Contribution to the mitogenome diversity in Delphacinae: Phylogenetic and ecological implications

We document the complete (or nearly complete) mitogenomes of 20 Delphacidae taxa, and together with 17 other delphacid mitogenomes currently in GenBank, to reconstruct the phylogeny of the Delphacinae and to investigate mitogenome differences among members of Delphacini, Tropidocephalini and Saccharosydnini. The mitogenomes of the 20 species encode the complete set of 37 genes usually found in animal mitogenomes. The length of complete mitogenomes in Delphacinae ranges from 15,531 to 16,231 bp. The gene order of all newly sequenced mitogenomes are identical, and the mitogenome gene order of Stenocranus matsumurai Metcalf in Stenocraninae has a transposition of tRNA^Thr. The two-clade system in Tropidocephalini was supported with high value (PP = 1, BS = 100), and the monophyly of Bambusiphaga was recovered in this study. Finally, we found that the host shift from plants with a C₃ to a C₄ photosynthetic pathway appears to have occurred independently in several clades.     

The Complete Mitochondrial Genome of two Tetragnatha Spiders (Araneae: Tetragnathidae): Severe Truncation of tRNAs and Novel Gene Rearrangements in Araneae

Mitogenomes can provide information for phylogenetic analysis and evolutionary biology. The Araneae is one of the largest orders of Arachnida with great economic importance. In order to develop mitogenome data for this significant group, we determined the complete mitogenomes of two long jawed spiders Tetragnatha maxillosa and T. nitens and performed the comparative analysis with previously published spider mitogenomes. The circular mitogenomes are 14578 bp long with A+T content of 74.5% in T. maxillosa and 14639 bp long with A+T content of 74.3% in T. nitens, respectively. Both the mitogenomes contain a standard set of 37 genes and an A+T-rich region with the same gene orientation as the other spider mitogenomes, with the exception of the different gene order by the rearrangement of two tRNAs (trnW and trnG). Most of the tRNAs lose TΨC arm stems and have unpaired amino acid acceptor arms. As interesting features, both trnS^AGN and trnS^UCN lack the dihydrouracil (DHU) arm and long tandem repeat units are presented in the A+T-rich region of both the spider mitogenomes. The phylogenetic relationships of 23 spider mitogenomes based on the concatenated nucleotides sequences of 13 protein-coding genes indicated that the mitogenome sequences could be useful in resolving higher-level relationship of Araneae. The molecular information acquired from the results of this study should be very useful for future researches on mitogenomic evolution and genetic diversities in spiders.     

Complete mitochondrial genome of the hemp borer,Grapholita delineana (Lepidoptera: Tortricidae): Gene variability and phylogeny among Grapholita

The mitochondrial genome (mitogenome) has been extensively used in phylogenetics and species-level evolutionary investigations. The lepidopteran family Tortricidae (leaf-roller moths), including the genus Grapholita, contains numerous species of economic importance, but for the majority of Grapholita species, their mitogenomes remain poorly studied. Here, we sequence and annotate the full mitogenome of Grapholita delineana, an important pest of hemp worldwide and compare it with the mitogenomes of two congeneric species available from GenBank. The G. delineana mitogenome is 15,599 bp long, including 37 typical mitochondrial genes and an A + T-rich region. Gene content, order and orientation are identical to other reported tortricid mitogenomes. Analyses of nucleotide diversity, Ka/Ks, genetic distance and number of variable sites together suggest that nad6 is the fastest-evolving gene among the mitochondrial PCGs of Grapholita. Our analyses indicate that Grapholita, as presently defined, is not monophyletic, confirming previous morphological and multiple-gene studies, using mitogenomic evidence. Our study provides information on comparative mitogenomics of Tortricidae especially Grapholita.     

