Evidence of major genes affecting resistance to bacterial cold water disease in rainbow trout using Bayesian methods of segregation analysis.

Bacterial cold water disease (BCWD) causes significant economic loss in salmonid aquaculture. We previously detected genetic variation for BCWD resistance in our rainbow trout population, and a family-based selection program to improve resistance was initiated at the National Center for Cool and Cold Water Aquaculture (NCCCWA). This study investigated evidence of major trait loci affecting BCWD resistance using only phenotypic data (without using genetic markers) and Bayesian methods of segregation analysis (BMSA). A total of 10,603 juvenile fish from 101 full-sib families corresponding to 3 generations (2005, 2007, and 2009 hatch years) of the NCCCWA population were challenged by intraperitoneal injection with Flavobacterium psychrophilum, the bacterium that causes BCWD. The results from single- and multiple-QTL models of BMSA suggest that 6 to 10 QTL explaining 83 to 89% of phenotypic variance with either codominant or dominant disease-resistant alleles plus polygenic effects may underlie the genetic architecture of BCWD resistance. This study also highlights the importance of polygenic background effects in the genetic variation of BCWD resistance. The polygenic heritability on the observed scale of survival status is slightly larger than that previously reported for rainbow trout BCWD resistance. These findings provide the basis for designing informative crosses for QTL mapping and carrying out genome scans for QTL affecting BCWD resistance in rainbow trout.

Response to selection for bacterial cold water disease resistance in rainbow trout.

A family-based selection program was initiated at the National Center for Cool and Cold Water Aquaculture in 2005 to improve resistance to bacterial cold water disease (BCWD) in rainbow trout. The objective of this study was to estimate response to 2 generations of selection. A total of 14,841 juvenile fish (BW = 3.1 g; SD = 1.1 g) from 230 full-sib families and 3 randomly mated control lines were challenged intraperitoneally with Flavobacterium psychrophilum, the bacterium that causes BCWD, and mortalities were observed for 21 d. Selection was applied to family EBV derived from a proportional-hazards frailty (animal) model while constraining rate of inbreeding to

Evidence of major genes affecting stress response in rainbow trout using Bayesian methods of complex segregation analysis.

As a first step toward the genetic mapping of QTL affecting stress response variation in rainbow trout, we performed complex segregation analyses (CSA) fitting mixed inheritance models of plasma cortisol by using Bayesian methods in large full-sib families of rainbow trout. To date, no studies have been conducted to determine the mode of inheritance of stress response as measured by plasma cortisol response when using a crowding stress paradigm and CSA in rainbow trout. The main objective of this study was to determine the mode of inheritance of plasma cortisol after a crowding stress. The results from fitting mixed inheritance models with Bayesian CSA suggest that 1 or more major genes with dominant cortisol-decreasing alleles and small additive genetic effects of a large number of independent genes likely underlie the genetic variation of plasma cortisol in the rainbow trout families evaluated. Plasma cortisol is genetically determined, with heritabilities of 0.22 to 0.39. Furthermore, a major gene with an additive effect of -42 ng/mL (approximately 1.0 genetic SD) is segregating in this rainbow trout broodstock population. These findings provide a basis for designing and executing genome-wide linkage studies to identify QTL for stress response in rainbow trout broodstock and markers for selective breeding.

Rainbow trout resistance to bacterial cold-water disease is moderately heritable and is not adversely correlated with growth.

The objectives of this study were to estimate the heritabilities for and genetic correlations among resistance to bacterial cold-water disease and growth traits in a population of rainbow trout (Oncorhynchus mykiss). Bacterial cold-water disease, a chronic disease of rainbow trout, is caused by Flavobacterium psychrophilum. This bacterium also causes acute losses in young fish, known as rainbow trout fry syndrome. Selective breeding for increased disease resistance is a promising strategy that has not been widely used in aquaculture. At the same time, improving growth performance is critical for efficient production. At the National Center for Cool and Cold Water Aquaculture, reducing the negative impact of diseases on rainbow trout culture and improving growth performance are primary objectives. In 2005, when fish averaged 2.4 g, 71 full-sib families were challenged with F. psychrophilum and evaluated for 21 d. Overall survival was 29.3% and family rates of survival varied from 1.5 to 72.5%. Heritability of postchallenge survival, an indicator of disease resistance, was estimated to be 0.35 +/- 0.09. Body weights at 9 and 12 mo posthatch and growth rate from 9 to 12 mo were evaluated on siblings of the fish in the disease challenge study. Growth traits were moderately heritable, from 0.32 for growth rate to 0.61 for 12-mo BW. Genetic and phenotypic correlations between growth traits and resistance to bacterial cold-water disease were not different from zero. These results suggest that genetic improvement can be made simultaneously for growth and bacterial cold-water disease resistance in rainbow trout by using selective breeding.

Genetic diversity and background linkage disequilibrium in the North American Holstein cattle population.

