programs in regions lacking baseline data. Biosecurity programs should be practical, feasible, cost-effective, mainly focused on prevention and early detection, and be built on the collaboration and coordination of government, nongovernment organizations, stakeholders, and local citizens for a rapid response.

A taxonomy-free diatom eDNA-based technique for assessing lake trophic level using lake sediments.

Anthropogenic eutrophication is one of the most pressing issues facing lakes globally. Our ability to manage lake eutrophication is hampered by the limited spatial and temporal extents of monitoring records, stemming from the time-consuming and expensive nature of physiochemical and biological monitoring. Diatom-based biomonitoring presents an alternative to traditional eutrophication monitoring, yet it is restricted by the high degree of taxonomic expertise required. Environmental DNA metabarcoding, while providing a promising substitute for diatom community enumeration, is plagued by inadequate taxonomic coverage of reference databases and methodological bias, limiting its use for biomonitoring. Here we show that taxonomy-free diatom-biomonitoring, in which environmental DNA metabarcoding data is utilised but not assigned to specific taxonomic classes, presents an accurate, fast, and relatively automated alternative to taxonomically assigned eutrophication biomonitoring. Our taxonomy-free index accounted for 85% of trophic level variability across 89 lakes and had the lowest average prediction error of the three approaches tested. By not relying on taxonomic identification or metabarcoding reference databases, taxonomy-free biomonitoring maintains diatom diversity that is lost in taxonomic assignment using molecular approaches. Furthermore, by utilising lake sediments, the approach outlined here presents a time-integrated estimation of lake trophic level and thus does not require time-consuming seasonal sampling. Taxonomy-free biomonitoring addresses the limitations of traditional physicochemical eutrophication monitoring and taxonomic biomonitoring alternatives and can be used to extend the spatial and temporal extents of eutrophication monitoring.

Resolving 500 Years of Anthropogenic Impacts in a Mesotrophic Lake: Nutrients Outweigh Other Drivers of Lake Change.

Interactions among multiple stressors, legacies of past perturbations, and the lack of historical information make it difficult to determine the influence of individual anthropogenic impacts on lakes and separate them from natural ecosystem variability. In the present study, we coupled paleolimnological approaches, historical data, and ecological experiments to disentangle the impacts of multiple long-term stressors on lake ecosystem structure and function. We found that the lake structure and function remained resistant to the impacts of catchment deforestation and erosion, and the introduction of several exotic fish species. Changes in ecosystem structure and function were consistent, with nutrient enrichment being the primary driver of change. Significant and sustained changes in the lake diatom community structure (and their nutrient requirements), bacterial community function, and paleolimnological proxies of ecosystem function coincided with nitrogen and phosphorus fertilizers in the catchment. The results highlight that the effects of increased nutrient inputs are much stronger than the influence of other, potentially significant, drivers of ecosystem change, and that the degree of nutrient impact can be underestimated by environmental monitoring due to its diffuse and accumulative nature. Delineating the effects of multiple anthropogenic drivers requires long-term records of both impacts and lake ecosystem change across multiple trophic levels.

Lake microbial communities are not resistant or resilient to repeated large-scale natural pulse disturbances.

Opportunities to study community-level responses to extreme natural pulse disturbances in unaltered ecosystems are rare. Lake sediment records that span thousands of years can contain well-resolved sediment pulses, triggered by earthquakes. These palaeorecords provide a means to study repeated pulse disturbances and processes of resistance (insensitivity to disturbance) and ecological resilience (capacity to regain structure, function and process). In this study, sedimentary DNA was extracted from a sediment core from Lake Paringa (New Zealand) that is situated in a near natural catchment. Metabarcoding and inferred functions were used to assess the lake microbial community over the past 1100 years - a period that included four major earthquakes. Microbial community composition and function differed significantly between highly perturbed (postseismic, ~50 years) phases directly after the earthquakes and more stable (interseismic, ~250 years) phases, indicating a lack of community resistance. Although community structure differed significantly in successive postseismic phases, function did not, suggesting potential functional redundancy. Significant differences in composition and function in successive interseismic phases demonstrate that communities are not resilient to large-scale natural pulse disturbances. The clear difference in structure and function, and high number of indicator taxa (responsible for driving differences in communities between phases) in the fourth interseismic phase probably represents a regime shift, possibly due to the two-fold increase in sediment and terrestrial biospheric organic carbon fluxes recorded following the fourth earthquake. Large pulse disturbances that enhance sediment inputs into lake systems may produce an underappreciated mechanism that destabilises lake ecosystem processes.

