Novel mitochondrial gene rearrangements pattern in the millipede Polydesmus sp. GZCS‐2019 and phylogenetic analysis of the Myriapoda

The subphylum Myriapoda included four extant classes (Chilopoda, Symphyla, Diplopoda, and Pauropoda). Due to the limitation of taxon sampling, the phylogenetic relationships within Myriapoda remained contentious, especially for Diplopoda. Herein, we determined the complete mitochondrial genome of Polydesmus sp. GZCS‐2019 (Myriapoda: Polydesmida) and the mitochondrial genomes are circular molecules of 15,036 bp, with all genes encoded on + strand. The A+T content is 66.1%, making the chain asymmetric, and exhibits negative AT‐skew (−0.236). Several genes rearrangements were detected and we propose a new rearrangement model: “TD (N\R) L + C” based on the genome‐scale duplication + (non‐random/random) loss + recombination. Phylogenetic analyses demonstrated that Chilopoda and Symphyla both were monophyletic group, whereas Pauropoda was embedded in Diplopoda to form the Dignatha. Divergence time showed the first split of Myriapoda occurred between the Chilopoda and other classes (Wenlock period of Silurian). We combine phylogenetic analysis, divergence time, and gene arrangement to yield valuable insights into the evolutionary history and classification relationship of Myriapoda and these results support a monophyletic Progoneata and the relationship (Chilopoda + (Symphyla + (Diplopoda + Pauropoda))) within myriapod. Our results help to better explain the gene rearrangement events of the invertebrate mitogenome and lay the foundation for further phylogenetic study of Myriapoda.     

The geological record and phylogeny of the Myriapoda

We review issues of myriapod phylogeny, from the position of the Myriapoda amongst arthropods to the relationships of the orders of the classes Chilopoda and Diplopoda. The fossil record of each myriapod class is reviewed, with an emphasis on developments since 1997. We accept as working hypotheses that Myriapoda is monophyletic and belongs in Mandibulata, that the classes of Myriapoda are monophyletic, and that they are related as (Chilopoda (Symphyla (Diplopoda + Pauropoda))). The most pressing challenges to these hypotheses are some molecular and developmental evidence for an alliance between myriapods and chelicerates, and the attraction of symphylans to pauropods in some molecular analyses. While the phylogeny of the orders of Chilopoda appears settled, the relationships within Diplopoda remain unclear at several levels. Chilopoda and Diplopoda have a relatively sparse representation as fossils, and Symphyla and Pauropoda fossils are known only from Tertiary ambers. Fossils are difficult to place in trees based on living forms because many morphological characters are not very likely to be preserved in the fossils; as a consequence, most diplopod fossils have been placed in extinct higher taxa. Nevertheless, important information from diplopod fossils includes the first documented occurrence of air-breathing, and the first evidence for the use of a chemical defense. Stem-group myriapods are unknown, but evidence suggests the group must have arisen in the Early Cambrian, with a major period of cladogenesis in the Late Ordovician and early Silurian. Large terrestrial myriapods were on land at least by mid-Silurian.     

The complete mitochondrial genome of Pauropus longiramus (Myriapoda: Pauropoda): implications on early diversification of the myriapods revealed from comparative analysis

