Phylogenomics of Bivalvia Using Ultraconserved Elements (UCEs) Reveal New Topologies for Pteriomorphia and Imparidentia.

Despite significant advances in phylogenetics over the past decades, the deep relationships within Bivalvia (phylum Mollusca) remain inconclusive. Previous efforts based on morphology or several genes have failed to resolve many key nodes in the phylogeny of Bivalvia. Advances have been made recently using transcriptome data, but the phylogenetic relationships within Bivalvia historically lacked consensus, especially within Pteriomorphia and Imparidentia. Here, we inferred the relationships of key lineages within Bivalvia using matrices generated from specifically designed ultraconserved elements (UCEs) with 16 available genomic resources and 85 newly sequenced specimens from 55 families. Our new probes (Bivalve UCE 2k v.1) for target sequencing captured an average of 849 UCEs with 1085-bp in mean length from in vitro experiments. Our results introduced novel schemes from six major clades (Protobranchina, Pteriomorphia, Palaeoheterodonta, Archiheterodonta, Anomalodesmata and Imparidentia), though some inner nodes were poorly resolved, such as paraphyletic Heterodonta in some topologies potentially due to insufficient taxon sampling. The resolution increased when analyzing specific matrices for Pteriomorphia and Imparidentia. We recovered three Pteriomorphia topologies different from previously published trees, with the strongest support for ((Ostreida + (Arcida + Mytilida)) + (Pectinida + (Limida + Pectinida))). Limida were nested within Pectinida, warranting further studies. For Imparidentia, our results strongly supported the new hypothesis of (Galeommatida + (Adapedonta + Cardiida)), while the possible non-monophyly of Lucinida was inferred but poorly supported. Overall, our results provide important insights into the phylogeny of Bivalvia and show that target enrichment sequencing of UCEs can be broadly applied to study both deep and shallow phylogenetic relationships.

Rapid Evolution of Host Repertoire and Geographic Range in a Young and Diverse Genus of Montane Butterflies.

Evolutionary changes in geographic distribution and larval host plants may promote the rapid diversification of montane insects, but this scenario has been rarely investigated. We studied rapid radiation of the butterfly genus Colias, which has diversified in mountain ecosystems in Eurasia, Africa, and the Americas. Based on a dataset of 150 nuclear protein-coding genetic loci and mitochondrial genomes, we constructed a time-calibrated phylogenetic tree of Colias species with broad taxon sampling. We then inferred their ancestral geographic ranges, historical diversification rates, and the evolution of host use. We found that the most recent common ancestor of Colias was likely geographically widespread and originated ~3.5 Ma. The group subsequently diversified in different regions across the world, often in tandem with geographic expansion events. No aspect of elevation was found to have a direct effect on diversification. The genus underwent a burst of diversification soon after the divergence of the Neotropical lineage, followed by an exponential decline in diversification rate toward the present. The ancestral host repertoire included the legume genera Astragalus and Trifolium but later expanded to include a wide range of Fabaceae genera and plants in more distantly related families, punctuated with periods of host range expansion and contraction. We suggest that the widespread distribution of the ancestor of all extant Colias lineages set the stage for diversification by isolation of populations that locally adapted to the various different environments they encountered, including different host plants. In this scenario, elevation is not the main driver but might have accelerated diversification by isolating populations.

Patterns of speciation in a parapatric pair of Saturnia moths as revealed by target capture.

The focus of this study has been to understand the evolutionary relationships and taxonomy of a widely distributed parapatric species pair of wild silk moths in Europe: Saturnia pavonia and Saturnia pavoniella (Lepidoptera: Saturniidae). To address species delimitation in these parapatric taxa, target enrichment and mtDNA sequencing was employed alongside phylogenetic, admixture, introgression, and species delimitation analyses. The dataset included individuals from both species close to and farther away from the contact zone as well as two hybrids generated in the lab. Nuclear markers strongly supported both S. pavonia and S. pavoniella as two distinct species, with hybrids forming a sister group to S. pavoniella. However, the Maximum Likelihood (ML) tree generated from mtDNA sequencing data presented a different picture, showing both taxa to be phylogenetically intermixed. This inconsistency is likely attributable to mitonuclear discordance, which can arise from biological factors (e.g., introgressive hybridization and/or incomplete lineage sorting). Our analyses indicate that past introgressions have taken place, but that there is no evidence to suggest an ongoing admixture between the two species, demonstrating that the taxa have reached full postzygotic reproductive isolation and hence represent two distinct biological species. Finally, we discuss our results from an evolutionary point of view taking into consideration the past climatic oscillations that have likely shaped the present dynamics between the two species. Overall, our study demonstrates the effectiveness of the target enrichment approach in resolving shallow phylogenetic relationships under complex evolutionary circumstances and that this approach is useful in establishing robust and well-informed taxonomic delimitations involving parapatric taxa.

