Black-boned Chickens Research Articles

Dynamic Transcriptome Profile Analysis of Mechanisms Related to Melanin Deposition in Chicken Muscle Development.

The pectoral muscle is an important component of skeletal muscle. The blackness of pectoral muscles can directly affect the economic value of black-boned chickens. Although the genes associated with melanogenesis in mammals and birds have been thoroughly investigated, only little is known about the key genes involved in muscle hyperpigmentation during embryonic development. Here, we analyzed melanin deposition patterns in the pectoral muscle of Yugan black-boned chickens and compared differentially expressed genes (DEGs) between the muscles of Wenchang (non-black-boned chickens) and Yugan black-boned chickens on embryonic days 9, 13, 17, and 21. Melanin pigments were found to gradually accumulate in the muscle fibers over time. Using RNA-seq, there were 40, 97, 169, and 94 genes were identified as DEGs, respectively, between Yugan black-boned chicken muscles and Wenchang chickens at embryonic day 9, 13, 17, and 21 stages (fold change ≥2.0, false discovery rate (FDR) < 0.05). Thirteen DEGs, such as MSTRG.720, EDNRB2, TYRP1, and DCT, were commonly identified among the time points observed. These DEGs were mainly involved in pigmentation, melanin biosynthetic and metabolic processes, and secondary metabolite biosynthetic processes. Pathway analysis of the DEGs revealed that they were mainly associated with melanogenesis and tyrosine metabolism. Moreover, weighted gene co-expression network analysis (WGCNA) was used to detect core modules and central genes related to melanogenesis in the muscles of black-boned chickens. A total of 24 modules were identified. Correlation analysis indicated that one of them (the orange module) was positively correlated with muscle pigmentation traits (r > 0.8 and p < 0.001). Correlations between gene expression and L* values of the breast muscle were investigated in Yugan and Taihe black-boned chickens after hatching. The results confirmed that EDNRB2, GPNMB, TRPM1, TYR, and DCT expression levels were significantly associated with L* values (p < 0.01) in black-boned chickens (p < 0.05). Our results suggest that EDNRB2, GPNMB, TRPM1, TYR, and DCT are the essential genes regulating melanin deposition in the breast muscle of black-boned chickens. MSTRG.720 is a potential candidate gene involved in melanin deposition in the breast muscles of Yugan black-boned chickens.

Characterization of the Coding Sequence of the MC1R (Melanocortin 1 Receptor) Gene of Ayam Cemani Black Chickens.

Plumage color is one of the most important traits characterizing chicken breeds. Black-boned chickens constitute a specific group of breeds with a unique phenotype. One of the representatives is the Indonesian Ayam Cemani. The extraordinary black phenotype results from a specific chromosomal rearrangement. We used complete CDS of crucial color-related gene MC1R, which plays a key role in melanin distribution but has not been previously studied in Ayam Cemani. It turned out that Ayam Cemani individuals possess a newly found non-synonymous mutation G355A resulting in amino acid substitution D119N. Together with the presence of G274A (E92K), the new missense variant enabled us to distinguish a new extended black allele at the E locus. All of the investigated birds were heterozygous in terms of the new mutation. Previous studies and our own results indicate a high level of genetic variation within the MC1R gene within and between chicken breeds. Besides the key mutations that make it possible to distinguish particular major alleles, there are also numerous substitutions that give haplotypes more characteristics for individual breeds.

Research Note: Transcriptome analysis of skeletal muscles of black-boned chickens, including 2 types (wild and mutated) of Taihe black-boned silky fowl and 1 type (wild) of Yugan black-boned chicken

