Long noncoding RNAs and mRNAs profiling in ovary during laying and broodiness in Taihe Black-Bone Silky Fowls (Gallus gallus Domesticus Brisson).

BACKGROUND: Broodiness significantly impacts poultry egg production, particularly notable in specific breeds such as the black-boned Silky, characterized by pronounced broodiness. An understanding of the alterations in ovarian signaling is essential for elucidating the mechanisms that influence broodiness. However, comparative research on the characteristics of long non-coding RNAs (lncRNAs) in the ovaries of broody chickens (BC) and high egg-laying chickens (GC) remains scant. In this investigation, we employed RNA sequencing to assess the ovarian transcriptomes, which include both lncRNAs and mRNAs, in eight Taihe Black-Bone Silky Fowls (TBsf), categorized into broody and high egg-laying groups. This study aims to provide a clearer understanding of the genetic underpinnings associated with broodiness and egg production. RESULTS: We have identified a total of 16,444 mRNAs and 18,756 lncRNAs, of which 349 mRNAs and 651 lncRNAs exhibited significantly different expression (DE) between the BC and GC groups. Furthermore, we have identified the cis-regulated and trans-regulated target genes of differentially abundant lncRNA transcripts and have constructed an lncRNA-mRNA trans-regulated interaction network linked to ovarian follicle development. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) annotation analyses have revealed that DE mRNAs and the target genes of DE lncRNAs are associated with pathways including neuroactive ligand-receptor interaction, CCR6 chemokine receptor binding, G-protein coupled receptor binding, cytokine-cytokine receptor interaction, and ECM-receptor interaction. CONCLUSION: Our research presents a comprehensive compilation of lncRNAs and mRNAs linked to ovarian development. Additionally, it establishes a predictive interaction network involving differentially abundant lncRNAs and differentially expressed genes (DEGs) within TBsf. This significantly contributes to our understanding of the intricate interactions between lncRNAs and genes governing brooding behavior.

Robust identification of interactions between heat-stress responsive genes in the chicken brain using Bayesian networks and augmented expression data.

Bayesian networks represent a useful tool to explore interactions within biological systems. The aims of this study were to identify a reduced number of genes associated with a stress condition in chickens (Gallus gallus) and to unravel their interactions by implementing a Bayesian network approach. Initially, one publicly available dataset (3 control vs. 3 heat-stressed chickens) was used to identify the stress signal, represented by 25 differentially expressed genes (DEGs). The dataset was augmented by looking for the 25 DEGs in other four publicly available databases. Bayesian network algorithms were used to discover the informative relationships between the DEGs. Only ten out of the 25 DEGs displayed interactions. Four of them were Heat Shock Proteins that could be playing a key role, especially under stress conditions, where maintaining the correct functioning of the cell machinery might be crucial. One of the DEGs is an open reading frame whose function is yet unknown, highlighting the power of Bayesian networks in knowledge discovery. Identifying an initial stress signal, augmenting it by combining other databases, and finally learning the structure of Bayesian networks allowed us to find genes closely related to stress, with the possibility of further exploring the system in future studies.

Genetic architecture of white striping in turkeys (Meleagris gallopavo).

White striping (WS) is a myopathy of growing concern to the turkey industry. It is rising in prevalence and has negative consequences for consumer acceptance and the functional properties of turkey meat. The objective of this study was to conduct a genome-wide association study (GWAS) and functional analysis on WS severity. Phenotypic data consisted of white striping scored on turkey breast fillets (N = 8422) by trained observers on a 0-3 scale (none to severe). Of the phenotyped birds, 4667 genotypic records were available using a proprietary 65 K single nucleotide polymorphism (SNP) chip. The SNP effects were estimated using a linear mixed model with a 30-SNP sliding window approach used to express the percentage genetic variance explained. Positional candidate genes were those located within 50 kb of the top 1% of SNP windows explaining the most genetic variance. Of the 95 positional candidate genes, seven were further classified as functional candidate genes because of their association with both a significant gene ontology and molecular function term. The results of the GWAS emphasize the polygenic nature of the trait with no specific genomic region contributing a large portion to the overall genetic variance. Significant pathways relating to growth, muscle development, collagen formation, circulatory system development, cell response to stimulus, and cytokine production were identified. These results help to support published biological associations between WS and hypoxia and oxidative stress and provide information that may be useful for future-omics studies in understanding the biological associations with WS development in turkeys.

Temporal dynamics of genetic parameters and SNP effects for performance and disorder traits in poultry undergoing genomic selection.