Wide-spread dispersal in a deep-sea brooding polychaete: the role of natural history collections in assessing the distribution in quill worms (Onuphidae, Annelida).

BACKGROUND: Modern integrative taxonomy-based annelid species descriptions are detailed combining morphological data and, since the last decades, also molecular information. Historic species descriptions are often comparatively brief lacking such detail. Adoptions of species names from western literature in the past led to the assumption of cosmopolitan ranges for many species, which, in many cases, were later found to include cryptic or pseudocryptic lineages with subtle morphological differences. Natural history collections and databases can aid in assessing the geographic ranges of species but depend on correct species identification. Obtaining DNA sequence information from wet-collection museum specimens of marine annelids is often impeded by the use of  formaldehyde and/or long-term storage in ethanol resulting in DNA degradation and cross-linking. RESULTS: The application of ancient DNA extraction methodology in combination with single-stranded DNA library preparation and target gene capture resulted in successful sequencing of a 110-year-old collection specimen of quill worms. Furthermore, a 40-year-old specimen of quill worms was successfully sequenced using a standard extraction protocol for modern samples, PCR and Sanger sequencing. Our study presents the first molecular analysis of Hyalinoecia species including the previously known species Hyalinoecia robusta, H. tubicloa, H. artifex, and H. longibranchiata, and a potentially undescribed species from equatorial western Africa morphologically indistinguishable from H. tubicola. The study also investigates the distribution of these five Hyalinoecia species. Reassessing the distribution of H. robusta reveals a geographical range covering both the Atlantic and the Indian Oceans as indicated by molecular data obtained from recent and historical specimens. CONCLUSION: Our results represent an example of a very wide geographical distribution of a brooding deep-sea annelid with a complex reproduction strategy and seemingly very limited dispersal capacity of its offspring, and highlights the importance of molecular information from museum specimens for integrative annelid taxonomy and biogeography.

Tangled webs and spider-flowers: Phylogenomics, biogeography, and seed morphology inform the evolutionary history of Cleomaceae.

PREMISE: Cleomaceae is an important model clade for studies of evolutionary processes including genome evolution, floral form diversification, and photosynthetic pathway evolution. Diversification and divergence patterns in Cleomaceae remain tangled as research has been restricted by its worldwide distribution, limited genetic sampling and species coverage, and a lack of definitive fossil calibration points. METHODS: We used target sequence capture and the Angiosperms353 probe set to perform a phylogenetic study of Cleomaceae. We estimated divergence times and biogeographic analyses to explore the origin and diversification of the family. Seed morphology across extant taxa was documented with multifocal image-stacking techniques and morphological characters were extracted, analyzed, and compared to fossil records. RESULTS: We recovered a well-supported and resolved phylogenetic tree of Cleomaceae generic relationships that includes 236 (~86%) species. We identified 11 principal clades and confidently placed Cleomella as sister to the rest of the family. Our analyses suggested that Cleomaceae and Brassicaceae diverged ~56 mya, and Cleomaceae began to diversify ~53 mya in the Palearctic and Africa. Multiple transatlantic disjunct distributions were identified. Seeds were imaged from 218 (~80%) species in the family and compared to all known fossil species. CONCLUSIONS: Our results represent the most comprehensive phylogenetic study of Cleomaceae to date. We identified transatlantic disjunctions and proposed explanations for these patterns, most likely either long-distance dispersals or contractions in latitudinal distributions caused by climate change over geological timescales. We found that seed morphology varied considerably but mostly mirrored generic relationships.

Assessment and validation of enrichment and target capture approaches to improve Mycobacterium tuberculosis WGS from direct patient samples.

