Katherine Jeppe

Genes involved in cysteine metabolism of Chironomus tepperi are regulated differently by copper and by cadmium

Freshwater invertebrates are often exposed to metal contamination, and changes in gene expression patterns can help understand mechanisms underlying toxicity and act as pollutant-specific biomarkers. In this study the expressions of genes involved in cysteine metabolism are characterized in the midge Chironomus tepperi during exposures to sublethal concentrations of cadmium and copper. These metals altered gene expression of the cysteine metabolism differently. Both metals decreased S-adenosylhomocysteine hydrolase expression and did not change the expression of S-adenosylmethionine synthetase. Cadmium exposure likely increased cystathionine production by up-regulating cystathionine-β-synthase (CβS) expression, while maintaining control level cysteine production via cystathionine-γ-lyase (CγL) expression. Conversely, copper down-regulated CβS expression and up-regulated CγL expression, which in turn could diminish cystathionine to favor cysteine production. Both metals up-regulated glutathione related expression (γ-glutamylcysteine synthase and glutathione synthetase). Only cadmium up-regulated metallothionein expression and glutathione S-transferase d1 expression was up-regulated only by copper exposure. These different transcription responses of genes involved in cysteine metabolism in C. tepperi point to metal-specific detoxification pathways and suggest that the transsulfuration pathway could provide biomarkers for identifying specific metals.

Separating the effects of water physicochemistry and sediment contamination on Chironomus tepperi (Skuse) survival, growth and development: a boosted regression tree approach.

More comprehensive ecological risk assessment procedures are needed as the unprecedented rate of anthropogenic disturbances to aquatic ecosystems continues. Identifying the effects of pollutants on aquatic ecosystems is difficult, requiring the individual and joint effects of a range of natural and anthropogenic factors to be isolated, often via the analysis of large, complicated datasets. Ecotoxicologists have traditionally used multiple regression to analyse such datasets, but there are inherent problems with this approach and a need to consider other potentially more suitable methods. Sediment pollution can cause a range of negative effects on aquatic animals, and these are used as the basis for toxicity bioassays to measure the biological impact of pollution and the success of remediation efforts. However, experimental artefacts can also lead to sediments being incorrectly classed as toxic in such studies. Understanding the influence of potentially confounding factors will help more accurate assessments of sediment pollution. In this study, we analysed standardised sediment bioassays conducted using the chironomid Chironomus tepperi, with the aim of modelling the impact of sediment toxicants and water physico-chemistry on four endpoints (survival, growth, median emergence day, and number of emerging adults). We used boosted regression trees (BRT), a method that has a number of advantages over multiple regression, to model bioassay endpoints as a function of water chemistry, sediment quality and underlying geology. Endpoints were generally influenced most strongly by water quality parameters and nutrients, although some metals negatively influenced emergence endpoints. Sub-lethal endpoints were generally better predicted than lethal endpoints; median emergence day was the most sensitive endpoint examined in this study, while the number of emerging adults was the least sensitive. We tested our modelling results by experimentally manipulating sediment and observing the impact on C. tepperi endpoints. For survival, experimental observations were accurately predicted by models, which highlighted the importance of conductivity and dissolved oxygen for this endpoint. In comparison, experimental median emergence day was poorly modelled, most likely due to the influence of a wider range of predictors identified as being important influences on this endpoint in models. To demonstrate how BRT model results compare to more traditional techniques, we analysed survival data using multiple regression. Both models yielded similar results, but boosted regression trees offer important advantages over multiple regression. Our results illustrate how boosted regression trees can be used to analyse complex ecotoxicological datasets, and reinforces the importance of water chemistry in sediment toxicology.

A multi-platform metabolomics approach demonstrates changes in energy metabolism and the transsulfuration pathway in Chironomus tepperi following exposure to zinc

Measuring biological responses in resident biota is a commonly used approach to monitoring polluted habitats. The challenge is to choose sensitive and, ideally, stressor-specific endpoints that reflect the responses of the ecosystem. Metabolomics is a potentially useful approach for identifying sensitive and consistent responses since it provides a holistic view to understanding the effects of exposure to chemicals upon the physiological functioning of organisms. In this study, we exposed the aquatic non-biting midge, Chironomus tepperi, to two concentrations of zinc chloride and measured global changes in polar metabolite levels using an untargeted gas chromatography-mass spectrometry (GC-MS) analysis and a targeted liquid chromatography-mass spectrometry (LC-MS) analysis of amine-containing metabolites. These data were correlated with changes in the expression of a number of target genes. Zinc exposure resulted in a reduction in levels of intermediates in carbohydrate metabolism (i.e., glucose 6-phosphate, fructose 6-phosphate and disaccharides) and an increase in a number of TCA cycle intermediates. Zinc exposure also resulted in decreases in concentrations of the amine containing metabolites, lanthionine, methionine and cystathionine, and an increase in metallothionein gene expression. Methionine and cystathionine are intermediates in the transsulfuration pathway which is involved in the conversion of methionine to cysteine. These responses provide an understanding of the pathways affected by zinc toxicity, and how these effects are different to other heavy metals such as cadmium and copper. The use of complementary metabolomics analytical approaches was particularly useful for understanding the effects of zinc exposure and importantly, identified a suite of candidate biomarkers of zinc exposure useful for the development of biomonitoring programs.

