Quantitative trait loci and genes associated with salmonid alphavirus load in Atlantic salmon: implications for pancreas disease resistance and tolerance.

Salmonid alphavirus infection results in pancreas disease causing severe economic losses for Atlantic salmon aquaculture. Knowledge about genes and pathways contributing to resistance is limited. A 54 K SNP panel was used to genotype 10 full-sibling families each consisting of ~ 110 offspring challenged with salmonid alphavirus subtype 3. Relative heart viral load was assessed at 4- and 10-weeks post-infection using quantitative PCR. A moderate genomic heritability of viral load at 4 weeks (0.15-0.21) and a high positive correlation with survival (0.91-0.98) were detected. Positions of QTL detected on chromosome 3 matched those for survival detected by other studies. The SNP of highest significance occurred in the 3' untranslated region of gig1, a fish-specific antiviral effector. Locus B of immunoglobulin heavy chain mapped to an area containing multiple SNPs with genome-wide association. Heart mRNA-seq comparing parr from families with high- versus low-genomic breeding value, and matching sample genotypes for SNPs, identified two eQTL for salmonid alphavirus load. Immune genes associated with trans-eQTL were numerous and spread throughout the genome. QTL regions contained several genes with known or predicted immune functions, some differentially expressed. The putative functional genes and variants identified could help improve marker-based selection for pancreas disease resistance.

Response of the Salmon Heart Transcriptome to Pancreas Disease: Differences Between High- and Low-Ranking Families for Resistance.

Pancreas disease caused by salmonid alphaviruses leads to severe losses in Atlantic salmon aquaculture. The aim of our study was to gain a better understanding of the biological differences between salmon with high and low genomic breeding values (H-gEBV and L-gEBV respectively) for pancreas disease resistance. Fish from H- and L-gEBV families were challenged by intraperitoneal injection of salmonid alphavirus or co-habitation with infected fish. Mortality was higher with co-habitation than injection, and for L- than H-gEBV. Heart for RNA-seq and histopathology was collected before challenge and at four- and ten-weeks post-challenge. Heart damage was less severe in injection-challenged H- than L-gEBV fish at week 4. Viral load was lower in H- than L-gEBV salmon after co-habitant challenge. Gene expression differences between H- and L-gEBV manifested before challenge, peaked at week 4, and moderated by week 10. At week 4, H-gEBV salmon showed lower expression of innate antiviral defence genes, stimulation of B- and T-cell immune function, and weaker stress responses. Retarded resolution of the disease explains the higher expression of immune genes in L-gEBV at week 10. Results suggest earlier mobilization of acquired immunity better protects H-gEBV salmon by accelerating clearance of the virus and resolution of the disease.

Whole genome duplication: challenges and considerations associated with sequence orthology assignment in Salmoninae.

To illustrate some of the challenges and considerations in assigning correct orthology necessary for any comparative genomic investigation among salmonids, sequence data from the non-coding regions of different chromosomes in three members of the subfamily Salmoninae, rainbow trout Oncorhynchus mykiss, Atlantic salmon Salmo salar and Arctic charr Salvelinus alpinus, were compared. By analysing c. 55 distinct loci, corresponding to c. 142 kbp sequence information per species, 18 duplicated patterns representative of the two sequential rounds of teleost-specific whole genome duplications (i.e. 3R and 4R WGD) were identified. Sequence similarities between the 4R paralogues were c. 90%, which was slightly lower than those of the 4R orthologues and c. 60% for the 3R products. Through careful examination of the sequence data, however, only 14 loci could reliably be assigned as true orthologues. Locus-specific trees were constructed through maximum parsimony, maximum likelihood and neighbour-joining methods and were rooted using the information from a close relative, lake whitefish Coregonus clupeaformis. All approaches generated congruent trees supporting the {Coregonus [Salmo (Oncorhynchus, Salvelinus)]} topology. The general phenotypic characteristics of sequences, however, were highly suggestive of the basal position of Oncorhynchus, raising the hypothesis of an accelerated rate of nucleotide evolution in this species.