Within-host Mycobacterium tuberculosis (Mtb) diversity may detect antibiotic resistance or predict tuberculosis treatment failure and is best captured through sequencing directly from sputum. Here, we compared three sample pre-processing steps for DNA decontamination and studied the yield of a new target enrichment protocol for optimal whole-genome sequencing (WGS) from direct patient samples. Mtb-positive NALC-NaOH-treated patient sputum sediments were pooled, and heat inactivated, split in replicates, and treated by either a wash, DNase I, or benzonase digestion. Levels of contaminating host DNA and target Mtb DNA were assessed by quantitative PCR (qPCR), followed by WGS with and without custom dsDNA target enrichment. The pre-treatment sample has a high host-to-target ratio of DNA (6,168 ± 1,638 host copies/ng to 212.3 ± 59.4 Mtb copies/ng) that significantly decreased with all three treatments. Benzonase treatment resulted in the highest enrichment of Mtb DNA at 100-fold compared with control (3,422 ± 2,162 host copies/ng to 11,721 ± 7,096 Mtb copies/ng). The custom dsDNA probe panel successfully enriched libraries from as little as 0.45 pg of Mtb DNA (100 genome copies). Applied to direct sputum the dsDNA target enrichment panel increased the percent of sequencing reads mapping to the Mtb target for all three pre-processing methods. Comparing the results of the benzonase sample sequenced both with and without enrichment, the percent of sequencing reads mapping to the Mtb increased to 90.95% from 1.18%. We demonstrate a low limit of detection for a new custom dsDNA Mtb target enrichment panel that has a favorable cost profile. The results also demonstrate that pre-processing to remove contaminating extracellular DNA prior to cell lysis and DNA extraction improves the host-to-Mtb DNA ratio but is not adequate to support average coverage WGS without target capture.

Phylogenomics sheds new light on the drivers behind a long-lasting systematic riddle: the figwort family Scrophulariaceae.

The figwort family, Scrophulariaceae, comprises c. 2000 species whose evolutionary relationships at the tribal level have proven difficult to resolve, hindering our ability to understand their origin and diversification. We designed a specific probe kit for Scrophulariaceae, targeting 849 nuclear loci and obtaining plastid regions as by-products. We sampled c. 87% of the genera described in the family and use the nuclear dataset to estimate evolutionary relationships, timing of diversification, and biogeographic patterns. Ten tribes, including two new tribes, Androyeae and Camptolomeae, are supported, and the phylogenetic positions of Androya, Camptoloma, and Phygelius are unveiled. Our study reveals a major diversification at c. 60 million yr ago in some Gondwanan landmasses, where two different lineages diversified, one of which gave rise to nearly 81% of extant species. A Southern African origin is estimated for most modern-day tribes, with two exceptions, the American Leucophylleae, and the mainly Australian Myoporeae. The rapid mid-Eocene diversification is aligned with geographic expansion within southern Africa in most tribes, followed by range expansion to tropical Africa and multiple dispersals out of Africa. Our robust phylogeny provides a framework for future studies aimed at understanding the role of macroevolutionary patterns and processes that generated Scrophulariaceae diversity.

Unmasking microsatellite deceptiveness and debunking hybridization with SNPs in four marine copepod species of Calanus.

Interspecific hybridization events are on the rise in natural systems due to climate change disrupting species barriers. Across taxa, microsatellites have long been the molecular markers of choice to identify admixed individuals. However, with the advent of high-throughput sequencing easing the generation of genome-wide datasets, incorrect reports of hybridization resulting from microsatellite technical artefacts have been uncovered in a growing number of taxa. In the marine zooplankton genus Calanus (Copepoda), whose species are used as climate change indicators, microsatellite markers have suggested hybridization between C. finmarchicus and C. glacialis, while other nuclear markers (InDels) never detected any admixed individuals, leaving the scientific community divided. Here, for the first time, we investigated the potential for hybridization among C. finmarchicus, C. glacialis, C. helgolandicus and C. hyperboreus using two large and independent SNP datasets. These were derived firstly from a protocol of target-capture applied to 179 individuals collected from 17 sites across the North Atlantic and Arctic Oceans, including sympatric areas, and second from published RNA sequences. All SNP-based analyses were congruent in showing that Calanus species are distinct and do not appear to hybridize. We then thoroughly re-assessed the microsatellites showing hybrids, with the support of published transcriptomes, and identified technical issues plaguing eight out of 10 microsatellites, including size homoplasy, paralogy, potential for null alleles and even two primer pairs targeting the same locus. Our study illustrates how deceptive microsatellites can be when applied to the investigation of hybridization.

