Interspecies comparison of selected pollution biomarkers in dreissenid spp. inhabiting pristine and moderately polluted sites.

Stress biomarkers, which can outline impacts of contaminants in aquatic biota at the biochemical level, are increasingly used as early warning tools in environmental monitoring. Reliable biomarker based assessment schemes, however, request appropriate knowledge of baseline levels of selected endpoints, and the potential influence of a range of natural influencing factors (both abiotic and biotic) as well. In this study, we examined the interspecies variability of various biomarkers (metallothioneins (MT), ethoxyresorufin-O-deethylase activity (EROD), lipid peroxidation (LPO), DNA strand breaks (DNA_sb), vitellogenin-like proteins (Vtg)) in Dreissena polymorpha and Dreissena bugensis inhabiting either pristine- or moderately impacted sites of Lake Balaton (Hungary). Levels of all biomarkers considered revealed low interspecies variability in the two dreissenid species at all sampling sites, with consistently higher (but statistically insignificant) values in Dreissena polymorpha. Levels of all biomarkers varied within the two investigated seasons, with significant influence of the reproduction cycle particularly on the levels of metallothioneins and vitellogenin-like proteins. Each biomarker considered was elevated by October, with significantly higher values in the mussels inhabiting harbours. Insignificant spatial and temporal variability in the general health indicators (condition index, total protein content) of dreissenids was observed, which, in parallel with evident rise in biomarker levels, apparently suggest that the anthropogenic impacts in harbours affect mussel fitness yet at sub organismal level. Our data might serve useful basis for future environmental monitoring surveys, especially in habitats where the progressive replacement of Dreissena polymorpha by Dreissena bugensis is taking place, as the interspecies variability in susceptibility to chemical stress of the two species is well comparable.

Seasonal and size-related variation of subcellular biomarkers in quagga mussels (Dreissena bugensis) inhabiting sites affected by moderate contamination with complex mixtures of pollutants.

The size-related differences in subcellular biomarker responses were assessed in Dreissena bugensis mussels inhabiting harbours moderately affected by pollution with complex mixtures of heavy metals and polycyclic aromatic hydrocarbons (PAHs). Adult D. bugensis samples were collected from three harbours of Lake Balaton (Hungary) characterized by moderate shipping activity, and as reference site, from a highly protected remote area of the lake. Biomarkers of exposure (metallothioneins (MTs), ethoxyresorufin-o-deethylase (EROD)), oxidative stress (lipid peroxidation (LPO), DNA strand breaks (DNAsb)) and possible endocrine disruption (vitellogenin-like proteins (VTG)) were analysed in whole-tissue homogenates of differently sized groups of mussels in relation to environmental parameters and priority pollutants (heavy metals and polycyclic aromatic hydrocarbons). Integrated biomarker response (IBR) indices were calculated for biomarker responses gained through in situ measurements to signalize critical sites and to better distinguish natural tendencies from biological effects of contaminants. Biomarker responses showed close positive correlation in case of MT, EROD, LPO, and DNAsb and negative correlation with VTG levels with mussel shell length in autumn, when higher levels of biomarkers appeared, possibly due to natural lifecycle changes of animals.

Neonicotinoid insecticides inhibit cholinergic neurotransmission in a molluscan (Lymnaea stagnalis) nervous system.

Neonicotinoids are highly potent and selective systemic insecticides, but their widespread use also has a growing impact on non-target animals and contaminates the environment, including surface waters. We tested the neonicotinoid insecticides commercially available in Hungary (acetamiprid, Mospilan; imidacloprid, Kohinor; thiamethoxam, Actara; clothianidin, Apacs; thiacloprid, Calypso) on cholinergic synapses that exist between the VD4 and RPeD1 neurons in the central nervous system of the pond snail Lymnaea stagnalis. In the concentration range used (0.01-1 mg/ml), neither chemical acted as an acetylcholine (ACh) agonist; instead, both displayed antagonist activity, inhibiting the cholinergic excitatory components of the VD4-RPeD1 connection. Thiacloprid (0.01 mg/ml) blocked almost 90% of excitatory postsynaptic potentials (EPSPs), while the less effective thiamethoxam (0.1 mg/ml) reduced the synaptic responses by about 15%. The ACh-evoked membrane responses of the RPeD1 neuron were similarly inhibited by the neonicotinoids, confirming that the same ACh receptor (AChR) target was involved. We conclude that neonicotinoids act on nicotinergic acetylcholine receptors (nAChRs) in the snail CNS. This has been established previously in the insect CNS; however, our data indicate differences in the background mechanism or the nAChR binding site in the snail. Here, we provide the first results concerning neonicotinoid-related toxic effects on the neuronal connections in the molluscan nervous system. Aquatic animals, including molluscs, are at direct risk while facing contaminated surface waters, and snails may provide a suitable model for further studies of the behavioral/neuronal consequences of intoxication by neonicotinoids.

