Cis-acting mutation affecting GJA5 transcription is underlying the Melanotic within-feather pigmentation pattern in chickens.

Melanotic (Ml) is a mutation in chickens that extends black (eumelanin) pigmentation in normally brown or red (pheomelanin) areas, thus affecting multiple within-feather patterns [J. W. Moore, J. R. Smyth Jr, J. Hered. 62, 215-219 (1971)]. In the present study, linkage mapping using a back-cross between Dark Cornish (Ml/Ml) and Partridge Plymouth Rock (ml+/ml+ ) chickens assigned Ml to an 820-kb region on chromosome 1. Identity-by-descent mapping, via whole-genome sequencing and diagnostic tests using a diverse set of chickens, refined the localization to the genomic region harboring GJA5 encoding gap-junction protein 5 (alias connexin 40) previously associated with pigmentation patterns in zebrafish. An insertion/deletion polymorphism located in the vicinity of the GJA5 promoter region was identified as the candidate causal mutation. Four different GJA5 transcripts were found to be expressed in feather follicles and at least two showed differential expression between genotypes. The results showed that Melanotic constitutes a cis-acting regulatory mutation affecting GJA5 expression. A recent study established the melanocortin-1 receptor (MC1R) locus and the interaction between the MC1R receptor and its antagonist agouti-signaling protein as the primary mechanism underlying variation in within-feather pigmentation patterns in chickens. The present study advances understanding the mechanisms underlying variation in plumage color in birds because it demonstrates that the activity of connexin 40/GJA5 can modulate the periodic pigmentation patterns within individual feathers.

Mutations Upstream of the TBX5 and PITX1 Transcription Factor Genes Are Associated with Feathered Legs in the Domestic Chicken.

Feathered leg is a trait in domestic chickens that has undergone intense selection by fancy breeders. Previous studies have shown that two major loci controlling feathered leg are located on chromosomes 13 and 15. Here, we present genetic evidence for the identification of candidate causal mutations at these loci. This was accomplished by combining classical linkage mapping using an experimental cross segregating for feathered leg and high-resolution identical-by-descent mapping using whole-genome sequence data from 167 samples of chicken with or without feathered legs. The first predicted causal mutation is a single-base change located 25 kb upstream of the gene for the forelimb-specific transcription factor TBX5 on chromosome 15. The second is a 17.7-kb deletion located ∼200 kb upstream of the gene for the hindlimb-specific transcription factor PITX1 on chromosome 13. These mutations are predicted to activate TBX5 and repress PITX1 expression, respectively. The study reveals a remarkable convergence in the evolution of the feathered-leg phenotype in domestic chickens and domestic pigeons, as this phenotype is caused by noncoding mutations upstream of the same two genes. Furthermore, the PITX1 causal variants are large overlapping deletions, 17.7 kb in chicken and 44 kb in pigeons. The results of the present study are consistent with the previously proposed model for pigeon that feathered leg is caused by reduced PITX1 expression and ectopic expression of TBX5 in hindlimb buds resulting in a shift of limb identity from hindlimb to more forelimb-like identity.

Changes in adipose tissue physiology during the first two weeks posthatch in chicks from lines selected for low or high body weight.

