Project description:In the present work, we evaluated the effects of membrane-initiated cortisol actions in vivo in the proteome of rainbow trout (Oncorhynchus mykiss) skeletal muscle. Quantitative iTRAQ analyses were performed to examine proteomic changes in rainbow trout stimulated with physiological concentrations of cortisol and cortisol-BSA. A total of 873 proteins were identified, among which 38 proteins were commonly and differentially expressed under both conditions. Functional clustering analysis revealed an upregulation of proteins associated with mitochondria, metal-binding and secreted proteins.
Project description:The rainbow trout (Oncorhynchus mykiss) is one of the most important aquaculture species worlwide. In this study, transcriptional profiling of skin by oligonucleotide microarray was applied to rainbow trout individuals infected with A. salmonicida, to identified enriched genes involved in pathogen response.
Project description:Proteomics represents a powerful tool for the analysis of fish spermatozoa, since these cells are transcriptionally inactive. The aim of the present study was to generate an inventory of the most prominent rainbow trout sperm proteins with the use of one-dimensional electrophoresis prefractionation combined with performance liquid chromatography electrospray ionization tandem mass spectrometry. This study provides the first in-depth analysis of the rainbow trout sperm proteome, with a total of 204 identified proteins. We found that rainbow trout spermatozoa are equipped with functionally diverse proteins related to energetic metabolism, signal transduction, protein turnover, transport, cytoskeleton, oxidative injures and stress and reproduction. The availability of a catalogue of rainbow trout sperm proteins provides a crucial tool for the understanding of fundamental molecular processes in fish spermatozoa for ongoing research in the development of novel markers of sperm quality and for the optimization of short- and long-term sperm preservation procedures.
Project description:The objective of this study was to identify and quantify proteomic profiles of spleen of rainbow trout Oncorhynchus mykiss. Specific pathogen free rainbow trout (mean length 15 ± 1 cm) were maintained in recirculating de-chlorinated water at 19±1 °C. Prior to the experiment, fish were distributed between 9 aquaria, 18 fish per aquarium. The test groups were infected by immersion of Yersinia ruckeri strains: CSF007-82 (biotype 1) and 7959-11 (biotype 2). The control group was immersed similar with sterile broth medium. There were 3 aquaria per each group (CSF007-82-infected, 7959-11-infected and control). Nine fish from infected and control fish groups were anaesthetized with MS-222 at 3, 9 and 28 days post exposure and sampled aseptically. Each spleen was washed three times with sterile phosphate-buffered saline containing a cocktail of mammalian protease inhibitors. Spleen samples were snap-frozen in liquid nitrogen and stored at –80 °C.
Project description:Background: Rainbow trout (Oncorhynchus mykiss) is a salmonid species with a complex life-history. Wild populations are naturally divided into freshwater residents and sea-run migrants. Migrants undergo an energy-demanding adaptation for life in seawater, known as smoltification, while freshwater residents display these changes in an attenuated magnitude and rate. Despite this, in seawater rainbow trout farming all animals are transferred to seawater. Under these circumstances, weeks after seawater transfer, a significant portion of the fish die (around 10%) or experience growth stunting (GS; around 10%), which represents an important profitability and welfare issue. The underlying causes leading to GS in seawater-transferred rainbow trout remain unknown. In this study, we aimed at characterising the GS phenotype in seawater-transferred rainbow trout using untargeted and targeted approaches. To this end, the liver proteome (LC-MS/MS) and lipidome (LC-MS) of GS and fast-growing phenotypes were profiled to identify molecules and processes that are characteristic of the GS phenotype. Moreover, the transcription, abundance or activity of key proteins and hormones related to osmoregulation (Gill Na+, K+–ATPase activity), growth (plasma IGF-I, and liver igf1, igfbp1b, ghr1 and ctsl) and stress (plasma cortisol) were measured using targeted approaches. Results: No differences in Gill Na+, K+–ATPase activity and plasma cortisol were detected between the two groups. However, a significant downregulation in plasma IGF-I and liver igf1 transcription pointed at this growth factor as an important pathomechanism for GS. Changes in the liver proteome revealed reactive-oxygen-species-mediated endoplasmic reticulum stress as a key mechanism underlying the GS phenotype. From the lipidomic analysis, key observations include a reduction in triacylglycerols and elevated amounts of cardiolipins, a characteristic lipid class associated with oxidative stress, in GS phenotype. Conclusion: While the triggers to the activation of endoplasmic reticulum stress are still unknown, data from this study point towards either an unresolved infection or a nutritional deficiency as underlying drivers of this phenotype.