Accurate genetic parameters are crucial for predicting breeding values and selection responses in breeding programs. Genetic parameters change with selection, reducing additive genetic variance and changing genetic correlations. This study investigates the dynamic changes in genetic parameters for residual feed intake (RFI), gain (GAIN), breast percentage (BP), and femoral head necrosis (FHN) in a broiler population that undergoes selection, both with and without the use of genomic information. Changes in single nucleotide polymorphism (SNP) effects were also investigated when including genomic information. The dataset containing 200,093 phenotypes for RFI, 42,895 for BP, 203,060 for GAIN, and 63,349 for FHN was obtained from 55 mating groups. The pedigree included 1,252,619 purebred broilers, of which 154,318 were genotyped with a 60K Illumina Chicken SNP BeadChip. A Bayesian approach within the GIBBSF90 + software was applied to estimate the genetic parameters for single-, two-, and four-trait models with sliding time intervals. For all models, we used genomic-based (GEN) and pedigree-based approaches (PED), meaning with or without genotypes. For GEN (PED), heritability varied from 0.19 to 0.2 (0.31 to 0.21) for RFI, 0.18 to 0.11 (0.25 to 0.14) for GAIN, 0.45 to 0.38 (0.61 to 0.47) for BP, and 0.35 to 0.24 (0.53 to 0.28) for FHN, across the intervals. Changes in genetic correlations estimated by GEN (PED) were 0.32 to 0.33 (0.12 to 0.25) for RFI-GAIN, -0.04 to -0.27 (-0.18 to -0.27) for RFI-BP, -0.04 to -0.07 (-0.02 to -0.08) for RFI-FHN, -0.04 to 0.04 (0.06 to 0.2) for GAIN-BP, -0.17 to -0.06 (-0.02 to -0.01) for GAIN-FHN, and 0.02 to 0.07 (0.06 to 0.07) for BP-FHN. Heritabilities tended to decrease over time while genetic correlations showed both increases and decreases depending on the traits. Similar to heritabilities, correlations between SNP effects declined from 0.78 to 0.2 for RFI, 0.8 to 0.2 for GAIN, 0.73 to 0.16 for BP, and 0.71 to 0.14 for FHN over the eight intervals with genomic information, suggesting potential epistatic interactions affecting genetic trait architecture. Given rapid genetic architecture changes and differing estimates between genomic and pedigree-based approaches, using more recent data and genomic information to estimate variance components is recommended for populations undergoing genomic selection to avoid potential biases in genetic parameters.

Genetic parameters are used to predict breeding values for individuals in breeding programs undergoing selection. However, inaccurate genetic parameters can cause breeding values to be biased, and genetic parameters can change over time due to multiple factors. This study aimed to investigate how genetic parameters changed over time in a broiler population using time intervals and observing the behavior of single nucleotide polymorphism (SNP) effects. We studied four traits related to production and disorders while also studying the impact of using genomic information on the estimates. Genetic variances showed an overall decreasing trend, whereas residual variances increased during each interval, resulting in decreasing heritability estimates. Genetic correlations between traits varied but with no major changes over time. Estimates tended to be lower when genomic information was included in the analysis. SNP effects showed changes over time, indicating changes to the genetic background of this population. Using outdated variance components in a population under selection may not represent the current population. Furthermore, when genomic selection is practiced, accounting for this information while estimating variance components is important to avoid biases.

Genome-wide association study of growth curve parameters reveals novel genomic regions and candidate genes associated with metatarsal bone traits in chickens.

The growth and development of chicken bones have an enormous impact on the health and production performance of chickens. However, the development pattern and genetic regulation of the chicken skeleton are poorly understood. This study aimed to evaluate metatarsal bone growth and development patterns in chickens via non-linear models, and to identify the genetic determinants of metatarsal bone traits using a genome-wide association study (GWAS) based on growth curve parameters. Data on metatarsal length (MeL) and metatarsal circumference (MeC) were obtained from 471 F2 chickens (generated by crossing broiler sires, derived from a line selected for high abdominal fat, with Baier layer dams) at 4, 6, 8, 10, and 12 weeks of age. Four non-linear models (Gompertz, Logistic, von Bertalanffy, and Brody) were used to fit the MeL and MeC growth curves. Subsequently, the estimated growth curve parameters of the mature MeL or MeC (A), time-scale parameter (b), and maturity rate (K) from the non-linear models were utilized as substitutes for the original bone data in GWAS. The Logistic and Brody models displayed the best goodness-of-fit for MeL and MeC, respectively. Single-trait and multi-trait GWASs based on the growth curve parameters of the Logistic and Brody models revealed 4 618 significant single nucleotide polymorphisms (SNPs), annotated to 332 genes, associated with metatarsal bone traits. The majority of these significant SNPs were located on Gallus gallus chromosome (GGA) 1 (167.433-176.318 Mb), GGA2 (96.791-103.543 Mb), GGA4 (65.003-83.104 Mb) and GGA6 (64.685-95.285 Mb). Notably, we identified 12 novel GWAS loci associated with chicken metatarsal bone traits, encompassing 35 candidate genes. In summary, the combination of single-trait and multi-trait GWASs based on growth curve parameters uncovered numerous genomic regions and candidate genes associated with chicken bone traits. The findings benefit an in-depth understanding of the genetic architecture underlying metatarsal growth and development in chickens.