Chickens from lines selected for low (LWS) or high (HWS) body weight (BW) differ in appetite and adiposity. Mechanisms associated with the predisposition to becoming obese are unclear. The objective of the experiment was to evaluate developmental changes in depot-specific adipose tissue during the first 2 wk posthatch. Subcutaneous (SQ), clavicular (CL), and abdominal (AB) depots were collected at hatch (DOH) and days 4 (D4) and 14 (D14) posthatch for histological and mRNA measurements. LWS chicks had decreased SQ fat mass on a BW basis with reduced adipocyte size from DOH to D4 and increased BW and fat mass with unchanged adipocyte size from D4 to D14. HWS chicks increased in BW from DOH to D14 and increased in fat mass in all three depots with enlarged adipocytes in the AB depot from D4 to D14. Meanwhile, CCAAT/enhancer-binding protein-α, neuropeptide Y, peroxisome proliferator-activated receptor-γ, and acyl-CoA dehydrogenase mRNAs differed among depots between lines at different ages. Plasma nonesterified fatty acids were greater in LWS than HWS at D4 and D14. From DOH to D4, LWS chicks mobilized SQ fat and replenished the reservoir through hyperplasia, whereas HWS chicks were dependent on hyperplasia and hypertrophy to maintain adipocyte size and depot mass. From D4 to D14, adipose tissue catabolism and adipogenesis slowed. Whereas LWS fat depots and adipocyte sizes remained stable, HWS chicks rapidly accumulated fat in CL and AB depots. Chicks predisposed to be anorexic or obese have different fat development patterns during the first 2 wk posthatch.

Genotyping by low-coverage whole-genome sequencing in intercross pedigrees from outbred founders: a cost-efficient approach.

BACKGROUND: Experimental intercrosses between outbred founder populations are powerful resources for mapping loci that contribute to complex traits i.e. quantitative trait loci (QTL). Here, we present an approach and its accompanying software for high-resolution reconstruction of founder mosaic genotypes in the intercross offspring from such populations using whole-genome high-coverage sequence data on founder individuals (~ 30×) and very low-coverage sequence data on intercross individuals (< 0.5×). Sets of founder-line informative markers were selected for each full-sib family and used to infer the founder mosaic genotypes of the intercross individuals. The application of this approach and the quality of the estimated genome-wide genotypes are illustrated in a large F2 pedigree between two divergently selected lines of chickens. RESULTS: We describe how we obtained whole-genome genotype data for hundreds of individuals in a cost- and time-efficient manner by using a Tn5-based library preparation protocol and an imputation algorithm that was optimized for this application. In total, 7.6 million markers segregated in this pedigree and, within each full-sib family, between 10.0 and 13.7% of these were fully informative, i.e. fixed for alternative alleles in the founders from the divergent lines, and were used for reconstruction of the offspring mosaic genotypes. The genotypes that were estimated based on the low-coverage sequence data were highly consistent (> 95% agreement) with those obtained using individual single nucleotide polymorphism (SNP) genotyping. The estimated resolution of the inferred recombination breakpoints was relatively high, with 50% of them being defined on regions shorter than 10 kb. CONCLUSIONS: A method and software for inferring founder mosaic genotypes in intercross offspring from low-coverage whole-genome sequencing in pedigrees from heterozygous founders are described. They provide high-quality, high-resolution genotypes in a time- and cost-efficient manner. The software is freely available at https://github.com/CarlborgGenomics/Stripes .

Short communication: Adipogenic, metabolic, and apoptotic marker mRNA in cellular fractions of adipose tissue from chickens predisposed to be anorexic or obese.

The body weight-selected lines of chickens are a model for understanding factors that predispose an individual to anorexia or obesity. The high body weight-selected (HWS) individuals are compulsive eaters that become obese whereas the low body weight-selected (LWS) are relatively lean and hypophagic. The objective of this study was to measure gene expression of various preadipocyte, proliferation, metabolic, and apoptotic markers in the stromal-vascular fraction and adipocytes from LWS and HWS adipose tissue. Although preadipocyte and proliferation markers were more highly expressed in the stromal-vascular fraction of LWS than HWS chicks, greater expression of granzyme-A and the presence of more annexin V-positive cells suggests that apoptosis may limit the adipogenic potential of adipocyte precursor cells and represent a novel mechanism that regulates the expansion of adipose tissue. Results provide insights on cellular mechanisms associated with adipose tissue development in the lean and obese state.

Anorexigenic effects of mesotocin in chicks are genetic background-dependent and are associated with changes in the paraventricular nucleus and lateral hypothalamus.

