Duck egg production, like that of laying hens, follows a typical low–peak–low cycle, reflecting the dynamics of the reproductive system. Post-peak, some ducks undergo a cessation of egg laying, indicative of a regression process in the oviduct. Notably, the magnum, being the longest segment of the oviduct, plays a crucial role in protein secretion. Despite its significance, few studies have investigated the molecular mechanisms underlying oviduct regression in ducks that have ceased laying eggs. In this study, we conducted single-cell transcriptome sequencing on the magnum tissue of Shaoxing ducks at 467 days of age, utilizing the 10× Genomics platform. This approach allowed us to generate a detailed magnum transcriptome map of both egg-laying and ceased-laying ducks. We collected transcriptome data from 13,708 individual cells, which were then subjected to computational analysis, resulting in the identification of 27 distinct cell clusters. Marker genes were subsequently employed to categorize these clusters into specific cell types. Our analysis revealed notable heterogeneity in magnum cells between the egg-laying and ceased-laying ducks, primarily characterized by variations in cells involved in protein secretion and extracellular matrix (ECM)-producing fibroblasts. Specifically, cells engaged in protein secretion were predominantly observed in the egg-laying ducks, indicative of their role in functional albumen deposition within the magnum, a phenomenon not observed in the ceased-laying ducks. Moreover, the proportion of THY1⁺ cells within the ECM-producing fibroblasts was found to be significantly higher in the egg-laying ducks (59%) compared to the ceased-laying ducks (24%). Similarly, TIMP4⁺ fibroblasts constituted a greater proportion of the ECM-producing fibroblasts in the egg-laying ducks (83%) compared to the ceased-laying ducks (58%). These findings suggest a potential correlation between the expression of THY1 and TIMP4 in ECM-producing fibroblasts and oviduct activity during functional reproduction. Our study provides valuable single-cell insights that warrant further investigation into the biological implications of fibroblast subsets in the degeneration of the reproductive tract. Moreover, these insights hold promise for enhancing the production efficiency of laying ducks.

The differences in earlobe color of Jiangshan black-bone chickens have been reported to be caused by the combined effects of melanin and collagen. In this study, we conducted LC-MS untargeted metabolomics and 16S rDNA diversity sequencing on the cecal contents of two types of earlobes: peacock green (Blue and Green group) and dark reddish-purple (Black group). The metabolomic sequencing identified a total of 747 differential metabolites (DMs), in which the metabolites were primarily enriched in tyrosine and tryptophan metabolism pathways between peacock green and dark reddish-purple earlobes. There were 15 different bacterial taxa among the three groups of earlobes at the genus level, and correlation analysis between metabolites and microbes revealed that the DMs between peacock green and dark reddish-purple earlobes were positively correlated with the different bacterial taxa. In short, there are differences in gut microbiota and metabolites between Jiangshan black-bone chickens with peacock green earlobes and those with dark reddish-purple earlobes. Our results suggest that the bacterial phyla Firmicutes and Bacteroidota may influence melanin synthesis by affecting tryptophan metabolism, induced by 5-Methoxyindoleacetate, and tyrosine metabolism, induced by maleylacetoacetic acid and maleic acid, leading to differences in earlobe color.

Chicken semen cryopreservation is crucial for utilizing high-quality cockerel genetics, but semen is highly sensitive to cryoinjury, leading to poor preservation outcomes. This study aimed to establish a theoretical foundation for selecting cockerels for semen cryopreservation through serum testing and to improve semen quality via DNA methylation editing. Semen and serum samples were collected from 102 Xiaoshan cockerels, with semen cryopreserved and thawed following standardized protocols. Post-thaw semen quality and serum testosterone (T) levels were assessed. Eight cockerels were selected based on motile sperm quality, and whole-genome bisulfite sequencing (WGBS) was used to analyze sperm DNA methylation. The results showed a significant positive correlation between serum T levels and sperm motility. There were notable differences in sperm motility and serum T levels between high-quality and low-quality semen groups but no differences in estradiol (E₂), superoxide dismutase (SOD), or glutathione peroxidase (GSH-Px) levels. A total of 217 differentially methylated regions (DMRs) and 116 differentially methylated genes (DMGs) were identified. Key genes such as PRKACB (protein kinase, cAMP-dependent, catalytic, beta) and ACSL1 (long-chain-fatty-acid--CoA ligase 1) were associated with sperm motility. These findings provide important insights for improving semen cryopreservation and contribute to breeding practices and the development of cryoprotectants.

The objective of this study was to identify the AlkB family genes in poultry using bioinformatics, and to explore their molecular characteristics, evolutionary relationships, and expression patterns to clarify their potential functions in poultry. (1) Methods: The study utilized the NCBI database to obtain chicken genome data, and screened and validated AlkB family members (ALKBH1-5, ALKBH8, and FTO) by hmmsearch and TBtools. MEGA 11.0 was used for phylogenetic analysis, PHYRE2 and I-TASSER predicted protein structures, and the String database was used to construct an interoperability network. Finally, the tissue expression profiles were analyzed by using The Human Protein Atlas online database and qRT-PCR. (2) Results: Phylogenetic analysis revealed distinct avian and mammalian clusters, with chicken AlkB proteins exhibiting low sequence homology but conserved 3D structures compared to mammals. Chromosomal synteny and conserved domains highlighted evolutionary divergence, with ALKBH4 lacking typical AlkB structural motifs. Protein interaction networks linked ALKBH1/2/3/5/8/FTO, underscoring functional coordination in poultry adaptation. Tissue-specific expression showed high AlkB levels in brain tissues, while ALKBH5 dominated in muscle. During differentiation, ALKBH3, ALKBH5, and FTO expression significantly increased during myoblast differentiation. (3) Conclusions: This study identified seven AlkB family genes in poultry, revealing their phylogenetic classification into two subfamilies, conserved structural domains, chromosomal synteny, and tissue-specific expression patterns.