Phylogenetic Divergence Times Research Articles

Phylogenetic and molecular dating analyses of Chinese endemic genus Dipelta (Caprifoliaceae) based on nuclear RAD‐Seq and chloroplast genome data

AbstractDipelta Maxim. (Caprifoliaceae) is a Tertiary relic genus endemic to China, which includes three extant species, Dipelta floribunda, Dipelta yunnanensis, and Dipelta elegans. Recent progress in the systematics and phylogeographics of Dipelta has greatly broadened our knowledge about its origin and evolution, however, conflicted phylogenetic relationships and divergence times have been reported and warrant further investigation. Here, we utilized chloroplast genomes and population‐level genomic data restriction site‐associated DNA‐single nucleotide polymorphisms (RAD‐SNPs) to evaluate the interspecific relationships, population genetic structure and demographic histories of this genus. Our results confirmed the sister relationship between D. elegans and the D. yunnanensis–D. floribunda group, but with cyto‐nuclear phylogenetic discordance observed in the latter. Coalescent simulations suggested that this discordance might be attributed to asymmetric “chloroplast capture” through introgressive hybridization between the two parapatric species. Our fossil‐calibrated plastid chronogram of Dipsacales and the coalescent modeling based on nuclear RAD‐SNPs simultaneously suggested that the three species of Dipelta diversified at the late Miocene, which may be related to the uplift of the eastern part of Qinghai–Tibet Plateau (QTP) and adjacent southwest China, and increasing Asian interior aridification since the late Miocene; while in the mid‐Pleistocene, the climatic transition and continuous uplift of the QTP, triggered allopatric speciation via geographical isolation for D. floribunda and D. yunnanensis regardless of bidirectional gene flow. Based on both plastid and nuclear genome‐scale data, our findings provide the most comprehensive and reliable phylogeny and evolutionary histories for Dipelta and enable further understanding of the origin and evolution of floristic endemisms of China.

Phylogeny, biogeography and ecological diversification of New Caledonian palms (Arecaceae).

The geographic origin and evolutionary mechanisms underpinning the rich and distinctive New Caledonian flora remain poorly understood. This is due to the complex geological past of the island and to the scarcity of well-resolved species-level phylogenies. Here, we infer phylogenetic relationships and divergence times of New Caledonian palms, which comprise 40 species. We use this framework to elucidate the biogeography of New Caledonian palm lineages, and to explore how extant species may have formed. A phylogenetic tree including 37 New Caledonian palm species and 77 relatives from tribe Areceae was inferred from 151 nuclear genes obtained by targeted sequencing. Fossil-calibrated divergence times were estimated, and ancestral ranges were inferred. Ancestral and extant ecological preferences in terms of elevation, precipitation and substrate were compared between New Caledonian sister species to explore their possible roles as drivers of speciation. New Caledonian palms form four well-supported clades, inside which relationships are well resolved. Our results support the current classification, but suggest that Veillonia and Campecarpus should be resurrected, and fail to clarify whether Rhopalostylidinae is sister to or nested in Basseliniinae. New Caledonian palm lineages derive from New Guinean and Australian ancestors, which reached the island through at least three independent dispersal events between the Eocene and Miocene. Palms then dispersed out of New Caledonia at least five times, mainly towards Pacific islands. Geographic and ecological transitions associated with speciation events differed across time and genera. Substrate transitions were more frequently associated with older than younger events. Neighbouring areas and a mosaic of local habitats shaped New Caledonia's palm flora, and the island played a significant role in generating palm diversity across the Pacific region. This new spatio-temporal framework will enable population-level ecological and genetic studies to further unpick the mechanisms underpinning New Caledonian palm endemism.

Comparative Analysis of Chloroplast Genomes of "Tiantai Wu-Yao" (Lindera aggregata) and Taxa of the Same Genus and Different Genera.

