Skeletal Development During Metamorphosis of the Rice Field Frog Hoplobatrachus rugulosus (Wiegmann, 1834) (Anura: Dicroglossidae)
Keywords:
ossification chronology, Anura, Dicroglossidae, ontogeny, double-stainingAbstract
The rice field frog (Hoplobatrachus rugulosus) is an important native species in Southeast Asian agricultural wetlands and a major commodity in the aquaculture industry. Despite its ecological and economic importance, and its use as an animal model in various studies, detailed data regarding its skeletal ontogeny remain scarce. This study addresses this gap by presenting a comprehensive chronological atlas of osteogenesis for H. rugulosus from Gosner stages 28 to 46. Using a cleared and double-staining technique with Alcian Blue and Alizarin Red S, we documented the transition from a primary cartilaginous framework to a mineralized skeleton. Premetamorphosis (stages 28–35) is characterized by a predominantly cartilaginous skeleton, with ossification initiating in the axial skeleton (neural arches of the vertebrae) at stage 34. Prometamorphosis (stages 36–41) is marked by the onset of cranial ossification (exoccipital) at stage 36, alongside the progression of ossification to the hindlimbs at stages 39–40. The final phase involves extensive ossification during the metamorphic climax (stages 42–46), where forelimb ossification initiates at stage 42. This is followed by the simultaneous ossification of the cranial elements (premaxilla, maxilla, and frontoparietal), along with completion of the urostyle at stage 46. The delayed cranial ossification in H. rugulosus is notable and diverges from the typical anuran pattern, suggesting that species-specific factors may influence skeletal developmental timing in this species. These findings provide an essential anatomical baseline for this significant species in Southeast Asia.
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