Sequential Bayesian Phylogenetic Inference.

The ideal approach to Bayesian phylogenetic inference is to estimate all parameters of interest jointly in a single hierarchical model. However, this is often not feasible in practice due to the high computational cost. Instead, phylogenetic pipelines generally consist of sequential analyses, whereby a single point estimate from a given analysis is used as input for the next analysis (e.g., a single multiple sequence alignment is used to estimate a gene tree). In this framework, uncertainty is not propagated from step to step, which can lead to inaccurate or spuriously confident results. Here, we formally develop and test a sequential inference approach for Bayesian phylogenetic inference, which uses importance sampling to generate observations for the next step of an analysis pipeline from the posterior distribution produced in the previous step. Our sequential inference approach presented here not only accounts for uncertainty between analysis steps, but also allows for greater flexibility in software choice (and hence model availability) and can be computationally more efficient than the traditional joint inference approach when multiple models are being tested. We show that our sequential inference approach is identical in practice to the joint inference approach only if sufficient information in the data is present (a narrow posterior distribution) and/or sufficiently many importance samples are used. Conversely, we show that the common practice of using a single point estimate can be biased, e.g., a single phylogeny estimate to transform an unrooted phylogeny into a time-calibrated phylogeny. We demonstrate the theory of sequential Bayesian inference using both a toy example and an empirical case study of divergence-time estimation in insects using a relaxed clock model from transcriptome data. In the empirical example, we estimate three posterior distributions of branch lengths from the same data (DNA character matrix with a GTR+Γ+I substitution model, an amino acid data matrix with empirical substitution models, and an amino acid data matrix with the PhyloBayes CAT-GTR model). Finally, we apply three different node-calibration strategies and show that divergence-time estimates are affected by both the data source and underlying substitution process to estimate branch lengths as well as the node-calibration strategies. Thus, our new sequential Bayesian phylogenetic inference provides the opportunity to efficiently test different approaches for divergence time estimation, including branch-length estimation from other software.

Phylogenomics, Lineage Diversification Rates, and the Evolution of Diadromy in Clupeiformes (Anchovies, Herrings, Sardines, and Relatives).

Migration independently evolved numerous times in animals, with a myriad of ecological and evolutionary implications. In fishes, perhaps the most extreme form of migration is diadromy, the migration between marine and freshwater environments. Key and longstanding questions are: how many times has diadromy evolved in fishes, how frequently do diadromous clades give rise to non-diadromous species, and does diadromy influence lineage diversification rates? Many diadromous fishes have large geographic ranges with constituent populations that use isolated freshwater habitats. This may limit gene flow among some populations, increasing the likelihood of speciation in diadromous lineages relative to non-diadromous lineages. Alternatively, diadromy may reduce lineage diversification rates if migration is associated with enhanced dispersal capacity that facilitates gene flow within and between populations. Clupeiformes (herrings, sardines, shads and anchovies) is a model clade for testing hypotheses about the evolution of diadromy because it includes an exceptionally high proportion of diadromous species and several independent evolutionary origins of diadromy. However, relationships among major clupeiform lineages remain unresolved and existing phylogenies sparsely sampled diadromous species, limiting the resolution of phylogenetically-informed statistical analyses. We assembled a phylogenomic dataset and used multi-species coalescent and concatenation-based approaches to generate the most comprehensive, highly-resolved clupeiform phylogeny to date, clarifying associations among several major clades and identifying recalcitrant relationships needing further examination. We determined that variation in rates of sequence evolution (heterotachy) and base-composition (non-stationarity) had little impact on our results. Using this phylogeny, we characterized evolutionary patterns of diadromy and tested for differences in lineage diversification rates between diadromous, marine, and freshwater lineages. We identified thirteen transitions to diadromy, all during the Cenozoic Era (ten origins of anadromy, two origins of catadromy, and one origin of amphidromy), and seven losses of diadromy. Two diadromous lineages rapidly generated non-diadromous species, demonstrating that diadromy is not an evolutionary dead-end. We discovered considerably faster transition rates out of diadromy than to diadromy. The largest lineage diversification rate increase in Clupeiformes was associated with a transition to diadromy, but we uncovered little statistical support for categorically faster lineage diversification rates in diadromous versus non-diadromous fishes. We propose that diadromy may increase the potential for accelerated lineage diversification, particularly in species that migrate long distances. However, this potential may only be realized in certain biogeographic contexts, such as when diadromy allows access to ecosystems in which there is limited competition from incumbent species.

