Forest top canopy bacterial communities are influenced by elevation and host tree traits.

BACKGROUND: The phyllosphere microbiome is crucial for plant health and ecosystem functioning. While host species play a determining role in shaping the phyllosphere microbiome, host trees of the same species that are subjected to different environmental conditions can still exhibit large degrees of variation in their microbiome diversity and composition. Whether these intra-specific variations in phyllosphere microbiome diversity and composition can be observed over the broader expanse of forest landscapes remains unclear. In this study, we aim to assess the variation in the top canopy phyllosphere bacterial communities between and within host tree species in the temperate European forests, focusing on Fagus sylvatica (European beech) and Picea abies (Norway spruce). RESULTS: We profiled the bacterial diversity, composition, driving factors, and discriminant taxa in the top canopy phyllosphere of 211 trees in two temperate forests, Veluwe National Parks, the Netherlands and Bavarian Forest National Park, Germany. We found the bacterial communities were primarily shaped by host species, and large variation existed within beech and spruce. While we showed that there was a core microbiome in all tree species examined, community composition varied with elevation, tree diameter at breast height, and leaf-specific traits (e.g., chlorophyll and P content). These driving factors of bacterial community composition also correlated with the relative abundance of specific bacterial families. CONCLUSIONS: While our results underscored the importance of host species, we demonstrated a substantial range of variation in phyllosphere bacterial diversity and composition within a host species. Drivers of these variations have implications at both the individual host tree level, where the bacterial communities differed based on tree traits, and at the broader forest landscape level, where drivers like certain highly plastic leaf traits can potentially link forest canopy bacterial community variations to forest ecosystem processes. We eventually showed close associations between forest canopy phyllosphere bacterial communities and host trees exist, and the consistent patterns emerging from these associations are critical for host plant functioning.

Environmental DNA highlights the influence of salinity and agricultural run-off on coastal fish assemblages in the Great Barrier Reef region.

Agricultural run-off in Australia's Mackay-Whitsunday region is a major source of nutrient and pesticide pollution to coastal and inshore ecosystems of the Great Barrier Reef. While the effects of run-off are well documented for the region's coral and seagrass habitats, the ecological impacts on estuaries, the direct recipients of run-off, are less known. This is particularly true for fish communities, which are shaped by the physico-chemical properties of coastal waterways that vary greatly in tropical regions. To address this knowledge gap, we used environmental DNA (eDNA) metabarcoding to examine fish assemblages at four locations (three estuaries and a harbour) subjected to varying levels of agricultural run-off during a wet and dry season. Pesticide and nutrient concentrations were markedly elevated during the sampled wet season with the influx of freshwater and agricultural run-off. Fish taxa richness significantly decreased in all three estuaries (F = 164.73, P = <0.001), along with pronounced changes in community composition (F = 46.68, P = 0.001) associated with environmental variables (largely salinity: 27.48% contribution to total variance). In contrast, the nearby Mackay Harbour exhibited a far more stable community structure, with no marked changes in fish assemblages observed between the sampled seasons. Among the four sampled locations, variation in fish community composition was more pronounced within the wet season (F = 2.5, P = 0.001). Notably, variation in the wet season was significantly correlated with agricultural contaminants (phosphorus: 6.25%, pesticides: 5.22%) alongside environmental variables (salinity: 5.61%, DOC: 5.57%). Historically contaminated and relatively unimpacted estuaries each demonstrated distinct fish communities, reflecting their associated catchment use. Our findings emphasise that while seasonal effects play a key role in shaping the community structure of fish in this region, agricultural contaminants are also important contributors in estuarine systems.

Multiple stressors unpredictably affect primary producers and decomposition in a model freshwater ecosystem.

