Conserved Gene Content Research Articles

The complete chloroplast genome sequences of five Miscanthus species, and comparative analyses with other grass plastomes

BackgroundMiscanthus has been considered as potentially high-yielding biomass crop with the low-nutrient requirements, high capability of Carbon mitigation and water-use efficiency. Miscanthus species have restricted to investigate functional genomics of their large genome size, which leads to challenges to genomics-aided crop improvement. Sequencing of the plastid genomes of Miscanthus is one of complementary tool making it available for genetic researches. ResultsThe complete chloroplast (cp) genomes of five Miscanthus species were sequenced using Illumina sequencing. Comparisons between the five chloroplast genome sequences revealed their conservation in gene content and order, including the IR/SC boundaries. However, some hotspots in coding regions and intergenic regions also occurred. Specifically a 13 bp deletion in the intergenic region of rps16-trnQ and a 17 bp deletion in rps4-trnT are shared by Miscanthus lutarioriparius L., Miscanthus sacchariflorus N. and Miscanthus × giganteus. In addition, a 39 bp deletion in trnL-trnF is shared by Miscanthus sinensis A. and Miscanthus floridulus W.. Comparisons of complete chloroplast genome of Miscanthus lutarioriparius to five published grass chloroplast genomes also reveal several hotspot regions with high polymorphism. Phylogenetic analysis based on complete chloroplast genomes and coding sequences of 77 protein genes suggest a sister relationship between the Eumiscanthus clade and the Triarrhena clade. ConclusionsThis report is the first comprehensive comparative analyses on Miscanthus species cp genome combining other published grass chloroplast genomes. The availability of these results will supplement valuable genetic information and framework for further phylogeography, population genetics and plastid genetic engineering researches in Miscanthus.sp.

Uncovering dynamic evolution in the plastid genome of seven Ligusticum species provides insights into species discrimination and phylogenetic implications

Ligusticum L., one of the largest members in Apiaceae, encompasses medicinally important plants, the taxonomic statuses of which have been proved to be difficult to resolve. In the current study, the complete chloroplast genomes of seven crucial plants of the best-known herbs in Ligusticum were presented. The seven genomes ranged from 148,275 to 148,564 bp in length with a highly conserved gene content, gene order and genomic arrangement. A shared dramatic decrease in genome size resulted from a lineage-specific inverted repeat (IR) contraction, which could potentially be a promising diagnostic character for taxonomic investigation of Ligusticum, was discovered, without affecting the synonymous rate. Although a higher variability was uncovered in hotspot divergence regions that were unevenly distributed across the chloroplast genome, a concatenated strategy for rapid species identification was proposed because separate fragments inadequately provided variation for fine resolution. Phylogenetic inference using plastid genome-scale data produced a concordant topology receiving a robust support value, which revealed that L. chuanxiong had a closer relationship with L. jeholense than L. sinense, and L. sinense cv. Fuxiong had a closer relationship to L. sinense than L. chuanxiong, for the first time. Our results not only furnish concrete evidence for clarifying Ligusticum taxonomy but also provide a solid foundation for further pharmaphylogenetic investigation.

Deciphering the organelle genomes and transcriptomes of a common ornamental plant Ligustrum quihoui reveals multiple fragments of transposable elements in the mitogenome

Ligustrum quihoui (L. quihoui) is an important hedge material for landscaping and also possesses medicinal value. To generate genomic resources for better understanding the evolutionary history of this important plant, the organelle genomes of L. quihoui are de novo assembled and functionally annotated. Compared with other Oleaceae species, the 163,069 bp chloroplast genome of L. quihoui exhibits a typical quadripartite structure with highly conserved gene content and gene order, while the 848,451 bp mitochondrial genome of L. quihoui exhibits highly divergent genome size and gene content. Codon usage analyses show that genes related with photosynthesis and mitochondrial respiratory chain show inconsistent codon biases. A total of 48,760 bp transposable elements (TEs) fragments and 41,887 bp chloroplast-like sequences are found in the L. quihoui mitochondrial genome. A striking discrepancy of RNA editing between the two organelle genomes is found in L. quihoui, in which 146 mitochondrial editing sites coexist with only 43 such sites in chloroplast. Based on DNA and RNA-Seq data, we propose that GTG may act as the start codon of mitochondrial rpl16 in Oleaceae species. Phylogenetic analysis based on chloroplast genome shows that L. quihoui and L. japonicum form a sister clade within the genus Ligustrum.

