Single and combined toxicity of gamma and ultraviolet B radiation in the aquatic macrophyte Lemna minor

Abstract

Aquatic plants in the natural environment are exposed to background levels of ionizing radiation originating from cosmic radiation and naturally occurring radioactive materials (NORM), as well as technologically enhanced naturally occurring radioactive materials (TENORM) and other anthropogenic sources from medical treatment, nuclear power production, and nuclear accidents. However, the in-depth understanding of how ionizing radiation leads to harmful effects in aquatic plants is limited and the toxicity mechanisms are rarely determined in detail. Ionizing radiation in the natural environment also co-occurs with other environmental factors, such as non-ionizing radiation (e.g. UVB radiation) and affects aquatic plants through combined toxicity. Research on how exposures of ionizing and non-ionizing radiation affect aquatic plants, alone in combination, is thus needed. This PhD study intended to characterize single and combined effects of ionizing radiation and non-ionizing radiation on the aquatic plant Lemna minor using γ-radiation and UVB radiation as prototypical stressors for ionizing and non-ionizing radiation. Three exposure experiments with L. minor were conducted with γ-radiation, UVB radiation, and their combination, based on the standardised toxicity test guideline (OECD TG221). Several biological effects were investigated through different endpoints, including reactive oxygen species (ROS) formation and DNA damage at the molecular level, inhibition of photosynthesis and mitochondrial dysfunction at the subcellular level, oxidative damage at the cellular level, and growth and reproductive inhibition at the individual and population levels. Quantitative real-time reverse transcription polymerase chain reaction (qPCR) was used to evaluate the expression of biomarker genes for key toxic cellular processes. The combined effects of γ-radiation and UVB radiation were assessed using the two-way analysis of variance (2W-ANOVA) to identify interactions between the stressors, whereas a modified independent action (IA) prediction model was used to identify the more exact nature of the combined toxicity (i.e. antagonism, additivity or synergy) at different levels of biological organisation. Correlational analysis was performed to identify potential causality between observations and to propose relevant toxicity pathways. The studies showed that γ-radiation led to dose rate-dependent and target-specific dependent modes of action (MoAs) in L. minor after exposure for 7 days. Low dose rates of γ-radiation (LOEDR: 1 to 14 mGy h-1) induced DNA damage, increased antioxidant and reduced pigment (chlorophyll a, chlorophyll b and carotenoid) levels, whereas higher dose rates (LOEDR: 24 to 47 mGy h-1) led to ROS formation, suppression of photosynthesis, inhibition of oxidative phosphorylation (OXPHOS) and reduction of plant growth. The UVB radiation also caused dose rate-dependent and target-specific responses in L. minor after exposure for 7 days. At low UVB irradiances (LOEI: 0.23 to 0.48 W m-2), UVB radiation induced ROS formation, inhibited photosynthesis, and suppressed plant growth. Higher irradiances (LOEI: 1.2 to 4 W m-2) caused a reduction in photosynthetic pigments (chlorophyll a and chlorophyll b), suppressed OXPHOS, DNA lesions such as cyclobutane pyrimidine dimers (CPDs). Compared to γ-radiation, UVB radiation exhibited different MoAs in L. minor, although they shared some common toxicity pathways. Co-exposures of γ-radiation and UVB radiation induced a plethora of combined effects across different levels of biological organisation, with the induction of oxidative stress and DNA damage considered the main toxicity mechanisms triggered. The combined effects observed ranged from non-interactive (additive) to interactive (antagonism and synergism) and were typically dose rate (irradiance)-dependent and target-specific. Observations of combined effects at the apical levels could to some degree be understood in terms of propagation of effects across different levels of biological organisation. The results from this thesis demonstrate that L. minor is a suitable model species to evaluate the toxicity of γ-radiation, UVB radiation and their combination. The suite of statistical and predictive models used improved characterisation of the combined effects observed. The proposed toxicity pathways from the work are anticipated to facilitate the development of data aggregation and visualisation frameworks such as Adverse Outcome Pathways (AOPs) for detailed assessment of single and joint effects of ionizing and non-ionizing radiation. The obtained knowledge can be integrated into future ecotoxicological studies to improve the understanding and prediction of the hazard of radiation in the environment due to anthropogenic activities.

Akvatiske planter i det naturlige miljøet er utsatt for bakgrunnsnivå av ioniserende stråling som stammer fra kosmisk stråling og naturlig forekommende radioaktive materialer (NORM), samt teknologisk endrede naturlig forekommende radioaktive materialer (TENORM) og andre menneskeskapte kilder fra medisinsk behandling, kjernekraftproduksjon og atomulykker. Den mekanistiske forståelsen av hvordan ioniserende stråling fører til skadelige effekter i akvatiske planter er fremdeles mangelfull og ofte lite anerkjent. Ioniserende stråling eksisterer imidlertid samtidig med andre miljøfaktorer i det naturlige miljøet, for eksempel sammen med ikke-ioniserende stråling som ultrafiolett stråling (UV-A og UV-B). UV stråling kan også påvirke akvatiske planter gjennom kombinert toksisitet. Forskning på hvordan eksponering av ioniserende og ikke-ioniserende stråling påvirker akvatiske planter, enkeltvis og i kombinasjon, er derfor viktige for å bedre kunnskapen om kombinasjonseffekter av ioniserende og ikke-ioniserende stråling. Hensikten med denne Phd-studien er å karakterisere enkelt- og kombinasjonseffektene av ioniserende stråling og ikke-ioniserende stråling i en akvatiske planten Lemna minor ved bruk av γ-stråling og UVB-stråling som prototypiske eksempler på ioniserende og ikke-ioniserende stråling.