Project description:Titanium dioxide (TiO2) based nanomaterials (NMs) are among the most produced NMs worldwide. When irradiated with light, particularly UV, TiO2 is photoactive, a property that is explored for several purposes. There is an increasingly number of reports on the negative effects of photoactivated TiO2 to non-target organisms. We have here studied the effect of a suite of reference type TiO2 NMs i.e. NM103, NM104, NM105 and compared these to the Bulk) with and without UV radiation to the oligochaete Enchytraeus crypticus. High-throughput gene expression was used to assess the molecular mechanisms, while also anchoring it to known effects at organism level (i.e., reproduction). Results showed that the photoactivity of TiO2 (UV exposed) played a major role in enhancing TiO2 toxicity, activating transcription of oxidative stress, lysosome damage and apoptosis mechanisms. For non-UV activated TiO2, where toxicity at organism level (reproduction) was lower, results showed the potential for long-term effects (i.e., mutagenic and epigenetic). NM specific mechanisms were identified: NM103 affected transcription and translation, NM104_UV negatively affected reproductive system/organs; and NM105_UV activated superoxide anion response. Results provided mechanistic information for UV-related phototoxicity of TiO2 materials and evidences of the potential long-term effects.
Project description:The effect of nanomaterials (NMs) is less understood in light of the implemented and existing methodologies for regular chemicals. To understand the mode of action of NMs is one of the alternatives to improve predictions and environmental risk assessment (ERA). In the present work the high-throughput gene expression tool (4x44K microarray for Enchytraeus crypticus) was used to investigate the mechanisms activated by Ni exposure. Ni nanoparticles (Ni-NPs) were investigated together with Ni-salt (NiNO3). Testing was done based on reproduction effect concentrations (EC20, EC50) using 3 and 7 days exposure periods.
Project description:The testing of NMs under the currently available standard toxicity tests does not cover many of the NMs specificities. One of the current recommended approaches forward lays on understanding the mechanisms of action as these can help predicting long term effects and safe-by-design production. Copper nanomaterials (Cu-NMs) usage has been highly increasing with the concern in terms of exposure, effect and associated risks. In the present study we used the high-throughput gene expression tool developed for Enchytraeus crypticus (44Kx4 Agilent microarray) to study to the effect of exposure to several copper forms. The copper treatments include two NMs (spherical and wires) and two copper-salt treatments (CuNO3 spiked and Cu field historical contamination). Testing was done based on reproduction effect concentrations (EC20, EC50) using 3 and 7 days exposure periods.
Project description:Negatives effects induced by exposure to ultra-violet (UV) radiation are well known. Nevertheless the modes of action of UV radiation are not well understood, in particular in soil invertebrates. In the present work, the effects of two UV doses (mimicking worst case scenarios in earth crust) on gene expression profile of Enchytraeus crypticus (Enchytraeidae, Oligochaeta) were investigated using the high-throughput 4 x 44K microarray developed for the species.
Project description:Zinc (Zn) is known to be relatively toxic to some soil-living invertebrates including the ecologically important enchytraeid worms. To reveal the molecular mechanisms of zinc toxicity we assessed the gene expression profile of Enchytraeus crypticus (Enchytraeidae), exposed to the reproduction effect concentrations EC10 and EC50, over 4 consecutive days, using a high-throughput microarray (species customized). Three main mechanisms of toxicity to Zn were observed: 1) Zn trafficking (upregulation of zinc transporters, a defence response to regulate the cellular zinc level), 2) oxidative stress (variety of defence mechanisms, triggered by Reactive Oxygen Species (ROS) generated by Zn), and 3) effects on the nervous system (possibly the primary lesion explaining the avoidance behaviour and also why enchytraeids are relatively susceptible to Zn). The adverse outcome at the organism level (reproduction EC50) could be predicted based on gene expression (male gonad development, oocyte maturation), with Zn at the EC50 affecting processes related to higher stress levels. The gene expression response was time-specific and reflected the cascade of events taking place over-time. The 1 to 4 days of exposure design was a good strategy to capture the sequence of events towards zinc adverse outcomes in E. crypticus.
Project description:Transcriptome assembly and microarray construction for Enchytraeus crypticus, a model oligochaete to assess stress response mechanisms derived from soil conditions
Project description:Silver nanomaterials (AgNMs) are broadly used in many products and also rate among the most studied nanoscaled materials. Their ecotoxicological impact in soil invertebrates has been covered, mostly using standard testing, where endpoints like survival and reproduction are assessed. The underlying molecular mechanisms have been assessed to a much less extent. Hence, we here assessed differentially expressed proteins (DEPs) and metabolites (DEMs) by high-throughput (HTP) techniques (HPLC-MS/MS with tandem mass tags for proteome analysis, as well as reversed-phase (RP)- or hydrophilic interaction liquid chromatography (HILIC) with mass spectrometric detection for metabolome analysis. The standard soil model Enchytraeus crypticus was exposed to Ag NM300K and soluble AgNO3, at the reproduction EC20 and EC50, in a time series of 0, 7, and 14 days. The impact was clearly larger after 14 days. Ag NM300K caused more upregulated DEPs/DEMs, and more so at the EC20, compared to the EC50, whereas AgNO3 caused a dose response increase of DEPs/DEMs. Similar pathways were activated, although often via opposite regulation (up vs down) of DEPs hence dissimilar mechanisms underlie the apical impact. Affected pathways include e.g. energy metabolism transport proteins, detoxifying enzymes, histidine (e.g. neurotransmission by gamma-aminobutyric acid (GABA)) and lipid metabolism. Uniquely affected by AgNO3 were catalase, malate dehydrogenase and ATP-citrate synthase, and by Ag NM300K were heat shock proteins (HSP70) and ferritin. The gene expression-based data in Adverse Outcome Pathway (AOP) was confirmed and additional key events were added. Evidences support that toxicity of Ag NM increases in longer-term exposure.
Project description:Gene expression can vary with the organisms' life stage. It is known that embryos can be more sensitive to toxicant exposure, as previously demonstrated for Enchytraeus crypticus (Oligochaeta) exposed to cadmium (Cd), known to cause embryotoxicity and hatching delay. It was shown that Ca enters embryos via the L-type Ca channels in the cocoon membrane, this being affected in Cd exposed embryos (Cd-Ca competition is well-known). In the present study, the embryotoxic mechanisms of Cd were studied via high-throughput gene expression for E. crypticus. Cocoons (1e2 days old), instead of the adult organism, were exposed in Cd spiked LUFA 2.2 soil during 1 day. Results showed that Cd affected Ca homeostasis which is implicated in several other molecular processes. Several of the major modulators of Cd toxicity (e.g., impaired gene expression, cell cycle arrest, DNA and mitochondrial damage) were identified in the embryos showing its relevancy as a model in ecotoxicogenomics. The draft Adverse Outcome Pathway was improved. Previously was hypothesized that gene regulation mechanisms were activated to synthesize more Ca channel proteins e this was confirmed here. Further, novel evidences were that, besides the extracellular competition, Cd competes intracellularly which causes a reduction in Ca efflux, and potentiates Cd embryotoxicity.