Archaeological and molecular evidence for ancient chickens in Central Asia.

The origins and dispersal of the chicken across the ancient world remains one of the most enigmatic questions regarding Eurasian domesticated animals. The lack of agreement concerning timing and centers of origin is due to issues with morphological identifications, a lack of direct dating, and poor preservation of thin, brittle bird bones. Here we show that chickens were widely raised across southern Central Asia from the fourth century BC through medieval periods, likely dispersing along the ancient Silk Road. We present archaeological and molecular evidence for the raising of chickens for egg production, based on material from 12 different archaeological sites spanning a millennium and a half. These eggshells were recovered in high abundance at all of these sites, suggesting that chickens may have been an important part of the overall diet and that chickens may have lost seasonal egg-laying.

MiRNA-21-5p induces chicken hepatic lipogenesis by targeting NFIB and KLF3 to suppress the PI3K/AKT signaling pathway.

The liver plays a critical role in metabolic activity and is the body's first immune barrier, and maintaining liver health is particularly important for poultry production. MicroRNAs (miRNAs) are involved in a wide range of biological activities due to their capacity as posttranscriptional regulatory elements. A growing body of research indicates that miR-21-5p plays a vital role as a modulator of liver metabolism in various species. However, the effect of miR-21-5p on the chicken liver is unclear. In the current study, we discovered that the fatty liver had high levels of miR-21-5p. Then the qPCR, Western blot, flow cytometry, enzyme-linked immunosorbent assay, dual-luciferase, and immunofluorescence assays were, respectively, used to determine the impact of miR-21-5p in the chicken liver, and it turned out that miR-21-5p enhanced lipogenesis, oxidative stress, and inflammatory responses, which ultimately induced hepatocyte apoptosis. Mechanically, we verified that miR-21-5p can directly target nuclear factor I B (NFIB) and kruppel-like factor 3 (KLF3). Furthermore, our experiments revealed that the suppression of NFIB promoted apoptosis and inflammation, and the KLF3 inhibitor accelerated lipogenesis and enhanced oxidative stress. Furthermore, the cotransfection results suggest that the PI3K/AKT pathway is also involved in the process of miRNA-21-5p-mediate liver metabolism regulation. In summary, our study demonstrated that miRNA-21-5p plays a role in hepatocyte lipogenesis, oxidative stress, inflammation, and apoptosis, via targeting NFIB and KLF3 to suppress the PI3K/AKT signal pathway in chicken.

miR-21-5p is a typical noncoding RNA that could inhibit messenger RNA expression by targeting the 3ʹ-untranslated region to participate in fatty liver-related disease formation and progression. We demonstrated that miRNA-21-5p plays a role in hepatocyte lipogenesis, oxidative stress, inflammation, and apoptosis, via targeting nuclear factor I B and kruppel-like factor 3 to suppress the PI3K/AKT signal pathway in chicken. This research established the regulatory network mechanisms of miR-21-5p in chicken hepatic lipogenesis and fatty liver syndrome.

Large-scale genome-wide SNP analysis reveals the rugged (and ragged) landscape of global ancestry, phylogeny, and demographic history in chicken breeds. / å¤§è§„æ¨¡å ¨åŸºå› ç»„SNPåˆ†æžæ­ç¤ºäº†é¸¡å“ç§çš„å ¨çƒç¥–å ˆã€ç§ç¾¤å‘å±•å’Œç§ç¾¤åŽ†å²çš„å¤æ‚(å’Œå¤šæ ·)çš„é—ä¼ å›¾è°±.

