Characterization of Four Complete Mitogenomes of Monolepta Species and Their Related Phylogenetic Implications.

Monolepta is one of the diverse genera in the subfamily Galerucinae, including 708 species and 6 sub-species worldwide. To explore the information on the mitogenome characteristics and phylogeny of the section "Monoleptites", especially the genus Monolepta, we obtained the newly completed mitochondrial genomes (mitogenomes) of four Monolepta species using high-throughput sequencing technology. The lengths of these four new mitochondrial genomes are 16,672 bp, 16,965 bp, 16,012 bp, and 15,866 bp in size, respectively. All four mitochondrial genomes include 22 transfer RNA genes (tRNAs), 13 protein-coding genes (PCGs), 2 ribosomal RNA genes (rRNAs), and one control region, which is consistent with other Coleoptera. The results of the nonsynonymous with synonymous substitution rates showed that ND6 had the highest evolution rate, while COI displayed the lowest evolution rate. The substitution saturation of three datasets (13 PCGs_codon1, 13 PCGs_codon2, 13 PCGs_codon3) showed that there was no saturation across all datasets. Phylogenetic analyses based on three datasets (ND1, 15 genes of mitogenomes, and 13 PCGs_AA) were carried out using maximum likelihood (ML) and Bayesian inference (BI) methods. The results showed that mitogenomes had a greater capacity to resolve the main clades than the ND1 gene at the suprageneric and species levels. The section "Monoleptites" was proven to be a monophyletic group, while Monolepta was a non-monophyletic group. Based on ND1 data, the newly sequenced species whose antennal segment 2 was shorter than 3 were split into several clades, while, based on the mitogenomic dataset, the four newly sequenced species had close relationships with Paleosepharia. The species whose antennal segment 2 was as long as 3 were split into two clades, which indicated that the characteristic of "antennal segment 2 as long as 3" of the true "Monolepta" evolved multiple times in several subgroups. Therefore, to explore the relationships among the true Monolepta, the most important thing is to perform a thorough revision of Monolepta and related genera in the future.

The complete mitochondrial genome of the Sideridis albicosta (Lepidoptera: Noctuoidea: Noctuidae).

The complete mitochondrial genome (mitogenome) of Sideridis albicosta is described in this study. The circular molecule is 15,320 bp in length and contains 13 protein-coding genes (PCGs), two rRNA genes, 22 tRNA genes, and one AT-rich region. Thirteen PCGs is 11,252 bp in total, encoding 3723 amino acids. All PCGs start with ATN, except for COI gene starting with CGA; 10 of the 13 PCGs use the typical stop codon TAA, whereas COI, COII, and ND4 stop with a single T. The rrnL and rrnS genes are 1377 bp and 783 bp in length, respectively. The 328 bp AT-rich region contains several structures characteristic of the lepidopterans. New phylogenetic analyses upon mitogenomics would provide us further insights on the taxonomy and phylogeny of Noctuoidea.

The complete mitochondrial genome of comma, polygonia c-aureum (Lepidoptera: Nymphalidae: Nymphalinae).

The mitochondrial genome (mitogenome) of Polygonia c-aureum (Lepidoptera: Nymphalidae: Nymphalinae) is determined to be 15,209 bp in length and shows AT bias (80.6%). Similar to other butterflies, it contains 37 typical mitochondrial genes and one AT-rich region (D-loop). All protein-coding genes (PCGs) started with ATN, except for cox1 gene with CGA(R), which is often found in other butterflies, and seven PCGs harbour the typical stop codon TAA, whereas cox1, cox2, nad3, nad5, nad4 and nad1 end with a single T. The rrnL and rrnS genes are 1332 bp and 773 bp in length, respectively. The 342 bp AT-rich region contains non-repetitive sequences, but harbour several features common to the lepidopterans, including the motif ATAGA followed by a 19-bp poly-T stretch and a microsatellite-like (TA)8 element preceded by the ATTTA motif. The complete mitogenome sequence provided here would be useful for further understanding the taxonomy and phylogeny of Nymphalinae.

The complete mitochondrial genome of the Orange Hairstreak Japonica lutea (Lepidoptera: Lycaenidae: Theclinae).

