Use of fish otoliths as a temporal biomarker of field uranium exposure.

This study aimed to determine uranium (U) pollution over time using otoliths as a marker of fish U contamination. Experiments were performed in field contamination (~20 µg L-1: encaged fish: 15d, 50d and collected wild fish) and in laboratory exposure conditions (20 and 250 µg L-1, 20d). We reported the U seasonal concentrations in field waterborne exposed roach fish (Rutilus rutilus), in organs and otoliths. Otoliths were analyzed by ICPMS and LA-ICP SF MS of the entire growth zone. Concentrations were measured on transects from nucleus to the edge of otoliths to characterize environmental variations of metal accumulation. Results showed a spatial and temporal variation of U contamination in water (from 51 to 9.4 µg L-1 at the surface of the water column), a high and seasonal accumulation in fish organs, mainly the digestive tract (from 1000 to 30,000 ng g-1, fw), the gills (from 1600 to 3200 ng g-1, fw) and the muscle (from 144 to 1054 ng g-1, fw). U was detected throughout the otolith and accumulation varied over the season from 70 to 350 ng g-1, close to the values measured (310 ng g-1) after high exposure levels in laboratory conditions. U in otoliths of encaged fish showed rapid and high U accumulation from 20 to 150 ng g-1. The U accumulation signal was mainly detected on the edge of the otolith, showing two U accumulation peaks, probably correlated to fish age, i.e. 2 years old. Surprisingly, elemental U and Zn signatures followed the same pattern therefore using the same uptake pathways. Laboratory, caging and field experiments indicated that otoliths were able to quickly accumulate U on the surface even for low levels and to store high levels of U. This study is an encouraging first step in using otoliths as a marker of U exposure.

Subcellular fractionation and chemical speciation of uranium to elucidate its fate in gills and hepatopancreas of crayfish Procambarus clarkii.

Knowledge of the organ and subcellular distribution of metals in organisms is fundamental for the understanding of their uptake, storage, elimination and toxicity. Detoxification via MTLP and MRG formation and chelation by some proteins are necessary to better assess the metal toxic fraction in aquatic organisms. This work focused on uranium, natural element mainly used in nuclear industry, and its subcellular fractionation and chemical speciation to elucidate its accumulation pattern in gills and hepatopancreas of crayfish Procambarus clarkii, key organs of uptake and detoxification, respectively. Crayfish waterborne exposure was performed during 4 and 10d at 0, 30, 600 and 4000 µg UL(-1). After tissue dissection, uranium subcellular fractionation was performed by successive ultracentrifugations. SEC-ICP MS was used to study uranium speciation in cytosolic fraction. The uranium subcellular partitioning patterns varied according to the target organ studied and its biological function in the organism. The cytosolic fraction accounted for 13-30% of the total uranium amount in gills and 35-75% in hepatopancreas. The uranium fraction coeluting with MTLPs in gills and hepatopancreas cytosols showed that roughly 55% of uranium remained non-detoxified and thus potentially toxic in the cytosol. Furthermore, the sum of uranium amount in organelle fractions and in the non-detoxified part of cytosol, possibly equivalent to available fraction, accounted for 20% (gills) and 57% (hepatopancreas) of the total uranium. Finally, the SEC-ICP MS analysis provided information on potential competition of U for biomolecules similar than the ones involved in endogenous essential metal (Fe, Cu) chelation.

An invitation to contribute to a strategic research agenda in radioecology.

With intentions of integrating a portion of their respective research efforts into a trans-national programme that will enhance radioecology, eight European organisations recently formed the European Radioecology ALLIANCE (www.er-alliance.org). The ALLIANCE is an Association open to other organisations throughout the world with similar interests in promoting radioecology. The ALLIANCE members recognised that their shared radioecological research could be enhanced by efficiently pooling resources among its partner organizations and prioritising group efforts along common themes of mutual interest. A major step in this prioritisation process was to develop a Strategic Research Agenda (SRA). An EC-funded Network of Excellence in Radioecology, called STAR (Strategy for Allied Radioecology), was formed, in part, to develop the SRA. This document is the first published draft of the SRA. The SRA outlines a suggested prioritisation of research topics in radioecology, with the goal of improving research efficiency and more rapidly advancing the science. It responds to the question: "What topics, if critically addressed over the next 20 years, would significantly advance radioecology?" The three Scientific Challenges presented within the SRA, with their 15 associated research lines, are a strategic vision of what radioecology can achieve in the future. Meeting these challenges will require a directed effort and collaboration with many organisations the world over. Addressing these challenges is important to the advancement of radioecology and in providing scientific knowledge to decision makers. Although the development of the draft SRA has largely been a European effort, the hope is that it will initiate an open dialogue within the international radioecology community and its stakeholders. This is an abbreviated document with the intention of introducing the SRA and inviting contributions from interested stakeholders. Critique and input for improving the SRA are welcomed via a link on the STAR website (www.star-radioecology.org).

