Characterization of Adverse Outcomes from Legacy-Contaminated Groundwater Exposure to Early Life Stages of Fathead Minnow.

Complex mixtures of chemicals present in groundwater at legacy-contaminated industrial sites can pose significant risks to adjacent surface waters. The combination of short-term molecular and chronic apical effect assessments is a promising approach to characterize the potential hazard of such complex mixtures. The objectives of this study were to: (1) assess the apical effects (survival, growth, development, and liver histopathology) after chronic exposure of early life stages (ELSs) of fathead minnows (FHM; Pimephales promelas) to contaminated groundwater from a legacy-contaminated pesticide manufacturing and packaging plant, and (2) identify possible molecular mechanisms responsible for these effects by comparing results to mechanistic outcomes previously determined by a short-term reduced transcriptome assay (EcoToxChips). This study revealed a significant increase in mortality and prevalence of spinal curvatures, as well as a significant reduction in the length of FHMs exposed to the groundwater mixtures in a concentration-dependent manner. There was an increasing trend in the prevalence of edema in FHMs, though not significantly different from controls. Additionally, no histopathological effects were observed in the liver of FHMs exposed to the groundwater mixtures. Short-term molecular outcomes determined in a parallel study were found to be informative of chronic apical outcomes, including cardiotoxicity, spinal deformities, and liver toxicity. Overall, the results observed in this study demonstrated that short-term transcriptomics analyses could support the hazard assessment of complex contaminated sites.

Time-course of oral toxicity to contaminated groundwater in male Sprague Dawley rats.

Assessing toxicity of complex mixtures of contaminants from industrial sites with historic and ongoing contamination remains a challenge for risk assessors. Groundwater from a pesticide packaging site in Canada containing a complex mixture of known and unknown contaminants was examined in male rats to determine the target organ toxicity. This study determined the time-course of toxicity (7, 14, 28, and 60 days) following ad libitum oral exposure to 0.05% v/v contaminated groundwater compared to tap water (control) in male Sprague Dawley rats (n=5 /group/time). Exposure to groundwater resulted in inflammation, indicated by a statistically significant increase in plasma lymphocyte and neutrophil counts on days 7 and 60, respectively, but a reduction in the plasma alpha 2 macroglobulin levels by day 60. Gonadotoxicity was indicated by a reduced Johnsen score (grading spermatogenesis) in all exposed groups at all time points, while seminiferous epithelial height was reduced on days 7, 14, and 28 compared to controls. Plasma testosterone was reduced in exposed groups on days 7 and 28, accompanied by elevated testicular lipid peroxidation at all time points compared to control. In contrast, lipid peroxidation in the lungs from exposed rats was elevated on days 7, 14, and 28. Plasma symmetric dimethylarginine was elevated on day 14 in the exposed group indicating renal impairment. Taken together, these results indicate that testes, kidney, immune and lung are target organs for the contaminated groundwater from this industrial site. The current study highlights the challenge in hazard assessment for complex mixtures and highlights the need for effects-directed analysis and the continued, albeit limited, use of animal models in toxicity testing.

Hazard assessment of complex legacy-contaminated groundwater mixtures using a novel approach method in adult fathead minnows.

Traditional risk assessment methods face challenges in the determination of drivers of toxicity for complex mixtures such as those present at legacy-contaminated sites. Bioassay-driven analysis across several levels of biological organization represents an approach to address these obstacles. This study aimed to apply a novel transcriptomics tool, the EcoToxChip, to characterize the effects of complex mixtures of contaminants in adult fathead minnows (FHMs) and to compare molecular response patterns to higher-level biological responses. Adult FHMs were exposed for 4 and 21 days to groundwater mixtures collected from a legacy-contaminated site. Adult FHM showed significant induction of micronuclei in erythrocytes, decrease in reproductive capacities, and some abnormal appearance of liver histology. Parallel EcoToxChip analyses showed a high proportion of upregulated genes and a few downregulated genes characteristic of compensatory responses. The three most enriched pathways included thyroid endocrine processes, transcription and translation cellular processes, and xenobiotics and reactive oxygen species metabolism. Several of the most differentially regulated genes involved in these biological pathways could be linked to the apical outcomes observed in FHMs. We concluded that molecular responses as determined by EcoToxChip analysis show promise for informing of apical outcomes and could support risk assessments of complex contaminated sites.

