Synchrotron XRF Analysis Identifies Cerium Accumulation Colocalized with Pharyngeal Deformities in CeO2 NP-Exposed Caenorhabditis elegans.

A combination of synchrotron radiation-based elemental imaging, in vivo redox status analysis, histology, and toxic responses was used to investigate the uptake, biodistribution, and adverse effects of Ce nanoparticles (CeO2 NP; 10 nm; 0.5-34.96 mg Ce L-1) or Ce(NO3)3 (2.3-26 mg Ce L-1) in Caenorhabditis elegans. Elemental mapping of the exposed nematodes revealed Ce uptake in the alimentary canal prior to depuration. Retention of CeO2 NPs was low compared to that of Ce(NO3)3 in depurated individuals. X-ray fluorescence (XRF) mapping showed that Ce translocation was confined to the pharyngeal valve and foregut. Ce(NO3)3 exposure significantly decreased growth, fertility, and reproduction, caused slightly reduced fecundity. XRF mapping and histological analysis revealed severe tissue deformities colocalized with retained Ce surrounding the pharyngeal valve. Both forms of Ce activated the sod-1 antioxidant defense, particularly in the pharynx, whereas no significant effects on the cellular redox balance were identified. The CeO2 NP-induced deformities did not appear to impair the pharyngeal function or feeding ability as growth effects were restricted to Ce(NO3)3 exposure. The results demonstrate the utility of integrated submicron-resolution SR-based XRF elemental mapping of tissue-specific distribution and adverse effect analysis to obtain robust toxicological evaluations of metal-containing contaminants.

Dose-dependent effects of gamma radiation on the early zebrafish development and gene expression.

Ionizing radiation from natural sources or anthropogenic activity has the potential to cause oxidative stress or genetic damage in living organisms, through the ionization and excitation of molecules and the subsequent production of free radicals and reactive oxygen species (ROS). The present work focuses on radiation-induced biological effects using the zebrafish (Danio rerio) vertebrate model. Changes in developmental traits and gene expression in zebrafish were assessed after continuous external gamma irradiation (0.4, 3.9, 15 and 38 mGy/h) with corresponding controls, starting at 2.5 hours post fertilization (hpf) and lasting through embryogenesis and the early larval stage. The lowest dose rate corresponded to recommended benchmarks at which adverse effects are not expected to occur in aquatic ecosystems (2-10 mGy/day). The survival observed at 96 hours post fertilization (hpf) in the 38 mGy/h group was significantly lower, while other groups showed no significant difference compared to controls. The total hatching was significantly lower from controls in the 15 mGy/h group and a delay in hatching onset in the 0.4 mGy/h group was observed. The deformity frequency was significantly increased by prolonged exposure duration at dose rates ≥ 0.4 mGy/h. Molecular responses analyzed by RNA-seq at gastrulation (5.5 hpf transcriptome) indicate that the radiation induced adverse effects occurred during the earliest stages of development. A dose-response relationship was found in the numbers of differentially regulated genes in exposure groups compared to controls at a total dose as low as 1.62 mGy. Ingenuity Pathway Analysis identified retinoic acid receptor activation, apoptosis, and glutathione mediated detoxification signaling as the most affected pathways in the lower dose rate (0.54 mGy/h), while eif2 and mTOR, i.e., involved in the modulation of angiogenesis, were most affected in higher dose rates (5.4 and 10.9 mGy/h). By comparing gene expression data, myc was found to be the most significant upstream regulator, followed by tp53, TNF, hnf4a, TGFb1 and cebpa, while crabp2b and vegfab were identified as most frequent downstream target genes. These genes are associated with various developmental processes. The present findings show that continuous gamma irradiation (≥ 0.54 mGy/h) during early gastrula causes gene expression changes that are linked to developmental defects in zebrafish embryos.

Aging and soil organic matter content affect the fate of silver nanoparticles in soil.