The complete mitochondrial genome of Japonica lutea is 15,225 bp in length, containing 37 typical insect genes: 13 protein-coding genes, 22 tRNA genes, 2 ribosomal RNA genes and a non-coding AT-rich region. All the 13 PCGs are initiated with ATN, except for COI gene which is started by CGA. Nine PCGs use the complete termination codon (TAN), whereas the COI, COII, ND4 and ND5 genes end with single nucleotide T. In total, 131 bp intergenic spacers and 31 bp overlapping sequences are interspersed throughout the whole genome. The two rRNA genes (lrRNA and srRNA) are 1327 bp and 770 bp in size, with their AT contents of 87.8% and 85.5%, respectively. All tRNAs display typical secondary cloverleaf structures except for tRNASer(AGN) which loses the DHU arm. The 401 bp long AT-rich region contains several features characteristic of the lepidopterans, such as the ATAGA motif followed by a 19 bp poly-T stretch, a microsatellite-like (AT)7 element preceded by the ATTTA motif and a 9 bp poly-A stretch.

Morphological characters are compatible with mitogenomic data in resolving the phylogeny of nymphalid butterflies (lepidoptera: papilionoidea: nymphalidae).

Nymphalidae is the largest family of butterflies with their phylogenetic relationships not adequately approached to date. The mitochondrial genomes (mitogenomes) of 11 new nymphalid species were reported and a comparative mitogenomic analysis was conducted together with other 22 available nymphalid mitogenomes. A phylogenetic analysis of the 33 species from all 13 currently recognized nymphalid subfamilies was done based on the mitogenomic data set with three Lycaenidae species as the outgroups. The mitogenome comparison showed that the eleven new mitogenomes were similar with those of other butterflies in gene content and order. The reconstructed phylogenetic trees reveal that the nymphalids are made up of five major clades (the nymphaline, heliconiine, satyrine, danaine and libytheine clades), with sister relationship between subfamilies Cyrestinae and Biblidinae, and most likely between subfamilies Morphinae and Satyrinae. This whole mitogenome-based phylogeny is generally congruent with those of former studies based on nuclear-gene and mitogenomic analyses, but differs considerably from the result of morphological cladistic analysis, such as the basal position of Libytheinae in morpho-phylogeny is not confirmed in molecular studies. However, we found that the mitogenomic phylogeny established herein is compatible with selected morphological characters (including developmental and adult morpho-characters).

The complete mitochondrial genome of Danaus chrysippus (Lepidoptera: Nymphalidae: Danainae).

The complete mitochondrial genome sequence of Danaus chrysippus (Lepidoptera: Nymphalidae: Danainae) was determined. The 15,236 bp long genome encodes 13 putative proteins, two ribosomal RNAs, 22 tRNAs and a non-coding AT-rich region. Its gene arrangement pattern is identical to most of other lepidopteran species. All protein-coding genes start with a typical ATN codon with the exception of COI gene which uses CGA as its initial codon; all PCGs terminate in the common stop TAA or TAG, except COI, COII, ND5 and ND4 which use single T as their stop codons. A total of 102 bp intergenic spacers and a total of 33 bp overlapping sequences are interspersed throughout the whole genome. The mitogenome harbors 22 txRNAs as those of most insect species and all tRNA genes evidence the typical clover leaf secondary structures with the exception of tRNAser (AGN) who loses its dihydrouridine (DHU) arm. The lrRNA and srRNA genes are 1339 and 783 bp, with the AT contents of 84.1 and 84.8%, respectively. The non-coding AT-rich region is 418 bp long, and contains the motif ATAGA followed by a 21-bp poly-T stretch and a microsatellite-like (AT)9 element preceded by the ATTTA motif.

The complete mitochondrial genomes of the Fenton's wood white, Leptidea morsei, and the lemon emigrant, Catopsilia pomona.