A shift from individual species to ecosystem services effect: Introducing the Eco-indicator Sensitivity Distribution (EcoSD) as an ecosystem services approach to redefining the species sensitivity distribution (SSD) for soil ecological risk assessment.

Incorporating the ecosystem services (ES) approach into soil ecological risk assessment (ERA) has been advocated over the years, but implementing the approach in ERA faces some challenges. However, several researchers have made significant improvements to the soil ERA, such as applying the species sensitivity distribution (SSD) to discern chemical effects on the soil ecosystem. Despite the considerable contributions of SSD to ERA, SSD fails to relate chemical impact on individual species to ES and account for functional redundancy as well as soil ecosystem complexity. Here, we introduce the Eco-indicator Sensitivity Distribution (EcoSD). An EcoSD fits ecological functional groups and soil processes, termed "eco-indicators," instead of individual species responses to a statistical distribution. These eco-indicators are related directly to critical ecosystem functions that drive ES. We derived an EcoSD for cadmium as a model chemical and estimated a soil ecosystem protection value (EcoPVSoil ) based on the eco-indicator dataset for cadmium from the literature. The EcoSD identified nitrogen cycling as the critical process disrupted by cadmium. A key advantage of EcoSD is that it identifies key ecological and chemical indicators of an ES effect. In doing so, it links chemical monitoring results to sensitive ecological functions. The estimated EcoPVSoil for cadmium was slightly more protective of the soil ecosystem than most regional soil values derived from this study's dataset and soil guideline values from the literature. Thus, EcoSD has proven to be a practical and valuable ES concept with the potential to serve as an initial step of the tiered ERA approach. Integr Environ Assess Manag 2023;00:1-14. © 2023 SETAC.

Characterization of molecular and apical effects of legacy-contaminated groundwater on early life stages of fathead minnows.

Mechanistic toxicology approaches represent a promising alternative to traditional live animal testing; however, the often-noted uncertainties concerning the linkages between effects observed at molecular and apical levels curtails the adoption of such approaches. The objective of this study was to apply a novel transcriptomics tool, EcoToxChips, to characterize the effects of complex mixtures of contaminants in fish and to compare molecular response patterns to higher-level biological responses including swimming behavior, deformities, and mortality. Fathead minnow (FHM) embryos were exposed for seven days to increasing concentrations of groundwater collected from moderate (MIAZ) and high (HIAZ) industrial activity zones of a legacy contaminated site. There was a concentration-dependent disruption of photo-dependent swimming responses associated with avoidance behavior patterns and spinal deformities (HIAZ and MIAZ), and an induction of pericardial edema and mortality (HIAZ-10%). Parallel EcoToxChip analyses showed a shift from a majority of upregulated genes at lower concentrations to a majority of downregulated genes at higher concentrations for both treatment conditions. Many of the significantly differentially regulated genes were involved in biological pathways including induction of oxidative stress, activating of several metabolic processes and growth, cell death, and inhibition of signal transduction signaling processes. Several contaminants present in the groundwater mixtures could have contributed to an exceedance of antioxidant system capacities that possibly led to the deformities, altered swimming behaviours, and mortality observed in FHMs. Therefore, molecular response patterns could be linked to apical outcomes observed in this study. Overall, the results observed in this study demonstrate that transcriptomics approaches such as the EcoToxChip system could be supportive of risk assessment of complex contaminated sites.

Integument colour change: Tracking delayed growth of Oppia nitens as a sub-lethal indicator of soil toxicity.