Pan-regional marine benthic cryptobiome biodiversity patterns revealed by metabarcoding Autonomous Reef Monitoring Structures.

Autonomous Reef Monitoring Structures (ARMS) have been applied worldwide to characterize the critical yet frequently overlooked biodiversity patterns of marine benthic organisms. In order to disentangle the relevance of environmental factors in benthic patterns, here, through standardized metabarcoding protocols, we analyse sessile and mobile (<2 mm) organisms collected using ARMS deployed across six regions with different environmental conditions (3 sites × 3 replicates per region): Baltic, Western Mediterranean, Adriatic, Black and Red Seas, and the Bay of Biscay. A total of 27,473 Amplicon Sequence Variants (ASVs) were observed ranging from 1,404 in the Black Sea to 9,958 in the Red Sea. No ASVs were shared among all regions. The highest number of shared ASVs was between the Western Mediterranean and the Adriatic Sea (116) and Bay of Biscay (115). Relatively high numbers of ASVs (103), mostly associated with the genus Amphibalanus, were also shared between the lower salinity seas (Baltic and Black Seas). We found that compositional differences in spatial patterns of rocky-shore benthos are determined slightly more by dispersal limitation than environmental filtering. Dispersal limitation was similar between sessile and mobile groups, while the sessile group had a larger environmental niche breadth than the mobile group. Further, our study can provide a foundation for future evaluations of biodiversity patterns in the cryptobiome, which can contribute up to 70% of the local biodiversity.

Beyond the visual: using metabarcoding to characterize the hidden reef cryptobiome.

In an era of coral reef degradation, our knowledge of ecological patterns in reefs is biased towards large conspicuous organisms. The majority of biodiversity, however, inhabits small cryptic spaces within the framework of the reef. To assess this biodiverse community, which we term the 'reef cryptobiome', we deployed 87 autonomous reef monitoring structures (ARMS), on 22 reefs across 16 degrees latitude of the Red Sea. Combining ARMS with metabarcoding of the mitochondrial cytochrome oxidase I gene, we reveal a rich community, including the identification of 14 metazoan phyla within 10 416 operational taxonomic units (OTUs). While mobile and sessile subsets were similarly structured along the basin, the main environmental driver was different (particulate organic matter and sea surface temperature, respectively). Distribution patterns of OTUs showed that only 1.5% were present in all reefs, while over half were present in a single reef. On both local and regional scales, the majority of OTUs were rare. The high heterogeneity in community patterns of the reef cryptobiome has implications for reef conservation. Understanding the biodiversity patterns of this critical component of reef functioning will enable a sound knowledge of how coral reefs will respond to future anthropogenic impacts.

Disentangling the complex microbial community of coral reefs using standardized Autonomous Reef Monitoring Structures (ARMS).

Autonomous Reef Monitoring Structures (ARMS) have been applied worldwide to describe eukaryotic cryptic reef fauna. Conversely, bacterial communities, which are critical components of coral reef ecosystem functioning, remain largely overlooked. Here we deployed 56 ARMS across the 2,000-km spread of the Red Sea to assay biodiversity, composition and inferred underlying functions of coral reef-associated bacterial communities via 16S rRNA gene sequencing. We found that bacterial community structure and diversity aligned with environmental differences. Indeed, sea surface temperature and macroalgae cover were key in explaining bacterial relative abundance. Importantly, taxonomic and functional alpha diversity decreased under more extreme environmental conditions (e.g., higher temperatures) in the southern Red Sea. This may imply a link between bacterial community diversity and functional capabilities, with implications for conservation management. Our study demonstrates the utility of ARMS to investigate the response of coral reef-associated bacterial communities to environmental change.

