1 Introduction

Duck occupies an important position in the global livestock and poultry industry and is one of the main sources of meat and down products. With the development of modern breeding, breeding work has significantly improved the growth rate and meat production performance of ducks, but it also brought about certain physiological development problems, such as shortening of the growth period accompanied by incomplete muscle development and incomplete feathers during slaughter (Chen et al., 2017). Therefore, on the basis of ensuring the results of genetic selection and breeding, it is of great significance to conduct in-depth research on the molecular mechanisms that affect duck growth and feather development. Recent studies have found that epigenetics provides a new perspective for understanding how genes and the environment work together on traits. Epigenetic mechanisms can regulate gene expression without changing DNA base sequences, resulting in phenotypic differences under conditions of constant genotype (Sepers et al., 2019).

Epigenetics mainly includes DNA methylation, histone covalent modification (such as acetylation, methylation, etc.), and non-coding RNA-mediated regulation of gene expression (Huang et al., 2025). DNA methylation usually occurs on the CpG dinucleotides in the promoter region of the gene, and the increased methylation level is often associated with gene silencing; post-translational modification of histones changes the degree of chromatin tightness, thereby affecting the accessibility of transcription factors to genes. In addition, non-coding RNAs such as microRNAs (miRNAs), long-chain non-coding RNAs (lncRNAs) and circular RNAs (circRNAs) can finely regulate gene function at the post-transcriptional or translational level by interacting with target mRNAs or proteins (Chen et al., 2017). Together, these epigenetic mechanisms form a complex regulatory network, which plays an important role in the processes of duck muscle growth, fat deposition, gonad development, and feather morphology construction.

Based on the above background, this study will review important research progress in the field of epigenetic regulation in recent years based on the two themes of duck growth and feather development. We will explore the role of epigenetic mechanisms in the growth process of ducks (including muscle and skeletal development, metabolic regulation, etc.) in the embryonic and growth fattening stages; focus on explaining the epigenetic regulation of duck feather development (hair follicle generation, feather differentiation and replacement); aim to systematically summarize the current research status of epigenetic regulation of duck growth and feather development, and provide a scientific basis for the future application of epigenetic principles in poultry production.

2 Overview of Duck Growth and Feather Development

2.1 Characteristics of growth and development stages

Ducks grow and develop through multiple stages in the embryonic and postnatal periods. Duck embryos can hatch out of the shell after developing in fertilized eggs for about 28 days. The embryo development speed and tissue differentiation of the fertilized eggs are affected together with the nutrition of the fertilized eggs and the external hatching environment. After being released from the shell, the meat duck can reach the market weight at about 58 weeks of age. This rapid growth is due to breeding improvement and efficient feeding, but is also closely related to endocrine regulation and epigenetic mechanisms. During the growth of ducks, the number of skeletal muscle fibers is mainly determined during the embryonic stage, while the growth and hypertrophy of muscle fibers are achieved after birth through satellite cell proliferation and fusion. The study found that the critical period of skeletal muscle development in the embryonic stage of duck (such as embryonic age of 13 to 19 days) corresponds to specific microRNA and gene expression peaks, indicating a developmental stage-specific gene regulation network. After the shelling, the muscle and adipose tissue of the duck chicks will change significantly with the increase of day age: the growth rate is the fastest before 4 weeks of age, and then the weight gain gradually slows down and matures (Chen and Li, 2024). In this process, endocrine factors such as the growth hormone (GH)-IGF axis play a role, and the epigenetic state of the gene (such as the methylation level of the growth-related gene promoter) is also changing dynamically (Cong et al., 2023).

2.2 Feather formation and periodic changes

The feather development of ducks has its own unique rules. Ducklings are covered with down feathers when they come out of their shells, which are mainly used for insulation. In subsequent growth, ducks will undergo a replacement process from down feather to child feather and then to adult feather, which is equivalent to the hair replacement cycle of mammals. Studies have shown that ducklings gradually grow primary flying feathers and body covering feathers within a few weeks after birth, and the primary down feathers are replaced with more functional juvenile feathers for simple flight and stronger insulation (Lu et al., 2024). As sexually mature, ducks also develop reproductive plumes, showing seasonal or gender-different feather morphology and color. Periodic growth and defecation of feathers (feathering) usually occurs 1 to 2 times a year in waterfowls such as ducks and others, and is mostly performed after the breeding season. Feather formation is driven by stem cell proliferation and differentiation within feather follicles (Figure 1). Feather development includes processes such as feather bud formation, feather axis and feather sheet differentiation, keratin deposition, etc., which are precisely controlled by a series of gene regulatory pathways, such as Wnt/β-catenin signal promoting feather primordial formation, BMP signal limiting feather spacing, and Shh signal mediating feather branch branches, etc. Feather development processes are also affected by epigenetic regulation. For example, the chromatin open state of the promoter of specific genes of feather stem cells is closely related to its differentiation potential. The color pattern of feathers also depends on the distribution of melanocytes in the embryonic stage and the expression of related genes, which may be affected by epigenetic factors (such as pigment gene promoter methylation, non-coding RNA regulation, etc.) (Twumasi et al., 2024). Therefore, feather development is a dynamic and complex process with clear stages and periodicity, and behind it involves the exquisite regulation of developmental biology and epigenetics.