Mesotocin (MT) decreases food intake and induces hyperthermia in chicks although hypothalamic mechanisms are unknown. The purpose of this study was thus to investigate effects of receptor antagonists and MT on feeding behavior and hypothalamic physiology. Intracerebroventricular injection of 2.5 nmol into broiler chicks was associated with decreased food intake for 180 min and water intake from 60 to 180 min. Cloacal temperatures were elevated in chicks injected with 0.156 and 0.625 nmol at 30 and 60 min, and up to 180 min in those injected with 2.5 nmol. MT also increased temperatures and decreased food and water intake in chicks from lines selected for low (LWS) or high (HWS) body weight with a higher dose threshold but longer food intake response in HWS chicks. An oxytocin receptor antagonist prevented MT-mediated changes in food intake but not water intake or temperature. Yohimbine, an α2-adrenergic receptor antagonist, did not affect food intake, temperature, or MT-mediated effects. MT increased c-Fos immunoreactivity in the paraventricular nucleus (PVN) and lateral hypothalamus (LH). Hypothalamic agouti-related peptide, corticotropin-releasing factor receptor sub-type 1, and melanocortin receptor 3 mRNAs increased in response to MT. There was increased MT mRNA in the LH and L-aromatic amino acid decarboxylase mRNA in the PVN of MT-injected chicks. In conclusion, MT induced anorexia and hyperthermia and reduced water intake. MT was associated with activation of the PVN and LH and differences in the mRNA abundance of some appetite-associated factors, thus implicating these nuclei and several signaling pathways in the effects observed.

Artificial Selection Response due to Polygenic Adaptation from a Multilocus, Multiallelic Genetic Architecture.

The ability of a population to adapt to changes in their living conditions, whether in nature or captivity, often depends on polymorphisms in multiple genes across the genome. In-depth studies of such polygenic adaptations are difficult in natural populations, but can be approached using the resources provided by artificial selection experiments. Here, we dissect the genetic mechanisms involved in long-term selection responses of the Virginia chicken lines, populations that after 40 generations of divergent selection for 56-day body weight display a 9-fold difference in the selected trait. In the F15 generation of an intercross between the divergent lines, 20 loci explained >60% of the additive genetic variance for the selected trait. We focused particularly on fine-mapping seven major QTL that replicated in this population and found that only two fine-mapped to single, bi-allelic loci; the other five contained linked loci, multiple alleles or were epistatic. This detailed dissection of the polygenic adaptations in the Virginia lines provides a deeper understanding of the range of different genome-wide mechanisms that have been involved in these long-term selection responses. The results illustrate that the genetic architecture of a highly polygenic trait can involve a broad range of genetic mechanisms, and that this can be the case even in a small population bred from founders with limited genetic diversity.

Growth patterns for three generations of an intercross between red junglefowl and chickens selected for low body weight.

Growth is a complex and dynamic process that may be measured at a specific point or over a period of time. Compared was the growth of male and female chickens over a three-generation period. Involved were red junglefowl (RJF; Gallus gallus), a line of White Plymouth Rock chickens (LWS; Gallus gallus domesticus) selected for low body weight, and their reciprocal F1 and F2 crosses. In both sexes, Gompertz's description of growth showed that RJF had significantly lower asymptotes, earlier inflection points, and faster growth rates than LWS. Heterosis for these measures was positive for asymptote and negative for growth rate and inflection point. The RJF commenced egg production at a significantly younger age and lower body weight than LWS. Although F1 and F2 reciprocal crosses were similar for body weight and for age at first egg, the F1 reciprocal crosses began lay at significantly younger ages than the F2 crosses and parental lines. When viewed on a physiological basis where age and body weight were simultaneously standardized, both parental lines and reciprocal F1 and F2 crosses had differing rapid and lag growth phases. Overall, sexual dimorphism increased in all populations from hatch to sexual maturity. The LWS males had a longer growth period consistent with their female counterparts who became sexually mature at older ages. Comprehensively, these results indicate additive and nonadditive genetic variation for distinct growth patterns and changes in resource allocation strategies over time.

