IgT, a primitive immunoglobulin class specialized in mucosal immunity.

Teleost fish are the most primitive bony vertebrates that contain immunoglobulins. In contrast to mammals and birds, these species are devoid of immunoglobulin A (IgA) or a functional equivalent. This observation suggests that specialization of immunoglobulin isotypes into mucosal and systemic responses took place during tetrapod evolution. Challenging that paradigm, here we show that IgT, an immunoglobulin isotype of unknown function, acts like a mucosal antibody. We detected responses of rainbow trout IgT to an intestinal parasite only in the gut, whereas IgM responses were confined to the serum. IgT coated most intestinal bacteria. As IgT and IgA are phylogenetically distant immunoglobulins, their specialization into mucosal responses probably occurred independently by a process of convergent evolution.

Rapid Identification of Rainbow Trout Adulteration in Atlantic Salmon by Raman Spectroscopy Combined with Machine Learning.

This study intends to evaluate the utilization potential of the combined Raman spectroscopy and machine learning approach to quickly identify the rainbow trout adulteration in Atlantic salmon. The adulterated samples contained various concentrations (0-100% w/w at 10% intervals) of rainbow trout mixed into Atlantic salmon. Spectral preprocessing methods, such as first derivative, second derivative, multiple scattering correction (MSC), and standard normal variate, were employed. Unsupervised algorithms, such as recursive feature elimination, genetic algorithm (GA), and simulated annealing, and supervised K-means clustering (KM) algorithm were used for selecting important spectral bands to reduce the spectral complexity and improve the model stability. Finally, the performances of various machine learning models, including linear regression, nonlinear regression, regression tree, and rule-based models, were verified and compared. The results denoted that the developed GA-KM-Cubist machine learning model achieved satisfactory results based on MSC preprocessing. The determination coefficient (R2) and root mean square error of prediction sets (RMSEP) in the test sets were 0.87 and 10.93, respectively. These results indicate that Raman spectroscopy can be used as an effective Atlantic salmon adulteration identification method; further, the developed model can be used for quantitatively analyzing the rainbow trout adulteration in Atlantic salmon.

Molecular characterization and expression analysis of four fish-specific CC chemokine receptors CCR4La, CCR4Lc1, CCR4Lc2 and CCR11 in rainbow trout (Oncorhynchus mykiss).

The chemokine and chemokine receptor networks regulate leukocyte trafficking, inflammation, immune cell differentiation, cancer and other biological processes. Comparative immunological studies have revealed that both chemokines and their receptors have expanded greatly in a species/lineage specific way. Of the 10 human CC chemokine receptors (CCR1-10) that bind CC chemokines, orthologues only to CCR6, 7, 9 and 10 are present in teleost fish. In this study, four fish-specific CCRs, termed as CCR4La, CCR4Lc1, CCR4Lc2 and CCR11, with a close link to human CCR1-5 and 8, in terms of amino acid homology and syntenic conservation, have been identified and characterized in rainbow trout (Oncorhynchus mykiss). These CCRs were found to possess the conserved features of the G protein-linked receptor family, including an extracellular N-terminal, seven TM domains, three extracellular loops and three intracellular loops, and a cytoplasmic carboxyl tail with multiple potential serine/threonine phosphorylation sites. Four cysteine residues known to be involved in forming two disulfide bonds are present in the extracellular domains and a DRY motif is present in the second intracellular loop. Signaling mediated by these receptors might be regulated by N-glycosylation, tyrosine sulfation, S-palmitoylation, a PDZ ligand motif and di-leucine motifs. Studies of intron/exon structure revealed distinct fish-specific CCR gene organization in different fish species/lineages that might contribute to the diversification of the chemokine ligand-receptor networks in different fish lineages. Fish-specific trout CCRs are highly expressed in immune tissues/organs, such as thymus, spleen, head kidney and gills. Their expression can be induced by the pro-inflammatory cytokines, IL-1ß, IL-6 and IFNγ, by the pathogen associated molecular patterns, PolyIC and peptidoglycan, and by bacterial infection. These data suggest that fish-specific CCRs are likely to have an important role in immune regulation in fish.

