DNA methylome and transcriptome identified Key genes and pathways involved in Speckled Eggshell formation in aged laying hens.

BACKGROUND: The quality of poultry eggshells is closely related to the profitability of egg production. Eggshell speckles reflect an important quality trait that influences egg appearance and customer preference. However, the mechanism of speckle formation remains poorly understood. In this study, we systematically compared serum immune and antioxidant indices of hens laying speckled and normal eggs. Transcriptome and methylome analyses were used to elucidate the mechanism of eggshell speckle formation. RESULTS: The results showed that seven differentially expressed genes (DEGs) were identified between the normal and speckle groups. Gene set enrichment analysis (GSEA) revealed that the expressed genes were mainly enriched in the calcium signaling pathway, focal adhesion, and MAPK signaling pathway. Additionally, 282 differentially methylated genes (DMGs) were detected, of which 15 genes were associated with aging, including ARNTL, CAV1, and GCLC. Pathway analysis showed that the DMGs were associated with T cell-mediated immunity, response to oxidative stress, and cellular response to DNA damage stimulus. Integrative analysis of transcriptome and DNA methylation data identified BFSP2 as the only overlapping gene, which was expressed at low levels and hypomethylated in the speckle group. CONCLUSIONS: Overall, these results indicate that aging- and immune-related genes and pathways play a crucial role in the formation of speckled eggshells, providing useful information for improving eggshell quality.

Structural characteristics of speckled chicken eggshells and their effect on reproductive performance.

Speckles are common on the brown eggshells of chicken eggs, especially for aged hens. They are important as they affect the consumer preference and economic value of eggs. The cause of speckles in eggshells is still unclear. In this study, we verified the difference of eggshell quality between speckled eggs and normal eggs. Structural characteristics of speckled eggshells were investigated using a scanning electron microscope. Results showed no significant difference in the eggshell quality between normal eggs and speckled eggs, except for a lower eggshell color-L value in the latter. More materials deposited between vertical crystal layer and cuticle layer in the speckled shell region, leading to the thicker eggshell than adjacent normal area. The relative content of protoporphyrin IX was significantly higher in the speckled area than in the adjacent normal area of the eggshells. In addition, there was no significant differences in productive and reproductive performance between hens that laid normal eggs or speckled eggs, except for a lower hatchability of the speckled eggs. In conclusion, we infer that the uneven distribution of eggshell pigment protoporphyrin IX leads to the formation of speckled eggs. This is the first study to establish the characteristics and causes of speckled eggshells. Moreover, this study provides novel insights into external egg quality and a foundation for the in-depth study of speckled eggshells.