Initial Phylotranscriptomic Confirmation of Homoplastic Evolution of the Conspicuous Coloration and Bufoniform Morphology of Pumpkin-Toadlets in the Genus Brachycephalus

The genus Brachycephalus is a fascinating group of miniaturized anurans from the Brazilian Atlantic Forest, comprising the conspicuous, brightly colored pumpkin-toadlets and the cryptic flea-toads. Pumpkin-toadlets are known to contain tetrodotoxins and therefore, their bright colors may perform an aposematic function. Previous studies based on a limited number of mitochondrial and nuclear-encoded markers supported the existence of two clades containing species of pumpkin-toadlet phenotype, but deep nodes remained largely unresolved or conflicting between data sets. We use new RNAseq data of 17 individuals from nine Brachycephalus species to infer their evolutionary relationships from a phylogenomic perspective. Analyses of almost 5300 nuclear-encoded ortholog protein-coding genes and full mitochondrial genomes confirmed the existence of two separate pumpkin-toadlet clades, suggesting the convergent evolution (or multiple reversals) of the bufoniform morphology, conspicuous coloration, and probably toxicity. In addition, the study of the mitochondrial gene order revealed that three species (B. hermogenesi, B. pitanga, and B. rotenbergae) display translocations of different tRNAs (NCY and CYA) from the WANCY tRNA cluster to a position between the genes ATP6 and COIII, showing a new mitochondrial gene order arrangement for vertebrates. The newly clarified phylogeny suggests that Brachycephalus has the potential to become a promising model taxon to understand the evolution of coloration, body plan and toxicity. Given that toxicity information is available for only few species of Brachycephalus, without data for any flea-toad species, we also emphasize the need for a wider screening of toxicity across species, together with more in-depth functional and ecological study of their phenotypes.

Mitogenomic phylogeny and fossil-calibrated mutation rates for all F- and M-type mtDNA genes of the largest freshwater mussel family, the Unionidae (Bivalvia)

The Unionidae represent an excellent model taxon for unravelling the drivers of freshwater diversity, but, phylogeographic studies on Southeast Asian taxa are hampered by lack of a comprehensive phylogeny and mutation rates for this fauna. We present complete female- (F) and male-type (M) mitogenomes of four genera of the Southeast Asian clade Contradentini+Rectidentini. We calculate substitution rates for the mitogenome, the 13 protein-coding genes, the two ribosomal units and three commonly used fragments (co1, nd1 and 16S) of both F- and M-mtDNA, based on a fossil-calibrated, mitogenomic phylogeny of the Unionidae. Phylogenetic analyses, including an M+F concatenated dataset, consistently recovers a monophyletic Gonideinae. Subfamily-level topology is congruent with that of a previous nuclear genomic study and with patterns in mitochondrial gene order, suggesting Unionidae F-type 2 as a synapomorphy of the Gonideinae. Our phylogeny indicates that the clades Contradentini+Rectidentini and Lamprotulini+Pseudodontini+Gonideini split in the early Cretaceous (~125 Mya), and that the crown group of Contradentini+Rectidentini originated in the late Cretaceous (~79 Mya). Most gonideine tribes originated during the early Palaeogene. Substitution rates were comparable to those previously published for F-type co1 and 16S for certain Unionidae and Margaritiferidae species (pairs).

Comprehensive Analyses of the Complete Mitochondrial Genome of Figulus binodulus (Coleoptera: Lucanidae)

Figulus binodulus Waterhouse is a small stag beetle distributed in East Asia. We determined the first mitochondrial genome of F. binodulus of which is 16,261-bp long including 13 protein-coding genes, two ribosomal RNA genes, 22 transfer RNAs, and a single large noncoding region of 1,717 bp. Gene order of F. binodulus is identical to the ancestral insect mitochondrial gene order as in most other stag beetle species. All of 22 tRNAs could be shaped into typical clover-leaf structure except trnSer1. Comparative analyses of 21 Lucanidae mitochondrial genomes was conducted in aspect of their length and AT-GC ratio. Nucleotide diversities analyses provide that cox1 and cox2 in Lucanidae are less diverse than those of Scarabaeoidea. Fifty simple sequence repeats (SSRs) were identified on F. binodulus mitochondrial genome. Comparative analysis of SSRs among five mitochondrial genomes displayed similar trend along with SSR types. Figulus binodulus was sister to all other available family Lucanidae species in the phylogenetic tree.