3 Summary of Epigenetic Regulation Mechanisms

3.1 The role of DNA methylation in gene expression regulation

DNA methylation is one of the earliest epigenetic markers discovered, referring to the addition of methyl groups on the cytosine (C) nucleotide in the DNA sequence, which usually occurs at the cytosine-guanine dinucleotide (CpG) site. On high-density CpG islands in gene promoter regions, methylation often causes transcription factors to fail to bind to DNA, thereby repressing gene transcription (Slawinska et al., 2020). Therefore, DNA methylation is often associated with gene silencing. Studies have shown that increased methylation levels can reduce the expression of muscle growth-related genes, which in turn affects the growth rate of meat poultry. DNA methylation is mediated by DNA methyltransferase (DNMT) family enzymes, in which DNMT1 is mainly responsible for maintaining methylation, and DNMT3A/3B is responsible for creating new methylation. Early in the embryo, most of the genome undergoes a "demethylation-reprogramming" process, which is then remethylated at specific sites in development (Wang and Ibeagha-Awemu, 2021). Different tissues and cell types have specific methylation maps, giving them different gene expression patterns. In poultry, DNA methylation is widely involved in the developmental regulation of muscle, fat, liver and other tissues. Studies have pointed out that changes in feeding environment (such as increased temperature) can lead to an increase in the methylation level of growth hormone receptor (GHR) and IGF1 gene promoter in duck liver, a decrease in gene expression, and ultimately inhibit growth.

3.2 Chromatin remodeling and histone modification

Eukaryotic DNA wraps histones to form chromatin unit nucleosomes, and their structural dynamic changes affect the accessibility of genes. Chromatin remodeling refers to the process of changing the conformation (open or tightening) of chromatin, which is achieved by the hydrolysis of the remodeling complex using ATP. There are two key mechanisms: one is to slide or remove nucleosomes to expose specific DNA regions to facilitate transcription; the other is to covalently modify histones to change the chromatin state. A variety of covalent modifications can occur at the N-terminal tail of histones. Different histone modifications have different effects on gene expression: for example, lysine acetylation at 4 (H3K4ac) and 9 (H3K9ac) of histone H3 are usually associated with active transcription because acetylation neutralizes histone positive charge, weakens DNA-histone binding, and relaxes chromatin; while modifications such as lysine trimethylation at 27 (H3K27me3) of histone H3 are associated with transcriptional repression because they recruit silencing complexes to coagulate chromatin. During development, chromatin remodeling and histone modification closely coordinate to regulate gene spatiotemporal expression patterns. In poultry studies, although data on histone modifications for ducks are limited, similar mechanisms can be speculated based on other species. Studies have found in chicken muscles that inducing exercise can cause increased levels of histone acetylation at certain sites in muscle cells, activate metabolism and growth-related genes. Chromatin status is also affected by the environment and metabolites, such as short-chain fatty acids (metabolism from intestinal flora) that inhibit histone deacetylase, leading to an increase in histone acetylation levels, thereby altering host tissue gene expression (Stoll et al., 2018).

3.3 Functions of non-coding RNA

In addition to the regulation of DNA and chromatin levels, the epigenetic mechanism of RNA levels cannot be ignored. Among them, small molecule non-coding RNA and long-chain non-coding RNA are research hotspots. MiRNA is a small class of about 20 to 24 nucleotides in length. It does not encode proteins and mainly mediates mRNA degradation or blocks translation through complementary binding to the 3’ non-translational region of mRNA (Dunislawska et al., 2022). Each miRNA can regulate the expression of multiple target genes, so miRNAs are involved in the fine regulation of the developmental process. In avians, many miRNAs have been found to be associated with muscle proliferation and differentiation, fat metabolism, and feather development. Studies have confirmed in chicken embryo muscle that overexpression of miR-27b can downregulate the MSTN gene, significantly accelerating myogenocyte proliferation and delaying differentiation (Zhang et al., 2021). In ducks, although specific miRNA function is less studied, it has been reported that miRNA expression profiles in muscle tissues of different developmental stages of duck embryos have been significantly changed, and some miRNAs may act as "promoting factors" or "inhibitor factors" to affect muscle growth. On the other hand, lncRNAs usually have a length of more than 200 nt and do not encode proteins, but can function in a variety of ways, including acting as competitive endogenous RNA sponges to bind miRNA, acting as transcriptional co-regulators, inducing three-dimensional conformational changes in chromatin, etc. lncRNA can also affect muscle and fat development. There have been studies that have identified hundreds of differentially expressed lncRNAs in primitive germ cells in early chicken embryos, which may regulate the development of gonads and germ cells.

4 Epigenetic Regulation in Duck Growth and Development

4.1 Epigenetic regulation of key growth genes

The growth rate and body size of animals are determined by a series of growth axis hormones and growth factor genes, such as growth hormone gene (GH), growth hormone receptor gene (GHR), islet-like growth factor 1 gene (IGF1), etc. The expression of these key genes is not only controlled by regulatory elements on the DNA sequence, but also by epigenetic mechanisms. In ducks, researchers found that differences in growth performance between different breeds and individuals are often accompanied by epigenetic differences in these genes (Xu et al., 2022). A comparison of genome-wide methylation of Shaoxing duck-protection population and breeding population found that 35 differential methylated genes are closely related to production traits such as meat and egg production, including key genes involved in growth and metabolism. This suggests that epigenetic variations may accumulate and fix during long-term breeding, resulting in line differences. In addition to the genetic background, the methylation dynamics of the promoters of endogenous genes in different growth periods will also affect gene expression. IGF1 is a key factor promoting growth. Studies have observed that the methylation level of the IGF1 gene promoter in duck liver changes during the transition stage from the end of embryonic development to the post-hull outbreak, which has an impact on endocrine regulation of post-natal growth. Under the influence of external factors, the epigenetic status of the growth gene will also change.