Myriapods are among the earliest arthropods and may have evolved to become part of the terrestrial biota more than 400 million years ago. A noticeable lack of mitochondrial genome data from Pauropoda hampers phylogenetic and evolutionary studies within the subphylum Myriapoda. We sequenced the first complete mitochondrial genome of a microscopic pauropod, Pauropus longiramus (Arthropoda: Myriapoda), and conducted comprehensive mitogenomic analyses across the Myriapoda. The pauropod mitochondrial genome is a circular molecule of 14,487 bp long and contains the entire set of thirty-seven genes. Frequent intergenic overlaps occurred between adjacent tRNAs, and between tRNA and protein-coding genes. This is the first example of a mitochondrial genome with multiple intergenic overlaps and reveals a strategy for arthropods to effectively compact the mitochondrial genome by overlapping and truncating tRNA genes with neighbor genes, instead of only truncating tRNAs. Phylogenetic analyses based on protein-coding genes provide strong evidence that the sister group of Pauropoda is Symphyla. Additionally, approximately unbiased (AU) tests strongly support the Progoneata and confirm the basal position of Chilopoda in Myriapoda. This study provides an estimation of myriapod origins around 555 Ma (95% CI: 444-704 Ma) and this date is comparable with that of the Cambrian explosion and candidate myriapod-like fossils. A new time-scale suggests that deep radiations during early myriapod diversification occurred at least three times, not once as previously proposed. A Carboniferous origin of pauropods is congruent with the idea that these taxa are derived, rather than basal, progoneatans.     

A phylogenetic analysis of Myriapoda (Arthropoda) using two nuclear protein-encoding genes

Nucleotide and inferred amino acid sequences from two nuclear protein-encoding genes, elongation factor-aα and RNA polymerase II, were obtained from 34 myriapods and 14 other arthropods to determine phylogenetic relationships among and within the myriapod classes. Phylogenetic analyses using maximum parsimony and maximum likelihood methods recovered all three represented myriapod classes (Chilopoda, Diplopoda, Symphyla) and all multiply sampled chilopod and diplopod orders, often with high node support. In contrast, relationships between classes and between orders were recovered less consistently and node support was typically lower. The temporal structure of phylogenetic diversification in Myriapoda may explain this apparent pattern of the phylogenetic recovery.     

Spermatozoa of the 'primitive type' in Scutigerella (Myriapoda, Symphyla)

The myriapod class Symphyla is of interest in that insects generally are assumed to be derived from symphylan-like ancestors. Male Symphyla form spermatophores that are picked up by the female. Both euspematozoa and paraspermatozoa are formed. In spite of the mode of fertilization their euspermatozoa were found to be of a kind that is typical of aquatic animals, so called 'primitive spermatozoa;' these are characterized by a short sperm head with a bilayered acrosome, a midpiece containing a few unmodified mitochondria, and a 9 + 2 flagellum. Scutigerella are unique among terrestrial arthropods in having 'primitive spermatozoa'; and together with horseshoe crabs they are only the second case in Arthropoda. Two further sperm plesiomorphies not found in other myriapods or insects are (1) the presence of a cytoplasmic canal housing the proximal flagellum and (2) the existence of microtubular triplets in centrioles, one of which acts as a basal body. Symphyla and Diplopoda both have a striated structure in the center of the subacrosomal material. The paraspermatozoa lack acrosome and nucleus but have a prominent crystal, a single mitochondrion, and two membrane systems. The structure of Scutigerella euspermatozoa is consistent with Symphyla being close to the stem group of Myriapoda plus Insecta.     

Chromosome studies in nine species of Austrian Symphyla (Myriapoda,Tracheata, Arthropoda)

The chromosomes of nine species of Austrian Symphyla, namely, Scutigerella causeyae, S. sp., S. sbordonii, S. palmonii, S. carpatica, S. seposita, Hanseniella nivea (fam. Scutigerellidae), Symphylella isabellae and S. vulgaris (fam. Scolopendrellidae) were studied using Giemsa-C-banding and fluorochrome staining techniques. Male diploid chromosome numbers range from 11 to 18. The mechanism of sex determination seems to be variable within this group: S. isabellae (2n=11) might have an XO-type, whereas the other species studied have even diploid chromosome numbers in the male sex. For S. causeyae, specimens with B-chromosomes and an obviously tetraploid form have been detected.     