Genetic mixing in conservation translocations increases diversity of a keystone threatened species, Bettongia lesueur.

Translocation programmes are increasingly being informed by genetic data to monitor and enhance conservation outcomes for both natural and established populations. These data provide a window into contemporary patterns of genetic diversity, structure and relatedness that can guide managers in how to best source animals for their translocation programmes. The inclusion of historical samples, where possible, strengthens monitoring by allowing assessment of changes in genetic diversity over time and by providing a benchmark for future improvements in diversity via management practices. Here, we used reduced representation sequencing (ddRADseq) data to report on the current genetic health of three remnant and seven translocated boodie (Bettongia lesueur) populations, now extinct on the Australian mainland. In addition, we used exon capture data from seven historical mainland specimens and a subset of contemporary samples to compare pre-decline and current diversity. Both data sets showed the significant impact of population founder source (whether multiple or single) on the genetic diversity of translocated populations. Populations founded by animals from multiple sources showed significantly higher genetic diversity than the natural remnant and single-source translocation populations, and we show that by mixing the most divergent populations, exon capture heterozygosity was restored to levels close to that observed in pre-decline mainland samples. Relatedness estimates were surprisingly low across all contemporary populations and there was limited evidence of inbreeding. Our results show that a strategy of genetic mixing has led to successful conservation outcomes for the species in terms of increasing genetic diversity and provides strong rationale for mixing as a management strategy.

Diversification and introgression in four chromosomal taxa of the Pearson's horseshoe bat (Rhinolophus pearsoni) group.

Chromosomal variation among closely related taxa is common in both plants and animals, and can reduce rates of introgression as well as promote reproductive isolation and speciation. In mammals, studies relating introgression to chromosomal variation have tended to focus on a few model systems and typically characterized levels of introgression using small numbers of loci. Here we took a genome-wide approach to examine how introgression rates vary among four closely related horseshoe bats (Rhinolophus pearsoni group) that possess different diploid chromosome numbers (2n = 42, 44, 46, and 60) resulting from Robertsonian (Rb) changes (fissions/fusions). Using a sequence capture we obtained orthologous loci for thousands of nuclear loci, as well as mitogenomes, and performed phylogenetic and population genetic analyses. We found that the taxon with 2n = 60 was the first to diverge in this group, and that the relationships among the three other taxa (2n = 42, 44 and 46) showed discordance across our different analyses. Our results revealed signatures of multiple ancient introgression events between the four taxa, with evidence of mitonuclar discordance in phylogenetic trees and reticulation events in their evolutionary history. Despite this, we found no evidence of recent and/or ongoing introgression between taxa. Overall, our results indicate that the effects of Rb changes on the reduction of introgression are complicated and that these may contribute to reproductive isolation and speciation in concert with other factors (e.g. phenotypic and genic divergence).

Phylogenomics reveals extreme gene tree discordance in a lineage of dominant trees: hybridization, introgression, and incomplete lineage sorting blur deep evolutionary relationships despite clear species groupings in Eucalyptus subgenus Eudesmia.

Eucalypts are a large and ecologically important group of plants on the Australian continent, and understanding their evolution is important in understanding evolution of the unique Australian flora. Previous phylogenies using plastome DNA, nuclear-ribosomal DNA, or random genome-wide SNPs, have been confounded by limited genetic sampling or by idiosyncratic biological features of the eucalypts, including widespread plastome introgression. Here we present phylogenetic analyses of Eucalyptus subgenus Eudesmia (22 species from western, northern, central and eastern Australia), in the first study to apply a target-capture sequencing approach using custom, eucalypt-specific baits (of 568 genes) to a lineage of Eucalyptus. Multiple accessions of all species were included, and target-capture data were supplemented by separate analyses of plastome genes (average of 63 genes per sample). Analyses revealed a complex evolutionary history likely shaped by incomplete lineage sorting and hybridization. Gene tree discordance generally increased with phylogenetic depth. Species, or groups of species, toward the tips of the tree are mostly supported, and three major clades are identified, but the branching order of these clades cannot be confirmed with confidence. Multiple approaches to filtering the nuclear dataset, by removing genes or samples, could not reduce gene tree conflict or resolve these relationships. Despite inherent complexities in eucalypt evolution, the custom bait kit devised for this research will be a powerful tool for investigating the evolutionary history of eucalypts more broadly.

