This study investigates the effectiveness of cytochrome c oxidase subunit 1 (COI) gene sequence divergence in distinguishing closely related animal species. The research analyzed over 13,000 congeneric species pairs across 11 animal phyla, focusing on sequence divergence in the COI gene. The results show that COI sequence divergence is generally sufficient to distinguish closely related species in most animal phyla, except for Cnidaria. The study found that sequence divergence in the COI gene is high in most animal groups, enabling species-level diagnosis. However, Cnidaria showed significantly lower divergence, likely due to their slow mitochondrial evolution. The study also found that COI divergence in fungi and protists is higher than in animals, suggesting that COI-based identification systems could be extended to these groups. The study concludes that COI divergence can serve as an effective tool for species recognition, as intraspecific divergence is typically low, and most congeneric species pairs show sufficient divergence for clear diagnosis. The study also addresses concerns about horizontal transfer of mitochondrial DNA and mitochondrial polymorphisms between closely related species, suggesting that such complications are rare. The study highlights the importance of mitochondrial and nuclear genome interactions in shaping mitochondrial gene diversity and suggests that selective sweeps may play a role in maintaining low intraspecific divergence. Overall, the study supports the use of COI-based DNA barcoding as a reliable method for species identification in the animal kingdom.This study investigates the effectiveness of cytochrome c oxidase subunit 1 (COI) gene sequence divergence in distinguishing closely related animal species. The research analyzed over 13,000 congeneric species pairs across 11 animal phyla, focusing on sequence divergence in the COI gene. The results show that COI sequence divergence is generally sufficient to distinguish closely related species in most animal phyla, except for Cnidaria. The study found that sequence divergence in the COI gene is high in most animal groups, enabling species-level diagnosis. However, Cnidaria showed significantly lower divergence, likely due to their slow mitochondrial evolution. The study also found that COI divergence in fungi and protists is higher than in animals, suggesting that COI-based identification systems could be extended to these groups. The study concludes that COI divergence can serve as an effective tool for species recognition, as intraspecific divergence is typically low, and most congeneric species pairs show sufficient divergence for clear diagnosis. The study also addresses concerns about horizontal transfer of mitochondrial DNA and mitochondrial polymorphisms between closely related species, suggesting that such complications are rare. The study highlights the importance of mitochondrial and nuclear genome interactions in shaping mitochondrial gene diversity and suggests that selective sweeps may play a role in maintaining low intraspecific divergence. Overall, the study supports the use of COI-based DNA barcoding as a reliable method for species identification in the animal kingdom.