The large amount of melanin deposited in Taihe black-boned silky fowl and other black-boned chicken breeds is a highly valued trait due to its desirable nutritional and functional properties, such as antiaging, immune-enhancing, and antifatigue properties. To identify the candidate genes and pathways potentially responsible for melanogenesis in Taihe black-boned silky fowl, digital gene expression tag (DGE-tag)-based transcriptome analyses were performed for 2 groups: wild-type Taihe black-boned silky fowl (TH-1245) vs. mutated Taihe black-boned silky fowl (BY-1245) and TH-1245 vs. wild-type Yugan black-boned chicken (YG-1245). In total, 430 and 765 differentially expressed genes (DEGs) were identified and 13 DEGs displaying different gene expression patterns between the 2 groups were considered valuable for further investigation, such as ANKRD1, MYOZ2, and MYOD1. Furthermore, 6 functionally grouped networks composed of 36 significant GO terms, mainly involved in muscle-related and signaling-related biological processes, were screened by functional enrichment network analysis. In addition, protein-protein interaction (PPI) network analysis identifies 2 top clusters containing 20 hub genes for 2 comparison groups. MYL1 and RPS14 were considered the most potential candidate genes among all hub genes. The Gene Set Enrichment Analysis (GSEA) results showed that the terms and pathways, such as muscle system process, arachidonic acid metabolism, melanogenesis, and tyrosine metabolism, may play important roles in the melanogenesis and further investigations were needed to clarify the relationships between these pathways and melanin. Overall, these results are helpful for furthering our understanding of melanogenesis in breast muscle of Taihe black-boned silky fowl and Yugan black-boned chicken.

Resveratrol alleviates heat-stress-induced impairment of the jejunal mucosa through TLR4/MAPK signaling pathway in black-boned chicken

Heat stress in chickens caused by high temperatures in summer is a serious issue faced by the poultry industry globally, which reduces product quality. The aim of this study is to investigate the role of resveratrol in alleviating heat stress injury and inflammatory response of jejunal mucosa in black-boned chickens through TLR4/MAPK signaling pathway. In total, 240 black-boned chickens (28-day old) were randomly divided into 4 treatment groups as follows. The normal temperature (NT) and normal temperature with resveratrol (NT+Res) groups received a basal diet without and with 400 mg/kg resveratrol, respectively, and treated at 24℃ ± 2℃, 24 h/d. The high temperature (HT) and high temperature with resveratrol (HT+Res) groups received basal diet without and with 400 mg/kg resveratrol, respectively, and treated at 37℃ ± 2℃ for 8 h/d and 24°C ± 2°C for the rest of the time for 12 d. The results revealed the heat-stress responses impaired the villous structure of the jejunum, causing a rough and uneven surface of the jejunal villus, and local intestinal villi were even more prone to rupture. However, resveratrol significantly improved the morphology and structure of jejunal mucosa under heat stress. Heat stress increased the mRNA levels of toll-like receptor 4 (TLR4), c-Jun, c-fos, caspase-3, and p38 (P < 0.05), reduced mRNA level of Bcl-2, and reduced the expression of tight junction proteins Occludin, ZO-1, and Claudin1 (P < 0.05) in the jejunal mucosa. However, resveratrol inhibited the TLR4/ mitogen-activated protein kinase (MAPK) signaling pathway via downregulating TLR4, c-Jun, p38, and caspase-3 (P < 0.05); upregulating Bcl-2 (P < 0.05); decreasing the protein levels of MKK3, p53, and myeloid differentiation factor 88 (MYD88); and increasing the protein levels of Occludin, ZO-1, and Claudin1. In addition, it reduced the levels of JNK and p38 proteins (P < 0.05) and inflammatory factors like tumor necrosis factor-α (TNF-α) in the jejunal mucosa of black-boned chickens under heat stress. In conclusion, resveratrol may play a regulatory role in heat-stress-induced damage and inflammatory response in the intestinal mucosa of black-boned chickens under heat stress.