Life-history diversity and its importance to population stability and persistence of a migratory fish: steelhead in two large North American watersheds.

Life-history strategies can buffer individuals and populations from environmental variability. For instance, it is possible that asynchronous dynamics among different life histories can stabilize populations through portfolio effects. Here, we examine life-history diversity and its importance to stability for an iconic migratory fish species. In particular, we examined steelhead (Oncorhynchus mykiss), an anadromous and iteroparous salmonid, in two large, relatively pristine, watersheds, the Skeena and Nass, in north-western British Columbia, Canada. We synthesized life-history information derived from scales collected from adult steelhead (N = 7227) in these watersheds across a decade. These migratory fishes expressed 36 different manifestations of the anadromous life-history strategy, with 16 different combinations of freshwater and marine ages, 7·6% of fish performing multiple spawning migrations, and up to a maximum of four spawning migrations per lifetime. Furthermore, in the Nass watershed, various life histories were differently prevalent through time - three different life histories were the most prevalent in a given year, and no life history ever represented more than 45% of the population. These asynchronous dynamics among life histories decreased the variability of numerical abundance and biomass of the aggregated population so that it was > 20% more stable than the stability of the weighted average of specific life histories: evidence of a substantial portfolio effect. Year of ocean entry was a key driver of dynamics; the median correlation coefficient of abundance of life histories that entered the ocean the same year was 2·5 times higher than the median pairwise coefficient of life histories that entered the ocean at different times. Simulations illustrated how different elements of life-history diversity contribute to stability and persistence of populations. This study provides evidence that life-history diversity can dampen fluctuations in population abundances and biomass via portfolio effects. Conserving genetic integrity and habitat diversity in these and other large watersheds can enable a diversity of life histories that increases population and biomass stability in the face of environmental variability.

How much does inbreeding contribute to the reduced fitness of hatchery-born steelhead (Oncorhynchus mykiss) in the wild?

Many declining populations are supplemented with captive-born individuals that are released directly into the wild. Because captive-born individuals can have lower fitness in the wild than their wild-born counterparts, a comprehensive understanding of the mechanisms responsible for the reduced fitness of these individuals is required for appropriate conservation and management decisions. Inbreeding among captive-born individuals is one plausible mechanism because captive breeding programs frequently use small numbers of breeders to create large numbers of siblings that are subsequently released together into the wild. We tested this hypothesis in a supplementation program for steelhead (Oncorhynchus mykiss) from the Hood River, Oregon, for which first-generation hatchery fish were demonstrated to have lower fitness in the wild than their wild-born counterparts. To determine the contribution of inbreeding to this fitness decline, we first assigned 11 run-years of hatchery steelhead (3005 fish) back to their broodstock parents (462 fish) using 8 polymorphic microsatellite loci. By combining pedigree analyses with species-specific estimates of genetic load, we found that inbreeding could at most account for a 1-4% reduction in the fitness of hatchery fish relative to wild fish. Thus, inbreeding alone cannot adequately explain the 15% average fitness decline observed in first-generation hatchery fish from this population.

Molecular characterization and expression analysis of the putative interleukin 6 receptor (IL-6Rα and glycoprotein-130) in rainbow trout (Oncorhynchus mykiss): salmonid IL-6Rα possesses a polymorphic N-terminal Ig domain with variable numbers of two repeats.