The objectives of this study were to 1) identify highly heterozygous Holstein bulls that are as unrelated as possible and widely used in the US dairy industry; 2) quantify the level of genetic diversity in US Holsteins; and 3) determine the extent of background linkage disequilibrium (BLD) and disease trait associated linkage disequilibrium (DLD) in the US Holstein population. Twenty-three Holstein bulls that are not closely related but were widely used in the US dairy industry were genotyped for 54 microsatellite loci. The genotyping was performed on automated DNA sequencers (PE Applied Biosystems, CA), following polymerase chain reaction amplification with fluorescent dye-labeled primers. The heterozygosity for the sampled population ranged from 0.43 to 0.80. This wide range of heterozygosity allows selection of the most heterozygous bulls to develop informative families for gene mapping studies. The degree of genetic diversity in this population is significant and allows selection for traits of economic importance. As expected, there is extensive linkage disequilibrium (LD) in the US Holstein population. About half of the syntenic marker pairs presented a typical pattern of LD produced by DLD. Most of the nonsyntenic marker pairs had a typical pattern of LD arising from BLD. These results suggest that the observed LD is not purely due to genetic drift and migration and that a portion might be due to DLD. This raises our hopes of successful fine-localization of genes for complex traits using LD mapping.

Genetic mapping of quantitative trait loci affecting susceptibility to Marek's disease virus induced tumors in F2 intercross chickens.

Marek's disease (MD) is a lymphoproliferative disease caused by the MD virus (MDV), which costs the poultry industry nearly $1 billion annually. To identify quantitative trait loci (QTL) affecting MD susceptibility, the inbred lines 6(3) (MD resistant) and 7(2) (MD susceptible) were mated to create more than 300 F2 chickens. The F2 chickens were challenged with MDV JM strain, moderately virulent) at 1 wk of age and assessed for MD susceptibility. The QTL analysis was divided into three stages. In stage 1, 65 DNA markers selected from the chicken genetic maps were typed on the 40 most MD-susceptible and the 40 most MD-resistant F2 chickens, and 21 markers residing near suggestive QTL were revealed by analysis of variance (ANOVA). In stage 2, the suggestive markers plus available flanking markers were typed on 272 F2 chickens, and three suggestive QTL were identified by ANOVA. In stage 3, using the interval mapping program Map Manager and permutation tests, two significant and two suggestive MD QTL were identified on four chromosomal subregions. Three to five loci collected explained between 11 and 23% of the phenotypic MD variation, or 32-68% of the genetic variance. This study constitutes the first report in the domestic chicken on the mapping of non-major histocompatibility complex QTL affecting MD susceptibility.

Mapping quantitative trait loci for binary traits using a heterogeneous residual variance model: an application to Marek's disease susceptibility in chickens.

A typical problem in mapping quantitative trait loci (QTLs) comes from missing QTL genotype. A routine method for parameter estimation involving missing data is the mixture model maximum likelihood method. We developed an alternative QTL mapping method that describes a mixture of several distributions by a single model with a heterogeneous residual variance. The two methods produce similar results, but the heterogeneous residual variance method is computationally much faster than the mixture model approach. In addition, the new method can automatically generate sampling variances of the estimated parameters. We derive the new method in the context of QTL mapping for binary traits in a F2 population. Using the heterogeneous residual variance model, we identified a QTL on chromosome IV that controls Marek's disease susceptibility in chickens. The QTL alone explains 7.2% of the total disease variation.

Non-association between Rfp-Y major histocompatibility complex-like genes and susceptibility to Marek's disease virus-induced tumours in 6(3) x 7(2) F2 intercross chickens.

Marek's disease (MD) is a lymphoproliferative disease caused by a member of the herpesvirus family, and the best understood genetic resistance to MD involves the chicken major histocompatibility complex (MHC) B-complex. Preliminary observations have suggested that MHC-like Rfp-Y genes might also influence the incidence of MD. This study describes the differentiation and definition of unique Rfp-Y genes in inbred lines 6(3) and 7(2), lines that possess identical B-complex genes, but that are resistant or susceptible to MD, respectively. To assess if Rfp-Y genes affect susceptibility to MD, 265 6(3) x 7(2) F2 chickens were challenged with the JM strain of MD virus at 1 week of age and were evaluated for MD lesions at up to 10 weeks of age. Genotyping of the F2 chickens for Rfp-Y haplotypes was performed by restriction fragment length polymorphism analysis of genomic DNA using TaqI and a B-FIV probe. Analysis of variance and interval mapping procedures were used to determine association between the Rfp-Y haplotypes and the phenotypic MD values of the F2 chickens. The cosegregation analysis of 265 F2 chickens indicated that there was no association between Rfp-Y haplotypes and MD susceptibility. Furthermore, the fact that the Rfp-Y haplotypes fit the 1:2:1 segregation ratio and the Rfp-Y allele frequencies did not differ significantly from 0.5 in the full population or in selected subpopulations (of either 40 MD-resistant or 39 MD-susceptible chickens) also indicated that Rfp-Y haplotypes do not significantly influence MD susceptibility. We conclude that Rfp-Y haplotypes do not play a major role in determining the genetic susceptibility to MD in 6(3) x 7(2) F2 White Leghorn chickens.