Growth is an important toxicity end-point in ecotoxicology but is rarely used in soil ecotoxicological studies. Here, we assessed the growth change of Oppia nitens when exposed to reference and heavy metal toxicants. To assess mite growth, we developed an image analysis methodology to measure colour spectrum changes of the mite integument at the final developmental stage, as a proxy for growth change. We linked the values of red, green, blue, key-black, and light colour of mites to different growth stages. Based on this concept, we assessed the growth change of mites exposed to cadmium, copper, zinc, lead, boric acid, or phenanthrene at sublethal concentrations in LUFA 2.2 soil for 14 days. Sublethal effects were detected after 7 days of exposure. The growth of O. nitens was more sensitive than survival and reproduction when exposed to copper (EC50growth = 1360 mg/kg compared to EC50reproduction = 2896 mg/kg). Mite growth sensitivity was within the same order of magnitude to mite reproduction when exposed to zinc (EC50growth = 1785; EC50reproduction = 1562 mg/kg). At least 25% of sublethal effects of boric acid and phenanthrene were detected in the mites but growth was not impacted when O. nitens were exposed to lead. Consistent with previous studies, cadmium was the most toxic metal to O. nitens. The mite growth pattern was comparable to mite survival and reproduction from previous studies. Mite growth is a sensitive toxicity endpoint, ecologically relevant, fast, easy to detect, and can be assessed in a non-invasive fashion, thereby complimenting existing O. nitens testing protocols.

Linking niche size and phylogenetic signals to predict future soil microbial relative abundances.

Bacteria provide ecosystem services (e.g., biogeochemical cycling) that regulate climate, purify water, and produce food and other commodities, yet their distribution and likely responses to change or intervention are difficult to predict. Using bacterial 16S rRNA gene surveys of 1,381 soil samples from the Biomes of Australian Soil Environment (BASE) dataset, we were able to model relative abundances of soil bacterial taxonomic groups and describe bacterial niche space and optima. Hold out sample validated hypothetical causal networks (structural equation models; SEM) were able to predict the relative abundances of bacterial taxa from environmental data and elucidate soil bacterial niche space. By using explanatory SEM properties as indicators of microbial traits, we successfully predicted soil bacterial response, and in turn potential ecosystem service response, to near-term expected changes in the Australian climate. The methods developed enable prediction of continental-scale changes in bacterial relative abundances, and demonstrate their utility in predicting changes in bacterial function and thereby ecosystem services. These capabilities will be strengthened in the future with growing genome-level data.

Rocks, lichens, and woody litter influenced the soil invertebrate density in upland tundra heath.

Soil invertebrates are an integral part of Arctic ecosystems through their roles in the breakdown of litter, soil formation, and nutrient cycling. However, studies examining soil invertebrates in the Arctic are limited and our understanding of the abiotic and biotic drivers of these invertebrate communities remains understudied. We examined differences in soil invertebrate taxa (mites, collembolans, enchytraeids) among several undisturbed upland tundra heath sites in Nunavut Canada and identified the drivers (vegetation and substrate cover, soil nutrients and pH) of the soil invertebrate community across these sites. Soil invertebrate densities were similar to that of other Arctic studies. While invertebrate communities were relatively consistent between our sites, cover of rocks, woody litter, and the lichen Alectoria nigricans had significant, positive influences on the density of all invertebrates studied. Mites and collembolans were more closely associated with cover of lichens, whereas enchytraeids were more closely associated with woody litter and rocks. Our results suggest that anthropogenic (e.g., resource exploration and extraction) and/or natural (e.g., climate change) disturbances that result in changes to the vegetation community and woody litter inputs will likely impact soil invertebrates and the ecosystem services they provide.

More than just a substrate for mites: Moss-dominated biological soil crust protected population of the oribatid mite, Oppia nitens against cadmium toxicity in soil.

Metal-impacted sites often need aggressive ecorestoration strategies to restore a functional plant-soil system. The use of biological soil crusts for soil stabilization, moisture retention and C and N input in disturbed and contaminated soils is becoming a more common ecorestoration practice. Biological soil crusts comprise cyanobacteria, fungi, lichens, and bryophytes (mostly moss). Moss-dominated BSCs provide significant N mineralization rate in most terrestrial ecosystems. Oribatid mites or moss mites dominate moss-dominated BSCs and provide essential ecosystem services such as decomposition and nutrient cycling. We hypothesized that moss-dominated BSCs would create a high-quality habitat niche for O. nitens to resist Cd-induced toxicity. Adult mites were exposed to Cd for 28 days in soil with or without BSCs that were aged for eight months. Cadmium toxicity to mites in soil without BSCs was 1.7 and 5.4times greater than in soil with BSCs, respectively for the mites reproduction and instantaneous population growth rate (PGRi). The moss-dominated BSC did not reduce Cd bioavailability in the mites but increased the mite's resilience to Cd toxicity, likely mediated by the trophic transfer of calcium from the BSC to the mites. Our work identifies a second mechanistic avenue by which BSCs are useful for ecorestoration, i.e., the improvement of soil invertebrate physiology to resist metal stress.

