QTLs associated with growth traits and abdominal fat weight and their interactions with gender and hatch in commercial meat-type chickens.

Associations between microsatellite markers and traits related to growth and fatness were investigated using resource broiler population. A sire-line x dam-line F1 male was backcrossed to 12 dam-line females to produce 24 sires and 47 dams of the backcross 1 (BC1) generation. These 71 parents were genotyped with 76 microsatellite markers. Following full-sib mating among the parents, 234 BC1-F2 progeny were phenotyped for five growth traits (body weight at 49 days from hatch, wog weight, front half weight, breast weight and tender weight) and abdominal fat weight. Maximum likelihood analysis was used to estimate the marker effects and to evaluate their statistical significance. Individual marker-trait analysis revealed 44 significant associations out of the 456 marker-trait combinations. Correction for multiple comparisons by controlling the false discovery rate (FDR) resulted in 12 significant associations at FDR = 10% with markers on chromosomes 1, 2, 5 and 13. Seventy-five percent of the 44 significant associations displayed no dependence on either hatch or gender; half of the remaining associations displayed dependence of the quantitative trait loci (QTL) effect on hatch x gender interaction. Thus, the analysed traits in this study may be dependent on external factors.

Assessment of genetic relationships in Cucurbita pepo (Cucurbitaceae) using DNA markers.

Cucurbita pepo (pumpkin, squash, gourd), an economically important species of the Cucurbitaceae, is extremely variable in fruit characteristics. The objective of the present study was to clarify genetic relationships across a broad spectrum of the C. pepo gene pool, with emphasis on domesticates, using AFLP, ISSR and SSR markers. Forty-five accessions were compared for presence or absence of 448 AFLP, 147 ISSR, and 20 SSR bands, their genetic distances (GDs) were estimated and UPGMA cluster analysis was conducted. The results obtained from these three marker systems were highly correlated (P << 0.001). Clustering was in accordance with the division of C. pepo into three subspecies, fraterna, texana and pepo, with the first two less distant to one another than to the last one. Within the clusters, sub-clustering occurred in accordance with fruit shape and size. The subsp. texana cluster consisted of six sub-clusters, one each for the representatives of its five cultivar-groups (Acorn, Crookneck, Scallop, Straightneck and Ovifera Gourd) and wild gourds. Within the subsp. pepo cluster, the representatives of two cultivar-groups (Zucchini and Orange Gourd) formed distinct sub-clusters and the representatives of two other groups (Cocozelle and Vegetable Marrow) tended to sub-cluster separately from one another but formed an assemblage with the representatives of the remaining group (Pumpkin). Within-group GDs were less than corresponding between-group GDs in nearly all comparisons. The smallest-fruited accession, 'Miniature Ball', appears to occupy a genetically central position within C. pepo.

Concentration of immnoglobulin G in plasma varies among 6C.7 recombinant congenic strains of chickens.

Chicken Lines 63 and 72 were inbred during selection for resistance or susceptibility to viral-induced tumors. A sandwich ELISA assay was adapted to define the milligrams per milliliter of Ig-gamma (IgG) in plasma from chickens of Lines 63 and 72, as well as 19 recombinant congenic strains (RCS). Each RCS resulted from a 7(2) x 6(3) F(1) and two backcross matings using 63 as the recurrent female line. The IgG levels in the RCS were evaluated after four to seven generations of sib-matings, when each RCS was becoming inbred and fixed for a different 12.5% of the 72 genome. In three generations approximately 24-wk-old chickens of Line 72 had higher levels of plasma IgG than chickens of Line 63 (P < 0.05). None of the RCS had repeatable IgG levels comparable to Line 7(2). However, in the last two generations, two of the 18 RCS had higher IgG levels than nine with low IgG levels (P < 0.05). There was no correlation between an IgG level of a RCS and resistance to Marek's disease. It was concluded that selected RCS may be useful for identifying genes that determine differences in IgG levels, as well as for understanding the relationship between genes, IgG levels, and other traits that differ between Lines 63 and 72.

Quantitative trait locus mapping in chickens by selective DNA pooling with dinucleotide microsatellite markers by using purified DNA and fresh or frozen red blood cells as applied to marker-assisted selection.

