# Characterization and Phylogenetic Analysis of the Complete Mitochondrial Genome of Celaenorrhinus victor (Lepidoptera: Hesperiidae)

## TL;DR

This study reports the complete mitochondrial genome of Celaenorrhinus victor and uses it to explore evolutionary relationships within skipper butterflies.

## Contribution

The paper provides the first complete mitogenome for Celaenorrhinus victor and contributes to resolving phylogenetic relationships in Hesperiidae.

## Key findings

- The mitogenome of C. victor is 15,180 bp with typical mitochondrial gene content and a strong A + T bias.
- Codon usage in Celaenorrhinus species is similar, with A + T mutational pressure influencing gene evolution.
- Phylogenetic analysis supports C. victor's placement in Celaenorrhinus but shows limited resolution for deeper hesperiid relationships.

## Abstract

Skipper butterflies (Hesperiidae) are diverse, yet mitochondrial genomic resources for many genera remain limited. Here, we report the complete mitochondrial genome of Celaenorrhinus victor and compare it with available congeners. The mitogenome is 15,180 bp long and contains the typical set of 37 mitochondrial genes and a control region, with a strong A + T bias that is common in insect mitochondria. Comparative analyses across Celaenorrhinus indicate broadly similar codon-usage profiles among species, and protein-coding genes show signals consistent with predominant purifying selection. Using a dataset of 66 complete mitogenomes (65 Hesperiidae plus one outgroup), we reconstructed a mitogenome-based phylogeny that supports the placement of C. victor within Celaenorrhinus and highlights relationships among close congeners. Deeper relationships among major hesperiid lineages show only moderate support in parts of the tree, suggesting that expanded taxon sampling and nuclear loci will be useful for refining the backbone phylogeny.

Background: Skipper butterflies (Hesperiidae) are a morphologically distinctive lineage within Papilionoidea, yet relationships among many groups remain difficult to resolve, and mitochondrial genomic resources remain limited for some tribes, including Celaenorrhinini. Methods: We sequenced and characterized the complete mitochondrial genome of Celaenorrhinus victor using Illumina short-read sequencing. Gene content and organization were annotated, codon-usage patterns were assessed across Celaenorrhinus using relative synonymous codon usage and multiple compositional/selection tests (ENC–GC3s, neutrality, and PR2 analyses), selective constraints were evaluated using Ka/Ks for 13 protein-coding genes, and phylogenetic relationships were inferred with a partitioned maximum-likelihood analysis of 66 complete hesperiid mitogenomes. Results: The circular mitogenome of C. victor is 15,180 bp and contains the typical 37 genes (13 protein-coding genes, 22 tRNAs, and two rRNAs) plus an A + T-rich control region, with an overall A + T content of 79.64%. Gene order and orientation match those of other Celaenorrhinus and hesperiid mitogenomes. All protein-coding genes use standard invertebrate mitochondrial start codons (with cox1 initiating with TTG) and terminate with complete TAA stop codons. Codon usage is strongly biased toward A/U-ending codons and is broadly similar among five sampled Celaenorrhinus mitogenomes; ENC–GC3s, neutrality, and PR2 analyses indicate a predominant influence of A + T-directed mutational pressure with additional effects beyond base composition. Ka/Ks values for all 13 protein-coding genes were <1, consistent with pervasive purifying selection; cox genes were the most conserved, whereas several NADH dehydrogenase subunit genes evolved comparatively faster. The phylogeny recovered monophyletic Celaenorrhinini and a well-supported Celaenorrhinus clade, placing C. victor as sister to Celaenorrhinus consanguineus, while deeper nodes among major hesperiid lineages showed only moderate support in parts of the tree. Conclusions: This study provides a new mitogenomic resource for Celaenorrhinini and a comparative reference for codon usage and selective constraints within Celaenorrhinus, supporting the placement of C. victor within Hesperiidae while highlighting remaining uncertainty at deeper hesperiid divergences.