Correlation between in vitro toxicity of pesticides and in vivo risk guidelines in support of complex operating site risk management: A meta-analysis.

In vitro cell systems can support hazard characterization and identify mechanisms involved in toxicity; however, using in vitro data for risk assessment still is challenging. As part of an effort to develop approaches for a complex operating site used for biocide packaging and distribution, we evaluated in vitro assays that could be used in a site management format. Across 66 studies, 108 pesticides were assessed on ten human-derived cell types at four endpoints. In vitro IC50s were compared to in vivo guidelines, NOEL/NOAELs, and ADIs using Spearman correlation and linear regression models. While human neuroblastoma cells (SH-SY5Y) were the most sensitive, HepG2 was the most used cell line in evaluating the toxicity of pesticides. Amongst the ten human cell lines, the IC50s derived from SH-SY5Y cells, using MTT-24 & 48 h (the most used assay) correlated (rho = 0.56-0.79; p < 0.05) with ADIs and NOEL/NOAELs. Although in vitro cell systems have some limitations, the correlation between in vitro data derived from SH-SY5Y cells and in vivo safety guidelines can provide site investigators with a tool to survey and prioritize areas and media of concern at complex operating sites impacted by pesticide mixtures.

Response addition is more protective of biogeochemical cycles of carbon and phosphorus compared to concentration addition.

In soils, enzymes are crucial to catalyzing reactions and cycling elements such as carbon (C), nitrogen (N), and phosphorus (P). Although these soil enzymes are sensitive to metals, they are often disregarded in risk assessments, and regulatory laws governing their existence are unclear. Nevertheless, there is a need to develop regulatory standards for metal mixtures that protect biogeochemical cycles because soil serve as a sink for metals and exposures occur as mixtures. Using a fixed ratio ray design, we investigated the effects of 5 single metals and 10 quinary mixtures of Zn, Cu, Ni, Pb, and Co metal oxides on two soil enzymes (i.e., acid phosphatases [ACP] and beta glucosidases [BGD]) in two acidic Canadian soils (S1: acid sandy forest soil, and S2: acid sandy arable soil), closely matched to EU REACH standard soils. Compared to BGD, ACP was generally the more sensitive enzyme to both the single metals and the metal mixtures. The effective concentration inhibiting 50% enzyme activity (EC50) estimates for single Cu (2.1-160.7 mmol kg-1) and Ni (12-272 mmol kg-1) showed that those were the most toxic to both enzymes in both soils. For metal mixtures, response addition (RA) was more conservative in predicting metal effects compared to concentration addition (CA). For both additivity models, antagonism was observed except at lower concentrations (≤10,000 mg/kg) where synergism was observed. At higher concentrations (>10,000 mg/kg), free and CaCl2 extractable Cu protected both enzymes against the toxicity of other metals in the mixture. The results suggest that assuming CA at concentrations less than EC50 does not protect biogeochemical cycling of C and P. And Cu in soil may protect soil enzymes from other toxic metals and thus may have an overall positive role.

Brassica napus Bacterial Assembly Processes Vary with Plant Compartment and Growth Stage but Not between Lines.