A comparison of two gene regions for assessing community composition of eukaryotic marine microalgae from coastal ecosystems.

Two gene regions commonly used to characterise the diversity of eukaryotic communities using metabarcoding are the 18S ribosomal DNA V4 and V9 gene regions. We assessed the effectiveness of these two regions for characterising diverisity of coastal eukaryotic microalgae communities (EMCs) from tropical and temperate sites. We binned amplicon sequence variants (ASVs) into the high level taxonomic groups: dinoflagellates, pennate diatoms, radial centric diatoms, polar centric diatoms, chlorophytes, haptophytes and 'other microalgae'. When V4 and V9 generated ASV abundances were compared, the V9 region generated a higher number of raw reads, captured more diversity from all high level taxonomic groups and was more closely aligned with the community composition determined using light microscopy. The V4 region did resolve more ASVs to a deeper taxonomic resolution within the dinoflagellates, but did not effectively resolve other major taxonomic divisions. When characterising these communities via metabarcoding, the use of multiple gene regions is recommended, but the V9 gene region can be used in isolation to provide high-level community biodiversity to reflect relative abundances within groups. This approach reduces the cost of sequencing multiple gene regions whilst still providing important baseline ecosystem function information.

Responses of the coral reef cryptobiome to environmental gradients in the Red Sea.

An essential component of the coral reef animal diversity is the species hidden in crevices within the reef matrix, referred to as the cryptobiome. These organisms play an important role in nutrient cycling and provide an abundant food source for higher trophic levels, yet they have been largely overlooked. Here, we analyzed the distribution patterns of the mobile cryptobiome (>2000 µm) along the latitudinal gradient of the Saudi Arabian coast of the Red Sea. Analysis was conducted based on 54 Autonomous Reef Monitoring Structures. We retrieved a total of 5273 organisms, from which 2583 DNA sequences from the mitochondrially encoded cytochrome c oxidase I were generated through sanger sequencing. We found that the cryptobiome community is variable over short geographical distances within the basin. Regression tree models identified sea surface temperature (SST), percentage cover of hard coral and turf algae as determinant for the number of operational taxonomic units present per Autonomous Reef Monitoring Structures (ARMS). Our results also show that the community structure of the cryptobiome is associated with the energy available (measured as photosynthetic active radiation), sea surface temperature, and nearby reef habitat characteristics (namely hard corals, turf and macroalgae). Given that temperature and reef benthic characteristics affect the cryptobiome, current scenarios of intensive climate change are likely to modify this fundamental biological component of coral reef functioning. However, the trajectory of change is unknow and can be site specific, as for example, diversity is expected to increase above SST of 28.5°C, and with decreasing hard coral and turf cover. This study provides a baseline of the cryptobenthic community prior to major coastal developments in the Red Sea to be used for future biodiversity studies and monitoring projects. It can also contribute to better understand patterns of reef biodiversity in a period where Marine Protected Areas are being discussed in the region.

Sedimentary ancient DNA reveals the impact of anthropogenic land use disturbance and ecological shifts on fish community structure in small lowland lake.

Freshwater fish biodiversity and abundance are decreasing globally. The drivers of decline are primarily anthropogenic; however, the causative links between disturbances and fish community change are complex and challenging to investigate. We used a suite of sedimentary DNA methods (droplet digital PCR and metabarcoding) and traditional paleolimnological approaches, including pollen and trace metal analysis, ITRAX X-ray fluorescence and hyperspectral core scanning to explore changes in fish abundance and drivers over 1390 years in a small lake. This period captured a disturbance trajectory from pre-human settlement through subsistence living to intensive agriculture. Generalized additive mixed models explored the relationships between catchment inputs, internal drivers, and fish community structure. Fish community composition distinctly shifted around 1350 CE, with the decline of a sensitive Galaxias species concomitant with early land use changes. Total fish abundance significantly declined around 1950 CE related to increases in ruminant bacterial DNA (a proxy for ruminant abundance) and cadmium flux (a proxy for phosphate fertilizers), implicating land use intensification as a key driver. Concurrent shifts in phytoplankton and zooplankton suggested that fish communities were likely impacted by food web dynamics. This study highlights the potential of sedDNA to elucidate the long-term disturbance impacts on biological communities in lakes.