Lindera aggregata is a species of the Lauraceae family, which has important medicinal, economic and ornamental values. In this study, we sequenced, assembled and annotated the chloroplast genome of L. aggregata and reannotated and corrected eight unverified annotations in the same genus. The chloroplast genomes taxa from Lindera and from different genera of Lauraceae were compared and analyzed, and their phylogenetic relationship and divergence time were speculated. All the 36 chloroplast genomes had typical quadripartite structures that ranged from 150,749 to 154,736 bp in total length. These genomes encoded 111-112 unique genes, including 78-79 protein-coding genes, 29-30 tRNA and 4 rRNA. Furthermore, there were 78-97 SSRs loci in these genomes, in which mononucleotide repeats were the most abundant; there were 24-49 interspersed repeats, and forward repeat types were the most frequent. The codon bias patterns of all species tended to use codons ending with A or U. Five and six highly variable regions were identified within genus and between genera, respectively, and three common regions (ycf1, ndhF-rpl32 and rpl32-trnL) were identified, which can be used as important DNA markers for phylogeny and species identification. According to the evaluation of the Ka/Ks ratio, most of the genes were under purifying selection, and only 10 genes were under positive selection. Finally, through the construction of the evolutionary tree of 39 chloroplast genomes, the phylogenetic relationship of Lauraceae was clarified and the evolutionary relationship of Lindera was revealed. The species of genus Lindera experienced rapid adaptive radiation from Miocene to Pleistocene. The results provided valuable insights for the study of chloroplast genomes in the Lauraceae family, especially in the genus Lindera.

Oligo-barcode illuminates holocentric karyotype evolution in Rhynchospora (Cyperaceae).

Holocentric karyotypes are assumed to rapidly evolve through chromosome fusions and fissions due to the diffuse nature of their centromeres. Here, we took advantage of the recent availability of a chromosome-scale reference genome for Rhynchospora breviuscula, a model species of this holocentric genus, and developed the first set of oligo-based barcode probes for a holocentric plant. These probes were applied to 13 additional species of the genus, aiming to investigate the evolutionary dynamics driving the karyotype evolution in Rhynchospora. The two sets of probes were composed of 27,392 (green) and 23,968 (magenta) oligonucleotides (45-nt long), and generated 15 distinct FISH signals as a unique barcode pattern for the identification of all five chromosome pairs of the R. breviuscula karyotype. Oligo-FISH comparative analyzes revealed different types of rearrangements, such as fusions, fissions, putative inversions and translocations, as well as genomic duplications among the analyzed species. Two rounds of whole genome duplication (WGD) were demonstrated in R. pubera, but both analyzed accessions differed in the complex chain of events that gave rise to its large, structurally diploidized karyotypes with 2n = 10 or 12. Considering the phylogenetic relationships and divergence time of the species, the specificity and synteny of the probes were maintained up to species with a divergence time of ~25 My. However, karyotype divergence in more distant species hindered chromosome mapping and the inference of specific events. This barcoding system is a powerful tool to study chromosomal variations and genomic evolution in holocentric chromosomes of Rhynchospora species.

Highly divergent sympatric lineages of Leptotila verreauxi (Aves: Columbidae) suggest a secondary contact area in the Isthmus of Tehuantepec, Mexico

Due to a complex geological and biotic history, the Isthmus of Tehuantepec (IT), has been long recognized as a driver for the evolutionary divergence of numerous lowland and highland taxa. Widely distributed in the lowlands of the American continent, the White-Tipped Dove (Leptotila verreauxi) is a polytypic species with 13 recognized subspecies. Four of these have been recorded in Mexico, and the distribution of three abuts at the IT, suggesting a contact zone. To estimate phylogenetic patterns, divergence times and genetic differentiation, we examined two mtDNA (ND2 and COI) and one nDNA (β-fibint 7) markers. We also used correlative ecological niche models (ENM) to assess whether ecological differences across the IT may have acted as a biogeographical boundary. We estimated paleodistributions during the Middle Holocene, Last Glacial Maximum and Last Interglacial, to evaluate the influence of climate changes on the distribution and demographic changes. Our results showed genetically distinct lineages that diverged approximately 2.5 million years ago. Climatic and ecological factors may have played a dual role in promoting differentiation, but also in the formation of a secondary contact zone in the southern IT. Our ecological niche comparisons indicated that the ecological niche of sympatric lineages at the IT are not identical, suggesting niches divergence; in addition, environmental niche models across the region indicated no abrupt biogeographic barriers, but the presence of regions with low suitability. These results suggest that genetic differentiation originated by a vicariant event probably related to environmental factors, favored the evolution of different ecological niches. Also, the absence of a biogeographic barrier but the presence of less suitable areas in the contact regions, suggest that secondary contact zones may be also maintained by climatic factors for the eastern group, but also by biotic interactions for the western group.