Freshwater ecosystems are affected by various stressors, such as contamination and exotic species, making them amongst the most imperilled biological systems on the planet. In Australia and elsewhere, copper is one of the most common metal contaminants in freshwater systems and the European carp (Cyprinus carpio L.) is one of the most pervasive and widespread invasive fish species. Copper (Cu) and carp can both directly affect primary production and decomposition, which are critical and interrelated nutrient cycling processes and ecosystem services. The aim of this study was to explore the direct and indirect effects of Cu and carp individually, and together on periphyton cover, chlorophyll a concentration, growth of the macrophyte Vallisneria spiralis L., and the decomposition of leaf litter and cotton strips in a controlled, factorial experiment in outdoor experimental ponds. In isolation, Cu reduced macrophyte growth and organic matter decomposition, while chlorophyll a concentrations and periphyton cover remained unchanged, possibly due to the Low-Cu concentrations in the overlying water. Carp addition alone had a direct negative effect on the biomass of aquatic plants outside protective cages, but also increased plant biomass inside the cages, periphyton cover and chlorophyll a concentrations. Leaf litter was more decomposed in the carp only ponds compared to controls, while there was no significant effect on cotton strip decomposition. Aquatic plants were absent in the Cu + carp ponds caused by the combined effects of Cu toxicity, carp disturbance and the increase in turbidity due to carp bioturbation. Increases in periphyton cover in Low-Cu + carp, while absence in the High-Cu + carp ponds, and differences in the decomposition of surface and buried cotton strips were not as predicted, which highlights the need for such studies to understand the complex interactions among stressors for environmental risk assessment.

Assessing the ecological impacts of NORM-contaminated scale on marine infauna using sediment microcosms.

Naturally occurring radioactive materials (NORMs) can be found in decommissioned oil and gas infrastructure (e.g. pipelines), including scales. The effects of NORM contaminants from offshore infrastructure on benthic macroorganisms remain poorly understood. To test the potential ecological effects of NORM-contaminated scale, we exposed a marine amphipod, a clam and a polychaete to marine sediments spiked with low level concentrations of barium sulfate scale retrieved from a decommissioned subsea pipe. Only amphipods were included in further analysis due to treatment mortalities of the clam and polychaete. Barium (Ba) and copper (Cu) were elevated in the seawater overlying the spiked sediments, although no sediment metals exceeded guidelines. 210Po was the only NORM detected in the overlying waters while both 210Po and 226Ra were significantly elevated in the scale-contaminated sediments when compared with the control sediments. The whole-body burden of Ba and 226Ra were significantly higher in the scale-exposed amphipods. Using experiment- and scale-specific parameters in biota dose assessments suggested potential dose rates may elicit individual and population level effects. Future work is needed to assess the biological impacts and effects of NORM scale at elevated levels above background concentrations and the accumulation of NORM-associated contaminants by marine organisms.

The challenge of implementing environmental DNA metabarcoding to detect elasmobranchs in a resource-limited marine protected area.

Elasmobranchs are threatened and eDNA metabarcoding is a powerful tool that can help efforts to better understand and conserve them. Nevertheless, the inter-calibration between optimal methodological practices and its implementation in resource-limited situations is still an issue. Based on promising results from recent studies, the authors applied a cost-effective protocol with parameters that could be easily replicated by any conservationist. Nonetheless, the results with fewer elasmobranchs detected than expected reveal that endorsed primers and sampling strategies still require further optimization, especially for applications in resource-limited conservation programmes.

Monitoring the Health of Coastal Environments in the Pacific Region-A Review.

Coastal areas provide important ecological services to populations accessing, for example, tourism services, fisheries, minerals and petroleum. Coastal zones worldwide are exposed to multiple stressors that threaten the sustainability of receiving environments. Assessing the health of these valuable ecosystems remains a top priority for environmental managers to ensure the key stressor sources are identified and their impacts minimized. The objective of this review was to provide an overview of current coastal environmental monitoring frameworks in the Asia-Pacific region. This large geographical area includes many countries with a range of climate types, population densities and land uses. Traditionally, environmental monitoring frameworks have been based on chemical criteria set against guideline threshold levels. However, regulatory organizations are increasingly promoting the incorporation of biological effects-based data in their decision-making processes. Using a range of examples drawn from across the region, we provide a synthesis of the major approaches currently being applied to examine coastal health in China, Japan, Australia and New Zealand. In addition, we discuss some of the challenges and investigate potential solutions for improving traditional lines of evidence, including the coordination of regional monitoring programs, the implementation of ecosystem-based management and the inclusion of indigenous knowledge and participatory processes in decision-making.

Characterizing the spatial distributions of soil biota at a legacy base metal mine using environmental DNA.