Biodiversity patterns of the coral reef cryptobiota around the Arabian Peninsula.

The Arabian Peninsula accounts for approximately 6% of the world's coral reefs. Some thrive in extreme environments of temperature and salinity. Using 51 Autonomous Reef Monitoring Structure (ARMS), a standardized non-destructive monitoring device, we investigated the spatial patterns of coral reef cryptobenthic diversity in four ecoregions around the Arabian Peninsula and analyzed how geographical and/or environmental drivers shape those patterns. The mitochondrial cytochrome c oxidase subunit I (COI) gene was used to identify Amplicon Sequence Variants and assign taxonomy of the cryptobenthic organisms collected from the sessile and mobile fractions of each ARMS. Cryptobenthic communities sampled from the two ecoregions in the Red Sea showed to be more diverse than those inhabiting the Arabian (Persian) Gulf and the Gulf of Oman. Geographic distance revealed a stronger relationship with beta diversity in the Mantel partial correlation than environmental distance. However, the two mobile fractions (106-500 µm and 500-2000 µm) also had a significant correlation between environmental distance and beta diversity. In our study, dispersal limitations explained the beta diversity patterns in the selected reefs, supporting the neutral theory of ecology. Still, increasing differences in environmental variables (environmental filtering) also had an effect on the distribution patterns of assemblages inhabiting reefs within short geographic distances. The influence of geographical distance in the cryptofauna assemblages makes these relevant, yet usually ignored, communities in reef functioning vulnerable to large scale coastal development and should be considered in ecosystem management of such projects.

Sudden peak in tetrodotoxin in French oysters during the summer of 2021: Source investigation using microscopy, metabarcoding and droplet digital PCR.

Tetrodotoxin (TTX) is a potent neurotoxin causing human intoxications from contaminated seafood worldwide and is of emerging concern in Europe. Shellfish have been shown to contain varying TTX concentrations globally, with concentrations typically higher in Pacific oysters Crassostrea gigas in Europe. Despite many decades of research, the source of TTX remains unknown, with bacterial or algal origins having been suggested. The aim of this study was to identify potential source organisms causing TTX contamination in Pacific oysters in French coastal waters, using three different techniques. Oysters were deployed in cages from April to September 2021 in an estuary where TTX was previously detected. Microscopic analyses of water samples were used to investigate potential microalgal blooms present prior or during the peak in TTX. Differences in the bacterial communities from oyster digestive glands (DG) and remaining flesh were explored using metabarcoding, and lastly, droplet digital PCR assays were developed to investigate the presence of Cephalothrix sp., one European TTX-bearing species in the DG of toxic C. gigas. Oysters analysed by liquid chromatography-tandem mass spectrometry contained quantifiable levels of TTX over a three-week period (24 June-15 July 2021), with concentrations decreasing in the DG from 424 µg/kg for the first detection to 101 µg/kg (equivalent to 74 to 17 µg/kg of total flesh), and trace levels being detected until August 13, 2021. These concentrations are the first report of the European TTX guidance levels being exceeded in French shellfish. Microscopy revealed that some microalgae bloomed during the TTX peak, (e.g., Chaetoceros spp., reaching 40,000 cells/L). Prokaryotic metabarcoding showed increases in abundance of Rubritaleaceae (genus Persicirhabdus) and Neolyngbya, before and during the TTX peak. Both phyla have previously been described as possible TTX-producers and should be investigated further. Droplet digital PCR analyses were negative for the targeted TTX-bearing genus Cephalothrix.

A paleoecological investigation of recent cyanobacterial blooms and their drivers in two contrasting lakes.

