Rostock, Germany
Rostock, Germany

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Rebl A.,Leibniz Institute for Farm Animal Biology | Verleih M.,Leibniz Institute for Farm Animal Biology | Kobis J.M.,Leibniz Institute for Farm Animal Biology | Kuhn C.,Institute For Fischerei | And 3 more authors.
Marine Biotechnology | Year: 2013

Thermal stress can pose a major challenge to salmonid fish. A 4x44K oligonucleotide microarray approach was used to screen for genetically determined variations of a temperature stress response during acclimation in fish gills, a highly specialized and complex organ responsible for gas and electrolyte exchange as well as excretion. The comparison addressed transcriptional changes in the local breeding strain BORN and imported (TCO) rainbow trout after graded 2-week acclimation to 8 and 23 °C. Besides well-characterized mediators of thermoregulation such as genes encoding cold-inducible RNA-binding protein and heat shock proteins, the present microarray study suggests several new candidate genes commonly regulated in gills of the two trout lines. Having identified the differential expression of thermoregulated genes as duplicated paralogues, they were subsequently validated in a gill cell model. Moreover, the comparison of transcriptome profiles provides evidence for distinctively employed expression patterns. The induction of genes encoding factors of the early innate immunity in BORN trout upon warming contrasts with the increased expression of adaptive immune genes in import trout. Cold acclimation induced genes assigned to the functional categories "cell death" and "ion channel activity" in import trout, but repressed "lipid metabolism." This manuscript provides an overview of the genes of the multifunctional gills in rainbow trout that are mandated after temperature change, suggesting links between the different temperature-dependent pathways and gene networks. © 2013 Springer Science+Business Media New York.


Kobis J.M.,Leibniz Institute for Farm Animal Biology | Rebl A.,Leibniz Institute for Farm Animal Biology | Kuhn C.,Institute For Fischerei | Korytar T.,Friedrich Loeffler Institute FLI | And 2 more authors.
Fish and Shellfish Immunology | Year: 2015

The complement system is one of the most ancient and most essential innate immune cascades throughout the animal kingdom. Survival of aquatic animals, such as rainbow trout, depends on this early inducible, efficient immune cascade. Despite increasing research on genes coding for complement components in bony fish, some complement-related genes are still unknown in salmonid fish. In the present study, we characterize the genes encoding complement factor D ( CFD), CD93 molecule ( CD93), and C-type lectin domain family 4, member M ( CLEC4M) from rainbow trout ( Oncorhynchus mykiss). Subsequently, we performed comprehensive and comparative expression analyses of 36 complement genes including CFD, CD93, and CLEC4M and further putative complement-associated genes to obtain general information about the functional gene interaction within the complement pathway in fish. These quantification analyses were conducted in liver, spleen and gills of healthy fish of two rainbow trout strains, selected for survival (strain BORN) and growth (Import strain), respectively. The present expression study clearly confirms for rainbow trout that liver represents the primary site of complement expression. Spleen and gills also express most complement genes, although the mean transcript levels were generally lower than in liver. The transcription data suggest a contribution of spleen and gills to complement activity. The comparison of the two rainbow trout strains revealed a generally similar complement gene expression. However, a significantly lower expression of numerous genes especially in spleen seems characteristic for the BORN strain. This suggests a strain-specific complement pathway regulation under the selected rearing conditions. © 2014 Elsevier Ltd.


Korytar T.,Friedrich Loeffler Institute | Jaros J.,Friedrich Loeffler Institute | Verleih M.,Leibniz Institute for Farm Animal Biology | Rebl A.,Leibniz Institute for Farm Animal Biology | And 4 more authors.
Fish and Shellfish Immunology | Year: 2013