The genetic architecture of embryonic developmental rate and genetic covariation with age at maturation in rainbow trout Oncorhynchus mykiss.

The genetic architecture underlying variation in embryonic developmental rate (DR) and genetic covariation with age of maturation (MAT) was investigated in rainbow trout Oncorhynchus mykiss. Highly significant additive parental effects and more limited evidence of epistatic effects on progeny hatching time were detected in three diallel sets of families. Genome scans with an average of 142 microsatellite loci from all 29 linkage groups in two families detected significant quantitative trait loci (QTL) for developmental rate on RT-8 and RT-30 with genome-wide and chromosome-wide effects, respectively. The QTL on linkage group RT-8 explained 23·7% of the phenotypic variation and supports results from previous studies. The co-localization of QTL for both DR and MAT to several linkage groups and the observation that alleles associated with faster developmental rate were found significantly more often in early maturing rather than typical and later maturing male ancestors supports the hypothesis of genetic covariation between DR and MAT. The maturation background and schedule of additional sires, however, did not have a consistent association with their progeny hatching times, suggesting that other genetic, environmental and physiological effects contribute to variation in these life-history traits.

Genetic mapping of the ornithine decarboxylase (odc) gene complex in rainbow trout (Oncorhynchus mykiss).

Ornithine decarboxylase (odc) and its associated gene complex, including antizymes of ODC and inhibitors of the antizymes, play a key role in regulating the overall polyamine levels (i.e., putrescine) in cells. Polyamine production levels are intricately coupled to mitotic and cellular turnover rates. Hence, these genes may be important candidates of growth regulation in vertebrates, if their chromosomal locations coincide with known quantitative trait locus (QTL) regions influencing growth traits. Here we report the genetic mapping of multiple duplicated forms of genes within this complex to previously known life-history and growth QTL regions in rainbow trout (Oncorhynchus mykiss). Specifically, duplicated copies of the 2 antizyme genes map to Om-8/9/24, while antizyme inhibitor and odc duplicates map to Om-27/31 and Om-14/25, respectively. All sets of paralogous mapping locations correspond to ancestrally duplicated synteny regions within the genome of rainbow trout (i.e., ancestral A, B, GH/I and M linkage groups).

Genomic organization of the IGF1, IGF2, MYF5, MYF6 and GRF/PACAP genes across Salmoninae genera.

Whole-genome duplication in the ancient ray-finned fish and subsequent tetraploidization in the ancestor to the salmonids have complicated genomic and candidate gene studies in these organisms as many genes with multiple copies are present throughout their genomes. In an attempt to identify genes with a potential influence on growth and development, we investigated the genomic positions of insulin-like growth factors 1 and 2 (IGF1, IGF2), myogenic factors 5 and 6 (MYF5, MYF6) and growth hormone-releasing factor/pituitary adenylate cyclase-activating polypeptide (GRF/PACAP) in three salmonid species: rainbow trout (Oncorhynchus mykiss), Atlantic salmon (Salmo salar) and Arctic charr (Salvelinus alpinus). Our results suggest a tight association between the IGF1, MYF5 and MYF6 genes in all three species. We further localized the duplicated copies of IGF1 to the homeologous linkage groups RT-7/15 in rainbow trout and AC-3/24 in Arctic charr, and the two copies of MYF6 to homeologous linkage groups AS-22/24 in Atlantic salmon. Localization of GRF/PACAP to RT-7, AS-31 and AC-27 and IGF2 to RT-27, AS-2 and AC-4 in rainbow trout, Atlantic salmon and Arctic charr respectively is consistent with previously reported homologies among these chromosomal segments identified using other genetic markers. However, localization of the second copy of GRF/PACAP to RT-19 and AC-14 and the duplicated copy of IGF2 to AC-19 suggest a possible new homology/homeology between these chromosomes. These results might also be an indication of a more ancient polyploidization event that occurred deep in the ray-finned fish lineage.

Genome organization of glutamine synthetase genes in rainbow trout (Oncorhynchus mykiss).