Cyanobacterial blooms are one of the most significant threats to global water security and freshwater biodiversity. Interactions among multiple stressors, including habitat degradation, species invasions, increased nutrient runoff, and climate change, are key drivers. However, assessing the role of anthropogenic activity on the onset of cyanobacterial blooms and exploring response variation amongst lakes of varying size and depth is usually limited by lack of historical records. In the present study we applied molecular, paleolimnological (trace metal, Itrax-µ-XRF and hyperspectral scanning, chronology), paleobotanical (pollen) and historical data to reconstruct cyanobacterial abundance and community composition and anthropogenic impacts in two dune lakes over a period of up to 1200 years. Metabarcoding and droplet digital PCR results showed very low levels of picocyanobacteria present in the lakes prior to about CE 1854 (1839-1870 CE) in the smaller shallow Lake Alice and CE 1970 (1963-1875 CE) in the larger deeper Lake Wiritoa. Hereafter bloom-forming cyanobacteria were detected and increased notably in abundance post CE 1984 (1982-1985 CE) in Lake Alice and CE 1997 (1990-2007 CE) in Lake Wiritoa. Currently, the magnitude of blooms is more pronounced in Lake Wiritoa, potentially attributable to hypoxia-induced release of phosphorus from sediment, introducing an additional source of nutrients. Generalized linear modelling was used to investigate the contribution of nutrients (proxy = bacterial functions), temperature, redox conditions (Mn:Fe), and erosion (Ti:Inc) in driving the abundance of cyanobacteria (ddPCR). In Lake Alice nutrients and erosion had a statistically significant effect, while in Lake Wiritoa nutrients and redox conditions were significant.

Characterizing carotenoids in cyanobacterial cultures - Opportunities and implications for paleolimnological studies.

Cyanobacterial blooms are increasing in frequency and intensity globally, impacting lake ecosystem health and posing a risk to human and animal health due to the toxins they can produce. Cyanobacterial pigments preserved in lake sediments provide a useful means of understanding the changes that have led to cyanobacterial blooms in lakes. However, there is some uncertainty as to whether specific carotenoids are unique to certain genera or types of cyanobacteria. To fill this knowledge gap, we analyzed pigments in 34 cyanobacteria cultures and applied the findings to sediments from three New Zealand lakes. The cyanobacterial carotenoids canthaxanthin, echinenone and zeaxanthin were detected in all cultures, whereas myxoxanthophyll was only detected in ten cultures (Microcoleus, Planktothrix and the picocyanobacteria cultures; Synechococcaceae). The sum of the individual carotenoid concentrations provided the strongest relationship with cyanobacterial biomass (R2 = 0.58) and could be used in paleolimnology studies to evaluate general cyanobacterial abundance. Ratios of canthaxanthin, zeaxanthin and myxoxanthophyll relative to echinenone indicated that carotenoid ratios could be used to differentiate picocyanobacteria and bloom-forming cyanobacteria, to some degree. High zeaxanthin/echinenone ratios were measured in picocyanobacteria and low zeaxanthin/echinenone ratios were measured in bloom-forming cyanobacteria. The zeaxanthin/echinenone ratio was applied to sediment core samples where the cyanobacterial community was also evaluated by 16S rRNA gene metabarcoding, with the zeaxanthin/echinenone ratios showing similar patterns to those observed in the cultures. The preliminary assessment described here suggests that zeaxanthin/echinenone ratios could provide a valuable paleoecological proxy for evaluating historical shifts in cyanobacterial communities and warrants further exploration.

Paleo-diatom records reveal ecological change not detected using traditional measures of lake eutrophication.

Lakes provide crucial ecosystem services and harbour unique and rich biodiversity, yet despite decades of research and management focus, cultural eutrophication remains a predominant threat to their health. Our ability to manage lake eutrophication is restricted by the lack of long-term monitoring records. To circumvent this, we developed a bio-indicator approach for inferring trophic level from lake diatom communities and applied this to sediment cores from two lakes experiencing eutrophication stress. Diatom indicators strongly predicted observed trophic levels, and when applied to sediment cores, diatom predicted trophic level reconstructions were consistent with monitoring data and land-use histories. However, there were significant recent shifts in diatom communities not captured by the diatom-based index or monitoring data, suggesting that conventional trophic level indices obscure important ecological change. New approaches, such as the one in this study, are critical to detect early changes in water quality and prevent the decline of lake ecosystems worldwide.

Deciphering the molecular signal from past and alive bacterial communities in aquatic sedimentary archives.