Genome-wide standing variation facilitates long-term response to bidirectional selection for antibody response in chickens.

BACKGROUND: Long-term selection experiments provide a powerful approach to gain empirical insights into adaptation, allowing researchers to uncover the targets of selection and infer their contributions to the mode and tempo of adaptation. Here we implement a pooled genome re-sequencing approach to investigate the consequences of 39 generations of bidirectional selection in White Leghorn chickens on a humoral immune trait: antibody response to sheep red blood cells. RESULTS: We observed wide genome involvement in response to this selection regime. Many genomic regions were highly differentiated resulting from this experimental selection regime, an involvement of up to 20% of the chicken genome (208.8 Mb). While genetic drift has certainly contributed to this, we implement gene ontology, association analysis and population simulations to increase our confidence in candidate selective sweeps. Three strong candidate genes, MHC, SEMA5A and TGFBR2, are also presented. CONCLUSIONS: The extensive genomic changes highlight the polygenic genetic architecture of antibody response in these chicken populations, which are derived from a common founder population, demonstrating the extent of standing immunogenetic variation available at the onset of selection.

Peripheral neuropeptide Y differentially influences adipogenesis and lipolysis in chicks from lines selected for low or high body weight.

Neuropeptide Y (NPY) stimulates appetite and promotes lipid deposition. We demonstrated a differential sensitivity in the food intake response to central NPY in chicks from lines selected for low (LWS) or high (HWS) body weight, but have not reported whether such differences exist in the periphery. At 5days, LWS and HWS chicks were intraperitoneally injected with 0 (vehicle), 60, or 120µg/kg BW NPY and subcutaneous adipose tissue and plasma were collected at 1, 3, 6, 12, and 24h (n=12). NPY injection increased glycerol-3-phosphate dehydrogenase (G3PDH) activity at 1 and 3h and reduced plasma non-esterified fatty acids (NEFAs) at 1 and 12h. G3PDH activity was greater in HWS than LWS while NEFAs were greater in LWS. At 1h, peroxisome proliferator-activated receptor gamma (PPARγ), CCAAT/enhancer binding protein (C/EBP)α, and microsomal triglyceride transfer protein (MTTP) mRNAs were reduced in NPY-injected chicks whereas NPY receptor 1 (NPYR1) was increased. Expression of stearoyl-CoA desaturase (SCD1) was increased by NPY at 1h in HWS but not LWS. PPARγ (3 and 6h), C/EBPß (3h), C/EBPα (6h) and NPYR1 and 2 (24h) mRNAs were greater in NPY- than vehicle-injected chicks. At several times, adipose triglyceride lipase, MTTP, perilipin 1, NPYR1, and NPYR2 mRNAs were greater in LWS than HWS, while expression of SCD1, glycerol-3-phosphate acyltransferase 3 and lipoprotein lipase was greater in HWS than LWS. Thus, NPY promotes fat deposition and inhibits lipolysis in chicks, with line differences indicative of greater rates of lipolysis in LWS and adipogenesis in HWS.

Stimulation of food intake after central administration of gonadotropin-inhibitory hormone is similar in genetically selected low and high body weight lines of chickens.

Gonadotropin-inhibitory hormone (GnIH), first isolated from the brain of the Japanese quail (Coturnix japonica), when centrally administered exerts orexigenic effects in birds. However, the precise mechanisms mediating this effect are poorly understood and limited information is available on this effect in models of body weight dysfunction. Thus, the purpose of the present study was to investigate appetite-associated effects of GnIH in chicks from lines that have been selected for either low or high body weight, and are anorexic or become obese, respectively. Central GnIH injection increased food intake in both lines with a similar magnitude of response. There was no effect on water intake. Hypothalamic GnIH mRNA was greater in the low than high weight lines and was greater in the fasted than fed chicks. GnIH receptor mRNA was similarly expressed in both lines, and was greater in fed than fasted chicks. Thus, although selection for body weight did not alter the effect of GnIH on feeding, fasting increased GnIH mRNA in both lines implying that it is an innate hunger factor.

