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The Taihe Black-Bone Silky Fowl ( Gallus gallus domesticus Brisson ) possesses significant value in terms of consumption, medicinal applications, and ornamental appeal, representing a precious genetic resource and traditional Chinese medicinal material. However, considerable variation exists within populations regarding egg-laying performance. This study integrates a whole-genome selection signal analysis (SSA) with a transcriptome analysis to identify genes associated with egg-laying traits in Taihe Black-Bone Silky Fowls. We identified 31 candidate genes under selection from the high-yield chicken (HC) and low-yield chicken (LC) groups. Additionally, through RNA-seq analysis, 257 common differentially expressed genes (DEGs) were identified from four comparative groups. Two overlapping genes- LPL and SETBP1 -were found in both the selected gene and DEG lists. These selected genes and DEGs were enriched in pathways related to ovarian development, including the lysosome pathway, the ECM-receptor interaction pathway, the TGF-beta signaling pathway, the Wnt signaling pathway, the PPAR signaling pathway, and the glycerolipid metabolism pathway. These research findings contribute to the breeding of Taihe Black-Bone Silky Fowls with high egg production traits and provide a theoretical foundation for exploring the regulatory mechanisms of avian reproduction.

Egg-laying performance is of great economic importance in poultry, but the underlying genetic mechanisms are still elusive. In this work, we conduct a multi-omics and multi-tissue integrative study in hens with distinct egg production, to detect the hub candidate genes and construct hub molecular networks contributing to egg-laying phenotypic differences. We identifiy three hub candidate genes as egg-laying facilitators: TFPI2, which promotes the GnRH secretion in hypothalamic neuron cells; CAMK2D, which promotes the FSHβ and LHβ secretion in pituitary cells; and OSTN, which promotes granulosa cell proliferation and the synthesis of sex steroid hormones. We reveal key endocrine factors involving egg production by inter-tissue crosstalk analysis, and demonstrate that both a hepatokine, APOA4, and an adipokine, ANGPTL2, could increase egg production by inter-tissue communication with hypothalamic-pituitary-ovarian axis. Together, These results reveal the molecular mechanisms of multi-tissue coordinative regulation of chicken egg-laying performance and provide key insights to avian reproductive regulation., (© 2024. The Author(s).)

The age of first egg (AFE) in chicken can affect early and even life-time egg production performance to some extent, and therefore is an important economic trait that affects production efficiency. To better understand the genetic patterns of AFE and other production traits including body weight at first egg (BWA), first egg weight (FEW), and total egg number from AFE to 58 wk of age (total-EN), we recorded the production performance of 2 widely used layer breeds, white leghorn (WL) and Rhode Island Red (RIR) and estimated genetic parameters based on pedigree and production data. The results showed that the heritability of AFE in both breeds ranged from 0.4 to 0.6, and AFE showed strong positive genetic and phenotypic correlations to BWA as well as FEW, while showing strong negative genetic and phenotypic correlations with total-EN. Furtherly, by genome-wide association analysis study (GWAS), we identified 12 and 26 significant SNPs to be related to AFE in the 2-layer breeds, respectively. A total of 18 genes were identified that could affect AFE based on the significant SNP annotations obtained, but there were no gene overlapped in the 2 breeds indicating the genetic foundation of AFE could differ from breed to breed. Our results provided a deeper understanding of genetic patterns and molecular basement of AFE in different breeds and could help in the selection of egg production traits., (Copyright © 2024. Published by Elsevier Inc.)

Compared to high-yield commercial laying hens, Chinese indigenous chicken breeds have poor egg laying capacity due to the lack of intensive selection. However, as these breeds have not undergone systematic selection, it is possible that there is a greater abundance of genetic variations related to egg laying traits. In this study, we assessed 5 egg number (EN) traits at different stages of the egg-laying period: EN1 (from the first egg to 23 wk), EN2 (from 23 to 35 wk), EN3 (from 35 to 48 wk), EN4 (from the first egg to 35 wk), and EN5 (from the first egg to 48 wk). To investigate the molecular mechanisms underlying egg number traits in a Chinese local chicken breed, we conducted a genome-wide association study (GWAS) using data from whole-genome sequencing (WGS) of 399 Laiwu Black chickens. We obtained a total of 3.01 Tb of raw data with an average depth of 7.07 × per individual. A total of 86 genome-wide suggestive or significant single-nucleotide polymorphisms (SNP) contained within a set of 45 corresponding candidate genes were identified and found to be associated with stages EN1-EN5. The genes vitellogenin 2 (VTG2), lipase maturation factor 1 (LMF1), calcium voltage-gated channel auxiliary subunit alpha2delta 3 (CACNA2D3), poly(A) binding protein cytoplasmic 1 (PABPC1), programmed cell death 11 (PDCD11) and family with sequence similarity 213 member A (FAM213A) can be considered as the candidate genes associated with egg number traits, due to their reported association with animal reproduction traits. Noteworthy, results suggests that VTG2 and PDCD11 are not only involved in the regulation of EN3, but also in the regulation of EN5, implies that VTG2 and PDCD11 have a significant influence on egg production traits. Our study offers valuable genomic insights into the molecular genetic mechanisms that govern egg number traits in a Chinese indigenous egg-laying chicken breed. These findings have the potential to enhance the egg-laying performance of chickens., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Resilience is the capacity of an animal to be minimally affected by disturbances or to rapidly return to its initial state before exposure to a disturbance. Resilient livestock are desired because of their improved health and increased economic profit. Genetic improvement of resilience may also lead to trade-offs with production traits. Recently, resilience indicators based on longitudinal data have been suggested, but they need further evaluation to determine whether they are indeed predictive of improved resilience, such as disease resilience. This study investigated different resilience indicators based on deviations between expected and observed egg production (EP) by exploring their genetic parameters, their possible trade-offs with production traits, and their relationships with antibody traits in chickens., Methods: Egg production in a nucleus breeding herd environment based on 1-week-, 2-week-, or 3-week-intervals of two purebred chicken lines, a white egg-laying (33,825 chickens) and a brown egg-laying line (34,397 chickens), were used to determine deviations between observed EP and expected average batch EP, and between observed EP and expected individual EP. These deviations were used to calculate three types of resilience indicators for two life periods of each individual: natural logarithm-transformed variance (ln(variance)), skewness, and lag-one autocorrelation (autocorrelation) of deviations from 25 to 83 weeks of age and from 83 weeks of age to end of life. Then, we estimated their genetic correlations with EP traits and with two antibody traits., Results: The most promising resilience indicators were those based on 1-week-intervals, as they had the highest heritability estimates (0.02-0.12) and high genetic correlations (above 0.60) with the same resilience indicators based on longer intervals. The three types of resilience indicators differed genetically from each other, which indicates that they possibly capture different aspects of resilience. Genetic correlations of the resilience indicator traits based on 1-week-intervals with EP traits were favorable or zero, which means that trade-off effects were marginal. The resilience indicator traits based on 1-week-intervals also showed no genetic correlations with the antibody traits, which suggests that they are not informative for improved immunity or vice versa in the nucleus environment., Conclusions: This paper gives direction towards the evaluation and implementation of resilience indicators, i.e. to further investigate resilience indicator traits based on 1-week-intervals, in breeding programs for selecting genetically more resilient layer chickens., (© 2024. The Author(s).)

Background: Egg-laying performance is economically important in poultry breeding programs. Crossbreeding between indigenous and elite commercial lines to exploit heterosis has been an upward trend in traditional layer breeding for niche markets. The objective of this study was to analyse the genetic background and to estimate the heterosis of longitudinal egg-laying traits in reciprocal crosses between an indigenous Beijing-You and an elite commercial White Leghorn layer line. Egg weights were measured for the first three eggs, monthly from 28 to 76 weeks of age, and at 86 and 100 weeks of age. Egg quality traits were measured at 32, 54, 72, 86, and 100 weeks of age. Egg production traits were measured from the start of lay until 43, 72, and 100 weeks of age. Heritabilities and phenotypic and genetic correlations were estimated. Heterosis was estimated as the percentage difference of performance of a crossbred from that of the parental average. Reciprocal cross differences were estimated as the difference between the reciprocal crossbreds as a percentage of the parental average., Results: Estimates of heritability of egg weights ranged from 0.29 to 0.75. Estimates of genetic correlations between egg weights at different ages ranged from 0.72 to 1.00. Estimates of heritability for cumulative egg numbers until 43, 72, and 100 weeks of age were around 0.15. Estimates of heterosis for egg weight and cumulative egg number increased with age, ranging from 1.0 to 9.0% and from 1.4 to 11.6%, respectively. From 72 to 100 weeks of age, crossbreds produced more eggs per week than the superior parent White Leghorn (3.5 eggs for White Leghorn, 3.8 and 3.9 eggs for crossbreds). Heterosis for eggshell thickness ranged from 2.7 to 6.6% when using Beijing-You as the sire breed. No significant difference between reciprocal crosses was observed for the investigated traits, except for eggshell strength at 54 weeks of age., Conclusions: The heterosis was substantial for egg weight and cumulative egg number, and increased with age, suggesting that non-additive genetic effects are important in crossbreds between the indigenous and elite breeds. Generally, the crossbreds performed similar to or even outperformed the commercial White Leghorns for egg production persistency., (© 2023. The Author(s).)