Evaluation of multixenobiotic resistance in dreissenid mussels as a screening tool for toxicity in freshwater sediments.

The multixenobiotic defense mechanism (MXR) in aquatic organisms was recognized as a first-line defense system, and its potential use as an early biomarker of exposure to environmental stress has raised attention in the last two decades. To evaluate the relevance of this biomarker in the freshwater mussel Dreissena polymorpha, we studied its responsiveness within laboratory exposures to contaminants sequestered in freshwater sediments affected by moderate anthropogenic impact. The effectiveness of this biomarker was assessed by comparing the MXR-transporter activities determined in bivalves first with toxicity scores recorded with the D. rerio embryo developmental assay. Both bioassays were applied in the sediment contact test format. As a second evaluation approach, MXR activities determined in exposed mussels were compared with sediment-contamination data integrated into toxic units on the basis of acute toxicity to Daphnia magna. In D. polymorpha subjected to acute exposure with moderately polluted sediments, we detected limited (22-33 %) but statistically significant induction of MXR activity. Mean MXR activities significantly correlated with TU values computed for test sediments. MXR activities in mussels showed strong positive correlation with the metal load of sediments and proved to be unrelated to the contamination with polycyclic aromatic compounds. MXR activity in laboratory-exposed mussels showed low variability within treatments and thus reliably reflected even low contaminant differences between the negative reference and moderately polluted harbor sediments. The strong correlation found in this study between the MXR-transporter activity in exposed mussels and environmentally realistic sediment contamination underscores the fairly good sensitivity of this biomarker in laboratory testing conditions to signal the bioavailability of sediment bound contaminants, and it may also anticipate even the incidence of toxicity to biota.

Ecotoxicological characterisation of sedimentation in the Kis-Balaton Water Protection System.

The main function of the Kis-Balaton Water Protection System is to retain nutrients and total suspended solids, thus protecting the water quality of Lake Balaton. In this paper, the toxic nature of the sediment in the 2nd reservoir of the KBWPS has been characterised, using a battery of tests: Vibrio fischeri acute bioassay on whole sediment samples, and V. fischeri bioassay on pore water and elutriate samples. The latest version of the V. fischeri bioluminescence inhibition was applied, the Flash assay which uses a kinetic mode and is able to detect the toxicity of solid, turbid/coloured samples. Whole sediment toxicity showed a clear spatial distribution of toxicity, in parallel with elutriate toxicity. However, no pore water toxicity was detected, leading to the conclusion that contaminants are not water soluble.

Effects of rotenone and other mitochondrial complex I inhibitors on the brine shrimp Artemia.

(Artemia) nauplii was used to asses the toxicity of rotenone, MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine), MP+ (1-methyl-4-phenylpyridinium) and the effect of L-DOPA co-treatment with rotenone. Rotenone had a dose dependent effect on mortality (LC50: 0.37 ± 0.04 µM mean ± S E, n = 24), while MPTP and MP+ proved to be toxic in millimolar range (LC50: 0.21 ± 0.09 mM and 0.20 ± 0.08 mM, respectively, n = 4). L-DOPA (50-200 µM) co-treatment increased the survival of the rotenone-treated animals (LC50: 0.51 ± 0.23 µM, 1.03 ± 0.66 µM, and 0.76 ± 0.52 µM, respectively). In the whole body tissue homogenates of Artemia, sublethal (up to 0.3 µM) concentrations of rotenone increased the glutathione S-transferase (GST) activity by up to 50 about percent (LC50: 53.3 ± 6.8 nM/min/mg protein, against 34.7 ± 3.6 nM/min/mg protein, n = 4). Nauplii treated in 100 mM L-DOPA and rotenone together showed further increase of GST activity all across the range of rotenone concentrations. These results on Artemia nauplii show similarities with other animal models, when complex I inhibitors were tested. Biochemical measurements suggest a protective role of L-DOPA by increasing the GST activity as part of the intracellular defences during toxin-evoked oxidative stress.