The worldwide chicken gene pool encompasses a remarkable, but shrinking, number of divergently selected breeds of diverse origin. This study was a large-scale genome-wide analysis of the landscape of the complex molecular architecture, genetic variability, and detailed structure among 49 populations. These populations represent a significant sample of the world's chicken breeds from Europe (Russia, Czech Republic, France, Spain, UK, etc.), Asia (China), North America (USA), and Oceania (Australia). Based on the results of breed genotyping using the Illumina 60K single nucleotide polymorphism (SNP) chip, a bioinformatic analysis was carried out. This included the calculation of heterozygosity/homozygosity statistics, inbreeding coefficients, and effective population size. It also included assessment of linkage disequilibrium and construction of phylogenetic trees. Using multidimensional scaling, principal component analysis, and ADMIXTURE-assisted global ancestry analysis, we explored the genetic structure of populations and subpopulations in each breed. An overall 49-population phylogeny analysis was also performed, and a refined evolutionary model of chicken breed formation was proposed, which included egg, meat, dual-purpose types, and ambiguous breeds. Such a large-scale survey of genetic resources in poultry farming using modern genomic methods is of great interest both from the viewpoint of a general understanding of the genetics of the domestic chicken and for the further development of genomic technologies and approaches in poultry breeding. In general, whole genome SNP genotyping of promising chicken breeds from the worldwide gene pool will promote the further development of modern genomic science as applied to poultry.

Adaptive expansion of ERVK solo-LTRs is associated with Passeriformes speciation events.

Endogenous retroviruses (ERVs) are ancient retroviral remnants integrated in host genomes, and commonly deleted through unequal homologous recombination, leaving solitary long terminal repeats (solo-LTRs). This study, analysing the genomes of 362 bird species and their reptilian and mammalian outgroups, reveals an unusually higher level of solo-LTRs formation in birds, indicating evolutionary forces might have purged ERVs during evolution. Strikingly in the order Passeriformes, and especially the parvorder Passerida, endogenous retrovirus K (ERVK) solo-LTRs showed bursts of formation and recurrent accumulations coinciding with speciation events over past 22 million years. Moreover, our results indicate that the ongoing expansion of ERVK solo-LTRs in these bird species, marked by high transcriptional activity of ERVK retroviral genes in reproductive organs, caused variation of solo-LTRs between individual zebra finches. We experimentally demonstrated that cis-regulatory activity of recently evolved ERVK solo-LTRs may significantly increase the expression level of ITGA2 in the brain of zebra finches compared to chickens. These findings suggest that ERVK solo-LTRs expansion may introduce novel genomic sequences acting as cis-regulatory elements and contribute to adaptive evolution. Overall, our results underscore that the residual sequences of ancient retroviruses could influence the adaptive diversification of species by regulating host gene expression.

Discrimination of distinct chicken M cell subsets based on CSF1R expression.

In mammals, a subset of follicle-associated epithelial (FAE) cells, known as M cells, conduct the transcytosis of antigens across the epithelium into the underlying lymphoid tissues. We previously revealed that M cells in the FAE of the chicken lung, bursa of Fabricius (bursa), and caecum based on the expression of CSF1R. Here, we applied RNA-seq analysis on highly enriched CSF1R-expressing bursal M cells to investigate their transcriptome and identify novel chicken M cell-associated genes. Our data show that, like mammalian M cells, those in the FAE of the chicken bursa also express SOX8, MARCKSL1, TNFAIP2 and PRNP. Immunohistochemical analysis also confirmed the expression of SOX8 in CSF1R-expressing cells in the lung, bursa, and caecum. However, we found that many other mammalian M cell-associated genes such as SPIB and GP2 were not expressed by chicken M cells or represented in the chicken genome. Instead, we show bursal M cells express high levels of related genes such as SPI1. Whereas our data show that bursal M cells expressed CSF1R-highly, the M cells in the small intestine lacked CSF1R and both expressed SOX8. This study offers insights into the transcriptome of chicken M cells, revealing the expression of CSF1R in M cells is tissue-specific.

Epigenetic regulation of H3K27me3 in laying hens with fatty liver hemorrhagic syndrome induced by high-energy and low-protein diets.

BACKGROUND: Fatty liver hemorrhagic syndrome (FLHS) in the modern poultry industry is primarily caused by nutrition. Despite encouraging progress on FLHS, the mechanism through which nutrition influences susceptibility to FLHS is still lacking in terms of epigenetics. RESULTS: In this study, we analyzed the genome-wide patterns of trimethylated lysine residue 27 of histone H3 (H3K27me3) enrichment by chromatin immunoprecipitation-sequencing (ChIP-seq), and examined its association with transcriptomes in healthy and FLHS hens. The study results indicated that H3K27me3 levels were increased in the FLHS hens on a genome-wide scale. Additionally, H3K27me3 was found to occupy the entire gene and the distant intergenic region, which may function as silencer-like regulatory elements. The analysis of transcription factor (TF) motifs in hypermethylated peaks has demonstrated that 23 TFs are involved in the regulation of liver metabolism and development. Transcriptomic analysis indicated that differentially expressed genes (DEGs) were enriched in fatty acid metabolism, amino acid, and carbohydrate metabolism. The hub gene identified from PPI network is fatty acid synthase (FASN). Combined ChIP-seq and transcriptome analysis revealed that the increased H3K27me3 and down-regulated genes have significant enrichment in the ECM-receptor interaction, tight junction, cell adhesion molecules, adherens junction, and TGF-beta signaling pathways. CONCLUSIONS: Overall, the trimethylation modification of H3K27 has been shown to have significant regulatory function in FLHS, mediating the expression of crucial genes associated with the ECM-receptor interaction pathway. This highlights the epigenetic mechanisms of H3K27me3 and provides insights into exploring core regulatory targets and nutritional regulation strategies in FLHS.