The complete mitochondrial genomes of Leptidea morsei Fenton (Lepidoptera: Pieridae: Dis-morphiinae) and Catopsilia pomona (F.) (Lepidoptera: Pieridae: Coliadinae) were determined to be 15,122 and 15,142 bp in length, respectively, with that of L. morsei being the smallest among all known butterflies. Both mitogenomes contained 37 genes and an A+T-rich region, with the gene order identical to those of other butterflies, except for the presence of a tRNA-like insertion, tRNA(Leu) (UUR), in C. pomona. The nucleotide compositions of both genomes were higher in A and T (80.2% for L. morsei and 81.3% for C. pomona) than C and G; the A+T bias had a significant effect on the codon usage and the amino acid composition. The protein-coding genes utilized the standard mitochondrial start codon ATN, except the COI gene using CGA as the initiation codon, as reported in other butterflies. The intergenic spacer sequence between the tRNA(Ser) (UCN) and ND1 genes contained the ATACTAA motif. The A+T-rich region harbored a poly-T stretch and a conserved ATAGA motif located at the end of the region. In addition, there was a triplicated 23 bp repeat and a microsatellite-like (TA)9(AT)3 element in the A+T-rich region of the L. morsei mitogenome, while in C. pomona, there was a duplicated 24 bp repeat element and a microsatellite-like (TA)9 element. The phylogenetic trees of the main butterfly lineages (Hesperiidae, Papilionidae, Pieridae, Nymphalidae, Lycaenidae, and Riodinidae) were reconstructed with maximum likelihood and Bayesian inference methods based on the 13 concatenated nucleotide sequences of protein-coding genes, and both trees showed that the Pieridae family is sister to Lycaenidae. Although this result contradicts the traditional morphologically based views, it agrees with other recent studies based on mitochondrial genomic data.

The complete mitochondrial genome of the Hybris subjacens (Neuroptera:Ascalaphidae).

The mitochondrial genome of Hybris subjacens (Neuroptera: Ascalaphidae) is a circular molecule of 15,873 bp in length, containing 37 typical animal mitochondrial genes: 13 protein-coding genes (PCGs), 2 ribosomal RNAs, 22 transfer RNAs and a non-coding AT-rich region. Its gene order and arrangement are identical to the common type found in most insect mitogenomes. All PCGs start with a typical ATN codon except for COI and ND1 which use CTT and TTG as their start codon, respectively; all PCGs terminate in the common stop codon TAA or TAG, except for the COI and ND5 which use single T as their stop codons. The non-coding AT-rich region is 1051 bp long, located between rrnS and tRNA(lle) genes. It contains some structures of repeated motifs and microsatellite-like elements characteristic of the neuropterids.

The first mitochondrial genome for the butterfly family Riodinidae (Abisara fylloides) and its systematic implications.

The Riodinidae is one of the lepidopteran butterfly families. This study describes the complete mitochondrial genome of the butterfly species Abisara fylloides, the first mitochondrial genome of the Riodinidae family. The results show that the entire mitochondrial genome of A. fylloides is 15 301 bp in length, and contains 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA genes and a 423 bp A+T-rich region. The gene content, orientation and order are identical to the majority of other lepidopteran insects. Phylogenetic reconstruction was conducted using the concatenated 13 protein-coding gene (PCG) sequences of 19 available butterfly species covering all the five butterfly families (Papilionidae, Nymphalidae, Peridae, Lycaenidae and Riodinidae). Both maximum likelihood and Bayesian inference analyses highly supported the monophyly of Lycaenidae+Riodinidae, which was standing as the sister of Nymphalidae. In addition, we propose that the riodinids be categorized into the family Lycaenidae as a subfamilial taxon. The Riodinidae is one of the lepidopteran butterfly families. This study describes the complete mitochondrial genome of the butterfly species Abisara fylloides, the first mitochondrial genome of the Riodinidae family. The results show that the entire mitochondrial genome of A. fylloides is 15 301 bp in length, and contains 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA genes and a 423 bp A+T-rich region. The gene content, orientation and order are identical to the majority of other lepidopteran insects. Phylogenetic reconstruction was conducted using the concatenated 13 protein-coding gene (PCG) sequences of 19 available butterfly species covering all the five butterfly families (Papilionidae, Nymphalidae, Peridae, Lycaenidae and Riodinidae). Both maximum likelihood and Bayesian inference analyses highly supported the monophyly of Lycaenidae+Riodinidae, which was standing as the sister of Nymphalidae. In addition, we propose that the riodinids be categorized into the family Lycaenidae as a subfamilial taxon.

The complete mitochondrial genome of Hebomoia glaucippe (Lepidoptera: Pieridae).

