miRNAs Predicted to Regulate Host Anti-viral Gene Pathways in IPNV-Challenged Atlantic Salmon Fry Are Affected by Viral Load, and Associated With the Major IPN Resistance QTL Genotypes in Late Infection

Infectious pancreatic necrosis virus (IPNV) infection has been a major problem in salmonid aquaculture. Marker-assisted selection of individuals with resistant genotype at the major IPN quantitative trait locus (IPN-QTL) has significantly reduced mortality in recent years. We have identified host miRNAs that respond to IPNV challenge in salmon fry that were either homozygous resistant (RR) or homozygous susceptible (SS) for the IPN-QTL. Small RNA-sequenced control samples were compared to samples collected at 1, 7, and 20 days post challenge (dpc). This revealed 72 differentially expressed miRNAs (DE miRNAs). Viral load (VL) was lower in RR vs. SS individuals at 7 and 20 dpc. However, analysis of miRNA expression changes revealed no differences between RR vs. SS individuals in controls, at 1 or 7 dpc, while 38 “high viral load responding” miRNAs (HVL-DE miRNAs) were identified at 20 dpc. Most of the HVL-DE miRNAs showed changes that were more pronounced in the high VL SS group than in the low VL RR group when compared to the controls. The absence of differences between QTL groups in controls, 1 and 7 dpc indicates that the QTL genotype does not affect miRNA expression in healthy fish or their first response to viral infections. The miRNA differences at 20 dpc were associated with the QTL genotype and could, possibly, contribute to differences in resistance/susceptibility at the later stage of infection. In silico target gene predictions revealed that 180 immune genes were putative targets, and enrichment analysis indicated that the miRNAs may regulate several major immune system pathways. Among the targets of HVL-DE miRNAs were IRF3, STAT4, NFKB2, MYD88, and IKKA. Interestingly, TNF-alpha paralogs were targeted by different DE miRNAs. Most DE miRNAs were from conserved miRNA families that respond to viral infections in teleost (e.g., miR-21, miR-146, miR-181, miR-192, miR-221, miR-462, miR-731, and miR-8159), while eight were species specific. The miRNAs showed dynamic temporal changes implying they would affect their target genes differently throughout disease progression. This shows that miRNAs are sensitive to VL and disease progression, and may act as fine-tuners of both immediate immune response activation and the later inflammatory processes

Life-long course and molecular characterization of the original Dutch family with epidermolysis bullosa simplex with muscular dystrophy due to a homozygous novel plectin point mutation

Plectin is one of the largest and most versatile cytolinker proteins known. Cloned and sequenced in 1991, it was later shown to have nonsense mutations in recessive epidermolysis bullosa with muscular dystrophy. A dominant mutation in the gene was found to cause epidermolysis bullosa simplex Ogna without muscular dystrophy. Here we report the DNA sequencing of the plectin gene (PLEC1) in a Dutch family originally described in 1972 as having epidermolysis bullosa with muscular dystrophy. The results revealed homozygosity for a new plectin nonsense mutation at position 13187 and its specific 8q24 marker haplotype profile. Western blotting of cultured fibroblasts and immunofluorescence microscopy of skin biopsy confirm that the plectin protein expression is grossly reduced or absent. A summary of the life-long clinical course of the two affected brothers homozygous for the new E1914X mutation is given.</p

Life-long course and molecular characterization of the original Dutch family with epidermolysis bullosa simplex with muscular dystrophy due to a homozygous novel plectin point mutation

Plectin is one of the largest and most versatile cytolinker proteins known. Cloned and sequenced in 1991, it was later shown to have nonsense mutations in recessive epidermolysis bullosa with muscular dystrophy. A dominant mutation in the gene was found to cause epidermolysis bullosa simplex Ogna without muscular dystrophy. Here we report the DNA sequencing of the plectin gene (PLEC1) in a Dutch family originally described in 1972 as having epidermolysis bullosa with muscular dystrophy. The results revealed homozygosity for a new plectin nonsense mutation at position 13187 and its specific 8q24 marker haplotype profile. Western blotting of cultured fibroblasts and immunofluorescence microscopy of skin biopsy confirm that the plectin protein expression is grossly reduced or absent. A summary of the life-long clinical course of the two affected brothers homozygous for the new E1914X mutation is given

Distribution of ancestral proto-Actinopterygian chromosome arms within the genomes of 4R-derivative salmonid fishes (Rainbow trout and Atlantic salmon)