Arginine alleviates Clostridium perfringens α toxin-induced intestinal injury in vivo and in vitro via the SLC38A9/mTORC1 pathway.

Introduction: Clostridium perfringens α toxin is a main virulence factor responsible for gut damage in animals. Arginine is a functional amino acid exhibiting significant immunoregulatory activities. However, the effects and immunoregulatory mechanisms of arginine supplementation on α toxin-induced intestinal injury remain unclear. Methods: In vivo, 256 male Arbor Acres chickens were randomly assigned to a 2×2 factorial arrangement, involving diet treatments (with or without 0.3% arginine supplementation) and immunological stress (with or without α toxin challenge). In vitro, IEC-6 cells were treated with or without arginine in the presence or absence of α toxin. Moreover, IEC-6 cells were transfected with siRNA targeting mTOR and SLC38A9 to explore the underlying mechanisms. Results and discussion: The results showed that in vivo, arginine supplementation significantly alleviated the α toxin-induced growth performance impairment, decreases in serum immunoglobulin (Ig)A and IgG levels, and intestinal morphology damage. Arginine supplementation also significantly reduced the α toxin-induced increase in jejunal proinflammatory cytokines interleukin (IL)-1ß, IL-6 and IL-17 mRNA expression. Clostridium perfringens α toxin significantly decreased jejunal mechanistic target of rapamycin (mTOR) and solute carrier family 38 member 9 (SLC38A9) mRNA expression, while arginine supplementation significantly increased mTOR and SLC38A9 mRNA expression. In vitro, arginine pretreatment mitigated the α toxin-induced decrease in cell viability and the increase in cytotoxicity and apoptosis. Arginine pretreatment also alleviated the α toxin-induced upregulation of mRNA expression of inflammation-related cytokines IL-6, C-X-C motif chemokine ligand (CXCL)10, CXCL11 and transforming growth factor-ß (TGF-ß), as well as apoptosis-related genes B-cell lymphoma-2 associated X protein (Bax), B-cell lymphoma-2 (Bcl-2), B-cell lymphoma-extra large (Bcl-XL) and cysteinyl aspartate specific proteinase 3 (Caspase-3) and the ratio of Bax to Bcl-2. Arginine pretreatment significantly increased the α toxin-induced decrease in mTOR, SLC38A9, eukaryotic translation initiation factor 4E (eIF4E)-binding protein 1 (4EBP1) and ribosomal protein S6 kinase (S6K) mRNA expression. Knockdown SLC38A9 and mTOR largely abrogated the positive effects of arginine pretreatment on α toxin-induced intracellular changes. Furthermore, SLC38A9 silencing abolished the increased mTOR mRNA expression caused by arginine pretreatment. In conclusion, arginine administration attenuated α toxin-induced intestinal injury in vivo and in vitro, which could be associated with the downregulation of inflammation via regulating SLC38A9/mTORC1 pathway.

Molecular characterization of the meq oncogene of Marek's disease virus in vaccinated Brazilian poultry farms reveals selective pressure on prevalent strains.

Marek's disease virus (MDV) has become an increasingly virulent pathogen in the poultry industry despite vaccination efforts to control it. Brazil has experienced a significant rise of Marek's disease (MD) outbreaks in recent years. Our study aimed to analyze the complete meq gene sequences to understand the molecular epidemiological basis of MD outbreaks in Brazilian vaccinated layer farms. We detected a high incidence rate of visceral MD (67.74%) and multiple circulating MDV strains. The most prevalent and geographically widespread genotype presented several clinical and molecular characteristics of a highly virulent strain and evolving under positive selective pressure. Phylogenetic and phylogeographic analysis revealed a closer relationship with strains from the USA and Japan. This study sheds light on the circulation of MDV strains capable of infecting vaccinated birds. We emphasize the urgency of adopting preventive measures to manage MDV outbreaks threatening the poultry farming industry.

Effect of heat stress on the hypothalamic expression profile of water homeostasis-associated genes in low- and high-water efficient chicken lines.