The Female-Specific W Chromosomes of Birds Have Conserved Gene Contents but Are Not Feminized.

Sex chromosomes are unique genomic regions with sex-specific or sex-biased inherent patterns and are expected to be more frequently subject to sex-specific selection. Substantial knowledge on the evolutionary patterns of sex-linked genes have been gained from the studies on the male heterogametic systems (XY male, XX female), but the understanding of the role of sex-specific selection in the evolution of female-heterogametic sex chromosomes (ZW female, ZZ male) is limited. Here we collect the W-linked genes of 27 birds, covering the three major avian clades: Neoaves (songbirds), Galloanserae (chicken), and Palaeognathae (ratites and tinamous). We find that the avian W chromosomes exhibit very conserved gene content despite their independent evolution of recombination suppression. The retained W-linked genes have higher dosage-sensitive and higher expression level than the lost genes, suggesting the role of purifying selection in their retention. Moreover, they are not enriched in ancestrally female-biased genes, and have not acquired new ovary-biased expression patterns after becoming W-linked. They are broadly expressed across female tissues, and the expression profile of the W-linked genes in females is not deviated from that of the homologous Z-linked genes. Together, our new analyses suggest that female-specific positive selection on the avian W chromosomes is limited, and the gene content of the W chromosomes is mainly shaped by purifying selection.

Tracing the Diploid Ancestry of the Cultivated Octoploid Strawberry

The commercial strawberry, Fragaria × ananassa, is a recent allo-octoploid that is cultivated worldwide. However, other than Fragaria vesca, which is universally accepted one of its diploid ancestors, its other early diploid progenitors remain unclear. Here, we performed comparative analyses of the genomes of five diploid strawberries, F. iinumae, F. vesca, F. nilgerrensis, F. nubicola, and F. viridis, of which the latter three are newly sequenced. We found that the genomes of these species share highly conserved gene content and gene order. Using an alignment-based approach, we show that F. iinumae and F. vesca are the diploid progenitors to the octoploid F. × ananassa, whereas the other three diploids that we analyzed in this study are not parental species. We generated a fully resolved, dated phylogeny of Fragaria, and determined that the genus arose ∼6.37 Ma. Our results effectively resolve conflicting hypotheses regarding the putative diploid progenitors of the cultivated strawberry, establish a reliable backbone phylogeny for the genus, and provide genetic resources for molecular breeding.

Genomic architecture of endogenous ichnoviruses reveals distinct evolutionary pathways leading to virus domestication in parasitic wasps

