Project description:An European eel-specific microarray platform was developed to identify genes involved in response to pollutants A comparative analysis of gene expression was conducted between European eel Anguilla anguilla individuals from high (Tiber river, Italy) and low pollution (Bolsena lake, Italy) environments. Gene expression profiling was performed using an European eel-specific oligo-DNA microarray of 14,913 probes based on single-colour detection (Cyanine-3 only). Microarrays were scanned with Agilent scanner G2565BA (barcode on the left, DNA on the back surface, scanned through the glass) at a resolution of 5 microns; all slides were scanned twice at two different sensitivity settings (XDRHi 100% and XDRLo 10%); the scanner software created a unique ID for each pair of XDR scans and saved it to both scan image files. Feature Extraction (FE) 9.5 used XDR ID to link the pairs of scans together automatically when extracting data. The signal left after all the FE processing steps have been completed is ProcessedSignal that contains the Multiplicatively Detrended, Background-Subtracted Signal.
Project description:An European eel-specific microarray platform was developed to identify genes involved in response to pollutants. A comparative analysis of gene expression was conducted between European eel Anguilla anguilla individuals from lowly-polluted Wijmeers pond at Uitbergen (Belgium), highly-polluted Hazewinkel pond at Willebroek (Belgium), extremely-polluted Dessel-Schotel canal at the locations of Schotel (Belgium) and low polluted Bolsena lake (Italy) environments.
Project description:An European eel-specific microarray platform was developed to identify genes involved in response to pollutants. A comparative analysis of gene expression was conducted between European eel Anguilla anguilla individuals from lowly-polluted Wijmeers pond at Uitbergen (Belgium), highly-polluted Hazewinkel pond at Willebroek (Belgium), extremely-polluted Dessel-Schotel canal at the locations of Schotel (Belgium) and low polluted Bolsena lake (Italy) environments. A comparative analysis of gene expression was conducted between European eel Anguilla anguilla individuals from lowly-polluted Wijmeers pond at Uitbergen (Belgium), highly-polluted Hazewinkel pond at Willebroek (Belgium), extremely-polluted Dessel-Schotel canal at the locations of Schotel (Belgium) and low polluted Bolsena lake (Italy) environments. Gene expression profiling was performed using an European eel-specific oligo-DNA microarray (GPL15124) of 14,913 probes based on single-colour detection (Cyanine-3 only). Microarrays were scanned with Agilent scanner G2565BA (barcode on the left, DNA on the back surface, scanned through the glass) at a resolution of 5 microns; all slides were scanned twice at two different sensitivity settings (XDRHi 100% and XDRLo 10%); the scanner software created a unique ID for each pair of XDR scans and saved it to both scan image files. Feature Extraction (FE) 9.5 used XDR ID to link the pairs of scans together automatically when extracting data. The signal left after all the FE processing steps have been completed is ProcessedSignal that contains the Multiplicatively Detrended, Background-Subtracted Signal.
Project description:We investigated salinity adaptation during the migration from freshwater to seawater of European eel (Anguilla anguilla) by examining the hypothesis that: The brain is the central organ for the co-ordination of environmental cues (day length, photoperiod, temperature and environmental salinity) with the anatomical and physiological adaptations which accompany pre-migrational morphogenesis and the osmoregulatory plasticity seen in post-migrational, salinity-adapted fish. We have characertised the mRNA expression profiles for the brains of fresh water and sea water adapted silver eel using a highly representative brain cDNA microarray. The array comprises 5760 cDNA clones from A.anguilla ranging from 0.5 -10 kb and an estimated redundancy of > 5 %.