Uranium-induced sensory alterations in the zebrafish Danio rerio.

The effect of chronic exposure to uranium ions (UO(2)(2+)) on sensory tissues including the olfactory and lateral line systems was investigated in zebrafish (Danio rerio) using scanning electron microscopy. The aim of this study was to determine whether exposure to uranium damaged sensory tissues in fish. The fish were exposed to uranium at the concentration of 250 µg l(-1) for 10 days followed by a depuration period of 23 days. Measurements of uranium uptake in different fish organs: olfactory rosettes and bulbs, brain, skin, and muscles, were also determined by ICP-AES and ICP-MS during the entire experimental period. The results showed that uranium displayed a strong affinity for sensory structures in direct contact with the surrounding medium, such as the olfactory and lateral line systems distributed on the skin. A decreasing gradient of uranium concentration was found: olfactory rosettes>olfactory bulbs>skin>muscles>brain. At the end of the experiment, uranium was present in non-negligible quantities in sensory tissues. In parallel, fish exposed to uranium showed severe sensory tissue alterations at the level of the olfactory and lateral line systems. In both sensory systems, the gross morphology was altered and the sensory hair cells were significantly damaged very early after the initiation of exposure (from the 3rd day). At the end of the experiment, after 23 days of depuration, the lateral line system still displayed slight tissue alterations, but approximately 80% of the neuromasts in this system had regenerated. In contrast, the olfactory system took more time to recover, as more than half of the olfactory rosettes observed remained destroyed at the end of the experiment. This study showed, for the first time, that uranium is able to damage fish sensory tissues to such an extent that tissue regeneration is delayed.

Induction of reactive oxygen species and algal growth inhibition by tritiated water with or without copper.

Tritium ((3) H) is a radioactive element of ecological concern because of its release into aquatic ecosystems from nuclear power plants. However, the acute and chronic effects of tritiated water (HTO) on aquatic organisms are poorly documented, as are its effects on oxidative stress. In addition, the effects of HTO in combination with other contaminants remain largely unexamined. Herein, we document the effect of HTO on a primary aquatic producer (Chlamydomonas reinhardtii) by measuring growth and oxidative stress using fluorimetric (H(2) DCF-DA) determination of Reactive Oxygen Species (ROS) production. The maximum cell density of the alga (1.65 × 10(6) cells mL(-1) ) was reduced by 23% (1.27 × 10(6) cells mL(-1) ) at the highest exposure tested (59 MBq mL(-1) HTO), whereas cells exposed to 0.9 MBq mL(-1) showed a significantly enhanced maximum cell density of 1.90 × 10(6) cells mL(-1) , an increase of 15%. With regard to oxidative stress, exposure to HTO (0.04, 0.16, and 2.8 MBq mL(-1) ) induced an early dose-dependent peak in ROS production after 14-15 min of exposure, followed by a slow decrease in ROS which stabilized after 60 min. Moreover, this study showed that the presence of HTO may influence the impact of other conventional, nonradioactive contaminants, such as copper, a well known oxidizing trace metal for aquatic organisms. A significant synergic effect of copper and HTO on ROS production was observed. This synergic effect on oxidative stress was shown to be linked to an enhanced copper uptake rate measured in the presence of HTO (> 4 times). We conclude that HTO should be considered as a sensitizer when in a mixture with other contaminants, especially through interactions on the antioxidant system of algae.

Investigating the embryo/larval toxic and genotoxic effects of γ irradiation on zebrafish eggs.

Eggs/larval of freshwater fish (Danio rerio) were exposed to low dose rates of external gamma radiation (from 1 to 1000 mGy d(-1)) over a 20-day period, with the objective of testing the appropriateness of the 10 mGy d(-1) guideline suggested by the IAEA. The present study examines different endpoints, mortality and hatching time and success of embryos as well as the genotoxicity of γ-irradiations (after 48 h). The 20-day embryo-larval bioassay showed an enhanced larval resistance to starvation after chronic exposure to γ irradiation (from low 1 mGy d(-1) to high dose rate 1000 mGy d(-1)) and an acceleration in hatching time. Gamma irradiation led to increased genotoxic damage Ito zebrafish egg (40-50% DNA in tail in Comet assay) from the lowest dose rate (1 mGy d(-1)). Possible mechanisms of γ radiotoxicity and implications for radioprotection are discussed.

Issues and practices in the use of effects data from FREDERICA in the ERICA Integrated Approach.

