Mirella Kanerva

Variation of basal EROD activities in ten passerine bird species--relationships with diet and migration status.

Inter-specific differences in animal defence mechanisms against toxic substances are currently poorly understood. The ethoxyresorufin-O-deethylase (EROD) enzyme plays an important role in defence against toxic chemicals in a wide variety of animals, and it is an important biomarker for environmental contamination. We compared basal hepatic EROD activity levels among ten passerine species to see if there is inter-specific variation in enzyme activity, especially in relation to their diet and migration status. Migratory insectivores showed higher EROD activity compared to granivores. We hypothesize that the variable invertebrate diet of migratory insectivores contains a wider range of natural toxins than the narrower diet of granivores. This may have affected the evolution of mixed function oxidases (MFO) system and enzyme activities. We further tested whether metabolic rates or relative liver size were associated with the variation in detoxification capacity. We found no association between EROD activity and relative (per mass unit) basal metabolic rate (BMR). Instead, EROD activity and relative liver mass (% of body mass) correlated positively, suggesting that a proportionally large liver also functions efficiently. Our results suggest that granivores and non-migratory birds may be more vulnerable to environmental contaminants than insectivores and migratory birds. The diet and migration status, however, are phylogenetically strongly connected to each other, and their roles cannot be fully separated in our analysis with only ten passerine species.

Oxidative status in nestlings of three small passerine species exposed to metal pollution.

Antioxidant defense has an important role in the protection of organisms against oxidative stress caused by reactive oxygen species (ROS). Many metals are capable of generating ROS and inducing oxidative damage, and may therefore lead to changes in oxidative regulation. We studied species-specific variation in the oxidative status of great tit (Parus major), blue tit (Cyanistes caeruleus) and pied flycatcher (Ficedula hypoleuca) nestlings in a vicinity of a non-ferrous smelter. Non-enzymatic (glutathione [tGSH], GSH:GSSG ratio, and carotenoids) and enzymatic (glutathione peroxidase [GP], glutathione-S-transferase [GST], superoxide dismutase [SOD], and catalase [CAT]) antioxidants were evaluated to determine the effects of metal exposure on the oxidative status of the birds. We found strong evidence of interspecific variation in CAT and SOD activities, whereas less variation was observed in parameters related to glutathione metabolism. Oxidative state (in terms of tGSH and GSH:GSSG) did not vary between species, suggesting that different species may employ different antioxidant pathways to achieve the same oxidative state. Oxidative status was only weakly related to metal exposure, and these associations were further obscured by species-specific environmental effects. Our results indicate that effects on oxidative status observed in one species cannot be generalized to other ones. Future work should attempt to incorporate species-specific biology and environmental context into assessments of contaminant impacts on oxidative regulation of passerine birds.

Warm acclimation and oxygen depletion induce species-specific responses in salmonids

ABSTRACT Anthropogenic activities are greatly altering the habitats of animals, whereby fish are already encountering several stressors simultaneously. The purpose of the current study was to investigate the capacity of fish to respond to two different environmental stressors (high temperature and overnight hypoxia) separately and together. We found that acclimation to increased temperature (from 7.7±0.02°C to 14.9±0.05°C) and overnight hypoxia (daily changes from normoxia to 63–67% oxygen saturation), simulating climate change and eutrophication, had both antagonistic and synergistic effects on the capacity of fish to tolerate these stressors. The thermal tolerance of Arctic char (Salvelinus alpinus) and landlocked salmon (Salmo salar m. sebago) increased with warm acclimation by 1.3 and 2.2°C, respectively, but decreased when warm temperature was combined with overnight hypoxia (by 0.2 and 0.4°C, respectively). In contrast, the combination of the stressors more than doubled hypoxia tolerance in salmon and also increased hypoxia tolerance in char by 22%. Salmon had 1.2°C higher thermal tolerance than char, but char tolerated much lower oxygen levels than salmon at a given temperature. The changes in hypoxia tolerance were connected to the responses of the oxygen supply and delivery system. The relative ventricle mass was higher in cold- than in warm-acclimated salmon but the thickness of the compact layer of the ventricle increased with the combination of warm and hypoxia acclimation in both species. Char had also significantly larger hearts and thicker compact layers than salmon. The results illustrate that while fish can have protective responses when encountering a single environmental stressor, the combination of stressors can have unexpected species-specific effects that will influence their survival capacity. Summary: Overnight hypoxia combined with warm acclimation increases the hypoxia tolerance of salmonids while upper critical temperature is reduced in a species-specific manner.

Resistance to oxidative damage but not immunosuppression by organic tin compounds in natural populations of Daubenton's bats (Myotis daubentonii).