Myriapod monophyly and relationships among myriapod classes based on nearly complete 28S and 18S rDNA sequences

Myriapods play a pivotal position in the arthropod phylogenetic tree. The monophyly of Myriapoda and its internal relationships have been difficult to resolve. This study combined nearly complete 28S and 18S ribosomal RNA gene sequences (3,826 nt in total) to estimate the phylogenetic position of Myriapoda and phylogenetic relationships among four myriapod classes. Our data set consists of six new myriapod sequences and homologous sequences for 18 additional species available in GenBank. Among the six new myriapod sequences, those of the one pauropod and two symphylans are very important additions because they were such difficult taxa to classify in past molecular-phylogenetic studies. Phylogenetic trees were constructed with maximum parsimony, maximum likelihood, and Bayesian analyses. All methods yielded moderate to strong support for the monophyly of Myriapoda. Symphyla grouped strongly with Pauropoda under all analytical conditions. The KH test rejected the traditional view of Dignatha and Progoneata, and the topology obtained here, though not significantly supported, was Diplopoda versus ((Symphyla + Pauropoda) + Chilopoda).     

Morphological study of the ovary in Hanseniella caldaria (Myriapoda; Symphyla): The position of oocyte-growth and evolution of ovarian structure in Arthropoda

In Arthropoda, the ovary is classified into Chelicerata-type and Mandibulata-type, based on the oocyte-growth position within the ovary. By contrast, oocytes of Diplopoda and Chilopoda grow within the hemocoelic space. However, as the position of oocyte-growth in Symphyla and Pauropoda has not been confirmed, whether the hemocoelic nature of oocyte-growth is common among myriapods remains ambiguous. This study described the ovarian structure of Hanseniella caldaria to reveal the oocyte-growth position in Symphyla. The oocyte is surrounded by the follicle epithelium, and the inner surface of the follicle epithelium, i.e., the space between follicle cells and oocytes, is lined with a basement membrane. The follicle epithelial layer continues to the ovarian epithelium via the follicle extension with a continuous layer of basement membrane. Data on the architecture of the follicle suggest that the follicle pouch opens to the hemocoel. Hence, the oocyte of H. caldaria grows within the hemocoelic space. Based on our findings in H. caldaria and previous studies in a millipede and in centipedes, the hemocoelic nature of oocyte-growth is considered as a common feature among myriapods and a synapomorphy of the Myriapoda for which morphological synapomorphies have been ambiguous.     

Myriapod (Arthropoda,Myriapoda) diversity and distribution in a floodplain forest of the Brazilian Pantanal

This study presents the diversity and distribution of myriapod species associated with different habitats in a seasonally flooded forest composed predominantly of Vochysia divergens Pohl. (Vochysiaceae) in the Brazilian Pantanal. We evaluated species richness distribution across habitats and over different seasonal periods, and also identified possible survival strategies employed during periodic flooding that occur within this forest. A total of 1505 myriapods were sampled, representing four classes and 15 species. Diplopoda and Chilopoda were the most abundant and rich taxa. Pauropoda and Symphyla were represented by only one species each. Species richness was distinctly distributed across habitats evaluated in this forest, as well as over the different seasonal periods. Survival strategies observed in Diplopoda and Chilopoda were primarily associated with vertical migration ranging from the edaphic environment to tree trunks and canopies, and horizontal migration associated with water level before and during the flood period. Species richness was considered low; however, the survival strategies, especially vertical and horizontal migration, demonstrate the adaptation of these species to the seasonal floods of this ecosystem.     

Myriapod metamerism and arthropod segmentation

Outstanding progress in understanding segmentation of tracheate arthropods (Atelocerata), i.e. Chilopoda, Diplopoda, Pauropoda, Symphyla and Insecta, has been gained through experimental studies carried out on a single, very derivative organism, i.e. Drosophila. We stress the need for a broader comparative approach. We have studied the segmental structure of the trunk in geophilomorph centipedes, where we can identify morphogenetic units of two, four, eight or 16 segments. Accordingly, we sketch an improved model for arthropod segmentation, with the following initial steps: (a) biochemical marking of a very few repetitive units (eosegments); (b) iterative duplications of this first periodicity, until the embryo acquires an array of biochemical markings matching the whole number of segments of the future larva or juvenile specimen; (c) transpatterning, stabilization and interpretation of this 'segmental' arrangement; (d) possible repatterning, to give a final repetitive pattern we define as metasegmental. Finally, we express some doubt about the homology between annelid and arthropod segmentation.     