The peritoneal cavity has been extensively used as a laboratory model of inflammation in many species, including the teleost fish. Although, the peritoneal cavity of rainbow trout (Oncorhynchus mykiss) was previously shown to contain a resident population of leukocytes, closer information about their exact composition and their functional response to pathogens is still missing. In the presented work, flow cytometric analysis using monoclonal antibodies was performed to characterize this cell population and evaluate its traffic during the first 72h after antigenic stimulation and infection with Aeromonas salmonicida. Obtained results indicate that the unstimulated peritoneal cavity represents rather a lymphoid niche, dominated by the IgM+ B cells. Expectedly, the composition changed rapidly after stimulation, which resulted in two complete changes of dominant cell type within first 72h post injection. While the first stage of inflammation was dominated by myeloid cells, lymphocytes predominated at the later time points, with IgM+ B cells representing more than two thirds of all cells. Later, the infection experiment elucidated the peritoneal infection and identified the key differences to the antigenic stimulation. Additionally, the data indicate that the resolution of the inflammation depends more on the bacterial clearance by myeloid cells than on regulation by lymphocytes. Taken together, obtained results represent the first complete description of the immune reaction protecting the peritoneal cavity of the fish and shed some light on the conservation of these processes during the evolution. © 2013 Elsevier Ltd.


Rebl A.,Leibniz Institute for Farm Animal Biology | Korytar T.,Institute For Immunologie | Kobis J.M.,Leibniz Institute for Farm Animal Biology | Verleih M.,Leibniz Institute for Farm Animal Biology | And 5 more authors.
Marine Biotechnology | Year: 2014

The fish gills represent a crucial organ for the communication with the aquatic environment. Transcriptional changes in gills of two hatchery rainbow trout strains in response to injection with the potent pathogen Aeromonas salmonicida were detected by global gene expression profiling using a 4×44K oligonucleotide microarray. Emphasis was placed on "day 3 postinfection" representing a decisive time point for the resolution of inflammation. The comparison of features and pathways differentially regulated in branchial tissues revealed that the local breeding strain BORN and imported American rainbow trout apply common and specific immune strategies. In gills of infected BORN trout, we observed a dynamic regulation of genes controlling NF-κB pathways and the induction of factors promoting the development of myeloid cells, whereas an increased expression of lysozyme and immunoglobulin genes was obvious in gills of infected import trout. In order to prove the relevance of the array-predicted candidates as well as well-known immune genes for gill immunity, a subsequent in vitro experiment was conducted. Altogether, we uncovered dynamic but moderate changes in the expression of a broad range of immune-relevant features implying the gill's involvement in pathogen defense strategies. © 2013 Springer Science+Business Media New York.


Verleih M.,Leibniz Institute for Farm Animal Biology | Rebl A.,Leibniz Institute for Farm Animal Biology | Kollner B.,Friedrich Loeffler Institute FLI | Korytar T.,Friedrich Loeffler Institute FLI | And 4 more authors.
Fish Physiology and Biochemistry | Year: 2010

The proline-rich protein 13 (PRR13) is reported to be a key regulator of the resistance to cytostatica by decreasing the copy number of the proapoptotic gene thrombospondin-1. We isolated and characterized the complete PRR13 gene sequence of rainbow trout (Oncorhynchus mykiss). The gene comprises four exons and three introns, the latter of comparatively short lengths (100-811 bp). The full-length PRR13 cDNA consists of 1,101 nucleotides, including an open reading frame of 563 bp, which is predicted to encode a 187 amino acid protein with a molecular mass of 18. 8 kDa. A continuous stretch of ten serine residues at the C-terminus is highly conserved and characteristic for vertebrate PRR13, but not for other known proline-rich proteins. Phylogenetic analyses suggest a clear separation of teleostean PRR13 proteins and those from mammalian and reptilian species. Comparison of the tissue-specific PRR13 mRNA abundance in two strains of the rainbow trout coastal form (TCO Steelhead II-WA vs. BORN Steelhead II-Germany) revealed an increased expression in the BORN trout in nearly all examined tissues. The major expression differences were detected in gill (2. 29-fold) and in liver tissue (2. 16-fold). Hence, the increased PRR13 expression in BORN trout might cause improved protection from natural cytostatica and therefore support our assumption that PRR13 is a candidate gene possibly involved in the varying ability of the two rainbow trout strains to handle environmental stress under local conditions of the Southern Baltic. © 2010 Springer Science+Business Media B.V.