Interleukin (IL)-6, the founding member of IL-6 family cytokines, plays non-redundant roles in hematopoiesis and acute phase responses. IL-6 signals via a specific private IL-6Rα and a common beta chain gp130. In this study, we have cloned both the IL-6Rα and gp130 in rainbow trout. The trout gp130 cDNA encodes 906 aa and is similar in size, extracellular domain structure (D1-D6) and presence of intracellular motifs important for signal transduction to tetrapod gp130s. The trout IL-6Rα cDNA encodes for 834 aa and is larger compared to tetrapod IL-6Rαs, as are other fish IL-6Rα molecules due to a large D1 domain. However, the cytokine-binding domain is well conserved across vertebrates, with four conserved cysteine residues in the N-terminal FNIII domain and a WSXWS motif in the C-terminal FNIII domain. Furthermore, a phylogenetic tree analysis confirmed that the reported fish IL-6Rα and gp130 molecules are orthologues to their tetrapod counterparts. The extra large D1 domain of the salmonid IL-6Rα molecules results partially from the insertions of two repetitive sequences of [TS]-[TF]-VSTTT-[ND]-TTSNG and TTVS-[AT]-IKD-[DG]-S-[KD]-N-[GR], respectively. Furthermore the numbers of repetitions of the two motifs were variable in different individuals and cell lines, and even in the same fish allelic polymorphism exists. Trout IL-6Rα was expressed at higher levels than gp130 in a number of tissues examined and the expression of both IL-6Rα and gp130 could be modulated by LPS and Poly I:C in the cell lines studied. The expression patterns of the receptors suggest that high level expression of IL-6Rα is critical for IL-6 responsiveness.

Rapid evolution of osmoregulatory function by modification of gene transcription in steelhead trout.

Populations experiencing sudden environmental change must be capable of rapidly evolving to survive. Here we explore changes in gene transcription as a mechanism for rapid adaptation at four osmoregulatory genes (CFTR I, NaK ATPase1αa, NaK ATPase1αb and GHRII) in anadromous steelhead trout versus a derived land-locked population after 14 generations. Transcription was measured before and after a 24-h saltwater challenge in pure and reciprocal hybrid offspring of fish from both populations reared in a common environment for two generations. Significant differences between the landlocked and migratory populations were observed, particularly in fresh water at the NaK ATPase1αa and GHRII genes, indicating rapid evolutionary change, possibly associated with reduced energy expenditure in the landlocked lake system. Phenotypic divergence analysis (Q (ST)) shows that the observed transcriptional differences deviate from neutral expectations. Some reciprocal crosses exhibited anomalous transcription consistent with sex-linked epistatic or genetic imprinting effects. Our results highlight unpredictable phenotypic outcomes of hybridization among locally adapted populations and the need to exercise caution when interbreeding populations for conservation purposes.

[Analysis of microsatellite variation in the rainbow trout (Parasalmo (Oncorhynchus) mykiss) from Kamchatka].

Genetic variation of Kamchatka rainbow trout Parasalmo (O.) mykiss was examined using 10 microsatellite DNA loci, and phylogeographic comparison with other representatives of the species across the distribution range was performed. It was demonstrated that Kamchatka populations differed from other geographic groups of rainbow trout in a number of microsatellite loci. These populations also displayed distinct clustering and were characterized by lower genetic diversity. Analysis of a set of 26 different microsatellite loci (personal and literature data) demonstrated that most of the populations within the Kamchatka region were separated from one another, characterized by marked geographic differentiation, and affiliation to certain river basins. In Kamchatka rainbow trout, with high degree of probability, three geographic clusters (northwestern, southwestern, and eastern) were identified. In general, analysis of microsatellite DNA supported the data on low genetic diversity of the Kamchatka group Parasalmo (O.) mykiss, based on the variation estimates for a number of genes of nuclear and mitochondrial DNA, and allozyme loci.

CD38 Defines a Subset of B Cells in Rainbow Trout Kidney With High IgM Secreting Capacities.

CD38 is a multifunctional molecule that functions both as a transmembrane signaling receptor and as an ectoenzyme with important roles in cell adhesion, calcium regulation and signal transduction. Within the B cell linage, CD38 is expressed in diverse murine B cell subsets, with highest levels in innate B cell subpopulations such as marginal zone (MZ) B cells or B1 cells. In humans, however, CD38 is transiently expressed on early lymphocyte precursors, is lost on mature B cells and is consistently expressed on terminally differentiated plasma cells. In the present work, we have identified two homologues of mammalian CD38 in rainbow trout (Oncorhynchus mykiss), designating them as CD38A and CD38B. Although constitutively transcribed throughout different tissues in homeostasis, both CD38A and CD38B mRNA levels were significantly up-regulated in head kidney (HK) in response to a viral infection. In this organ, after the generation of a specific monoclonal antibody (mAb) against CD38A, the presence of CD38A+ populations among IgM+ B cells and IgM- leukocytes was investigated by flow cytometry. Interestingly, the percentage of IgM+CD38A+ B cells increased in response to an in vitro stimulation with inactivated Aeromonas salmonicida. Finally, we demonstrated that HK IgM+CD38A+ B cells had an increased IgM secreting capacity than that of cells lacking CD38A on the cell surface, also showing increased transcription levels of genes associated with B cell differentiation. This study strongly suggests a role for CD38 on the B cell differentiation process in teleosts, and provides us with novel tools to discern between B cell subsets in these species.