The ERICA Integrated Approach requires that a risk assessment screening dose rate is defined for the risk characterisation within Tiers 1 and 2. At Tier 3, no numerical screening dose rate is used, and the risk characterisation is driven by methods that can evaluate the possible effects of ionising radiation on reproduction, mortality and morbidity. Species sensitivity distribution has been used to derive the ERICA risk assessment predicted no-effect dose rate (PNEDR). The method used was based on the mathematical processing of data from FRED (FASSET radiation effects database merged with the EPIC database to form FREDERICA) and resulted in a PNEDR of 10 microGy/h. This rate was assumed to ascribe sufficient protection of all ecosystems from detrimental effects on structure and function under chronic exposure. The value was weighed against a number of points of comparison: (i) PNEDR values obtained by application of the safety factor method, (ii) background levels, (iii) dose rates triggering effects on radioactively contaminated sites and (iv) former guidelines from literature reviews. In Tier 3, the effects analysis must be driven by the problem formulation and is thus highly case specific. Instead of specific recommendations on numeric values, guidance on the sorts of methods that may be applied for refined effect analysis is provided and illustrated.

Modelling the propagation of effects of chronic exposure to ionising radiation from individuals to populations.

This study evaluated the potential effect of ionising radiation on population growth using simple population models and parameter values derived from chronic exposure experiments in two invertebrate species with contrasting life-history strategies. In the earthworm Eisenia fetida, models predicted increasing delay in population growth with increasing gamma dose rate (up to 0.6 generation times at 11 mGy h(-1)). Population extinction was predicted at 43 mGy h(-1). In the microcrustacean Daphnia magna, models predicted increasing delay in population growth with increasing alpha dose rate (up to 0.8 generation times at 15.0 mGy h(-1)), only after two successive generations were exposed. The study examined population effects of changes in different individual endpoints (including survival, number of offspring produced and time to first reproduction). Models showed that the two species did not respond equally to equivalent levels of change, the fast growing daphnids being more susceptible to reduction in fecundity or delay in reproduction than the slow growing earthworms. This suggested that susceptibility of a population to ionising radiation cannot be considered independent of the species' life history.

Increased effects of internal alpha irradiation in Daphnia magna after chronic exposure over three successive generations.

A 70-day experiment was performed with Daphnia magna exposed to waterborne Am-241 on a range of concentrations (from 0.4 to 40 Bq ml(-1)) in order to test chronic effects of internal alpha irradiation on respiration, somatic growth and reproduction over three successive generations. Changes in Am-241 concentrations were followed in the water and in daphnid tissues, eggs and cuticles. Corresponding average dose rates of 0.3, 1.5 and 15 mGy h(-1) were estimated. This study confirmed that oxygen consumption increased significantly in the first generation (F0) after 6 days of exposure to a dose rate >or=1.5 mGy h(-1). Consequences were limited to a reduction in body length (5%) and dry mass of females (16%) and eggs (8%) after 23 days of exposure, while mortality and fecundity remained unaffected. New cohorts were started with neonates of broods 1 and 5, to examine potential consequences of the reduced mass of offspring for subsequent exposed generations. Results strongly contrasted with those observed in F0. At the highest dose rate, an early mortality of 38-90% affected juveniles while survivors showed delayed reproduction and reduced fecundity in F1 and F2. At 0.3 and 1.5 mGy h(-1), mortality ranged from 31 to 38% of daphnids depending on dose rate, but was observed only in generation F1 started with neonates of the brood 1. Reproduction was affected through a reduction in the proportion of breeding females, occurring in the first offspring generation at 1.5 mGy h(-1) (to 62% of total daphnids) and in the second generation at 0.3 mGy h(-1) (to 69% of total daphnids). Oxygen consumption remained significantly higher at dose rates >or=0.3 mGy h(-1) than in the control in almost every generation. Body size and mass continued decreasing in relation to dose rate, with a significant reduction in mass ranging from 15% at 0.3 mGy h(-1) to 27% at 15 mGy h(-1) in the second offspring generation.

Effects of chronic internal alpha irradiation on physiology, growth and reproductive success of Daphnia magna.

Daphnids were chronically exposed to waterborne Am-241, an alpha-emitting radionuclide, ranging in concentration from 0.4 to 40 Bq ml(-1). Am-241 amounts were monitored in the medium, daphnid tissues and cuticles. Corresponding average dose rates of 0.02, 0.11 and 0.99 mGy h(-1) were calculated for whole organisms with internal alpha-radiation contributing 99% of total dose rates. Effects of internal alpha irradiation on respiration and ingestion rates, adult, egg and neonate individual dry masses, fecundity and larval resistance to starvation were examined in 23-day experiments. Daphnids showed increased respiratory demand after 23 days at the highest dose rate, suggesting increased metabolic cost of maintenance due to coping with alpha radiological stress. Although no effect was detected on ingestion rates between contaminated and control daphnids, exposure to dose rates of 0.11 mGy h(-1) or higher, resulted in a significant 15% reduction in body mass. Fecundity remained unchanged over the 23-day period, but individual masses of eggs and neonates were significantly smaller compared to the control. This suggested that increased metabolic expenditure in chronically alpha-radiated daphnids came at the expense of their energy investment per offspring. As a consequence, neonates showed significantly reduced resistance to starvation at every dose rate compared to the control. Our observations are discussed in comparison with literature results reported for cadmium, a chemical toxicant which affects feeding activity and strongly reduces individual energy uptake.