Complete subunit sequences, structure and evolution of the 6 x 6-mer hemocyanin from the common house centipede, Scutigera coleoptrata.

Hemocyanins are large oligomeric copper-containing proteins that serve for the transport of oxygen in many arthropod species. While studied in detail in the Chelicerata and Crustacea, hemocyanins had long been considered unnecessary in the Myriapoda. Here we report the complete molecular structure of the hemocyanin from the common house centipede Scutigera coleoptrata (Myriapoda: Chilopoda), as deduced from 2D-gel electrophoresis, MALDI-TOF mass spectrometry, protein and cDNA sequencing, and homology modeling. This is the first myriapod hemocyanin to be fully sequenced, and allows the investigation of hemocyanin structure-function relationship and evolution. S. coleoptrata hemocyanin is a 6 x 6-mer composed of four distinct subunit types that occur in an approximate 2 : 2 : 1 : 1 ratio and are 49.5-55.5% identical. The cDNA of a fifth, highly diverged, putative hemocyanin was identified that is not included in the native 6 x 6-mer hemocyanin. Phylogenetic analyses show that myriapod hemocyanins are monophyletic, but at least three distinct subunit types evolved before the separation of the Chilopoda and Diplopoda more than 420 million years ago. In contrast to the situation in the Crustacea and Chelicerata, the substitution rates among the myriapod hemocyanin subunits are highly variable. Phylogenetic analyses do not support a common clade of Myriapoda and Hexapoda, whereas there is evidence in favor of monophyletic Mandibulata.     

Novel rearrangements of arthropod mitochondrial DNA detected with long-PCR: applications to arthropod phylogeny and evolution

Rearrangements of mitochondrial DNA gene order have been suggested as a tool for defining the pattern of evolutionary divergence in arthropod taxa. We have employed a combination of highly conserved insect-based polymerase chain reaction (PCR) primers with long-PCR to survey 14 noninsect arthropods for mitochondrial gene rearrangements. The size of the amplified fragments was used to order the primer containing genes. Five chelicerates exhibit amplicons that are consistent with the presumptive ancestral arthropod mtDNA gene order. These five species comprise two soft ticks, two prostriate hard ticks, and an opilionid. Six other chelicerates, all metastriate hard ticks, have a different arrangement that was originally discovered by this procedure and has been previously detailed in a complete mtDNA sequence. Three new arthropod mtDNA gene arrangements are described here. They were discovered in a terrestrial crustacean (Isopoda) and two myriapods (Chilopoda, centipede; Diplopoda, millipede). These rearrangements include major realignments of some of the large coding regions and two possible new positions for the tRNA(Met) (M) gene in arthropods. The long-PCR approach affords an opportunity to quickly screen divergent taxa for major rearrangements. Taxa exhibiting rearrangements can be targeted for DNA sequencing of gene boundaries to establish the details of the mtDNA organization.     

The complete mitochondrial genome of Parafronurus youi (Insecta: Ephemeroptera) and phylogenetic position of the Ephemeroptera

The first complete mitochondrial genome of a mayfly, Parafronurus youi (Arthropoda: Insecta: Pterygota: Ephemeroptera: Heptageniidae), was sequenced using a long PCR-based approach. The genome is a circular molecule of 15,481 bp in length, and encodes the set of 38 genes. Among them, 37 genes are found in other conservative insect mitochondrial genomes, and the 38(th) unique gene is trnM-like (trnM2). The duplication-random loss model can be used to explain one of the translocations at least. The A+T content of the control region is 57%, the lowest proportion detected so far in Hexapoda. Based on the nucleotide dataset and the corresponding amino acid dataset of 12 protein-coding genes, Bayesian inference and maximum likelihood analyses yielded stable support for the relationship of the three basal clades of winged insects as Ephemeroptera+(Odonata+Neoptera).     