Egg products from indigenous chickens have growing market shares as consumers are pursuing differentiation in egg consumption. The genetic improvement in egg production performance of those breeds is crucial for increasing the economic profit. This study aimed to estimate genetic parameters for egg production and clutch-related traits in indigenous Beijing-You chickens for understanding the genetic architecture and exploring proper biological traits for selection. Data on traits including age at first egg (AFE), egg number (EN), average clutch length (ACL), maximum clutch length (MCL), number of clutches (NC) and pauses (NP), and average pause length (APL) were collected from 4 generations of purebred Beijing-You chickens based on the 43-wk and 66-wk of individual egg production record. The heritabilities, genetic and phenotypic correlations were analyzed by the DMU software with the restricted maximum likelihood method in a multivariate animal model. The results showed that the AFE of Beijing-You chickens was 174.45 d of age, and its heritability was as high as 0.62. The heritability was 0.26 for EN43 and 0.18 for EN66. The clutch traits including ACL, MCL, NC, and NP were moderate to high heritable (h 2  = 0.15-0.39), but APL was very low heritable (h 2  = 0.05). Genetic correlations were high between AFE and EN (r G(AFE, EN43)  = -0.79, r G(AFE, EN66)  = -0.39), whereas low between AFE and ACL (r G(AFE, ACL43)  = -0.08, r G(AFE, ACL66)  = 0.01) and MCL (r G(AFE, MCL)  = -0.07). EN had higher correlations with ACL (r G(EN43, ACL43)  = 0.59, r G(EN66, ACL66)  = 0.40) than that with MCL (r G(EN43, MCL43)  = 0.56, r G(EN66, MCL66)  = 0.32). The heritability for ACL43 (h 2  = 0.38) was higher than that for MCL43 (h 2  = 0.33). ACL43 had a positive correlation with EN66 (r G(ACL43, EN66)  = 0.62). These results indicated that the egg production of whole laying period could be improved by early selection for AFE and ACL at the same time in Beijing-You chickens., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Matrix metalloproteinases are a group of proteases involved in the regulation of ovarian follicular development and ovulation. Among the different MMPs, MMP13 is known to play an important role in reproduction. Therefore, this study aimed to screen the molecular genetic markers of the MMP13 gene that affect the egg-laying performance of Chahua chickens. Polymerase chain reaction (PCR) and sequencing were performed in the 5' regulation region of the MMP13 gene to detect loci significantly related to the egg-laying performance of Chahua chickens. A double fluorescence reporting system, quantitative reverse transcription PCR (RT-qPCR), and Western blotting were used to study whether gene expression was regulated by identified sites, providing a theoretical basis to improve egg production in Chahua chickens. The results revealed six single nucleotide polymorphisms (SNPs; A-1887T, T-1889C, A-1890T, T-2252C, T-2329C, and C-2360A) in the promoter region of the MMP13 gene. Further analysis revealed that hens with T -1890 -C -1889 -T -1887 /T -1890 -C -1889 -T -1887 (mutant type, MT) had an earlier age at first egg (AFE) than hens with A -1890 -T -1889 -A -1887 /A -1890 -T -1889 -A -1887 (wild type, WT; p < 0.05). RT-qPCR showed that the relative expression level of the MMP13 gene in the ovarian tissues of individuals with the mutation was higher than that of individuals with the wild gene ( p < 0.05). Western blot results confirmed higher levels of the MMP13 protein in MT ovaries compared to those in WT ovaries. Thus, this study suggests that mutation sites on the MMP13 promoter may affect gene expression. In conclusion, the MMP13 gene in Chahua chickens may be significant for egg-laying performance, and the polymorphism in its promoter region could be used as a molecular marker to improve egg-laying performance.

Background: Floor eggs, which are defined as eggs that hens lay off-nest, are a major issue in cage-free layer poultry systems. They create additional work for farmers because they must be collected by hand. They are also usually soiled or broken, which results in economic losses. Nonetheless, knowledge about the genetics of nesting behavior is limited. The aim of this study was to estimate genetic parameters for traits related to nest preference for laying and to time spent in the nests used for laying (laying duration)., Methods: Two pure lines of laying hens were studied: 927 Rhode Island Red and 980 White Leghorn. Electronic nests were used to record the nesting behavior of these hens in floor pens from 24 to 64 weeks of age. Nest preference was studied based on the mean distance between nests used for laying and the percentage of nests used for laying. Laying duration was studied based on mean laying duration, mean duration in the nest before laying, and mean duration in the nest after laying. Genetic parameters were estimated for each line using a restricted maximum-likelihood method applied to a pedigree-based multi-trait animal model., Results: Estimates of genetic parameters were similar for the two lines. Estimates of heritability ranged from 0.18 to 0.37 for nest preference traits and from 0.54 to 0.70 for laying duration traits. Estimates of genetic correlations of these traits with clutch number or mean oviposition time were favorable. Positive genetic correlations were estimated between nest preference and laying rate in the nests or nest acceptance for laying (+ 0.06 to + 0.37)., Conclusions: These results show that genetics influences traits related to nest preference and laying duration. Selecting hens that have no preference for particular nests and spend little time laying in the nests could help optimize nest use, reduce their occupation rate, and thus decrease the incidence of floor eggs in cage-free systems. Genetic correlations of these traits with other traits of interest related to hen welfare and egg quality have yet to be estimated., (© 2023. The Author(s).)

The number of egg-laying is an important indicator of reproduction performance in poultry breeding. To investigate the relationship between the function of Angiotensin-converting enzyme (ACE) and egg-laying performance of Taihang chicken, the mRNA and protein expression and single nucleotide polymorphism (SNP) of ACE were detected. Analysis of ACE bioinformatics and association analysis of polymorphisms were then performed. The polymorphisms analysis of ACE showed that three SNP loci (g.5066812A>C, g.5080076G>A, and g.5072728A>G) were detected in 800 Taihang chickens with egg-laying records. Association analysis of egg-laying found that ACE g.5066812A>C mutation was significantly associated with the egg-laying performance of Taihang chickens (P < 0.05), and the individuals with the g.5066812A>C mutation showed significantly increasing egg-laying. The mRNA expression was significantly higher in individuals with the AA genotype mutation than those with the AC and CC genotypes (P < 0.01), and the expression of ACE protein levels was consistent with the mRNA expression. Bioinformatics analysis indicated that these mutations affected the secondary and tertiary structure of ACE. This study provides new insights into ACE affecting chicken egg production and some basis for improving the egg production rate of Taihang chickens., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Duration of fertility (DF) is an important economic trait in poultry production because it has a strong effect on chick output. Various criteria or traits to assess DF on individual hens have been reported but they are affected by many nongenetic factors. Thus, a reliable definition and associated genetic parameters are needed. Because egg production is also vital in chicken breeding, knowledge of the relationship between DF and laying performance is needed for designing selection programs., Methods: We used five traits that consider both fertility and embryonic livability to delineate DF. Phenotypic and genetic analyses were completed for 2094 hens, with measurements of DF at 35 and 60 weeks of age and hatching egg production at 400 days of age (HEP400). The selection differentials for DF and HEP400 were evaluated., Results: DF is largely independent of the number of oviposited eggs in the peak laying period but both egg production and DF naturally decline with age. The heritability of the five DF traits ranged from 0.11 to 0.13 at 35 weeks of age and increased slightly in the later laying period, ranging from 0.14 to 0.17 (except for efficient duration, time between insemination and the first unhatched egg). Estimates of the genetic correlation for a given trait measured at the two ages were moderate (0.37-0.44), except for efficient duration. However, number of viable embryos depends strongly on egg production. Estimates of genetic correlations of fertility duration day (FDD) at both ages with HEP were weak. Selection for FDD improved DF but without a significant change in laying performance. Selection for increased HEP400 did not contribute to DF improvement., Conclusions: Although estimates of heritability of the five traits related to DF were low, selection to improve DF based on any one of them is possible. Among these, FDD is an effective selection criterion when the eggs are collected for approximately two weeks after insemination. The best selection procedure for DF improvement would involve multiple measurements at various ages. FDD is independent of laying performance and can be incorporated into a breeding program with egg production to improve reproductive efficiency., (© 2022. The Author(s).)