Lake sediments accumulate information on biological communities thus acting as natural archives. Traditionally paleolimnology has focussed on fossilized remains of organisms, however, many organisms do not leave fossil evidence, meaning major ecosystem components are missing from environmental reconstructions. Many paleolimnology studies now incorporate molecular methods, including investigating microbial communities using environmental DNA (eDNA), but there is uncertainty about the contribution of living organisms to molecular inventories. In the present study, we obtained DNA and RNA inventories from sediment spanning 700 years to investigate the contribution of past and active communities to the molecular signal from sedimentary archives. Additionally, a droplet digital PCR (ddPCR) targeting the 16S ribosomal RNA (16S rRNA) gene of the photosynthetic cyanobacterial genera Microcystis was used to explore if RNA signals were from legacy RNA. We posit that the RNA signal is a mixture of legacy RNA, dormant cells, living bacteria and modern-day trace level contaminants that were introduced during sampling and preferentially amplified. The presence of legacy RNA was confirmed by the detection of Microcystis in sediments aged to ~200 years ago. Recent comparisons between 16S rRNA gene metabarcoding and traditional paleo proxies showed that past changes in bacterial communities can be reconstructed from sedimentary archives. The recovery of RNA in the present study has provided new insights into the origin of these signals. However, caution is required during analysis and interpretation of 16S rRNA gene metabarcoding data especially in recent sediments were there are potentially active bacteria.

First transcriptome of the copepod Gladioferens pectinatus subjected to chronic contaminant exposures.

Contaminants are often at low concentrations in ecosystems and their effects on exposed organisms can occur over long periods of time and across multiple generations. Alterations to subcellular mechanistic pathways in response to exposure to contaminants can provide insights into mechanisms of toxicity that methods measuring higher levels of biological may miss. Analysis of the whole transcriptome can identify novel mechanisms of action leading to impacts in exposed biota. The aim of this study was to characterise how exposures to copper, benzophenone and diclofenac across multiple generations altered molecular expression pathways in the marine copepod Gladioferens pectinatus. Results of the study demonstrated differential gene expression was observed in cultures exposure to diclofenac (569), copper (449) and benzophenone (59). Pathways linked to stress, growth, cellular and metabolic processes were altered by exposure to all three contaminants with genes associated with oxidative stress and xenobiotic regulation also impacted. Protein kinase functioning, cytochrome P450, transcription, skeletal muscle contraction/relaxation, mitochondrial phosphate translocator, protein synthesis and mitochondrial methylation were all differentially expressed with all three chemicals. The results of the study also suggested that using dimethyl sulfoxide as a dispersant influenced the transcriptome and future research may want to investigate it's use in molecular studies. Data generated in this study provides a first look at transcriptomic response of G. pectinatus exposed to contaminants across multiple generations, future research is needed to validate the identified biomarkers and link these results to apical responses such as population growth to demonstrate the predictive capacity of molecular tools.

Using metabarcoding and droplet digital PCR to investigate drivers of historical shifts in cyanobacteria from six contrasting lakes.

The frequency and intensity of cyanobacterial blooms is increasing worldwide. Multiple factors are implicated, most of which are anthropogenic. New Zealand provides a useful location to study the impacts of human settlement on lake ecosystems. The first humans (Polynesians) arrived about 750 years ago. Following their settlement, there were marked landscape modifications which intensified after European settlement about 150 years ago. The aims of this study were to reconstruct cyanobacterial communities in six lakes over the last 1000 years and explore key drivers of change. Cyanobacterial environmental DNA was extracted from sediment cores and analysed using metabarcoding and droplet digital PCR. Cyanobacteria, including potentially toxic or bloom forming species, were already present in these lakes prior to human arrival, however their overall abundance was low. Total cyanobacteria abundance and richness increased in all lakes after European settlement but was very pronounced in four lakes, where bloom-forming taxa became dominant. These shifts occurred concomitant with land-use change. The catchment of one deteriorated lake is only moderately modified, thus the introduction of non-native fish is posited as the key factor driving this change. The paleolimnological approach used in this study has enabled new insights into timing and potential causes of changes in cyanobacterial communities.