Identification of Rfp-Y (Mhc-like) haplotypes in chickens of Cornell lines N and P.

Two strains of chickens selected for differential Marek's disease (MD) resistance or susceptibility were studied for the presence of the recently described Rfp-Y major histocompatibility complex (Mhc-like) haplotypes. MD resistant chickens from line N were fixed for the classical Mhc B21 haplotype, whereas MD susceptible line P chickens were fixed for the B19 haplotype. The Rfp-Y haplotypes were identified by restriction fragment polymorphism (RFP) analysis using enzymes and Mhc probes for B-G, B-L beta II, and B-FIV. In addition an Rfp-L beta III clone was developed that differentiated Rfp-L beta from B-L beta genes. Three Rfp-Y haplotypes, defined for both class I and class II Mhc-like loci, were identified in line N (Rfp-Y5, Rfp-Y7, and Rfp-Y8) and in line P (Rfp-Y5, Rfp-Y8, and Rfp-Y9), respectively. The Rfp-Y7, Rfp-Y8, and Rfp-Y9 haplotypes have not been described previously. The Rfp-Y5 haplotype was most frequent (0.70) in line N, but existed in low frequency (approximately 0.04) in line P; the Rfp-Y9 haplotype was most frequent in line P (0.63), but was absent in line N. The Rfp-Y haplotypes-segregated in a Mendelian fashion in each line based on analysis of progeny from Rfp-Y heterozygous matings. The frequency of recombination between the Rfp-Y F and L loci was estimated to be less than 0.25%.

Mapping functional chicken genes: an alternative approach.

Functional genes were selected for linkage analysis mapping using the East Lansing (EL) reference population ¿[Jungle Fowl (JF) x White Leghorn (WL)] x WL¿. The approach used was based on the identification of DNA sequence polymorphisms in the introns of those genes found in JF and WL. Deoxyribonucleic acid sequence analysis revealed single base substitutions in introns of six Type I marker genes: adenylate kinase 1 (AK1), aldolase B (ALDOB), a lysosomal membrane protein gene (LAMP1), vitellogenin 2 (VTG2), apolipoprotein A1 (APOA1), and creatine kinase B (CKB). Transitions or transversions were found in introns of AK1, ALDOB, LAMP1, VTG2, APOA1, and CKB. A transversion in the intron of the JF allele of AK1 generated a unique BspHI cleavage site. The design of polymerase chain reaction (PCR) primers based on the site of base substitution led to the specific amplification of the JF allele in the remaining five genes. A size polymorphism in the PCR production derived from iron response element binding protein (IREBP) distinguished the JF from the WL allele. Linkage analysis of the EL reference population revealed that these candidate genes were located in the following EL linkage groups (E) or chromosomes (Chrom) of the chicken genome: AK1 (E41), VTG2 (E43), APOA1 (E49), CKB (E07), LAMP1 (E01), ALDOB (Chrom Z), and IREBP (Chrom Z). Provided that a base substitution can be found in the parents of the reference population, this PCR-based approach can be used to map any cloned candidate gene. This approach will lead to further information on synteny of the chicken genome with cognate genes of mammalian species.

Development of a genetic map of the chicken with markers of high utility.

Microsatellites are tandem duplications with a simple motif of one to six bases as the repeat unit. Microsatellites provide an excellent opportunity for developing genetic markers of high utility because the number of repeats is highly polymorphic, and the assay to score microsatellite polymorphisms is quick and reliable because the procedure is based on the polymerase chain reaction (PCR). We have identified 404 microsatellite-containing clones of which 219 were suitable as microsatellite markers. Primers for 151 of these microsatellites were developed and used to detect polymorphisms in DNA samples extracted from the parents of two reference populations and three resource populations. Sixty, 39, 46, 49, and 61% of the microsatellites exhibited length polymorphisms in the East Lansing reference population, the Compton reference population, resource population No. 1 (developed to identify resistance genes to Marek's disease), resource population No. 2 (developed to identify genes involved in abdominal fat), and resource population No. 3 (developed to identify genes involved in production traits), respectively. The 91 microsatellites that were polymorphic in the East Lansing reference population were genotyped and 86 genetic markers were eventually mapped. In addition, 11 new random amplified polymorphic DNA (RAPD) markers and 24 new markers based on the chicken CR1 element were mapped. The addition of these markers increases the total number of markers on the East Lansing genetic map to 273, of which 243 markers are resolved into 32 linkage groups. The map coverage within linkage groups is 1,402 cM with an average spacing of 6.7 cM between loci. The utility of the genetic map is greatly enhanced by adding 86 microsatellite markers. Based on our current map, approximately 2,550 cM of the chicken genome is within 20 cM of at least one microsatellite marker.