Benefits and Limits of Phasing Alleles for Network Inference of Allopolyploid Complexes.

Accurately reconstructing the reticulate histories of polyploids remains a central challenge for understanding plant evolution. Although phylogenetic networks can provide insights into relationships among polyploid lineages, inferring networks may be hindered by the complexities of homology determination in polyploid taxa. We use simulations to show that phasing alleles from allopolyploid individuals can improve phylogenetic network inference under the multispecies coalescent by obtaining the true network with fewer loci compared to haplotype consensus sequences or sequences with heterozygous bases represented as ambiguity codes. Phased allelic data can also improve divergence time estimates for networks, which is helpful for evaluating allopolyploid speciation hypotheses and proposing mechanisms of speciation. To achieve these outcomes in empirical data, we present a novel pipeline that leverages a recently developed phasing algorithm to reliably phase alleles from polyploids. This pipeline is especially appropriate for target enrichment data, where depth of coverage is typically high enough to phase entire loci. We provide an empirical example in the North American Dryopteris fern complex that demonstrates insights from phased data as well as the challenges of network inference. We establish that our pipeline (PATÉ: Phased Alleles from Target Enrichment data) is capable of recovering a high proportion of phased loci from both diploids and polyploids. These data may improve network estimates compared to using haplotype consensus assemblies by accurately inferring the direction of gene flow, but statistical non-identifiability of phylogenetic networks poses a barrier to inferring the evolutionary history of reticulate complexes.

Phylogenomics resolves the higher-level phylogeny of herbivorous eriophyoid mites (Acariformes: Eriophyoidea).

BACKGROUND: Eriophyoid mites (Eriophyoidea) are among the largest groups in the Acariformes; they are strictly phytophagous. The higher-level phylogeny of eriophyoid mites, however, remains unresolved due to the limited number of available morphological characters-some of them are homoplastic. Nevertheless, the eriophyoid mites sequenced to date showed highly variable mitochondrial (mt) gene orders, which could potentially be useful for resolving the higher-level phylogenetic relationships. RESULTS: Here, we sequenced and compared the complete mt genomes of 153 eriophyoid mite species, which showed 54 patterns of rearranged mt gene orders relative to that of the hypothetical ancestor of arthropods. The shared derived mt gene clusters support the monophyly of eriophyoid mites (Eriophyoidea) as a whole and the monophylies of six clades within Eriophyoidea. These monophyletic groups and their relationships were largely supported in the phylogenetic trees inferred from mt genome sequences as well. Our molecular dating results showed that Eriophyoidea originated in the Triassic and diversified in the Cretaceous, coinciding with the diversification of angiosperms. CONCLUSIONS: This study reveals multiple molecular synapomorphies (i.e. shared derived mt gene clusters) at different levels (i.e. family, subfamily or tribe level) from the complete mt genomes of 153 eriophyoid mite species. We demonstrated the use of derived mt gene clusters in unveiling the higher-level phylogeny of eriophyoid mites, and underlines the origin of these mites and their co-diversification with angiosperms.

The size diversity of the Pteridaceae family chloroplast genome is caused by overlong intergenic spacers.

BACKGROUND: While the size of chloroplast genomes (cpDNAs) is often influenced by the expansion and contraction of inverted repeat regions and the enrichment of repeats, it is the intergenic spacers (IGSs) that appear to play a pivotal role in determining the size of Pteridaceae cpDNAs. This provides an opportunity to delve into the evolution of chloroplast genomic structures of the Pteridaceae family. This study added five Pteridaceae species, comparing them with 36 published counterparts. RESULTS: Poor alignment in the non-coding regions of the Pteridaceae family was observed, and this was attributed to the widespread presence of overlong IGSs in Pteridaceae cpDNAs. These overlong IGSs were identified as a major factor influencing variations in cpDNA size. In comparison to non-expanded IGSs, overlong IGSs exhibited significantly higher GC content and were rich in repetitive sequences. Species divergence time estimations suggest that these overlong IGSs may have already existed during the early radiation of the Pteridaceae family. CONCLUSIONS: This study reveals new insights into the genetic variation, evolutionary history, and dynamic changes in the cpDNA structure of the Pteridaceae family, providing a fundamental resource for further exploring its evolutionary research.