Transcriptomic, 16S ribosomal ribonucleic acid and network pharmacology analyses shed light on the anticoccidial mechanism of green tea polyphenols against Eimeria tenella infection in Wuliangshan black-boned chickens

BackgroundEimeria tenella is an obligate intracellular parasitic protozoan that invades the chicken cecum and causes coccidiosis, which induces acute lesions and weight loss. Elucidating the anticoccidial mechanism of action of green tea polyphenols could aid the development of anticoccidial drugs and resolve the problem of drug resistance in E. tenella.MethodsWe constructed a model of E. tenella infection in Wuliangshan black-boned chickens, an indigenous breed of Yunnan Province, China, to study the efficacy of green tea polyphenols against the infection. Alterations in gene expression and in the microbial flora in the cecum were analyzed by ribonucleic acid (RNA) sequencing and 16S ribosomal RNA (rRNA) sequencing. Quantitative real-time polymerase chain reaction was used to verify the host gene expression data obtained by RNA sequencing. Network pharmacology and molecular docking were used to clarify the interactions between the component green tea polyphenols and the targeted proteins; potential anticoccidial herbs were also analyzed.ResultsTreatment with the green tea polyphenols led to a reduction in the lesion score and weight loss of the chickens induced by E. tenella infection. The expression of matrix metalloproteinase 7 (MMP7), MMP1, nitric oxide synthase 2 and ephrin type-A receptor 2 was significantly altered in the E. tenella infection plus green tea polyphenol-treated group and in the E. tenella infection group compared with the control group; these genes were also predicted targets of tea polyphenols. Furthermore, the tea polyphenol (-)-epigallocatechin gallate acted on most of the targets, and the molecular docking analysis showed that it has good affinity with interferon induced with helicase C domain 1 protein. 16S ribosomal RNA sequencing showed that the green tea polyphenols had a regulatory effect on changes in the fecal microbiota induced by E. tenella infection. In total, 171 herbs were predicted to act on two or three targets in MMP7, MMP1, nitric oxide synthase 2 and ephrin type-A receptor 2.ConclusionsGreen tea polyphenols can directly or indirectly regulate host gene expression and alter the growth of microbiota. The results presented here shed light on the mechanism of action of green tea polyphenols against E. tenella infection in chickens, and have implications for the development of novel anticoccidial products.Graphical

Transcriptome Profile Analysis Identifies Candidate Genes for the Melanin Pigmentation of Skin in Tengchong Snow Chickens.

Tengchong Snow chickens are one of the most precious, black-boned chickens in Yunnan province and usually produce black meat. However, we found a small number of white meat traits in the chicken population during feeding. In order to determine the pattern of melanin deposition and the molecular mechanism of formation in the Tengchong Snow chicken, we measured the luminance value (L value) and melanin content in the skin of black meat chickens (Bc) and white meat chickens (Wc) using a color colorimeter, ELISA kit, and enzyme marker. The results showed that the L value of skin tissues in black meat chickens was significantly lower than that of white meat chickens, and the L value of skin tissues gradually increased with an increase in age. The melanin content of skin tissues in black meat chickens was higher than that of white meat chickens, and melanin content in the skin tissues gradually decreased with an increase in age, but this difference was not significant (p > 0.05); the L value of skin tissues in black meat chickens was negatively correlated with melanin content, and the correlation coefficient was mostly above -0.6. In addition, based on the phenotypic results, we chose to perform the comparative transcriptome profiling of skin tissues at 90 days of age. We screened a total of 44 differential genes, of which 32 were upregulated and 12 were downregulated. These DEGs were mainly involved in melanogenesis, tyrosine metabolism and RNA transport. We identified TYR, DCT, and EDNRB2 as possible master effector genes for skin pigmentation in Tengchong Snow black meat chickens through DEGs analysis. Finally, we measured the mRNA of TYR, DCT, MC1R, EDNRB2, GPR143, MITF, and TYRP1 genes through a quantitative real-time polymerase chain reaction (qPCR) and found that the mRNA of all the above seven genes decreased with increasing age. In conclusion, our study initially constructed an evaluation system for the black-boned traits of Tengchong Snow chickens and found key candidate genes regulating melanin deposition, which could provide an important theoretical basis for the selection and breeding of black-boned chickens.

