Mayflies of the family Neoephemeridae are widespread in the Holarctic and Oriental regions, and its phylogenetic position is still unstable in the group Furcatergalia (mayflies with fringed gills). In the present study, we determined the complete mitogenomes of two species, namely Potamanthellus edmundsi and Pulchephemera projecta, of this family. The lengths of two mitogenomes were 15,274 bp and 16,031 bp with an A + T content of 73.38% and 73.07%, respectively. Two neoephemerid mitogenomes had a similar gene size, base composition, and codon usage of protein-coding genes (PCGs), and the sequenced gene arrangements were consistent with the putative ancestral insect mitogenomes as understood today. The most variable gene of Furcatergalia mitogenomes was ND2, while the most conserved gene was COI. Meanwhile, the analysis of selection pressures showed that ND6 and ATP8 exhibited a relaxed purifying selection, and COI was under the strongest purifying selection. Phylogenetic trees reconstructed based on two concatenated nucleotide datasets using both maximum likelihood (ML) and Bayesian inference (BI) estimations yielded robust identical topologies. These results corroborated the monophyly of seven studied families and supported the family Leptophlebiidae as being of the basal lineage of Furcatergalia. Additionally, the sister-group relationship of Caenidae and Neoephemeridae was well supported. Methodologically, our present study provides a general reference for future phylogenetic studies of Ephemeroptera at the mitogenome level.
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.
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