Phylogenetic analysis and divergence time estimation of Lycium species in China based on the chloroplast genomes.

BACKGROUND: Lycium is an economically and ecologically important genus of shrubs, consisting of approximately 70 species distributed worldwide, 15 of which are located in China. Despite the economic and ecological importance of Lycium, its phylogeny, interspecific relationships, and evolutionary history remain relatively unknown. In this study, we constructed a phylogeny and estimated divergence time based on the chloroplast genomes (CPGs) of 15 species, including subspecies, of the genus Lycium from China. RESULTS: We sequenced and annotated 15 CPGs in this study. Comparative analysis of these genomes from these Lycium species revealed a typical quadripartite structure, with a total sequence length ranging from 154,890 to 155,677 base pairs (bp). The CPGs was highly conserved and moderately differentiated. Through annotation, we identified a total of 128-132 genes. Analysis of the boundaries of inverted repeat (IR) regions showed consistent positioning: the junctions of the IRb/LSC region were located in rps19 in all Lycium species, IRb/SSC between the ycf1 and ndhF genes, and SSC/IRa within the ycf1 gene. Sequence variation in the SSC region exceeded that in the IR region. We did not detect major expansions or contractions in the IR region or rearrangements or insertions in the CPGs of the 15 Lycium species. Comparative analyses revealed five hotspot regions in the CPG: trnR(UCU), atpF-atpH, ycf3-trnS(GGA), trnS(GGA), and trnL-UAG, which could potentially serve as molecular markers. In addition, phylogenetic tree construction based on the CPG indicated that the 15 Lycium species formed a monophyletic group and were divided into two typical subbranches and three minor branches. Molecular dating suggested that Lycium diverged from its sister genus approximately 17.7 million years ago (Mya) and species diversification within the Lycium species of China primarily occurred during the recent Pliocene epoch. CONCLUSION: The divergence time estimation presented in this study will facilitate future research on Lycium, aid in species differentiation, and facilitate diverse investigations into this economically and ecologically important genus.

Thirteen complete chloroplast genomes of the costaceae family: insights into genome structure, selective pressure and phylogenetic relationships.

BACKGROUND: Costaceae, commonly known as the spiral ginger family, consists of approximately 120 species distributed in the tropical regions of South America, Africa, and Southeast Asia, of which some species have important ornamental, medicinal and ecological values. Previous studies on the phylogenetic and taxonomic of Costaceae by using nuclear internal transcribed spacer (ITS) and chloroplast genome fragments data had low resolutions. Additionally, the structures, variations and molecular evolution of complete chloroplast genomes in Costaceae still remain unclear. Herein, a total of 13 complete chloroplast genomes of Costaceae including 8 newly sequenced and 5 from the NCBI GenBank database, representing all three distribution regions of this family, were comprehensively analyzed for comparative genomics and phylogenetic relationships. RESULT: The 13 complete chloroplast genomes of Costaceae possessed typical quadripartite structures with lengths from 166,360 to 168,966 bp, comprising a large single copy (LSC, 90,802 - 92,189 bp), a small single copy (SSC, 18,363 - 20,124 bp) and a pair of inverted repeats (IRs, 27,982 - 29,203 bp). These genomes coded 111 - 113 different genes, including 79 protein-coding genes, 4 rRNA genes and 28 - 30 tRNAs genes. The gene orders, gene contents, amino acid frequencies and codon usage within Costaceae were highly conservative, but several variations in intron loss, long repeats, simple sequence repeats (SSRs) and gene expansion on the IR/SC boundaries were also found among these 13 genomes. Comparative genomics within Costaceae identified five highly divergent regions including ndhF, ycf1-D2, ccsA-ndhD, rps15-ycf1-D2 and rpl16-exon2-rpl16-exon1. Five combined DNA regions (ycf1-D2 + ndhF, ccsA-ndhD + rps15-ycf1-D2, rps15-ycf1-D2 + rpl16-exon2-rpl16-exon1, ccsA-ndhD + rpl16-exon2-rpl16-exon1, and ccsA-ndhD + rps15-ycf1-D2 + rpl16-exon2-rpl16-exon1) could be used as potential markers for future phylogenetic analyses and species identification in Costaceae. Positive selection was found in eight protein-coding genes, including cemA, clpP, ndhA, ndhF, petB, psbD, rps12 and ycf1. Maximum likelihood and Bayesian phylogenetic trees using chloroplast genome sequences consistently revealed identical tree topologies with high supports between species of Costaceae. Three clades were divided within Costaceae, including the Asian clade, Costus clade and South American clade. Tapeinochilos was a sister of Hellenia, and Parahellenia was a sister to the cluster of Tapeinochilos + Hellenia with strong support in the Asian clade. The results of molecular dating showed that the crown age of Costaceae was about 30.5 Mya (95% HPD: 14.9 - 49.3 Mya), and then started to diverge into the Costus clade and Asian clade around 23.8 Mya (95% HPD: 10.1 - 41.5 Mya). The Asian clade diverged into Hellenia and Parahellenia at approximately 10.7 Mya (95% HPD: 3.5 - 25.1 Mya). CONCLUSION: The complete chloroplast genomes can resolve the phylogenetic relationships of Costaceae and provide new insights into genome structures, variations and evolution. The identified DNA divergent regions would be useful for species identification and phylogenetic inference in Costaceae.