Sewage sludge application on soils represents an important potential source of silver nanoparticles (Ag NPs) to terrestrial ecosystems, and it is thus important to understand the fate of Ag NPs once in contact with soil components. Our aim was to compare the behavior of three different forms of silver, namely silver nitrate, citrate stabilized Ag NPs (5nm) and uncoated Ag NPs (19nm), in two soils with contrasting organic matter content, and to follow changes in binding strength over time. Soil samples were spiked with silver and left to age for 2h, 2 days, 5 weeks or 10 weeks before they were submitted to sequential extraction. The ionic silver solution and the two Ag NP types were radiolabeled so that silver could be quantified by gamma spectrometry by measuring the (110m)Ag tracer in the different sequential extraction fractions. Different patterns of partitioning of silver were observed for the three forms of silver. All types of silver were more mobile in the mineral soil than in the soil rich in organic matter, although the fractionation patterns were very different for the three silver forms in both cases. Over 20% of citrate stabilized Ag NPs was extractible with water in both soils the first two days after spiking (compared to 1-3% for AgNO(3) and uncoated Ag NPs), but the fraction decreased to trace levels thereafter. Regarding the 19nm uncoated Ag NPs, 80% was not extractible at all, but contrary to AgNO(3) and citrate stabilized Ag NPs, the bioaccessible fraction increased over time, and by day 70 was between 8 and 9 times greater than that seen in the other two treatments. This new and unexpected finding demonstrates that some Ag NPs can act as a continuous source of bioaccessible Ag, while AgNO(3) is rapidly immobilized in soil.

Effects of different gamma exposure regimes on reproduction in the earthworm Eisenia fetida (Oligochaeta).

Ecological risk assessment of ionising radiation requires knowledge about the responses of individuals and populations to chronic exposures, including situations when exposure levels change over time. The present study investigated processes such as recovery and the adaptive response with respect to reproduction endpoints in the earthworm Eisenia fetida exposed to (60)Co γ-radiation. Furthermore, a crossed experiment was performed to investigate the influence of F0 parental and F1 embryonic irradiation history on the response of irradiated or non-irradiated F1 offspring. Recovery: The sterility induced by sub-chronic exposure at 17 m Gy/h (accumulated dose: 25 Gy) was temporary, and 8 weeks after irradiation the worms had regained their reproductive capacity (number of viable offspring produced per adult per week). Adaptive response: Adult worms were continuously exposed at a low priming dose rate of 0.14 mGy/h for 12 weeks (accumulated dose: 0.24 Gy), followed by 14 weeks exposure at a challenge dose rate of 11 mGy/h. The results suggest a lack of adaptive response, since there were no significant differences in the effects on reproduction capacity between the primed and the unprimed groups after challenge doses ranging from 7.6 to 27 Gy. Crossed experiment: The effects of exposure at 11 mGy/h for 21 weeks on growth, sexual maturation and reproduction of offspring, derived either from parent worms and cocoons both exposed at 11 mGy/h, or from non-irradiated parents and cocoons (total accumulated dose 44 and 38 Gy, respectively) were compared. There were no significant differences between the two exposed offspring groups for any of the endpoints. The reproduction capacity was very low for both groups compared to the controls, but the reproduction seemed to be maintained at the reduced level, which could indicate acclimatisation or stabilisation. Finally, parental and embryonic exposures at 11 mGy/h did not affect reproduction in the F1 offspring as adults.

Induction and repair of DNA strand breaks and oxidised bases in somatic and spermatogenic cells from the earthworm Eisenia fetida after exposure to ionising radiation.

