ABSTRACT: Insects of the order Phasmatodea are mainly distributed in the tropics and subtropics and are best known for their remarkable camouflage as plants. In this study, we sequenced three complete mitochondrial genomes from three different families: Orestes guangxiensis, Peruphasma schultei, and Phryganistria guangxiensis. The lengths of the three mitochondrial genomes were 15,896 bp, 16,869 bp, and 17,005 bp, respectively, and the gene composition and structure of the three stick insects were identical to those of the most recent common ancestor of insects. The phylogenetic relationships among stick insects have been chaotic for a long time. In order to discuss the intra- and inter-ordinal relationship of Phasmatodea, we used the 13 protein-coding genes (PCGs) of 85 species for maximum likelihood (ML) and Bayesian inference (BI) analyses. Results showed that the internal topological structure of Phasmatodea had a few differences in both ML and BI trees and long-branch attraction (LBA) appeared between Embioptera and Zoraptera, which led to a non-monophyletic Phasmatodea. Consequently, after removal of the Embioptera and Zoraptera species, we re-performed ML and BI analyses with the remaining 81 species, which showed identical topology except for the position of Tectarchus ovobessus (Phasmatodea). We recovered the monophyly of Phasmatodea and the sister-group relationship between Phasmatodea and Mantophasmatodea. Our analyses also recovered the monophyly of Heteropterygidae and the paraphyly of Diapheromeridae, Phasmatidae, Lonchodidae, Lonchodinae, and Clitumninae. In this study, Peruphasma schultei (Pseudophasmatidae), Phraortes sp. YW-2014 (Lonchodidae), and species of Diapheromeridae clustered into the clade of Phasmatidae. Within Heteropterygidae, O. guangxiensis was the sister clade to O. mouhotii belonging to Dataminae, and the relationship of (Heteropteryginae + (Dataminae + Obriminae)) was recovered.