Bifenthrin Causes Toxicity in Urban Stormwater Wetlands: Field and Laboratory Assessment Using Austrochiltonia (Amphipoda)

Stormwater wetlands are engineered to accumulate sediment and pollutants from stormwater and provide environmental value to urban environments. Therefore, contaminated sediment risks causing toxicity to aquatic fauna. This research identifies contaminants of concern in urban wetland sediments by assessing sediment toxicity using the amphipod Austrochiltonia subtenuis. Sediments from 98 wetlands were analyzed for contaminants, and laboratory bioassays were performed with A. subtenuis. Wild Austrochiltonia spp. were also collected from wetlands to assess field populations. Random forest modeling was used to identify the most important variables predicting survival, growth, and field absence of Austrochiltonia spp. Bifenthrin was the most frequently detected pesticide and also the most important predictor of Austrochiltonia spp. responses. Copper, permethrin, chromium, triclosan, and lead were also important. The median lethal effect concentration (LC50) of bifenthrin to laboratory-based A. subtenuis (1.09 (±0.08) μg/gOC) exposed to wetland sediments was supported by a bifenthrin-spiked sediment experiment, indicating A. subtenuis is a suitable test species. Furthermore, Austrochiltonia spp. were absent from all sites that exceeded the calculated bifenthrin LC50, demonstrating the impact of this contaminant on wild populations. This research demonstrates the sensitivity of Austrochiltonia spp. to urban sediment contamination and identifies bifenthrin as a contaminant of concern in urban wetlands.

Toxicant mixtures in sediment alter gene expression in the cysteine metabolism of Chironomus tepperi

Sediment contamination can pose risks to the environment, and sediment toxicity tests have been developed to isolate the impact of sediment from other factors. Mixtures of contaminants often occur in sediments, and traditional endpoints used in toxicity testing, such as growth, reproduction, and survival, cannot discern the cause of toxicity from chemical mixtures because of complex interactions. In urban waterways, the synthetic pyrethroid bifenthrin and the metal copper are commonly found in mixtures, so the present study was designed to investigate how these contaminants cause toxicity in mixtures. To investigate this, Chironomus tepperi was exposed to environmentally relevant concentrations of copper and bifenthrin-spiked sediments in a 2-way factorial mixture for 5 d. Growth and expression profiles of cysteine metabolism genes were measured after exposure. Growth increased at low copper concentrations, decreased at high copper concentrations, and was unaffected by bifenthrin exposures. Copper exposures induced possible cellular repair by upregulating S-adenosylmethionine synthetase expression and downregulating expression of S-adenosylhomocysteine hydrolase and cystathionine-β-synthase. Metallothionein upregulation was also observed. Bifenthrin exposure altered cysteine metabolism to a lesser extent, downregulating cystathionine-β-synthase and γ-glutamylcysteine synthase. Synergistic, antagonistic, and dose-dependent interactions were observed, and there was evidence of conflicting modes of action and limited substrate production. These findings demonstrate how contextual gene expression changes can be sensitive and specific identifiers of toxicant exposure in mixtures. Environ Toxicol Chem 2017;36:691-698.

Potentially Toxic Concentrations of Synthetic Pyrethroids Associated with Low Density Residential Land Use

Trace organic compounds associated with human activity are now ubiquitous in the environment. As the population becomes more urbanised and the use of pesticides and person care products continues to increase, urban waterways are likely to receive higher loads of trace organic contaminants with unknown ecological consequences. To establish the extent of trace organic contamination in urban runoff, concentrations of emerging chemicals of concern were determined in sediments from 99 urban wetlands in and around Melbourne, Australia between February and April, 2015. As a preliminary estimation of potential risks to aquatic biota, we compared measured concentrations with thresholds for acute and chronic toxicity, and modelled toxic units as a function of demographic and land use trends. The synthetic pyrethroid insecticide bifenthrin was common and widespread, and frequently occurred at concentrations likely to cause toxicity to aquatic life. Personal care products DEET and triclosan were common and widely distributed, while the herbicides diuron and prometryn, and the fungicides pyrimethanil and trifloxystrobin occurred less frequently. Toxic unit modelling using random forests found complex and unexpected associations between urban land uses and trace organic concentrations. Synthetic pyrethroid insecticides were identified as emerging compounds of concern, particularly bifenthrin. In contrast with previous surveys, the highest bifenthrin concentrations were associated with lower housing and population density, implicating low-density residential land use in bifenthrin contamination. We discuss the implications for pesticide regulation and urban wetland management in a global context.

