Molecular and morphological identification of yellow tonguefish ( Symphurus williamsi ) and darkcheek tonguefish ( Symphurus chabanaudi ) at the southern and entrance to the Gulf of California

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Abstract
The genus Symphurus is a group of demersal fish with high diversity and taxonomic challenges during their early development. This study addressed the morphological and molecular identification of Symphurus larvae in La Bahía de la Paz and the entrance to the Gulf of California. Zooplankton samples were collected during the day and night at 58 stations in October 2022. Based on morphological characteristics such as meristics and pigmentation, seven Symphurus morphotypes were pre‐classified and genetically analysed using the mitochondrial genes cytochrome c oxidase subunit I (COI) and 16S rRNA (16S). Genetic analysis identified four morphotypes: three of them corresponded to a single species, Symphurus williamsi , with 100% similarity in the 16S gene, while the other morphotype was identified as Symphurus chabanaudi , with 100% similarity in both markers (COI and 16S). The pigmentation patterns described after molecular identification were consistent and allowed differentiation between the two species. S. williamsi presented three to five strongly stellate melanophores on the dorsal side and five to six melanophores on the ventral side during the preflexion to postflexion stages, while S. chabanaudi had three prominent melanophores in the central part of the notochord (lateral line), as well as a series of small melanophores positioned on the dorsal and ventral sides. S. williamsi larvae were located throughout the entire entrance to the Gulf of California, mainly in Transitional Water, while Symphurus chabanaudi was collected only in La Bahía de la Paz in Gulf of California Water. These results demonstrated that the integration of genetic and morphological tools is fundamental in taxonomic studies of ichthyoplankton, particularly for the identification of species in early developmental stages that exhibit high morphological similarity. In addition, water masses might be a signal of spawning habitat of different species.

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