DNA barcoding and morphology of tongue sole species(family: cynoglossidae) in the Mekong delta

Abstract

Accurate species identification in tongue soles (Family: Cynoglossidae) is hindered by overlapping morphological traits, the absence of robust taxonomic keys, and discrepancies among reference sequences in public genetic databases. This study aimed to improve taxonomic resolution of tongue soles in the Mekong Delta by integrating morphological analysis and DNA barcoding using the mitochondrial cytochrome c oxidase subunit I (COI) gene. A total of 63 specimens were collected from local markets and landing sites across the Mekong Delta (Viet Nam), representing eight species from two genera Cynoglossus and Paraplagusia. Three species (C. macrolepidotus, C. oligolepis, C. itinus) are newly recorded in the region. Morphometric and meristic traits, along with key diagnostic features, were analyzed. These characters were used to construct a taxonomic key to support reliable field-based species identification. COI barcoding produced 33 sequences, with intraspecific Kimura 2- parameter (K2P) < 0.01 and interspecific distances ranging from 0.134 to 0.239, confirming a clear barcode gap. The integrative approach clarified species boundaries among morphologically similar taxa and genetic misidentification, especially C. macrolepidotus, C. oligolepis, and C. arel. Several inconsistencies in GenBank and BOLD were detected and corrected, including the first validated COI sequences of C. feldmanni. A COI-based neighbor-joining phylogenetic tree supported species-level monophyly but revealed unresolved relationships at the genus level, suggesting potential paraphyly of Cynoglossus. These findings highlight the value of combining morphological and molecular tools to resolve taxonomic ambiguities, improve DNA reference libraries, and support biodiversity monitoring of tongue soles