This study investigated the application prospects of environmental DNA (eDNA) technology for studying the species diversity of cetaceans. It involves extracting eDNA from water samples in the Xisha sea area using universal primers for amplification targeting cetacean mitochondrial 12S rRNA and 16S rRNA. High-throughput sequencing was conducted. The results were combined with visual observation data to discuss the potential of eDNA technology in cetacean species diversity studies. These results indicated that the 4 universal primers were effective for cetacean identification. Using these primers, 5 cetacean species were detected in samples from 19 stations in the Xisha sea area. The identified species included the pan-tropical spotted dolphin (Stenella attenuata), spinner dolphin (Stenella longirostris), Fraser's dolphin (Lagenodelphis hosei), Eden's whale (Balaenoptera edeni edeni), and short-finned pilot whale (Globicephala macrorhynchus). The cetaceans observed during sampling included the pantropical spotted dolphin, spinner dolphin, and Risso's dolphin (Grampus griseus). The dominant species detected by both methods were consistent, and the environmental DNA (eDNA) technology identified species that were not visually observed. The detection results of primers Cet-12S and Marver3 covered 5 cetacean species and 17 sites where cetaceans were detected, indicating that the combined use of primers targeting different gene segments is advantageous for improving detection effectiveness. There were no significant differences among the 4 primers in terms of the detected cetacean sequences and number of species. Cet-12S exhibited the highest proportion of cetacean sequences and species, accounting for 33.0% and 21.1% of the total population, respectively. In contrast, the other primers detected much lower proportions of cetacean sequences and species, ranging from 0.2% to 0.6% and 2.0% to 4.1%, respectively. In addition, the number of nonspecifically amplified sequences and species detected by Cet-12S was significantly lower than that of the other 3 primers, indicating its high specificity as a universal primer for cetacean eDNA. Compared to visual observation, eDNA technology has advantages such as high sensitivity, efficiency, and cost-effectiveness, making it well suited for studying the species diversity of cetaceans. This study enhanced our understanding of cetacean species diversity in the Xisha Sea area, offering technical insights for cetacean conservation and research.