Cocktail effects and synergistic interactions of chemicals in mixtures are an area of great concern to both the public and regulatory authorities. The main concern is whether some chemicals can ...enhance the effect of other chemicals, so that they jointly exert a larger effect than predicted. This phenomenon is called synergy. Here we present a review of the scientific literature on three main groups of environmentally relevant chemical toxicants: pesticides, metal ions and antifouling compounds. The aim of the review is to determine 1) the frequency of synergy, 2) the extent of synergy, 3) whether any particular groups or classes of chemicals tend to induce synergy, and 4) which physiological mechanisms might be responsible for this synergy. Synergy is here defined as mixtures with minimum two-fold difference between observed and predicted effect concentrations using Concentration Addition (CA) as a reference model and including both lethal and sub-lethal endpoints. The results showed that synergy occurred in 7%, 3% and 26% of the 194, 21 and 136 binary pesticide, metal and antifoulants mixtures included in the data compilation on frequency. The difference between observed and predicted effect concentrations was rarely more than 10-fold. For pesticides, synergistic mixtures included cholinesterase inhibitors or azole fungicides in 95% of 69 described cases. Both groups of pesticides are known to interfere with metabolic degradation of other xenobiotics. For the four synergistic metal and 47 synergistic antifoulant mixtures the pattern in terms of chemical groups inducing synergy was less clear. Hypotheses in terms of mechanisms governing these interactions are discussed. It was concluded that true synergistic interactions between chemicals are rare and often occur at high concentrations. Addressing the cumulative rather than synergistic effect of co-occurring chemicals, using standard models as CA, is therefore regarded as the most important step in the risk assessment of chemical cocktails.
The herbicide, glyphosate, has been shown to stimulate growth in a range of species when applied at doses of 5–60
g a.e.
ha
−1, corresponding to realistic spray drift events. This study investigates ...growth of shoot parameters over time to detect whether the glyphosate induced growth increase was sustained and had a final effect on reproduction. The results showed that an actual biomass growth rate increase took place within the first week after spraying with glyphosate doses <60
g a.e.
ha
−1. This initial growth boost kept treated plants larger than untreated plants for up to six weeks, but at harvest there was no significant difference between control plants and treated plants. Possible effects of glyphosate hormesis on the competitive ability of spray drift affected plants are discussed.
Glyphosate induced hormesis in barley is not sustained over time.
The global biodiversity crisis emphasizes our need to understand how different stressors (climatic, chemical, parasitic, etc.) interact and affect biological communities. We provide a comprehensive ...meta-analysis investigating joint effects of chemical and parasitic stressors for 1064 chemical-parasitic combinations using the Multiplicative model on mortality of arthropods. We tested both features of the experimental setup (control mortality, stressor effect level) and the chemical mode of action, host and parasite phylogeny, and parasite-host interaction traits as explanatory factors for deviations from the reference model. Synergistic interactions, defined as higher mortality than predicted, were significantly more frequent than no interactions or antagony. Experimental setup significantly affected the results, with studies reporting high (> 10%) control mortality or using low stressor effects (< 20%) being more synergistic. Chemical mode of action played a significant role for synergy, but there was no effects of host and parasite phylogeny, or parasite-host interaction traits. The finding that experimental design played a greater role in finding synergy than biological factors, emphasize the need to standardize the design of mixed stressor studies across scientific disciplines. In addition, combinations testing more biological traits e.g. avoidance, coping, and repair processes are needed to test biology-based hypotheses for synergistic interactions.
Azole fungicides are known inhibitors of the important enzyme class cytochrome P450 monooxygenases (CYPs), thereby influencing the detoxification of co-occurring substances via biotransformation. ...This synergism in mixtures containing an azole has mostly been studied by effect measurements, while the underlying mechanism has been less well investigated. In this study, six azole fungicides (cyproconazole, epoxiconazole, ketoconazole, prochloraz, propiconazole, and tebuconazole) were selected to investigate their synergistic potential and their CYP inhibition strength in the aquatic invertebrate Gammarus pulex. The strobilurin fungicide azoxystrobin was chosen as co-occurring substrate, and the synergistic potential was measured in terms of internal concentrations of azoxystrobin and associated biotransformation products (BTPs). Azoxystrobin is biotransformed by various reactions, and 18 BTPs were identified. By measuring internal concentrations of azoxystrobin and its BTPs with high-resolution tandem mass spectrometry in the presence and absence of azole fungicides followed by toxicokinetic modeling, we showed that the inhibition of CYP-catalyzed biotransformation reactions indeed played a role for the observed synergism. However, synergism was only observed for prochloraz at environmentally realistic concentrations. Increased uptake rate constants, an increase in the total internal concentration of azoxystrobin and its BTPs, in vivo assays for measuring CYP activities, and G. pulex video-tracking suggested that the 2-fold increase in bioaccumulation, and, thereby, the raised toxicity of azoxystrobin in the presence of prochloraz is not only caused by inhibited biotransformation but even more by increased azoxystrobin uptake induced by hyperactivity.