BackgroundPolydnaviruses (PDVs) are mutualistic endogenous viruses inoculated by some lineages of parasitoid wasps into their hosts, where they facilitate successful wasp development. PDVs include the ichnoviruses and bracoviruses that originate from independent viral acquisitions in ichneumonid and braconid wasps respectively. PDV genomes are fully incorporated into the wasp genomes and consist of (1) genes involved in viral particle production, which derive from the viral ancestor and are not encapsidated, and (2) proviral segments harboring virulence genes, which are packaged into the viral particle. To help elucidating the mechanisms that have facilitated viral domestication in ichneumonid wasps, we analyzed the structure of the viral insertions by sequencing the whole genome of two ichnovirus-carrying wasp species, Hyposoter didymator and Campoletis sonorensis.ResultsAssemblies with long scaffold sizes allowed us to unravel the organization of the endogenous ichnovirus and revealed considerable dispersion of the viral loci within the wasp genomes. Proviral segments contained species-specific sets of genes and occupied distinct genomic locations in the two ichneumonid wasps. In contrast, viral machinery genes were organized in clusters showing highly conserved gene content and order, with some loci located in collinear wasp genomic regions. This genomic architecture clearly differs from the organization of PDVs in braconid wasps, in which proviral segments are clustered and viral machinery elements are more dispersed.ConclusionsThe contrasting structures of the two types of ichnovirus genomic elements are consistent with their different functions: proviral segments are vehicles for virulence proteins expected to adapt according to different host defense systems, whereas the genes involved in virus particle production in the wasp are likely more stable and may reflect ancestral viral architecture. The distinct genomic architectures seen in ichnoviruses versus bracoviruses reveal different evolutionary trajectories that have led to virus domestication in the two wasp lineages.

Recent advances in understanding mitochondrial genome diversity.

Ever since its discovery, the double-stranded DNA contained in the mitochondria of eukaryotes has fascinated researchers because of its bacterial endosymbiotic origin, crucial role in encoding subunits of the respiratory complexes, compact nature, and specific inheritance mechanisms. In the last few years, high-throughput sequencing techniques have accelerated the sequencing of mitochondrial genomes (mitogenomes) and uncovered the great diversity of organizations, gene contents, and modes of replication and transcription found in living eukaryotes. Some early divergent lineages of unicellular eukaryotes retain certain synteny and gene content resembling those observed in the genomes of alphaproteobacteria (the inferred closest living group of mitochondria), whereas others adapted to anaerobic environments have drastically reduced or even lost the mitogenome. In the three main multicellular lineages of eukaryotes, mitogenomes have pursued diverse evolutionary trajectories in which different types of molecules (circular versus linear and single versus multipartite), gene structures (with or without self-splicing introns), gene contents, gene orders, genetic codes, and transfer RNA editing mechanisms have been selected. Whereas animals have evolved a rather compact mitochondrial genome between 11 and 50 Kb in length with a highly conserved gene content in bilaterians, plants exhibit large mitochondrial genomes of 66 Kb to 11.3 Mb with large intergenic repetitions prone to recombination, and fungal mitogenomes have intermediate sizes of 12 to 236 Kb.

The Jekyll and Hyde Symbiont: Could Wolbachia Be a Nutritional Mutualist?

The most common intracellular symbiont on the planet-Wolbachia pipientis-is infamous largely for the reproductive manipulations induced in its host. However, more recent evidence suggests that this bacterium may also serve as a nutritional mutualist in certain host backgrounds and for certain metabolites. We performed a large-scale analysis of conserved gene content across all sequenced Wolbachia genomes to infer potential nutrients made by these symbionts. We review and critically evaluate the prior research supporting a beneficial role for Wolbachia and suggest future experiments to test hypotheses of metabolic provisioning.

Clustered Core- and Pan-Genome Content on Rhodobacteraceae Chromosomes.

In Bacteria, chromosome replication starts at a single origin of replication and proceeds on both replichores. Due to its asymmetric nature, replication influences chromosome structure and gene organization, mutation rate, and expression. To date, little is known about the distribution of highly conserved genes over the bacterial chromosome. Here, we used a set of 101 fully sequenced Rhodobacteraceae representatives to analyze the relationship between conservation of genes within this family and their distance from the origin of replication. Twenty-two of the analyzed species had core genes clustered significantly closer to the origin of replication with representatives of the genus Celeribacter being the most apparent example. Interestingly, there were also eight species with the opposite organization. In particular, Rhodobaca barguzinensis and Loktanella vestfoldensis showed a significant increase of core genes with distance from the origin of replication. The uneven distribution of low-conserved regions is in particular pronounced for genomes in which the halves of one replichore differ in their conserved gene content. Phage integration and horizontal gene transfer partially explain the scattered nature of Rhodobacteraceae genomes. Our findings lay the foundation for a better understanding of bacterial genome evolution and the role of replication therein.