With climate change, selection for water efficiency and heat resilience are vitally important. We undertook this study to determine the effect of chronic cyclic heat stress (HS) on the hypothalamic expression profile of water homeostasis-associated markers in high (HWE)- and low (LWE)-water efficient chicken lines. HS significantly elevated core body temperatures of both lines. However, the amplitude was higher by 0.5-1°C in HWE compared to their LWE counterparts. HWE line drank significantly less water than LWE during both thermoneutral (TN) and HS conditions, and HS increased water intake in both lines with pronounced magnitude in LWE birds. HWE had better feed conversion ratio (FCR), water conversion ratio (WCR), and water to feed intake ratio. At the molecular level, the overall hypothalamic expression of aquaporins (AQP8 and AQP12), arginine vasopressin (AVP) and its related receptor AVP2R, angiotensinogen (AGT), angiotensin II receptor type 1 (AT1), and calbindin 2 (CALB2) were significantly lower; however, CALB1 mRNA and AQP2 protein levels were higher in HWE compared to LWE line. Compared to TN conditions, HS exposure significantly increased mRNA abundances of AQPs (8, 12), AVPR1a, natriuretic peptide A (NPPA), angiotensin I-converting enzyme (ACE), CALB1 and 2, and transient receptor potential cation channel subfamily V member 1 and 4 (TRPV1 and TRPV4) as well as the protein levels of AQP2, however it decreased that of AQP4 gene expression. A significant line by environment interaction was observed in several hypothalamic genes. Heat stress significantly upregulated AQP2 and SCT at mRNA levels and AQP1 and AQP3 at both mRNA and protein levels, but it downregulated that of AQP4 protein only in LWE birds. In HWE broilers, however, HS upregulated the hypothalamic expression of renin (REN) and AVPR1b genes and AQP5 proteins, but it downregulated that of AQP3 protein. The hypothalamic expression of AQP (5, 7, 10, and 11) genes was increased by HS in both chicken lines. In summary, this is the first report showing improvement of growth performances in HWE birds. The hypothalamic expression of several genes was affected in a line- and/or environment-dependent manner, revealing potential molecular signatures for water efficiency and/or heat tolerance in chickens.

Integrated transcriptome and proteome analysis reveals the unique molecular features and nutritional components on the muscles in Chinese Taihe black-bone silky fowl chicken.

The Taihe Black-Bone silky fowl chicken (BB-sfc) is a renowned dietary and medicinal chicken globally recognized for its high nutritional and medicinal value. Compared to the local Black-Bone black-feathered chicken (BB-bfc), the Taihe silky fowl chicken has higher levels of amino acids, trace elements, and unsaturated fatty acids in their muscles, which offer anti-aging, anti-cancer, and immune enhancing benefits. Despite this, the unique nutritional components, genes, and proteins in Taihe silky fowl chicken muscles are largely unknown. Therefore, we performed a comprehensive transcriptome and proteome analysis of muscle development between BB-sfc and BB-bfc chickens using RNA-Seq and TMT-based quantitative proteomics methods. RNA-Seq analysis identified 286 up-regulated genes and 190 down-regulated genes in BB-sfc chickens, with oxidoreductase activity and electron transfer activity enriched in up-regulated genes, and phospholipid homeostasis and cholesterol transporter activity enriched in down-regulated genes. Proteome analysis revealed 186 significantly increased and 287 significantly decreased proteins in Taihe BB-sfc chicken muscles, primarily affecting mitochondrial function and oxidative phosphorylation, crucial for enhancing muscle antioxidant capacity. Integrated transcriptome and proteome analysis identified 6 overlapped up-regulated genes and 8 overlapped down-regulated genes in Taihe silky fowl chicken, related to improved muscle antioxidant status. Taken together, this research provides a comprehensive database of gene expression and protein information in Taihe Black-Bone silky fowl chicken muscles, aiding in fully exploring their unique economic value in the future.

Chicken UFL1 Restricts Avian Influenza Virus Replication by Disrupting the Viral Polymerase Complex and Facilitating Type I IFN Production.