Characterizing the distribution of biota in response to contaminants is a critical element of site risk assessments. In this study we investigated the spatial distributions of biota and soil chemistry data in surface soil from Sunny Corner, a legacy base metal sulfide mine, Australia. Our results showed that copper (Cu), zinc (Zn), arsenic (As) and lead (Pb) in the surface soil exceeded Australian national soil quality guidelines and posed risks to the environment. Environmental (e)DNA metabarcoding of prokaryote and eukaryote composition confirmed the suggestion of environmental risk posed by these elements collectively explaining 72.9 % and 60.5 % of the total variation in the composition of soil prokaryotes and eukaryotes, respectively. Prokaryotic taxa from the phyla Gemmatimonadetes, Verrucomicrobia and Deinococcus-Thermus showed similar spatial patterns to As and Pb, and were positively correlated. Eukaryotic taxa from the phylum Chlorophyta had similar positive correlations with As and Pb in the soil. In contrast, Amoebozoa and Cercozoa, were sensitive to metals and metalloids, having higher relative abundances in soils with lower concentrations of contaminants. Our study shows that metabarcoding is a promising ecological approach for rapid, large scale assessment of contaminated and potentially impacted sites.

Towards reproducible metabarcoding data: Lessons from an international cross-laboratory experiment.

Advances in high-throughput sequencing (HTS) are revolutionizing monitoring in marine environments by enabling rapid, accurate and holistic detection of species within complex biological samples. Research institutions worldwide increasingly employ HTS methods for biodiversity assessments. However, variance in laboratory procedures, analytical workflows and bioinformatic pipelines impede the transferability and comparability of results across research groups. An international experiment was conducted to assess the consistency of metabarcoding results derived from identical samples and primer sets using varying laboratory procedures. Homogenized biofouling samples collected from four coastal locations (Australia, Canada, New Zealand and the USA) were distributed to 12 independent laboratories. Participants were asked to follow one of two HTS library preparation workflows. While DNA extraction, primers and bioinformatic analyses were purposefully standardized to allow comparison, many other technical variables were allowed to vary among laboratories (amplification protocols, type of instrument used, etc.). Despite substantial variation observed in raw results, the primary signal in the data was consistent, with the samples grouping strongly by geographical origin for all data sets. Simple post hoc data clean-up by removing low-quality samples gave the best improvement in sample classification for nuclear 18S rRNA gene data, with an overall 92.81% correct group attribution. For mitochondrial COI gene data, the best classification result (95.58%) was achieved after correction for contamination errors. The identified critical methodological factors that introduced the greatest variability (preservation buffer, sample defrosting, template concentration, DNA polymerase, PCR enhancer) should be of great assistance in standardizing future biodiversity studies using metabarcoding.

The use of diversity indices for local assessment of marine sediment quality.

Diversity indices are commonly used to measure changes in marine benthic communities. However, the reliability (and therefore suitability) of these indices for detecting environmental change is often unclear because of small sample size and the inappropriate choice of communities for analysis. This study explored uncertainties in taxonomic density and two indices of community structure in our target region, Japan, and in two local areas within this region, and explored potential solutions. Our analysis of the Japanese regional dataset showed a decrease in family density and a dominance of a few species as sediment conditions become degraded. Local case studies showed that species density is affected by sediment degradation at sites where multiple communities coexist. However, two indices of community structure could become insensitive because of masking by community variability, and small sample size sometimes caused misleading or inaccurate estimates of these indices. We conclude that species density is a sensitive indicator of change in marine benthic communities, and emphasise that indices of community structure should only be used when the community structure of the target community is distinguishable from other coexisting communities and there is sufficient sample size.

Impact assessment of ephemeral discharge of contamination downstream of two legacy base metal mines using environmental DNA.

