Gene arrangement in the mitochondrial genome (mitogenome) has been regarded as an important evolutionary event that is useful as a phylogenetic signal. The mountainous duskywing, Erynnis montanus, belongs to a lepidopteran family Hesperiidae. We sequenced 15,530-bp long complete mitogenome of the species. The genome has the typical gene content of animals (13 protein-coding genes, two rRNA genes, 22 tRNA genes, and one major non-coding A+T-rich region). Further, E. montanus mitogenome also contained a high A/T content in the whole genome (81.7%) and the CGA (arginine) as the start codon for the COI gene, as typical in lepidopteran mitogenome. However, unlike other lepidopteran species, including two sequenced skippers, the E. montanus mitogenome has a unique arrangement tRNASer-tRNAAsn, instead of the tRNAAsn-tRNASer found unanimously in other lepidopteran species, providing a new gene arrangement in Lepidoptera. Such rearrangement probably was likely caused by duplication of gene block tRNASer-tRNAAsn and subsequent random loss of tRNAAsn in the first copy and tRNASer in the second copy, resulting in the arrangement tRNASer-tRNAAsn. Considering current phylogenetic relationships among available lepidopteran groups in connection with lepidopteran gene arrangement the new gene arrangement found in E. montanus seems to be apomorphy, requiring cautious interpretation as a phylogenetic signal.

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 complete mitochondrial genome sequence of the nerippe fritillary butterfly, Argynnis nerippe, which is listed as an endangered species in Korea, is described with an emphasis on the A+T-rich region. The 15,140-bp long circular molecule consisted of 13 protein-coding genes, two rRNA genes, 22 tRNA genes and one control region, known in insect as the A+T-rich region, as found in typical metazoans. The 329-bp long A+T-rich region located between srRNA and tRNAMet possessed the highest A/T content (95.7%) than any other region of the genome. Along with the several conserved sequences found typically in the lepidopteran insects the genome contained one tRNAMet-like and tRNALeu(UUR) -like sequence in the A+T-rich region.

The phylogenetic relationships among the Nymphalidae (Lepidoptera: Papilionoidea) have been controversial. The present study sequenced approximately 1,099 bp from cytochrome oxidase subunit I (COI), 1,336 ~ 1,551 bp from 16S ribosomal RNA (16S rRNA), and 1,066 bp from elongation factor-1 alpha (EF-1α) in 80 species belonging to seven subfamilies (Linmenitidinae, Heliconiinae, Nymphalinae, Apaturinae, Libytheinae, Satyrinae, and Danainae) of Nymphalidae, along with those of six lycaenid species as outgroups. The average base compositions for the three genes (COI, 16S rRNA, and EF-1α) are as follows: A (30.6%, 38.8%, and 25.8%), G (14.7, 5.2%, and 23.6%), T (39.8%, 45.2%, and 23.4%), and C (14.9%, 10.8%, and 27.3%). This result shows the A/T bias in the mitochondrial genes, but not for the nuclear EF-1α. Between the two mitochondrial genes, the 16S rRNA gene evidenced a significantly higher A/T content than was detected in the COI gene. These sequences were subjected to phylogenetic reconstruction via Bayesian Inference (BI) and Maximum Likelihood (ML) algorithms. Both analyses concordantly supported the subfamilial relationships of (((((Linmenitidinae + Heliconiinae) + (Nymphalinae + Apaturinae)) + Libytheinae) + Satyrinae) + Danainae), along with highly supported monophyletics of tribes within subfamilies. This result is largely consistent with a previous study performed with a large sequence information and morphological characters, except for the position of Libytheinae, which was suggested to be the basal lineage 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).

