Emergent synergistic lysosomal toxicity of chemical mixtures in molluscan blood cells (hemocytes)

The problem of effective assessment of risk posed by complex mixtures of toxic chemicals in the environment is a major challenge for government regulators and industry. The biological effect of the individual contaminants, where these are known, can be measured; but the problem lies in relating toxicity to the multiple constituents of contaminant cocktails. The objective of this study was to test the hypothesis that diverse contaminant mixtures may cause a greater toxicity than the sum of their individual parts, due to synergistic interactions between contaminants with different intracellular targets. Lysosomal membrane stability in hemocytes from marine mussels was used for in vitro toxicity tests; and was coupled with analysis using the isobole method and a linear additive statistical model. The findings from both methods have shown significant emergent synergistic interactions between environmentally relevant chemicals (i.e., polycyclic aromatic hydrocarbons, pesticides, biocides and a surfactant) when exposed to isolated hemocytes as a mixture of 3 & 7 constituents. The results support the complexity-based hypothesis that emergent toxicity occurs with increasing contaminant diversity, and raises questions about the validity of estimating toxicity of contaminant mixtures based on the additive toxicity of single components. Further experimentation is required to investigate the potential for interactive effects in mixtures with more constituents (e.g., 50 –100) at more environmentally realistic concentrations in order to test other regions of the model, namely, very low concentrations and high diversity. Estimated toxicant diversity coupled with tests for lysosomal damage may provide a potential tool for determining the toxicity of estuarine sediments, dredge spoil or contaminated soil

Effect of Sub-Lethal Exposure to Ultraviolet Radiation on the Escape Performance of Atlantic Cod Larvae (Gadus morhua)

The amount of ultraviolet (UV) radiation reaching the earth's surface has increased due to depletion of the ozone layer. Several studies have reported that UV radiation reduces survival of fish larvae. However, indirect and sub-lethal impacts of UV radiation on fish behavior have been given little consideration. We observed the escape performance of larval cod (24 dph, SL: 7.6±0.2 mm; 29 dph, SL: 8.2±0.3 mm) that had been exposed to sub-lethal levels of UV radiation vs. unexposed controls. Two predators were used (in separate experiments): two-spotted goby (Gobiusculus flavescens; a suction predator) and lion's mane jellyfish (Cyanea capillata; a “passive" ambush predator). Ten cod larvae were observed in the presence of a predator for 20 minutes using a digital video camera. Trials were replicated 4 times for goby and 5 times for jellyfish. Escape rate (total number of escapes/total number of attacks ×100), escape distance and the number of larvae remaining at the end of the experiment were measured. In the experiment with gobies, in the UV-treated larvae, both escape rate and escape distance (36%, 38±7.5 mm respectively) were significantly lower than those of control larvae (75%, 69±4.7 mm respectively). There was a significant difference in survival as well (UV: 35%, Control: 63%). No apparent escape response was observed, and survival rate was not significantly different, between treatments (UV: 66%, Control: 74%) in the experiment with jellyfish. We conclude that the effect and impact of exposure to sub-lethal levels of UV radiation on the escape performance of cod larvae depends on the type of predator. Our results also suggest that prediction of UV impacts on fish larvae based only on direct effects are underestimations

Multi-Drug Resistance Transporters and a Mechanism-Based Strategy for Assessing Risks of Pesticide Combinations to Honey Bees

Annual losses of honey bee colonies remain high and pesticide exposure is one possible cause. Dangerous combinations of pesticides, plant-produced compounds and antibiotics added to hives may cause or contribute to losses, but it is very difficult to test the many combinations of those compounds that bees encounter. We propose a mechanism-based strategy for simplifying the assessment of combinations of compounds, focusing here on compounds that interact with xenobiotic handling ABC transporters. We evaluate the use of ivermectin as a model substrate for these transporters. Compounds that increase sensitivity of bees to ivermectin may be inhibiting key transporters. We show that several compounds commonly encountered by honey bees (fumagillin, Pristine, quercetin) significantly increased honey bee mortality due to ivermectin and significantly reduced the LC50 of ivermectin suggesting that they may interfere with transporter function. These inhibitors also significantly increased honey bees sensitivity to the neonicotinoid insecticide acetamiprid. This mechanism-based strategy may dramatically reduce the number of tests needed to assess the possibility of adverse combinations among pesticides. We also demonstrate an in vivo transporter assay that provides physical evidence of transporter inhibition by tracking the dynamics of a fluorescent substrate of these transporters (Rhodamine B) in bee tissues. Significantly more Rhodamine B remains in the head and hemolymph of bees pretreated with higher concentrations of the transporter inhibitor verapamil. Mechanism-based strategies for simplifying the assessment of adverse chemical interactions such as described here could improve our ability to identify those combinations that pose significantly greater risk to bees and perhaps improve the risk assessment protocols for honey bees and similar sensitive species