4.2 The epigenetic mechanism of muscle growth and fat deposition

Skeletal muscle and adipose tissue are important economic traits that affect the production performance of meat ducks, and the growth and development of both are also deeply regulated by epigenetics. In terms of muscle growth, dynamic changes in DNA methylation are highly correlated with the muscle development stage. The study combined genome-wide methylation sequencing and transcriptome analysis to screen out differential methylated regions (DMRs) and genes during skeletal muscle development in ducks during embryonic period, and found that they are enriched in key signaling pathways for muscle formation. This shows that during the process of duck embryonic muscle cells turning from proliferation to differentiation, specific genome sites undergo methylation status changes, which in turn regulates downstream gene networks and affects the formation and maturation of muscle fibers. Secondly, histone modification is also important in regulating muscle growth genes. The promoters of muscle differentiation markers are usually accompanied by an increase in activated histone markers, while cell cycle genes obtain inhibitory markers when differentiated. Such epigenetic "switches" ensure that myocytes exit proliferate and enter differentiation in time. In duck muscle development, similar histone modification regulatory mechanisms can be speculated to exist. Again, non-coding RNAs are involved in epigenetic regulation of muscle development. Many miRNAs are rich in expression in muscle tissue and have specific developmental stages. For example, miR-206, miR-133, etc. are significantly upregulated during the embryonic muscle differentiation period, helping to promote myotubes.

Epigenetics is also a key regulator of fat deposition. The formation of adipose tissue is regulated by peroxisome proliferator-activated receptor (PPAR) pathways, and epigenetics can affect the activity of these genes. From the late stage of duck embryos to fattening, the regulation of expression of lipid metabolism genes in the liver and muscle is associated with changes in DNA methylation/demethylation. Recent studies have parsed the mRNA N6-methyladenine (m6A) modification map of duck embryonic muscle tissue for the first time, and found that m6A RNA methylation can affect fatty acid β oxidation and lipid metabolism-related pathways, thereby regulating fat deposition (Gu et al., 2022). m6A is an emerging epigenetic mechanism that regulates gene function by affecting mRNA stability and translation efficiency, showing an important role in duck muscle and fat development.

4.3 The influence of environmental factors on the epigenetic status of growth

What is unique about epigenetics is its plasticity, and environmental factors can have a lasting impact on gene function through epigenetic pathways. During the growth of a duck, the external environment (including the embryonic and growth stage environment) will "program" its epigenetic state. Embryo phase temperature is a typical factor. Appropriate incubation temperatures are essential for normal development, and temperature deviations from the optimal range can cause an embryonic epigenetic response. Studies have shown that increasing the temperature of avian embryos by 1 °C will change the embryonic development rhythm and induce dynamic changes in DNA methyltransferase gene expression. In duck embryo tests, higher than normal hatching temperatures have been shown to significantly increase the activity of enzymes such as DNMT1 and DNMT3A in the embryo muscles and liver (Yan et al., 2015), indicating that temperature rise activates the embryonic epigenetic enzyme system, which may lead to changes in genome-wide methylation levels. This temperature-induced epigenetic change may have long-term effects on the growth of duck chicks after hatching, such as better heat tolerance but decreased weight gain. Nutritional level is also an important environmental factor. Nutrients (such as folic acid, choline, vitamin B12, methionine, etc.) can provide methyl donors, which directly affects the DNA methylation process. In poultry, both maternal nutrition and nutrient injections during incubation may alter the epigenetic status of the embryo. Stress factors (such as high temperature, density, transportation, etc.) also affect growth through epigenetic effects during the growth period of ducks. Long-term heat stress can lead to imbalance of stress axis hormones in ducks, and it is accompanied by changes in promoter methylation of stress-related genes (such as HSP heat shock protein gene), causing adaptive adjustments to the stress response ability of ducks to only high temperatures (Massimino et al., 2021). In addition, the impact of the microbial environment on growth is becoming increasingly important. Intestinal flora can produce metabolites such as short-chain fatty acids, which can change the histone acetylation state of the host liver and muscle cells by inhibiting histone deacetylation enzymes, thereby regulating the expression of metabolic genes.