Identification, characterization and genetic mapping of TLR1 loci in rainbow trout (Oncorhynchus mykiss).

Induction of innate immune pathways is critical for early anti-microbial defense but there is limited understanding of how teleosts recognize microbial molecules and activate these pathways. In mammals, Toll-like receptors (TLR) 1 and 2 form a heterodimer involved in recognizing peptidoglycans and lipoproteins of microbial origin. Herein, we identify and describe the rainbow trout (Oncorhynchus mykiss) TLR1 gene ortholog and its mRNA expression. Two TLR1 loci were identified from a rainbow trout bacterial artificial chromosome (BAC) library using DNA sequencing and genetic linkage analyses. Full length cDNA clone and direct sequencing of four BACs revealed an intact omTLR1 open reading frame (ORF) located on chromosome 14 and a second locus on chromosome 25 that contains a TLR1 pseudogene. The duplicated trout loci exhibit conserved synteny with other fish genomes that extends beyond the TLR1 gene sequences. The omTLR1 gene includes a single large coding exon similar to all other described TLR1 genes, but unlike other teleosts it also has a 5' UTR exon and intron preceding the large coding exon. The omTLR1 ORF is predicted to encode an 808 amino-acid protein with 69% similarity to the Fugu TLR1 and a conserved pattern of predicted leucine-rich repeats (LRR). Phylogenetic analysis grouped omTLR1 with other fish TLR1 genes on a separate branch from the avian TLR1 and mammalian TLR1, 6 and 10. omTLR1 expression levels in rainbow trout anterior kidney leukocytes were not affected by the human TLR2/6 and TLR2/1 agonists diacylated lipoprotein (Pam(2)CSK(4)) and triacylated lipoprotein (Pam(3)CSK(4)). However, due to the lack of TLR6 and 10 genes in teleost genomes and up-regulation of TLR1 mRNA in response to LPS and bacterial infection in other fish species we hypothesize an important role for omTLR1 in anti-microbial immunity. Therefore, the identification of a TLR2 ortholog in rainbow trout and the development of assays to measure ligand binding and downstream signaling are critical for future elucidation of omTLR1 functions.

Identification of suppressor of cytokine signalling (SOCS) 6, 7, 9 and CISH in rainbow trout Oncorhynchus mykiss and analysis of their expression in relation to other known trout SOCS.

Four new members of the SOCS family of molecules in rainbow trout (Oncorhynchus mykiss), CISH and SOCS6, 7 and 9, are described for the first time in this species. The genes had a wide tissue distribution in trout, and were detected in gills, skin, muscle, liver, spleen, head kidney, intestine and brain, with brain having the highest expression levels. Stimulation of a rainbow trout leucocyte cell line, RTS-11, (mononuclear/macrophage-like cells) with LPS or Poly I:C had no effect on the expression of these genes, although in both cases the previously identified SOCS1-3 genes were up-regulated. Similarly, stimulation of RTS-11 or RTG-2 (fibroblasts) cells with the trout recombinant cytokines IFN-gamma or IL-1beta had no effect on CISH or SOCS6, 7 and 9 expression. However, PMA stimulation did impact on SOCS6 and SOCS9 expression, and LPS stimulation of primary cultures or bacterial infection (Yersinia ruckeri) increased significantly CISH expression (as well as SOCS1 and SOCS2 or SOCS3 respectively). It is apparent that the type II SOCS genes (CISH, SOCS1-3) are particularly relevant to immune regulation in fish, although the intriguing expansion of the SOCS4/5 subgroup in fish requires further investigation as to their role and functional divergence.