De Novo Mutation Rates in Sticklebacks.

Mutation rate is a fundamental parameter in population genetics. Apart from being an important scaling parameter for demographic and phylogenetic inference, it allows one to understand at what rate new genetic diversity is generated and what the expected level of genetic diversity is in a population at equilibrium. However, except for well-established model organisms, accurate estimates of de novo mutation rates are available for a very limited number of organisms from the wild. We estimated mutation rates (µ) in two marine populations of the nine-spined stickleback (Pungitius pungitius) with the aid of several 2- and 3-generational family pedigrees, deep (>50×) whole-genome resequences and a high-quality reference genome. After stringent filtering, we discovered 308 germline mutations in 106 offspring translating to µ = 4.83 × 10-9 and µ = 4.29 × 10-9 per base per generation in the two populations, respectively. Up to 20% of the mutations were shared by full-sibs showing that the level of parental mosaicism was relatively high. Since the estimated µ was 3.1 times smaller than the commonly used substitution rate, recalibration with µ led to substantial increase in estimated divergence times between different stickleback species. Our estimates of the de novo mutation rate should provide a useful resource for research focused on fish population genetics and that of sticklebacks in particular.

Shrinkage-based Random Local Clocks with Scalable Inference.

Molecular clock models undergird modern methods of divergence-time estimation. Local clock models propose that the rate of molecular evolution is constant within phylogenetic subtrees. Current local clock inference procedures exhibit one or more weaknesses, namely they achieve limited scalability to trees with large numbers of taxa, impose model misspecification, or require a priori knowledge of the existence and location of clocks. To overcome these challenges, we present an autocorrelated, Bayesian model of heritable clock rate evolution that leverages heavy-tailed priors with mean zero to shrink increments of change between branch-specific clocks. We further develop an efficient Hamiltonian Monte Carlo sampler that exploits closed form gradient computations to scale our model to large trees. Inference under our shrinkage clock exhibits a speed-up compared to the popular random local clock when estimating branch-specific clock rates on a variety of simulated datasets. This speed-up increases with the size of the problem. We further show our shrinkage clock recovers known local clocks within a rodent and mammalian phylogeny. Finally, in a problem that once appeared computationally impractical, we investigate the heritable clock structure of various surface glycoproteins of influenza A virus in the absence of prior knowledge about clock placement. We implement our shrinkage clock and make it publicly available in the BEAST software package.

Phylogeny and diversification of genus Sanicula L. (Apiaceae): novel insights from plastid phylogenomic analyses.