Complete Chloroplast Genomes and the Phylogenetic Analysis of Three Native Species of Paeoniaceae from the Sino-Himalayan Flora Subkingdom.

Paeonia delavayi var. lutea, Paeonia delavayi var. angustiloba, and Paeonia ludlowii are Chinese endemics that belong to the Paeoniaceae family and have vital medicinal and ornamental value. It is often difficult to classify Paeoniaceae plants based on their morphological characteristics, and the limited genomic information has strongly hindered molecular evolution and phylogenetic studies of Paeoniaceae. In this study, we sequenced, assembled, and annotated the chloroplast genomes of P. delavayi var. lutea, P. delavayi var. angustiloba, and P. ludlowii. The chloroplast genomes of these strains were comparatively analyzed, and their phylogenetic relationships and divergence times were inferred. These three chloroplast genomes exhibited a typical quadripartite structure and were 152,687-152,759 bp in length. Each genome contains 126-132 genes, including 81-87 protein-coding genes, 37 transfer RNAs, and 8 ribosomal RNAs. In addition, the genomes had 61-64 SSRs, with mononucleotide repeats being the most abundant. The codon bias patterns of the three species tend to use codons ending in A/U. Six regions of high variability were identified (psbK-psbL, trnG-UCC, petN-psbM, psbC, rps8-rpl14, and ycf1) that can be used as DNA molecular markers for phylogenetic and taxonomic analysis. The Ka/Ks ratio indicates positive selection for the rps18 gene associated with self-replication. The phylogenetic analysis of 99 chloroplast genomes from Saxifragales clarified the phylogenetic relationships of Paeoniaceae and revealed that P. delavayi var. lutea, P. delavayi var. angustiloba, and P. ludlowii are monophyletic groups and sisters to P. delavayi. Divergence time estimation revealed two evolutionary divergences of Paeoniaceae species in the early Oligocene and Miocene. Afterward, they underwent rapid adaptive radiation from the Pliocene to the early Pleistocene when P. delavayi var. lutea, P. delavayi var. angustiloba, and P. ludlowii formed. The results of this study enrich the chloroplast genomic information of Paeoniaceae and reveal new insights into the phylogeny of Paeoniaceae.

Comparative transcriptomes of four Elm species provide insights into the genetic features and adaptive evolution of Ulmus spp.

Ulmus spp. have a wide distribution, strong adaptability, and high application value. However, due to the outbreak of Dutch elm disease and a lack of nuclear genome data, studies on Ulmus spp. are lacking. In this study, we obtained full-length transcriptomes from four elm species, with an average of 64,529 transcripts per species. Among these transcripts, an average of 6410 long noncoding RNAs were obtained per species. We performed SSR site detection and primer design and obtained an average of 20,650 SSR primer pairs per elm species. Fisher’s exact test revealed a large difference in the enrichment of functional genes shared between Ulmus parvifolia and three other elms, which represented 31 Gene Ontology terms, four Kyoto Encyclopedia of Genes and Genomes pathways, and two transcription factors. Phylogenetic evolution and divergence time analyses indicated that U. parvifolia diverged from the three other elms ca. 13.98 Mya; the divergence time of Ulmus macrocarpa, Ulmus davidiana, and Ulmus pumila was ca. 10.84–7.25 Mya. Furthermore, we identified 22 genes that were positively selected, and transcriptional profiling analysis showed that the expression levels of those genes showed marked changes under high temperature. These positively selected genes are widely involved in the ubiquitin system, nucleotide excision repair, protein modification, circadian rhythm regulation, light stimulus response, abiotic stimulus response, MAPK signaling, and plant pathogenicity interactions; they may play an important role in the adaptation of elms to environmental changes. This study provides a basis for the study of adaptive evolution, genetic diversity and functional gene development in Ulmus spp.

