# The complete mitochondrial genomes of the two species of Astyanax (Characiformes: Acestrorhamphidae) that occur in cenotes of the Yucatán Peninsula karst aquifer: comparative analyses and their taxonomic implications

## TL;DR

This paper analyzes the mitochondrial genomes of two Astyanax fish species from Yucatán cenotes and finds they are likely the same species.

## Contribution

The study provides complete mitochondrial genome sequences for two Astyanax species and challenges their current taxonomic classification.

## Key findings

- The mitochondrial genomes of Astyanax altior and Astyanax bacalarensis are nearly identical in length, composition, and gene arrangement.
- Species delimitation analyses reject the hypothesis that A. altior and A. bacalarensis are distinct species.
- The study suggests the two species likely represent the same species-level lineage.

## Abstract

Despite their high diversity and widespread distribution throughout Neotropical freshwaters, published genomic resources for fishes of the genus Astyanax are rather limited. This situation is exemplified by Astyanax altior and Astyanax bacalarensis, the two species in the genus found in cenotes of the Yucatán Peninsula karst aquifer, a unique yet vulnerable hydrologic system in southeastern Mexico. Distinguishing between these two species based on external morphology, however, is not straightforward, making their purportedly allopatric distributions the most conclusive way to tell them apart. Therefore, testing the current taxonomy is warranted.

To address the perceived deficiency in available genomic data and to test the hypothesis that A. altior and A. bacalarensis are different species, we generated novel complete mitochondrial genome sequences for both species and analyzed them in a comparative framework. We used a four-step next-generation sequencing protocol consisting of genomic DNA extraction, library preparation, sequencing, and bioinformatic analysis to sequence, assemble and annotate the resulting mitogenomes. Descriptive analyses were used to characterize the mitogenomes, while comparative analyses were used to shed light on the evolutionary relationships and species limits. The complete mitochondrial genomes of A. altior and A. bacalarensis are almost identical in length, composition, and general gene arrangement, and follow the overall genomic organization of most teleosts. The degree of genetic differentiation between A. altior and A. bacalarensis was found to be minimal and the hypothesis that they are different species was rejected by our species delimitation analyses.

Besides reporting novel mitogenomic data for a highly diverse group of fishes with limited representation in genomic repositories, our results strongly support previous suspicions that A. altior and A. bacalarensis correspond to the same species-level lineage and would therefore represent synonyms. Despite our findings, we defer synonymizing them until a proper revisionary study can corroborate this evolutionary and taxonomic hypothesis.

The online version contains supplementary material available at 10.1007/s11033-025-10788-6.