Eeva-Riikka Vehniäinen

Photoinduced lethal and sublethal toxicity of retene, a polycyclic aromatic hydrocarbon derived from resin acid, to coregonid larvae.

A comparative investigation on the acute phototoxicity of retene to vendace (Coregonus albula) and whitefish (C. lavaretus), both having pelagial larvae in spring, was conducted. To test the concept of early warning of sublethal biomarkers in relation to lethality to posthatch stages, we examined the effects of ultraviolet-B (UV-B) and retene on the levels of cytochrome P4501A (CYP1A) and heat shock protein 70 (HSP70) by exposing the animals to elevated levels of these factors for 48 and 72 h, respectively. Whereas UV-B and retene on their own were not lethal, simultaneous retene and UV-B exposure caused very high mortality to both species. The median lethal concentration (LC50; i.e., the concentration at which half of the larvae died) of retene as a precursor was 41 g/L for vendace and 15 to 16 microg/L, depending on the UV-B dose, for whitefish. Retene evoked substantial induction of CYP1A in larvae of both species, and UV-B induced CYP1A in whitefish. In vendace, no effect on HSP70 levels by any factor was observed. In whitefish, however, UV-B radiation and water retene alone upregulated HSP70, but no additive response was detected. The CYPIA is a biomarker of exposure to retene in both species. The HSP70 is an early warning signal of UV-B exposure in whitefish. As a species, vendace appears to be more resistant than whitefish to the phototoxicity of retene, as indicated by the higher tolerance.

Fluence Rate or Cumulative Dose? Vulnerability of Larval Northern Pike (Esox lucius) to Ultraviolet Radiation

Newly hatched larvae of northern pike were exposed in the laboratory to four fluence rates of ultraviolet radiation (UVR; 290–400 nm) over three different time periods, resulting in total doses ranging from 3.0 ± 0.2 to 63.0 ± 4.4 kJ·m−2. Mortality and behavior of the larvae were followed for 8–12 days, and growth measured at the end of the experiment. Also, the principle of reciprocity—that the UVR‐induced mortality depends on the cumulative dose, independent of fluence rate—was tested. Fluence rates higher than 1480 ± 150 mW·m−2 caused mortality and growth retardation. The highest fluence rate (3040 ± 210 mW·m−2) caused 100% mortality in 5 days. All fluence rates caused behavioral disorders, which led to death at fluence rates higher than 1480 mW·m−2. Reciprocity failure occurred with the lowest and highest dose (550 ± 45 and 3040 ± 210 mW·m−2, respectively). The results show that fluence rate is of primary importance when assessing the UVR‐related risk.

UV-B exposure causes DNA damage and changes in protein expression in northern pike (Esox lucius) posthatched embryos.

The ongoing anthropogenically caused ozone depletion and climate change has increased the amount of biologically harmful UV‐B radiation, which is detrimental to fish in embryonal stages. The effects of UV‐B radiation on the levels and locations of DNA damage manifested as cyclobutane pyrimidine dimers (CPDs), heat shock protein 70 (HSP70) and p53 protein in newly hatched embryos of pike were examined. Pike larvae were exposed in the laboratory to current and enhanced doses of UV‐B radiation. UV‐B exposure caused the formation of CPDs in a fluence rate‐dependent manner, and the CPDs were found deeper in the tissues with increasing fluence rates. UV‐B radiation induced HSP70 in epidermis, and caused plausible p53 activation in the brain and epidermis of some individuals. Also at a fluence rate occurring in nature, the DNA damage in the brain and eyes of pike and changes in protein expression were followed by severe behavioral disorders, suggesting that neural molecular changes were associated with functional consequences.

More accuracy to the EROD measurements--the resorufin fluorescence differs between species and individuals.