The BG line was originated from the cross between 2 Chinese indigenous breeds, Dongxiang blue eggshell, and Jiangshan black-bone, and has been bred to combine dark heavy black-bone body and high production of blue-shell eggs, into single dual-purpose line. Full-pedigree hens from 2 generations, G4 (n = 441) and G5 (n = 464), were reared in the same single-cage laying facility in 2019-2020 and 2020-2021, respectively. Starting from the first egg of each hen, its daily egg production was recorded until 300 days-of-age. Up to 7 "no-egg" days were considered normal laying breaks between clutches, whereas laying cessation of 8 or more days was considered Pause, and the laying pattern of each hen was assigned either with Pause or No-Pause. The other traits included PsDays: number of Pause days; AFE: age at first egg; EN300: eggs laid until 300 d; %L300: total laying rate (EN300/[300-AFE]); %Lnet: net laying rate (EN300/[300-AFE-PsDays]); ClLng: average clutch length; EW200 and EW300: average egg weight around 200 d and 300 d. Estimates of heritability (h 2 ) of each trait, and phenotypic and genetic correlation between traits, were calculated in each generation using the animal model. Heritability estimates were calculated also by regressing the means of full-sib G5 hens on their G4 parents' means. Mean overall laying rate of all G4 hens was low (%L300 = 57%) because 53% of them had Pause in their laying pattern. In G5, incidence of Pause was higher (75%) due to a 3-wk cold stress, with mean %L300 = 54%. However, significant estimates of heritability and genetic correlations suggest that selection for low PsDays will reduce the incidence of Pause in BG hens and elevate the line's mean laying rate towards %L300 = 70%, as the No-Pause hens in G5. PsDays-free laying rate (%Lnet) was found to be highly correlated with the significantly heritable (h 2 ≈0.4) clutch length (ClLng). Selection index combining the genetically independent low PsDays and high ClLng is expected to maximize egg production improvement in the BG line, and in similar populations derived from indigenous breeds., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Egg production is an essential indicator of poultry fertility. The ovary is a crucial organ involved in egg production; however, little is known about the key genes and signaling pathways involved in the whole egg-laying cycle of hens. In order to explore the mechanism of egg production at different stages of the egg-laying process, ovarian tissues from four chickens were randomly selected for transcriptome analysis at each of the three ages (145 d, 204 d, and 300 d in the early, peak, and late stages of egg laying). A total of 12 gene libraries were constructed, and a total of 8433 differential genes were identified from NH145d vs. NH204d, NH145d vs. NH300d and NH300d vs. NH204d (Ninghai 145-day-old, Ninghai 204-day-old, and Ninghai 300-day-old), with 1176, 1653 and 1868 up-regulated genes, and 621, 1955 and 1160 down-regulated genes, respectively. In each of the two comparison groups, 73, 1004, and 1030 differentially expressed genes were found to be co-expressed. We analyzed the differentially expressed genes and predicted nine genes involved in egg production regulation, including LRP8, BMP6, ZP4, COL4A1, VCAN, INHBA, LOX, PTX3 , and IHH , as well as several essential egg production pathways, such as regulation adhesion molecules (CAMs), calcium signaling pathways, neuroactive ligand-receptor interaction, and cytokine-cytokine receptor interaction. Transcriptional analysis of the chicken ovary during different phases of egg-lay will provide a useful molecular basis for study of the development of the egg-laying ovary.

The cuticle formed in the uterus is the outermost layer as the first defense line of eggshell against microbial invasions in most avian species, and analyzing its genetic regulation and influencing factors are of great importance to egg biosecurity in poultry production worldwide. The current study compared the uterine transcriptome and proteome of laying hens producing eggs with good and poor cuticle quality (GC and PC, the top and tail of the cuticle quality distribution), and identified several genes involved with eggshell cuticle quality (ESCQ). Overall, transcriptomic analysis identified 53 differentially expressed genes (DEGs) between PC versus GC group hens, among which 25 were up-regulated and 28 were down-regulated. No differences were found in the uterine proteome. Several DEGs, including PTGDS, PLCG2, ADM and PRLR related to uterine functions and reproductive hormones, were validated by qPCR analysis. Egg quality measurements between GC and PC hens showed GC hens had longer laying interval between two consecutive ovipositions (25.64 ± 1.23 vs 24.94 ± 1.12 h) and thicker eggshell thickness (352.01 ± 23.04 vs 316.20 ± 30.58 μm) (P < 0.05). Apart from eggshell traits, other egg quality traits didn't differ. The result demonstrated eggshell and cuticle deposition duration in the uterus is one of the major factors affecting ESCQ in laying hens. PTGDS, PLCG2, ADM and PRLR genes were discovered and might play crucial roles in cuticle deposition by regulating the uterine muscular activities and secretion function. The findings in the present study provide new insights into the genetic regulation of cuticle deposition in laying hens and establish a foundation for further investigations., (© 2021. The Author(s).)

Background: As cage-free production systems become increasingly popular, behavioral traits such as nesting behavior and temperament have become more important. The objective of this study was to estimate heritabilities for frequency of perching and proportion of floor eggs and their genetic correlation in two Rhode Island Red lines., Results: The percent of hens observed perching tended to increase and the proportion of eggs laid on the floor tended to decrease as the test progressed. This suggests the ability of hens to learn to use nests and perches. Under the bivariate repeatability model, estimates of heritability in the two lines were 0.22 ± 0.04 and 0.07 ± 0.05 for the percent of hens perching, and 0.52 ± 0.05 and 0.45 ± 0.05 for the percent of floor eggs. Estimates of the genetic correlation between perching and floor eggs were - 0.26 ± 0.14 and - 0.19 ± 0.27 for the two lines, suggesting that, genetically, there was some tendency for hens that better use perches to also use nests; but the phenotypic correlation was close to zero. Random regression models indicated the presence of a genetic component for learning ability., Conclusions: In conclusion, perching and tendency to lay floor eggs were shown to be a learned behavior, which stresses the importance of proper management and training of pullets and young hens. A significant genetic component was found, confirming the possibility to improve nesting behavior for cage-free systems through genetic selection.

Ovomucoid is a major egg white protein which is considered as the most dominant allergen in chicken eggs. Owing to the difficulty of separating ovomucoid from egg whites, researchers have adopted genetic deletion for development of hypoallergenic eggs. Previously, we used CRISPR/Cas9 to establish chickens with ovomucoid gene (OVM) mutations, but it remained unknown whether such hens could produce eggs at maturity. Here, we have reported on eggs laid by OVM-targeted hens. Except for watery egg whites, the eggs had no evident abnormalities. Real-time PCR revealed alternative splicing of OVM mRNA in hens, but their expression was limited. Immunoblotting detected neither mature ovomucoid nor ovomucoid-truncated splicing variants in egg whites. Sixteen chicks hatched from 28 fertilized eggs laid by OVM-targeted hens, and fourteen of the sixteen chicks demonstrated healthy growth. Taken together, our results demonstrated that OVM knockout could almost completely eliminate ovomucoid from eggs, without abolishing fertility. Thus, the eggs developed in this study have potential as a hypoallergenic food source for most patients with egg allergies., (Copyright © 2020. Published by Elsevier Inc.)

The molecular mechanisms underlying reproductive aging in avian species are poorly understood. Previous studies have shown the importance of mechanistic target of rapamycin (mTOR) signaling pathway in aging. In this study, we investigated the relationship between the mTOR signaling pathway and ovarian aging in the peak phase and late phase of egg production in laying hens. The egg production rate and egg quality were tracked for 5 consecutive weeks in 30-week-old and 70-week-old Dawu Jinfeng hens (N = 30/group). During the peak phase (week 35) and late phase (week 75), antioxidant and immune indicators were detected by enzyme-linked immunosorbent assay, and mTOR signaling-related genes (CLIP-170, GRB10, LIPIN-1, ATG1, 4E-BP1, S6K, PKC, RHO, and SGK1) were detected in the follicles by quantitative reverse transcription-PCR technology. The protein expression of key genes (mTOR, PKC, 4EBP1) was evaluated by Western blot analysis. The egg production rate and the antioxidant indexes superoxide dismutase and glutathione peroxidase and the levels of total antioxidant capacity and immunoglobulins (IgM and IgG) were significantly higher at week 35 than those at week 75 (P < 0.01), while malondialdehyde levels were significantly lower (P < 0.01). At week 75, there were fewer follicles in the different stages of development than were detected at week 35. The number of white follicles (large and small) and primary follicles were significantly higher at week 75 than those detected at week 35 (P < 0.01). The mRNA expression of avTOR, CLIP-170, GRB10, LIPIN-1, 4E-BP1, S6K, RHO, and SGK genes in small white follicles (SWF), large white follicles (LWF), F3, F1, and ovary at week 75 was lower than that in the hens at week 35 (P < 0.05). The mRNA expression in small yellow follicle (SYF) was significantly higher than that at week 35 (P < 0.05), while the mRNA expression of ULK1 in SWF, LWF, F3, F1, and ovary at week 75 was higher than that of hens at week 35 (P＜0.01), and SYF was lower (P < 0.05). Treatment of chicken granulosa cells with the mTOR agonist MHY1485 significantly enhanced granulocyte proliferation (P < 0.01) and inhibited apoptosis (P < 0.01) and significantly increased avTOR, S6K, 4E-BP1, and PKC gene expression (P < 0.01). The protein expression levels of mTOR, S6K, p-mTOR, and p-S6K were consistent with mRNA expression levels. The mTOR activity is age-specific, and a compensatory enhancement of the mTOR signaling cascade can regulate ovarian follicular development in aged laying hens., (Copyright © 2020. Published by Elsevier Inc.)