Thai Local Chicken Breeds, Chee Fah and Fah Luang, Originated from Chinese Black-Boned Chicken with Introgression of Red Junglefowl and Domestic Chicken Breeds

Knowledge of the genetic characteristics, origin, and local adaptation of chickens is essential to identify the traits required for chicken breeding programs. Chee Fah and Fah Luang are black-boned chicken breeds reared in Chiang Rai, Thailand. Chickens are an important part of the local economy and socio-culture; however, the genetic diversity, characteristics, and origins of these two breeds have been poorly studied. Here, we investigated the genetic diversity, gene pool, and origin of the Chee Fah and Fah Luang chickens using mitochondrial DNA D-loop (mtDNA D-loop) sequencing and microsatellite genotyping, as well as habitat suitability analysis using maximum entropy modeling. The MtDNA D-loop sequencing and microsatellite genotype analyses indicated that the Chee Fah and Fah Luang chickens shared haplogroups A, B, and CD with Chinese black-boned chickens. Gene pool analysis revealed that the Chee Fah and Fah Luang chickens have distinct genetic patterns compared to Thai domestic chickens and red junglefowl. Some gene pools of red junglefowl and other Thai domestic chickens were observed within the Chee Fah and Fah Luang chicken gene pool structures, suggesting genetic exchange. The data indicate that the Chee Fah and Fah Luang chickens originated from Chinese indigenous black-boned chicken breeds and experienced crossbreeding/hybridization and introgression with red junglefowl and other domestic breeds during domestication. Interestingly, the Chee Fah and Fah Luang chickens from Chiang Rai shared the same allelic gene pool, which was not shared with the Chee Fah and Fah Luang chickens from Mae Hong Son, suggesting at least two gene pool origins in the Chee Fah and Fah Luang chicken populations. Alternatively, different gene pools in the Chee Fah and Fah Luang chickens from different localities might be caused by differences in environmental factors, especially elevation.

Time-Course Transcriptome Landscape of Bursa of Fabricius Development and Degeneration in Chickens

The bursa of Fabricius (BF) is a target organ for various pathogenic microorganisms; however, the genes that regulate BF development and decline have not been fully characterized. Therefore, in this study, histological sections of the BF were obtained from black-boned chickens at 7 (N7), 42 (N42), 90 (N90) and 120 days (N120) of age, and the differential expression and expression trends of the BF at different stages were analyzed by transcriptome analysis. The results showed that the growth of the BF progressively matured with age, followed by gradual shrinkage and disappearance. Transcriptome differential analysis revealed 5914, 5513, 4575, 577, 530 and 66 differentially expressed genes (DEG) in six different comparison groups: N7 vs. N42, N7 vs. N90, N7 vs. N120, N42 vs. N90, N42 vs. N120 and N90 vs. N120, respectively. Moreover, we performed transcriptomic analysis of the time series of BF development and identified the corresponding stages of biological process enrichment. Finally, quantitative real-time polymerase chain reaction (qRT-PCR) was used to validate the expression of the 16 DEGs during bursal growth and development. These results were consistent with the transcriptome results, indicating that they reflect the expression of the BF during growth and development and that these genes reflect the characteristics of the BF at different times of development and decline. These findings reflect the characteristics of the BF at different time intervals.

Transcriptomics and Metabolomics Analysis of the Ovaries of High and Low Egg Production Chickens.

Simple SummaryThe ovarian tissues of different breeds of hens during egg production were investigated through transcriptomics and metabolomics to provide a more comprehensive understanding of the molecular mechanisms of the ovary during egg production. Four genes involved in egg production were predicted by the transcriptome, including P2RX1, INHBB, VIPR2, and FABP3, and several close metabolites associated with reproduction were identified in the metabolome, including 17α-hydroxyprogesterone, iloprost, spermidine and adenosine. Correlation analysis of specific differential genes and differential metabolites identified important gene-metabolite pairs VIPR2–Spermidine and P2RX1–Spermidine in the reproductive process.Egg production is a pivotal indicator for evaluating the fertility of poultry, and the ovary is an essential organ for egg production and plays an indispensable role in poultry production and reproduction. In order to investigate different aspects of egg production mechanisms in different poultry, in this study we performed a metabolomic analysis of the transcriptomic combination of the ovaries of two chicken breeds, the high-production Ninghai indigenous chickens and the low-production Wuliangshan black-boned chickens, to analyze the biosynthesis and potential key genes and metabolic pathways in the ovaries during egg production. We predicted four genes in the transcriptomic that are associated with egg production, namely P2RX1, INHBB, VIPR2, and FABP3, and identified three important pathways during egg production, “Calcium signaling pathway”, “Neuroactive ligand–receptor interaction” and “Cytokine–cytokine receptor interaction”, respectively. In the metabolomic 149 significantly differential metabolites were identified, 99 in the negative model and 50 in the positive model, of which 17α-hydroxyprogesterone, iloprost, spermidine, and adenosine are important metabolites involved in reproduction. By integrating transcriptomics and metabolomics, the correlation between specific differential genes and differential metabolites identified important gene-metabolite pairs “VIPR2-Spermidine” and “P2RX1-Spermidine” in egg production. In conclusion, these data provide a better understanding of the molecular differences between the ovaries of low- and high-production hens and provide a theoretical basis for further studies on the mechanics of poultry egg production.