Masilko J.,University of South Bohemia | Zajic T.,University of South Bohemia | Hlavac D.,University of South Bohemia | Sampels S.,University of South Bohemia | And 2 more authors.
Aquaculture International | Year: 2016

The objective of this study on common carp (Cyprinus carpio) was to obtain and predict the first cross-validated data of the fat content on market size carps using a non-invasive or non-destructive method in situ. The carps (1680 ± 388 g; n = 136) used were from a semi-intensive system and were on a different diet (cereal, pelleted and extruded diet). For the evaluation of the fat content, a Fish Fatmeter FM 692 from Distell.com. (FFM) and a manual measurement of back fat height using a digital calliper were used. For the prediction model, the following basic body measurements (variables) were used: total body length, body length, body height, the width of the body, and the circuit of the body. The body weight, weight of intestines, weight of gonads, weight of hepatopancreas, and fillet yield (%) were measured, and the Fulton coefficient was calculated. The study was focussed on evaluating the applicability of these methods and the accuracy of the obtained result, respectively. Results showed that all the rapid methods had a strong correlation. Multiple regression models with forward selection of variables were used throughout. The final prediction model between predicted and observed values for the fat content for FFM and calliper being adjusted index of determination is shown here (Radj 2= 0.88; 5 variables and Radj 2= 0.91; 7 variables), respectively. © 2016 Springer International Publishing Switzerland


Verleih M.,Leibniz Institute for Farm Animal Biology | Rebl A.,Leibniz Institute for Farm Animal Biology | Kollner B.,Friedrich Loeffler Institute FLI | Korytar T.,Friedrich Loeffler Institute FLI | And 4 more authors.
Gene | Year: 2013

The iron-sulfur cluster protein ISCU is a scaffold protein tasked with the building and mediation of iron-sulfur [Fe-S]-clusters. These are crucial for [Fe-S]-enzymes, which are involved in essential biological cell processes like metabolism or ion transport. Analysis of ISCU in rainbow trout (Oncorhynchus mykiss) and maraena whitefish (Coregonus maraena) revealed the existence of two gene variants in each of the two salmonids. This study presents the characterization of the duplicated ISCU cDNA sequences in both species as well as the comparative functional analysis of the genes in healthy and affected fish of two rainbow trout strains differing in trait robustness under regional aquaculture conditions. Coding sequences of trout ISCUA and ISCUB genes are spanning over five exons. Open reading frames (ORF) of trout (ISCUA: 495. bp, ISCUB: 498. bp) and whitefish (ISCUA and ISCUB: 495. bp) genes encode for evolutionary highly conserved proteins and share 72% sequence similarity with human ISCU.Transcriptome analyses comparing healthy fish of the local rainbow trout strain BORN and the import strain TCO revealed strain-specific expression patterns for ISCU. Expression analyses by quantitative RT-PCR indicated remarkable differences between the transcript level of the gene variants ISCUA and ISCUB. Moderate temperature challenge (8. °C and 23. °C) suggests a generally higher transcript level of the two gene variants at 8. °C in the liver, spleen, and gill of both strains. However, no remarkable differences between the strains occurred in the temperature-dependent ISCU gene expression profiles. The experimental infection with Aeromonas salmonicida resulted in a different ISCU gene expression in the gill and trunk kidney of both strains after two weeks, suggesting a specific role of the scaffold gene in rainbow trout strain BORN, regarding the recovery after infection. Although results partially reflect the expected strain- and tissue-specific ISCUA and ISCUB regulation in rainbow trout, the data do not support the assumed association of ISCU with the trait robustness. © 2012 Elsevier B.V.


