Yolk metabolomics reveals candidate compounds associated with egg specific density and hatchability in white layer breeder hens.

The present study aimed to evaluate the influence of egg geometry, weight loss, and specific density (SD) on hatchability in older white layer breeder hens. In addition, based on the positive correlation observed between fertility and egg SD, we hypothesized that the mineral and metabolomic composition of the yolk may be associated with specific density. A total of 8,874 eggs from five Lohmann LSL Lite breeder flocks (52-64 weeks) were analyzed. We found that digital image analysis is an adequate method for evaluating the egg shape index, whereas Archimedes' principle is suitable for determining SD. Although we observed an influence of shape index and weight loss on hatchability, the clearest findings were related to SD. Eggs within the upper 50% SD range (1075-1110g/L) exhibited higher hatchability than those with lower SD, primarily due to reduced infertility and lower early embryonic mortality. Low-SD eggs showed increased contamination rates. Yolk mineral concentrations did not differ significantly between SD groups, except for the Na:K ratio. However, the N;Ka ratio showed minimal predictive value, indicating that mineral composition alone has limited explanatory power for eggshell density. Untargeted metabolomics identified 310 metabolites in yolk, but only melatonin glucuronide and dihydroxytetradecanoic acid met the significance criteria (p<0.05; fold change > 2). This study provides a precise way to evaluate egg geometry and SD. Furthermore, using a large dataset, it demonstrates that eggs with low SD have reduced hatchability due to infertility and early mortality, likely linked to contamination. Although the Na:K ratio, melatonin glucuronide, and dihydroxytetradecanoic acid showed limited predictive performance when evaluated individually, their identification highlights potential biochemical differences associated with SD and supports their further investigation as candidate markers.