DNA barcode reference database and gap analysis for biomonitoring Hong Kong's marine animals

DNA-based taxonomic identification relies on comprehensive DNA reference libraries, but uneven barcode coverage poses a global challenge. Here we determined the gaps in BOLD and GenBank for marine animals in Hong Kong's subtropical waters. We compiled 170,529 sequences from four marker genes [cytochrome c oxidase subunit I (COI), 12S ribosomal RNA (rRNA), 16S rRNA, and 18S rRNA] representing 4074 marine animal species spanning 17 phyla. Gap analysis highlighted variations in COI barcode coverage across phyla, ranging from 0 % in Ctenophora, Gnathostomulida, Hemichordata, Phoronida and Platyhelminthes to 90.44 % in Chordata. The average coverage of COI (53.24 % in BOLD and 58.47 % in GenBank) was substantially higher than that of the other three rRNA genes (19.46–32.25 %). In total, 4392 Barcode Index Numbers (BINs) were assigned to 2169 species, of which 41.13 % were taxonomically concordant and 50.71 % were displayed multiple BINs, indicating the presence of cryptic species and the potential for undiscovered diversity. Overall, this study provided a valuable DNA barcode library for marine animals in Hong Kong, revealed that many marine animals lack a barcode, and many species may be undescribed, highlighting the need for enhanced reference libraries. We also outlined a strategy for filling the gaps in the database. Utilizing the local reference database can significantly enhance molecular-based biodiversity assessment and improve the identification of cryptic (morphologically similar) and invasive species, thereby providing more informed support for coastal management and conservation efforts in Hong Kong waters and the surrounding areas of China's Hong Kong-Zhuhai-Macau Greater Bay Area.