Comparative analysis of morphological atlas at different HH stages of embryonic development for SPF ducks and SPF...

Avian embryos are widely used models in developmental biology and biomedical research, yet systematic comparative morphology across specific pathogen-free (SPF) poultry species remains limited. We investigated the embryonic development of three SPF duck breeds Shanma (HSM), Shaoxing (HBW), and Jinding (HJD) and SPF White Leghorn chickens (BM8 line) under controlled SPF conditions. Fertilized eggs were incubated at standardized temperature and humidity, and embryonic growth was monitored daily. Developmental stages were classified according to the Hamburger-Hamilton (HH) system and grouped into early, middle, and late periods. Morphological, internal, and growth characteristics were recorded using in ovo and ex ovo chorioallantoic membrane (CAM) imaging, enabling the construction of comprehensive developmental atlases, and morphological analyses were subsequently performed. Early-stage embryos exhibited conserved features, including somite formation, neural tube development, and limb bud initiation. Divergence between species became apparent from the middle stages, with duck embryos showing accelerated beak elongation, broader craniofacial structures, and species-specific limb patterning compared with chickens. Vascular development, ocular and integumentary pigmentation, and appendage differentiation progressed in a stage-dependent manner. Across duck breeds, morphology was largely similar, with minor variations in growth rate and timing. Late-stage development was characterized by growth stabilization, skeletal maturation, and yolk and albumen utilization. Ducks required 7-8 more days of incubation than chickens, reflecting interspecies differences in developmental tempo. This study establishes a multi-dimensional reference framework based on comprehensive morphological observations, providing breed-and stage-specific embryonic developmental atlases for SPF ducks and chickens. By incorporating multiple observational perspectives, the atlases enable more intuitive visualization of dynamic developmental changes, particularly in structures such as the yolk sac. These resources provide valuable references for developmental biology, disease modeling, and vaccine research, and further broaden the applicability of SPF poultry in biomedical research.