Detecting copper toxicity in sediments: from the subindividual level to the population level

Summary 1.Sediments accumulate chemicals that can be toxic to biota and often contribute to aquatic ecosystem decline. Measuring mortality in laboratory-bred organisms is a common way to assess sediment toxicity. However, mortality-based responses of resilient laboratory organisms may not reflect indigenous macroinvertebrate responses, which can be relatively more sensitive to sediment toxicants. A possible solution is to also measure responses at the sub-individual level. 2.Several organism responses to sediment copper toxicity were assessed in a field-based microcosm. Responses of laboratory-bred chironomids and snails deployed in microcosms were compared at sub-individual (metabolomic and gene expression), individual (survival and dry weight) and population (reproduction) levels, and contrasted to the abundance of colonizing macroinvertebrates in the microcosms. 3.Colonizing macroinvertebrate abundance showed a range of sensitivities based on EC50 (effect dose 50% change). Chironomidae made up 94.5% of the microcosm macroinvertebrates, with Paratanytarsus the most sensitive genus (EC50: 89 mg/kg copper) and Procladius the least sensitive (EC50: 681 mg/kg). 4.Survival of laboratory-bred organisms was the least sensitive response, comparable to decreased abundance of the least sensitive macroinvertebrate. Juvenile production in the snail, Potamopyrgus antipodarum, was the most sensitive population level response (EC50: 121 mg/kg), in contrast the snail Physella acuta was relatively more tolerant (EC50: 298 mg/kg). 5.Changes in sub-individual responses (gene expression and metabolite abundance) in laboratory-bred chironomid, Chironomus tepperi, were evident at 60 mg/kg. These changes likely reflect the direct effects of copper exposure and represent metal-specific responses. 6.Synthesis and applications. We showed that copper toxicity in sediments could be readily detected through changes in gene expression and metabolites in laboratory-bred chironomids exposed in field-based microcosms. These responses were more sensitive than mortality, and detected copper levels that caused microcosm chironomid populations to decline. These novel approaches will provide managers with new tools to better assess sediment toxicity. This article is protected by copyright. All rights reserved.

Sensitive community responses of microbiota to copper in sediment toxicity test

Sediment contamination is widespread and can be toxic to aquatic ecosystems and impair human health. Despite their significant ecological function, meio- and microbiota in aquatic ecosystems have been poorly studied in conventional sediment ecotoxicity tests because of the difficulty in sample collecting and identification. In the present study, a novel DNA metabarcoding method was used to assess the effects of spiked copper (Cu) on benthic eukaryotic and prokaryotic communities in laboratory sediment toxicity tests with macroinvertebrates, the chironomid Chironomus tepperi and the amphipod Austrochiltonia subtenuis. In addition to the obvious toxic effects to experimental animals, microbiota (bacteria, protists, algae, and fungi) were significantly altered by spiked Cu in the sediments. The phylogenetic diversity of eukaryotic communities was decreased after spiked-Cu exposure. Even a low-spiked Cu treatment (125 mg/kg) altered structures of eukaryotic and prokaryotic communities in the amphipod experiment. The present study demonstrates that measuring microbiota communities will expand our understanding of the influences of contaminants on aquatic ecosystems. Particularly, the alterations of phylogenetic biodiversity of eukaryotic communities and the structure of sedimentary communities are sensitive indicators for sediment contamination, which can be incorporated in the monitoring and assessment of sediment quality. Environ Toxicol Chem 2018;37:599-608.

Metabolomic Profiles of a Midge (Procladius villosimanus, Kieffer) Are Associated with Sediment Contamination in Urban Wetlands

Metabolomic techniques are powerful tools for investigating organism-environment interactions. Metabolite profiles have the potential to identify exposure or toxicity before populations are disrupted and can provide useful information for environmental assessment. However, under complex environmental scenarios, metabolomic responses to exposure can be distorted by background and/or organismal variation. In the current study, we use LC-MS (liquid chromatography-mass spectrometry) and GC-MS (gas chromatography-mass spectrometry) to measure metabolites of the midge Procladius villosimanus inhabiting 21 urban wetlands. These metabolites were tested against common sediment contaminants using random forest models and metabolite enrichment analysis. Sediment contaminant concentrations in the field correlated with several P. villosimanus metabolites despite natural environmental and organismal variation. Furthermore, enrichment analysis indicated that metabolite sets implicated in stress responses were enriched, pointing to specific cellular functions affected by exposure. Methionine metabolism, sugar metabolism and glycerolipid metabolism associated with total petroleum hydrocarbon and metal concentrations, while mitochondrial electron transport and urea cycle sets associated only with bifenthrin. These results demonstrate the potential for metabolomics approaches to provide useful information in field-based environmental assessments.