Mining and processing metalliferous ores can degrade the environment well beyond the footprint of the mine, particularly where on-site containment and post-mining remediation has been insufficient to prevent releases of solid and aqueous mine wastes. In this study, we investigated the potential of sediment and water chemistry coupled with environmental (e)DNA metabarcoding to evaluate discrete and cumulative ecological impacts of two legacy base metal (copper (Cu), zinc (Zn), lead (Pb)) mines (Peelwood and Cordillera) which discharge metals via ephemeral tributaries into perennial Peelwood Creek. Although the two mine streams exceeded Australian guidelines for sediment and freshwater quality for Cu, Zn and Pb, Peelwood Creek had relatively low sediment and water metal concentrations, suggesting a low potential for environmental toxicity. Although sediment and water chemistry defined the extent of biological impacts, metabarcoding showed that Peelwood and Cordillera mines had discrete impacts and Peelwood mine was the main source of contamination of Peelwood Creek. Metabarcoding showed that prokaryotes can be good indicators of metal contamination whereas eukaryotes did not reflect contamination impacts in Peelwood Creek. Metabarcoding results showed that benthic communities downstream of Cordillera mine were less impacted than those below Peelwood mine, suggesting that Peelwood mine should be considered for further remediation.

Saline mine-water alters the structure and function of prokaryote communities in shallow groundwater below a tropical stream.

Bacteria and archaea (prokaryotes) are vital components for maintaining healthy function of groundwater ecosystems. The prokaryotic community composition and associated putative functional processes were examined in a shallow sandy aquifer in a wet-dry tropical environment. The aquifer had a contaminated gradient of saline mine-water, which primarily consisted of elevated magnesium (Mg2+) and sulfate (SO42-), although other major ions and trace metals were also present. Groundwaters were sampled from piezometers, approximately 2 m in depth, located in the creek channel upstream and downstream of the mine-water influence. Sampling occurred during the dry-season when only subsurface water flow was present. Next generation sequencing was used to analyse the prokaryote assemblages using 16S rDNA and metabolic functions were predicted with FAPROTAX. Significant changes in community composition and functional processes were observed with exposure to mine-waters. Communities in the exposed sites had significantly lower relative abundance of methanotrophs such as Methylococcaceae and methanogens (Methanobacteriaceae), but higher abundance in Nitrososphaeraceae, associated with nitrification, indicating potentially important changes in the biogeochemistry of the exposed sites. The changes were most strongly correlated with concentrations of SO42-, Mg2+ and Na+. This knowledge allows an assessment of the risk of mine-water contamination to groundwater ecosystem function and aids mine-water management.

Application of environmental DNA for assessment of contamination downstream of a legacy base metal mine.

Acid Rock Drainage (ARD) from legacy mines can negatively impact the biota in sediments and waters for tens of kilometers downstream. Here we used environmental (e)DNA metabarcoding to assess the impacts of metal contaminants on biota in sediment and water downstream of a legacy base metal sulfide mine in southeastern Australia, as exemplar of similar mines elsewhere. Concentrations of metals in water were below Australian water quality guideline values at 20 km downstream for copper (Cu), 40 km downstream for zinc (Zn) and 10 km downstream for lead (Pb). Sediment metal concentrations were below national guideline concentrations at 10 km downstream for Cu, 60 km downstream for Zn and 20 km downstream for Pb. In contrast, metabarcoding showed that biological communities from sediment samples at 10 km and 20 km downstream were similar to sites close to the mine and thus indicative of being impacted, despite metal concentrations being relatively low. As we illustrate, when combined with sediment and water chemistry, metabarcoding can provide more ecological robust perspective on the downstream effects of legacy mines, capturing the sensitivities of a diverse range of organisms.

Metabarcoding Reveals Changes in Benthic Eukaryote and Prokaryote Community Composition along a Tropical Marine Sediment Nickel Gradient.