Background
Many plants contain phytotoxic alkaloids to deter herbivorous pests and grazing animals. Alkaloids include quinolizidine and indole alkaloids found in the lupin (
Lupinus
spp.), an ...ornamental flower and emerging protein crop, as well as pyrrolizidine alkaloids in the ragwort (
Senecio jacobaea
), an invasive, weed-like flower. When lupins and ragworts are present in large densities in fields, there is a concern that alkaloids may leach into freshwater environments in amounts that may affect non-target organisms, such as
Daphnia magna
. This study aimed to investigate (i) the acute toxicity of alkaloids (gramine, heliotrine, lupanine, lupinine, monocrotaline, monocrotaline N-oxide, senecionine and sparteine) in
D. magna
, (ii) the contribution of these individual alkaloids to lupin plant extract toxicity, (iii) the longer term reproductive effects of a representative alkaloid, sparteine, and conclude with (iv) a tentative risk assessment for the sum of alkaloids measured in soil and surface waters.
Results
The alkaloids exhibited toxicity, with 48 h EC
50
values in the range of 5.6 to > 100 mg/L. The 48 h EC
50
of the
Lupinus angustifolius
plant extract was 1.38 mg/L, which was far more toxic than the simulated extract where lethality was < 10% at 10 mg/L after 48 h. Hence, non-measured compounds may have contributed to the joint toxicity. Daphnid mothers exposed to > 2.5 mg/L sparteine produced significantly fewer and smaller offspring during the 21-day exposure, making chronic effects occur at concentrations approximately 10-fold lower than the 48 h EC
50
for sparteine. The risk assessment of cumulated alkaloids measured in drain, running and pond waters showed a potential risk, particularly for stagnant pond water, where concentrations were severalfold higher than in the drain and running waters.
Conclusions
The results highlight that natural toxins may contribute to poor chemical quality of natural waters, and that natural toxins from upcoming crops or invasive weeds should be considered in aquatic risk assessments.
The first step in xenobiotic detoxification in aquatic invertebrates is mainly governed by the cytochrome P450 mixed function oxidase system. The ability to measure cytochrome P450 activity provides ...an important tool to understand macroinvertebrates’ responses to chemical stressors. However, measurements of P450 activity in small aquatic invertebrates have had variable success and a well characterized assay is not yet available. The general lack of success has been scarcely investigated and it is therefore the focus of the present work. In particular, the suitability of the substrate selected for the assay, the sensitivity of the assay and the possible inhibition/attenuation of enzymatic activity caused by endogenous substances were investigated. 7-ethoxycoumarin-O-dealkylation activity of Daphnia magna, Chironomus riparius larvae and Hyalella azteca was assessed in vivo and in vitro and possible inhibition of enzymatic activity by macroinvertebrates homogenate was investigated. Activities of D. magna and C. riparius larvae measured in vivo were 1.37 ± 0.08 and 2.2 ± 0.2 pmol h⁻¹ organism⁻¹, respectively, while activity of H. azteca could not be detected. In vitro activity could be measured in C. riparius larvae only (500–1000 pmol h⁻¹ mg microsomal protein⁻¹). The optimization of the in vitro assay has been especially long and resource consuming and particularly for D. magna, substances that inhibited cytochrome P450 activity seemed to be released during tissue homogenization preventing activity measurements in vitro. We therefore recommend testing the P450 inhibition potential of homogenate preparations prior to any investigation of P450 activity in vitro in macroinvertebrates.
Applying organic wastes such as manure or sewage sludge on agricultural fields improves soil physical structure and fertility, while giving purpose to otherwise disposed and burned waste products. ...Yet, xenobiotics, and especially pharmaceuticals and personal care products, may be present in these wastes and thereby enter the soil ecosystem. To investigate their occurrence and their potential effect on soil fauna, the CRUCIAL experimental fields, Denmark, were used. Here we combine, for the first time, a chemical wide-scope suspect screening analysis with a study of soil nematode faunal response on soils, which were fertilized with cattle manure, sewage sludge, human urine, and mineral fertilizers over 14 years. The suspect screening analysis was performed with a nanoflow ultra-high performance liquid chromatography coupled with high-resolution tandem mass spectrometry. The abundance of soil indigenous nematodes was determined in each treatment at two time-points, followed by laboratory tests with the nematode species
Caenorhabditis elegans
to assess its reproduction capacity in the soils. A total of 12 pharmaceuticals and personal care products were identified as being up- or down-concentrated in the different treatments compared to controls (mineral fertilizers): 8 and 2 were found to be up-concentrated in sludge- and urine-treated soils, respectively. Our results, however, showed a significantly greater abundance of nematodes in soils amended with sludge and manure, while
C. elegans
had the lowest reproduction capacity in the controls. The reproduction capacity tests did not reveal any chronic toxicity. A principal component analysis showed a clear correlation between nematode abundance and soil organic matter, water content, nutrient contents, and porosity. Overall, urban and animal wastes enhanced soil quality. Pharmaceuticals were detected, but adverse effects on the measured endpoints could not be demonstrated. Although further investigations are needed to examine other associated risks, recycling nutrients from urban areas into fertilizers can contribute to a more sustainable fertilization strategy.