Molecular phylogenetics and biogeography reveal the origin of cherries (Prunus subg. Cerasus, Rosaceae)

Abstract Cherries (subg. Cerasus) belong to Prunus s.l. (Rosaceae) and are widely distributed in temperate and subtropical regions of the Northern Hemisphere. Given its ornamental and edible functions, it is a hugely preferred germplasm resource for human societies. However, the interspecific phylogenetic relationships, divergence times, and routes of this subgenus remain unclear. To reveal its enigmatic and intricate evolutionary history, we newly sequenced 12 subg. Cerasus species, reconstructed the plastome phylogeny with 24 previously published subg. Cerasus species. This study supports that subg. Cerasus split into five clades, among which P. mahaleb is the basal group of subg. Cerasus; P. campanulata is genetically distantly related to P. cerasoides and is not recommended for treatment as a variety of the latter; P. subhirtella is involved in the speciation of P. yedoensis. Molecular dating and ancestral area reconstructions infer that subg. Cerasus originated in the Eastern Mediterranean–Central Asia before 11.57 Mya (95% HPD = 5.34–19.1 Mya) and subsequently spread into the Qinghai-Tibetan Plateau (QTP) and adjacent regions. The first dispersal event of modern subg. Cerasus was facilitated by ‘out-of-QTP’, colonizing North America across the Bering Land Bridge and re-dispersing into Europe from the Late Miocene to Pliocene and forming the initial distribution pattern of extant subg. Cerasus in the Pliocene; The end of the Quaternary glacial period witnessed a second dispersal event, and core subg. Cerasus population diversity flourished radially in its separate refugia during the Late Pleistocene.

Phylogenetic relationships and historical biogeography of Otus species based on mitochondrial cytochrome b gene sequence

Abstract This study analyzed mitochondrial cytochrome b gene sequences to infer Otus species’ phylogenetic relationships and historical biogeography. The researchers used Maximum Parsimony and Maximum Likelihood methods to generate the phylogenetic trees. They constructed a Bayesian MCC Chronogram Tree to determine the Otus species’ age and speciation events. Paleoclimate and paleogeology maps were also used to correlate the branching out of Otus species with geological time periods and climatic conditions. The results showed five major clusters or clades for the core taxon group (Otus) and one for the sister group (Tyto). The Bayesian tree showed seven major clades for Otus, with the first speciation occurrence dating back to 21.54 mya during the Early Miocene epoch. Most of the core taxon species speciated during the Pliocene Period, with some exceptions. The paleoclimate data indicated that Otus clades branched out during the Icehouse climate state, with the first branching out occurring during the Coolhouse climatic state. The paleogeologic data revealed that the African and American (North and South) continents were already distinct and divided by the Atlantic Ocean during the first branching out of the Otus clade. The distribution areas of Otus species were also mapped using updated Wallace’s zoogeographic realms. Overall, this study provides a better understanding of the phylogenetic history of Otus species and their diversification in relation to geological and climatic events. However, the authors caution that all phylogenetic trees, evolutionary studies, and divergence time estimates should be considered hypotheses with significant uncertainty.

Evolutionary origins of the lampriform pelagic radiation

Abstract Ray-finned fishes, which compose nearly half of living vertebrate diversity, provide an excellent system for studying the evolution of novel body forms. Lampriformes is a species-poor lineage of acanthomorph ray-finned fishes that has evolved two very different and highly specialized body plans suited to life in pelagic oceanic habitats: the deep, round-bodied bathysomes and the ribbon-like taeniosomes. Here, we present a new phylogenetic hypothesis and divergence time estimates for lampriform fishes based on an updated morphological dataset and DNA sequences from nuclear genes for all but one of the living lampriform families and 55% of recognized extant genera. Our analyses resolve two major clades in Lampriformes: the Bathysomi and the Taeniosomi. A time calibrated phylogeny shows that the origin of living lampriforms coincides with the aftermath of the Cretaceous–Palaeogene extinction and that anatomically modern pelagic morphotypes evolved 10 Myr after the start of the Palaeogene.