Ethoxyresorufin-O-deethylase (EROD) activity is a biomarker of exposure to planar aromatic hydrocarbons, and it is often measured from the S9 fraction. The effect of the liver S9 fraction of seven boreal freshwater fish species on the fluorescence of resorufin was studied. The S9 fractions diminished resorufin fluorescence by 40-80%, and there were large differences between species. Thus, using a resorufin standard curve without the S9 fraction leads to a large underestimation of the EROD activity. Therefore a microwell plate EROD method was developed that takes into account the effect of each sample on resorufin fluorescence. At least two mechanisms were involved in the decrease of the fluorescence: opaqueness of the sample, and enzymes (DT-diaphorase and plausibly NADPH-CYP450 oxidoreductase) that reduce resorufin to a non-fluorescent form.

Recovery of Historically Contaminated Watercourse Polluted by the Chemical Wood Industry: EROD Activity in Fish as Biomarker

Despite outstanding process alterations over decades, pulp- and paper-mill-contaminated sediments and continuing exposure by the effluents may still have effects on biota. In this study, ecotoxicological impacts in the boreal watercourse were analyzed by measuring ethoxyresorufin-O-deethylase (EROD) induction from wild fish populations and from experimentally exposed fish. In order to assess the role of sediment-borne chemicals, juvenile rainbow trout (Oncorhynchus mykiss) were exposed in the laboratory to the surface sediments of Lake Vatianjärvi and Southern Lake Saimaa, both watercourses impacted by the chemical wood industry for approximately a century. Hepatic EROD activity was also measured from roach (Rutilus rutilus) and perch (Perca fluviatilis) caught in Lake Vatianjärvi. Increased EROD activity was not observed in wild fish caught in Lake Vatianjärvi nor in rainbow trout exposed to the sediment of Lake Vatianjärvi, but it was observed in rainbow trout exposed to the sediment of Southern Lake Saimaa, probably explained by critically high concentrations of retene. The results of both the laboratory and field study indicate the absence of EROD-inducing compounds of the previously heavily contaminated Lake Vatianjärvi.

Retene causes multifunctional transcriptomic changes in the heart of rainbow trout (Oncorhynchus mykiss) embryos

Fish are particularly sensitive to aryl hydrocarbon receptor (AhR)-mediated developmental toxicity. The molecular mechanisms behind these adverse effects have remained largely unresolved in salmonids, and for AhR-agonistic polycyclic aromatic hydrocarbons (PAHs). This study explored the cardiac transcriptome of rainbow trout (Oncorhynchus mykiss) eleuteroembryos exposed to retene, an AhR-agonistic PAH. The embryos were exposed to retene (nominal concentration 32 μg/L) and control, their hearts were collected before, at and after the onset of the visible signs of developmental toxicity, and transcriptomic changes were studied by microarray analysis. Retene up- or down-regulated 122 genes. The largest Gene Ontology groups were signal transduction, transcription, apoptosis, cell growth, cytoskeleton, cell adhesion/mobility, cardiovascular development, xenobiotic metabolism, protein metabolism, lipid metabolism and transport, and amino acid metabolism. Together these findings suggest that retene affects multiple signaling cascades in the heart of rainbow trout embryos, and potentially disturbs processes related to cardiovascular development and function.

Multixenobiotic resistance efflux activity in Daphnia magna and Lumbriculus variegatus

Multixenobiotic resistance is a phenomenon in which ATP-binding cassette (ABC) family proteins transfer harmful compounds out of cells. Daphnia magna and Lumbriculus variegatus are model species in aquatic ecotoxicology, but the presence and activity of ABC proteins have not been well described in these species. The aim of this work was to study the presence, activity, and inhibition of ABC transport proteins in D. magna and L. variegatus. The presence of abcb1 and abcc transcripts in 8-9-day-old D. magna was investigated by qRT-PCR. The activity of MXR in D. magna and L. variegatus was explored by influx of the fluorescent ABC protein substrates rhodamine B and calcein-AM, with and without the model inhibitors verapamil (unspecific ABC inhibitor), reversin 205 (ABCB1 inhibitor) and MK571 (ABCC inhibitor). Juvenile D. magna possessed all examined abcb and abcc transcripts, but only reversin 205 inhibited MXR activity. The MXR activity in L. variegatus was inhibited by MK571, and to a lesser extent by verapamil, whereas reversin 205 seemed to stimulate the transport activity. Whereas calcein-AM worked better as an MXR substrate in D. magna, rhodamine B was a better substrate for L. variegatus MXR activity measurements. This is the first report on MXR activity in the order Lumbriculida, subclass Oligochaeta, and class Clitellata.