Mitochondrial Genome Diversity in Collembola: Phylogeny, Dating and Gene Order

Collembola (springtails) are an early diverging class of apterygotes, and mark the first substantial radiation of hexapods on land. Despite extensive work, the relationships between major collembolan lineages are still debated and, apart from the Early Devonian fossil Rhyniella praecursor, which demonstrates their antiquity, the time frame of springtail evolution is unknown. In this study, we sequence two new mitochondrial genomes and reanalyze all known Collembola mt-genomes, including selected metagenomic data, to produce an improved phylogenetic hypothesis for the group, develop a tentative time frame for their differentiation, and provide a comprehensive overview of gene order diversity. Our analyses support most taxonomically recognized entities. We find support for an Entomobryomorpha + Symphypleona clade, while the position of Neelipleona could not be assessed with confidence. A Silurian time frame for their basal diversification is recovered, with an indication that divergence times may be fairly old overall. The distribution of mitochondrial gene order indicates the pancrustacean arrangement as plesiomorphic and dominant in the group, with the exception of the family Onychiuridae. We distinguished multiple instances of different arrangements in individual genomes or small clusters. We further discuss the opportunities and drawbacks associated with the inclusion of metagenomic data in a classic study on mitochondrial genome diversity.

Precise annotation of tick mitochondrial genomes reveals multiple STR variation and one transposon-like element

In this study, we used long-PCR amplification combined with Next-Generation Sequencing (NGS) to obtain complete mitochondrial genomes of individual ticks and performed precise annotation of these genomes. These annotations were confirmed by the PacBio full-length transcriptome data to cover both entire strands of the mitochondrial genomes without any gaps or overlaps. Based on these annotations, most of our findings were consistent with those from previous studies. Moreover, two important findings were reported for the first time, contributing fundamental knowledge to mitochondrial biology. The first was the discovery of a transposon-like element that may reveal the mechanisms of mitochondrial gene order rearrangement and genomic structural variation. Another finding was that Short Tandem Repeat (STRs) are the dominant variation type causing mitochondrial sequence diversity within an individual tick, insect, mouse and human. Comparisons between interindividual and intraindividual variation showed that polynucleotides and STRs with longer repeat units had the same variation pattern. Particularly, mitochondria containing deleterious mutations can accumulate in cells and deleterious STR mutations irreversibly change the proteins made from their mRNAs. Therefore, we proposed that deleterious STR mutations in mitochondria cause aging and diseases. This finding helped to ultimately reveal the mechanisms of mitochondrial DNA variation and its consequences (e.g., aging and diseases) in animals. Our study will give rise to the reconsideration of the importance of STRs and a unified study of STR variation with longer and shorter repeat units (particularly polynucleotides) in both nuclear and mitochondrial genomes. The complete mitochondrial genome sequence of Dermacentor silvarum is available at the NCBI GenBank database under the accession number MN347015 and the raw data is available at the NCBI SRA database under the accession number SRP178347.

Comparative Mitochondrial Genome Analysis of the Tobacco Endophytic Fungi Leptosphaerulina Chartarum and Curvularia Trifolii and the Phylogenetic Implications

In tobacco plants, symbiont endophytic fungi are widely distributed in all tissues where they play important roles. It is therefore important to determine the species distribution and characteristics of endophytic fungi in tobacco. Here, two parasitic fungi Leptosphaerulina chartarum and Curvularia trifolii were isolated and identified from normal tobacco tissue. We sequenced the mitogenomes of these two species and analysed their features, gene content, and evolutionary histories. The L. chartarum and C. trifolii mitochondrial genomes were 68,926 bp and 59,100 bp long circular molecules with average GC contents of 28.60% and 29.31%, respectively. The L. chartarum mitogenome contained 36 protein coding genes, 26 tRNA genes, and 2 rRNA genes (rrnL and rrnS), which were located on both strands. The C. trifolii mitogenome contained 26 protein coding genes, 29 tRNA genes, and 2 rRNA genes (rrnL and rrnS). The L. chartarum 26 tRNAs ranged from 70 bp to 84 bp in length, whereas the 29 tRNAs in C. trifolii ranged from 71 bp to 85 bp. L. chartarum and C. trifolii mtDNAs had an identical mitochondrial gene order and orientation and were phylogenetically identified as sisters. These data therefore provide an understanding of the gene content and evolutionary history of species within Pleosporales.