Comparative Analysis of the Transcriptome and Distribution of Putative SNPs in Two Rainbow Trout (Oncorhynchus mykiss) Breeding Strains by Using Next-Generation Sequencing.

Selective breeding can significantly improve the establishment of sustainable and profitable aquaculture fish farming. For rainbow trout (Oncorhynchus mykiss), one of the main aquaculture coldwater species in Europe, a variety of selected hatchery strains are commercially available. In this study, we investigated the genetic variation between the local Born strain, selected for survival, and the commercially available Silver Steelhead strain, selected for growth. We sequenced the transcriptome of six tissues (gills, head kidney, heart, liver, spleen, and white muscle) from eight healthy individuals per strain, using RNA-seq technology to identify strain-specific gene-expression patterns and single nucleotide polymorphisms (SNPs). In total, 1760 annotated genes were differentially expressed across all tissues. Pathway analysis assigned them to different gene networks. We also identified a set of SNPs, which are heterozygous for one of the two breeding strains: 1229 of which represent polymorphisms over all tissues and individuals. Our data indicate a strong genetic differentiation between Born and Silver Steelhead trout, despite the relatively short time of evolutionary separation of the two breeding strains. The results most likely reflect their specifically adapted genotypes and might contribute to the understanding of differences regarding their robustness toward high stress and pathogenic challenge described in former studies.

Single nucleotide polymorphism discovery in rainbow trout by deep sequencing of a reduced representation library.

BACKGROUND: To enhance capabilities for genomic analyses in rainbow trout, such as genomic selection, a large suite of polymorphic markers that are amenable to high-throughput genotyping protocols must be identified. Expressed Sequence Tags (ESTs) have been used for single nucleotide polymorphism (SNP) discovery in salmonids. In those strategies, the salmonid semi-tetraploid genomes often led to assemblies of paralogous sequences and therefore resulted in a high rate of false positive SNP identification. Sequencing genomic DNA using primers identified from ESTs proved to be an effective but time consuming methodology of SNP identification in rainbow trout, therefore not suitable for high throughput SNP discovery. In this study, we employed a high-throughput strategy that used pyrosequencing technology to generate data from a reduced representation library constructed with genomic DNA pooled from 96 unrelated rainbow trout that represent the National Center for Cool and Cold Water Aquaculture (NCCCWA) broodstock population. RESULTS: The reduced representation library consisted of 440 bp fragments resulting from complete digestion with the restriction enzyme HaeIII; sequencing produced 2,000,000 reads providing an average 6 fold coverage of the estimated 150,000 unique genomic restriction fragments (300,000 fragment ends). Three independent data analyses identified 22,022 to 47,128 putative SNPs on 13,140 to 24,627 independent contigs. A set of 384 putative SNPs, randomly selected from the sets produced by the three analyses were genotyped on individual fish to determine the validation rate of putative SNPs among analyses, distinguish apparent SNPs that actually represent paralogous loci in the tetraploid genome, examine Mendelian segregation, and place the validated SNPs on the rainbow trout linkage map. Approximately 48% (183) of the putative SNPs were validated; 167 markers were successfully incorporated into the rainbow trout linkage map. In addition, 2% of the sequences from the validated markers were associated with rainbow trout transcripts. CONCLUSION: The use of reduced representation libraries and pyrosequencing technology proved to be an effective strategy for the discovery of a high number of putative SNPs in rainbow trout; however, modifications to the technique to decrease the false discovery rate resulting from the evolutionary recent genome duplication would be desirable.

Characterization and expression of NADPH oxidase in LPS-, poly(I:C)- and zymosan-stimulated trout (Oncorhynchus mykiss W.) macrophages.

In vertebrates, the generation of superoxide reactive oxygen species (ROS) via activation of the Nox/Duox family of NADPH oxidases is a prototypical feature of the pathogen-induced defensive responses of activated professional phagocytes. To understand the role of the rainbow trout (Oncorhynchus mykiss) Phox oxidase from a phylogenetic and functional perspective we describe the cloning, sequencing and expression analysis of multiple NADPH components in cultured macrophages. Phylogenetic analyses support the notion of the emergence of Phox-related components before the diversification of basal euteleosts and add to the limited collection of teleost NADPH oxidases. Expression studies using lipopolysaccharide, polyinosine-polycytidylic acid and zymosan to mimic the onset of inflammatory responses in trout macrophages suggest differences in regulation of the NADPH complex throughout the maturation/differentiation period of culture and between different treatments.