Holobiont bacterial community assembly processes are an essential element to understanding the plant microbiome. To elucidate these processes, leaf, root, and rhizosphere samples were collected from eight lines of Brassica napus in Saskatchewan over the course of 10 weeks. We then used ecological null modeling to disentangle the community assembly processes over the growing season in each plant part. The root was primarily dominated by stochastic community assembly processes, which is inconsistent with previous studies that suggest of a highly selective root environment. Leaf assembly processes were primarily stochastic as well. In contrast, the rhizosphere was a highly selective environment. The dominant rhizosphere selection process leads to more similar communities. Assembly processes in all plant compartments were dependent on plant growth stage with little line effect on community assembly. The foundations of assembly in the leaf were due to the harsh environment, leading to dominance of stochastic effects, whereas the stochastic effects in the root interior likely arise due to competitive exclusion or priority effects. Engineering canola microbiomes should occur during periods of strong selection assuming strong selection could promote beneficial bacteria. For example, engineering the microbiome to resist pathogens, which are typically aerially born, should focus on the flowering period, whereas microbiomes to enhance yield should likely be engineered postflowering as the rhizosphere is undergoing strong selection. IMPORTANCE In order to harness the microbiome for more sustainable crop production, we must first have a better understanding of microbial community assembly processes that occurring during plant development. This study examines the bacterial community assembly processes of the leaf, root, and rhizosphere of eight different lines of Brassica napus over the growing season. The influence of growth stage and B. napus line were examined in conjunction with the assembly processes. Understanding what influences the assembly processes of crops might allow for more targeted breeding efforts by working with the plant to manipulate the microbiome when it is undergoing the strongest selection pressure.

Positron-emitting radiotracers spatially resolve unexpected biogeochemical relationships linked with methane oxidation in Arctic soils.

Arctic soils are marked by cryoturbic features, which impact soil-atmosphere methane (CH4 ) dynamics vital to global climate regulation. Cryoturbic diapirism alters C/N chemistry within frost boils by introducing soluble organic carbon and nutrients, potentially influencing microbial CH4 oxidation. CH4 oxidation in soils, however, requires a spatio-temporal convergence of ecological factors to occur. Spatial delineation of microbial activity with respect to these key microbial and biogeochemical factors at relevant scales is experimentally challenging in inherently complex and heterogeneous natural soil matrices. This work aims to overcome this barrier by spatially linking microbial CH4 oxidation with C/N chemistry and metagenomic characteristics. This is achieved by using positron-emitting radiotracers to visualize millimeter-scale active CH4 uptake areas in Arctic soils with and without diapirism. X-ray absorption spectroscopic speciation of active and inactive areas shows CH4 uptake spatially associates with greater proportions of inorganic N in diapiric frost boils. Metagenomic analyses reveal Ralstonia pickettii associates with CH4 uptake across soils along with pertinent CH4 and inorganic N metabolism associated genes. This study highlights the critical relationship between CH4 and N cycles in Arctic soils, with potential implications for better understanding future climate. Furthermore, our experimental framework presents a novel, widely applicable strategy for unraveling ecological relationships underlying greenhouse gas dynamics under global change.

A sustainable colloidal material with sorption and nutrient-supply capabilities for in situ groundwater bioremediation.

Microbial degradation of subsurface organic contaminants is often hindered by the low availability of both contaminants and nutrients, especially phosphorus (P). The use of activated carbon and traditional P fertilizers to overcome these challenges has proved ineffective; therefore, we sought to find an innovative and effective solution. By heating bone meal-derived organic residues in water in a closed reactor, we synthesized nonporous colloids composed of aromatic and aliphatic structures linked to P groups. X-ray absorption near edge spectroscopy analysis revealed that the materials contain mostly bioavailable forms of P (i.e., adsorbed P and magnesium-bearing brushite). The capacity of the materials to adsorb organic contaminants was investigated using benzene and batch isotherm experiments. The adsorption isotherms were fitted to the linearized Freundlich model; isotherm capacity (logKF ) values for the materials ranged between 1.6 and 2.8 µg g-1 . These results indicate that the colloidal materials have a high affinity for organic contaminants. This, coupled with their possession of bioavailable P, should make them effective amendments for in situ groundwater bioremediation. Also, the materials' chemical properties suggest that they are not recalcitrant, implying that they will not become potential contaminants when released into the environment.

Are structural and functional endpoints of soil communities similarly affected by metal mixtures? - A terrestrial model ecosystem approach.

