The phylogenetic relationships among the Nymphalidae (Lepidoptera: Papilionoidea) have been controversial in several perspective. The present study sequenced a total of ~ 3,500 bp from cytochrome oxidase subunit I (COI), 16S ribosomal RNA (16S rRNA), and elongation factor-1 alpha (EF-1α) in 80 nymphalid species belonging to seven subfamilies (Linmenitidinae, Heliconiinae, Nymphalinae, Apaturinae, Libytheinae, Satyrinae, and Danainae), along with those of six lycaenid species as outgroups. Phylogenetic analyses via Bayesian Inference (BI) and Maximum Likelihood (ML) algorithms concordantly supported the subfamilial relationships of (((((Linmenitidinae + Heliconiinae) + (Nymphalinae + Apaturinae)) + Libytheinae) + Satyrinae) + Danainae), with high nodal support for monophyletic subfamilies and tribes. This result is largely consistent with a previous study performed with a substantially large sequence information and morphological characters, except for the position of Libytheinae that has previously been placed as the sister to all reminder of Nymphalidae.

The phylogenetic relationships among true butterfly families (superfamily Papilionoidea) have been a matter of substantial controversy, and that debate has led to several competing hypotheses. Two of the most compelling of those hypotheses involve the relationships of (Nymphalidae + Lycaenidae) + (Pieridae + Papilionidae) and (((Nymphalidae + Lycaenidae) + Pieridae) + Papilionidae). In this study, approximately 3,500 nucleotide sequences from cytochrome oxidase subunit I (COI), 16S ribosomal RNA (16S rRNA), and elongation factor-1 alpha (EF-1α) were sequenced from 83 species belonging to four true butterfly families, along with those of eight outgroup species belonging to the skipper family (superfamily Hesperioidea). These sequences were subjected to phylogenetic reconstruction via Bayesian Inference (BI), Maximum Likelihood (ML), and Maximum Parsimony (MP) algorithms. All phylogenetic analyses among the four true butterfly families strongly indicated a sister relationship between the Nymphalidae and Lycaenidae on one hand, and relatively strongly indicated a sister relationship between the Pieridae and Papilionidae on another hand, thus supporting the hypothesis: (Nymphalidae + Lycaenidae) + (Pieridae + Papilionidae).