Hypothalamic differences in expression of genes involved in monoamine synthesis and signaling pathways after insulin injection in chickens from lines selected for high and low body weight.

Long-term selection for juvenile body weight from a common founder population resulted in two divergent chicken lines (low-weight selected line (LWS), high-weight selected line (HWS)) that display distinct food intake and blood glucose responses to exogenous neuropeptides and insulin. The objective of this study was to elucidate putative targets affecting food intake and energy homeostasis by sequencing hypothalamic RNA from LWS and HWS chickens after insulin injection. Ninety-day-old female LWS and HWS chickens were injected with either vehicle or insulin and hypothalamus collected at 1 h postinjection. Through RNA sequencing, a total of 361 differentially expressed genes (DEGs) were identified. There was greater expression of genes, mainly tyrosine hydroxylase (TH), L-aromatic amino acid decarboxylase (DDC), and vesicular monoamine transporter (VMAT), involved in serotonin and dopamine biosynthesis and signaling in LWS than in HWS vehicle-injected chickens. In contrast, after insulin injection, these genes were more highly expressed in HWS than in LWS. We identified 90 single nucleotide polymorphisms (SNPs) existing only in the HWS and 121 SNPs specific to LWS and 5119 SNPs close to fixation (with absolute frequency difference ≥0.9). Four were located in genes encoding enzymes associated with serotonergic and dopaminergic pathways, such as DDC, TH, and solute carrier family 18, member 2 (VMAT). These data implicate differences in biogenic amines such as serotonin and dopamine in hypothalamic physiology between the chicken lines, and these differences might be associated with polymorphisms during long-term selection. Changes in serotonergic and dopaminergic signaling pathways in response to insulin injection suggest a role in whole-body energy homeostasis.

Selection for divergent body size alters rates of embryonic skeletal muscle formation and muscle gene expression patterns.

The impact of divergent selection for body size on embryogenesis is poorly understood. The objective of this experiment was to document skeletal muscle development during embryogenesis in two lines of chickens that display divergent growth as adults. Results reveal that after 54 generations of opposing selection from a common founder population, the embryos from the low weight select (LWS) line develop more rapidly during early embryogenesis than those from the high weight select (HWS) line. Muscle formation during the late embryonic period is more rapid and extensive in the HWS embryo than in the LWS contemporary. Isolated muscle progenitors from embryonic day 10 HWS embryos proliferated more rapidly, forming fibers sooner with a larger size than the LWS cells. The limited myogenic capacity of the LWS progenitor cells is not attributed to altered patterns of expression of Pax7, Pax3 or the myogenic regulatory factor genes. Members of the fibroblast growth factor family are potent mitogens and inhibitors of myoblast differentiation. Transcript abundance of FGF2 and FGF4 was measured in cultures of HWS and LWS progenitors as a function of time. The pattern of expression of FGF4 was similar between HWS and LWS with a large increase between days 1 and 3 followed by a reduction at day 5 of culture. Expression of FGF2 in LWS muscle cells did not change while a significant reduction in FGF2 expression was observed by day 5 in the HWS. Our results indicate that divergent selection for postnatal growth has altered embryonic development.

Delayed access of low body weight-selected chicks to food at hatch is associated with up-regulated pancreatic glucagon and glucose transporter gene expression.