Phylogeography of two codistributed terrestrial snails in the Maputaland–Pondoland–Albany biodiversity hotspot, South Africa

Abstract We investigated the evolutionary patterns of two forest-dwelling endemic terrestrial snails (Gittenedouardia spadicea and Maizania wahlbergi) in the Maputaland–Pondoland–Albany biodiversity hotspot in South Africa to examine the degree of phylogeographical congruence between the two species. Phylogenetic relationships and divergence time estimations within each species were inferred using the combined mitochondrial DNA (mtDNA) sequence dataset for the cytochrome c oxidase subunit I (COI) and large subunit ribosomal RNA (16S rRNA). In addition, the rapidly evolving COI datasets were used to infer intraspecific genetic structure and population differentiation within both species. Phylogeographical concordance factor (PCF) analysis was used to statistically estimate the degree of congruence between the two species at localities where both species were sampled. Phylogenetically, G. spadicea exhibited two clades that diverged during the Plio/Pleistocene, while M. wahlbergi formed a single shallow clade that showed Pleistocene divergence. The haplotype networks for the COI locus demonstrated evidence of geographical and genetic isolation within both species. PCF analysis demonstrated partial phylogeographical concordance, suggesting that the two species may have been affected by similar ancient climatic events. Traits other than poor dispersal capabilities and limited gene flow may also influence how a species responds to fluctuating climate change.

Genome-Wide Identification of Trachinotus ovatus Antimicrobial Peptides and Their Immune Response against Two Pathogen Challenges.

Golden pompano, Trachinotus ovatus, as a highly nutritious commercially valuable marine fish, has become one of the preferred species for many fish farmers due to its rapid growth, wide adaptability, and ease of feeding and management. However, with the expansion of aquaculture scale, bacterial and parasitic diseases have also become major threats to the golden pompano industry. This study, based on comparative genomics, shows the possibility of preferential evolution of freshwater fish over marine fish by analyzing the phylogenetic relationships and divergence times of 14 marine fish and freshwater fish. Furthermore, we identified antimicrobial peptide genes from 14 species at the genomic level and found that the number of putative antimicrobial peptides may be related to species evolution. Subsequently, we classified the 341 identified AMPs from golden pompano into 38 categories based on the classification provided by the APD3. Among them, TCP represented the highest proportion, accounting for 23.2% of the total, followed by scolopendin, lectin, chemokine, BPTI, and histone-derived peptides. At the same time, the distribution of AMPs in chromosomes varied with type, and covariance analysis showed the frequency of its repeat events. Enrichment analysis and PPI indicated that AMP was mainly concentrated in pathways associated with disease immunity. In addition, our transcriptomic data measured the expression of putative AMPs of golden pompano in 12 normal tissues, as well as in the liver, spleen, and kidney infected with Streptococcus agalactiae and skin infected with Cryptocaryon irritans. As the infection with S. agalactiae and C. irritans progressed, we observed tissue specificity in the number and types of responsive AMPs. Positive selection of AMP genes may participate in the immune response through the MAPK signaling pathway. The genome-wide identification of antimicrobial peptides in the golden pompano provided a complete database of potential AMPs that can contribute to further understanding the immune mechanisms in pathogens. AMPs were expected to replace traditional antibiotics and be developed into targeted drugs against specific bacterial and parasitic pathogens for more precise and effective treatment to improve aquaculture production.

Time‐calibrated phylogeny of Daphne (Thymelaeaceae): Pre‐mediterranean temporal origin of the sclerophyllous Daphne gnidium

AbstractThe sclerophyllous syndrome is characterized by well‐defined traits (evergreen coriaceous leaves, inconspicuous flowers, and fleshy fruits). It has been hypothesized that lineages displaying the sclerophyllous syndrome show lower speciation rates than non‐sclerophyllous lineages after the establishment of the mediterranean climate. Daphne gnidium displays sclerophyllous traits and some differentiation into three subspecies (gnidium, mauritanica, maritima), but the spatio‐temporal origin of this taxonomic group is unknown due to the lack of a time‐calibrated phylogeny of the whole genus. Here, we inferred phylogenetic relationships and divergence times of Daphne (32 species, 238 samples) and other genera of Thymelaeaceae (16 genera, 38 species, 34 samples) using the internal transcribed spacer (ITS), which revealed that the current circumscription of Daphne is profoundly polyphyletic because some species are nested within the genera Wikstroemia and Edgeworthia. In contrast, D. gnidium formed a well‐supported clade as recognized in taxonomic accounts (subgenus Spachia). We found morphological and phylogenetic support for Daphne mauritanica as a monophyletic lineage sister to D. gnidium. Divergence between D. gnidium and D. mauritanica appears to have predated the establishment of seasonally dry conditions, which supports a pre‐mediterranean temporal origin of the lineage. A phylogeographical analysis within D. gnidium based on 66 nrDNA (ITS) and 84 cpDNA (rps16, trnV) sequences agreed with the low differentiation of the species in the Pleistocene despite its large distribution range. Altogether, D. gnidium illustrates one more example of the sclerophyllous syndrome with no speciation after the onset of the mediterranean climate.