During avian influenza virus (AIV) infection, host defensive proteins promote antiviral innate immunity or antagonize viral components to limit viral replication. UFM1-specific ligase 1 (UFL1) is involved in regulating innate immunity and DNA virus replication in mammals, but the molecular mechanism by which chicken (ch)UFL1 regulates AIV replication is unclear. In this study, we first identified chUFL1 as a negative regulator of AIV replication by enhancing innate immunity and disrupting the assembly of the viral polymerase complex. Mechanistically, chUFL1 interacted with chicken stimulator of IFN genes (chSTING) and contributed to chSTING dimerization and the formation of the STING-TBK1-IRF7 complex. We further demonstrated that chUFL1 promoted K63-linked polyubiquitination of chSTING at K308 to facilitate chSTING-mediated type I IFN production independent of UFMylation. Additionally, chUFL1 expression was upregulated in response to AIV infection. Importantly, chUFL1 also interacted with the AIV PA protein to inhibit viral polymerase activity. Furthermore, chUFL1 impeded the nuclear import of the AIV PA protein and the assembly of the viral polymerase complex to suppress AIV replication. Collectively, these findings demonstrate that chUFL1 restricts AIV replication by disrupting the viral polymerase complex and facilitating type I IFN production, which provides new insights into the regulation of AIV replication in chickens.

Parental betaine supplementation promotes gosling growth with epigenetic modulation of IGF gene family in the liver.

Betaine is widely used as a feed additive in the chicken industry to promote laying performance and growth performance, yet it is unknown whether betaine can be used in geese to improve the laying performance of goose breeders and the growth traits of offspring goslings. In this study, laying goose breeders at 39 wk of age were fed basal (Control, CON) or betaine-supplemented diets at low (2.5 g/kg, LBT) or high (5 g/kg, HBT) levels for 7 wk, and the breeder eggs laid in the last week were collected for incubation. Offspring goslings were examined at 35 and 63 d of age. The laying rate tended to be increased (P = 0.065), and the feed efficiency of the breeders was improved by betaine supplementation, while the average daily gain of the offspring goslings was significantly increased (P < 0.05). Concentrations of insulin-like growth factor 2 (IGF-2) in serum and liver were significantly increased in the HBT group (P < 0.05), with age-dependent alterations of serum T3 levels. Concurrently, hepatic mRNA expression of the IGF gene family was significantly increased in goslings derived from betaine-treated breeders (P < 0.05). A higher ratio of proliferating cell nuclear antigen (PCNA)-immunopositive nuclei was found in the liver sections of the HBT group, which was confirmed by significantly upregulated hepatic expression of PCNA mRNA and protein (P < 0.05). Moreover, hepatic expression of thyroxine deiodinase type 1 (Dio1) and thyroid hormone receptor ß (TRß) was also significantly upregulated in goslings of the HBT group (P < 0.05). These changes were associated with significantly higher levels of global DNA 5-mC methylation, together with increased expression of methyl transfer genes (P < 0.05), including betaine-homocysteine methyltransferase (BHMT), glycine N-methyltransferase (GNMT), and DNA (cytosine-5-)-methyltransferase 1 (DNMT1). The promoter regions of IGF-2 genes, as well as the predicted TRß binding site on the IGF-2 gene, were significantly hypomethylated (P < 0.05). These results indicate that gosling growth can be improved by dietary betaine supplementation in goose breeders via epigenetic modulation of the IGF gene family, especially IGF-2, in the liver.

The goose industry plays important roles in economics, cultures, and ecosystems, yet the low laying and growth rates of many indigenous breeds hinders the development of the goose farming. Betaine, an important methyl donor, is commonly used as a feed additive in livestock and poultry to enhance animal growth. Dietary supplementation of betaine in laying hens or gestational sows has been reported to promote the growth of their offspring. Here, we sought to investigate whether and how dietary betaine supplementation affects the growth and development of offspring goslings. In this study, goose breeders, both male and female, were fed a basal diet supplemented respectively with 0, 2.5, or 5 g/kg betaine for 7 wk. Goslings hatched from the breeder eggs of different groups were raised under the same standard condition for assessing the growth performance. Parental betaine increases the growth rate of offspring goslings with decreased DNA methylation on the IGF-2 gene promoter and increased expression of the IGF-2 gene in the liver. These results provide scientific evidence for the inter-generational effect of betaine on gosling growth.

Identification of key differentially methylated genes in regulating muscle development and intramuscular fat deposition in chickens.

Muscle development and intramuscular fat (IMF) deposition are intricate physiological processes characterized by multiple gene expressions and interactions. In this research, the phenotypic variations in the breast muscle of Jingyuan chickens were examined at three different time points: 42, 126, and 180 days old. Differential expression analysis and weighted gene co-expression network analysis (WGCNA) were performed to identify differentially methylated genes (DMGs) responsible for regulating muscle development and IMF deposition. The findings indicate a significant increase in breast muscle weight (BMW), myofiber diameter, and cross-sectional area, as well as IMF content, in correlation with the progressive number of growing days in Jingyuan chickens. The findings also revealed that 380 hypo-methylated and 253 hyper-methylated DMGs were identified between the three groups of breast muscle. Module gene and DMG association analysis identified m6A methylation-mediated multiple DMGs associated with muscle development and fat metabolism. In vitro cell modeling analysis reveals stage-specific differences in the expression of CUBN, MEGF10, BOP1, and BMPR2 during the differentiation of myoblasts and intramuscular preadipocytes. Cycloleucine treatment significantly inhibited the expression levels of CUBN, BOP1, and BMPR2, and promoted the expression of MEGF10. These results suggest that m6A methylation-mediated CUBN, MEGF10, BOP1, and BMPR2 can serve as potential candidate genes for regulating muscle development and IMF deposition, and provide an important theoretical basis for further investigation of the functional mechanism of m6A modification involved in adipogenesis.

