# Assembly, Characterization, and Phylogenetic Insights from the Complete Mitochondrial Genome of Cleisthenes herzensteini (Pleuronectiformes: Pleuronectidae)

## TL;DR

This study sequenced the complete mitochondrial genome of Cleisthenes herzensteini, a declining flatfish species, to better understand its genetics and evolutionary relationships.

## Contribution

This is the first complete mitochondrial genome sequencing and analysis for Cleisthenes herzensteini, providing new genetic resources for conservation and phylogenetic studies.

## Key findings

- The mitochondrial genome of Cleisthenes herzensteini is 17,171 bp long with a typical A + T bias.
- It shares a close evolutionary relationship with Dexistes rikuzenius and forms a distinct clade with other flatfish species.
- The genome exhibits unique features like incomplete stop codons and specific codon usage patterns.

## Abstract

The pointhead flounder is an important fish in the Northwest Pacific, but its population has dropped sharply due to overfishing and habitat damage. To help protect this species, we need to better understand its genetic makeup. In this study, we decoded the complete set of genetic material contained in its mitochondria—the energy-producing parts of the cell—which is useful for tracking evolution and family relationships among fish. We found that its mitochondrial genetic structure is similar to that of other related flatfish, but also has some unique features, such as preferred patterns in its genetic code and certain incomplete stop signals in genes. By comparing it with other species, we confirmed its close evolutionary relationship with another flatfish, Dexistes rikuzenius, and clarified its place within the broader flatfish family tree. This genetic information provides a valuable resource for scientists and conservationists, supporting future efforts in species identification, population monitoring, and the development of protection strategies for this declining fishery resource.

Cleisthenes herzensteini is a commercially important demersal fish in the Northwest Pacific. However, the resource stock of this species has undergone a drastic decline due to overfishing and habitat degradation. As a representative taxon for benthic adaptation in the order Pleuronectiformes, the molecular mechanisms underlying its specialized phenotypic traits remain poorly elucidated. Furthermore, population-level studies focusing on the mitochondrial genome of Cleisthenes herzensteini are currently scarce. Given that the mitochondrial genome serves as an ideal genetic tool for deciphering species evolution and population genetics, sequencing of its mitogenome will help fill critical gaps in genetic resources and provide essential support for species conservation and phylogenetic research. In this study, we sequenced, assembled, and annotated its complete mitochondrial genome. The circular mitogenome is 17,171 bp in length and exhibits a typical A + T bias (54.04%). Repeat sequence analysis identified 35 dispersed repeats. Codon usage analysis revealed that leucine was the most frequently encoded amino acid, with CUU being the preferred codon. Several protein-coding genes possessed incomplete stop codons (T--/TA-), and a nucleotide preference for A and C was observed at the third codon position. Phylogenetic reconstruction based on mitogenomes from 23 species supported the monophyly of the order Pleuronectiformes. C. herzensteini showed the closest relationship with Dexistes rikuzenius, forming a distinct clade alongside Hippoglossoides dubius and Limanda aspera. These results provide essential genetic resources for understanding the evolution and population genetics of C. herzensteini and related flatfishes. According to the investigation, this study represents the first report on the sequencing and analysis of the complete mitochondrial genome of the Cleisthenes herzensteini. This not only fills the gap in mitochondrial genetic information for this species but also provides a reference for subsequent investigations into the phylogenetic relationships and evolutionary processes within the family Pleuronectidae.