The acute toxicity of organic tin compounds (OTCs) has been studied in detail. However, due to their complex nature, very little is known about species-specific methods of accumulation and consequences for food-webs. Chironomids, on which e.g. Daubentons bats feed, may act as vectors for the transport of organic tin compounds from aquatic to terrestrial ecosystems. Bats are prone to environmental toxins because of their longevity and their ecological role as top predators. Organic tin compounds are associated with increased formation of reactive oxygen species and associated oxidative damage as well as suppression of immune function. The present paper investigates whether the OTC, tributyltin (TBT) and its metabolite, dibutyltin (DBT), accumulate in natural populations of Daubentons bats and whether TBT-associated effects are seen in general body condition, redox balance, redox enzyme activities, associated oxidative damage of red blood cells and complement function. We discovered the concentration of bat fur DBT correlated with local marine sediment TBT concentrations. However, we did not find a correlation between the explanatory factors, bat fur DBT and marine sediment TBT concentrations, and several physiological and physical response variables apart from complement activity. Higher DBT concentrations resulted in weaker complement activity and thus a weaker immune response. Although the observed physiological effects in the present study were not strongly correlated to butyltin concentrations in fur or sediment, the result is unique for natural populations so far and raises interesting questions for future ecotoxicological studies.

Seasonal Variation in the Regulation of Redox State and Some Biotransformation Enzyme Activities in the Barn Swallow (Hirundo rustica L.)

Little is known of the normal seasonal variation in redox state and biotransformation activities in birds. In long-distance migratory birds, in particular, seasonal changes could be expected to occur because of the demands of migration and reproduction. In this study, we measured several redox parameters in the barn swallow (Hirundo rustica L.) during the annual cycle. We captured the wintering barn swallows before spring migration in South Africa, and we captured the barn swallows that arrived in spring, bred in summer, and migrated in autumn in Finland. The redox status and biotransformation activities of barn swallows varied seasonally. Wintering birds in South Africa had high biotransformation activities and appeared to experience oxidative stress, whereas in spring and summer, they showed relatively low redox (superoxide dismutase [SOD], catalase [CAT], and glutathione reductase [GR]) and biotransformation enzyme activities. Autumn birds had very low biotransformation enzyme activities and low indication of oxidative stress but high activity of some redox enzymes (GR and glucose 6-phosphate dehydrogenase [G6PDH]). High activities of some redox enzymes (SOD, GR, and G6PDH) seem to be related to migration, whereas low activities of some redox enzymes (SOD, CAT, and GR) may be associated with breeding. Barn swallows in South Africa may experience pollution-related oxidative stress, which may hamper interpretation of normal seasonal variation in redox parameters.

Plasma carotenoid levels are not directly related to heavy metal exposure or reproductive success in three insectivorous passerines

The authors measured plasma carotenoid levels in three insectivorous bird species, the pied flycatcher (Ficedula hypoleuca), the blue tit (Cyanistes caeruleus), and the great tit (Parus major) in metal-polluted and unpolluted sites around a copper smelter. Their aim was to determine whether there was interspecific, age-related, or yearly variation in carotenoid levels and their responses to the ambient pollution level. The three bird species showed qualitatively and quantitatively similar carotenoid profiles, with lutein being always the predominant plasma carotenoid. Ficedula hypoleuca nestlings showed higher plasma carotenoid levels in the unpolluted than in the polluted area, whereas no significant differences were found in F. hypoleuca females or in nestlings or females of the Parids. Yearly comparison of plasma concentrations (only for P. major nestlings) showed, however, that levels varied nonsynchronously between polluted and unpolluted sites among the breeding seasons, with the yearly variation being higher in the polluted area. The survival of nestlings did not depend on their plasma carotenoid concentrations. The two Parid species showed lower nestling survival at sites with high metal exposure levels. The authors conclude that plasma carotenoid levels showed a marked spatial, temporal, and age-related variation, but were not directly related to heavy metal exposure or the reproductive success of birds.

The effects of the painkiller diclofenac and hypoxia on gene transcription and antioxidant system in the gills of three-spined stickleback

Aquatic organisms face multiple stressors in natural ecosystems. More and more often painkillers are detected in surface waters since their prescription has increased worldwide within the last years. Here we examined the effects of the non-steroidal anti-inflammatory drug (NSAID) diclofenac and hypoxia on three-spined sticklebacks (Gasterosteus aculeatus). We exposed sticklebacks to an environmentally relevant concentration of diclofenac (1μg/L) for 14days, to 24h of hypoxia (2.0mg O2/L), and a combination of both. Hypoxia and diclofenac both can be associated with oxidative stress in fish, but it is unclear whether they would act synergistically. Expression analysis of genes related to antioxidant response, hypoxia response, and chemical metabolism in gills showed that diclofenac alone had little effect, while the combination of hypoxia and diclofenac affected transcript levels most, indicating synergistic effects of these stressors. Of the antioxidant enzymes, only superoxide dismutase activity remained unchanged by treatments, while glutathione peroxidase (GPx) was the most affected antioxidant response on both the transcript and activity levels. Our results suggest that diclofenac may lead to suppressed catalase (CAT) activity but increased GPx activity, probably as compensatory mechanism to remove increasing H2O2 in the gills, and that this response is not affected by hypoxia. The activities of lactate dehydrogenase, CAT, and GPx also showed temporal variability during treatments, which can be attributable to tissue-specific circadian rhythms. Our study shows how responses to NSAIDs and hypoxia can interact in fish, suggesting that getting more insight into temporal variation and about the different levels of regulation of environmental responses is necessary in future studies.