Two complete mitochondrial genomes (mitogenomes) of the endangered, lycaenid butterflies, Spindasis takanonis and Protantigius superans (Lepidoptera: Lycaenidae), were sequenced. Each 15,349 bp and 15,248 bp-long genome contained both the lepidopteran specific gene arrangement that differ from the most common arrangement of insects by the movement of tRNAMet to a position 5’-upstream of tRNAIle. Neither of the species have typical COI start codon. Instead, the CGA (arginine) sequence that is commonly present in all other lepidopterans was also found in both lycaenids. The possible binding site for the transcription termination peptide, TACTA sequence, also was well retained in both species. The high A+T-content, which is a characteristic of insect mitogenomes was well reflected in the genomes in the form of higher frequency of codons with A/T nucleotides, severe A/T bias in 3rd codon position, and extremely high A/T content in the A+T-rich region. The 19 bp-long poly-T stretch and the downstream conserved motif ATAG, which were suggested previously to function as a structural signal for minor-strand mtDNA replication, was also well conserved in the A+T-rich region of both lycaenids. Phylogenetic analysis among lepidopteran superfamilies supported the relationships of either (((((Bombycoidea + Geometroidea) + Noctuoidea) + Papilionoidea) + Pyraloidea) + Tortricoidea) by concatenated amino acid sequence or (((((Bombycoidea + Geometroidea) + Noctuoidea) + Pyraloidea) + Papilionoidea) + Tortricoidea) by concatenated nucleotide sequences of 1st and 2nd codon positions of 13 protein-coding genes, two rRNA genes, and 22 tRNA genes, revealing fluctuating positions of Papilionoidea and Pyraloidea between the two data sets.

There has been a substantial controversy on the phylogenetic relationships among butterfly families and several competing phylogenetic hypothesis have been suggested. Among them the relationships of (((Nymphalidae + Lycaenidae) + Pieridae) + Papilionidae) has been further widely accepted. In this study, we sequenced EF1-α, COI, and 16S rRNA from 62 species belonging to four true butterfly families, Papilionoidea. Phylogenetic analyses using BI, ML, and MP showed that the traditionally recognizable families were strongly supported as monophyletic groups, with the exception of Nymphalidae, wherein the singly included species of Danainae was placed as basal lineage of the Nymphalidae + Lycaenidae group. Phylogenetic relationships among families supported the sister group relationship of Nymphalidae and Lycaenidae strongly by all analyses and placed Papilionidae as the most basal lineage of the Papilionoidea. On the other hand, the relationships of Nymphalidae and Lycaenidae group to Pieridae were either unresolved, revealing trichotomy, or the relationships of (((Nymphalidae + Lycaenidae) + Pieridae) + Papilionidae) as previously supported by several morphological and molecular works supported. Detailed within-family relationships among some genera also are shown in the presentation.

The 15,389-bp long complete mitogenome of the endangered red-spotted apollo butterfly, Parnassius bremeri (Lepidoptera: Papilionidae) was determined. This genome has a gene arrangement identical to those of all other sequenced lepidopteran insects, which have the gene order of tRNAMet, tRNAIle, and tRNAGln at the beginning. Due to the uncertainty the start codon for COI gene in insect has been discussed extensively. We propose the CGA sequence as the start codon for COI gene in lepidopteran insects, based on complete mitogenome sequences of lepidopteran insects including our P. bremerii and additional sequences of the COI start region from a diverse taxonomic range of lepidopteran species (a total of 51 species belonging to 15 families). As has been suggested in other sequenced lepidopteran insects the 18 bp-long poly-T stretch and the downstream conserved motif ATAGA that were previously suggested to serve as a structural signal for minor-strand mtDNA replication also was found at the 3’-end region of the P. bremerii A+T-rich region. In an extensive search to find out tRNA-like structure in the A+T-rich region, each one tRNATrp-like sequence and tRNALeu (UUR)-like sequence were found in the P. bremeri A+T-rich region, and most of other sequenced lepidopteran insects were shown to have tRNA-like structure within the A+T-rich region, thereby indicating that such feature is frequent in the lepidopteran A+T-rich region. Phylogenetic analysis using the concatenated 13 amino acid sequences and nucleotide sequences of PCGs of the four macrolepidopteran suferfamilies together with Tortricoidea and Pyraloidea well recovered a monophyly of Papilionoidea and a monophyly of Bombycoidea. However, Geometroidea and Noctuoidea were unexpectedly clustered as one group and placed this group to the sister group to Bombycoidea, instead of Papilionoidea in most analyses.

