| Summary: | More studies are using mitochondrial genomes of insects to explore the sequence variability, evolutionary traits, monophyly of groups and phylogenetic relationships. Controversies remain on the classification of the Mileewinae and the phylogenetic relationships between Mileewinae and other subfamilies remain ambiguous. In this study, we present two newly completed mitogenomes of Mileewinae (<i>Mileewa rufivena</i> Cai and Kuoh 1997 and <i>Ujna puerana</i> Yang and Meng 2010) and conduct comparative mitogenomic analyses based on several different factors. These species have quite similar features, including their nucleotide content, codon usage of protein genes and the secondary structure of tRNA. Gene arrangement is identical and conserved, the same as the putative ancestral pattern of insects. All protein-coding genes of <i>U. puerana</i> began with the start codon ATN, while 5 <i>Mileewa</i> species had the abnormal initiation codon TTG in <i>ND5</i> and <i>ATP8</i>. Moreover, <i>M. rufivena</i> had an intergenic spacer of 17 bp that could not be found in other mileewine species. Phylogenetic analysis based on three datasets (PCG123, PCG12 and AA) with two methods (maximum likelihood and Bayesian inference) recovered the Mileewinae as a monophyletic group with strong support values. All results in our study indicate that Mileewinae has a closer phylogenetic relationship to Typhlocybinae compared to Cicadellinae. Additionally, six species within Mileewini revealed the relationship (<i>U. puerana</i> + (<i>M. ponta</i> + (<i>M. rufivena</i> + <i>M. alara</i>) + (<i>M. albovittata</i> + <i>M. margheritae</i>))) in most of our phylogenetic trees. These results contribute to the study of the taxonomic status and phylogenetic relationships of Mileewinae.
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