Mitochondrial Phylogenomics and Genome Evolution in Anura: Insights From Structure and Gene Order Rearrangements

Mitochondrial genome (mitogenome) has been proposed as a powerful molecular marker for phylogenetic reconstruction, evolutionary genomics, and population-level studies due to its maternal inheritance, relatively conserved structure, and elevated evolutionary rates. Anurans (frogs and toads), comprising 7693 species and ~90% of extant amphibian diversity, exhibit extensive mitogenome variability, yet the evolutionary significance of this structural diversity remains poorly understood. Using 277 anuran mitogenomes (spanning 35 families) and outgroup comparisons, phylogenomic reconstructions and gene order evolution analyses were conducted. Phylogenetic analyses resolved five major clades: Leiopelmatoidea (basal lineage), Discoglossoidea, Pipoidea, Pelobatoidea, and the crown group Neobatrachia. Archaeobatrachians emerged as a paraphyletic group, with sequential divergences of the first four clades. Neobatrachia was monophyletic, with Heleophrynidae and Sooglossidae occupying basal positions. Subsequent diversification revealed two major neobatrachian subclades: one uniting Calyptocephalellidae with Myobatrachoidea and Hyloidea, and another comprising Ranoidea, a topology consistent with recent phylogenomic studies but highlighting persistent conflicts within Hyloidea and Ranoidea. Divergence time estimation traced the origin of Anura to the Early-Late Triassic boundary, with major neobatrachian radiations occurring from the Late Cretaceous through the Early Neogene. CREx-based comparative analyses identified 58 distinct gene arrangement patterns, including lineage-specific synapomorphic rearrangements that were phylogenetically mapped. Our study provides a robust mitogenomic framework for Anura, reconciling deep phylogenetic discordance while revealing novel patterns of gene order evolution. These findings establish critical foundations for future investigation into the mechanisms driving mitogenomic structure diversification and its interplay with the evolutionary success of this ecologically vital vertebrate group.