BACKGROUND: The genus Sanicula L. is a unique perennial herb that holds important medicinal values. Although the previous studies on Sanicula provided us with a good research basis, its taxonomic system and interspecific relationships have not been satisfactorily resolved, especially for those endemic to China. Moreover, the evolutionary history of this genus also remains inadequately understood. The plastid genomes possessing highly conserved structure and limited evolutionary rate have proved to be an effective tool for studying plant phylogeny and evolution. RESULTS: In the current study, we newly sequenced and assembled fifteen Sanicula complete plastomes. Combined with two previously reported plastomes, we performed comprehensively plastid phylogenomics analyses to gain novel insights into the evolutionary history of this genus. The comparative results indicated that the seventeen plastomes exhibited a high degree of conservation and similarity in terms of their structure, size, GC content, gene order, IR borders, codon bias patterns and SSRs profiles. Such as all of them displayed a typical quadripartite structure, including a large single copy region (LSC: 85,074-86,197 bp), a small single copy region (SSC: 17,047-17,132 bp) separated by a pair of inverted repeat regions (IRs: 26,176-26,334 bp). And the seventeen plastomes had similar IR boundaries and the adjacent genes were identical. The rps19 gene was located at the junction of the LSC/IRa, the IRa/SSC junction region was located between the trnN gene and ndhF gene, the ycf1 gene appeared in the SSC/IRb junction and the IRb/LSC boundary was located between rpl12 gene and trnH gene. Twelve specific mutation hotspots (atpF, cemA, accD, rpl22, rbcL, matK, ycf1, trnH-psbA, ycf4-cemA, rbcL-accD, trnE-trnT and trnG-trnR) were identified that can serve as potential DNA barcodes for species identification within the genus Sanicula. Furthermore, the plastomes data and Internal Transcribed Spacer (ITS) sequences were performed to reconstruct the phylogeny of Sanicula. Although the tree topologies of them were incongruent, both provided strong evidence supporting the monophyly of Saniculoideae and Apioideae. In addition, the sister groups between Saniculoideae and Apioideae were strongly suggested. The Sanicula species involved in this study were clustered into a clade, and the Eryngium species were also clustered together. However, it was clearly observed that the sections of Sanicula involved in the current study were not respectively recovered as monophyletic group. Molecular dating analysis explored that the origin of this genus was occurred during the late Eocene period, approximately 37.84 Ma (95% HPD: 20.33-52.21 Ma) years ago and the diversification of the genus was occurred in early Miocene 18.38 Ma (95% HPD: 10.68-25.28 Ma). CONCLUSION: The plastome-based tree and ITS-based tree generated incongruences, which may be attributed to the event of hybridization/introgression, incomplete lineage sorting (ILS) and chloroplast capture. Our study highlighted the power of plastome data to significantly improve the phylogenetic supports and resolutions, and to efficiently explore the evolutionary history of this genus. Molecular dating analysis explored that the diversification of the genus occurred in the early Miocene, which was largely influenced by the prevalence of the East Asian monsoon and the uplift of the Hengduan Mountains (HDM). In summary, our study provides novel insights into the plastome evolution, phylogenetic relationships, taxonomic framework and evolution of genus Sanicula.

The evolution of ephemeral flora in Xinjiang, China: insights from plastid phylogenomic analyses of Brassicaceae.

BACKGROUND: The ephemeral flora of northern Xinjiang, China, plays an important role in the desert ecosystems. However, the evolutionary history of this flora remains unclear. To gain new insights into its origin and evolutionary dynamics, we comprehensively sampled ephemeral plants of Brassicaceae, one of the essential plant groups of the ephemeral flora. RESULTS: We reconstructed a phylogenetic tree using plastid genomes and estimated their divergence times. Our results indicate that ephemeral species began to colonize the arid areas in north Xinjiang during the Early Miocene and there was a greater dispersal of ephemeral species from the surrounding areas into the ephemeral community of north Xinjiang during the Middle and Late Miocene, in contrast to the Early Miocene or Pliocene periods. CONCLUSIONS: Our findings, together with previous studies, suggest that the ephemeral flora originated in the Early Miocene, and species assembly became rapid from the Middle Miocene onwards, possibly attributable to global climate changes and regional geological events.

Dynamics and timing of diversification events of ciliated eukaryotes from a large phylogenomic perspective.