Integrative analysis of transcriptomics and metabolomics to reveal the melanogenesis pathway of muscle and related meat characters in Wuliangshan black-boned chickens

BackgroundMelanin is an important antioxidant in food and has been used in medicine and cosmetology. Chicken meat with high melanin content from black-boned chickens have been considered a high nutritious food with potential medicinal properties. The molecular mechanism of melanogenesis of skeletal muscle in black-boned chickens remain poorly understood. This study investigated the biological gene-metabolite associations regulating the muscle melanogenesis pathways in Wuliangshan black-boned chickens with two normal boned chicken breeds as control.ResultsWe identified 25 differentially expressed genes and 11 transcription factors in the melanogenesis pathways. High levels of the meat flavor compounds inosine monophosphate, hypoxanthine, lysophospholipid, hydroxyoctadecadienoic acid, and nicotinamide mononucleotide were found in Wuliangshan black-boned chickens.ConclusionIntegrative analysis of transcriptomics and metabolomics revealed the dual physiological functions of the PDZK1 gene, involved in pigmentation and/or melanogenesis and regulating the phospholipid signaling processes in muscle of black boned chickens.

Research Note: Fine mapping of sequence variants associated with body weight of Lueyang black-boned chicken in the CCKAR gene

Cholecystokinin A receptor (CCKAR) is a key receptor mediating satiety. Previous studies found that decreased expression of CCKAR attenuated satiety, and thus contributed to the high-growth of broiler chickens. The objective of this study is to map sequence variants associated with the growth of chickens in the CCKAR. The CCKAR and upstream 1.4 kb genomic sequences were resequenced to find out all sequence variants using 35 Lueyang black-boned chickens (LBC). Haplotypes were reconstructed using the PHASE program. Linkage disequilibrium between variants was analyzed using the Haploview software. Associations of 33 tag SNPs that captured 89% of all variants with body weight of LBC (n = 675) at 16 (BW16), 20 (BW20) weeks of age and the onset (BWOEP) of egg production were tested using linear mixed models. A total of 126 SNPs were found and formed 41 haplotypes in 35 resequenced samples. Average length of haplotype blocks is 129 bp, indicating that LBC maintains low linkage disequilibrium at the CCKAR locus. Eleven of 33 tag SNPs were significantly associated with BW16, but not with BW20 and BWOEP. These significantly associated variants were most (8/11) distributed in a 2 kb region (chr4:73206169-73208244) around the Exon3. They together with 33 captured variants potentially disrupted binding sites of 471 transcription factors. Twelve variants can disrupt appetite (FOXO1) or lipid metabolism-related TF (AR and C/EBP) motifs. This study recognized chr4:73206169-73208244 as a key region harboring functional variants affecting the growth of chickens.