Crossbreeding advantage in hybrids compared with their parents, termed heterosis, has been exhaustively exploited in chicken breeding over the last century. Reports for crossbreeding of elite laying chickens covering rearing and laying period remain infrequent. In this study, resource populations of Rhode Island Red (RIR) and White Leghorn (WL) pure-bred chickens were reciprocally crossed to generate 4 distinct groups that were evaluated for prelaying growth, egg production, and egg quality. Birds monitored for prelaying growth consists of 105 (RIR), 131 (WL), 207 (RIR × WL) and 229 (WL × RIR), and 30 pullets from each group were evaluated. Egg laying records were collected from 102, 89, 147, and 191 hens in the 4 populations, respectively. In addition, expression of 5 candidate genes for egg production in the ovarian follicles was measured by RT-qPCR. Results showed that BW of hatched chicks in the WL line was higher than the other populations. However, the 2 crossbreds grew faster than WL purebred throughout the prelaying period. Low to medium heterosis was observed for BW and body length before the onset of lay. White Leghorn and the hybrids commenced laying earlier than RIR pullets and egg production traits were favorable in the crossbreds compared with purebreds. Heterosis for egg number and clutch size was moderate in WL × RIR but low in RIR × WL hens. Expression of antimullerian hormone gene was high in WL and RIR × WL hybrids, suggesting WL parent-specific enhancing dominant expression. Shell weight was higher in the crossbreds than purebreds at 52 wk of age, but RIR hens laid eggs with higher shell ratio than the other populations (P < 0.05). Conversely, WL and the hybrids had higher eggshell strength than RIR birds (P < 0.05). Eggshell strength was the only egg quality trait that showed heterosis above 10% in WL × RIR hybrids at 32 and 52 wk of age. White Leghorn × RIR hens demonstrated higher percent heterosis for economic traits than birds of the reciprocal hybrid. This means that RIR breed is a better dam than a sire line for growth, egg laying, and egg quality traits., (Copyright © 2020. Published by Elsevier Inc.)

The abundance of miR-1b-3p in the chicken ovary is greater after sexual maturation. In the present study there was assessment of whether a single nucleotine polymorphism (SNP) led to an alteration in expression of reproductive traits. The miR-1b-3p abundance was greatest in ovarian follicles The SNP site of rs737028527 (G > A), located in the 734 bp upstream region of pre-miR-1b-3p, was identified in three different chicken breeds. Results from an association analysis of chicken egg-laying traits indicated the SNP was associated with age at first egg production (AFE) and egg number at 32 and 48 weeks (E32, E48; P < 0.01). Hens with genotype AA had an earlier AFE and greater E32 and E48 than hens with other genotypes. The abundance of mature miR-1b-3p in the hens with the GG genotype was larger than those with the AA genotype (P < 0.01), and the luciferase activity of GG genotype promoter was also greater in birds with the AA than GG genotype (P < 0.05). There was inhibition of the production of the transcription factor bound by the specificity protein 1 (Sp1) as a result of the G-to-A mutation, and the luciferase activity of the GG, but not AA, genotype was markedly increased by Sp1. In conclusion, the SNP, rs737028527 (G> A), affected the abundance of mature miR-1b-3p by Sp1 and was associated with chicken egg-laying traits. Data from the present study allow for an increased understanding of the functions and regulation of miR-1b-3p in ovarian follicle development of hens., Competing Interests: Declaration of Competing Interest All authors agreed with the final version of the manuscript and all aspects of the work., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Daidzein (DA) is a kind of isoflavone that is extracted primarily from soy plants and that has become increasingly popular as a dietary supplement. The objective of this study was to evaluate the effects of dietary DA supplementation for laying breeder hens on laying performance, reproductive organ development, hatching performance of seed eggs, and growth performance of offspring and to investigate the underlying molecular mechanisms. A total of 180 55-week-old laying breeder hens were randomly divided into 2 treatment groups and, after 3 weeks of acclimation, were fed either a control diet (CON) or a DA-supplemented diet (DAS, CON+30 mg/kg DA) for a total of 12 weeks. DAS treatment improved the laying rate, luteinizing hormone (LH) levels, and small yellow follicle (SYF) numbers without negative effects on the hatchability of breeder eggs or the growth performance of offspring. High-throughput RNA sequencing was utilized to identify differentially expressed genes in the SYF granulosa layer in the two groups. Transcriptome analysis showed that 161 genes (fold change ≥2 or ≤0.5; P-value<0.05) were significantly differentially expressed between the two groups, including 139 upregulated genes and 22 downregulated genes. Gene ontology (GO) functional annotation analysis revealed potential genes, processes and pathways involved in cell proliferation and differentiation related to the improvement of laying performance stimulated by DA. Dietary DA supplementation for laying breeder hens improved laying performance and reproductive performance with no negative impacts on hatchability or offspring growth. A series of differentially expressed genes in SYF granulosa cells were significantly upregulated in the DAS group relative to the CON group. This study provides insight into the genetic architecture of the transcriptome of the SYF granulosa layer in layer breeding hens and proposes candidate genes that respond to dietary DA., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The objective of this study was to examine the expression profiles of follistatin (FST) and its associated molecules (MSTN, INHA, INHBB, INHBA, ACVR2A, and ACVR2B) in the oviduct of laying hens at 3 hr and 20 hr post-ovulation (p.o., n = 5; 35 weeks old), molting (n = 5; 60 weeks old), and non-laying (n = 4; 35-60 weeks old) hens and also to localize the FST by using immunohistochemistry assay. Expression of FST was significantly higher (p < .05), and MSTN was lower in the uterus of laying hens around 15-20 hr p.o. (during eggshell formation), however, their expressions in the magnum remain unchanged across different physiological stages of hens. FST was mainly expressed in the luminal and glandular epithelium of the uterine tissues, and their expression intensity was highest in laying hens during the eggshell mineralization. There was a relatively increased expression of INHA in the magnum of laying hens around 3 hr p.o. as compared to non-laying and molting hens. At the same time (3 hr p.o.), there was a significant (p < .05) decrease in the expression of the INHBB, ACVR2A, and ACV2B. These results indicate that follistatin may regulate the differentiation of uterine luminal and glandular epithelium during eggshell biomineralization., (© 2020 Japanese Society of Animal Science.)

Background: At sexual maturity, the liver of laying hens undergoes many metabolic changes to support vitellogenesis. In published transcriptomic approaches, hundreds of genes were reported to be overexpressed in laying hens and functional gene annotation using gene ontology tools have essentially revealed an enrichment in lipid and protein metabolisms. We reanalyzed some data from a previously published article comparing 38-week old versus 10-week old hens to give a more integrative view of the functions stimulated in the liver at sexual maturity and to move beyond current physiological knowledge. Functions were defined based on information available in Uniprot database and published literature., Results: Of the 516 genes previously shown to be overexpressed in the liver of laying hens, 475 were intracellular (1.23-50.72 fold changes), while only 36 were predicted to be secreted (1.35-66.93 fold changes) and 5 had no related information on their cellular location. Besides lipogenesis and protein metabolism, we demonstrated that the liver of laying hens overexpresses several clock genes (which supports the circadian control of liver metabolic functions) and was likely to be involved in a liver/brain/liver circuit (neurotransmitter transport), in thyroid and steroid hormones metabolisms. Many genes were associated with anatomical structure development, organ homeostasis but also regulation of blood pressure. As expected, several secreted proteins are incorporated in yolky follicles but we also evidenced that some proteins are likely participating in fertilization (ZP1, MFGE8, LINC00954, OVOCH1) and in thyroid hormone maturation (CPQ). We also proposed that secreted proteins (PHOSPHO1, FGF23, BMP7 but also vitamin-binding proteins) may contribute to the development of peripheral organs including the formation of medullar bones to provide labile calcium for eggshell formation. Thirteen genes are uniquely found in chicken/bird but not in human species, which strengthens that some of these genes may be specifically related to avian reproduction., Conclusions: This study gives additional hypotheses on some molecular actors and mechanisms that are involved in basic physiological function of the liver at sexual maturity of hen. It also revealed some additional functions that accompany reproductive capacities of laying hens, and that are usually underestimated when using classical gene ontology approaches.

The number of days (DN) when hens lay fertile eggs as well as the number of fertile eggs (FN) were produced after a single artificial insemination (AI), including the two duration of fertility (DF) traits. Indeed, they are the key production performance that associates with the production cost of hatching egg when its determination the interval between successive artificial inseminations. However, the relevant genes response for regulating the DF has not been uncovered yet. Therefore, we performed a weighted gene co-expression network analysis (WGCNA) to investigate the insight into co-expression gene modules on DF process in hens. The total mRNA was extracted from the utero-vaginal junction (UVJ, with the sperm storage function in hen's oviduct which is the biological basis for DF) of 20 hens with several levels of DF traits, and performed transcriptome sequences of mRNA. As a result, three co-expression gene modules were identified to be highly correlated with DF traits. Moreover, the expression changes of top 5 hub genes in each module with DF traits were further confirmed in other 20 hens by RT-PCR. These findings highlighted the co-expression modules and their affiliated genes as playing important roles in the regulation of DF traits.