Soils are habitat to a variety of flora and fauna in a linked ecosystem which provides essential ecosystem services. In soil, metals can accumulate at high concentrations, because of anthropogenic activities, leading to toxic effects, threatening the ecosystem and the services it provides. In most real-world contamination scenarios, metals occur as complex mixtures which can interact and produce different toxicity than predicted from individual metal data. Current regulatory guidelines are based on single species responses to individual metals and ignore indirect effects inherent to the inter-linked nature of ecosystems. Also, the evaluation of anthropogenic impacts to the soil communities is usually measured through structural endpoints (e.g. abundance) disregarding functional measurements (e.g. organic matter decomposition rates), which are often seen as tightly related, and thus, similarly affected. In this study we tested three mixture ratios of five metal oxides (lead, copper, nickel, zinc, cobalt) at three dose levels (Low, Med, High) in a terrestrial model ecosystem experiment and measured structural and functional endpoints. Exposure to metal mixtures for 16 weeks did not affect the microarthropod community, but produced severe effects on soil microbial activity (PNR and DHA) reducing activity below 50% compared to control levels, in all dosed treatments. Metal contamination also significantly affected feeding activity and organic matter decomposition, but effects were not as pronounced as on microbial activity. Data suggest that, in the risk assessment of metals and their mixtures, effects on ecosystem structure and functions must be considered to provide adequate environmental protection.

Soil Buffering Capacity Can Be Used To Optimize Biostimulation of Psychrotrophic Hydrocarbon Remediation.

Effective bioremediation of hydrocarbons requires innovative approaches to minimize phosphate precipitation in soils of different buffering capacities. Understanding the mechanisms underlying sustained stimulation of bacterial activity remains a key challenge for optimizing bioremediation-particularly in northern regions. Positron emission tomography (PET) can trace microbial activity within the naturally occurring soil structure of intact soils. Here, we use PET to test two hypotheses: (1) optimizing phosphate bioavailability in soil will outperform a generic biostimulatory solution in promoting hydrocarbon remediation and (2) oligotrophic biostimulation will be more effective than eutrophic approaches. In so doing, we highlight the key bacterial taxa that underlie aerobic and anaerobic hydrocarbon degradation in subarctic soils. In particular, we showed that (i) optimized phosphate bioavailability outperformed generic biostimulatory solutions in promoting hydrocarbon degradation, (ii) oligotrophic biostimulation is more effective than eutrophic approaches, and (iii) optimized biostimulatory solutions stimulated specific soil regions and bacterial consortia. The knowledge gleaned from this study will be crucial in developing field-scale biodegradation treatments for sustained stimulation of bacterial activity in northern regions.

The role of monodentate tetrahedral borate complexes in boric acid binding to a soil organic matter analogue.

Boron is an essential plant micronutrient responsible for several important functions. Boron availability in soils may be influenced by binding with soil organic matter (SOM), particularly with aromatic diol and polyphenol groups on SOM. The mechanism by which aromatic diols bind boron, however, remains unclear. The objective of this work is to further investigate interaction between boric acid and varying concentrations of an aromatic, polyphenolic SOM analogue (tannic acid at 5, 10 and 20 g L-1) from pH = 5-9. UV/Visible spectroscopy showed boric acid enhanced tannic acid deprotonation at pH = 7.0 and 9.0, resulting in singly deprotonated tannic acid subunits. Attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR) showed boric acid/tannic acid binding for all concentrations at pH = 7 and 9, whereas binding at pH = 5.0 was observed only at 20 g L-1 tannic acid. Uncomplexed boron species were not evident at pH = 9.0, but were detectable at pH = 7.0 at lower tannic acid concentrations and prevalent at pH = 5.0, qualitatively indicating binding affinity increases from pH = 5.0 to 9.0. ATR-FTIR results indicated tetrahedral coordination of boron upon complexation to tannic acid with a monodentate mechanism. These results collectively highlight a transition of solution planar boric acid to a tetrahedral, monodentate coordination with a single phenol group in tannic acid polyphenols. This contrasts with previous spectroscopic studies, which indicated bidentate tetrahedral or monodentate trigonal planar orientations prevail at aromatic diol sites. This work presents a previously unobserved boric acid coordination mechanism to an SOM analogue and, therefore, may better inform prediction and modeling of boron behavior in soils.

Community effect concentrations as a new concept to easily incorporate community data in environmental effect assessment of complex metal mixtures.