The phenomenon that toxins can be stimulatory at low concentrations to plants has been known now for more than 50 years under the designation of hormesis. Although up to now still largely ...unexplained, the plant growth enhancing effects of chemicals is gaining increased attention in relation to increasing crop production. However, the use of chemical hormesis has been hampered by insufficient predictability of the effects. The reasons for this lack of adequate predictability may be manifold, and knowledge of the possible causes of variations is needed if chemical hormesis is to have practical implications in the field.
With the objective of determining the causes of variability of hormesis, experiments were conducted under controlled conditions to investigate the impact of environmental factors (temperature, light and nutrient availability) on plant growth stimulation by the natural phytotoxin parthenin. A small scale test system using
Lactuca sativa as test species was used to study the expression and progression of parthenin hormesis under various conditions in whole-plant concentration–response experiments.
Results showed that parthenin hormesis is constrained by manipulations of experimental conditions that affect the general plant growth patterns. Parthenin induced hormesis only took place at below maximal but still at good growth conditions for root elongation in
L. sativa. If growth conditions were changed to promote root elongation, the hormetic response diminished or simply disappeared. The same applied for conditions that repressed root elongation. Variation was mainly in the amplitude of hormesis, rather than in the concentration range of hormetic effects. Studying the progression of hormesis for a longer period showed that hormesis remained impaired only at conditions promoting root elongation, while hormesis was merely delayed at poor growth conditions. Hence, the effect of experimental conditions on parthenin hormesis was principally explainable by the effect of experimental factors on growth rate and development of
L. sativa. Based on this, it is questionable if parthenin or other phytotoxin induced hormesis can ultimately be used to predictably increase crop yield under the conditions encountered in the field. The potential might rather be for greenhouse crops, where growth conditions can be more carefully regulated.
Dynamic models for toxic effects toxicokinetic–toxicodynamic (TKTD) models are increasingly used in the analysis of toxicity data for single-chemical exposure. However, these models also offer a ...natural extension to the effects of chemical mixtures. Here, we demonstrate how a simple model for the energy budget (DEBkiss) can be used to interpret the effects of cadmium and fluoranthene, in both single and mixed exposure, on the nematode Caenorhabditis elegans. The data for all time points and all end points (growth and reproduction) are combined into a single coherent framework. These modeling results are compared to a more traditional independent-action approach based on the dose–response curves for a single end point at a single time point. The analysis with DEBkiss does not lead to a radically different interpretation of the mixture effects, both indicating an antagonistic interaction in the mixture. The DEBkiss analysis does, however, provide much more insight into the relevant dynamic processes underlying the toxic effect on the organism and allows for the generation of mechanistic hypotheses that can be used to guide further research.
A wealth of studies has investigated how chemical sensitivity is affected by temperature, however, almost always under different constant rather than more realistic fluctuating regimes. Here we ...compared how the nematode Caenorhabditis elegans responds to copper at constant temperatures (8-24°C) and under fluctuation conditions of low (±4°C) and high (±8°C) amplitude (averages of 12, 16, 20°C and 16°C respectively). The DEBkiss model was used to interpret effects on energy budgets. Increasing constant temperature from 12-24°C reduced time to first egg, life-span and population growth rates consistent with temperature driven metabolic rate change. Responses at 8°C did not, however, accord with this pattern (including a deviation from the Temperature Size Rule), identifying a cold stress effect. High amplitude variation and low amplitude variation around a mean temperature of 12°C impacted reproduction and body size compared to nematodes kept at the matching average constant temperatures. Copper exposure affected reproduction, body size and life-span and consequently population growth. Sensitivity to copper (EC50 values), was similar at intermediate temperatures (12, 16, 20°C) and higher at 24°C and especially the innately stressful 8°C condition. Temperature variation did not increase copper sensitivity. Indeed under variable conditions including time at the stressful 8°C condition, sensitivity was reduced. DEBkiss identified increased maintenance costs and increased assimilation as possible mechanisms for cold and higher copper concentration effects. Model analysis of combined variable temperature effects, however, demonstrated no additional joint stressor response. Hence, concerns that exposure to temperature fluctuations may sensitise species to co-stressor effects seem unfounded in this case.
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