Methods for analysing oxidised DNA lesions [formamidopyrimidine glycosylase (Fpg)-sensitive sites] in coelomocytes and spermatogenic cells from the earthworm Eisenia fetida using the Fpg-modified comet assay were established. The DNA integrity (SSBs = strand breaks plus alkali labile sites and Fpg-sensitive sites) in cells from E. fetida continuously exposed to (60)Co gamma-radiation (dose rates 0.18-43 mGy/h) during two subsequent generations (F0 and F1) were measured and related to effects on reproduction end points which have already been reported. The data suggest a slight increase of Fpg-sensitive sites in spermatogenic cells from worms exposed at 11 mGy/h in the F0 generation but not in F1, whereas reduced reproduction had been observed at dose rates at or >4 mGy/h in F0 and at 11 mGy/h in F1. Using acute X-rays (41.9 Gy/h), dose-response relationships were established for SSBs in coelomocytes and spermatogenic cells exposed in vitro. In vivo DNA repair was studied by measuring the decrease in damage (SSBs and Fpg-sensitive sites) in coelomocytes and spermatogenic cells isolated from worms at different times (0-6 h) after acute X-ray exposure (4 Gy). SSBs were repaired in coelomocytes following biphasic kinetics, i.e. with a fast and a slow half-life (t(1/2)) of 36 min (95%) and 6.7 h (5%), respectively. Fpg-sensitive sites were repaired at considerably lower rates (t(1/2) = 4-5 h). In spermatogenic cells, SSB repair during the first hour was observed but a half-life could not be estimated. Repair of Fpg-sensitive sites could not be determined. In general, a reduced repair of Fpg-sensitive sites suggests a higher potential for accumulation of oxidised lesions, compared to SSBs, in earthworms exposed to radiation and other environmental contaminants. This is the first study comparing DNA damage with reproduction in earthworms exposed to ionising radiation.

Silver nanoparticle exposure causes apoptotic response in the earthworm Lumbricus terrestris (Oligochaeta).

In terrestrial ecotoxicology there is a serious lack of data for potential hazards posed by engineered nanoparticles (ENPs). This is partly due to complex interactions between ENPs and the soil matrix, but also to the lack of suitable toxicological end points in organisms that are exposed to ENPs in a relevant manner. Earthworms are key organisms in terrestrial ecosystems, but so far only physiological end points of low sensitivity have been used in ecotoxicity studies with ENPs. We exposed the earthworm Lumbricus terrestris to silver nanoparticles and measured their impact on apoptosis in different tissues. Increased apoptotic activity was detected in a range of tissues both at acute and sublethal concentrations (down to 4 mg/kg soil). Comparing exposure in water and soil showed reduced bioavailability in soil reflected in the apoptotic response. Apoptosis appears to be a sensitive end point and potentially a powerful tool for quantifying environmental hazards of ENPs.

Neutron activation of engineered nanoparticles as a tool for tracing their environmental fate and uptake in organisms.

Studies regarding the environmental impact of engineered nanoparticles (ENPs) are hampered by the lack of tools to localize and quantify ENPs in water, sediments, soils, and organisms. Neutron activation of mineral ENPs offers the possibility of labeling ENPs in a way that avoids surface modification and permits both localization and quantification within a matrix or an organism. Time-course experiments in vivo also may be conducted with small organisms to study metabolism and exposure, two aspects currently lacking in ecotoxicological knowledge about ENPs. The present report explains some of the prerequisites and advantages of neutron activation as a tool for studying ENPs in environmental samples and ecologically relevant organisms, and it demonstrates the suitability of neutron activation for Ag, Co/Co3O4, and CeO2 nanoparticles. In a preliminary experiment with the earthworm Eisenia fetida, the dietary uptake and excretion of a Co nanopowder (average particle size, 4 nm; surface area, 59 m2/g) were studied. Cobalt ENPs were taken up to a high extent during 7 d of exposure (concentration ratios of 0.16-0.20 relative to the ENP concentration in horse manure) and were largely retained within the worms for a period of eight weeks, with less than 20% of absorbed ENPs being excreted. Following dissection of the worms, 60Co was detected in spermatogenic cells, cocoons, and blood using scintillation counting and autoradiography. The experimental opportunities that neutron activation of ENPs offer are discussed.