Egg production is determined by the function of ovary and is regulated by the hypothalamic-pituitary-ovary axis. The mechanism by which the ovary regulates egg production, however, is still poorly understood. The purpose of this study is to compare the transcriptome difference in ovary of relatively greater and lesser egg producing chickens, and to screen candidate genes related to egg production. A RNA sequencing was performed to analyze and compare the mRNA in ovarian tissues of relatively greater and lesser egg producing chickens. A total of 4 431 new genes expressed in the chicken ovary were mined. There were 305 differentially expressed genes (DEGs) identified between the relatively greater and lesser egg producing hens. Gene ontology analysis identified five candidate genes related to egg production, including ZP2, WNT4, AMH, IGF1, and CYP17A1 genes. Tissue expression profiles indicated these five candidate genes were highly expressed in chicken ovarian tissues, indicating a potential role in regulating chicken ovarian function and egg production. The KEGG analysis indicated the neuroactive ligand-receptor interaction pathway might have an important function in regulation of egg production. In addition, four known pathways related to reproduction were detected, including the calcium signaling, wnt signaling pathway, focal adhesion, and cytokine-cytokine receptor interaction pathways. Results of the present study indicate gene expression differences in the ovarian tissues of relatively greater and lesser egg producing chickens, and identified five important candidate genes related to egg production, which provided a theoretical basis for improving egg production of Jinghai Yellow Chickens., (Copyright © 2019. Published by Elsevier B.V.)

Egg production is an important economic trait in poultry, and it is of great significance to study the key genes and functional SNPs that affect egg laying performance. Follicle-stimulating hormone (FSH) plays an important physiological role in the reproductive performance of humans and animals by binding to its receptor (FSHR). Studies have shown that there are many transcriptional regulatory elements in the 5' flanking region of the FSHR gene that interact with transcription factors to regulate FSHR transcription. In this study, DNA sequencing was used to identify SNPs in the FSHR promoter sequence in both Dongxiang and Suken chickens. To detect the activity of the chicken FSHR gene promoter, we analysed the characteristics of the sequence and constructed three deletion vectors. We confirmed that the region (-18/-544) was the core promoter. Furthermore, five polymorphisms, including a 200-bp indel at -869, C-1684T, C-1608T, G-368A and T-238A, were detected in both the Dongxiang and Suken chickens. The age at first egg (AFE) for different genotype of -869 indel in Suken chicken was significantly different (p < 0.01). For SNP C-1684T in Dongxiang chickens, the CC genotype had higher egg number at 43 weeks of age (E43) than that of the TC genotype (p < 0.05). For SNP C-1684T in Suken chickens, the TC genotype had higher AFE than that of the CC genotype (p < 0.05). For SNP C-1608T in Suken chickens, the CC genotype had higher AFE than that of the TC genotype (p < 0.05). For SNP G-368A in Suken chickens, the AG genotype had higher AFE than that of the GG genotype (p < 0.05)., (© 2019 The Authors. Reproduction in Domestic Animals Published by Blackwell Verlag GmbH.)

MicroRNAs (miRNAs) are a class of endogenous non-coding RNAs that have an important role in post-transcriptional regulation. Previous analysis of the chicken ovary transcriptome indicated there was a differential abundance of miR-26a-5p in sexually mature and immature hens. In this study, ten single nucleotide polymorphism (SNP) sites in miR-26a-5p primary transcripts (pri-miR-26a-5p) were identified using Sanger sequencing on PCR products in three chicken breeds, among which, the rs316365438(C > T)SNP was associated with age at first egg (AFE) and total number of eggs at 32 weeks (E32). The hens with the TT genotype of the rs316365438(C > T)SNP had an earlier AFE and greater E32 than those with CC and CT genotypes. Due to the presence of linkage disequilibrium (LD) in SNPs, there were only two dominant haplotypes. The two dominant haplotypes with different free-energy and secondary structure were identified and the overexpression vectors were constructed. Overexpression of mature miR-26a-5p from TCTACGCACG haplotype was greater than that from the TTTATATACA haplotype. In summary, the polymorphisms of pri-miR-26a-5p are associated with chicken egg production traits and affect the transcript abundance of the mature miR-26a-5p., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Clutch traits were proposed as a more detailed description of egg-laying patterns than simple total egg production. In this study, egg production of 23,809 Rhode Island Red (RIR) and 22,210 White Leghorn (WL) hens was described in terms of number of clutches, average and maximum clutch size, age at first egg, total saleable egg production, and percentage of egg defects. Genetic parameters were estimated using a six-trait animal model. Of the phenotyped birds, 1433 RIR hens and 1515 WL hens were genotyped with line specific 50K Affymetrix Axiom single nucleotide polymorphism chips to perform genome-wide association analyses. Moderate heritabilities were estimated for clutch traits of 0.20 to 0.42 in the RIR line and 0.29 to 0.41 in the WL line. Average and maximum clutch size was positively genetically correlated with total saleable egg number in both lines. Genome-wide association analysis identified seven regions that were associated with egg production in the RIR line and 12 regions in the WL line. The regions identified were line and trait specific, except for one region on chromosome 6 from 28 to 29 Mb that influenced number of clutches and maximum and average clutch size in WL hens. Regions associated with egg production identified here overlapped with 260 genes, with some strong positional candidates based on gene ontology including WASH1, which is involved in oocyte maturation, NPVF, involved in regulation of follicle-stimulating hormone secretion, and FOXO3, involved in oocyte maturation and ovulation from the ovarian follicle. Confirmation of the role of these genes in regulation of egg production pattern will require further studies.

Eggshell is the outermost calcified covering of an egg that protects it from microbial invasion and physical damage, and is critical for egg quality. However, understanding of the genes/proteins and the biological pathways regulating the eggshell formation is still obscure. We hypothesized that the transcriptomic analysis of the chicken uteri using RNA-sequencing may reveal novel genes and biological pathways involved in the eggshell biomineralization. RNA-sequence analysis using uteri of laying hens at 15-20 h post-ovulation (layers, n = 3) and non-laying (non-layers, n = 3) hens was carried out. About 229 differentially expressed genes (DEGs) were up-regulated in the layers compared to the non-layers. Kyoto Encyclopedia of Genes and Genomes (KEGG) and Ingenuity Pathway Analysis (IPA) revealed more than ten novel genes and biological pathways related to calcium transport and mineralization in the uterus. Based on the enriched pathways and molecular function analysis, 12 DEGs related to eggshell mineralization were further analyzed in the uteri of layers (3 h and 15-20 h post-ovulation), non-layers and molters using qPCR. Expressions of OC-116 (regulator of mineralization), OTOP2 (modulator of cellular calcium influx), CALCB (intracellular release of Ca-ions), STC2 (increases alkaline phosphatase activity), and ATP2C2 (cellular import of Ca-ions) were significantly higher in the uteri of laying hen at 15-20 h post-ovulation. This study identified the involvement of novel genes and their proposed biological pathways in the regulation of eggshell formation.

Background: In this work we have determined molecular signatures of oviduct epithelial and progenitor cells. We have proposed a panel of selected marker genes, which correspond with the phenotype of oviduct cells of a laying hen (Gallus gallus domesticus) and quail (Coturnix japonica). We demonstrated differences in characteristics of those cells, in tissue and in vitro, with respect to different anatomical and functional parts of the oviduct (infundibulum (INF), distal magnum (DM, and proximal magnum (PM)). The following gene expression signatures were studied: (1) oviduct markers (estrogen receptor 1, ovalbumin, and SPINK7 - ovomucoid), (2) epithelial markers (keratin 5, keratin 14, and occludin) and (3) stem-like/progenitor markers (CD44 glycoprotein, LGR5, Musashi-1, and sex determining region Y-box 9, Nanog homebox, OCT4/cPOUV gene encoding transcription factor POU5F3)., Results: In chicken, the expression of oviduct markers increased toward the proximal oviduct. Epithelial markers keratin14 and occludin were high in distal oviduct and decreased toward the proximal magnum. In quail oviduct tissue, the gene expression pattern of oviduct/epithelial markers was similar to chicken. The markers of progenitors/stemness in hen oviduct (Musashi-1 and CD44 glycoprotein) had the highest relative expression in the infundibulum and decreased toward the proximal magnum. In quail, we found significant expression of four progenitor markers (LGR5 gene, SRY sex determining region Y-box 9, OCT4/cPOUV gene, and CD44 glycoprotein) that were largely present in the distal oviduct. After in vitro culture of oviduct cells, the gene expression pattern has changed. High secretive potential of magnum-derived cells diminished by using decreased abundance of mRNA. On the other hand, chicken oviduct cells originating from the infundibulum gained ability to express OVM and OVAL. Epithelial character of the cells was maintained in vitro. Among progenitor markers, both hen and quail cells expressed high level of SOX9, LGR5 and Musashi-1., Conclusion: Analysis of tissue material revealed gradual increase/decrease pattern in majority of the oviduct markers in both species. This pattern changed after the oviductal cells have been cultured in vitro. The results can provide molecular tools to validate the phenotype of in vitro biological models from reproductive tissue.

