Transgenerational effects of low dose radiation and in combination with secondary stressors
Living organisms can be exposed to ionizing radiation from various radionuclides occurring in the environment naturally and from nuclear accidents, nuclear waste and other anthropogenic activity. There are major gaps in knowledge on multigenerational and transgenerational effects of low dose ionizing radiation in the environment as well as the combined effects of ionizing radiation and other environmental factors (chemical, physical, biological). Therefore, the Centre of Environmental Radioactivity (CERAD), the Centre of Excellence has been established at the Norwegian University of Life Sciences (NMBU) in collaboration with the Norwegian Radiation Protection Authority (NRPA/DSA) and the institutions NMI, NIPH and NIVA and funded by the Norwegian Research Council (2013-2023, project 223268/F50). The goal of CERAD and its advisory board of outstanding international researchers is to generate new scientific data on the harmful effects of radiation. This study will investigate effects / toxicokinetics and toxicodynamics of ionizing radiation exposure in zebrafish during sensitive life stages (gametogenesis and embryogenesis) alone or in combination with other environmental factors. Particular focus will be placed on linking reproduction, immune responses and cancer with the genome and epigenome in four generations of fish. The NMBU VetBio group members, who possess competence in working with zebrafish (completed FELASA) will expose zebrafish to gamma radiation emitted by a Co-60 radiation source (FIGARO). The effects of radiation alone and combined with other environmental stressors (such as e.g. UV radiation, bacterial pathogen, Uranium and plastics) will be studied over four consecutive generations of fish. Results from the experiment will be of national and international significance and will provide more knowledge about how radiation alone or in combination with other stressors affects the health of animals and humans. This will improve the ability for assessing the radiological impact and managing the risk associated with radioactivity in the environment. It will be used 700 zebra fish in the experiments. The experiment is classified as severe. There will be used as few animals as possible in the in the radiation exposure and in the bacterial pathogen challenge experiments.
Etterevaluering
The Norwegian Food Safety Authority must retrospectively assess all projects involving severe procedures.
Begrunnelse for etterevalueringen
The experimental goals were largely reached. Thee reprotoxic, genetic and transgenerational epigenetic effects of radiation exposure during sensitive developmental stages in the zebrafish were analyzed and a multigenerational experiment was performed. Exposures to the bacterial pathogens and environmental stressors in the F1 generation were not performed due to the corona pandemic in 2020 and 2021, a reduced capacity at the NMBU zebrafish laboratory and a severe accident and long-term injury to the project manager.
A total of 700 animals were used, of which 300 were exposed to radiation in F0. Of these 300, 104 fish exposed to the higher dose rates of radiation (4 fish in the 10 mGy/h group and 100 fish in the 40 mGy/h group) did not develop sexual organs or were not able to reproduce, and this group's severity load was termed as "moderate". The rest of the fish used in this experiment were offspring of irradiated fish or controls and were classified as mildly harmed.
Fish health was recorded daily instead of weekly, during the exposure experiment and up to 40 days post fertilization to reduce animal suffering. Video recording could have been included to monitor the fish more closely and reduce the burden on the animals. The transport and movement of animals should be shortened. For exposures to bacterial pathogens, a facility with a flow-through system needs to be available. To further reduce the harm on the animals, fish could be monitored daily during the entire transgenerational experiment. Fish which developed an adverse condition not attributed to radiation exposure were quarantined immediately upon registration, monitored or euthanized according to the SOP of the NMBU zebrafish laboratory if reaching "the point of no return".
Humane endpoints could be improved by checking the water quality in each housing tank instead of the main water tank in the system, to detect any possible variation of water quality from tank to tank. Automatized feeding and elimination of parasites in the zebrafish facility could have also improved fish health and experiments. Iced water and an overdose of tricaine methanesulfonate (MS-222) was considered fast and least painful to fish in this experiment.
A total of 700 animals were used, of which 300 were exposed to radiation in F0. Of these 300, 104 fish exposed to the higher dose rates of radiation (4 fish in the 10 mGy/h group and 100 fish in the 40 mGy/h group) did not develop sexual organs or were not able to reproduce, and this group's severity load was termed as "moderate". The rest of the fish used in this experiment were offspring of irradiated fish or controls and were classified as mildly harmed.
Fish health was recorded daily instead of weekly, during the exposure experiment and up to 40 days post fertilization to reduce animal suffering. Video recording could have been included to monitor the fish more closely and reduce the burden on the animals. The transport and movement of animals should be shortened. For exposures to bacterial pathogens, a facility with a flow-through system needs to be available. To further reduce the harm on the animals, fish could be monitored daily during the entire transgenerational experiment. Fish which developed an adverse condition not attributed to radiation exposure were quarantined immediately upon registration, monitored or euthanized according to the SOP of the NMBU zebrafish laboratory if reaching "the point of no return".
Humane endpoints could be improved by checking the water quality in each housing tank instead of the main water tank in the system, to detect any possible variation of water quality from tank to tank. Automatized feeding and elimination of parasites in the zebrafish facility could have also improved fish health and experiments. Iced water and an overdose of tricaine methanesulfonate (MS-222) was considered fast and least painful to fish in this experiment.