Ciliophora, an exceptionally diverse lineage of unicellular eukaryotes, exhibits a remarkable range of species richness across classes in the ciliate Tree of Life. In this study, we have acquired transcriptome and genome data from 40 representative species in seven ciliate classes. Utilizing 247 genes and 105 taxa, we devised a comprehensive phylogenomic tree for Ciliophora, encompassing over 60 % of orders and constituting the most extensive dataset of ciliate species to date. We established a robust phylogenetic framework that encompasses ambiguous taxa and the major classes within the phylum. Our findings support the monophyly of each of two subphyla (Postciliodesmatophora and Intramacronucleata), along with three subclades (Protocruzia, CONTHREEP, and SAPML) nested within Intramacronucleata, and elucidate evolutionary positions among the major classes within the phylum. Drawing on the robust ciliate Tree of Life and three constraints, we estimated the radiation of Ciliophora around 1175 Ma during the middle of the Proterozoic Eon, and most of the ciliate classes diverged from their sister lineage during the latter half of this period. Additionally, based on the time-calibrated tree and species richness pattern, we investigated net diversification rates of Ciliophora and its classes. The global net diversification rate for Ciliophora was estimated at 0.004979 species/Ma. Heterogeneity in net diversification rates was evident at the class level, with faster rates observed in Oligohymenophorea and Spirotrichea than other classes within the subclades CONTHREEP and SAPML, respectively. Notably, our analysis suggests that variations in net diversification rates, rather than clade ages, appear to contribute to the differences in species richness in Ciliophora at the class level.

Scalable Bayesian Divergence Time Estimation With Ratio Transformations.

Divergence time estimation is crucial to provide temporal signals for dating biologically important events from species divergence to viral transmissions in space and time. With the advent of high-throughput sequencing, recent Bayesian phylogenetic studies have analyzed hundreds to thousands of sequences. Such large-scale analyses challenge divergence time reconstruction by requiring inference on highly correlated internal node heights that often become computationally infeasible. To overcome this limitation, we explore a ratio transformation that maps the original $N-1$ internal node heights into a space of one height parameter and $N-2$ ratio parameters. To make the analyses scalable, we develop a collection of linear-time algorithms to compute the gradient and Jacobian-associated terms of the log-likelihood with respect to these ratios. We then apply Hamiltonian Monte Carlo sampling with the ratio transform in a Bayesian framework to learn the divergence times in 4 pathogenic viruses (West Nile virus, rabies virus, Lassa virus, and Ebola virus) and the coralline red algae. Our method both resolves a mixing issue in the West Nile virus example and improves inference efficiency by at least 5-fold for the Lassa and rabies virus examples as well as for the algae example. Our method now also makes it computationally feasible to incorporate mixed-effects molecular clock models for the Ebola virus example, confirms the findings from the original study, and reveals clearer multimodal distributions of the divergence times of some clades of interest.

Phylogenetics and biogeography of the olive family (Oleaceae).

BACKGROUND AND AIMS: Progress in the systematic studies of the olive family (Oleaceae) during the last two decades provides the opportunity to update its backbone phylogeny and to investigate its historical biogeography. We additionally aimed to understand the factors underlying the disjunct distribution pattern between East Asia and both West Asia and Europe that is found more commonly in this family than in any other woody plant families. METHODS: Using a sampling of 298 species out of ca. 750, the largest in a phylogenetic study of Oleaceae thus far, and a set of 36 plastid and nuclear markers, we reconstructed and dated a new phylogenetic tree based on maximum likelihood and Bayesian methods and checked for any reticulation events. We also assessed the relative support of four competing hypotheses [Qinghai-Tibet Plateau uplift (QTP-only hypothesis), climatic fluctuations (Climate-only hypothesis), combined effects of QTP uplift and climate (QTP-Climate hypothesis), and no effects (Null hypothesis)] in explaining these disjunct distributions. KEY RESULTS: We recovered all tribes and subtribes within Oleaceae as monophyletic, but uncertainty in the position of tribe Forsythieae remains. Based on this dataset, no reticulation event was detected. Our biogeographic analyses support the QTP-Climate hypothesis as the likely main explanation for the East-West Eurasian disjunctions in Oleaceae. Our results also show an earlier origin of Oleaceae at ca. 86 Mya and the role of Tropical Asia as a main source of species dispersals. CONCLUSION: Our new family-wide and extensive phylogenetic tree highlights both the stable relationships within Oleaceae, including the polyphyly of the genus Chionanthus, and the need for further systematic studies within the family's largest and most under-sampled genera (Chionanthus and Jasminum). Increased sampling will also help to fine-tune biogeographic analyses across spatial scales and geological times.