Transcription factors (TFs) encoded by SALL1 and SALL3 genes play central roles in the regulation of ovarian development in hens. The present study aimed to examine polymorphisms of these two genes in Chinese Dagu chickens, and to identify the effects of TFs on the laying performance. Among the population, two novel single-nucleotide polymorphisms (SNPs) were identified by single-strand conformation polymorphism (SSCP) in the amplicons of the candidate genes. The effect of the SNP (729C > A) in exon 2 of SALL1 gene on egg production at 43, 57, and 66 weeks and EW at 30 and 43 weeks were the most significant in the 360 samples (P < 0.05). Moreover, for the SNP 1014T > A (in exon 2 of SALL3), the TT genotype was significantly correlated with higher egg production and EW (P < 0.05). Furthermore, four combined genotypes were reconstructed based on the two SNPs. The combined genotype TATT was correlated with the highest egg production at 43-66 weeks and with higher EW at 30, 43 weeks (P < 0.05). The polymorphisms of the two TFs studied are potential molecular genetic markers for chicken breeding, which might help in understanding the genetic structure of laying performance and improving these traits directly by marker-assisted selection (MAS)., (© 2018 Wiley Periodicals, Inc.)

The aim of the study was to evaluate changes in laying patterns depending on the age of different genotypes of laying hens. In the experiment, six genotypes were evaluated (brown-egg hens Bovans Brown, Bovans Sperwer and Isa Sussex, white-egg hens Dekalb White, and laying hens with tinted shells Moravia Barred and Moravia BSL) in three periods during the laying cycle (the onset of lay between 20 and 26 weeks of age, the middle from 36 to 42 weeks of age and the end of lay between 64 and 70 weeks of age). A significant interaction between genotype and age was apparent in mean sequence length (P<0.001), length of the prime sequence (P<0.001), mean number of sequences (P<0.001) and mean time of oviposition (P˂0.001). The longest lag during the course of the experiment was with Moravia BSL, which was more than 3h; the shortest lag was observed in Bovans Brown, which was less than 1h. The mean time of oviposition was also affected by genotype (P˂0.001). Bovans Brown laid their eggs approximately 3.5h after the lights came on, whereas Moravia BSL laid their eggs almost 6h after the lights came on. Egg weight increased with age (P˂0.001), and the smallest differences in egg weight were with ISA Sussex (5g), whereas the biggest differences were with Moravia BSL (10g)., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Estrogen receptors α (ESR1) and β (ESR2) play central roles in folliculogenesis and therefore in reproductive biology. In the present study, two single nucleotide polymorphisms (SNPs) were identified in the ESR1 and ESR2 genes using PCR-single strand conformation polymorphism (PCR-SSCP) and DNA sequencing. One of the identified SNPs, a T1101C transition located within exon 4 of the ESR1 gene, was significantly associated with hen-housed egg production (HHEP) at 30, 43, 57 and 66 weeks of age (P<0.05), and egg weight (EW) at 30 weeks (P<0.05). Another SNP, a G1755A transition leading to a non-synonymous substitution (valine 459-to-isoleucine) located within exon 8 of the ESR2 gene, was also markedly correlated with the HHEP at 30, 43, 57 and 66 weeks of age (P<0.05), and EW at 30 weeks (P<0.05). A greater proportion of the additive variance was explained by the SNPs for most of the associated egg production traits (>1%). Furthermore, the results of the combined genotype-based association analysis supported the finding that the two SNPs were associated with the traits under a study. Taken together, our findings suggest that the two sequence variations in the ESR1 and ESR2 genes may provide promising genetic markers for the early selection and prediction of advantageous phenotypes in chicken breeding.

Although non-destructive deformation is relevant for assessing eggshell strength, few long-term selection experiments are documented which use non-destructive deformation as a selection criterion. This study used restricted maximum likelihood-based methods with a four-trait animal model to analyze the effect of non-destructive deformation on egg production, egg weight and sexual maturity in a two-way selection experiment involving 17 generations of White Leghorns. In the strong shell line, corresponding to the line selected for low non-destructive deformation values, the heritability estimates were 0.496 for non-destructive deformation, 0.253 for egg production, 0.660 for egg weight and 0.446 for sexual maturity. In the weak shell line, corresponding to the line selected for high non-destructive deformation values, the heritabilities were 0.372, 0.162, 0.703 and 0.404, respectively. An asymmetric response to selection was observed for non-destructive deformation, egg production and sexual maturity, whereas egg weight decreased for both lines. Using non-destructive deformation to select for stronger eggshell had a small negative effect on egg production and sexual maturity, suggesting the need for breeding programs to balance selection between eggshell traits and egg production traits. However, the analysis of the genetic correlation between non-destructive deformation and egg weight revealed that large eggs are not associated with poor eggshell quality., (© 2016 Japanese Society of Animal Science.)

Shell quality is one of the most important traits for improvement in layer chickens. Proper consideration of repeated records can increase the accuracy of estimated breeding values and thus genetic improvement of shell quality. The objective of this study was to compare different models for genetic evaluation of the collected data. For this study, 81,646 dynamic stiffness records on 21,321 brown egg layers and 93,748 records on 24,678 white egg layers from 4 generations were analyzed. Across generations, data were collected at 2 to 4 ages (at approximately 26, 42, 65, and 86 wk), with repeated records at each age. Seven models were compared, including 5 repeatability models with increasing complexity, a random regression model, and a multitrait model. The models were compared using Akaike Information Criteria with significance testing of nested models with a Log Likelihood Ratio test. Estimates of heritability were 0.31-0.36 for the brown line and 0.23-0.26 for the white line, but repeatability was higher for the model with age-specific permanent environment effects (0.59 for both lines) than for the model with an overall permanent environmental effects (0.47 for the brown and 0.41 for the white line). The model that allowed for permanent environmental effect within age and heterogeneous residual variance between ages resulted in improved fit compared to the traditional model that fits single permanent environment and residual effects, but was inferior in fit and predictive ability to the full multiple-trait model. The random regression model had better fit to the data than repeatability models but slightly worse than the multiple-trait model. For traits with repeated records at different ages, repeatability within and across ages as well as genetic correlations should be considered while choosing the number of records collected per individual as well as the model for genetic evaluation.

To identify molecular markers and candidate genes associated with reproductive traits, a genome-wide analysis was performed in Jinghai Yellow Chickens to analyze body weight at first oviposition (BWF), age at first oviposition (AFE), weight of the egg at first oviposition (FEW), egg weight at the age of 300 days (EW300), number of eggs produced by 300 days of age (EN300), egg hatchability (HA) and multiple selection index for egg production (MSI). The results showed that seven single nucleotide polymorphisms (SNPs) were associated with reproductive traits (P<1.80E-6, Bonferroni correction). The P-values of the seven SNPs were 5.62E-10, 3.45E-08, 9.76E-07, 8.90E-07, 1.12E-06, 1.42E-07 and 1.48E-07, respectively. These SNPs were located in close proximity to or within the sequence of the five candidate genes, including FAM184B, TTL, RGS1, FBLN5 and PCNX. An additional 46 SNPs that could be associated with reproductive traits were identified (P<3.59E-5, Bonferroni correction). Identification of the candidate genes as well as genome-wide SNPs that may be associated with reproductive traits will greatly advance the understanding of the genetic basis and molecular mechanisms underlying reproductive traits and may have practical significance in breeding programs for the improvements of reproductive traits in the Jinghai Yellow Chicken., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Egg number (EN), egg laying rate (LR) and age at first egg (AFE) are important production traits related to egg production in poultry industry. To better understand the knowledge of genetic architecture of dynamic EN during the whole laying cycle and provide the precise positions of associated variants for EN, LR and AFE, laying records from 21 to 72 weeks of age were collected individually for 1,534 F2 hens produced by reciprocal crosses between White Leghorn and Dongxiang Blue-shelled chicken, and their genotypes were assayed by chicken 600 K Affymetrix high density genotyping arrays. Subsequently, pedigree and SNP-based genetic parameters were estimated and a genome-wide association study (GWAS) was conducted on EN, LR and AFE. The heritability estimates were similar between pedigree and SNP-based estimates varying from 0.17 to 0.36. In the GWA analysis, we identified nine genome-wide significant loci associated with EN of the laying periods from 21 to 26 weeks, 27 to 36 weeks and 37 to 72 weeks. Analysis of GTF2A1 and CLSPN suggested that they influenced the function of ovary and uterus, and may be considered as relevant candidates. The identified SNP rs314448799 for accumulative EN from 21 to 40 weeks on chromosome 5 created phenotypic differences of 6.86 eggs between two homozygous genotypes, which could be potentially applied to the molecular breeding for EN selection. Moreover, our finding showed that LR was a moderate polygenic trait. The suggestive significant region on chromosome 16 for AFE suggested the relationship between sex maturity and immune in the current population. The present study comprehensively evaluates the role of genetic variants in the development of egg laying. The findings will be helpful to investigation of causative genes function and future marker-assisted selection and genomic selection in chickens.