Phylogenetic analysis and temporal diversification of the tribe Alsineae (Caryophyllaceae) with the description of three new genera, Hesperostellaria, Reniostellaria and Torreyostellaria.

Alsineae are one of the most taxonomically difficult tribes in Caryophyllaceae and consist of over 500 species distributed in the northern temperate zone. Recent phylogenetic results have improved our understanding on the evolutionary relationships among Alsineae members. Nevertheless, there are still some unresolved taxonomic and phylogenetic problems at the generic level, and the evolutionary history of major clades within the tribe was unexplored to date. In this study, we carried out phylogenetic analyses and divergence time estimation of Alsineae using the nuclear ribosomal internal transcribed spacer (nrITS) and four plastid regions (matK, rbcL, rps16, trnL-F). The present analyses yielded a robustly supported phylogenetic hypothesis of the tribe. Our results showed that the monophyletic Alsineae are strongly supported to be the sister of Arenarieae, and the inter-generic relationships within Alsineae were mostly resolved with strong support. Both molecular phylogenetic and morphological evidence supported the Asian species Stellaria bistylata and the two North American species Pseudostellaria jamesiana and Stellaria americana all should be recognized as new monotypic genera respectively, and three new genera Reniostellaria, Torreyostellaria, and Hesperostellaria were thereby proposed here. Additionally, molecular and morphological evidence also supported the proposal of the new combination Schizotechium delavayi. Nineteen genera were accepted within Alsineae and a key to these genera was provided. Molecular dating analysis suggested that Alsineae splitted from its sister tribe at ca. 50.2 million-years ago (Ma) during the early Eocene and began to diverge at ca. 37.9 Ma during the late Eocene, and divergent events within Alsineae occurred mainly since the late Oligocene. Results from the present study provide insights into the historical assembly of herbaceous flora in northern temperate regions.

Phylogenomics, evolution of trophic traits and divergence times of hemiodontid fishes (Ostariophysi: Characiformes)

The South American characiform family Hemiodontidae comprises five genera and 34 species. The family lacks comprehensive phylogenetic hypotheses resolving its species relationships. The studies that addressed these questions exhibited a narrow taxon sampling or used single-locus markers. Herein we surveyed hundreds of ultraconserved elements (UCEs) loci to provide the first molecular phylogenetic hypothesis and divergence time estimates for hemiodontids encompassing all its genera and most species (27 of the 34 valid species). We also tracked the history of the protractile upper jaw in the genera Argonectes and Bivibranchia across the recovered phylogenies through ancestral state reconstruction. Our results corroborate the monophyly of Hemiodontidae and the genera Argonectes and Bivibranchia in all phylogenetic methods with maximum clade support. The genera Anodus and Hemiodus were not monophyletic because Anodus elongatus was sister to the monotypic Micromischodus instead of A. orinocensis, and H. immaculatus did not form a clade with its other congeners, but instead was sister to the clade including Anodus and Micromischodus. All remaining species of Hemiodus were placed together into a monophyletic group, where they were arranged into four major subclades. The relationship in the family is summarised as: (Bivibranchia, (Argonectes, ((H. immaculatus, (Anodus, Micromischodus)), Hemiodus clade))), in discordance with the morphological phylogeny that placed all genera monophyletic and resolved the family as: ((Anodus, Micromischodus), (Hemiodus, (Argonectes, Bivibranchia))). The origin of Hemiodontidae was estimated from the Late Cretaceous to the Middle Paleogene, with the mean age in the Paleocene, while the origin of most hemiodontid genera except Bivibranchia occurred in the Miocene. Unordered parsimony and likelihood reconstruction indicates that Argonectes and Bivibranchia developed their protractile upper jaw independently.

