Development of the axial skeleton and skeletal abnormalities of Atlantic halibut (Hippoglossus hippoglossus) from first feeding through metamorphosis

Larval Atlantic halibut (Hippoglossus hippoglossus) were sampled from a commercial hatchery to observe skeletal abnormalities throughout development and to develop a metamorphic stage definition utilizing order of vertebral ossifications in combination with ossification events of the cranium. Using a whole mount cartilage and bone staining technique, skeletal abnormalities were classified and quantified into 21 types and the order of ossification events determined. During metamorphic development, stage 5 fish lacked calcified vertebral elements. Stage 6 was characterized by ossification of the exterior perimeter of vertebrae and attachment points of neural and hemal spines within the prehemal and posterior hemal regions. Further calcification of the vertebral column extends cephalad and caudad from the origin of ossification during stage 7. In stage 8, the vertebral column and caudal fin bones are weakly ossified but showing full ossification by stage 9. There was a high correlation between age (R2 = 0.964), standard length (R2 = 0.996), and myotome height (R2 = 0.994) with stage. Rapid development occurring over a short period of time while in the prometamorphic stages of development, may give rise to the presence of serious skeletal abnormalities in later development. The highest frequency of skeletal abnormalities occurred in the prehemal region, followed by the hemal region despite developmental stage. Hypertrophic vertebrae in the prehemal region are present in earlier developing larvae, and pre- to pro-metamorphic. Skeletal abnormalities commonly begin during stage 6 and 7, when the majority of the vertebral elements are ossifying. Studies on the immunohistochemistry and biochemical processes involved in bone growth may be useful to determine the causative factors of skeletal abnormalities.