Unlike mammals, bony fish appear to possess multiple genes encoding glutamine synthetase (GS), the nitrogen metabolism enzyme responsible for the conversion of glutamate and ammonia into glutamine at the expense of ATP. This study reports on the development of genetic markers for each of the four isoforms identified thus far in rainbow trout (Oncorhynchus mykiss) and their genome localization by linkage mapping. We found that genes coding for GS01, GS02, GS03, and GS04 map to four different linkage groups in the trout genome, namely RT-24, RT-23, RT-08, and RT-13, respectively. Linkage groups RT-23 and RT-13 appear to represent distinct chromosomes sharing duplicated marker regions, which lends further support to the previous suggestion that GS02 and GS04 may be duplicate gene copies that evolved from a whole-genome duplication in the trout ancestor. In contrast, there is at present no further evidence that RT-24 and RT-08 share ancestrally homologous segments and additional genomic studies will be needed to clarify the evolutionary origin of genes coding for GS01 and GS03.

Heritability of sudden death syndrome and its associated correlations to ascites and body weight in broilers.

(1) Genetic parameters for the sudden death syndrome (SDS) were estimated in meat-type chickens. Data were collected over 11 generations of selection for body weight within two distinct breeds (Cornish and White Rock). (2) The animal model was used exclusively with linear methods (LM) to estimate genetic parameters. Heritability (h2) of SDS on the liability scale was 0.30 +/- 0.002 and 0.25 +/- 0.002 in the Cornish and White Rock breeds, respectively. (3) A positive genetic correlation (r(g)) with ascites (AS) was determined (approximately 0.3 +/- 0.006). However, it was not possible to estimate the rg of SDS with body weight because of the low prevalence of the defect trait studied (1.8% in the Cornish and 1-5% in the White Rock). (4) Heritability of SDS calculated using male records only was 0.45 +/- 0.009 and 0.35 +/- 0.009, and r(g) with body weight was 0.30 +/- 0.010 and 0.27 +/- 0.009, in the Cornish and White Rock breeds, respectively. (5) In conclusion, the heart defect investigated was heritable with a positive genetic correlation with AS and body weight.

Marker-assisted estimation of quantitative genetic parameters in rainbow trout, Oncorhynchus mykiss.

Estimation of quantitative genetic parameters conventionally requires known pedigree structure. However, several methods have recently been developed to circumvent this requirement by inferring relationship structure from molecular marker data. Here, two such marker-assisted methodologies were used and compared in an aquaculture population of rainbow trout (Oncorhynchus mykiss). Firstly a regression-based model employing estimates of pairwise relatedness was applied, and secondly a Markov Chain Monte Carlo (MCMC) procedure was employed to reconstruct full-sibships and hence an explicit pedigree. While both methods were effective in detecting significant components of genetic variance and covariance for size and spawning time traits, the regression model resulted in estimates that were quantitatively unreliable, having both significant bias and low precision. This result can be largely attributed to poor performance of the pairwise relatedness estimator. In contrast, genetic parameters estimated from the reconstructed pedigree showed close agreement with ideal values obtained from the true pedigree. Although not significantly biased, parameters based on the reconstructed pedigree were underestimated relative to ideal values. This was due to the complex structure of the true pedigree in which high numbers of half-sibling relationships resulted in inaccurate partitioning of full-sibships, and additional unrecognized relatedness between families.

Estimation of genetic parameters for ascites syndrome in broiler chickens.

Genetic parameters for the ascites syndrome (AS) were estimated for meat-type chickens. Data had been collected over 11 generations of selection for body weight and other traits within two distinct breeds (Cornish and White Rock). Linear methods (LM) were used to estimate genetic parameters and also to analyze a binary measure of survival. Survival analyses (SA) were also conducted to estimate the effects of various factors influencing the incidence of AS by evaluating the number of days that the birds survived. The animal model was used exclusively with linear methods. Heritabilities (h2) on the liability scale were 0.12 +/- 0.02 and 0.22 +/- 0.01 in the White Rock and Cornish breeds, respectively; however, the genetic correlation (r(g)) with body weight was not possible to estimate due to the low prevalence of the defect trait studied (1.5% in the Cornish and 1.1% in the White Rock). Because males are more prone to AS, the h2 using the male records only were 0.22 +/- 0.017 and 0.41 +/- 0.009, and the r(g) were 0.35 +/- 0.007 and 0.22 +/- 0.009 in the dam and sire populations, respectively. In conclusion, the heart defect investigated was heritable and had a positive genetic correlation with body weight.