The goal of this study was to incorporate community data into the effect assessment of environmental and regulatory relevant metal mixtures. In this experiment three fixed mixture ratios (Canadian soil quality guideline ratio - CSQG; Agricultural, residential and Loamy ratio - ARL; and Sudbury ratio - SUD) were tested in a natural community microcosm with 11 doses for each mixture ratio. The effect of metal mixtures on the community was measured using the community effect concentration (EC) concept which assumes that as contamination increases, the community similarity between test and control treatments decreases producing a dose response curve allowing the calculation of community effect concentrations. In regulatory mixture ratios (CSQG and ARL) community EC10s were four times higher than regulatory thresholds and current regulation might be overprotective of the microarthropod communities in some soils. For the contaminated site ratio (SUD), the field dose in the contaminated site corresponded to a community EC20 and if metal concentrations were reduced by 1TU, (from 3.1TU to 2.1TU) effects would be below a community EC10. Overall, the community EC concept was successfully applied and has the potential for inclusion in risk assessment schemes as a measure of community response.

Does habitat quality matter to soil invertebrates in metal-contaminated soils?

This study investigated the influence of habitat quality (HQ) on the reproduction and bioenergetics (energy reserve and metabolic enzyme activities) of the oribatid mite, Oppia nitens, in response to cadmium (Cd). In the baseline toxicity test, Cd elevated the carbohydrate reserve of adult mites at intermediate Cd concentrations (88 and 175 mg Cd kg-1) but without a change in lipid and protein reserve across 0-700 mg Cd kg-1. The activities of glucose metabolism enzymes, glucose-6-phosphate dehydrogenase (G6PDH) and pyruvate kinase (PK) were inhibited in the mites at 700 mg Cd kg-1. Adult mites reared in high HQ soils had higher reproduction relative to mites from low HQ soils when exposed to Cd in OECD soil, but there was no difference in bioenergetics between mites from low and high HQ soils. Hence, HQ significantly (p = 0.024) influenced the reproduction of mites (i.e., juvenile production) irrespective of the Cd concentration in the OECD soil but did not significantly affect the bioenergetics of the mites. We suggest that habitat quality's effect could be more significant than metal concentration on the biological fitness (juvenile production) of O. nitens in metal-contaminated soils.

Is assuming additivity of single-metal toxicity thresholds a conservative approach to assessing risk of ecotoxicity from elevated soil concentrations of cobalt, copper, and nickel at contaminated sites?

Multiple metal-impacted soils are often realistic scenarios for risk assessments, but tools to address these are currently lacking. The objective of this work was to evaluate whether assuming concentration addition (CA) of metal mixture effects was conservative for prospective risk assessment of soils that were elevated mainly in Ni and Cu and somewhat with Co, Pb, or As. Observed whole mixture toxicity for field soils with aged metal mixtures was compared to the expected whole mixture toxicity, assuming additivity of prospective single-metal thresholds ("toxic units") for the mixture components. Bioavailability-adjusted single-metal toxicity thresholds expected for those field soils were the median hazard concentration affecting 5% of species (HC5-50) from the predicted no-effect concentration (PNEC) calculator and calculated from the species-specific dose-response multiple linear relationships (MLRs), all from the European Union Registration, Evaluation, Authorisation and Restriction of Chemicals (EU REACH) dossiers for metals. Generic single-metal toxicity thresholds were based on Canadian Council of Ministers of the Environment soil quality guidelines (CCME SQGs) for agricultural soils. Observed toxicity thresholds were from the community-based risk assessments conducted for Port Colborne and Sudbury, Ontario, Canada. Mostly, prospective single-metal toxicity thresholds were protective relative to the observed toxicity, although that was species or ecological process dependent. The bioavailability-adjusted single-metal thresholds were less conservative than the CCME SQG method, even though the former is based on site-specific EC10 values, and the latter is based on generic EC25 values. When within-site variability in soil properties was used to calculate the 5th and 95th CI for the HC5 sum of toxic units (∑TUs), CA was conservative for far fewer endpoints. In addition, the prospective ∑TUs were more conservative predictions of the observed whole mixture toxicities for Port Colborne soils than for Sudbury soils. The most appropriate balance of accuracy and conservatism for identifying low-level risk of the whole mixtures in these soils appeared to be the bioavailability-adjusted HC5-50, which was applicable to many endpoints and 2 quite different exposure concentration ratios. Integr Environ Assess Manag 2021;17:753-766. © 2020 SETAC.