Linking sub-cellular biomarkers to embryo aberrations in the benthic amphipod Monoporeia affinis.

To adequately assess and monitor environmental status in the aquatic environment a broad approach is needed that integrates physical variables, chemical analyses and biological effects at different levels of the biological organization. Embryo aberrations in the Baltic Sea key species Monoporeia affinis can be induced by both metals and organic substances as well as by hypoxia, increasing temperatures and malnutrition. This amphipod has therefore been used for more than three decades as a biological effect indicator in monitoring and assessment of chemical pollution and environmental stress. However, little is known about the sub-cellular mechanisms underlying embryo aberrations. An improved mechanistic understanding may open up the possibility of including sub-cellular alterations as sensitive warning signals of stress-induced embryo aberrations. In the present study, M. affinis was exposed in microcosms to 4 different sediments from the Baltic Sea. After 88-95 days of exposure, survival and fecundity were determined as well as the frequency and type of embryo aberrations. Moreover, oxygen radical absorption capacity (ORAC) was assayed as a proxy for antioxidant defense, thiobarbituric acid reactive substances (TBARS) level as a measure of lipid peroxidation and acetylcholinesterase (AChE) activity as an indicator of neurotoxicity. The results show that AChE and ORAC can be linked to the frequency of malformed embryos and arrested embryo development. The occurrence of dead broods was significantly associated with elevated TBARS levels. It can be concluded that these sub-cellular biomarkers are indicative of effects that could affect Darwinian fitness and that oxidative stress is a likely mechanism in the development of aberrant embryos in M. affinis.

Bacteria-mediated effects of antibiotics on Daphnia nutrition.

In polluted environments, contaminant effects may be manifested via both direct toxicity to the host and changes in its microbiota, affecting bacteria-host interactions. In this context, particularly relevant is exposure to antibiotics released into environment. We examined effects of the antibiotic trimethoprim on microbiota of Daphnia magna and concomitant changes in the host feeding. In daphnids exposed to 0.25 mg L(-1) trimethoprim for 24 h, the microbiota was strongly affected, with (1) up to 21-fold decrease in 16S rRNA gene abundance and (2) a shift from balanced communities dominated by Curvibacter, Aquabacterium, and Limnohabitans in controls to significantly lower diversity under dominance of Pelomonas in the exposed animals. Moreover, decreased feeding and digestion was observed in the animals exposed to 0.25-2 mg L(-1) trimethoprim for 48 h and then fed 14C-labeled algae. Whereas the proportion of intact algal cells in the guts increased with increased trimethoprim concentration, ingestion and incorporation rates as well as digestion and incorporation efficiencies decreased significantly. Thus, antibiotics may impact nontarget species via changes in their microbiota leading to compromised nutrition and, ultimately, growth. These bacteria-mediated effects in nontarget organisms may not be unique for antibiotics, but also relevant for environmental pollutants of various nature.

Feeding activity and xenobiotics modulate oxidative status in Daphnia magna: implications for ecotoxicological testing.

To apply biomarkers of oxidative stress in laboratory and field settings, an understanding of their responses to changes in physiological rates is important. The evidence is accumulating that caloric intake can increase production of reactive oxygen species and thus affect background variability of oxidative stress biomarkers in ecotoxicological testing. This study aimed to delineate effects of food intake and xenobiotics on oxidative biomarkers in Daphnia magna. Antioxidant capacity measured as oxygen radical absorbance capacity (ORAC) and lipid peroxidation assayed as thiobarbituric acid reactive substances (TBARS) were measured. Food intake was manipulated by varying food densities or by exposing the animals to chemicals inhibiting feeding rate (pharmaceutical haloperidol and pesticide lindane). Feeding rate proved to affect both protein, ORAC, and TBARS in unexposed daphnids. However, there was no significant effect of feeding rate on the protein-specific ORAC values. Both substances affected individual protein and ORAC levels and changed their relationship to feeding rate. Our results show that inhibition of feeding rate influenced the interpretation of biomarker response and further emphasize the importance of understanding (1) baseline variability in potential biomarkers due to variations in metabolic state and (2) the contribution of feeding rate on toxic response of biomarkers.

Are pharmaceuticals with evolutionary conserved molecular drug targets more potent to cause toxic effects in non-target organisms?

The ubiquitous use of pharmaceuticals has resulted in a continuous discharge into wastewater and pharmaceuticals and their metabolites are found in the environment. Due to their design towards specific drug targets, pharmaceuticals may be therapeutically active already at low environmental concentrations. Several human drug targets are evolutionary conserved in aquatic organisms, raising concerns about effects of these pharmaceuticals in non-target organisms. In this study, we hypothesized that the toxicity of a pharmaceutical towards a non-target invertebrate depends on the presence of the human drug target orthologs in this species. This was tested by assessing toxicity of pharmaceuticals with (miconazole and promethazine) and without (levonorgestrel) identified drug target orthologs in the cladoceran Daphnia magna. The toxicity was evaluated using general toxicity endpoints at individual (immobility, reproduction and development), biochemical (RNA and DNA content) and molecular (gene expression) levels. The results provide evidence for higher toxicity of miconazole and promethazine, i.e. the drugs with identified drug target orthologs. At the individual level, miconazole had the lowest effect concentrations for immobility and reproduction (0.3 and 0.022 mg L-1, respectively) followed by promethazine (1.6 and 0.18 mg L-1, respectively). At the biochemical level, individual RNA content was affected by miconazole and promethazine already at 0.0023 and 0.059 mg L-1, respectively. At the molecular level, gene expression for cuticle protein was significantly suppressed by exposure to both miconazole and promethazine; moreover, daphnids exposed to miconazole had significantly lower vitellogenin expression. Levonorgestrel did not have any effects on any endpoints in the concentrations tested. These results highlight the importance of considering drug target conservation in environmental risk assessments of pharmaceuticals.