The aim of this study was to determine whether the egg-laying phase and estrogen affect the induction of cytotoxic cells in response to avian infectious bronchitis (IB) virus at early stage of infection in the oviduct. Attenuated IB virus (aIBV group) or its vehicle (control group) was introduced to the oviductal magnum lumen of White Leghorn hens in the laying and molting phase, as well as molting hens injected with estradiol benzoate (M-EB hens) or corn oil (M-oil hens). Oviductal isthmus and uterus were collected 24h after injection. The frequency of CD8(+) and TCRγδ(+) T cells expression was examined by immunohistochemistry, followed by image analysis. The expression of the genes of toll-like receptor 7 (TLR7), natural killer cell receptor (BNK), cytotoxic substances (granzyme, perforin), and cytokines (CXCL12, CX3CL1, and IFNγ) were examined by real-time polymerase chain reaction analysis. The frequency of CD8(+) and TCRγδ(+) T cells in the isthmus, and CD8(+) cells in the uterus was significantly higher in the aIBV group compared to the control group of laying and M-EB hens. The expression of all the genes examined in this study in the isthmus, and CX3CL1 and IFNγ expression in the uterus was significantly higher in the aIBV group in the laying and M-EB hens. These results suggested that infection with IB virus causes an immune response involving the influx of cytotoxic cells and upregulation of cytokines in the isthmus and uterus at early stage of infection. This response was stronger during the laying phase compared to the molting phase, probably due to the effect of estrogen., (Copyright © 2015 Elsevier B.V. All rights reserved.)

One of the main concerns for poultry producers is how to maintain egg uniformity and stability in size and weight following the rapid growth during the early laying period. In this study, we aimed to investigate the increase in egg weight with advancing hen age, and to estimate genetic parameters of these increment traits in 2 pure lines of chickens (i.e., 2,010 White Leghorn and 1,200 brown-egg dwarf hens), using the restricted maximum likelihood method with the DMU procedure. We collected age at first egg (AFE), first egg weight (FEW) and kept records of egg weight per 10 wk from 30 to 60 wk of age. Meanwhile, the increments of egg weight were calculated for the evaluation of age-dependent dynamic changes. The increment of egg weight gained dramatically before 30 wk of age and became slower with the advance of age. Heritability estimates of AFE were larger than 0.32, and the low to moderate genetic correlations between AFE and FEW were observed in the 2 lines. The FEW showed high variation level compared with egg weights at later ages in the 2 lines, and had moderate heritability estimates in White Leghorns (0.20) and dwarf hens (0.33). Egg weights at different ages were highly heritable in the 2 lines (h(2) ≥ 0.35), and had strong genetic and phenotypic correlations among different ages. The estimates of heritability for most increment traits were low to moderate, especially those increments for 10-wk intervals ranging from 0.00 to 0.14. The genetic correlations among 3 consecutive egg weight increments for 10-wk intervals were low to moderate. Our results in the 2 lines should provide important insights into the genetic architecture of increment traits and offer some suggestions for producing uniform and stable eggs in response to advancing age., (©2014 Poultry Science Association Inc.)

Discovery of genes with large effects on economically important traits has for many years been of interest to breeders. The development of SNP panels which cover the whole genome with high density and, more importantly, that can be genotyped on large numbers of individuals at relatively low cost, has opened new opportunities for genome-wide association studies (GWAS). The objective of this study was to find genomic regions associated with egg production and quality traits in layers using analysis methods developed for the purpose of whole genome prediction. Genotypes on over 4500 birds and phenotypes on over 13,000 hens from eight generations of a brown egg layer line were used. Birds were genotyped with a custom 42K Illumina SNP chip. Recorded traits included two egg production and 11 egg quality traits (puncture score, albumen height, yolk weight and shell colour) at early and late stages of production, as well as body weight and age at first egg. Egg weight was previously analysed by Wolc et al. (2012). The Bayesian whole genome prediction model--BayesB (Meuwissen et al. 2001) was used to locate 1 Mb regions that were most strongly associated with each trait. The posterior probability of a 1 Mb window contributing to genetic variation was used as the criterion for suggesting the presence of a quantitative trait locus (QTL) in that window. Depending upon the trait, from 1 to 7 significant (posterior probability >0.9) 1 Mb regions were found. The largest QTL, a region explaining 32% of genetic variance, was found on chr4 at 78 Mb for body weight but had pleiotropic effects on other traits. For the other traits, the largest effects were much smaller, explaining <7% of genetic variance, with regions on chromosomes 2, 12 and 17 explaining above 5% of genetic variance for albumen height, shell colour and egg production, respectively. In total, 45 of 1043 1 Mb windows were estimated to have a non-zero effect with posterior probability > 0.9 for one or more traits., (© 2014 Blackwell Verlag GmbH.)

Different single-strand conformation polymorphism (SSCP) patterns of the 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR) gene amplified by six pairs of primers were sequenced. Comparisons among the six nucleotide sequences of three genotypes revealed three base substitutions (G-789-A, C-937-G, and A-2316-C) in the HMGCR gene. The genotypes of single nucleotide polymorphism 1 (SNP1) and SNP3 were significantly associated with the number of eggs (P < 0.01), and the different genotypes of SNP2 did not significantly influence the number of eggs. The genotypes of all three SNPs were not related to the hatching rate of fertilized eggs (P > 0.05). Of the three SNPs of the HMGCR gene, those genotypes with lower plasma very low-density lipoprotein (VLDL) concentrations had higher egg production. The 300-day egg production of the six studied haplotypes showed three levels (P < 0.01). The ADE and ACE haplotypes showed lower egg production than the other haplotypes. The ADE haplotype had the highest plasma VLDL concentration, and the ACE haplotype had the lowest plasma VLDL concentration. We concluded that specific variants of the HMGCR gene are associated with chicken egg production, and haplotypes with higher and lower plasma VLDL concentrations showed lower egg production than other haplotypes.

1. Two experiments were conducted to determine the influence of the frizzle gene (F) on the production and quality of chicken eggs kept on village farms in Ghana. 2. In the first experiment, 144 pullets, 72 each of Ff and ff pullets from F1 birds, were compared in a randomised complete block design (RCBD), with three villages and 4 batches of hatch as blocks and the two feather genotypes (Ff and ff) as the treatments. 3. The Ff pullets were superior (P < 0.05) to their ff counterparts in terms of the number of eggs per clutch, egg mass and hen-housed and hen-d rates of lay, whereas age at first egg was lower (P < 0.05) in ff layers compared to Ff ones. 4. The eggs of the F1 heterozygous frizzle (Ff) layers had higher values for albumen height, Haugh unit and yolk height compared with eggs from their normal feathered counterparts. 5. In the second experiment, 144 pullets, 48 each of FF, Ff and ff pullets, were compared in a RCBD, with three villages and 4 batches of hatch as blocks and the two feather genotypes (Ff and ff) as the treatments. 6. The F2 birds with genotypes Ff and FF were better than their ff counterparts in terms of the number of eggs per clutch, egg mass, and hen-housed and hen-d rates of lay. Age at first egg was significantly lower in ff layers compared to FF and Ff ones. 7. The Haugh unit value was higher in the homozygous and heterozygous frizzles compared to the normal feathered ones. 8. The presence of the frizzle gene (F) in egg type chickens led to an improvement in egg production and egg quality traits in village chickens, and the cross-breeding scheme evaluated in this project could contribute to improved productivity.

A breed development strategy aimed at making exotic layers (Lohmann Brown) more productive under tropical environment using thermo-regulatory genes is underway at Akate Farms in Kumasi, Ghana. The present experiment was carried out to find out the effect of the genes on egg production in hot and humid environments. Three genetic groups comprising naked-neck, frizzle and their normally feathered sibs were obtained after successive generations of crossing between naked-neck and frizzle cocks and Lohmann brown hens. A total of 270 18-week-old pullets, 90 each of the 3 groups, were selected randomly and assigned to a completely randomized design experiment with 3 replicates, with 30 birds in each replicate group and kept up to a period of 72 weeks. The birds were kept in a partitioned open-sided deep-litter house constructed with sandcrete blocks with 30 pullets in each compartment. They were fed ad libitum with layer diets containing 18 % crude protein and 2,800 kcal ME/kg. Results obtained showed that the crossbred naked-neck and frizzle phenotypes produced eggs at a significantly (P < 0.05) higher rates than their normally feathered sibs and also out-performed their normally feathered sibs in other egg production parameters measured, even though they all segregated from similar parents. This is an indication of the favourable effect of the genes on egg production under hot and humid environments.