The Southeast Asia and Melanesia region has extensive nickel (Ni)-rich lateritic regoliths formed from the tropical weathering of ultramafic rocks. As the global demand for Ni continues to rise, these lateritic regoliths are increasingly being exploited for their economic benefit. Mining of these regoliths contributes to the enrichment of coastal sediments in trace metals, especially Ni. The present study used high-throughput sequencing (metabarcoding) to determine changes in eukaryote (18s v7 recombinant DNA [rDNA] and diatom-specific subregion of the 18s v4 rDNA) and prokaryote (16s v4 rDNA) community compositions along a sediment Ni concentration gradient offshore from a large lateritized ultramafic regolith in New Caledonia (Vavouto Bay). Significant changes in the eukaryote, diatom, and prokaryote community compositions were found along the Ni concentration gradient. These changes correlated most with the dilute-acid extractable concentration of Ni in the sediments, which explained 26, 23, and 19% of the variation for eukaryote, diatom, and prokaryote community compositions, respectively. Univariate analyses showed that there was no consistent change in indices of biodiversity, evenness, or richness. Diatom richness and diversity did, however, decrease as sediment acid extractable-Ni concentrations increased. Threshold indicator taxa analysis was conducted separately for each of the 3 targeted genes to detect changes in taxa whose occurrences decreased or increased along the acid extractable-Ni concentration gradient. Based on these data, 46 mg acid extractable-Ni/kg was determined as a threshold value where sensitive species began to disappear. In the case of the estuarine sediments offshore from lateritized ultramafic regolith in New Caledonia, this is recommended as an interim threshold value until further lines of evidence can contribute to a region-specific Ni sediment quality guideline value. Environ Toxicol Chem 2021;40:1894-1907. © 2021 SETAC.

A weight-of-evidence approach for identifying potential sources of untreated sewage inputs into a complex urbanized catchment.

The Hawkesbury-Nepean River (HNR) is the largest catchment in the Sydney region and is undergoing unprecedented population growth. The HNR system receives a mix of anthropogenic inputs such as treated sewage, stormwater and agricultural runoff. Combined, these can diminish the ecological system health and pose potential concerns to human health. Of particular concern are inputs of untreated sewage, that can occur due to a range of different reasons including illegal point source discharges, failure of the sewerage network, and overloading of wastewater treatment plants during storm events. Here, we present findings of an intensive assessment across the HNR catchment where we used a weight-of-evidence (WOE) approach to identify untreated sewage contamination in surface waters against the background of treated effluent and diffuse inputs during post high flow conditions. Total nitrogen and phosphorus concentrations were used to assess treated effluent and diffuse inputs, and microbial analysis, including both culture-based traditional methods for E. coli and enterococci and qPCR analysis of Bacteroides and Lachnospiraceae, were used to assess raw sewage contamination. Despite a background of diffuse inputs from recent high flow events and the influence of treated wastewater, we found no gradient of faecal contamination along the HNR system or its tributaries. We observed two sites with evidence of untreated sewage contamination, where the human markers Bacteroides and Lachnospiraceae qPCR copy numbers were high. The biological and chemical approaches suggested these latter two hotspots originate from an industrial runoff source and possibly from a dry weather sewage leak. Our findings demonstrate the potential of a WOE approach in the assessment of human faecal signal in an urban river that can also pinpoint small sources of contamination as a strategy that can reshape the way monitoring is performed and the chemical end-points chosen to provide pertinent information on the potential risks to aquatic system health.

Ecosystems monitoring powered by environmental genomics: A review of current strategies with an implementation roadmap.

A decade after environmental scientists integrated high-throughput sequencing technologies in their toolbox, the genomics-based monitoring of anthropogenic impacts on the biodiversity and functioning of ecosystems is yet to be implemented by regulatory frameworks. Despite the broadly acknowledged potential of environmental genomics to this end, technical limitations and conceptual issues still stand in the way of its broad application by end-users. In addition, the multiplicity of potential implementation strategies may contribute to a perception that the routine application of this methodology is premature or "in development", hence restraining regulators from binding these tools into legal frameworks. Here, we review recent implementations of environmental genomics-based methods, applied to the biomonitoring of ecosystems. By taking a general overview, without narrowing our perspective to particular habitats or groups of organisms, this paper aims to compare, review and discuss the strengths and limitations of four general implementation strategies of environmental genomics for monitoring: (a) Taxonomy-based analyses focused on identification of known bioindicators or described taxa; (b) De novo bioindicator analyses; (c) Structural community metrics including inferred ecological networks; and (d) Functional community metrics (metagenomics or metatranscriptomics). We emphasise the utility of the three latter strategies to integrate meiofauna and microorganisms that are not traditionally utilised in biomonitoring because of difficult taxonomic identification. Finally, we propose a roadmap for the implementation of environmental genomics into routine monitoring programmes that leverage recent analytical advancements, while pointing out current limitations and future research needs.