Kobis J.M.,Leibniz Institute for Farm Animal Biology | Rebl A.,Leibniz Institute for Farm Animal Biology | Kuhn C.,Institute For Fischerei | Goldammer T.,Leibniz Institute for Farm Animal Biology
Molecular Biology Reports | Year: 2013

The spleen plays a crucial role in innate and adaptive immunity in bony fish. Consequently, this organ is well suited to assess the immune competence of an organism and hence provides useful information for comparison and classification of production traits of food fish, such as robustness and susceptibility. To gain information about differences in the basal splenic transcriptome activity, healthy rainbow trout of the commercially important strain TCO and the local selection strain BORN have been compared in a holistic expression analysis using the GRASP 16K cDNA microarray. Nearly all differentially expressed genes (n = 807) in the spleen were on a lower expression level (n = 802) in BORN trout compared to the TCO strain. Global gene ontology analysis revealed that most genes are involved in fundamental biological processes like cellular growth, vesicular trafficking and energy metabolism. Surprisingly, only 7 % of splenic differentially expressed genes are associated with functions of the immune system like TLR signaling, acute phase response and complement system. MARCH3 is one lower expressed gene of interest in BORN trout. This gene, coding for an E3 ubiquitin ligase, is involved in three metabolic functions: immune system, vesicular trafficking and ubiquitination. Since MARCH genes are furthermore differently regulated in the two strains after viral infection and assumed to be potentially active in regulation of immune receptors in fish, MARCH3 was chosen for a closer structural analysis. In concert with the data interpretation of the achieved comparative transcriptome analysis for the involved rainbow trout strains, we provide the full mRNA sequence of trout MARCH3 and its hypothetical protein structure. © 2012 Springer Science+Business Media Dordrecht.


PubMed | Institute For Fischerei and Leibniz Institute for Farm Animal Biology
Type: | Journal: SpringerPlus | Year: 2014

Creatine plays an important role in the cell as an energy buffer. As the energy system is a basic element of the organism it may possibly contribute to differences between rainbow trout strains selected for the traits growth and robustness, respectively. The cDNA sequences of creatine-related genes encoding glycine amidinotransferase (GATM), guanidinoacetate N-methyltransferase (GAMT), creatine kinase muscle-type (CKM) and creatine transporter 1 (CT1, encoded by gene solute carrier family 6, member 8 (SLC6A8)) were characterized in rainbow trout. Transcripts of the respective genes were quantified in kidney, liver, brain and skeletal muscle in both trout strains that had been acclimated to different temperatures. Several differences between the compared trout strains were found as well as between temperatures indicating that the energy system may contribute to differences between both strains. In addition to that, the expression data showed clear differences between the creatine system in rainbow trout and mammals, as the spatial distribution of the enzyme-encoding gene expression was clearly different from the patterns described for mammals. In rainbow trout, creatine synthesis seems to take place to a big extent in the skeletal muscle.


Verleih M.,Leibniz Institute for Farm Animal Biology | Rebl A.,Leibniz Institute for Farm Animal Biology | Kollner B.,Friedrich Loeffler Institute FLI | Korytar T.,Friedrich Loeffler Institute FLI | And 3 more authors.
Molecular Biology Reports | Year: 2012

The Ca2+-binding protein regucalcin (RGN) is crucial for the regulation of Ca2+ ion homeostasis and signal transduction of cells. It is involved in the regulation of Ca2+-dependent protein kinases and Ca2+ pump enzymes in cell membranes. Comparative transcriptome analysis in healthy fish of two aquacultured rainbow trout (Oncorhynchus mykiss) lines (BORN, TCO) varying in susceptibility to environmental stress identified significant differences in the expression of the RGN gene. Therefore, we firstly determined the full genomic DNA and cDNA sequence of RGN gene from rainbow trout and comparatively investigated the complete cDNA sequence in another salmonid fish dedicated for local aquaculture, the maraena whitefish (Coregonus marena). The sequence coding region translates for proteins of 298 and 299 amino acids (aa), respectively, indicating a high conservation of RGN proteins (95.7% aa identity) between the two related salmonids. In the second place, we generated RGN gene expression profiles after pathogen (Aeromonas salmonicidae subsp. salmonicida) and temperature (8 and 23°C) challenge in the two rainbow trout lines using salmon microarrays and quantitative RT-PCR. The profiles not only verified initially detected gene expression differences, they also display a tissue specific gene expression in dependence from the stressor and time. The differences in gene expression support our assumption that RGN might play a role in recovery of rainbow trout after environmental stress. © Springer Science+Business Media B.V. 2011.

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