Evolutionary dynamics of the chromatophore genome in three photosynthetic Paulinella species

The thecate amoeba Paulinella is a valuable model for understanding plastid organellogenesis because this lineage has independently gained plastids (termed chromatophores) of alpha-cyanobacterial provenance. Plastid primary endosymbiosis in Paulinella occurred relatively recently (90–140 million years ago, Mya), whereas the origin of the canonical Archaeplastida plastid occurred >1,500 Mya. Therefore, these two events provide independent perspectives on plastid formation on vastly different timescales. Here we generated the complete chromatophore genome sequence from P. longichromatophora (979,356 bp, GC-content = 38.8%, 915 predicted genes) and P. micropora NZ27 (977,190 bp, GC-content = 39.9%, 911 predicted genes) and compared these data to that from existing chromatophore genomes. Our analysis suggests that when a basal split occurred among photosynthetic Paulinella species ca. 60 Mya, only 35% of the ancestral orthologous gene families from the cyanobacterial endosymbiont remained in chromatophore DNA. Following major gene losses during the early stages of endosymbiosis, this process slowed down significantly, resulting in a conserved gene content across extant taxa. Chromatophore genes faced relaxed selection when compared to homologs in free-living alpha-cyanobacteria, likely reflecting the homogeneous intracellular environment of the Paulinella host. Comparison of nucleotide substitution and insertion/deletion events among different P. micropora strains demonstrates that increases in AT-content and genome reduction are ongoing and dynamic processes in chromatophore evolution.

The molecular characterization of fixed inversions breakpoints unveils the ancestral character of the Drosophila guanche chromosomal arrangements

Cytological studies revealed that the number of chromosomes and their organization varies across species. The increasing availability of whole genome sequences of multiple species across specific phylogenies has confirmed and greatly extended these cytological observations. In the Drosophila genus, the ancestral karyotype consists of five rod-like acrocentric chromosomes (Muller elements A to E) and one dot-like chromosome (element F), each exhibiting a generally conserved gene content. Chromosomal fusions and paracentric inversions are thus the major contributors, respectively, to chromosome number variation among species and to gene order variation within chromosomal element. The subobscura cluster of Drosophila consists in three species that retain the genus ancestral karyotype and differ by a reduced number of fixed inversions. Here, we have used cytological information and the D. guanche genome sequence to identify and molecularly characterize the breakpoints of inversions that became fixed since the D. guanche-D. subobscura split. Our results have led us to propose a modified version of the D. guanche cytological map of its X chromosome, and to establish that (i) most inversions became fixed in the D. subobscura lineage and (ii) the order in which the four X chromosome overlapping inversions occurred and became fixed.

Functional and evolutionary genomic inferences in Populus through genome and population sequencing of American and European aspen

The Populus genus is one of the major plant model systems, but genomic resources have thus far primarily been available for poplar species, and primarily Populus trichocarpa (Torr. & Gray), which was the first tree with a whole-genome assembly. To further advance evolutionary and functional genomic analyses in Populus, we produced genome assemblies and population genetics resources of two aspen species, Populus tremula L. and Populus tremuloides Michx. The two aspen species have distributions spanning the Northern Hemisphere, where they are keystone species supporting a wide variety of dependent communities and produce a diverse array of secondary metabolites. Our analyses show that the two aspens share a similar genome structure and a highly conserved gene content with P. trichocarpa but display substantially higher levels of heterozygosity. Based on population resequencing data, we observed widespread positive and negative selection acting on both coding and noncoding regions. Furthermore, patterns of genetic diversity and molecular evolution in aspen are influenced by a number of features, such as expression level, coexpression network connectivity, and regulatory variation. To maximize the community utility of these resources, we have integrated all presented data within the PopGenIE web resource (PopGenIE.org).