Mitochondrial DNA Genomes Organization and Phylogenetic Relationships Analysis of Eight Anemonefishes (Pomacentridae: Amphiprioninae)

Anemonefishes (Pomacentridae Amphiprioninae) are a group of 30 valid coral reef fish species with their phylogenetic relationships still under debate. The eight available mitogenomes of anemonefishes were used to reconstruct the molecular phylogenetic tree; six were obtained from this study (Amphiprion clarkii, A. frenatus, A. percula, A. perideraion, A. polymnus and Premnas biaculeatus) and two from GenBank (A. bicinctus and A. ocellaris). The seven Amphiprion species represent all four subgenera and P. biaculeatus is the only species from Premnas. The eight mitogenomes of anemonefishes encoded 13 protein-coding genes, two rRNA genes, 22 tRNA genes and two main non-coding regions, with the gene arrangement and translation direction basically identical to other typical vertebrate mitogenomes. Among the 13 protein-coding genes, A. ocellaris (AP006017) and A. percula (KJ174497) had the same length in ND5 with 1,866 bp, which were three nucleotides less than the other six anemonefishes. Both structures of ND5, however, could translate to amino acid successfully. Only four mitogenomes had the tandem repeats in D-loop; the tandem repeats were located in downstream after Conserved Sequence Block rather than the upstream and repeated in a simply way. The phylogenetic utility was tested with Bayesian and Maximum Likelihood methods using all 13 protein-coding genes. The results strongly supported that the subfamily Amphiprioninae was monophyletic and P. biaculeatus should be assigned to the genus Amphiprion. Premnas biaculeatus with the percula complex were revealed to be the ancient anemonefish species. The tree forms of ND1, COIII, ND4, Cytb, Cytb+12S rRNA, Cytb+COI and Cytb+COI+12S rRNA were similar to that 13 protein-coding genes, therefore, we suggested that the suitable single mitochondrial gene for phylogenetic analysis of anemonefishes maybe Cytb. Additional mitogenomes of anemonefishes with a combination of nuclear markers will be useful to substantiate these conclusions in future studies.     

The evolution of mitochondrial genomes in modern frogs (Neobatrachia): nonadaptive evolution of mitochondrial genome reorganization

Background

Although mitochondrial (mt) gene order is highly conserved among vertebrates, widespread gene rearrangements occur in anurans, especially in neobatrachians. Protein coding genes in the mitogenome experience adaptive or purifying selection, yet the role that selection plays on genomic reorganization remains unclear. We sequence the mitogenomes of three species of Glandirana and hot spots of gene rearrangements of 20 frog species to investigate the diversity of mitogenomic reorganization in the Neobatrachia. By combing these data with other mitogenomes in GenBank, we evaluate if selective pressures or functional constraints act on mitogenomic reorganization in the Neobatrachia. We also look for correlations between tRNA positions and codon usage.

Results

Gene organization in Glandirana was typical of neobatrachian mitogenomes except for the presence of pseudogene trnS (AGY). Surveyed ranids largely exhibited gene arrangements typical of neobatrachian mtDNA although some gene rearrangements occurred. The correlation between codon usage and tRNA positions in neobatrachians was weak, and did not increase after identifying recurrent rearrangements as revealed by basal neobatrachians. Codon usage and tRNA positions were not significantly correlated when considering tRNA gene duplications or losses. Change in number of tRNA gene copies, which was driven by genomic reorganization, did not influence codon usage bias. Nucleotide substitution rates and d_N/d_S ratios were higher in neobatrachian mitogenomes than in archaeobatrachians, but the rates of mitogenomic reorganization and mt nucleotide diversity were not significantly correlated.

Conclusions

No evidence suggests that adaptive selection drove the reorganization of neobatrachian mitogenomes. In contrast, protein-coding genes that function in metabolism showed evidence for purifying selection, and some functional constraints appear to act on the organization of rRNA and tRNA genes. As important nonadaptive forces, genetic drift and mutation pressure may drive the fixation and evolution of mitogenomic reorganizations.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-691) contains supplementary material, which is available to authorized users.     

The complete mitochondrial genome of Mahanta tanyae compared with other zygaenoid moths (Lepidoptera: Zygaenoidea)