Chickens selected for low (LWS) and high (HWS) juvenile body weight (BW) for 55 generations differ in BW by 10-fold at selection age. High (HWR) and low (LWR) body weight-relaxed lines have been random-bred since the 46th generation. Our objective was to evaluate the developmental and nutritional regulation of pancreatic mRNA abundance of pancreatic and duodenal homeobox 1 (PDX1), preproinsulin (PPI), preproglucagon (PPG), and glucose transporter 2 (GLUT2). At day of hatch (DOH) and days 1, 3, 7, and 15 (D1, 3, 7 and 15, respectively), pancreas was collected and real time PCR was performed in Experiment 1. In Experiment 2, HWS and LWS were fed or delayed access to food for 72 h post-hatch, and pancreas collected at D15. There was an interaction of line and age for GLUT2 (P=0.001), PPI (P<0.0001), PPG (P=0.034), and PDX1 (P<0.0001). Expression was greater in chicks from LWR and LWS than HWR and HWS. There was an interaction of line and nutrition on PPG (P<0.0001) and GLUT2 (P=0.001) mRNA, where expression was similar among chicks that were fed but greater in LWS than HWS when chicks were delayed access to food. Thus, the first two weeks is important for maturation of pancreatic endocrine function. Long-term selection for BW is associated with differences in pancreas development, and delaying access to food at hatch may have persisting effects on glucose regulatory function.

Housing systems interacting with sex and genetic line affect broiler growth and carcass traits.

Housing systems used in the production of poultry meat vary worldwide dependent on climate, land availability, and other resources essential for production. Reported here are comparisons between pen and cage rearing (the housing system, denoted HS: ), line crosses LC: ), two native Chinese lines (EM males were mated to Y1 and Y2 and their offspring denoted as EMY1 and EMY2), and sex in determining broiler traits. At hatch, 320 males and 320 females from each LC (giving a total of 1,280 chicks) were randomly assigned within each subgroup to 16 battery pens. There were 4 replicates for each combination of LC by sex. On d 28, half of the chicks were transferred to indoor floor pens, and the others were raised in single cages from d 29 to 91. Weekly body weights, livability, and feed conversion ratios ( FCR: ) were obtained to d 91, the age at which the broilers were slaughtered for carcass measurements. The caged males and females were heavier (P < 0.05) than their penned counterparts (2,292 vs 2,219 g). Except for females from line EMY1 (94.9%), the livability for each unit from 1 to 28 d, and 29 to 91 d was greater than 95%. Penned EMY2 broilers had the highest FCR (3.02), whereas penned EMY1 broilers had the lowest FCR (2.96) among the housing systems by LC combinations (P < 0.05). Caged chickens had thicker subcutaneous fat (7.24 mm), a higher percentages of abdominal fat (5.01%) and liver mass (3.13%) , but lower eviscerated carcass (60.63%) and breast muscle weights (pectoralis major and minor, 17.10%). Males were heavier and had higher percentages of leg muscle (boneless drum plus thigh, 24.22%) and heart muscle (1.08%) than the females (P < 0.05). However, the females had thicker subcutaneous fat (7.19 mm) and higher percentages of carcass weight (87.28%), breast muscle (18.11%), abdominal fat (6.54%), and liver mass (3.15%) than males. Penned females had the highest percentage of breast muscle (18.94%), and caged females had the highest percentage of liver mass (3.72%). Females of EMY1 had the highest percentage of breast muscle (18.40%). Generally, the housing system employed and the sex of the broilers greatly affect the carcass traits.

Insulin-induced hypoglycemia associations with gene expression changes in liver and hypothalamus of chickens from lines selected for low or high body weight.