Comparative genomic studies suggest that the modern day assemblage of ray-finned fishes have descended from an ancestral grouping of fishes that possessed 12-13 linkage groups. All jawed vertebrates are postulated to have experienced two whole genome duplications (WGD) in their ancestry (2R duplication). Salmonids have experienced one additional WGD (4R duplication event) compared to most extant teleosts which underwent a further 3R WGD compared to other vertebrates. We describe the organization of the 4R chromosomal segments of the proto-rayfinned fish karyotype in Atlantic salmon and rainbow trout based upon their comparative syntenies with two model species of 3R ray-finned fishes. Results: Evidence is presented for the retention of large whole-arm affinities between the ancestral linkage groups of the ray-finned fishes, and the 50 homeologous chromosomal segments in Atlantic salmon and rainbow trout. In the comparisons between the two salmonid species, there is also evidence for the retention of large whole-arm homeologous affinities that are associated with the retention of duplicated markers. Five of the 7 pairs of chromosomal arm regions expressing the highest level of duplicate gene expression in rainbow trout share homologous synteny to the 5 pairs of homeologs with the greatest duplicate gene expression in Atlantic salmon. These regions are derived from proto-Actinopterygian linkage groups B, C, E, J and K. Conclusion: Two chromosome arms in Danio rerio and Oryzias latipes (descendants of the 3R duplication) can, in most instances be related to at least 4 whole or partial chromosomal arms in the salmonid species. Multiple arm assignments in the two salmonid species do not clearly support a 13 proto-linkage group model, and suggest that a 12 proto-linkage group arrangement (i.e., a separate single chromosome duplication and ancestral fusion/fissions/recombination within the putative G/H/I groupings) may have occurred in the more basal soft-rayed fishes. We also found evidence supporting the model that ancestral linkage group M underwent a single chromosome duplication following the 3R duplication. In the salmonids, the M ancestral linkage groups are localized to 5 whole arm, and 3 partial arm regions (i.e., 6 whole arm regions expected). Thus, 3 distinct ancestral linkage groups are postulated to have existed in the G/H and M lineage chromosomes in the ancestor of the salmonids

A description of the origins, design and performance of the TRAITS-SGP Atlantic salmon Salmo salar L. cDNA microarray

The origins, design, fabrication and performance of an Atlantic salmon microarray are described. The microarray comprises 16 950 Atlantic salmon-derived cDNA features, printed in duplicate and mostly sourced from pre-existing expressed sequence tag (EST) collections [SALGENE and salmon genome project (SGP)] but also supplemented with cDNAs from suppression subtractive hybridization libraries and candidate genes involved in immune response, protein catabolism, lipid metabolism and the parr–smolt transformation. A preliminary analysis of a dietary lipid experiment identified a number of genes known to be involved in lipid metabolism. Significant fold change differences (as low as 1.2x) were apparent from the microarray analysis and were confirmed by quantitative real-time polymerase chain reaction analysis. The study also highlighted the potential for obtaining artefactual expression patterns as a result of cross-hybridization of similar transcripts. Examination of the robustness and sensitivity of the experimental design employed demonstrated the greater importance of biological replication over technical (dye flip) replication for identification of a limited number of key genes in the studied system. The TRAITS (TRanscriptome Analysis of Important Traits of Salmon)–salmon genome project microarray has been proven, in a number of studies, to be a powerful tool for the study of key traits of Atlantic salmon biology. It is now available for use by researchers in the wider scientific community

The major histocompatibility class I locus in Atlantic salmon (Salmo salar L.) : polymorphism, linkage analysis and protein modelling

Abstract We show that a.s. all of the connected components of the Wired Spanning Forest are recurrent, proving a conjecture of Benjamini, Lyons, Peres and Schramm. Our analysis relies on a simple martingale involving the effective conductance between the endpoints of an edge in a uniform spanning tree. We believe that this martingale is of independent interest and will find further applications in the study of uniform spanning trees and forests

Lack of genetic differentiation between anadromous and resident sympatric brown trout (Salmo trutta) in a Normandy population

The level of genetic differentiation between anadromous and non-andromous trout (Salmo trutta L.) coexisting in a Norman river (Oir river) was examined at 15 microsatellites markers. Despite a large amount of variation, no genetic differentiation was found between forms. Four domesticated stocks were also genotyped with the same loci. Highly significant differences were found with natural samples allowing to exclude a recent stocking effect on the Oir population. We conclude that the resident and anadromous trout form a single naturally reproducing unit in this river system. However, this conclusion could reflect particular ecological or demographic conditions in the Oir basin and needs to be validated in other hydrographical and ecological contexts.Le niveau de différenciation génétique entre les truites anadromes et résidentes (Salmo trutta L.) coexistant dans une rivière normande (l'Oir) a été mesuré à l'aide de 15 locus microsatellites. Malgré une variabilité génétique importante, aucune différenciation génétique entre les deux formes n'a été décelée. Quatre souches domestiques ont été génotypées avec les mêmes locus. Une différenciation génétique hautement significative a été trouvée entre ces souches et les échantillons naturels permettant d'exclure un effet récent du repeuplement sur le secteur étudié. Les truites résidentes et anadromes de ce secteur semblent donc provenir d'une même unité de reproduction. Cette conclusion pourrait être propre à ce bassin et devra être confirmée sur d'autres bassins hydrographiques correspondant à d'autres contextes démographiques et écologiques