The complete mitochondrial genome (mitogenome) of Mahanta tanyae was sequenced and extensively compared with all seven additionally reported zygaenoid mitogenomes. The M. tanyae mitogenome is circular, double-stranded, and 15,323 bp long. Gene content, gene order, and orientation are all typical of Lepidoptera, despite the existence of gene rearrangements for some other zygaenoid mitogenomes. Comparative analyses further showed that the incomplete termination codon T is consistently recognized in the mitochondrial cox1, cox2 and nad4 genes of all zygaenoid species, as well as in the nad5 gene in two limacodid species. Among 13 protein-coding genes, nad6 exhibits the highest evolutionary rate. The structure for each tRNA is highly conserved, including loss of the dihydorouidine (DHU) arm in trnS1 (AGN), but remarkable nucleotide variation exists, primarily in the pseudouridine (TψC) loops. Interestingly, in four species of Zygaenidae, the anticodons for trnS1 (AGN) are consistently UCU, instead of the routinely used codon GCU, in all three species of Limacodidae. In the intergenic region between trnS2 and nad1, a short sequence before the motif “ATACTAA” is present in the M. tanyae mitogenome that is unique among reported zygaenoid mitogenomes. In the A + T-rich region between the motif “ATTTA” and the microsatellite (AT)_n element, some nucleotides were present for most zygaenoid mitogenomes, which is, to our knowledge, rare even in reported lepidopteran mitogenomes. Phylogenetic analyses based on the combined 37 mitochondrial genes confirmed the position of M. tanyae in Limacodidae of the Zygaenoidea.     

Two novel mitogenomes of Dipodidae species and phylogeny of Rodentia inferred from the complete mitogenomes

Two novel mitogenomes of Eozapus setchuanus (KJ648495) and Sicista concolor (KJ648496) were reported and their total lengths were 16,630 bp and 16,493 bp, respectively. Both mitogenomes which were analogous to other rodent mitogenomes, contained 13 protein-coding genes, 22 tRNAs, 2 rRNAs, and a control region. Specifically, the ND2 gene of S. concolor has three amino acids lesser than that of two other Dipodidae species (E. setchuanus and Jaculus jaculus) due to a premature termination codon in the 3′ end. We detected a tandem repeat cluster of 221 bp and 274 bp in the control region of S. concolor and E. setchuanus, respectively. Along with phylogenetic relationship analysis, we speculated that the tandem repeats in control regions might be common in Dipodinae species. Our phylogenetic analysis using concatenated mitochondrial gene datasets suggested five suborder and 16 family monophyletic groups in 54 rodent taxa sampled and strongly supported a basal position of the squirrel-related clade (PP = 1; BP = 100). Dipodidae had a sister-group relationship with Muroidea, and Sicistinae was in the base of Dipodidae clade. The complete mitochondrial genomes showed high resolution in deep-level phylogenetic relationship reconstructions of Rodentia.     

Evolutionary relatedness of mackerels of the genus Scomber based on complete mitochondrial genomes: Strong support to the recognition of Atlantic Scomber colias and Pacific Scomber japonicus as distinct species

Mackerels of the genus Scomber are commercially important species, but their taxonomic status is still controversial. Although previous phylogenetic data support the recognition of Atlantic Scomber colias and Pacific Scomber japonicus as separate species, it is only based on the analysis of partial mitochondrial and nuclear DNA sequences. In an attempt to shed light on this relevant issue, we have determined the complete mitochondrial DNA sequence of S. colias, S. japonicus, and Scomber australasicus. The total length of the mitogenomes was 16,568 bp for S. colias and 16,570 bp for both S. japonicus and S. australasicus. All mitogenomes had a gene content (13 protein-coding, 2 rRNAs, and 22 tRNAs) and organization similar to that observed in Scomber scombrus and most other vertebrates. The major noncoding region (control region) ranged between 865 and 866 bp in length and showed the typical conserved blocks. Phylogenetic analyses revealed a monophyletic origin of Scomber species with regard to other scombrid fish. The major finding of this study is that S. colias and S. japonicus were significantly grouped in distinct lineages within Scomber cluster, which phylogenetically constitutes evidence that they may be considered as separate species. Additionally, molecular data here presented provide a useful tool for evolutionary as well as population genetic studies.     