Project description:Ecotoxicogenomic studies face the problem of the lack of gene sequence information available for toxicologically relevant non-model pollution sentinel species. In this sense, next generation sequencing technologies allow obtaining deep de novo sequence information cost-effectively. We have thus employed the platform GS-FLX of Roche technologies, in order to sequence the multitissue transcriptome of a fish species under severe population decline, the European eel (Anguilla anguilla), using normalized cDNA (Evrogen). The European eel could be successfully employed in pollution biomonitoring studies as it is a euryhaline teleost that is quite resistant to chemical exposure. We have completed a one whole plate sequencing-run using the Titanium kit of Roche from which a total of 7.8x105 reads were obtained with a length average of 303.53 bp. Annotated genes, for instance, could be further assessed as biomarkers of exposure to specific chemical compounds in marine, estuarine and river waters. Designed tool will be thus useful, in the study of: a.- the mode of action of chemical compounds in active monitoring studies using caged eels, b.- the physiological processes that are specific to freshwater eels such as the one of sexual development and reproduction, c.- reasons that could explain the disappearance of the species from European waters. In the first context, a caging experiment was performed to measure the putative effects of a paper industry effluent on eel hepatic transcription levels. For this purpose, eels were caged upstream and downstream to the SMURFFIT-KAPPA paper industry in Iurreta (Biscay, Basque Country). Thus, hepatic transcription profiles reflect the stress levels suffered by eels.
Project description:Many enigmas surround different aspects of freshwater eel biology and life cycle. In the same way different hypothesis about why eels are disappearing from European continental waters have been proposed. One such proposal defends that poor fat accumulation of eels, due to pollution in continental waters, might be stopping eels from reproductive migration to the Sargasso Sea. Thus, habitat deterioration could be blamed for the decline in the catchment potential of the European rivers. In this context, and with the aim to study the mode of action of environmentally relevant chemicals in eels, the multi tissue transcriptome was sequenced (454 Titanium Roche) in order to design a high density custom oligonucleotide microarray (eArray, Agilent). To validate this tool a laboratory experiment was carried to analyze the gene trasncrition profiles related to chemical compounds released from pulp and paper mills; 100 μg/L mercury and 150 μg/L β-sitosterol (only Hg data is presented here). 20 yellow European eel (Anguilla anguilla) elvers (11.7±5.35 g) obtained from a local farm (Acuivas SL, Usurbil, Gipuzkoa) were exposed for 3, 6 and 9 days. Pyrosequencing allowed the design and construction of a 60K microarray platform containing 3923 gene signatures identified through BlastN analysis and 7212 sequences annotated through BlastX coupled to Blast2Go analysis. Additional 226 sequences were incorporated from NCBI databases and 3551 from the information available in EeelBase in 2011. Two probes were generated per sequence and they were spotted twice in the array. Hepatic gene expression profiling of the exposed eels indicated that Hg significantly down-regulated (LIMMA, adj. p<0.005) only gene signatures related to selenoprotein W-1 (SeW), something typically described in mammals exposed to methyl-mercury. Increasing the adj. p value to <0.05, 116 genes were significantly regulated (38 down-regulated and 79 up-regulated). Among them, we found additional selenoproteins such as ROS metabolism related genes; glutathione peroxidases (gpx1 & gpx4b) and thioredoxine which were up-regulated. In addition, complement system genes (C3 & C4b) were also up-regulated. Studying enriched Go pathways (p<0.005) and in relation with lipid homeostasis we observed that the following pathways were enriched after exposure to Hg: fatty acids degradation and metabolism of arachidonic acid, linoleic acid, ether lipids, alpha linolenic acid, and glycerophospholipids. In addition, among the top 10 significantly enriched KEGG pathways, p53 signalling, apoptosis, and MAPK signalling were present, suggesting possible effects on cell cycle regulation. In summary, transcriptome pyrosequencing and subsequently designed microarray provided the molecular tools to successfully study the gene transcription profiles of toxic chemical compounds such as Hg in European eel tissues. In addition to study the molecular modes of action of specific chemical compounds, the developed gene expression microarray will be useful in active monitoring of the quality of freshwater environments using caged sentinel eels. This study was funded by Basque Government (SAIOTEK S-PE09UN32; Consolidated Research Groups IT810-13) and UPV/EHU (UFI 11/37). Technical and human support provided by SGIker (UPV/EHU) is acknowledged.