Abstract
Background:Onychostoma virgulatum is an endemic freshwater fish in South China, first described as a new species in 2009. However, little is known about this species and no complete mitochondrial genomes of O. virgulatum has been reported to date. This study seeks to elucidate the characteristics of the mitochondrial genome of O. virgulatum and investigate the phylogenetic relationships within the Acrossocheilinae subfamily, particularly among the genera Onychostoma, Acrossocheilus, and Folifer. Methods: The mitochondrial genome of O. virgulatum was sequenced and assembled. We analyzed its sequence length, nucleotide composition, and evolutionary relationships within the Acrossocheilinae by incorporate data from 58 previously published mitochondrial genomes. Results: The complete circular sequence is 16,606 bp in length and contains 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes, and a typical control region (D-loop), all arranged in a typical order. The genomic base composition is biased toward A+T content (56.5%), with 31.4% A, 25.1% T, 27.4% C, and 16.1% G. Among about 30 Acrossocheilina species, the nonsynonymous (Ka) to synonymous substitutions (Ks) for all 13 protein-coding genes (PCGs) are significantly less than 1, suggesting strong negative or purifying selection in these species. The phylogenetic trees inferred from the mitogenome and 13 PCGs of 58 Acrossocheilinae sequences consistently indicate that: (1) O. virgulatum shares the closest genetic relationship with Onychostoma barbatulum; (2) Acrossocheilinae species are clustered into three major clades, with neither Acrossocheilus nor Onychostoma forming monophyletic groups. Conclusions: This study provides new insights into the taxonomy and phylogenetic relationships of Acrossocheilinae, particularly O. virgulatum, contributing to a better understanding of the systematics, origin, and evolution of this subfamily.