5 Epigenetic Regulation of Duck Feather Development

5.1 Gene regulation in feather formation and growth

Feather formation is a highly programmed developmental process that requires precise gene regulation networks. The most in-depth research is the role of various signaling pathways in feather follicle morphogenesis. In chicken feather development models, the Wnt signal is considered as the first signal to initiate feather primordial formation; subsequent Eda (ectodermal development factor) and FGF (fibroblast growth factor) signals induce dermal papillary aggregation; BMP signal acts as antagonist to limit the size and interfoetal width of the primary axis of the feather. The spatial and temporal activation of these gene pathways laid the basic morphology of feathers (Ji et al., 2023). So, how does epigenetic involve it? In the chromatin state of the promoter of feather-related genes, opening or closing will determine whether the gene can be expressed in time. It is speculated that the LEF1 gene promoter may have epigenetic markers such as histone acetylation in the feather bud stage to ensure its transcriptional activity. At the same time, switching of gene expression at different stages may also be achieved through DNA methylation regulation. When a signaling pathway completes its phased operation, the rise in methylation levels of its key gene promoter can silence it, allowing the next pathway to dominate. When feather buds differentiate into feather branches (small branches), some early signals may be required, and the maintenance of this inhibition may involve DNA methylation (Chen et al., 2024). Although there is currently a lack of research on epigenetic maps during duck feather development, it is speculated based on data from other species that many genes that regulate feather morphology are also epigenetic. For example, there was a study comparing the feather tissue expression profiles of different feather color lines of ducks, and it was found that multiple genes that control feather morphology and pigment had expression differences between lines (Figure 2) (Twumasi et al., 2024). These gene expression differences are not entirely caused by mutations in the coding sequence, but are more likely to be caused by epigenetic regulation, such as promoter methylation differences or distal enhancer chromatin accessibility.

5.2 Epigenetic characteristics of feather follicle stem cells

There is a group of stem cells in the feather follicles located in the nipple and base areas of the hair follicle, responsible for the continuous growth and periodic regeneration of the feathers. These stem cells have the potential for multidirectional differentiation and can produce different types of cells such as feather epidermal cells, medulla cells, and feather branches. The fate decision of hair follicle stem cells is closely related to their epigenetic status. When stem cells maintain an undifferentiated state, genes related to proliferation and self-renewal in their genome remain active, while differentiation-related genes are inhibited. This is often reflected in epigenetics, i.e., the widespread presence of histone activation markers and open chromatin structures in stem cells, whereas the promoter of differentiation marker genes carries inhibitory histone modifications (such as H3K27me3) or higher DNA methylation. When hair follicles enter the growth phase and stem cells begin to activate and differentiate into new feathers, their epigenetic map changes. Studies have observed in mammalian hair follicle stem cells that activation of α smooth muscle actin (α-SMA)-positive dermal papillary cells is an important link in the restart of the hair follicle cycle, and this process involves histone demethylation to open up chromatin and express specific genes of dermal papillary stem cells. In duck feather follicles, a similar mechanism can be inferred: when the rest period turns to the growth phase, some inhibitory epigenetic markers in the hair follicle stem cells are removed, and the stem cells are activated and proliferated, inducing the growth of new feathers. In addition to DNA and histone levels, epigenetics of hair follicle stem cells also involve the role of non-coding RNA. There is evidence that some lncRNAs are highly expressed in hair follicle stem cells, which may maintain stem cells undifferentiated by affecting chromatin status (Raghuwanshi et al., 2017). In addition, signals in the hair follicle microenvironment ("indomain") can also alter stem cell behavior through epigenetic. Cell-cell signals (such as BMP, FGF) at the papillary site of feathers can induce acetylation or demethylation of specific gene promoters in stem cells, thereby initiating a differentiation procedure.

5.3 Non-coding RNA regulation of feather morphology and pigmentation deposition

The final morphology and pigmentation of feathers are controlled by multiple genes and are also affected by epigenetic levels such as non-coding RNA. In particular, miRNA and lncRNA have attracted increasing attention in regulating pigment cell function and keratin formation. In terms of feather pigmentation, the synthesis of melanin is determined by enzymes such as tyrosinase (TYR), tyrosinase-related proteins (TRP-1, TRP-2), and its expression in feather follicle melanocytes is subject to complex regulation. Studies have compared the skin transcriptomes of different feather colors in the hybrid offspring of ducks, and found that the expression differences of many pigment-related genes may be caused by upstream non-coding RNA. In particular, long chain non-coding RNAs play the role of "palette" in it. In addition to lncRNA, miRNA is also involved in the formation of feather pattern. It has been reported that in pupil cells with high expression of feather keratin, some miRNAs are downregulated to relieve the inhibition of the keratin gene, thereby ensuring large-scale synthesis of keratin (Dunislawska et al., 2022). If these miRNA levels are abnormal, phenotypes such as fragile feathers and structural abnormalities may appear.

6 Case Analysis

6.1 IGF1 gene methylation and growth performance improvement

IGF1 gene methylation and growth performance have improved. Islet-like growth factor 1 (IGF1) is an important hormone on the growth axis and can promote the growth of bones and muscles. Improving IGF1 expression is often considered conducive to growth performance. However, the expression of the IGF1 gene is not only regulated by pituitary growth hormone, but also by the epigenetic status of its promoter region. A typical case comes from a study on "hot programming" of poultry embryos: Cong et al. (2023) conducted intermittent high-temperature treatment of broiler embryos in the middle of hatching, and found that the growth rate of broiler chickens after hatching high-temperature embryos decreased. Mechanistic analysis showed that the expression of IGF1 gene in the liver of these chickens was significantly lower than that of the control group, while the DNA methylation level in the IGF1 promoter region was significantly increased (Cong et al., 2023). That is to say, the high-temperature embryonic environment induced abnormal methylation of the IGF1 promoter, inhibiting the normal transcription of IGF1, and thus leading to growth restriction. This finding shows that by regulating the epigenetic status of the IGF1 gene, it can affect the growth performance of animals. For ducks, although no detailed studies have been conducted similarly to high temperature treatments, it is speculated that moderate environmental interventions during embryonic stage may regulate duck growth by affecting IGF1 methylation. If the "epigenetic programming" method is used, temperature/nutritional stimulation is given at a critical stage of duck embryo development, reducing the methylation of the IGF1 gene promoter, thereby promoting high IGF1 expression, which may show faster growth and higher feed conversion after incubation.