Adaptive evolution characteristics of mitochondrial genomes in genus Aparapotamon (Brachyura, Potamidae) of freshwater crabs

BackgroundAparapotamon, a freshwater crab genus endemic to China, includes 13 species. The distribution of Aparapotamon spans the first and second tiers of China's terrain ladder, showing great altitudinal differences. To study the molecular mechanisms of adaptive evolution in Aparapotamon, we performed evolutionary analyses, including morphological, geographical, and phylogenetic analyses and divergence time estimation. We sequenced the mitogenomes of Aparapotamon binchuanense and Aparapotamon huizeense for the first time and resequenced three other mitogenomes of Aparapotamon grahami and Aparapotamon gracilipedum. These sequences were combined with NCBI sequences to perform comparative mitogenome analysis of all 13 Aparapotamon species, revealing mitogenome arrangement and the characteristics of protein-coding and tRNA genes.ResultsA new species classification scheme of the genus Aparapotamon has been detected and verified by different aspects, including geographical, morphological, phylogenetics and comparative mitogenome analyses. Imprints from adaptive evolution were discovered in the mitochondrial genomes of group A, including the same codon loss at position 416 of the ND6 gene and the unique arrangement pattern of the tRNA-Ile gene. Multiple tRNA genes conserved or involved in adaptive evolution were detected. Two genes associated with altitudinal adaptation, ATP8 and ND6, which experienced positive selection, were identified for the first time in freshwater crabs.ConclusionsGeological movements of the Qinghai-Tibet Plateau and Hengduan Mountains likely strongly impacted the speciation and differentiation of the four Aparapotamon groups. After some group A species dispersed from the Hengduan Mountain Range, new evolutionary characteristics emerged in their mitochondrial genomes, facilitating adaptation to the low-altitude environment of China's second terrain tier. Ultimately, group A species spread to high latitudes along the upper reaches of the Yangtze River, showing faster evolutionary rates, higher species diversity and the widest distribution.

Mitochondrial genome analysis, phylogeny and divergence time evaluation of Strixaluco (Aves, Strigiformes, Strigidae).

Background Prior research has shown that the European peninsulas were the main sources of Strixaluco colonisation of Northern Europe during the late glacial period. However, the phylogenetic relationship and the divergence time between S.aluco from Leigong Mountain Nature Reserve, Guizhou Province, China and the Strigiformes from overseas remains unclear. The mitochondrial genome structure of birds is a covalent double-chain loop structure that is highly conserved and, thus, suitable for phylogenetic analysis. This study examined the phylogenetic relationship and divergence time of Strix using the whole mitochondrial genome of S.aluco. New information In this study, the complete mitochondrial genome of Strixaluco, with a total length of 18,632 bp, is reported for the first time. A total of 37 genes were found, including 22 tRNAs, two rRNAs, 13 protein-coding genes and two non-coding control regions. Certain species of Tytoninae were used as out-group and PhyloSuite software was applied to build the ML-tree and BI-tree of Strigiformes. Finally, the divergence time tree was constructed using BEAST 2.6.7 software and the age of Miosurniadiurna fossil-bearing sediments (6.0-9.5 Ma) was set as internal correction point. The common ancestor of Strix was confirmed to have diverged during the Pleistocene (2.58-0.01 Ma). The combined action of the dramatic uplift of the Qinling Mountains in the Middle Pleistocene and the climate oscillation of the Pleistocene caused Strix divergence between the northern and southern parts of mainland China. The isolation of glacial-interglacial rotation and glacier refuge was the main reason for the divergence of Strixuralensis and S.aluco from their common ancestor during this period. This study provides a reference for the evolutionary history of S.aluco.

Phylogeny and Evolutionary Timescale of Muscidae (Diptera: Calyptratae) Inferred from Mitochondrial Genomes.