Mitochondrial genome sequence evolution in Chlamydomonas

The mitochondrial genomes of the Chlorophyta exhibit significant diversity with respect to gene content and genome compactness; however, quantitative data on the rates of nucleotide substitution in mitochondrial DNA, which might help explain the origin of this diversity, are lacking. To gain insight into the evolutionary forces responsible for mitochondrial genome diversification, we sequenced to near completion the mitochondrial genome of the chlorophyte Chlamydomonas incerta, estimated the evolutionary divergence between Chlamydomonas reinhardtii and C. incerta mitochondrial protein-coding genes and rRNA-coding regions, and compared the relative evolutionary rates in mitochondrial and nuclear genes. Synonymous and nonsynonymous substitution rates do not differ significantly between the mitochondrial and nuclear protein-coding genes. The mitochondrial rRNA-coding regions, however, are evolving much faster than their nuclear counterparts, and this difference might be explained by relaxed functional constraints on the mitochondrial translational apparatus due to the small number of proteins synthesized in Chlamydomonas mitochondria. Substitution rates at synonymous sites in a nonstandard mitochondrial gene (rtl) and at intronic and synonymous sites in nuclear genes expressed at low levels suggest that the mutation rate is similar in these two genetic compartments. Potential evolutionary forces shaping mitochondrial genome evolution in Chlamydomonas are discussed.     

The complete nucleotide sequence of the mitochondrial genome of the oriental fruit fly, Bactrocera dorsalis (Diptera: Tephritidae)

The complete mitochondrial genome of the oriental fruit fly Bactrocera dorsalis s.s. has been sequenced, and is here described and compared with the homologous sequences of Bactrocera oleae and Ceratitis capitata. The genome is a circular molecule of 15,915 bp, and encodes the set of 37 genes generally found in animal mitochondrial genomes. The structure and organization of the molecule is typical and similar to the two closely related species B. oleae and C. capitata, although it presents an interesting case of putative intra-molecular recombination. The relevance of the growing comparative dataset of tephritid complete mitochondrial genomes is discussed in relation to the possibility to develop robust assays for species discrimination in quarantine and agricultural monitoring practices, as well as basic phylogeography/population genetic studies.     

The Complete Mitochondrial Genome of the Stalk-Eyed Bug Chauliops fallax Scott,and the Monophyly of Malcidae (Hemiptera: Heteroptera)

Chauliops fallax Scott, 1874 (Hemiptera: Heteroptera: Malcidae: Chauliopinae) is one of the most destructive insect pests of soybean and rice fields in Asia. Here we sequenced the complete mitochondrial genome of this pest. This genome is 15,739 bp long, with an A+T content of 73.7%, containing 37 typical animal mitochondrial genes and a control region. All genes were arranged in the same order as most of other Heteroptera. A remarkable strand bias was found for all nine protein coding genes (PCGs) encoded by the majority strand were positive AT-skew and negative GC-skew, whereas the reverse were found in the remaining four PCGs encoded by the minority strand and two rRNA genes. The models of secondary structures for the two rRNA genes of sequenced true bugs and Lygaeoidea were predicted. 16S rRNA consisted of six domains (domain III is absent as in other known arthropod mitochondrial genomes) and 45 helices, while three domains and 27 helices for 12S rRNA. The control region consists of five subregions: a microsatellite-like region, a tandem repeats region and other three motifs. The unusual intergenic spacer between tRNA-H and ND4 only found in the species of Lygaeoidea, not in other heteropteran species, may be the synapomorphy of this superfamily. Phylogenetic analyses were carried out based on all the 13 PCGs showed that Chauliopinae was the sister group of Malcinae and the monophyly of Lygaeoidea.