6.2 miRNA-Mediated MSTN gene expression and muscle development

Myostatin encoded by the myostatin gene (MSTN) is a negative regulator that limits muscle overgrowth. The function of inhibiting MSTN significantly increases muscle mass. At the epigenetic level, the regulation of MSTN expression by microRNA is a classic case. Zhang et al. (2021) found in chicken muscle cells that miR-27b-3p can complement the mRNA of the MSTN gene, thereby inhibiting MSTN translation. They verified through dual fluorescence reporter experiments that MSTN is the direct target gene of miR-27b-3p. Further cellular function assays showed that upregulation of miR-27b-3p significantly promoted the proliferation of primary cells of chicken skeletal muscle (myoblasts) and inhibited their premature differentiation. This is consistent with the effect of inhibiting MSTN: because MSTN usually limits myocyte count through cell cycle inhibition and prodifferentiation signals, miR-27b-mediated decrease in MSTN cancels a portion of the inhibition, and cells therefore proliferate more. This case clearly demonstrates how miRNAs affect muscle development by acting on specific genes. For ducks, given that the function of MSTN is highly conserved in vertebrates, it can be inferred that similar regulatory axes exist in duck muscle cells (Miretti et al., 2013). For example, if the duck's miR-27 family members are highly expressed in skeletal muscle, they are likely to also target MSTN, thereby regulating the growth rate and muscle fiber size of the duck's muscle. Future studies can perform miRNA sequencing on duck muscle tissue, identify miRNAs associated with the MSTN pathway and verify their function. If miRNAs like miR-27b-3p that have significant regulatory effects on MSTN are found in ducks, then increasing the expression of this miRNA through molecular breeding methods or inhibiting the mRNA of MSTN through antisense oligonucleotide technology will be expected to obtain more muscle-developed duck species.

6.3 lncRNA-FeatherLnc regulates feather pigmentation

The color of feathers is determined by the deposition of biopigments (mainly melanin and flavonoids, etc.) in the feathers. The formation process is complex and regulated by multiple genes. In recent years, the role of long-chain non-coding RNA in the regulation of pigment deposition has been gradually revealed. A representative case is skin melanin deposition in black-feathered chicken species. Zhang et al. (2022) identified a lncRNA that plays an important role in the melanin deposition process, named LMEP. In black-feathered chicken skin, LMEP is highly expressed, which promotes upregulation of pigment synthases such as tyrosinase (TYR) through interaction with proteins related to melanin production pathway, resulting in an increase in melanin content (Figure 3).

Figure 3 Histomorphological examination of YS and BS in Xichuan black chicken. Differences in skin melanination and histomorphology between YS (a) and BS (b). Arrows indicate melanin (Adopted from Zhang et al., 2022)

This study provides an example for understanding lncRNA regulating feather/skin pigmentation. Among ornamental duck varieties, lncRNA regulation technology can be used to cultivate new varieties with specific patterns. In production practice, the color of the feather also affects the value of the commodity. For example, all white feathers are beneficial to the appearance of the carcass. If a certain lncRNA is found to inhibit melanin deposition, it can be considered to enhance its expression to cultivate white feather varieties (Li et al., 2024). Of course, feather pigmentation is a complex trait, and it also involves multiple links such as enzyme activity and pigment transport. lncRNA is just one link.

7 External Factors Affecting the Epigenetic Status of Ducks

7.1 Programming epigenetic states by the embryonic environment

7.1.1 Effects of temperature changes on DNA methylation and feather development

Embryo development is highly sensitive to temperature, and temperature changes not only affect development speed, but may also leave "memory" through epigenetic pathways. In duck embryos, it was found that increasing the incubation temperature from 37.8 °C to 38.8 °C will upregulate the expression of methylation-related genes such as DNMT1 and DNMT3A in the early embryo (1 to 10 days) and enhance the activity of DNA methyltransferase. This alteration of enzyme activity means that embryonic DNA methylation may increase, especially in certain gene promoter regions. Accordingly, these altered gene expression programs may affect the phenotype after incubation. For example, duck chicks in the high-temperature treatment group tend to be smaller when they emerge from the shell, with a decrease in growth potential, and may have better heat tolerance. This is because the methylation changes induced by embryos inhibit some growth-promoting genes and activate some stress-responsive genes (Massimino et al., 2021). Temperature also affects the epigenetics of feather development. Conversely, appropriate temperature reduction (cold stimulation) is considered beneficial in some cases: studies have shown that transient cryogenic treatment during the embryo phase can reduce metabolic rates and fat deposition in avian offspring, which may involve epigenetic reprogramming of metabolic regulatory genes, such as increased demethylation of the promoter of PGC-1α gene leads to upregulation of its expression and promotes lipid utilization (Gajendran and Veeramani, 2022).