Mitochondrial DNA diversity and demographic history of Black-boned chickens in China

Black-boned chickens (Gallus domesticus, herein abbreviated BBCs) are well known for their unique appearance and medicinal properties and have a long breeding history in China. However, the genetic diversity and demographic history of BBCs remain unclear. In this study, we analyzed 844 mitochondrial DNA D-loop sequences, including 346 de novo sequences and 498 previously published sequences from 20 BBC breeds. We detected a generally high level of genetic diversity among the BBCs, with average haplotype and nucleotide diversities of 0.917 ± 0.0049 and 0.01422, respectively. Nucleotide diversity was highest in populations from Southwest China (0.01549 ± 0.00026), particularly in Yunnan Province (0.01624 ± 0.00025). Significant genetic divergence was detected between most breeds, particularly between Yunnan chickens and those from all other provinces. Haplogroups F and G had the highest levels of genetic diversity and were restricted to Southwest China, particularly Yunnan Province. Based on neutrality tests and mismatch distribution analyses, we did not obtain evidence for rapid population expansions and observed similar demographic histories in BBCs and local non-BBCs. Our results suggest that Chinese BBCs have complex breeding histories and may be selected in situ from local domestic chickens. These results improve our understanding of the genetic heritage and breeding histories of these desirable chickens.

Identification of key microRNAs affecting melanogenesis of breast muscle in Muchuan black-boned chickens by RNA sequencing

ABSTRACT 1. Melanin content is considered an important indicator of meat quality in black-boned chickens, which have a high market value. To understand the complex physiological processes underlying muscle melanogenesis in this chicken, differentially expressed miRNAs (DEMs) were detected between black muscle (BM) and white muscle (WM) of chickens using high-throughput sequencing technology. Six small RNA libraries were constructed, and more than 16.75 million clean reads were obtained for each library. 2. A total of 582 known miRNAs and 65 novel miRNAs were identified from the six chicken sequence libraries. A total of 19 DEMs were identified between the two groups, of which nine were upregulated and 10 were downregulated. Furthermore, the DEMs were predicted to target 572 genes. 3. Certain DEMs (such as miR-204, miR-133b, and miR-12 229-3p) and their target genes may play an important role in muscle melanogenesis of chickens. These findings provide a foundation for clarifying the miRNA regulatory mechanisms involved in muscle pigmentation in avian species.

Investigating structural impact of a valine to isoleucine substitution on anti-Müllerian hormone in silico and genetic association of the variant and AMH expression with egg production in chickens

Anti-Müllerian hormone (AMH) acts in maintaining orderly cyclic recruitment of early follicles, suggesting that it is a promising candidate for influencing animal reproductive efficiency. This study aimed to elucidate the effect of a missense mutation of Val566Ile on the structure of AMH protein and the genetic association of Val566Ile and AMH expression with egg production in chickens. Structural perturbations of Val566Ile were predicted by homology modeling. The association of the variant with the number of eggs was tested using a quantitative trait transmission disequilibrium test model. AMH expression in granulosa cells in Lueyang black-boned chickens was compared with that in Nick chickens. The Val566 of AMH is a non-conservative amino acid among mammals and birds, but its hydrophobicity is completely conservative. The substitution of Val566 for Ile566 potentially disrupted hydrogen bonds and solvent accessibility of 22 residues and created a short α-helix in the C terminus of AMH. Despite having striking structure-disrupting potential, the variant was not statistically associated with the number of eggs (P>0.05) in the Lueyang black-boned chickens. We did not detect differential expression of AMH between Lueyang black-boned chickens and Nick chickens (P>0.05). These results confirmed the structural impact of Val566Ile, but suggested that Val566Ile and AMH expression might not be the major genetic determinants for egg production in Lueyang black-boned chickens.

Identification and Characterisation of Alternative Splice Variants of Hoxb9 and Their Correlation with Melanogenesis in the Black-Boned Chicken

ABSTRACT Homeobox B9 (HOXB9) gene has been demonstrated to be associated with melanogenesis in chicken plumage by high-throughput RNA sequencing. In this study, we cloned and characterised HOXB9 in black-boned chickens. Two alternative splice variants (HOXB9-1 and HOXB9-2) were identified in chicken feather bulbs. Expression analysis of HOXB9 in 11 different chicken tissues by RT-PCR indicated that the two transcripts were only expressed in the kidney, abdominal fat, feather bulbs, skin, and small intestine. No HOXB9-1 or HOXB9-2 transcripts were detected in the breast muscle or the ovary. The two HOXB9 variants were expressed at significantly different levels in black feather bulbs and white feather bulbs (p<0.01), and in black skin compared with white skin (p<0.01). The results indicate that HOXB9-1 and HOXB9-2 may be involved in melanin formation in the plumage and skin. The expression of HOXB9-2 was higher in white than in black muscles (p<0.05), HOXB9-2 may play a role in muscle melanogenesis. Our results suggest that HOXB9, which is partially regulated by alternative splicing, may be involved in the process of melanogenesis in the black-boned chicken.