Comparative Analysis of Mitochondrial Genomes of Five Aphid Species (Hemiptera: Aphididae) and Phylogenetic Implications

Insect mitochondrial genomes (mitogenomes) are of great interest in exploring molecular evolution, phylogenetics and population genetics. Only two mitogenomes have been previously released in the insect group Aphididae, which consists of about 5,000 known species including some agricultural, forestry and horticultural pests. Here we report the complete 16,317 bp mitogenome of Cavariella salicicola and two nearly complete mitogenomes of Aphis glycines and Pterocomma pilosum. We also present a first comparative analysis of mitochondrial genomes of aphids. Results showed that aphid mitogenomes share conserved genomic organization, nucleotide and amino acid composition, and codon usage features. All 37 genes usually present in animal mitogenomes were sequenced and annotated. The analysis of gene evolutionary rate revealed the lowest and highest rates for COI and ATP8, respectively. A unique repeat region exclusively in aphid mitogenomes, which included variable numbers of tandem repeats in a lineage-specific manner, was highlighted for the first time. This region may have a function as another origin of replication. Phylogenetic reconstructions based on protein-coding genes and the stem-loop structures of control regions confirmed a sister relationship between Cavariella and pterocommatines. Current evidence suggest that pterocommatines could be formally transferred into Macrosiphini. Our paper also offers methodological instructions for obtaining other Aphididae mitochondrial genomes.     

Characterization of the complete mitochondrial genome of Phodopus roborovskii (Rodentia: Cricetidae) and systematic implications for Cricetinae phylogenetics

Phodopus roborovskii (subfamily Cricetinae) is widely distributed in the northern arid regions of China. This study reports its complete mitochondrial genome (mitogenome) for the first time. The complete sequence was 16,273 bp long, including 13 protein-coding genes, 2 ribosomal RNAs, 22 transfer RNAs, and 1 major noncoding region. The base composition and codon usage were described. The putative origin of replication for the light strand (O_L) of P. roborovskii was approximately 45 bp long and was highly conserved in the stem-loop and adjacent sequences, but the starting sequence of replication varied between genera among Rodentia. We analyzed the three domains of the D-loop region, and the results indicated that the central domain had higher G + C content and lower A + T content than two peripheral domains. Phylogenetic analyses indicated high resolution in four main divergent clades using mitogenomes data within Cricetidae. Within Cricetinae clade, P. roborovskii was at basal position which was in line with previous researches, and it shared a common ancestor with other extant hamsters. This work validated previous molecular and karyotype researches using mitogenomes data, and provided a set of useful data on phylogeny and molecular evolution in Cricetidae species.     

Molecular population genetics of a host‐associated sibling species complex of phytophagous ladybird beetles (Coleoptera: Coccinellidae: Epilachninae)

To clarify evolutionary relationships and assess the intensity and persistence of reproductive isolation because of host fidelity in three closely related host‐associated phytophagous ladybird beetles (Henosepilachna pustulosa, Henosepilachna niponica and Henosepilachna yasutomii), we determined sequences of part of the nuclear DNA internal transcribed spacer‐II (ITS‐2) region (388 bp long) for 70 individuals from seven populations, and part of the mitochondrial cytochrome c oxidase subunit I (COI) gene (985 bp long) for 280 individuals from 28 populations across the three species. We failed to detect any polymorphic sites in the ITS‐2 region. We detected 59 COI haplotypes, among which two were shared by different species. While a haplotype network and a phylogenetic gene tree of the COI haplotypes did not support monophyly for any of the three species, the results of AMOVA did not invalidate the classification of these beetles into three species. The isolation‐with‐migration analytic (IMa) test suggested that gene flow between allopatric species (i.e., H. pustulosa versus H. niponica; H. pustulosa versus H. yasutomii) was well prevented, probably because of their limited dispersal power. On the other hand, the IMa test detected a low level of unidirectional gene flow between the sympatric species H. niponica and H. yasutomii, from the former to the latter. This result was consistent with a survival rate of F1 hybrids between the two species that is higher on the host plant of H. yasutomii than on the host plant of H. niponica. High F_ST values among two sympatric and three nearly sympatric population pairs of H. niponica and H. yasutomii, however, indicated that gene flow between the two species has been quite restricted even if it has occurred. Because no reproductive barriers other than the difference in host plants is known for sympatric H. niponica and H. yasutomii, our results suggest that host differentiation and associated ecological divergence function as an effective reproductive barrier between the two species.     