Project description:The objective of this study was to identify and quantify proteomic profiles of head kidney of rainbow trout Oncorhynchus mykiss. Specific pathogen free rainbow trout (mean length 15 ± 1 cm) were maintained in recirculating de-chlorinated water at 19±1 °C. Prior to the experiment, fish were distributed between 9 aquaria, 18 fish per aquarium. The test groups were infected by immersion of Yersinia ruckeri strains: CSF007-82 (biotype 1) and 7959-11 (biotype 2). The control group was immersed similar with sterile broth medium. There were 3 aquaria per each group (CSF007-82-infected, 7959-11-infected and control). Nine fish from infected and control fish groups were anaesthetized with MS-222 at 3, 9 and 28 days post exposure and sampled aseptically. Each head kidney was washed three times with sterile phosphate-buffered saline containing a cocktail of mammalian protease inhibitors. Head kidney samples were snap-frozen in liquid nitrogen and stored at –80 °C.
Project description:Stocking density is considered as a key factor determining the productivity of fish aquaculture systems. The transcriptomic response to crowding stress is, however, still poorly investigated. We aimed at the identification of potential biomarker genes via microarray analyses to get insight into molecular pathways modulated through density-induced stress in farmed rainbow trout Oncorhynchus mykiss. Transcriptome profiling in liver, kidney, and gills was complemented with behaviarol observation and analysis of classical plasma parameters. Individuals of two trout strains were exposed for eight days to definite stocking densities, 1 kg/m³ (low density); 10 kg/m³ (moderate); 18 kg/m³ (elevated); and 35 kg/m³ (high). Whereas stocking density had no significant effect on cortisol levels, plasma glucose levels were elevated in trout kept at high density. Pathway enrichment analyses confirmed the upregulation of HIF1a signaling in liver contributing to glucose homeostasis during stress conditions, while mTOR and PI3K/AKT signaling pathways were downregulated. Further perturbed hepatic pathways were involved in protein ubiquitination and the biosynthesis of cholesterol, retinol and glutathione. Three stocking density conditions were investigated: an uncrowded âmoderateâ density (MD: 10 kg trout/m³) , an elevated density (ED: 18 kg/m³ ), and high density (HD: 35 kg/m³). The experiment was performed twice with two strains of Steelhead rainbow trout (Troutlodge and Born trout), randomly assigned to identical glass tanks with MD (30 and 34 individuals), ED (60 and 64 individuals), and HD (120 and 140 individuals). Trout were sampled 8 d after experimental onset.
Project description:In the present study the combination of protein fractionation by one-dimensional gel electrophoresis and high performance liquid chromatography electrospray ionization tandem mass spectrometry was used to characterize the rainbow trout seminal plasma proteome. Our results led to the creation of a catalogue of rainbow trout seminal plasma proteins and significantly contributed to the current knowledge regarding the protein composition of fish seminal plasma. The major proteins of rainbow trout seminal plasma, such as transferrin, apolipoproteins, complement C3, serum albumin, hemopexin-like protein, alpha-1-antiproteinase-like and precerebellin-like protein were recognized as acute phase proteins (proteins which plasma concentration changes in response to inflammation). This study provides the basis for further functional studies of fish seminal plasma proteins, as well as for the identification of novel biomarkers for sperm quality.
Project description:We have constructed a rainbow trout high-density oligonucleotide microarray by using all the available tentative consensus (TC) sequences from the Rainbow Trout Gene Index database (The Computational Biology and Functional Genomics Lab., Dana Farber Cancer Institute and Harvard School of Public Health). The Rainbow Trout Gene Index integrates research data from all available international rainbow trout genomic research projects. The newly designed microarray incorporates 37,394 unique transcript-specific oligonucleotide probes, 60-mer long each. The microarray was printed according to our design by Agilent Technologies using the 4 X 44-design format and contains 1417 Agilent control spots. The performance of the new microarray platform was evaluated by analyzing gene expression associated with the rainbow trout vitellogenesis-induced muscle atrophy. These chips can be ordered from Agilent using design number 016320. This microarray is anticipated to open new avenues of research that will aid in the development of novel strategies to enhance growth efficiency and quality in salmonid species. Keywords: Development of an oligo-array for rainbow trout
Project description:The objective of this study was to identify and quantify proteomic profiles of intestine of rainbow trout (Oncorhynchus mykiss). Specific pathogen free rainbow trout (mean length 15 ± 1 cm) were maintained in recirculating de-chlorinated water at 19±1 °C. Prior to the experiment, fish were distributed between aquaria. The test groups were infected by immersion of Yersinia ruckeri CSF007-82 (biotype 1) and 7959-11 (biotype 2) strains. The control group was immersed similar with sterile broth medium. Fish were anaesthetized and sampled aseptically at different time points. Each intestine was washed three times with sterile phosphate-buffered saline containing a cocktail of mammalian protease inhibitors. Intestinal mucosa was scraped with a sterile large scalpel blade. Intestinal samples were snap-frozen in liquid nitrogen and stored at –80 °C.