Transcriptome profile analysis identifies candidate genes for the melanin pigmentation of breast muscle in Muchuan black-boned chicken.

Melanin-based coloration in the meat of black-boned chicken is a major economic issue in China. Variation in the pigmentation (hypopigmentation) of chicken muscle causes direct economic losses every year. To determine the molecular mechanisms involved in the melanogenesis of muscle tissue, this study used high-throughput sequencing to compare differences in the transcriptome between black (BM) and white (WM) chicken breast muscles. We constructed 6 cDNA libraries from BM and WM groups in Muchuan black-boned chickens. A comparison between the BM and WM groups revealed 264 differentially expressed genes, of which 152 were upregulated, whereas 112 were downregulated in black muscle. Gene ontology and a Kyoto Encyclopedia of Genes and Genomes pathway analysis identified several differentially enriched biological functions and processes of the 2 muscles. Seven promising candidate genes [PMEL, Ras-related protein RAB29, and 5 solute carrier superfamily genes: SLC6A9, SLC38A4, SLC22A5, SLC35F3, and SLC16A3] may play an important role in the melanogenesis of chicken muscle. Our data provide a valuable resource for identifying genes whose functions are critical for muscle melanogenesis, and will assist studies of the molecular mechanisms of melanogenesis regulation in chicken muscle.

Transcriptome Profile Analysis of Mechanisms of Black and White Plumage Determination in Black-Bone Chicken

Background/Aims: Melanin is a major and ubiquitous component of plumage colouration, and patterns of melanin pigmentation in birds are extremely varied. However, the molecular mechanism of pigmentation in avian plumage is still largely unknown. Methods: To elucidate the molecular mechanisms involved in the formation of black and white plumage, this study takes advantage of high-throughput sequencing technology to compare differences in the transcriptome between black and white chicken feather bulbs. In total, we constructed six cDNA libraries from black (Group B) and white (Group W) feather bulbs in the dorsal plumage of Muchuan black-boned chickens. Results: A comparison between Groups B and W revealed 61 differentially expressed genes, with 47 displaying higher, and 14 displaying lower, levels of expression in white feather bulbs. Our results revealed a set of candidate genes and two potential metabolic pathways involved in black-bone chicken plumage melanogenesis. These include four homeobox genes (HOXB9, HOXC8, HOXA9, and HOXC 9), two glutathione (GSH) metabolism-related genes (CHAC1 and GPX3), and the transforming growth factor beta (TGF-β) signalling pathway. Two known genes, TYR and MITF, were also shown to play a role in melanin formation. Conclusion: our data provide a valuable resource for discovering genes important in plumage melanin formation and will help further elucidate the molecular mechanisms for black and white plumage.

Genetic variation in the MITF promoter affects skin colour and transcriptional activity in black-boned chickens

ABSTRACT1. Microphthalmia-associated transcription factor (MITF) plays a pivotal role in melanocyte development by regulating the transcription of major pigmentation enzymes (e.g. TYR, TYRP1 and DCT). A single-nucleotide polymorphism (SNP), c.-638T>C, was identified in the MITF promoter, and genotyping of a population (n = 426) revealed that SNP c.-638T>C was associated with skin colour in black-boned chickens.2. Individuals with genotypes CC and TC exhibited greater MTIF expression than those with genotype TT. Luciferase assays also revealed that genotype CC and TC promoters had higher activity levels than genotype TT. Expression of melanogenesis-related gene (TYR) was higher in the skin of chickens with the CC and CT genotype compared to TT chickens (P < 0.05).3. Transcription factor-binding site analyses showed that the c.-638C allele contains a putative binding site for transcription factor sterol regulatory element-binding transcription factor 2, aryl hydrocarbon receptor nuclear translocator, transcription factor binding to IGHM enhancer 3 and upstream transcription factor 2. In contrast, the c.-638T allele contains binding sites for Sp3 transcription factor and Krüppel-like factor 1.4. It was concluded that MITF promoter polymorphisms affected chicken skin colour. SNP c.-638T>C could be used for the marker-assisted selection of skin colour in black-boned chicken breeding.

