Comparative Mitogenomic Analysis of Three Chionea Species (Tipulomorpha: Limoniidae): Insights into Phylogenetic Relationships and Selection Pressure

Abstract

Chionea is classified within the Tipuloidea superfamily and predominantly inhabits cold regions. However, its phylogenetic relationships remain contentious. In this study, the first three mitogenomes of Chionea (Diptera: Limoniidae) sampled in northeastern China (Jilin region) were sequenced, and their phylogenetic relationships were reconstructed by integrating these sequences with 30 additional Tipuloidea mitogenomes retrieved from NCBI. Unlike other Tipuloidea species, which are predominantly distributed in relatively warmer regions, this research investigates whether positive selection has acted on the mitogenomes of these three Chionea species due to environmental pressures, thereby elucidating key evolutionary drivers for Chionea. The three mitogenomes of Chionea exhibit characteristic features typical of insect mitogenomes, comprising 13 protein-coding genes (PCGs), 2 ribosomal RNA genes (16S rRNA and 12S rRNA), 22 transfer RNA genes (tRNA), and a single non-coding control region (D-loop). Notably, the secondary structure of trnS1 lacks the DHU arm in all three samples, and UUA (Leu) emerges as the most frequently utilized codon. Furthermore, the COX2 and ND5 genes utilize incomplete stop codons "T". Utilizing these 13 PCGs, we reconstructed the internal phylogenetic relationships within Tipuloidea, revealing that Chionea tianhuashana and C. sphaerae form sister branches, while (C. tianhuashana + C. sphaerae) constitutes a sister branch to C. crassipes. Moreover, our analysis confirms the monophyly of Tipulidae, Tipula, and Nephrotoma as well as the polyphyly of Tipulinae, Chioneinae, and Limoniidae. In the branch site model analysis, three positively selected sites were detected when Chionea was designated as the foreground branches: COX3 (at position 242), ND5 (at position 535), and ND6 (at position 138).

Keywords: Chionea; crane fly; mitochondrial genome; phylogenetic relationship; selective stress analysis.

Figure 2

The RSCU of C. tianhuashana, C. sphaerae and C. crassipes. The X-axis shows all possible codon combinations, including synonymous codons, while the Y-axis indicates the corresponding RSCU values, with each codon represented by a distinct color.

Figure 1

Mitogenome map of Chionea tianhuashana, Chionea sphaerae and Chionea crassipes. The outermost ring displays features encoded by the positive strand, while the adjacent ring corresponds to the negative strand. tRNAs are annotated using standard abbreviations. The third ring visualizes GC content, and the innermost ring (violet and green) indicates GC skew.

Figure 3

ML and BI trees for all Tipulomorpha were constructed using 13 PCGs. Different subfamilies and genera are color-coded, while the three outgroup species are grouped under a single color. The GenBank accession numbers for each species are provided following their respective names.