House flies (Muscidae) comprise the most species-rich family of the muscoid grade with over 5000 described species worldwide, and they are abundant in various terrestrial and aquatic ecosystems. The high number of species, varied appearances, complex feeding habits, and wide distributions have hindered researchers from understanding their phylogeny and evolutionary history. Here, we newly sequenced fifteen mitochondrial genomes and reconstructed the phylogenetic relationships and divergence time among eight subfamilies of Muscidae (Diptera). The best phylogenetic tree, which was inferred by IQ-Tree, recovered the monophyly for seven out of eight subfamilies (except for Mydaeinae). Based on phylogenetic analyses and morphological characteristics, we prefer the subfamily status of Azeliinae and Reinwardtiinae, and separate Stomoxyinae from Muscinae. Genus Helina Robineau-Desvoidy, 1830 was synonymized with Phaonia Robineau-Desvoidy, 1830. The divergence time estimation indicated Muscidae originated at 51.59 Ma (early Eocene). Most subfamilies had originated around 41 Ma. We provided a mtgenomic viewpoint on the phylogenetic relationships and divergence time estimation of Muscidae.

Origin and diversification of pheretimoid megascolecid earthworms in the Japanese Archipelago as revealed by mitogenomic phylogenetics

Megascolecid earthworms of the pheretimoid group are dominant detritivores of soil ecosystems in the Japanese Archipelago and East Asia. However, their diversity and phylogenetic relationships are poorly understood. We assembled whole mitogenome sequences for 197 megascolecid earthworms collected throughout Japan to study the phylogenetic relationships, phylogeography, divergence times, and diversification of important morphological characteristics among pheretimoid earthworms. Using 197 mitogenome sequences and 24 published mitogenome sequences from the East Asian mainland (221 sequences in total), we constructed a maximum likelihood tree and found that the pheretimoid earthworms currently assigned to Amynthas, Metaphire, Duplodicodrilus, and Manus are involved in the most senior genus Amynthas; thus, Amynthas can be treated as the sole genus encompassing all of the above genera. Within the Amynthas group, we identified three major lineages that led to four groups of endemic species in Japan. These lineages originated from different lineages on the East Asian mainland and Taiwan Island, indicating multiple colonization events from the East Asian mainland by different ancestral lineages, possibly after the Miocene. We also assembled nuclear ribosomal DNA sequences encompassing the 18S to 28S rRNA genes. The nuclear gene tree showed major groups consistent with the mitogenome tree except for different (and not well-resolved) relationships among major clades. Our molecular data covered 115–158 native and 7 non-native Amynthas group species in Japan in terms of DNA-based species delimitation. Our findings provide a basis for understanding the evolutionary relationships among diversified megascolecid earthworms in the Amynthas group in Japan and adjacent regions.

Integrative taxonomy of the stalk‐eyed bug genus Chauliops (Heteroptera: Malcidae: Chauliopinae) reveals orogeny‐driven speciation

AbstractEconomically significant bean pests of the genus Chauliops are species rich in the areas surrounding the Qinghai–Tibet Plateau and provide an excellent system for speciation studies. Here, an integrative taxonomic approach, employing morphological analyses, population genetic methods, and multiple molecular species delimitation methods, was used to clarify the taxonomy of Chauliops in East and Southeast Asia. Four new species (Chauliops parahorizontalis Li & Bu, sp. nov., Chauliops albida Li & Bu, sp. nov., Chauliops bicoloripes Li & Bu, sp. nov., and Chauliops paraconica Li & Bu, sp. nov.) were described, which increases the number of Chauliops species in this area from six to 10; a key for Chauliops species is also provided. Phylogenetic analysis and divergence time estimation revealed that Chauliops was divided into four clades: Clade A (Chauliops bisontula + [Chauliops horizontalis + C. parahorizontalis sp. nov.]), Clade B (C. albida sp. nov. and C. bicoloripes sp. nov.), Clade C (Chauliops quaternaria and Chauliops zhengi), and Clade D (Chauliops fallax + [Chauliops conica + C. paraconica sp. nov.]). Two species diversification events of Chauliops estimated to have occurred 7–1 million years ago (Ma) and 25–13 Ma were detected. These speciation events were consistent with the two historical uplift events of the Qinghai–Tibet Plateau, suggesting that orogeny might have provided opportunities for the diversification of Chauliops species on the southeastern margin of the Qinghai–Tibet Plateau. Our findings show that population genetic analyses can be used to delimit related species and that orogeny is a key driver of species diversification on the southeastern margin of the Qinghai–Tibet Plateau.