Chickens selected for low (LWS) or high (HWS) body weight for more than 56 generations now have a 10-fold difference in body weight at 56 days of age and correlated responses in appetite and glucose regulation. The LWS chickens are lean and some are anorexic, while the HWS are compulsive feeders and have a different threshold sensitivity of food intake and blood glucose to both central and peripheral insulin, respectively. We previously demonstrated that at 90-days of age, insulin-induced hypoglycemia was associated with reduced glucose transporter expression in the liver of both lines, and differences in expression of neuropeptide Y (NPY) and NPY receptor sub-type genes between LWS and HWS in the hypothalamus. The objective of this study was to determine effects of insulin-induced hypoglycemia on gene expression in the hypothalamus and liver of early post-hatch LWS and HWS chicks. On day 5 post-hatch chicks from each line were fasted for 3h and injected intraperitoneally with insulin or vehicle. At 1h post-injection, chicks were euthanized, blood glucose was measured, and hypothalamus and liver were removed. Total RNA was isolated and real time PCR performed. Insulin injection was associated with a more pronounced reduction in blood glucose in HWS compared with LWS chicks (two-way interaction; P<0.05). Aromatic L-amino acid decarboxylase, NPY, and NPY receptor sub-types 2 and 5 mRNA quantities were greater in LWS than HWS chicks in the hypothalamus (P<0.05), whereas pro-opiomelanocortin mRNA was greater in the hypothalamus of HWS than LWS (P<0.05). In the liver, glucose transporter 1, 2 and 3 (GLUT 1, 2 and 3, respectively) mRNA abundance was greater in HWS than LWS chicks (P<0.05). Compared to the vehicle, insulin treatment was associated with an increase in tryptophan hydroxylase 2 mRNA in the hypothalamus of both lines (P=0.02). In the liver of both lines, insulin treatment was associated with decreased (P=0.01) GLUT2 mRNA and increased (P=0.01) GLUT1 mRNA, compared to vehicle-treated chicks. Results suggest that NPY-associated factors and glucose transporters are differentially-expressed between LWS and HWS chickens and that HWS chicks display greater sensitivity to exogenous insulin during the early post-hatch period.

Replication and explorations of high-order epistasis using a large advanced intercross line pedigree.

Dissection of the genetic architecture of complex traits persists as a major challenge in biology; despite considerable efforts, much remains unclear including the role and importance of genetic interactions. This study provides empirical evidence for a strong and persistent contribution of both second- and third-order epistatic interactions to long-term selection response for body weight in two divergently selected chicken lines. We earlier reported a network of interacting loci with large effects on body weight in an F(2) intercross between these high- and low-body weight lines. Here, most pair-wise interactions in the network are replicated in an independent eight-generation advanced intercross line (AIL). The original report showed an important contribution of capacitating epistasis to growth, meaning that the genotype at a hub in the network releases the effects of one or several peripheral loci. After fine-mapping of the loci in the AIL, we show that these interactions were persistent over time. The replication of five of six originally reported epistatic loci, as well as the capacitating epistasis, provides strong empirical evidence that the originally observed epistasis is of biological importance and is a contributor in the genetic architecture of this population. The stability of genetic interaction mechanisms over time indicates a non-transient role of epistasis on phenotypic change. Third-order epistasis was for the first time examined in this study and was shown to make an important contribution to growth, which suggests that the genetic architecture of growth is more complex than can be explained by two-locus interactions only. Our results illustrate the importance of designing studies that facilitate exploration of epistasis in populations for obtaining a comprehensive understanding of the genetics underlying a complex trait.

Delayed feeding after hatch caused compensatory increases in blood glucose concentration in fed chicks from low but not high body weight lines.

This experiment used 2 lines of chickens that have been selected 54 generations for either low (LWS) or high (HWS) 8-wk BW from the same founder population, sublines (HWR and LWR) in which selection was relaxed in generation 43 in the selected lines, and crosses (HL and LH) made from generation 54 of HWS and LWS. For 8-wk BW, the difference between lines LWS and HWS in generation 54 was approximately 10-fold, whereas for the relaxed contemporary lines they were approximately 7-fold. Three trials were designed to measure developmental, nutritional, and genetic aspects of blood glucose homeostasis during the first 2 wk posthatch. In trial 1, we measured BW, whole blood glucose (BG), and weights (relative to BW) of liver, pancreas, and yolk sac of chicks fed from day of hatch to d 15. In trial 2, we compared those traits in chicks feed-delayed 72 h posthatch and in chicks without feed delay. In trial 3, we evaluated the effect of a 16-h fast on BW and BG on d 3, 8, and 15. There were higher levels of BG in HWS than LWS, and males than females in the fed state. Delayed access to feed for 72 h after hatch was associated with a dramatic reduction in BG. Feeding triggered a compensatory response whereby LWS displayed greater BG but smaller pancreases (% BW; d 15), compared with the controls. There were maternal effects for BW in both fed and fasted states and the reciprocal crosses exhibited heterosis for BG in the fasted state. These results show that chickens selected for high or low BW differ in BG regulation during the early posthatch period.