7.1.2 The epigenetic effect of humidity and light intervention on embryonic development

In addition to temperature, hatching humidity and light conditions are also important components of the embryonic environment and may also affect duck embryo development through epigenetic mechanisms. Humidity mainly affects the weight loss rate and air chamber size of the embryo egg, thereby affecting the respiration and metabolism of the embryo. Under low humidity environments, embryos will be in mild hypoxia and dehydrated states, which may activate the stress response genes of the embryo. The promoters of some hypoxia-responsive genes (such as HIF1A) may undergo demethylation under hypoxic conditions, allowing stronger expression to aid embryo adaptation (Dunislawska et al., 2022). Although this improves hatching or stress resistance, it may also change the development process and affect feather growth. Light is another external signal. Traditional hatching is mostly conducted in the dark, but studies have shown that fertilized eggs do not respond to light during hatching. Especially in the late incubation stage, light can pass through the eggshell to a certain extent to affect the embryo's circadian rhythm and pineal gland development. If a regular light-dark cycle stimulation is given in hatching, the biological clock genes of duck embryos may be "programmed". The promoters of these genes (such as Clock, BMAL1) and related clock-controlled gene regulatory elements may undergo epigenetic changes, resulting in different behavioral and metabolic patterns of ducklings after birth.

7.1.3 Programming effect of nutritional supplementation in embryonic stage on the later representative obsogenetics

Duck embryos rely entirely on nutrients inside the breeding eggs for development. The yolk and protein provided by the mother not only contain a large amount of protein, lipids and vitamins, but also contains some cofactors required for epigenetic regulation (such as methyl donors). Therefore, the nutritional composition of female duck feed directly affects the epigenetic status of the embryo. For example, levels of folic acid, choline, etc. in maternal diets will affect the basal level of embryonic DNA methylation (Nie et al., 2019). In addition to direct nutrient replenishment of embryos through the mother is also a hot topic in research. Injecting specific nutrients (such as sugars, amino acids, vitamins) into eggs during the mid-embryo period can improve the growth and intestinal development of ducklings after hatching (Bodin et al., 2019). The mechanism behind it can be partially attributed to epigenetics. Intraocular arginine supplementation may activate the embryonic mTOR pathway, thereby promoting muscle protein synthesis by changing the histone acetylation status of downstream metabolic genes. Although this type of epigenetic regulation is indirect, it cannot be ignored. The application of some plant extracts and functional additives in the embryonic stage may also bring about epigenetic effects. It should be emphasized that the epigenetic programming role of nutritional supplementation in embryonic stage is "double-edged". Excessive nutrition may lead to early development of embryonic adipose tissue, and offspring have "metabolic memory" and prefer fat deposition; insufficient nutrition may lead to slow growth of offspring through epigenetic epigenetics. Therefore, nutritional regulation should be carried out based on specific goals. For example, to cultivate lean meat ducks, yolk lipid utilization can be moderately limited during the embryo, which promotes increased metabolic rate in offspring - this can be achieved by giving mild stress or metabolic stimulation in the embryo, but should be controlled within a safe range to avoid negative effects.

7.2 Effects of feeding conditions on growth and feather development

As an important external factor affecting duck growth and feather development, feeding conditions act on epigenetic mechanisms through nutrient supply, environmental stress and management measures, thereby regulating gene expression and phenotypic expression. Changes in feed composition have a significant effect on the growth performance and feather quality of ducks. Studies have shown that methionine supplementation in feed can improve the growth rate and feather quality of ducks, especially during brooding (1 to 14 days old) and 35 days old (Zeng et al., 2015). Methionine, as a methyl donor, may regulate the expression of genes related to growth and feather development by affecting DNA methylation levels. Second, stress factors such as temperature fluctuations and transportation also have significant effects on the epigenetic status of ducks. Research has found that environmental stress can change epigenetic markers by activating stress-related signaling pathways, thereby affecting the expression of growth-related genes. In addition, drinking water management and microecological intervention are also considered important means to regulate the epigenetic status of ducks. Research points out that adding probiotics to drinking water or regulating water quality can affect the composition of the intestinal microbiota, and then affect the epigenetic regulatory mechanism through the intestinal-brain-axial pathway, promoting duck growth and feather development.

8 Commentary Summary

8.1 The importance of epigenetic regulation in duck growth and feather development

The epigenetic regulatory mechanism runs through the entire growth and feather development process of ducks, and has an important impact on gene function and trait performance. Compared with traditional genetic variation, epigenetics is more dynamically plastic, which allows ducks to make adaptive adjustments in the face of environmental changes and also provides possible ways for artificial intervention. In terms of growth, DNA methylation and non-coding RNA regulate the expression of key growth genes (such as IGF1 and MSTN), which directly affects the growth rate and body composition of ducks. In terms of feather development, chromatin state and long-chain non-coding RNA determine the degree of differentiation and pigmentation of feather stem cells, shaping the rich and diverse feather morphology of ducks (Zhang et al., 2022a; Lu et al., 2024). It can be said that without epigenetic participation, it is impossible to fully explain the complex process of ducks developing from fertilized eggs to mature individuals. For ducks, their production performance (growth rate, feed conversion efficiency, carcass quality) and appearance characteristics (feather color, feather quality) are all epigenetically regulated to varying degrees. Therefore, in poultry genetic breeding and breeding management, the important role of epigenetic factors should be fully paid attention to.