Mitochondrial and Plastid Genomes from Coralline Red Algae Provide Insights into the Incongruent Evolutionary Histories of Organelles.

Mitochondria and plastids are generally uniparentally inherited and have a conserved gene content over hundreds of millions of years, which makes them potentially useful phylogenetic markers. Organelle single gene-based trees have long been the basis for elucidating interspecies relationships that inform taxonomy. More recently, high-throughput genome sequencing has enabled the construction of massive organelle genome databases from diverse eukaryotes, and these have been used to infer species relationships in deep evolutionary time. Here, we test the idea that despite their expected utility, conflicting phylogenetic signal may exist in mitochondrial and plastid genomes from the anciently diverged coralline red algae (Rhodophyta). We generated complete organelle genome data from five coralline red algae (Lithothamnion sp., Neogoniolithon spectabile, Renouxia sp., Rhodogorgon sp., and Synarthrophyton chejuensis) for comparative analysis with existing organelle genome data from two other species (Calliarthron tuberculosum and Sporolithon durum). We find strong evidence for incongruent phylogenetic signal from both organelle genomes that may be explained by incomplete lineage sorting that has maintained anciently derived gene copies or other molecular evolutionary processes such as hybridization or gene flow during the evolutionary history of coralline red algae.

The first plastid genome of a filamentous taxon \u2018Bangia\u2019 sp. OUCPT-01 in the Bangiales

Red algae are important primary photosynthetic organisms. The Bangiales comprise a morphologically diverse order of red algae. Until now, complete plastid genomes of the Bangiales were only mapped for foliose species. To date, no filamentous plastomes have been published. The aim of this study was to determine and analyze the complete plastid genome of the filamentous marine species ‘Bangia’ sp. OUCPT-01. It is a circular molecule, 196,913 bps in length with a guanine-cytosine (GC) content of 33.5%. It has a quadripartite structure with two single copy regions separated by two direct non-identical repeats. It has 205 protein-coding genes, 37 tRNAs, and 6 rRNAs. Therefore, it has a high coding capacity and is highly similar to other Bangiales species in terms of content and structure. In particular, it reveals that the genera in the Bangiales have highly conserved gene content and plastome synteny. This plastome and existing data provide insights into the phylogenetic relationships among the Bangiales genera of the Rhodophyta. According to its plastid- and mitochondrial genomes, ‘Bangia 2′ is a sister group to Porphyra. However, the position of Wildemania schizophylla in the Bangiales is still controversial. Our results show that the Bangiales divergence time was ~225 million years ago.

The PL6-Family Plasmids of Haloquadratum Are Virus-Related.

Plasmids PL6A and PL6B are both carried by the C23T strain of the square archaeon Haloquadratum walsbyi, and are closely related (76% nucleotide identity), circular, about 6 kb in size, and display the same gene synteny. They are unrelated to other known plasmids and all of the predicted proteins are cryptic in function. Here we describe two additional PL6-related plasmids, pBAJ9-6 and pLT53-7, each carried by distinct isolates of Haloquadratum walsbyi that were recovered from hypersaline waters in Australia. A third PL6-like plasmid, pLTMV-6, was assembled from metavirome data from Lake Tyrell, a salt-lake in Victoria, Australia. Comparison of all five plasmids revealed a distinct plasmid family with strong conservation of gene content and synteny, an average size of 6.2 kb (range 5.8–7.0 kb) and pairwise similarities between 61–79%. One protein (F3) was closely similar to a protein carried by betapleolipoviruses while another (R6) was similar to a predicted AAA-ATPase of His 1 halovirus (His1V_gp16). Plasmid pLT53-7 carried a gene for a FkbM family methyltransferase that was not present in any of the other plasmids. Comparative analysis of all PL6-like plasmids provided better resolution of conserved sequences and coding regions, confirmed the strong link to haloviruses, and showed that their sequences are highly conserved among examples from Haloquadratum isolates and metagenomic data that collectively cover geographically distant locations, indicating that these genetic elements are widespread.