Different origins of paralogues of salmonid TNR1 and TNFR2: Characterisation and expression analysis of four TNF receptor genes in rainbow trout Oncorhynchus mykiss.

Mammalian TNFR1 and TNFR2 bind TNFα and TNFß, and provide key communication signals to a variety of cell types during development and immune responses that are crucial for cell survival, proliferation and apoptosis. In teleost fish TNFß is absent but TNFα has been expanded by the third whole genome duplication (3R WGD) and again by a 4R WGD in some lineages, leading to the four TNFα paralogues known in salmonids. Two paralogues for each of TNFR1 and TNFR2 have been cloned in rainbow trout in this study and are present in other salmonid genomes. Whilst the TNFR2 paralogues were generated via the 4R salmonid WGD, the TNFR1 paralogues arose from a local en bloc duplication. Functional diversification of TNFR paralogues was evidenced by differential gene expression and modulation, upstream ATGs affecting translation, ATTTA motifs in the 3'-UTR regulating mRNA stability, and post-translational modification by N-glycosylation. Trout TNFR are highly expressed in immune tissues/organs, and other tissues, in a gene- and tissue-specific manner. Furthermore, their expression is differentially modulated by PAMPs and cytokines in a cell type- and stimulant-specific manner. Such findings suggest an important role of the TNF/TNFR axis in the immune response and other physiological processes in fish.

A novel, functional, and highly divergent sex hormone-binding globulin that may participate in the local control of ovarian functions in salmonids.

A cDNA encoding for a novel rainbow trout SHBG was identified and characterized. Phylogenetic analysis showed that this novel SHBG, named SHBGb, was a highly divergent paralog of the classical SHBG (SHBGa) form previously known in vertebrates including zebrafish, seabass, and rainbow trout. Using all available sequences, no SHBGb-like sequence could be identified in any fish species besides Atlantic salmon. Rainbow trout SHBGa and SHBGb share only 26% sequence identity at the amino acid level and exhibit totally distinct tissue distribution, thus demonstrating a functional shift of SHBGb. Indeed, shbga mRNA was predominantly expressed in liver and spleen but could not be detected in the ovary, whereas shbgb had a predominant ovarian expression but could not be detected in liver. Despite its high divergence, rainbow trout SHBGb expressed in COS-7 cells could bind estradiol and testosterone with high affinity and specificity. Both rainbow trout shbgb mRNA and proteins were localized to the granulosa cells of vitellogenic ovarian follicles, whereas SHBGb immunoreactivity was also found in theca cells. Finally, shbgb ovarian mRNA expression exhibited a significant drop between late vitellogenesis and oocyte maturation at a time when ovarian aromatase (cyp19a) gene expression and estradiol circulating levels exhibited a dramatic decrease. Together, these observations show that SHBGb is a functional and highly divergent SHBG paralog probably arising from a salmonid-specific duplication of the shbg gene.

Genetic characterisation of farmed rainbow trout in Norway: intra- and inter-strain variation reveals potential for identification of escapees.