Young evolutionary origins of dioecy in the genus Asparagus.

PREMISE: Dioecy (separate sexes) has independently evolved numerous times across the angiosperm phylogeny and is recently derived in many lineages. However, our understanding is limited regarding the evolutionary mechanisms that drive the origins of dioecy in plants. The recent and repeated evolution of dioecy across angiosperms offers an opportunity to make strong inferences about the ecological, developmental, and molecular factors influencing the evolution of dioecy, and thus sex chromosomes. The genus Asparagus (Asparagaceae) is an emerging model taxon for studying dioecy and sex chromosome evolution, yet estimates for the age and origin of dioecy in the genus are lacking. METHODS: We use plastome sequences and fossil time calibrations in phylogenetic analyses to investigate the age and origin of dioecy in the genus Asparagus. We also review the diversity of sexual systems present across the genus to address contradicting reports in the literature. RESULTS: We estimate that dioecy evolved once or twice approximately 2.78-3.78 million years ago in Asparagus, of which roughly 27% of the species are dioecious and the remaining are hermaphroditic with monoclinous flowers. CONCLUSIONS: Our findings support previous work implicating a young age and the possibility of two origins of dioecy in Asparagus, which appear to be associated with rapid radiations and range expansion out of Africa. Lastly, we speculate that paleoclimatic oscillations throughout northern Africa may have helped set the stage for the origin(s) of dioecy in Asparagus approximately 2.78-3.78 million years ago.

Challenge accepted: Evolutionary lineages versus taxonomic classification of North American shrub willows (Salix).

PREMISE: The huge diversity of Salix subgenus Chamaetia/Vetrix clade in North America and the lack of phylogenetic resolution within this clade has presented a difficult but fascinating challenge for taxonomists to resolve. Here we tested the existing taxonomic classification with molecular tools. METHODS: In this study, 132 samples representing 46 species from 22 described sections of shrub willows from the United States and Canada were analyzed and combined with 67 samples from Eurasia. The ploidy levels of the samples were determined using flow cytometry and nQuire. Sequences were produced using a RAD sequencing approach and subsequently analyzed with ipyrad, then used for phylogenetic reconstructions (RAxML, SplitsTree), dating analyses (BEAST, SNAPPER), and character evolution analyses of 14 selected morphological traits (Mesquite). RESULTS: The RAD sequencing approach allowed the production of a well-resolved phylogeny of shrub willows. The resulting tree showed an exclusively North American (NA) clade in sister position to a Eurasian clade, which included some North American endemics. The NA clade began to diversify in the Miocene. Polyploid species appeared in each observed clade. Character evolution analyses revealed that adaptive traits such as habit and adaxial nectaries evolved multiple times independently. CONCLUSIONS: The diversity in shrub willows was shaped by an evolutionary radiation in North America. Most species were monophyletic, but the existing sectional classification could not be supported by molecular data. Nevertheless, monophyletic lineages share several morphological characters, which might be useful in the revision of the taxonomic classification of shrub willows.

Characterization of the organellar genomes of Mazzaella laminarioides and Mazaella membranacea (Gigartinaceae, Rhodophyta).

Mazzaella, a genus with no genomic resources available, has extensive distribution in the cold waters of the Pacific, where they represent ecologically and economically important species. In this study, we aimed to sequence, assemble, and annotate the complete mitochondrial and chloroplast genomes from two Mazzaella spp. and characterize the intraspecific variation among them. We report for the first time seven whole organellar genomes (mitochondria: OR915856, OR947465, OR947466, OR947467, OR947468, OR947469, OR947470; chloroplast: OR881974, OR909680, OR909681, OR909682, OR909683, OR909684, OR909685) obtained through high-throughput sequencing for six M. laminarioides sampled from three Chilean regions and one M. membranacea. Sequenced Mazzaella mitogenomes have identical gene number, gene order, and genome structure. The same results were observed for assembled plastomes. A total of 52 genes were identified in mitogenomes, and a total of 235 genes were identified in plastomes. Although the M. membranacea plastome included a full-length pbsA gene, in all M. laminarioides samples, the pbsA gene was split in three open reading frames (ORFs). Within M. laminarioides, we observed important plastome lineage-specific variations, such as the pseudogenization of the two hypothetical protein-coding genes, ycf23 and ycf45. Nonsense mutations in the ycf23 and ycf45 genes were only detected in the northern lineage. These results are consistent with phylogenetic reconstructions and divergence time estimation using concatenated coding sequences that not only support the monophyly of M. laminarioides but also underscore that the three M. laminarioides lineages are in an advanced stage of divergence. These new results open the question of the existence of still undisclosed species in M. laminarioides.