Genomic, spatial and morphometric data for discrimination of four species in the Mediterranean Tamus clade of yams (Dioscorea, Dioscoreaceae).

BACKGROUND AND AIMS: Among the numerous pantropical species of the yam genus, Dioscorea, only a small group occurs in the Mediterranean basin, including two narrow Pyrenean endemics (Borderea clade) and two Mediterranean-wide species (D. communis and D. orientalis, Tamus clade). However, several currently unrecognized species and infraspecific taxa have been described in the Tamus clade due to significant morphological variation associated with D. communis. Our overarching aim was to investigate taxon delimitation in the Tamus clade using an integrative approach combining phylogenomic, spatial and morphological data. METHODS: We analysed 76 herbarium samples using Hyb-Seq genomic capture to sequence 260 low-copy nuclear genes and plastomes, together with morphometric and environmental modelling approaches. KEY RESULTS: Phylogenomic reconstructions confirmed that the two previously accepted species of the Tamus clade, D. communis and D. orientalis, are monophyletic and form sister clades. Three subclades showing distinctive geographic patterns were identified within D. communis. These subclades were also identifiable from morphometric and climatic data, and introgression patterns were inferred between subclades in the eastern part of the distribution of D. communis. CONCLUSIONS: We propose a taxonomy that maintains D. orientalis, endemic to the eastern Mediterranean region, and splits D. communis sensu lato into three species: D. edulis, endemic to Macaronesia (Canary Islands and Madeira); D. cretica, endemic to the eastern Mediterranean region; and D. communis sensu stricto, widespread across western and central Europe. Introgression inferred between D. communis s.s. and D. cretica is likely to be explained by their relatively recent speciation at the end of the Miocene, disjunct isolation in eastern and western Mediterranean glacial refugia and a subsequent westward recolonization of D. communis s.s. Our study shows that the use of integrated genomic, spatial and morphological approaches allows a more robust definition of species boundaries and the identification of species that previous systematic studies failed to uncover.

Comprehensive phylogenomic time tree of bryophytes reveals deep relationships and uncovers gene incongruences in the last 500 million years of diversification.

PREMISE: Bryophytes form a major component of terrestrial plant biomass, structuring ecological communities in all biomes. Our understanding of the evolutionary history of hornworts, liverworts, and mosses has been significantly reshaped by inferences from molecular data, which have highlighted extensive homoplasy in various traits and repeated bursts of diversification. However, the timing of key events in the phylogeny, patterns, and processes of diversification across bryophytes remain unclear. METHODS: Using the GoFlag probe set, we sequenced 405 exons representing 228 nuclear genes for 531 species from 52 of the 54 orders of bryophytes. We inferred the species phylogeny from gene tree analyses using concatenated and coalescence approaches, assessed gene conflict, and estimated the timing of divergences based on 29 fossil calibrations. RESULTS: The phylogeny resolves many relationships across the bryophytes, enabling us to resurrect five liverwort orders and recognize three more and propose 10 new orders of mosses. Most orders originated in the Jurassic and diversified in the Cretaceous or later. The phylogenomic data also highlight topological conflict in parts of the tree, suggesting complex processes of diversification that cannot be adequately captured in a single gene-tree topology. CONCLUSIONS: We sampled hundreds of loci across a broad phylogenetic spectrum spanning at least 450 Ma of evolution; these data resolved many of the critical nodes of the diversification of bryophytes. The data also highlight the need to explore the mechanisms underlying the phylogenetic ambiguity at specific nodes. The phylogenomic data provide an expandable framework toward reconstructing a comprehensive phylogeny of this important group of plants.

Interrogating Genomic-Scale Data to Resolve Recalcitrant Nodes in the Spider Tree of Life.