We analyzed a portion of mitochondrial COI gene sequences (658 bp) to investigate the genetic diversity and geographic variation of the swallowtail butterfly, Papilioxuthus L., and the cabbage butterfly, Pieris rapae (Lepidoptera: Papilionidae). P. xuthus showed a moderate level of sequence divergence (0.91% at maximum) in 15 haplotypes, whereas P. rapae showed a moderate to high level of sequence divergence (1.67% at maximum) in 30 haplotypes, compared with other relevant studies. Analyses of population genetic structure showed that most populations are not genetically differentiated in both species. The distribution pattern of both species appears to be consistent with category IV of the phylogeographic pattern sensu Avise (Avise et al. 1987): a phylogenetic continuity, an absence of regional isolation of mtDNA clones, and extensive distribution of close clones. The observed pattern of genetic diversity and geographic variation of the two butterfly species seems to reflect the abundant habitats, abundant host plants, and flying abilities in connection with the lack of historical biogeographic barriers.

In this study, we determined the complete mitochondrial genome of the jewel beetle, Chrysochroa fulgidissima (Coleoptera: Buprestidae), from four overlapping fragments. The 15,592-bp long C. fulgidissima mitogenome exhibits a gene arrangement and content identical to the most common type in insects. The start codon of the C. fulgidissima COI gene is unusual, in that no typical ATN codon is available. The 875-bp A+T-rich region is the shortest among the coleopteran mitogenomes that have thus far been sequenced in their entirety. The most unusual feature of the genome is the presence of three tRNA-like sequences within the A+T-rich region: two tRNALeu(UUR)-like sequences and one tRNAAsnlike sequence. These sequence stretches evidence the proper anticodon sequence and the potential to form secondary structures, but also harbor many mismatches in the stems. Phylogenetic analysis using a concatenation of 13 amino acid sequences of protein-coding genes among the available sequenced species of coleopteran superfamilies (Buprestoidea and Elateroidea belonging to the infraorder Elateriformnia, and Chrysomeloidea and Tenebrioroidea belonging to the infraorder Cucujoiformia) by Bayesian inference, maximum-parsimony analyses, and maximum-likelihood analysis unexpectedly revealed a lack of support for monophyletic Elateriformia.

The 15,389-bp long complete mitochondrial genome (mitogenome) of the redspotted apollo butterfly, Parnassius bremeri (Lepidoptera: Papilionidae) was determined. This genome has a gene arrangement identical to those of all other sequenced lepidopteran insects, but differs from the most common type, as the result of the movement of tRNAMet to a position 5’-upstream of tRNAIle. No typical start codon of the P. bremeri COI gene is available. Instead, a tetranucleotide, TTAG, which is found at the beginning context of all sequenced lepidopteran insects was tentatively designated as the start codon for P. bremeri COI gene. All protein-coding genes (PCGs), but COII (T) have complete termination codon TAA or TAG. One of the most unusual feature of the P. bremeri mitochondrial genome is the presence of two tRNA-like structures, such as the tRNATrp-like sequence and tRNALeu (UUR)-like sequence with proper anticodon and clover-leaf structures in the 514-bp long A+T-rich region. Furthermore, the A+T-rich region possesses three sequences that have the potential to form stem-and-loop structures, flanked by the conserved sequences, "TA(A)TA" at the 5’ end and "G(A)nT’ at the 3’ end. After more genomic and phylogenetic analyses are performed, further detailed information will be available.

We determined the complete mitochondrial genome (mitogenome) of the jewel beetle, Chrysochroa fulgidissima (Coleoptera: Buprestidae) from two overlapping fragments and subsequent sub fragments. The 15,592-bp long C. fulgidissima mitogenome contains gene arrangement and content identical to the most common arrangement found in insects. Most individual C. fulgidissima mitochondrial (mt) genes were well within the range found in the respective genes of other insects. The 875-bp A+T-rich region is shortest among the coleopteran mitogenomes sequenced in their entirety. The region is interesting in that it contains several stem-and-loop structures and tRNA-like structure found in the A+T-rich regions of other insect mitogenomes. As seen in other insect motogenomes the start codon of C. fulgidissima COI gene also is unusual because no typical start codon is available. Three of the 13 protein-coding genes have incomplete termination codon T or TA. All tRNA formed stable stem-and-loop structure, except for tRNASer(AGN), the DHU arm of which formed a simple loop as seen in many other metazoan mt tRNASer(AGN).