The effects of treated effluents on the intensity of papillomatosis and HSP70 expression in roach

Epidermal papillomatosis in fish has been proposed as an indicator of environmental stress but experimental evidence of connection between contaminants and papillomatosis in fish is scarce. We studied changes in the intensity of epidermal papillomatosis and the expression of heat shock protein 70 (HSP70) in roach, Rutilus rutilus, exposed to treated pulp mill and municipal effluents. In male roach, the increase in papillomatosis intensity was higher in fish exposed to 15% than in fish exposed to 1.5% concentration of municipal effluent. No differences were observed in papillomatosis development in females, or in HSP70 expression. In all the experiments conducted, the increasing effect of effluents seemed to be more pronounced in male fish suggesting that sex-related factors affected the intensity of papillomatosis after exposure to effluents. The present results indicate that environmentally relevant concentrations of municipal effluents may be contributing to the development of papillomatosis in fish.

Partitioning of nanoparticle‐originated dissolved silver in natural and artificial sediments

Sediments are believed to be a major sink for silver nanoparticles (AgNPs) in the aquatic environment, but there is a lack of knowledge about the environmental effects and behavior of AgNPs in sediments. The release of highly toxic Ag+ through dissolution of AgNPs is one mechanism leading to toxic effects in sediments. We applied an ultrasound-assisted sequential extraction method to evaluate the dissolution of AgNPs and to study the partitioning of dissolved Ag in sediments. Silver was spiked into artificial and 2 natural sediments (Lake Höytiäinen sediment and Lake Kuorinka sediment) as silver nitrate (AgNO3 ), uncoated AgNPs, or polyvinylpyrrolidone-coated AgNPs (PVP-AgNPs). In addition, the total body burdens of Ag in the sediment-dwelling oligochaete Lumbriculus variegatus were assessed over a 28-d exposure period. The dissolution rate was found to be similar between the uncoated AgNP and PVP-AgNP groups. In all sediments, dissolved Ag was mainly bound to the residual fraction of the sediment, followed by iron and manganese oxides or natural organic matter. In Lake Kuorinka sediment, dissolved Ag that originated from PVP-AgNPs was relatively more bioaccessible, also resulting in higher total body burden in L. variegatus than that from uncoated AgNPs or AgNO3 . In artificial sediment and Lake Höytiäinen sediment, AgNO3 was significantly more bioaccessible than AgNPs. Our results highlight the importance of sediment properties and AgNP surface chemistry when evaluating the environmental exposure of AgNPs. Environ Toxicol Chem 2017;36:2593-2601.

Different Sensitivities of Whitefish (Coregonus Lavaretus) and Northern Pike (Esox Lucius) Eleutheroembryos to Photoinduced Toxicity of Polycyclic Aromatic Hydrocarbons

The photoinduced toxicity of two polycyclic aromatic hydrocarbons (PAHs), retene (RET) and pyrene (PYR), to the eleutheroembryos of whitefish and northern pike was studied. Fish were exposed to three concentrations of RET and PYR, and irradiated with ultraviolet radiation (UVR) or visible light for 3 h on two consecutive days. UVR covered the absorption maxima of RET and PYR at UVB and UVA, the daily UVR doses were 30 and 28 kJ m−2, respectively. After 72 h, mortality and behavioral abnormalities were observed. Survivors were analyzed for the proteins of CYP1A, CYP3A, Hsp70, and the activity of glutathione reductase (GR). While neither PAHs nor UVR on their own were lethal, simultaneous exposure caused mortality. The whitefish were more sensitive than the pike to the photoinduced toxicity, and PYR was much more phototoxic than RET. RET induced Hsp70 in both species, and PYR induced GR without UVR in pike.