The mitochondrial genome of Hebomoia glaucippe (Lepidoptera: Pieridae) is a circular molecule of 15,701 bp in length, containing 37 typical mitochondrial genes and an AT-rich region. Its gene arrangement pattern is identical with those of other butterfly species. All protein-coding genes start with ATN start codon except for the cox1 gene, which uses CGA as the initiation codon. A total of 109 bp intergenic spacers and a total of 38 bp overlapping sequences are interspersed throughout the whole genome. The 899-bp long AT-rich region is the second longest among the completely sequenced lepidopteran insects, and contains the motif ATAGA followed by an 19-bp poly-T stretch, two microsatellite-like (AT)(10) elements, a duplicated 271-bp repeat element, and two extra tRNA(Leu)(UUR)-like genes.

Complete mitochondrial genome of the Common Evening Brown, Melanitis leda Linnaeus (Lepidoptera: Nymphalidae: Satyrinae).

The complete mitochondrial genome (mitogenome) of Melanitis leda (Lepidoptera: Nymphalidae: Satyrinae) is a circular molecule of 15,122 bp in length, containing 37 typical animal mitochondrial genes and 1 control region, known in insects as the AT-rich region. Its gene content and order are identical to all other available butterfly mitogenomes. All protein-coding genes (PCGs) start with a typical ATN initiation codon, except for COI, which is initiated by the CGA codon as observed in other butterfly species. A total of 97 bp of intergenic spacers are interspersed in 11 regions, ranging in size from 1 to 45 bp. The 314-bp-long AT-rich region is the smallest of all the butterfly corresponding regions available and contains some conserved structures similar to those found in other butterfly mitogenomes, including the motif ATAGA followed by a 19-bp poly-T stretch and a microsatellite-like (AT)6 element preceded by the ATTTA motif.

Complete mitogenome of the Painted Jezebel, Delias hyparete Linnaeus (Lepidoptera: Pieridae) and its comparison with other butterfly species.

In the present study, we report the first complete mitochondrial genome (mitogenome) of the Painted Jezebel, Delias hyparete. The mitogenome of Delias hyparete is 15 186 bp in length, and has typical sets of 37 genes: 13 protein-coding genes (PCGs), 2 ribosomal RNAs, 22 transfer RNAs and a non-coding A+T-rich region. All protein-coding genes are initiated by ATN codons, except for COI, which is tentatively designated by the CGA codon, as observed in other butterfly species. A total of 10 PCGs harbored the complete termination codon TAA or TAG, while the COI, COII and ND5 genes ended at a single T residue. All 22 tRNA genes show typical clover structures, with the exception of the tRNA(Ser(AGN)) which lacks the dihydrouridine (DHU) stem and is instead replaced by a simple loop. Thirteen intergenic spacers totaling 153 bp, and 13 overlapping regions totaling 46 bp are scattered throughout the whole genome. The 377 bp long of D. hyparete A+T-rich region is not comprised of large repetitive sequences, but harbors several features characteristic of the lepidopteran insects, including the motif ATAGA followed by an 18 bp poly-T stretch, a microsatellite-like (AT)(5) element preceded by the ATTTA motif, an 10 bp polyA-like stretch (AAAAATAAAA) present immediately upstream tRNA(Met).

Complete mitochondrial genome of the five-dot sergeant Parathyma sulpitia (Nymphalidae: Limenitidinae) and its phylogenetic implications.

The complete mitochondrial genome of the Parathyma sulpitia (Lepidoptera, Nymphalidae, Limenitidinae) was determined. The entire mitochondrial DNA (mtDNA) molecule was 15 268 bp in size. Its gene content and organization were the same as those of other lepidopteran species, except for the presence of the 121 bp long intergenic spacer between trnS1(AGN) and trnE. The 13 protein-coding genes (PCGs) started with the typical ATN codon, with the exception of the cox1 gene that used CGA as its initial codon. In addition, all protein-coding genes terminated at the common stop codon TAA, except the nad4 gene which used a single T as its terminating codon. All 22 tRNA genes possessed the typical clover leaf secondary structure except for trnS1(AGN), which had a simple loop with the absence of the DHU stem. Excluding the A+T-rich region, the mtDNA genome of P. sulpitia harbored 11 intergenic spacers, the longest of which was 121 bp long with the highest A+T content (100%), located between trnS1(AGN) and trnE. As in other lepidopteran species, there was an 18-bp poly-T stretch at the 3'-end of the A+T-rich region, and there were a few short microsatellite-like repeat regions without conspicuous macro-repeats in the A+T-rich region. The phylogenetic analyses of the published complete mt genomes from nine Nymphalidae species were conducted using the concatenated sequences of 13 PCGs with maximum likelihood and Bayesian inference methods. The results indicated that Limenitidinae was a sister to the Heliconiinae among the main Nymphalidae lineages in this study, strongly supporting the results of previous molecular data, while contradicting speculations based on morphological characters.