The mitogenomes of three beetles (Coleoptera: Polyphaga: Cucujiformia): New gene rearrangement and phylogeny

The mitochondrial genome (mitogenome) has been extensively used for studying phylogenetic relationships at different taxonomic levels. Several molecular analyses have been performed, but the phylogenetic relationships among infraorders in Polyphaga have not been well resolved. In this work, three nearly complete mitogenomes of Coleoptera, Sitophilus oryzae, Oryzaephilus surinamensis and Callosobruchus chinensis, were determined. The O. surinamensis and S. oryzae mitogenomes harbor gene content typical of other Polyphaga mitogenomes, while a gene rearrangement (trnQ) was found in the C. chinensis mitogenome. The mitogenomes of these three Coleoptera species each consist of approximately 13 protein-coding genes, 22 tRNA genes, two rRNA genes and one A + T-rich region. Phylogenetic analysis within Polyphaga was carried out based on mitochondrial data. The phylogenetic results within Polyphaga support the basal position of Cyphon sp., which belonged to Scirtoidea, Elateriformia. Within Cucujiformia, monophyletic Curculionoidea, Chrysomeloidea and Tenebrionoidea were confirmed.     

The complete mitochondrial genomes of two vent squat lobsters,Munidopsis lauensis and M. verrilli: Novel gene arrangements and phylogenetic implications

Hydrothermal vents are considered as one of the most extremely harsh environments on the Earth. In this study, the complete mitogenomes of hydrothermal vent squat lobsters, Munidopsis lauensis and M. verrilli, were determined through Illumina sequencing and compared with other available mitogenomes of anomurans. The mitogenomes of M. lauensis (17,483 bp) and M. verrilli (17,636 bp) are the largest among all Anomura mitogenomes, while the A+T contents of M. lauensis (62.40%) and M. verrilli (63.99%) are the lowest. The mitogenomes of M. lauensis and M. verrilli display novel gene arrangements, which might be the result of three tandem duplication–random loss (tdrl) events from the ancestral pancrustacean pattern. The mitochondrial gene orders of M. lauensis and M. verrilli shared the most similarities with S. crosnieri. The phylogenetic analyses based on both gene order data and nucleotide sequences (PCGs and rRNAs) revealed that the two species were closely related to Shinkaia crosnieri. Positive selection analysis revealed that eighteen residues in seven genes (atp8, Cytb, nad3, nad4, nad4l, nad5, and nad6) of the hydrothermal vent anomurans were positively selected sites.     

The complete mitochondrial genome of Tuta absoluta (Lepidoptera: Gelechiidae) and genetic variation in two newly invaded populations in China

Tuta absoluta (Meyrick) (Lepidoptera: Gelechiidae) is a devastating invasive pest worldwide, causing severe damage to tomatoes. Recently, it has been recorded in the northwestern and southwestern parts of China. Here, the mitogenomes and genetic variation of two newly invaded T. absoluta populations in Xinjiang and Yunnan, were determined. The results showed that the complete mitogenome size of T. absoluta is 15298 bp for the individual from Xinjiang and 15296 bp for the individual from Yunnan, which were both longer than the reported mitogenome from Spain (15290 bp). The mitogenome sequences of individuals collected from three locations showed high levels of sequence similarity, except for 8 polymorphic sites, which were in genes cox2 (1 site), cox3 (2 sites), cob (1 site), atp6 (1 site), nad1 (2 sites) and nad5 (1 site). Tuta absoluta mitogenomes share many features with other 6 Gelechiidae mitogenomes, except for several differences in the start and stop codons of protein-coding genes and the length of intergenic spacers. Seven partial mitochondrial genes (cox1, cox2, cox3, atp6, cob, nad1, and nad5) were used for genetic variation analysis, and significant population differentiation was found between the two populations based on cox2, atp6, nad1, and nad5. The complete mitogenomes and sensitive mitochondrial gene markers reported here provide useful data for further population genetics study of this pest.     