Many large, half-sib sire families are an integral component of chicken genetic improvement programs. These family structures include a sufficient number of individuals for mapping quantitative trait loci (QTL) at high statistical power. However, realizing this statistical power through individual or selective genotyping is yet too costly to be feasible under current genotyping methodologies. Genotyping costs can be greatly reduced through selective DNA pooling, involving densitometric estimates of marker allele frequencies in pooled DNA samples. When using dinucleotide microsatellite markers, however, such estimates are often confounded by overlapping "shadow" bands and can be confounded further by differential amplification of alleles. In the present study a shadow correction procedure provided accurate densitometric estimates of allele frequency for dinucleotide microsatellite markers in pools made from chicken purified DNA samples, fresh blood samples, and frozen-thawed blood samples. In a retrospective study, selective DNA pooling with thawed blood samples successfully identified two QTL previously shown by selective genotyping to affect resistance in chickens to Marek's disease. It is proposed that use of selective DNA pooling can provide relatively low-cost mapping and use in marker-assisted selection of QTL that affect production traits in chickens.

DNA microsatellites linked to quantitative trait loci affecting antibody response and survival rate in meat-type chickens.

Selection for immune response parameters may lead to improved general disease resistance. Because disease resistance and immune response are hard-to-measure quantitative traits with low to moderate heritability, they may respond more efficiently to marker-assisted selection (MAS) than to phenotypic selection. To detect DNA markers linked to quantitative trait loci (QTL) associated with immune response, a resource half-sib family of 160 backcross (BC1) and intercross (F2) birds was derived from a cross between two meat-type lines divergently selected for high or low antibody (Ab) response to Escherichia coli. By using 25 microsatellite DNA markers covering approximately 25% of the chicken genome, initial genotyping of 40% of the resource family was followed by complete genotyping of the entire family with four suggestive markers. Three of these markers exhibited significant association with immune response: (1) ADL0146 on Chromosome 2 associated with Ab to SRBC and Newcastle disease virus (NDV), (2) ADL0290 on linkage group 31 affecting Ab to NDV, and (3) ADL0298 on linkage group 34 associated with Ab to E. coli and survival. The family was also genotyped with five linked markers from two of the suggested regions, and interval mapping was applied. The results confirmed the significant effects, suggested the location of the QTL, and confirmed the genetic association between immune responses and disease resistance. These findings support the idea of improving poultry immunocompetence by MAS.

The dynamics of antibody response to Escherichia coli vaccination in meat-type chicks.

The dynamics of serum antibody (Ab) response in young broilers were studied in lines divergently selected for high (HC) or low (LC) Ab response to Escherichia coli vaccination at an early age, and their cross (HL). Chicks were divided into three vaccination-age (VA) groups: 8, 10, and 12 d of age (VA8, VA10, and VA12, respectively). Antibody response was determined five times for each chick, at 6, 8, 10, 12, and 14 d postvaccination (dPV). The effects of line, VA, and dPV on Ab titers were highly significant. The HC and LC chicks exhibited the highest and lowest mean titers, respectively, in all VA groups. The HL chicks exhibited midparent Ab values for all VA and dPV combinations, indicating additive inheritance of early Ab production. In LC, the highest mean Ab titer was obtained on Day 26 (14 dPV of the VA12 group), whereas in HC, the same titer had already been obtained on Day 18 (VA8-10 dPV and VA10-8 dPV combinations). The VA8 and VA12 chicks differed markedly in their Ab titer dynamics curves, and the VA10 chicks exhibited an intermediate curve. The three VA groups exhibited a similar change in Ab level from 6 to 10 dPV, but they differed in Ab change from 10 to 14 dPV. This significant dPV x VA interaction suggests that the VA12 and VA10, but not VA8, chicks maintained the capability to produce persisting Ab.

Microsatellite polymorphism between and within broiler populations.

Two independent broiler chicken populations were genotyped with microsatellite markers to determine genetic polymorphisms within and among broiler populations. Birds were genotyped with primers from the US Poultry Genome Mapping Kits 1 and 2. The 59 primer sets selected for this study provided wide genomic coverage. All 59 primer sets amplified a polymerase chain reaction product in Population L, whereas 57 primer sets produced a product in Population C. The average allele number per line per microsatellite was 2.8 and 2.9 for Populations L and C, respectively. Considering the 57 primer pairs generating product in both lines, 72.3% of the total alleles were unique to one or the other population. This study illustrates the high polymorphism level in broiler populations of microsatellites amplified from primers developed from Red Jungle Fowl or White Leghorn sequences.