8.2 Necessity and significance of deepening research

The current epigenetic research on ducks is still relatively limited, far less in-depth than that of chickens and other models of birds. However, as shown in this review, some preliminary evidence has revealed the existence of epigenetic regulation of ducks and its effects on productive traits. In order to better utilize this knowledge to serve the duck raising industry, related research must be further deepened. On the one hand, it is necessary to draw an epigenetic map of the entire genome of duck, including DNA methylation groups, histone modification groups and open chromatin regions in different tissues and different periods, to find key regulatory nodes. This helps identify epigenetic markers closely related to growth and feather traits, providing new indicators for breeding. On the other hand, functional studies should be carried out to verify the causal relationship of specific epigenetic modifications on duck traits through gene editing or pharmacological means. For example, using gene editing technology to knock out specific miRNAs in duck embryos and observe their effects on muscle and feather development can clarify the function of this miRNA in epigenetic regulation. These in-depth research will deepen our understanding of duck developmental biology and also provide new tools and new ideas for cultivating high-yield and high-quality duck species.

8.3 Future development direction and cooperation initiatives

The epigenetic research of ducks is in its infancy and requires multidisciplinary collaboration and new technology application. Future research directions include but are not limited to: multiomics integrated analysis: combining genome, transcriptome and epigeneticomic data to build a complex network model for the regulation of duck growth and development, and identify key regulatory factors. Interaction between transgenerational genetics and epigenetics: Study the cumulative effect of epigenetic variation in poultry breeding selection, explore whether certain epigenetic markers can serve as auxiliary breeding indicators, and how epigenetic interacts with gene mutations to influence traits. Environmental-Epigenetic Interaction Research: Systematically evaluate the changes in epigenetic markers of ducks under different environmental conditions, find beneficial variants and try to solidify these variants to improve breeds. To achieve the above goals, scientific research institutions, breeding enterprises and breeding units should strengthen cooperation. At the basic research level, a duck epigenetic research alliance can be established to share duck genomic resources and epigenetic data, strengthen communication with researchers of model species such as chickens, and learn from research experience. At the application level, industrial experimental cooperation can be carried out, such as the company provides experimental scenarios for different feeding conditions, and scientific researchers conduct biological sample collection and epigenetic analysis to screen out best practice plans. Only through the combination of industry, academia and research can laboratory discoveries be converted into productivity in the farm faster.

Acknowledgements

The authors gratefully acknowledge the support provided by Cai R.X. and thank the two peer reviewers for their suggestions.

Conflict of Interest Disclosure

The authors confirm that the study was conducted without any commercial or financial relationships and could be interpreted as a potential conflict of interest.

References

Bodin L., Secula A., Chapuis H., Cornuez A., Lessire M., Cobo E., Marie-Louise S., Bonnefont C., Barrieu J., Mercerand F., Bravo C., Manse H., Bourhis M., Martin X., Pitel F., Brun J., and Morisson M., 2019, Dietary methionine deficiency reduces laying performances of female common ducks and impacts traits of interest of their mule ducklings, Poultry Science, 98(9): 4035-4045.

https://doi.org/10.3382/ps/pez315

Chen C., Chang Y., Jiang T., Yue Z., Liu T., Lu J., Yu Z., Lin J., Vu T., Huang T., Harn H., Ng C., Wu P., Chuong C., and Li W., 2024, Conserved regulatory switches for the transition from natal down to juvenile feather in birds, Nature Communications, 15: 48303.

https://doi.org/10.1038/s41467-024-48303-3

Chen X., Ge K., Wang M., Zhang C., and Geng Z., 2017, Integrative analysis of the Pekin duck (Anas anas) microRNAome during feather follicle development, BMC Developmental Biology, 17(1): 12.

https://doi.org/10.1186/s12861-017-0153-1

Chen J., and Li X., 2024, Delayed plumage maturation in juvenile males of the white-bellied shortwing (Brachypteryx leucophrys), International Journal of Molecular Zoology, 14(3): 141-153.

Cong W., Han W., Liu J., Zhu J., Liu X., and Yang M., 2023, Embryonic thermal manipulation leads growth inhibition and reduced hepatic insulin-like growth factor1 expression due to promoter DNA hypermethylation in broilers, Poultry Science, 102(4): 102562.

Dunislawska A., Pietrzak E., Sławińska A., Siwek M., and Bednarczyk M., 2022, Pre-hatching and post-hatching environmental factors related to epigenetic mechanisms in poultry, Animals, 12(3): 332.

https://doi.org/10.3390/ani12030332

Gajendran K., and Veeramani P., 2022, Intensive Duck Rearing, in Duck Production and Management Strategies, Springer.

https://doi.org/10.1007/978-981-16-6100-6_7

Gu L., Zhang S., Li B., Jiang Q., Xu T., Huang Y., Lin D., Xing M., Huang L., Zheng X., Wang F., Chao Z., and Sun W., 2022, m6A and miRNA jointly regulate the development of breast muscles in duck embryonic stages, Frontiers in Veterinary Science, 9: 933850.

https://doi.org/10.3389/fvets.2022.933850

Huang Z., Hu L., Liu Z., Jin J., Xu Z., Zuo B., and Zhu Q., 2025, The functions and regulatory mechanisms of histone modifications in skeletal muscle development and disease, International Journal of Molecular Sciences, 26(8): 3644.

https://doi.org/10.3390/ijms26083644

Ji G., Zhang M., Tu Y., Liu Y., Shan Y., Ju X., Zou J., Shu J., Sheng Z., and Li H., 2023, Molecular regulatory mechanisms in chicken feather follicle morphogenesis, Genes, 14(8): 1646.