Genome-scale data sets are converging on robust, stable phylogenetic hypotheses for many lineages; however, some nodes have shown disagreement across classes of data. We use spiders (Araneae) as a system to identify the causes of incongruence in phylogenetic signal between three classes of data: exons (as in phylotranscriptomics), noncoding regions (included in ultraconserved elements [UCE] analyses), and a combination of both (as in UCE analyses). Gene orthologs, coded as amino acids and nucleotides (with and without third codon positions), were generated by querying published transcriptomes for UCEs, recovering 1,931 UCE loci (codingUCEs). We expected that congeners represented in the codingUCE and UCEs data would form clades in the presence of phylogenetic signal. Noncoding regions derived from UCE sequences were recovered to test the stability of relationships. Phylogenetic relationships resulting from all analyses were largely congruent. All nucleotide data sets from transcriptomes, UCEs, or a combination of both recovered similar topologies in contrast with results from transcriptomes analyzed as amino acids. Most relationships inferred from low-occupancy data sets, containing several hundreds of loci, were congruent across Araneae, as opposed to high occupancy data matrices with fewer loci, which showed more variation. Furthermore, we found that low-occupancy data sets analyzed as nucleotides (as is typical of UCE data sets) can result in more congruent relationships than high occupancy data sets analyzed as amino acids (as in phylotranscriptomics). Thus, omitting data, through amino acid translation or via retention of only high occupancy loci, may have a deleterious effect in phylogenetic reconstruction.

A large population sample of African HIV genomes from the 1980s reveals a reduction in subtype D over time associated with propensity for CXCR4 tropism.

We present 109 near full-length HIV genomes amplified from blood serum samples obtained during early 1986 from across Uganda, which to our knowledge is the earliest and largest population sample from the initial phase of the HIV epidemic in Africa. Consensus sequences were made from paired-end Illumina reads with a target-capture approach to amplify HIV material following poor success with standard approaches. In comparisons with a smaller 'intermediate' genome dataset from 1998 to 1999 and a 'modern' genome dataset from 2007 to 2016, the proportion of subtype D was significantly higher initially, dropping from 67% (73/109), to 57% (26/46) to 17% (82/465) respectively (p < 0.0001). Subtype D has previously been shown to have a faster rate of disease progression than other subtypes in East African population studies, and to have a higher propensity to use the CXCR4 co-receptor ("X4 tropism"); associated with a decrease in time to AIDS. Here we find significant differences in predicted tropism between A1 and D subtypes in all three sample periods considered, which is particularly striking the 1986 sample: 66% (53/80) of subtype D env sequences were predicted to be X4 tropic compared with none of the 24 subtype A1. We also analysed the frequency of subtype in the envelope region of inter-subtype recombinants, and found that subtype A1 is over-represented in env, suggesting recombination and selection have acted to remove subtype D env from circulation. The reduction of subtype D frequency over three decades therefore appears to be a result of selective pressure against X4 tropism and its higher virulence. Lastly, we find a subtype D specific codon deletion at position 24 of the V3 loop, which may explain the higher propensity for subtype D to utilise X4 tropism.

Adaptive potential of Coffea canephora from Uganda in response to climate change.

Understanding vulnerabilities of plant populations to climate change could help preserve their biodiversity and reveal new elite parents for future breeding programmes. To this end, landscape genomics is a useful approach for assessing putative adaptations to future climatic conditions, especially in long-lived species such as trees. We conducted a population genomics study of 207 Coffea canephora trees from seven forests along different climate gradients in Uganda. For this, we sequenced 323 candidate genes involved in key metabolic and defence pathways in coffee. Seventy-one single nucleotide polymorphisms (SNPs) were found to be significantly associated with bioclimatic variables, and were thereby considered as putatively adaptive loci. These SNPs were linked to key candidate genes, including transcription factors, like DREB-like and MYB family genes controlling plant responses to abiotic stresses, as well as other genes of organoleptic interest, such as the DXMT gene involved in caffeine biosynthesis and a putative pest repellent. These climate-associated genetic markers were used to compute genetic offsets, predicting population responses to future climatic conditions based on local climate change forecasts. Using these measures of maladaptation to future conditions, substantial levels of genetic differentiation between present and future diversity were estimated for all populations and scenarios considered. The populations from the forests Zoka and Budongo, in the northernmost zone of Uganda, appeared to have the lowest genetic offsets under all predicted climate change patterns, while populations from Kalangala and Mabira, in the Lake Victoria region, exhibited the highest genetic offsets. The potential of these findings in terms of ex situ conservation strategies are discussed.

HybPhaser identifies hybrid evolution in Australian Thelypteridaceae.