Detection of RFLP markers associated with antibody response in meat-type chickens: haplotype/genotype, single-band, and multiband analyses of RFLP in the major histocompatibility complex.

Improving disease resistance in poultry by direct selection or by selecting for immune response is hardly feasible due to the quantitative nature of these traits, their low heritability, and the difficulties associated with reliable measurements. In this situation, marker-assisted selection (MAS) is expected to be a more effective breeding approach. The major histocompatibility complex (MHC), known to affect immune response and disease resistance, was examined as a set of candidate genes for association between DNA markers and antibody response. Backcross (BC1) and F2 families were generated from a cross between lines divergently selected for high or low antibody response to Escherichia coli vaccination. Restriction fragment length polymorphism (RFLP) analysis of the highly polymorphic MHC class IV (B-G) region suggested an association with antibody response to several antigens (E. coli, SRBC, NDV). The multiband data generated with the class IV probe were used to compare the efficacies of three alternative analyses: "single-band" (carriers versus noncarriers of each RFLP band separately), "multiband" (multiple regression on all RFLP bands), and "genotype" (determined from family analysis of RFLP patterns/haplotypes). Groups of birds identified by the "multiband" analysis were identical to the haplotype-based genotypes, suggesting that the laborious step of haplotype determination can be omitted without unduly sacrificing power of analysis.

Major histocompatibility complex (MHC) related cDNA probes associated with antibody response in meat-type chickens.

The major histocompatibility complex (MHC) region was examined as a set of candidate genes for association between DNA markers and antibody response. Intercross F2 families of chickens were generated from a cross between high (HC) and low (LC) Escherichia coli(i) antibody lines. Restriction fragment length polymorphism (RFLP) analysis was conducted by using three MHC-related cDNA probes: chicken MHC class IV (B-G), chicken MHC class I (B-F), and human MHC-linked Tap2. Association between RFLP bands and three antibody response traits (E. coli, sheep red blood cells and Newcastle disease virus) were determined by two methods: by statistically analyzing each band separately and also by analyzing all bands obtained from the three probes by using multiple regression analysis to account for the multiple comparisons. The MHC class IV probe was the highest in polymorphisms but had the lowest number of bands associated with antibody response. The MHC class I probe yielded 15 polymorphic bands of which four exhibited association with antibody response traits. The Tap2 probe yielded 20 different RFLP bands of which five were associated with antibody production. Some Tap2 bands were associated with multiple antibody response traits. The multiband analysis of the three probes' bands revealed more significant effects than the analysis of each band separately. This study illustrates the efficacy of using multiple MHC region probes as candidate markers for quantitative trait loci (QTLs) controlling antibody response in chickens.

High resolution mapping and identification of new quantitative trait loci (QTL) affecting susceptibility to Marek's disease.

Marek's disease (MD) is a lymphoproliferative disease of chickens that costs the poultry industry approximately $1 billion annually. Genetic resistance to MD is gaining increased attention to augment vaccinal control as disease outbreaks occur more frequently. Previously, analysis of a 272 F2 White Leghorn resource population measured for many MD traits and genotyped for 78 microsatellite markers revealed two and four quantitative trait loci (QTL) with significant and suggestive association, respectively, to one or more MD associated traits. Additional genetic markers have since been scored on the MD resource population to increase QTL resolution and genome coverage. Saturation of four of the QTL regions with 17 markers revealed five new QTL while 32 markers extended the genome coverage by 400 + CM and uncovered three more QTL. QTL analysis by single-point and interval mapping algorithms agreed well when marker saturation was approximately 20 CM or less. Currently 127 genetic markers cover approximately 68% of the genome that contain up to 14 MD QTL associated to one or more MD trait; seven at the significant level and seven at the suggestive level. Individually each QTL accounts for 2-10% of the variation and, in general, resistance was dominant although the resistant allele may come from either parental line. This study suggests that a limited number of genomic regions play a major role in the genetic control of MD resistance. Markers linked to these loci may be useful for selection of MD resistant stock by the poultry industry following verification of the association within their breeding populations.

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.

Genetic differences and heritability of antibody response to Escherichia coli vaccination in young broiler chicks.

