Project description:Changes in soil properties (e.g. pH, organic matter content, granulometry) can influence chemical toxicity to organisms and act alone as stressors. Previous studies on Enchytraeus albidus showed that changes in soil properties caused effects on reproduction and avoidance behavior and also oxidative stress. In addition, results at the transcritptomic level indicated changes in gene expression profile due to soil properties changes. In this study, E. albidus was exposed to modified versions of the artificial standard OECD soil (different pH, OM and clay content) in different exposure times (2, 4 and 8 days). The gene expression profile was characterized using a class comparison statistical analysis. Results indicated that the transcriptional response was time dependent, with different genes being affected at different time points. Results also showed some genes (and biological functions) being affected in a soil specific way.
Project description:The soil oligochaete Enchytraeus albidus is a standard test organism used in biological testing for Environmental Risk Assessment (ERA). Although effects are known at acute and chronic level, through survival, reproduction and avoidance behaviour endpoints, very little is known at the sub-cellular and molecular levels. In this study, the effects of soil properties (clay, organic matter and pH) and of the chemicals copper and phenmedipham were studied on gene expression in E. albidus during exposure periods of 2 days, 4 days and 3 weeks, using DNA microarrays based on a normalised cDNA library for this test species (Amorim et al., 2011) The main objectives of this study were: 1) to assess changes in gene expression of E. albidus over time, and 2) to identify molecular markers for natural and chemical exposures. Results showed a clear influence of exposure time in gene expression. Transcriptional responses to phenmedipham were seen at 2 days while the responses to copper and the different soils were more pronounced at 4 days of exposure. Some genes were differentially expressed in a stress specific manner and, in general, the responses were related with effects at energy metabolism and cell growth level.
Project description:Transcriptional responses in Enchytraeus albidus (Oligochaeta): comparison between cadmium and zinc exposure and linkage to reproduction effects
Project description:Changes in soil properties (e.g. pH, organic matter content, granulometry) can influence chemical toxicity to organisms and act alone as stressors. Previous studies on Enchytraeus albidus showed that changes in soil properties caused effects on reproduction and avoidance behavior and also oxidative stress. In addition, results at the transcritptomic level indicated changes in gene expression profile due to soil properties changes. In this study, E. albidus was exposed to modified versions of the artificial standard OECD soil (different pH, OM and clay content) in different exposure times (2, 4 and 8 days). The gene expression profile was characterized using a class comparison statistical analysis. Results indicated that the transcriptional response was time dependent, with different genes being affected at different time points. Results also showed some genes (and biological functions) being affected in a soil specific way. Fluorescently labelled cDNA, from enchytraeids exposed during 2, 4 and 8 days OECD standard soil (Cy3) and to the different exposure conditions (modified OECD soil) (Cy5), was synthesized for microarray analysis and hybridizations were performed. After scanning (Agilent Microarray Scanner from Agilent Technologies), spots were identified and ratios quantified using Quantarray (Packard Biochip Technologies). Statistical analysis of the microarrays was performed using BRB Array Tools version 3.8.1 Stable Release (http://linus.nci.nih.gov/BRB-ArrayTools.html). After background subtraction, the replicated spots within each array were averaged, and microarrays were normalized using loess method (Smith and Speed, 2003). Statistical class comparison between groups of arrays was performed between each exposure condition and the respective “control” (exposure in OECD standard soil, for each time of exposure) using two-sample T-test and 95% of confidence level for the assessment of differentially expressed genes.
Project description:Due to its antimicrobial activity, silver nanoparticles (Ag-NPs) are among the most used NPs worldwide, yet little information is available regarding their effects, particularly in soil dwelling organisms. Enchytraeids (Oligochaeta) are important members of the soil fauna which actively contribute to the acceleration of organic matter decomposition and nutrient recycling processes. Hence, for hazard and risk assessment it is important to provide toxicity data for these organisms and to understand more in regard to the mode of action of Ag-NPs within organism. To study this we conducted toxicity experiments using the OECD standard guideline, testing Ag-NPs and AgNO3, having assessed survival, reproduction and differential gene expression. Population toxicity responses were assessed showing higher toxicity for the AgNO3. In an attempt to understand the mode of action we performed transcription profiling using the microarray. Gene expression profile of Enchytraeus albidus was analysed after 2 days of exposure to 100 and 200 mg/kg of two silver forms (nanoparticles and salt_silver nitrate) in OECD soil. Three biological replicates per test treatment and control (clean OECD soil) were used.
Project description:Background: Molecular mechanisms of response to pesticides are scarce and information on such responses from soil invertebrates is almost inexistent. Enchytraeus albidus (Oligochaeta) is a standard soil ecotoxicology model species for which effects of many pesticides are known on survival, reproduction and avoidance behaviour. With the recent microarray development additional information can be retrieved on the molecular effects. Methodology/Principal Findings: Experiments were performed to investigate the transcription responses of E. albidus when exposed to three pesticides – dimethoate (insecticide), atrazine (herbicide) and carbendazim (fungicide) – in a range of concentrations that inhibited reproduction by 10%, 20%, 50% and 90% (EC10, EC20, EC50 and EC90, respectively). The goal of this study was to further identify key biological processes affected by each compound and if dose-related. All three pesticides significantly affected biological processes like translation, regulation of the cell cycle or general response to stress. Intracellular signalling and microtubule-based movement were affected by dimethoate and carbendazim whereas atrazine affected lipid and steroid metabolism (also by dimethoate) or carbohydrate metabolism (also by carbendazim). Response to DNA damage/DNA repair was exclusively affected by carbendazim. Conclusions: Changes in gene expression were significantly altered after 2 days of exposure in a dose-related manner. The mechanisms of response were comparable with the ones for mammals, suggesting across species conserved modes of action. The present results indicate the potential of using gene expression in risk assessment and the advantage as early markers.
Project description:The soil oligochaete Enchytraeus albidus is a standard test organism used in biological testing for Environmental Risk Assessment (ERA). Although effects are known at acute and chronic level, through survival, reproduction and avoidance behaviour endpoints, very little is known at the sub-cellular and molecular levels. In this study, the effects of soil properties (clay, organic matter and pH) and of the chemicals copper and phenmedipham were studied on gene expression in E. albidus during exposure periods of 2 days, 4 days and 3 weeks, using DNA microarrays based on a normalised cDNA library for this test species (Amorim et al., 2011) The main objectives of this study were: 1) to assess changes in gene expression of E. albidus over time, and 2) to identify molecular markers for natural and chemical exposures. Results showed a clear influence of exposure time in gene expression. Transcriptional responses to phenmedipham were seen at 2 days while the responses to copper and the different soils were more pronounced at 4 days of exposure. Some genes were differentially expressed in a stress specific manner and, in general, the responses were related with effects at energy metabolism and cell growth level. Fluorescently labelled cDNA, from enchytraeids exposed during 4 days and 3 weeks to control soil (LUFA 2.2) and to the different exposure conditions was synthesized for microarray analysis and hybridizations were performed. After scanning, spots were identified and ratios quantified using the QuantArray (Packard Biochip Technologies). Statistical analysis of the microarrays was performed using limmaGUI package (1.18.0) (Smyth, 2005) in the R (2.8.0) software environment (http://www.R-project.org/). After being submitted to local background subtraction, microarrays were normalized using global loess method. To statistically evaluate the differential gene expression between the different conditions, a gene-per-gene linear model (limma – linear model for microarray analysis) and empirical Bayes methods were applied. The results were then corrected for multiple testing using the Benjamini-Hochberg’s method (adjusted p<0.05 was considered significant) (Benjamini and Hochberg, 1995).