Comparative Mitogenome Analyses of Subgenera and Species Groups in Epeorus (Ephemeroptera: Heptageniidae)

SIMPLE SUMMARY: As one of the most species-rich genera of Ephemeroptera, Epeorus Eaton, 1881, was found to be widely distributed in Holarctic and Oriental regions, and nine subgenera have been reported. Previous phylogenetic studies of Epeorus were mainly focused on morphological characters or several gene fragments. Here, 15 mitogenomes of Epeorus are sequenced and the comparative mitogenome analysis of six subgenera is performed. The gene rearrangement of trnI-trnM-trnQ-NCR-ND2 was first found in the genus. In addition, differences in genetic composition and codon usage between the species with this kind of rearrangement and other Epeorus species were observed. Phylogenetic analyses show that three subgenera, Proepeorus, Belovius and Iron, are not monophyletic groups, and our results imply that gill structures are not always appropriate for the classification of subgenera in Epeorus. ABSTRACT: Epeorus Eaton, 1881 is a diverse mayfly genus in Heptageniidae comprising more than 100 species which are further divided into nine subgenera and several species groups. However, the classification and the phylogenetic relationships among them are still uncertain. Here, 15 complete mitochondrial genomes of Epeorus were sequenced and compared together with six available ones of same genus in the NCBI database. Based on morphological classification, the 21 mitogenomes were classified into six subgenera (Proepeorus, Epeorus s.str., Belovius, Iron, Caucasiron and Siniron) and four species groups (G1, G2, montanus and longimanus). Among all analyzed mitogenomes, the gene rearrangement of trnI-trnM-trnQ-NCR-ND2 was first found occurring in three species of group G1, whereas the gene block trnI-trnM-trnQ-trnM-ND2 was observed in all other mitogenomes of Epeorus. Furthermore, the genetic composition and codon usage of species in group G1 were also significantly different from all other Epeorus species, except group longimanus. The intergenic spacer between trnA and trnR, which has the stem-loop secondary structure, occurred in all 21 mitogenomes, and the sequences of stems and loops were conserved within species groups. Furthermore, the phylogenetic analyses strongly support the monophyly of all species groups, although three of six recognized subgenera Proepeorus, Belovius, and Iron, were shown as the non-monophyletic groups.