Fermented Green Vegetable Soybean Meal Increases Dressed Carcass Meat and Bone Weight in Black-Boned Chickens (Fah Luang Chicken, Gallus gallus)

To use effectively waste green vegetable soyabean (edamame) and to improve the dressed carcass meat in black-boned chickens (Fah Luang chicken, Gallus gallus), fermented edamame meal was fed to black-boned chickens from 4 to 16 weeks of age. At 4 weeks of age, 20 birds (10 male and 10 female birds) were used to measure the initial weights of dressed carcass meat and each bone part. Another 60 birds were divided into 2 groups with 3 replicates of 10 chickens (5 male and 5 female birds) each. The control group received a basal formula diet only and the experimental group was fed 40 g fermented edamame meal per bird for a period of 30 min before being fed the basal formula diet. The weights of whole body, each dressed carcass muscle part and each bone part increased in the chickens fed fermented edamame meal (p<0.05). The feed cost per Kg body weight gain decreased (p<0.05). Income over feed cost per Kg body weight gain, as well as income over feed cost and strain per bird, increased in the fermented edamame meal group (p<0.05). These facts suggest that the feeding of fermented edamame meal to black-boned chicken induces the chicken’s production due to increase of dressed carcass muscle, cheap feed cost (USD/Kg body weight gain) and high income (USD/bird), suggesting the potential for commercialization of waste edamame meal as a feed ingredient for black-boned chickens.

Resveratrol modulates intestinal morphology and HSP70/90, NF-κB and EGF expression in the jejunal mucosa of black-boned chickens on exposure to circular heat stress.

The aim of this study was to investigate whether supplementation with resveratrol could alleviate intestinal injuries and to explore how resveratrol regulates heat shock protein (HSP)70, HSP90, nuclear factor kappa B (NF-κB) and epidermal growth factor (EGF) expression in the jejunal mucosa of black-boned chickens under circular heat stress. A total of 300 black-boned chicks of 42-d-old were randomly assigned to five treatment groups. The positive control chickens were kept in a normal-temperature (NT, 24 ± 2 °C) chamber and fed with a basal diet. The other four groups were kept in a circular high-temperature (HT, 37 ± 2 °C) chamber for 8 h and fed a basal diet supplemented with 0, 200, 400, or 600 mg per kg of resveratrol for 15 days. The results showed that the heat-stress responses damaged the villus structures of the jejunum and ileum, resulting in shortened intestinal villi, deepened crypts, and a reduced villus height to crypt depth (V/C) ratio and decreased the numbers of goblet cells and lymphocytes. Heat stress also caused increased mRNA and protein expression of HSP70, HSP90 and NF-κB, and reduced EGF in the jejunal mucosa. Dietary supplementation with 400 mg per kg of resveratrol improved the villus morphology, increased the numbers of goblet cells and lymphocytes, attenuated the mRNA overexpression of HSP70, HSP90 and NF-κB on the 6th, 10th and 15th day of heat stress (P < 0.05), and activated the expression of EGF (P < 0.05) in the jejunal mucosa. Resveratrol reduced protein expression of HSP70, HSP90 and NF-κB in the jejunal villi after 15 days of heat stress, and increased EGF expression from the lamina propria toward the epithelial cells of the villi. These results suggest that dietary resveratrol offers a potential nutritional strategy to improve the intestinal morphology and alleviate jejunum mucosa injuries by modulating the mRNA and protein expression of HSPs, and the epithelial growth factor and transcription factor in black-boned chickens subjected to circular heat stress.