Complete mitogenome of the Lesser Purple Emperor Apatura ilia (Lepidoptera: Nymphalidae: Apaturinae) and comparison with other nymphalid butterflies.

The complete mitochondrial genome of Apatura ilia (GenBank accession no. JF437925) was determined as a circular DNA molecule of 15 242 bp, with common genes of 13 putative proteins, 2 rRNAs, and 22 tRNAs and of the same gene arrangement as in other sequenced lepidopterans. All protein-coding genes had the typical start codon ATN, except for the COI's using CGA as its start codon as previously demonstrated in other lepidopteran species. The comparison of the nucleotide sequences of the A. ilia mitogenome with ten other Nymphalidae species showed nearly identical gene orientation and arrangement, with only a few alterations in non-coding fragments. The nucleotide composition and codon frequency all fell into the range estimated for the order Lepidoptera. The A. ilia mitochondrial genome had the canonical set of 22 tRNA genes folded in the typical cloverleaf structure, with an unique exception of tRNA(Ser) (AGN). The mitochondrial genes from A. ilia were overlapped in a total of 33 bp at 9 locations, as well as interleaved with a total of 155 bp intergenic spacers, spread over 12 regions with the size ranging from 1 to 49 bp. Furthermore, the spacer between ND6 and Cyt b harbored a microsatellite-like repeat (TA)(23) not found in other completely sequenced nymphalid genomes. The 403 bp AT-rich region harbored two conserved motifs (ATAGA, ATTTA), a 21 bp polyT stretch, a 10 bp poly-A region, along with two microsatellite-like repeats ( (TA)(10) and (TA)(7)), as detected in other nymphalid butterflies.

Complete mitochondrial genome of the laced fritillary Argyreus hyperbius (Lepidoptera: Nymphalidae).

We investigated the complete mitochondrial genome (mitogenome) of Argyreus hyperbius. The 15156 bp long genome harbored the gene content (13 protein coding genes, 22 tRNA genes, 2 rRNA genes and an A+T-rich region) and the gene arrangement was identical to all known lepidopteran mitogenomes. Mitogenome sequence nucleotide organization and codon usage analyses showed that the genome had a strong A+T bias, accounting for A+T content of 80.8%, with a small negative AT skew (-0.019). Eleven intergenic spacers totaling 96 bp, and 14 overlapping regions totaling 34 bp were scattered throughout the whole genome. As has been observed in other lepidopteran species, 12 of the 13 protein-coding genes (PCGs) were initiated by ATN codons, while the COI gene was tentatively designated by the CGA codon. A total of 11 PCGs harbored the complete termination codon TAA, while the COI and COII genes ended at a single T residue. All of the 22 tRNA genes showed typical clover structures except that the tRNA(Ser(AGN)) lacks the dihydrouridine (DHU) stem which is replaced by a simple loop. The intergenic spacer sequence between the tRNA(Ser(AGN)) and ND1 also contained the ATACTAA motif, which is conserved in all other lepidopterans as well. Additionally, the 349 bp A+T-rich region was not comprised of large tandem repetitive sequences, but harbored a few structures common to other lepidopteran insects, such as the motif ATAGA followed by a 20 bp poly-T stretch, a microsatellite-like (AT)(9) element preceded by the ATTTA motif, and a 5 bp poly-A site present immediately upstream of tRNA(Met). The mitochondrial genomic sequence features found in this study not only contribute to genetic diversity information of the group, but also are useful in future studies of the endangered nymphalid butterfly in population genetic dynamics, species conservation, phylogeography and evolution.