Whole-Genome Analysis Illustrates Global Clonal Population Structure of the Ubiquitous Dermatophyte Pathogen Trichophyton rubrum.

Dermatophytes include fungal species that infect humans, as well as those that also infect other animals or only grow in the environment. The dermatophyte species Trichophyton rubrum is a frequent cause of skin infection in immunocompetent individuals. While members of the T. rubrum species complex have been further categorized based on various morphologies, their population structure and ability to undergo sexual reproduction are not well understood. In this study, we analyze a large set of T. rubrum and T. interdigitale isolates to examine mating types, evidence of mating, and genetic variation. We find that nearly all isolates of T. rubrum are of a single mating type, and that incubation with T. rubrum “morphotype” megninii isolates of the other mating type failed to induce sexual development. While the region around the mating type locus is characterized by a higher frequency of SNPs compared to other genomic regions, we find that the population is remarkably clonal, with highly conserved gene content, low levels of variation, and little evidence of recombination. These results support a model of recent transition to asexual growth when this species specialized to growth on human hosts.

Plastid Genome Evolution in the Early-Diverging Legume Subfamily Cercidoideae (Fabaceae).

The subfamily Cercidoideae is an early-branching legume lineage, which consists of 13 genera distributed in the tropical and warm temperate Northern Hemisphere. A previous study detected two plastid genomic variations in this subfamily, but the limited taxon sampling left the overall plastid genome (plastome) diversification across the subfamily unaddressed, and phylogenetic relationships within this clade remained unresolved. Here, we assembled eight plastomes from seven Cercidoideae genera and conducted phylogenomic-comparative analyses in a broad evolutionary framework across legumes. The plastomes of Cercidoideae all exhibited a typical quadripartite structure with a conserved gene content typical of most angiosperm plastomes. Plastome size ranged from 151,705 to 165,416 bp, mainly due to the expansion and contraction of inverted repeat (IR) regions. The order of genes varied due to the occurrence of several inversions. In Tylosema species, a plastome with a 29-bp IR-mediated inversion was found to coexist with a canonical-type plastome, and the abundance of the two arrangements of isomeric molecules differed between individuals. Complete plastome data were much more efficient at resolving intergeneric relationships of Cercidoideae than the previously used selection of only a few plastid or nuclear loci. In sum, our study revealed novel insights into the structural diversification of plastomes in an early-branching legume lineage, and, thus, into the evolutionary trajectories of legume plastomes in general.

Multiple losses of photosynthesis and convergent reductive genome evolution in the colourless green algae Prototheca

Autotrophic eukaryotes have evolved by the endosymbiotic uptake of photosynthetic organisms. Interestingly, many algae and plants have secondarily lost the photosynthetic activity despite its great advantages. Prototheca and Helicosporidium are non-photosynthetic green algae possessing colourless plastids. The plastid genomes of Prototheca wickerhamii and Helicosporidium sp. are highly reduced owing to the elimination of genes related to photosynthesis. To gain further insight into the reductive genome evolution during the shift from a photosynthetic to a heterotrophic lifestyle, we sequenced the plastid and nuclear genomes of two Prototheca species, P. cutis JCM 15793 and P. stagnora JCM 9641, and performed comparative genome analyses among trebouxiophytes. Our phylogenetic analyses using plastid- and nucleus-encoded proteins strongly suggest that independent losses of photosynthesis have occurred at least three times in the clade of Prototheca and Helicosporidium. Conserved gene content among these non-photosynthetic lineages suggests that the plastid and nuclear genomes have convergently eliminated a similar set of photosynthesis-related genes. Other than the photosynthetic genes, significant gene loss and gain were not observed in Prototheca compared to its closest photosynthetic relative Auxenochlorella. Although it remains unclear why loss of photosynthesis occurred in Prototheca, the mixotrophic capability of trebouxiophytes likely made it possible to eliminate photosynthesis.