Characterization of the complete mitochondrial genome of Japanagallia spinosa and Durgades nigropicta (Hemiptera: Cicadellidae: Megophthalminae)

In this study, we determined and analyzed the complete mitochondrial genomes (mitogenomes) of Japanagallia spinosa and Durgades nigropicta (Hemiptera: Megophthalminae). The circular genome were 15,655 bp long in J. spinosa (GenBank: KY123686) and 15,974 bp long in D. nigropicta (GenBank: KY123687). The J. spinosa and D. nigropicta mitogenomes both contained 37 genes and the gene order was similar to that in other leafhoppers. All of the protein-coding genes started with ATN. In the J. spinosa mitogenome, the nad3, nad4L, and cytb genes used TAG as a stop codon, the atp8 and nad1 genes used TGA, and the cox2 gene used a single T. However, in the D. nigropicta mitogenome, three genes used a single T as the stop codon, whereas the nad3 gene used TAG. We predicted the secondary structures of the rRNAs in J. spinosa and D. nigropicta. The secondary structure of rrnL comprised six domains (domain III is absent in arthropods) with 42 helices and that of rrnS comprised three structural domains with 26 helices. Comparisons of J. spinosa and D. nigropicta detected some differences in H577 and H673. We determined the structural organization of the control regions in the mitogenomes of leafhoppers, where three types of repeat regions were found in most. The phylogenetic relationships between J. spinosa and D. nigropicta with related lineages were reconstructed using Bayesian inference and maximum likelihood analyses. The monophyly of each superfamily considered in this study was confirmed by the clades in the phylogenetic tree. And in this study, Cicadellidae was resolved as monophyletic by the phylogenetic analysis. This mitogenome information for J. spinosa and D. nigropicta could facilitate future studies of mitogenomic diversity and the evolution of related insect lineages.     

Comparative analysis of mitochondrial genomes of Nirvanini and Evacanthini (Hemiptera: Cicadellidae) reveals an explicit evolutionary relationship

Mitogenomes of five leafhopper species, Chudania hellerina and Concaveplana rufolineata in Nirvanini, Carinata rufipenna, Evacanthus danmainus and E. heimianus representing Evacanthini, were sequenced. The lengths of these five mitogenomes range from 15,044 (C. hellerina) to 15,680 bp (E. heimianus). All five mitogenomes exhibit similar base composition, gene size and codon usage of protein-coding genes. All 22 tRNA genes have typical cloverleaf secondary structures, except for trnS1 (AGN) which appears to lack the dihydrouridine arm. The two included Nirvanini species employ the anticodon TCT instead of the commonly used GCT in trnS1 (AGN). Genes nad2, atp8 and nad6 were highly variable while cox1 and cob showed the lowest nucleotide diversity. Phylogenetic analyses of two concatenated nucleotide datasets, incorporating the newly sequenced taxa and other available membracoid mitogenomes, recovered each included leafhopper subfamily as monophyletic with evacanthine tribes Nirvanini and Evacanthini forming monophyletic sister clades. A relationship among Evacanthinae, Cicadellinae and Typhlocybinae received moderate branch support.     

Homologous expression,purification and characterization of a novel high-alkaline and thermal stable lipase from Burkholderia cepacia ATCC 25416

The lipase secreted by Burkholderia cepacia ATCC 25416 was particularly attractive in detergent and leather industry due to its specific characteristics of high alkaline and thermal stability. The lipase gene (lipA), lipase chaperone gene (lipB), and native promoter upstream of lipA were cloned. The lipA was composed of 1095 bp, corresponding to 364 amino acid residues. The lipB located immediately downstream of lipA was composed of 1035 bp, corresponding to 344 amino acid residues. The lipase operon was inserted into broad host vector pBBRMCS1 and electroporated into original strain. The homologous expression of recombinant strain showed a significant increase in the lipase activity. LipA was purified by three-step procedure of ammonium sulfate precipitation, phenyl-sepharose FF and DEAE-sepharose FF. SDS-PAGE showed the molecular mass of the lipase was 33 kDa. The enzyme optimal temperature and pH were 60 °C and 11.0, respectively. The enzyme was stable at 30–70 °C. After incubated in 70 °C for 1 h, enzyme remained 72% of its maximal activity. The enzyme exhibited a good stability at pH 9.0–11.5. The lipase preferentially hydrolyzed medium-chain fatty acid esters. The enzyme was strongly activated by Mg²⁺, Ca²⁺, Cu²⁺, Zn²⁺, Co²⁺, and apparently inhibited by PMSF, EDTA and also DTT with SDS. The enzyme was compatible with various ionic and non-ionic surfactants as well as oxidant H₂O₂. The enzyme had good stability in the low- and non-polar solvents.