Overall nutrient intake of preschool hyperactive and normal boys.

Studies examining the relationship between sugar intake of hyperactive children and behavior problems have reported inconsistent results. We hypothesized that if the problem behaviors of attention deficit-disordered children with hyperactivity (ADDH) are exacerbated by food, then this might be due to differences in their background nutrition relative to non-ADDH children. Parents and day care workers of 24 ADDH and 27 non-ADDH preschool-aged boys were trained to keep food diaries for 21 days. Analysis of the last 14 days for each child revealed no differences in overall nutrient intake between the two groups. On the other hand, for 5 ADDH and 3 non-ADDH children the within-subject correlations between daily behavior and sugar intake were significant. We concluded that nutrition-behavior interactions are more likely a function of idiosyncratic sensitivities, rather than a general tendency for ADDH children to eat differently from non-ADDH children.

Dietary replacement in preschool-aged hyperactive boys.

A 10-week study was conducted in which all food was provided for the families of 24 hyperactive preschool-aged boys whose parents reported the existence of sleep problems or physical signs and symptoms. A within-subject crossover design was used, and the study was divided into three periods: a baseline period of 3 weeks, a placebo-control period of 3 weeks, and an experimental diet period of 4 weeks. The experimental diet was broader than those studied previously in that it eliminated not only artificial colors and flavors but also chocolate, monosodium glutamate, preservatives, caffeine, and any substance that families reported might affect their specific child. The diet was also low in simple sugars, and it was dairy free if the family reported a history of possible problems with cow's milk. According to the parental report, more than half of the subjects exhibited a reliable improvement in behavior and negligible placebo effects. In addition, several nonbehavioral variables tended to improve while the children received the experimental diet, particularly halitosis, night awakenings, and latency to sleep onset.

Physical signs and symptoms in preschool-age hyperactive and normal children.

Although little research has been done on the topic, it is commonly believed that attention deficit disorder with hyperactivity (ADDH) is often associated with elevated levels of physical complaints. Three studies were conducted to evaluate the prevalence of physical signs and symptoms (primarily gastrointestinal, respiratory, and dermal) in preschool-age ADDH and non-ADDH children. In the first two studies, the parents of ADDH children reported consistently greater frequency of such signs and symptoms than did the parents of non-ADDH children. In the third study, parents kept daily logs for 21 days, in which they recorded three times each day the presence or absence of each sign and symptom. In this study, there also was a remarkable increase in the frequency of reported physical signs and symptoms for the ADDH children. Several possible mechanisms for these findings are discussed.

Sleep disturbance in preschool-aged hyperactive and nonhyperactive children.

In spite of inadequate laboratory demonstrations of sleep problems in children with attention deficit disorder with hyperactivity, the belief persists that such problems exist. Sleep restlessness is, in fact, one of the criteria in the Diagnostic and Statistical Manual of Mental Disorders, ed 3, definition of attention deficit disorder with hyperactivity, and sleep problems are listed on two major checklists often used for describing the symptoms of this disorder. In a series of three studies, sleep problems were investigated in preschool-aged children with attention deficit disorder relative to control children without the disorder. Results of the first two studies demonstrated clearly that parents of hyperactive children considered their children to have many more sleep problems than did parents of the control children. Parental daily documentation, which is less likely to be affected by reporting bias, was used in the third study. Although the results of the third study supported the finding of increased frequency of night wakings in these children, there was no difference in total sleep time or sleep onset latency between the two groups. Two other significant group differences (enuresis and night sweats) were primarily due to subgroups of children with attention deficit disorder and hyperactivity. The greater number of sleep wakings, which disrupt parents' sleep, may be responsible for the clinical reports that these children are poor sleepers.