Comparative analysis of inverted repeats of polypod fern (Polypodiales) plastomes reveals two hypervariable regions

BackgroundFerns are large and underexplored group of vascular plants (~ 11 thousands species). The genomic data available by now include low coverage nuclear genomes sequences and partial sequences of mitochondrial genomes for six species and several plastid genomes.ResultsWe characterized plastid genomes of three species of Dryopteris, which is one of the largest fern genera, using sequencing of chloroplast DNA enriched samples and performed comparative analysis with available plastomes of Polypodiales, the most species-rich group of ferns. We also sequenced the plastome of Adianthum hispidulum (Pteridaceae). Unexpectedly, we found high variability in the IR region, including duplication of rrn16 in D. blanfordii, complete loss of trnI-GAU in D. filix-mas, its pseudogenization due to the loss of an exon in D. blanfordii. Analysis of previously reported plastomes of Polypodiales demonstrated that Woodwardia unigemmata and Lepisorus clathratus have unusual insertions in the IR region. The sequence of these inserted regions has high similarity to several LSC fragments of ferns outside of Polypodiales and to spacer between tRNA-CGA and tRNA-TTT genes of mitochondrial genome of Asplenium nidus. We suggest that this reflects the ancient DNA transfer from mitochondrial to plastid genome occurred in a common ancestor of ferns. We determined the marked conservation of gene content and relative evolution rate of genes and intergenic spacers in the IRs of Polypodiales. Faster evolution of the four intergenic regions had been demonstrated (trnA- orf42, rrn16-rps12, rps7-psbA and ycf2-trnN).ConclusionsIRs of Polypodiales plastomes are dynamic, driven by such events as gene loss, duplication and putative lateral transfer from mitochondria.

Polymorphic duplicate genes and persistent non-coding sequences reveal heterogeneous patterns of mitochondrial DNA loss in salamanders

BackgroundMitochondria are the site of the citric acid cycle and oxidative phosphorylation (OXPHOS). In metazoans, the mitochondrial genome is a small, circular molecule averaging 16.5 kb in length. Despite evolutionarily conserved gene content, metazoan mitochondrial genomes show a diversity of gene orders most commonly explained by the duplication-random loss (DRL) model. In the DRL model, (1) a sequence of genes is duplicated in tandem, (2) one paralog sustains a loss-of-function mutation, resulting in selection to retain the other copy, and (3) the non-functional paralog is eventually deleted from the genome. Despite its apparent role in generating mitochondrial gene order diversity, little is known about the tempo and mode of random gene loss after duplication events. Here, we determine mitochondrial gene order across the salamander genus Aneides, which was previously shown to include at least two DRL-mediated rearrangement events. We then analyze these gene orders in a phylogenetic context to reveal patterns of DNA loss after mitochondrial gene duplication.ResultsWe identified two separate duplication events that resulted in mitochondrial gene rearrangements in Aneides; one occurred at the base of the clade tens of millions of years ago, while the other occurred much more recently (i.e. within a single species), resulting in gene order polymorphism and paralogs that are readily identifiable. We demonstrate that near-complete removal of duplicate rRNA genes has occurred since the recent duplication event, whereas duplicate protein-coding genes persist as pseudogenes and duplicate tRNAs persist as functionally intact paralogs. In addition, we show that non-coding DNA duplicated at the base of the clade has persisted across species for tens of millions of years.ConclusionsThe evolutionary history of the mitochondrial genome, from its inception as a bacterial endosymbiont, includes massive genomic reduction. Consistent with this overall trend, selection for efficiency of mitochondrial replication and transcription has been hypothesized to favor elimination of extra sequence. Our results, however, suggest that there may be no strong disadvantage to extraneous sequences in salamander mitochondrial genomes, although duplicate rRNA genes may be deleterious.