Phylogeographic History of Endangered Hokuriku Salamander, Hynobius takedai (Amphibia: Caudata).

Knowledge of the phylogeographic history of organisms is valuable for understanding their evolutionary processes. To the best of our knowledge, the phylogeographic structure of Hokuriku salamander, Hynobius takedai, an endangered species, remains unclear. This study aimed to elucidate the phylogeographic history of H. takedai, which is expected to be strongly influenced by paleogeographic events. Phylogenetic analysis based on partial sequences of the mitochondrial DNA cytochrome b gene confirmed the genetic independence of H. takedai, and the divergence time with closely related species was estimated to be from the Late Pliocene to the Early Pleistocene. In the phylogenetic tree, two clades were identified within H. takedai, and their haplotypes were found in samples collected from the west and east of the distribution range. These intraspecific divergences were strongly influenced by geohistorical subdivisions of the current major distribution areas in the Middle Pleistocene. One clade was further subdivided and its formation may have been influenced by sea level changes in the Late Pleistocene.

Phylogeography of the freshwater crab Potamon persicum (Decapoda: Potamidae): an ancestral ring species?

The Zagros Mountains, characterized by complex topography and three large drainage systems, harbor the endemic freshwater crab Potamon persicum in Iran. Our study delves into the evolutionary history of P. persicum, utilizing two mitochondrial and one nuclear marker. We collected 214 specimens from 24 localities, identifying 21 haplotypes grouped into two major evolutionary lineages. Substantial differentiation exists between drainage systems and lineages. Historical demographic analysis revealed a significant decrease in population size during the late Holocene, accompanied by a recent population bottleneck. Species distribution modeling has revealed eastward shifts in suitable habitats between the last glacial maximum and the present day. Following the last glacial maximum, habitat fragmentation occurred, resulting in the establishment of small populations. These smaller populations are more vulnerable to climatic and geological events, thereby limiting gene flow and accelerating genetic differentiation within species. Historical biogeographic analysis traced the origin of P. persicum to the western Zagros Mountains, with major genetic divergence occurring during the Pleistocene. Our genetic analyses suggest that P. persicum may have shown a genetic pattern similar to a classical ring species before the Pleistocene. The Namak Lake sub-basin could have served as a contact zone where populations did not interbreed but were connected through gene flow in a geographic ring. Currently, genetic separation is evident between basins, indicating that P. persicum in the Zagros Mountains is not a contemporary ring species. Also, our biogeographical analysis estimated that range evolution may have been driven initially by dispersal, and only during the late Pleistocene by vicariance.

A global phylogeny of turtles reveals a burst of climate-associated diversification on continental margins.

Living turtles are characterized by extraordinarily low species diversity given their age. The clade's extensive fossil record indicates that climate and biogeography may have played important roles in determining their diversity. We investigated this hypothesis by collecting a molecular dataset for 591 individual turtles that, together, represent 80% of all turtle species, including representatives of all families and 98% of genera, and used it to jointly estimate phylogeny and divergence times. We found that the turtle tree is characterized by relatively constant diversification (speciation minus extinction) punctuated by a single threefold increase. We also found that this shift is temporally and geographically associated with newly emerged continental margins that appeared during the Eocene-Oligocene transition about 30 million years before present. In apparent contrast, the fossil record from this time period contains evidence for a major, but regional, extinction event. These seemingly discordant findings appear to be driven by a common global process: global cooling and drying at the time of the Eocene-Oligocene transition. This climatic shift led to aridification that drove extinctions in important fossil-bearing areas, while simultaneously exposing new continental margin habitat that subsequently allowed for a burst of speciation associated with these newly exploitable ecological opportunities.