Analysis of H5N8 influenza virus infection in chicken with mApple reporter genes in vivo and in vitro.

H5N8 highly pathogenic avian influenza virus (HPAIV) has caused huge losses to the global poultry industry and critically threatens public health. Chickens are the important host for the transmission. However, the distribution of H5N8 avian influenza virus (AIV) in chicken and the infected cell types are limitedly studied. Therefore, in this study, we detected viral replication and infection by generating recombinant H5N8 AIV expressing an easily tracked mApple fluorescent reporter. The results showed that recombinant viruses passaged four times in chicken embryos successfully expressed mApple proteins detected by fluorescence microscopy and WB, which verified that the constructed recombinant viruses were stable. Compared to parental virus, although recombinant virus attenuated for replication in MDCK cells, it can still replicate effectively, and form visible plaques. Importantly, the experiments on infection of chicken PBMCs in vitro showed a strong correlation between mApple positivity rate and NP positivity rate (r = 0.7594, P =0.0176), demonstrating that mApple reporter could be used as an indicator to accurately reflect AIV infection. Then we infected monocytes/macrophages in PBMCs in vitro and detected the mApple positive percentage was 55.1%-80.4%, which confirmed the chicken primary monocytic/macrophages are important target cells for avian influenza virus infection. In chicken, compared with parental virus, the recombinant virus-infected chickens had lower viral titers in oropharyngeal cloacal and organs, but it can cause significant pathogenicity in chicken and the mortality rate was approximately 66%. In addition, the results of bioluminescent imaging showed that the fluorescence in the lungs was strongest at 5 days post-infection (DPI). Finally, we discovered the mApple positive expression in chicken lung immune cells (CD45+ cells), especially some T cells (CD4 and CD8 T cells) also carrying mApple, which indicates that the H5N8 AIV showed a tropism for immune cells including chicken T cells causing potentially aggressive against cellular immunity. We have provided a simple visualization for further exploration of H5N8 AIV infected chicken immune cells, which contributes to further understanding pathogenic mechanism of H5N8 AIV infection in chicken.

[Air Microbial Contamination and Risk of Respiratory Exposure of Workers in Chicken Farms].

Animal farms are important sources of microbial contamination in the air environment. However, there are few reports on the time-regularity characteristics of airborne microbial contamination in farms. In the context of this situation, a study was conducted for more than 80 weeks using 16S rRNA gene amplicon sequencing to characterize the bacterial distribution and respiratory exposure in the farm air and fecal environment, respectively, taking a layer farm as an example. The results showed that 16S rRNA concentrations in air and manure samples ranged from 6.08×105-4.90×106 copies·m-3 and 4.27×108-1.15×1010 copies·g-1, respectively. The mean values of airborne bacterial concentrations were significantly higher in winter than in summer, whereas the biodiversity showed the opposite trend. The dominant bacterial phylum in both air and manure in the layer farm was Firmicutes. During the investigated time, the top three dominant genera in the air were relatively stable, in the order of Lactobacillus, Bacteroides, and Faecalibacterium, whereas the dominant genera in feces fluctuated with the increase in breeding time. The correlation between the community structure of bacteria and pathogenic bacteria in both air and manure was not significant, but the concentrations of both target microorganisms in different media were significantly correlated. The bioaerosolization index of bacteria in manure showed an increasing trend with increasing breeding time, whereas the opposite trend was observed for pathogenic bacteria. In this case, [Ruminococcus]_torques_group, Bacteroides, and Faecalibacterium were the top three pathogenic genera that were the most prone to aerosolization. There were seasonal differences in bacterial respiratory exposures of chicken farm workers, with mean intake values of 2.54×107 copies·d-1 and 2.87×105 copies·d-1 for bacteria and pathogenic bacteria, respectively. The results of this study will provide a scientific basis for systematically assessing the contamination characteristics and potential health risks of airborne microorganisms on farms and for developing corresponding industry standards for occupational exposure and prevention and control measures.