https://doi.org/10.3390/genes14081646

Li R., Li D., Xu S., Zhang P., Zhang Z., He F., Li W., Sun G., Jiang R., Li Z., Tian Y., Liu X., and Kang X., 2024, Whole-transcriptome sequencing reveals a melanin-related ceRNA regulatory network in the breast muscle of Xichuan black-bone chicken, Poultry Science, 103: 103539.

https://doi.org/10.1016/j.psj.2024.103539

Lu J., Yu Z., Lin J., Chang Y.M., Jiang T.X., Yue Z., Liu T.Y., Vu T.D., Huang T.Y., Harn H.I., Ng C.S., Wu P., Chuong C.M., and Li W.H., 2024, Conserved regulatory switches for the transition from natal down to juvenile feather in birds, Nature Communications, 15: 4174.

https://doi.org/10.1038/s41467-024-48303-3

Lu Y., Zhou J., Li F., Cao H., Zhang X., Yu D., He Z., Ji H., Lv K., Wu G., and Yu M., 2023, The integration of genome-wide DNA methylation and transcriptomics identifies the potential genes that regulate the development of skeletal muscles in ducks, International Journal of Molecular Sciences, 24(20): 15476.

https://doi.org/10.3390/ijms242015476

Massimino W., Andrieux C., Biasutti S., Chartrin P., Métayer-Coustard S., Marty-Gasset N., Diot C., Davail S., and Hérault F., 2021, Impacts of embryonic thermal programming on the expression of genes involved in foie gras production in mule ducks, Frontiers in Physiology, 12: 779689.

https://doi.org/10.3389/fphys.2021.779689

Miretti S., Martignani E., Accornero P., and Baratta M., 2013, Functional effect of miR‑27b on myostatin expression: a relationship in Piedmontese cattle with double‑muscled phenotype, BMC Genomics, 14: 194.

https://doi.org/10.1186/1471-2164-14-194

Nie Q., Sun Y., and Lee J., 2019, DNA methylation in poultry: a review, Livestock Science, 220: 72-80.

https://doi.org/10.1016/j.livsci.2018.12.005

Raghuwanshi S., Dahariya S., Kandi R., Gutti U., Undi R., Sharma D., Sahu I., Kovuru N., Yarla N., Saladi R., and Gutti R., 2017, Epigenetic mechanisms: role in hematopoietic stem cell lineage commitment and differentiation, Current Drug Targets, 19(14): 1683-1695.

https://doi.org/10.2174/1389450118666171122141821

Sepers B., van den Heuvel K., Lindner M., Viitaniemi H., Husby A., and van Oers K., 2019, Avian ecological epigenetics: pitfalls and promises, Journal of Ornithology, 160(4): 1183-1203.

https://doi.org/10.1007/s10336-019-01684-5

Slawinska A., Dunislawska A., and Siwek M., 2020, Hepatic DNA methylation in response to early stimulation of microbiota with Lactobacillus synbiotics in broiler chickens, Genes, 11(5): 579.

https://doi.org/10.3390/genes11050579

Stoll J., He S., and Lagana S., 2018, Microbiota derived short chain fatty acids promote histone crotonylation in the colon, Nature, 563(7731): 17-21.

https://doi.org/10.1038/s41586-018-0634-8

Twumasi G., Wang H., Xi Y., Zhao X., Chen Y., Li J., Zhang Z., and Wu Q., 2024, Genome-wide association studies reveal candidate genes associated with pigmentation patterns of single feathers of Tianfu Nonghua ducks, Animals, 14(1): 85.

https://doi.org/10.3390/ani14010085

Wang M., and Ibeagha‑Awemu E. M., 2021, Impacts of epigenetic processes on the health and productivity of livestock, Frontiers in Genetics, 11: 613636.

https://doi.org/10.3389/fgene.2020.613636

Xu L., Shi Z., Li H., Zhang Y., Wang X., and Chen Q., 2022, Genome-wide DNA methylation differences between conservation and breeding populations of Shaoxing ducks, Heliyon, 8(11): e11555.

https://doi.org/10.1016/j.heliyon.2022.e11555

Yan X. P., Liu H. H., Liu J. Y., Chen Z. Q., and Sun B. L., 2015, Evidence in duck for supporting alteration of incubation temperature may have influence on methylation of genomic DNA, Poultry Science, 94(10): 2537-2545.

https://doi.org/10.3382/ps/pev217

Zeng Q., Zhang Q., Chen X., Doster A., Murdoch R., Makagon M., Gardner A., and Applegate T., 2015, Effect of dietary methionine content on growth performance, carcass traits, and feather growth of Pekin duck from 15 to 35 days of age, Poultry Science, 94(7): 1592-1599.

https://doi.org/10.3382/ps/pev117

Zhang G., He M., Wu P., Zhang X., Zhou K., Li T., Zhang T., Xie K., Dai G., and Wang J., 2021, MicroRNA‑27b‑3p targets the myostatin gene to regulate myoblast proliferation and is involved in myoblast differentiation, Cells, 10(2): 423.

https://doi.org/10.3390/cells10020423

Zhang P., Cao Y., Fu Y., Zhu H., Xu S., Zhang Y., Li W., Sun G., Jiang R., Han R., Li H., Li G., Tian Y., Liu X., Kang X., and Li D., 2022, Revealing the regulatory mechanism of lncRNA‑LMEP on melanin deposition based on high‑throughput sequencing in Xichuan chicken skin, Genes, 13(11): 2143.

https://doi.org/10.3390/genes13112143