Broiler chicken lines, selected divergently for high (HC) or low (LC) antibody titer to Escherichia coli vaccination at an early age, were evaluated for antibody response at the S5 and S9 generations of selection. The full-pedigreed populations consisted of about 300 and 400 chicks per line in S5 and S9, respectively. At S5, all chicks were vaccinated at 10 d of age (VA10) and antibody titer was determined twice for each chick, at 8 and 12 d postvaccination (dPV). At S9, each line was divided into two equal groups; in the HC line, one group was vaccinated at 8 d of age (VA8), and the other at 10 d of age (VA10), whereas in the LC line, one group was VA10 and the other was VA12. Antibody titers were determined twice for each chick, 8 and 10 dPV. The effects of line, age at vaccination (VA), and days for antibody development (dPV) were tested, and the heritability of antibody titer was estimated for each line-VA-dPV set of data. The HC and LC lines differed significantly in the maturation process of their immune systems. The percentage of chicks with detectable antibody at 18 d of age (VA10-8 dPV) among HC chicks was significantly higher than among LC chicks (85 vs 48% in S5 and 96 vs 63% in S9). In S9, 90% of the HC chicks had already responded at 16 d of age and 100% at 18 d of age, whereas among the LC chicks, only 62% were positive at 18 d of age, increasing to no more than 98% at 22 d of age. The results demonstrates that selection of antibody titer to E. coli vaccination at 20 d of age actually affects the earliest age of immune response, as the immune system of the HC chicks matures earlier than that of the LC chicks. The HC S9 chicks at 8 dPV exhibited a fourfold higher antibody titer than their LC 8 counterparts. This difference further increased at 10 dPV, indicating that the lines differed not only in the level of antibody at a specific age, but also in their rate of antibody titer development. The highest estimate of heritability was very similar in both lines (0.44 and 0.42 in HC and LC, respectively). However, in the HC line this heritability was exhibited at 18 d of age, and only at 22 d in the LC line. Thus, both lines have a similar amount of genetic variation for early immune response, but in the HC line this variation is fully expressed 4 d earlier than in the LC line. These results suggest that selection for high or low antibody response in young chicks results in early or late antibody production, respectively. To maximize the efficiency of selection for early immune response, one must determine the best vaccination age and timing of antibody evaluation in any given population, and these values must be revalidated and updated as selection proceeds.

Response of three class-IV major histocompatibility complex haplotypes to Eimeria acervulina in meat-type chickens.

1. The importance of MHC genes and background genes in controlling disease resistance, including resistance to avian coccidiosis, has not been clarified in meat-type chickens. 2. The role of class IV MHC genes in resistance to Eimeria acervulina was assessed in F2 progeny of a cross between 2 meat-type lines, selected divergently for immune response to Escherichia coli. 3. Disease susceptibility was assessed by lesion score, body weight, packed cell volume and carotene absorption. 4. Chickens with the "K" class IV MHC haplotype had lower lesion scores than chickens with "F" and "A" haplotypes. 5. Plasma carotene concentrations were higher in chickens with "K" haplotype and lower in chickens with "F" and "A" haplotypes whereas body weight and packed cell volume were less sensitive measures of Eimeria infection. 6. Eimeria acervulina resistance appears to be associated with MHC class IV genes; information about MHC haplotypes may be useful in selecting for increased resistance of meat-type chickens to coccidiosis.

Parental effect on the humoral immune response to Escherichia coli and Newcastle disease virus in young broiler chicks.

Genetic and environmental variables influence animal resistance to disease infection. In addition, maternal effects were also found in studies with egg-type chicken lines. In our laboratory, meat-type chicken lines were divergently selected for either early or late maturation of the immune system, based on family and individual antibody responsiveness at 10 d of age. The high-antibody (HC) and low-antibody (LC) lines differed significantly in the early immune response to Escherichia coli, to Newcastle disease virus (NDV) vaccination, and to several other immune functions. Reciprocal crosses between the HC and LC lines were performed over 2 yr at three different locations. Immune responses to E. coli and NDV vaccination provided separate estimates of maternal and paternal effects. Dam effect on immune response to E. coli vaccine was significantly larger than sire effect; the antibody titer in both reciprocal crosses was intermediate between the parental lines, but the mean titer of the HC x LC cross was significantly lower than that of the LC x HC cross. Similar, but not significant, ranking of crosses was observed for the response to NDV. Evidently, the level of the offspring humoral immune response was more a dam than a sire effect.