Growth of White Leghorn Chicken Immune Organs after Long-Term Divergent Selection for High or Low Antibody Response to Sheep Red Blood Cells.

Long-term divergent selection from a common founder population for a single trait-antibody response to sheep erythrocytes 5 days post-injection-has resulted in two distinct lines of White Leghorn chickens with a well-documented difference in antibody titers: high (HAS)- and low (LAS)-antibody selected lines. Subpopulations-high (HAR)- and low (LAR)-antibody relaxed-were developed from generation 24 of the selected lines to relax selection. The objective of the current experiment was to determine if this long-term selection and relaxation of selection impacted the growth of two organs important to chicken immunity: the spleen and the bursa of Fabricius. Spleens and bursae were obtained from ten chickens per line at nine timepoints (E18, D0, D6, D13, D20, D35, D49, D63, and D91) throughout their rapid growth phase and presented as a percent of body weight. Significance was set at p ≤ 0.05. For the spleen, all lines consistently increased in size relative to body weight to D49, followed by a consistent decline. All lines had a similar growth pattern, but HAS spleens grew faster than LAS spleens. For the bursa, LAS was smaller than the other three lines as an embryo and also smaller than HAS through D63. In the selected lines, bursa weight peaked at D35, whereas the relaxed lines peaked at D49. By D91, there was no difference between lines. Artificial and natural selection, represented by the long-term selected and relaxed antibody lines, resulted in differences in the growth patterns and relative weights of the spleen and bursa of Fabricius.

Complex genetic architecture of the chicken Growth1 QTL region.

The genetic complexity of polygenic traits represents a captivating and intricate facet of biological inheritance. Unlike Mendelian traits controlled by a single gene, polygenic traits are influenced by multiple genetic loci, each exerting a modest effect on the trait. This cumulative impact of numerous genes, interactions among them, environmental factors, and epigenetic modifications results in a multifaceted architecture of genetic contributions to complex traits. Given the well-characterized genome, diverse traits, and range of genetic resources, chicken (Gallus gallus) was employed as a model organism to dissect the intricate genetic makeup of a previously identified major Quantitative Trait Loci (QTL) for body weight on chromosome 1. A multigenerational advanced intercross line (AIL) of 3215 chickens whose genomes had been sequenced to an average of 0.4x was analyzed using genome-wide association study (GWAS) and variance-heterogeneity GWAS (vGWAS) to identify markers associated with 8-week body weight. Additionally, epistatic interactions were studied using the natural and orthogonal interaction (NOIA) model. Six genetic modules, two from GWAS and four from vGWAS, were strongly associated with the studied trait. We found evidence of both additive- and non-additive interactions between these modules and constructed a putative local epistasis network for the region. Our screens for functional alleles revealed a missense variant in the gene ribonuclease H2 subunit B (RNASEH2B), which has previously been associated with growth-related traits in chickens and Darwin's finches. In addition, one of the most strongly associated SNPs identified is located in a non-coding region upstream of the long non-coding RNA, ENSGALG00000053256, previously suggested as a candidate gene for regulating chicken body weight. By studying large numbers of individuals from a family material using approaches to capture both additive and non-additive effects, this study advances our understanding of genetic complexities in a highly polygenic trait and has practical implications for poultry breeding and agriculture.