BACKGROUND: The rainbow trout (Oncorhynchus mykiss) is one of the most important aquaculture species in the world, and Norway is one of the largest producers. The present study was initiated in response to a request from the Norwegian police authority to identify the farm of origin for 35 escaped rainbow trout captured in a fjord. Eleven samples, each consisting of approximately 47 fish, were collected from the three farms operating in the fjord where the escapees were captured. In order to gain a better general understanding of the genetic structure of rainbow trout strains used in Norwegian aquaculture, seven samples (47 fish per sample) were collected from six farms located outside the region where the escapees were captured. All samples, including the escapees, were genotyped with 12 microsatellite loci. RESULTS: All samples displayed considerable genetic variability at all loci (mean number of alleles per locus per sample ranged from 5.4-8.6). Variable degrees of genetic differentiation were observed among the samples, with pair-wise FST values ranging from 0-0.127. Self-assignment tests conducted among the samples collected from farms outside the fjord where the escapees were observed gave an overall correct assignment of 82.5%, demonstrating potential for genetic identification of escapees. In the "real life" assignment of the 35 captured escapees, all were excluded from two of the samples included as controls in the analysis, and 26 were excluded from the third control sample. In contrast, only 1 of the escapees was excluded from the 11 pooled samples collected on the 3 farms operating in the fjord. CONCLUSION: Considerable genetic variation exists within and among rainbow trout strains farmed in Norway. Together with modern statistical methods, this will provide commercial operators with a tool to monitor breeding and fish movements, and management authorities with the ability to identify the source of escapees. The data generated in this study were used by the Norwegian police to initiate an investigation of the company operating the three farms in the fjord where escapees were observed.

Heat shock protein (HSP70) RNA expression differs among rainbow trout (Oncorhynchus mykiss) clonal lines.

Heat shock protein 70 (HSP70, 70 kDa) is the most commonly expressed protein in response to thermal stress. The extent of its expression is associated with differences in environmental temperatures. We investigated the heat shock response in red blood cells collected from one-year-old rainbow trout (Oncorhynchus mykiss). Three different clonal lines of rainbow trout (Arlee, OSU and Whale Rock) were utilized, originating from habitats that likely experienced different thermal profile. The relative expression of HSP70 from blood cells treated at 13 degrees C, 16 degrees C, 18 degrees C, 20 degrees C, 22 degrees C, and 24 degrees C was quantified using real-time PCR. The use of red blood cells allows for the control and replication of HSP70 expression patterns. Relative expression of HSP70 differed significantly among the three clonal lines. The Arlee line had the lowest HSP70 response of the three clonal lines at any temperature; indicating a heritable difference. Maximum expression of HSP70 occurred at 22 degrees C in the OSU line and at 24 degrees C in the Whale Rock line. The discovery of variation in HSP70 expression among the clonal lines indicates that future studies to map the genetic control of HSP70 expression differences are possible.

The Arctic charr (Salvelinus alpinus) genome and transcriptome assembly.

Arctic charr have a circumpolar distribution, persevere under extreme environmental conditions, and reach ages unknown to most other salmonids. The Salvelinus genus is primarily composed of species with genomes that are structured more like the ancestral salmonid genome than most Oncorhynchus and Salmo species of sister genera. It is thought that this aspect of the genome may be important for local adaptation (due to increased recombination) and anadromy (the migration of fish from saltwater to freshwater). In this study, we describe the generation of a new genetic map, the sequencing and assembly of the Arctic charr genome (GenBank accession: GCF_002910315.2) using the newly created genetic map and a previous genetic map, and present several analyses of the Arctic charr genes and genome assembly. The newly generated genetic map consists of 8,574 unique genetic markers and is similar to previous genetic maps with the exception of three major structural differences. The N50, identified BUSCOs, repetitive DNA content, and total size of the Arctic charr assembled genome are all comparable to other assembled salmonid genomes. An analysis to identify orthologous genes revealed that a large number of orthologs could be identified between salmonids and many appear to have highly conserved gene expression profiles between species. Comparing orthologous gene expression profiles may give us a better insight into which genes are more likely to influence species specific phenotypes.

Species Complex: Classification and Conservation in American Environmental History.

How does the classification of biological organisms shape efforts to conserve them? This essay addresses this key question through the scientific, administrative, and legal histories of steelhead and rainbow trout. Members of the diverse salmon family, these two fish have different life histories and physical appearances, but since the 1930s scientists have considered them the same species. Over the past 150 years, however, their histories diverged. Today, rainbow trout are bred by the millions in hatcheries and are among the world's most common and widespread fish, while steelhead are listed as threatened or endangered all along the West Coast of the United States. Their remarkable story shows that conservation is not merely a political struggle over things that exist in nature; it is a perennial competition to prove the existence and define the very nature of those things that are the focus of such struggles. Biological taxonomy and classification are central to these debates, as they are to environmental history and the history of science more generally.