Horizon Scan of the Belt and Road Initiative.

The Belt and Road Initiative (BRI) represents the largest infrastructure and development project in human history, and presents risks and opportunities for ecosystems, economies, and communities. Some risks (habitat fragmentation, roadkill) are obvious, however, many of the BRI's largest challenges for development and conservation are not obvious and require extensive consideration to identify. In this first BRI Horizon Scan, we identify 11 frontier issues that may have large environmental and social impacts but are not yet recognised. More generally, the BRI will increase China's participation in international environmental governance. Thus, new cooperative modes of governance are needed to balance geopolitical, societal, and environmental interests. Upgrading and standardising global environmental standards is essential to safeguard ecological systems and human societies.

Environmental DNA can act as a biodiversity barometer of anthropogenic pressures in coastal ecosystems.

Loss of biodiversity from lower to upper trophic levels reduces overall productivity and stability of coastal ecosystems in our oceans, but rarely are these changes documented across both time and space. The characterisation of environmental DNA (eDNA) from sediment and seawater using metabarcoding offers a powerful molecular lens to observe marine biota and provides a series of 'snapshots' across a broad spectrum of eukaryotic organisms. Using these next-generation tools and downstream analytical innovations including machine learning sequence assignment algorithms and co-occurrence network analyses, we examined how anthropogenic pressures may have impacted marine biodiversity on subtropical coral reefs in Okinawa, Japan. Based on 18 S ribosomal RNA, but not ITS2 sequence data due to inconsistent amplification for this marker, as well as proxies for anthropogenic disturbance, we show that eukaryotic richness at the family level significantly increases with medium and high levels of disturbance. This change in richness coincides with compositional changes, a decrease in connectedness among taxa, an increase in fragmentation of taxon co-occurrence networks, and a shift in indicator taxa. Taken together, these findings demonstrate the ability of eDNA to act as a barometer of disturbance and provide an exemplar of how biotic networks and coral reefs may be impacted by anthropogenic activities.

Social Barriers in Ecological Landscapes: The Social Resistance Hypothesis.

Across animal societies, individuals invest time and energy in social interactions. The social landscape that emerges from these interactions can then generate barriers that limit the ability of individuals to disperse to, and reproduce in, groups or populations. Therefore, social barriers can contribute to the difference between the physical capacity for movement through the habitat and subsequent gene flow. We call this contributing effect 'social resistance'. We propose that social resistance can act as an agent of selection on key life-history strategies and promote the evolution of social strategies that facilitate effective dispersal. By linking landscape genetics and social behaviour, the social resistance hypothesis generates predictions integrating dispersal, connectivity, and life-history evolution.

The effect of dissolved nickel and copper on the adult coral Acropora muricata and its microbiome.

The potential impacts of mining activities on tropical coastal ecosystems are poorly understood. In particular, limited information is available on the effects of metals on scleractinian corals which are foundation species that form vital structural habitats supporting other biota. This study investigated the effects of dissolved nickel and copper on the coral Acropora muricata and its associated microbiota. Corals collected from the Great Barrier Reef were exposed to dissolved nickel (45, 90, 470, 900 and 9050 µg Ni/L) or copper (4, 11, 32 and 65 µg Cu/L) in flow through chambers at the National Sea Simulator, Townsville, Qld, Australia. After a 96-h exposure DNA metabarcoding (16S rDNA and 18S rDNA) was undertaken on all samples to detect changes in the structure of the coral microbiome. The controls remained healthy throughout the study period. After 36 h, bleaching was only observed in corals exposed to 32 and 65 µg Cu/L and very high nickel concentrations (9050 µg Ni/L). At 96 h, significant discolouration of corals was only observed in 470 and 900 µg Ni/L treatments, the highest concentrations tested. While high concentrations of nickel caused bleaching, no changes in the composition of their microbiome communities were observed. In contrast, exposure to copper not only resulted in bleaching, but altered the composition of both the eukaryote and bacterial communities of the coral's microbiomes. Our findings showed that these effects were only evident at relatively high concentrations of nickel and copper, reflecting concentrations observed only in extremely polluted environments. Elevated metal concentrations have the capacity to alter the microbiomes which are inherently linked to coral health.