In an earlier study, serum levels of natural antibody isotypes IgM- and IgG-binding keyhole limpet hemocyanin were found to be indicative for survival through the laying period of hens and therefore considered as promising traits for future implementation in breeding programs for higher survival of layers. In the present study, we first estimated the genetic parameters for the two isotypes at 20, 40 and 65 weeks of age (IgM20, IgM40 and IgM65; IgG20, IgG40 and IgG65). Pooled genetic parameters were estimated from the total population of 2504 hens from nine purebred layer lines, with line included in the model to account for admixture. Moderate heritabilities (0.14-0.44) indicated that selection for isotype titers is feasible, especially for IgM. Secondly, associations between 1022 SNP markers and the above-mentioned six immunological traits were estimated in 650 genotyped hens from the nine lines. The association study was performed across lines to detect markers that are closer to the QTL and have the same phase of association in the entire population. Forty-three significant associations between SNPs and isotype titers were detected. The SNPs of interleukins IL10 and IL19 were associated with both isotypes; SNPs of tripartite motif containing 33 (TRIM33) and IL6 showed significant association with IgG20 and IgM20 respectively; SNPs of heat shock protein 90kDa alpha (cytosolic), class B member 1 (HSP90AB1) were associated with IgG titers at older ages. Some detected SNPs were also reported associated with other immune and behavioral traits., (© 2012 The Authors, Animal Genetics © 2012 Stichting International Foundation for Animal Genetics.)

Random regression models allow for analysis of longitudinal data, which together with the use of genomic information are expected to increase accuracy of selection, when compared with analyzing average or total production with pedigree information. The objective of this study was to estimate variance components for egg production over time in a commercial brown egg layer population using genomic relationship information. A random regression reduced animal model with a marker-based relationship matrix was used to estimate genomic breeding values of 3,908 genotyped animals from 6 generations. The first 5 generations were used for training, and predictions were validated in generation 6. Daily egg production up to 46 wk in lay was accumulated into 85,462 biweekly (every 2 wk) records for training, of which 17,570 were recorded on genotyped hens and the remaining on their nongenotyped progeny. The effect of adding additional egg production data of 2,167 nongenotyped sibs of selection candidates [16,037 biweekly (every 2 wk) records] to the training data was also investigated. The model included a 5th order Legendre polynomial nested within hatch-week as fixed effects and random terms for coefficients of quadratic polynomials for genetic and permanent environmental components. Residual variance was assumed heterogeneous among 2-wk periods. Models using pedigree and genomic relationships were compared. Estimates of residual variance were very similar under both models, but the model with genomic relationships resulted in a larger estimate of genetic variance. Heritability estimates increased with age up to mid production and decreased afterward, resulting in an average heritability of 0.20 and 0.33 for pedigree and genomic models. Prediction of total egg number was more accurate with the genomic than with the pedigree-based random regression model (correlation in validation 0.26 vs. 0.16). The genomic model outperformed the pedigree model in most of the 2-wk periods. Thus, results of this study show that random regression reduced animal models can be used in breeding programs using genomic information and can result in substantial improvements in the accuracy of selection for trajectory traits.

Avian beta-defensins (AvBDs), also known as gallinacins, are small cationic peptides having three cysteine disulfide bonds between their cysteine residues. They play essential roles in the innate immune system as well as stimulate proliferation of epithelial cells and fibroblasts. Although we found the avian homolog of human beta-defensin 11 to be highly expressed in chicks treated with the diethylstilbestrol (DES, a synthetic estrogen agonist), little is known about the hormonal and transcriptional regulation of AvBD-11 in the chicken oviduct and its expression in cancerous ovaries of chickens. Results of this study of young chicks revealed that DES induced AvBD-11 mRNA and protein in the oviduct, specifically luminal and glandular epithelial cells. In addition, microRNA-1615 was discovered to influence AvBD-11 expression via its 3'-UTR which suggests post-transcriptional regulation of AvBD-11 expression in chickens. Furthermore, we compared the expression patterns of the AvBD-11 gene in normal and cancerous ovaries from laying hens which are models for human epithelial ovarian cancer. Our results demonstrated that AvBD-11 is most abundant in the glandular epithelium of endometrioid-type ovarian tumors, but not normal ovaries of laying hens. Collectively, these results suggest that AvBD-11 is an estrogen-induced gene during oviduct development and that it may be used as a biomarker for diagnosis of ovarian cancer and for monitoring effects of therapeutics on progression of ovarian carcinogenesis., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

1. The aim of this study was to compare the changes in the production and in the body and egg composition of 45 TETRA SL brown egg layers and 45 TETRA BLANCA white egg layers during the first egg-laying period. 2. Changes in the body composition of the hens were followed in vivo by means of computed tomography (CT) four-weekly, between 20 and 72 weeks of age. The measurements covered the whole body of the hens using overlapping 10 mm slice thicknesses on a Siemens Somatom Emotion 6 multislice CT scanner. 3. The yolk, albumen and shell ratio of the eggs, produced on the days of the CT measurements by the hens, were determined and their composition was analysed chemically. 4. The body fat content of the hens increased continuously until 44 weeks of age and plateaued thereafter in both genotypes. However, the body fat content of the white egg layers was always higher than that of the brown egg layers. 5. The yolk ratio and the dry matter and crude fat content of the eggs of white egg layers were higher than the brown egg layers throughout the experiment. 6. Moderate correlations were observed in both genotypes between the body fat content of the hens and egg yolk ratio of their eggs.

Egg production curves describe the laying patterns of hen populations over time. The objectives of this study were to fit the weekly egg production rate of selected and nonselected lines of a White Leghorn hen population, using nonlinear and segmented polynomial models, and to study how the selection process changed the egg-laying patterns between these 2 lines. Weekly egg production rates over 54 wk of egg production (from 17 to 70 wk of age) were measured from 1,693 and 282 laying hens from one selected and one nonselected (control) genetic line, respectively. Six nonlinear and one segmented polynomial models were gathered from the literature to investigate whether they could be used to fit curves for the weekly egg production rate. The goodness of fit of the models was measured using Akaike's information criterion, mean square error, coefficient of determination, graphical analysis of the fitted curves, and the deviations of the fitted curves. The Logistic, Yang, Segmented Polynomial, and Grossman models presented the best goodness of fit. In this population, there were significant differences between the parameter estimates of the curves fitted for the selected and nonselected lines, thus indicating that the effect of selection changed the shape of the egg production curves. The selection for egg production was efficient in modifying the birds' egg production curve in this population, thus resulting in genetic gain from the 5th to the 54th week of egg laying and improved the peak egg production and the persistence of egg laying.

White Leghorn chickens were selected for 36 generations for high (HAS) or low (LAS) antibody response to SRBC 5 d after an intravenous challenge. The aim of this study was to investigate possible changes in reproductive soundness resulting from that selection. Age and BW at onset of lay (first egg), along with weight of the first egg, were recorded on 45 hens from each line. Intensity of lay was measured as the number of ovulations within a 15-d period over 15 sequential intervals (total 225 d). Three cycles of fertility also were assessed, coinciding with early, middle, and late production stages. For fertility of males and females within a line to be independently evaluated, roosters and hens were mated by artificial insemination to an unrelated control line of White Plymouth Rocks. Twenty roosters from each antibody line were considered, as well as the 45 hens. Pooled semen from the control line was used for mating the hens from the antibody lines. Hens from the LAS line commenced lay at a younger age (11.67±3.53 d; P<0.001), lighter BW (-169.46±40.20 g; P<0.001), and with greater intensity (2.68±0.25%; P=0.001) than those from the HAS line. Any differences in intensity thereafter were trivial between lines (P=0.42), with intensity decreasing sharply toward the end of the 7-mo production period in both lines. Length of fertility differed between hens of the antibody lines during the first cycle (3.35±0.85 d; P=0.002) and between roosters during the first (3.58±1.06 d; P=0.02) and second (3.38±1.07 d; P=0.03) cycles, with chickens from the LAS line having the longer length of fertility in both sexes. A correlated response in reproductive soundness to divergent selection for antibody response was observed. This may in part be due to differences in resource allocations, with particular impact on duration of fertility.

We estimated genetic parameters for egg production in different periods by means of random regression models, aiming at selection based on partial egg production from a generation of layers. The production was evaluated for each individual by recording the number of eggs produced from 20 to 70 weeks of age, with partial records taken every three weeks for a total of 17 periods. The covariance functions were estimated with a random regression model by the restricted maximum likelihood method. A model composed of third-order polynomials for the additive effect, ninth-order polynomials for the permanent environment, and a residual variance structure with five distinct classes, was found to be most suitable for adjusting the egg production data for laying hens. The heritability estimates varied from 0.04 to 0.14. The genetic correlations were all positive, varying from 0.10 to 0.99. Selection applied in partial egg production periods will result in greater genetic profit for the adjacent periods. However, as the distance in time between periods increases, selection becomes less efficient. Selection based on the second period (23 to 25 weeks of age), where greater heritability was estimated, would note benefit the final egg-laying cycle periods.