Amoya chusanensis and Wuhanlinigobius polylepis are endemic goby species (Gobiidae, Perciformes) in Chinese waters, with A. chusanensis occurring in the East China Sea and W. polylepis distributed in waters off Shanghai and Hainan. We applied mitochondrial COI gene sequencing to larval specimens collected from the Yangtze River estuary in August 2017, confirming species identity for both endemic gobies. This study presents the first detailed morphological descriptions of their larvae, supported by high-resolution photographic documentation, and compares their ontogenetic development with related gobiid species. Comparative analysis revealed that the melanin pigmentation pattern along the ventral body margin—extending from the anus to the posterior terminus of the notochord—exhibits species-specific characteristics and serves as a diagnostic criterion for larval identification. These findings establish a novel approach for early developmental research on endemic Chinese fishes and provide scientific basis for conservation strategies targeting their nursery grounds.