Hypoxia and the pharmaceutical diclofenac influence the circadian responses of three-spined stickleback.

Pollution with low concentrations of pharmaceuticals, especially when combined with low-oxygen conditions (hypoxia), is a threat to aquatic ecosystems worldwide. The non-steroidal anti-inflammatory drug diclofenac is commonly detected in wastewater effluents, and has potential to accumulate in the bile of fish. Diclofenac has been shown to activate aryl hydrocarbon receptor (AHR), which induces transcription in the metabolic enzyme cytochrome P450 1a (cyp1a). Previously, crosstalk has been shown to occur between AHR and hypoxia inducible factor 1 (HIF-1). In addition, both of these transcription factors interact with the proteins regulating circadian (24-h) rhythms in vertebrates. Yet little is known about the significance of these interactions during simultaneous exposure to chemicals and hypoxia in fish in vivo. We exposed wild-caught three-spined sticklebacks (Gasterosteus aculeatus) to diclofenac (1 μg/L, 14 days), hypoxia (2.0 mg/L, up to 24h) and the combination of both. We then analyzed markers of chemical biotransformation (EROD activity, cyp1a and ahr mRNA levels), glycolysis (lactate dehydrogenase (LDH) enzyme activity, ldh and enolase 1a mRNA levels), and the transcription of core circadian clock genes clock and period 1 in liver tissue. Samples were taken at three time points during the light period in order to address disturbances in the circadian variation of metabolic processes. The results show that mRNA levels and LDH activity tended to be lowest before the dark period, but this pattern was disturbed by hypoxia and diclofenac. Diclofenac and hypoxia co-exposure induced EROD activity more strongly than diclofenac exposure alone, while cyp1a mRNA level was increased also by hypoxia and diclofenac alone. LDH activity and mRNA expression showed a clear time-dependent response during hypoxia, which is consistent with the previously suggested decreased accumulation of HIF-1 during the dark period. Furthermore, LDH activity and transcription was disturbed by diclofenac, indicating important effects of environmental pollutants in disturbing natural acclimation. This study demonstrates the need for more studies to understand the potential disturbances in endogenous rhythms caused by environmental pollution in natural populations.

Interspecific variation in redox status regulation and immune defence in five bat species: the role of ectoparasites.

Harmful reactive oxygen species (ROS) produced during metabolism and immune responses are neutralized in part by a powerful enzymatic antioxidant system. Inter-species variability in the baseline activity of antioxidant enzymes may be explained by a variety of life history traits. For instance, ectoparasites can elicit repeated immune responses, thus increasing the production of reactive oxygen species. The bat species studied so far have been acknowledged to have effective antioxidant defences. However, interspecific comparisons within the clade do not exist. The present study compares the antioxidant defence and immune function activities in five northern boreal bat species relative to their ectoparasite prevalence and intensity (wing mites and louse flies) to reveal inter-species differences. Antioxidant enzyme and immune defense activities, which differ between species, are positively associated, with total ectoparasite (mites and bat flies) frequencies, total ROS, and protein carbonylation in Daubenton’s bats, but enzyme activities are also independently influenced by sampling date with activities increasing towards the autumn. Antioxidant activities are also positively associated with total reactive oxygen species and oxidative damage (protein carbonylation) in the Daubenton’s bat. Our results suggest that antioxidant activities are associated with ecological factors such as parasite load and season, and we consider it likely that these may partly explain the observed interspecific variation.

Antioxidative defense and oxidative stress in ringed seals (Pusa hispida) from differently polluted areas.

High contaminant levels detected in Baltic seals have been associated with various health effects. In this study several parameters related to antioxidative defense and oxidative stress (concentrations of reduced and oxidised glutathione, lipid hydroperoxide and vitamin E, activities of glutathione reductase, peroxidase and S-transferase, glucose-6-phosphate dehydrogenase, catalase, and superoxidedismutase) were measured in the livers of ringed seals from the Baltic Sea and from a less contaminated reference area, Svalbard, Norway. Seals were caught during two different time periods 1996-1997 and 2002-2007, which represent different levels of contamination. No signs of oxidative damage were found in the Baltic seals. However, glutathione metabolism was enhanced in the ringed seals from the Baltic Sea compared to the seals from Svalbard. The adaptation to dive where repetitive ischemia/reperfusion occurs naturally may contribute to the resistance of oxidative stress and to the capacity to increase enzymatic antioxidant defense in phocid seals. This could explain the similarities in oxidative stress levels despite the differences in antioxidant responses between the ringed seals from the Baltic Sea and Svalbard.