Hybridisation has been proposed to play an important role in fern evolution, but has been difficult to investigate. This study explores the utility of target sequence capture and read-to-reference phasing of putative hybrids to investigate the role of evolutionary reticulation in ferns using Australian Thelypteridaceae as a model. The bioinformatics workflow HybPhaser was used to assess divergence between alleles, phase sequence reads to references and construct accessions resembling parental haplotypes. These accessions were included in phylogenetic and network analyses to detect hybrids and infer their parentage. This approach identified two novel hybrid lineages in Thelypteridaceae, one occurring between two different genera (Abacopteris and Christella), and another as part of a complex of Christella. In addition, hybrid phasing successfully reduced conflicting data and improved overall resolution in the Thelypteridaceae phylogeny, highlighting the power of this approach for reconstructing evolutionary history in reticulated lineages.

Evolutionary relationships of anglerfishes (Lophiiformes) reconstructed using ultraconserved elements.

The macroevolutionary consequences of evolving in the deep-sea remain poorly understood and are compounded by the fact that convergent adaptations for living in this environment makes elucidating phylogenetic relationships difficult. Lophiiform anglerfishes exhibit extreme habitat and predatory specializations, including the use of a fin-spine system as a luring device and unique reproductive strategies where parasitic males attach and fuse to females. Despite their notoriety for these odd characteristics, evolutionary relationships among these fishes remain unclear. We sought to clarify the evolutionary history of Lophiiformes using data from 1000 ultraconserved elements and phylogenomic inference methods with particular interest paid to the Ceratioidei (deep-sea anglerfishes) and Antennarioidei (frogfishes and handfishes). At the suborder level, we recovered similar topologies in separate phylogenomic analyses: The Lophioidei (monkfishes) are the sister group to the rest of the Lophiiformes, Ogcocephaloidei (batfishes) and Antennarioidei (frogfishes) form a sister group, and Chaunacioidei (coffinfishes) and Ceratioidei (deep-sea anglerfishes) form a clade. The relationships we recover within the ceratioids disagree with most previous phylogenetic investigations, which used legacy phylogenetic markers or morphology. We recovered non-monophyletic relationships in the Antennarioidei and proposed three new families based on molecular and morphological evidence: Histiophrynidae, Rhycheridae, and Tathicarpidae. Antennariidae was re-evaluated to include what was known as Antennariinae, but not Histiophryninae. Non-bifurcating signal in splits network analysis indicated reticulations among and within suborders, supporting the complicated history of the Lophiiformes previously found with morphological data. Although we resolve relationships within Antennarioidei, Ceratioidei relationships remain somewhat unclear without better taxonomic sampling.

Cryptic species diversity in a widespread neotropical tree genus: The case of Cedrela odorata.

PREMISE: Reconciling the use of taxonomy to partition morphological variation and describe genetic divergence within and among closely related species is a persistent challenge in phylogenetics. We reconstructed phylogenetic relationships among Cedrela odorata (Meliaceae) and five closely allied species to test the genetic basis for the current model of species delimitation in this economically valuable and threatened genus. METHODS: We prepared a nuclear species tree with the program SNPhylo and 16,000 single-nucleotide polymorphisms from 168 Cedrela specimens. Based on clades present and ancestral patterns ADMIXTURE, we designed nine species delimitation models and compared each model to current taxonomy with Bayes factor delimitation. Timing of major lineage divergences was estimated with the program SNAPP. RESULTS: The resulting analysis revealed that modern C. odorata evolved from two genetically distinct ancestral sources. All species delimitation models tested better fit the data than the model representing current taxonomic delimitation. Models with the greatest marginal likelihoods separated Mesoamerican C. odorata and South American C. odorata into two species and lumped C. angustifolia and C. montana as a single species. We estimated that Cedrela diversified in South America within the last 19 million years following one or more dispersal events from Mesoamerican lineages. CONCLUSIONS: Our analyses show that the present taxonomic understanding within the genus obscures divergent lineages in C. odorata due in part to morphological differentiation and taxonomic distinctions that are not predictably associated with genetic divergence. A more accurate application of taxonomy to C. odorata and related species may aid in its conservation, management, and restoration efforts.

Phylogenomic discordance suggests polytomies along the backbone of the large genus Solanum.

PREMISE: Evolutionary studies require solid phylogenetic frameworks, but increased volumes of phylogenomic data have revealed incongruent topologies among gene trees in many organisms both between and within genomes. Some of these incongruences indicate polytomies that may remain impossible to resolve. Here we investigate the degree of gene-tree discordance in Solanum, one of the largest flowering plant genera that includes the cultivated potato, tomato, and eggplant, as well as 24 minor crop plants. METHODS: A densely sampled species-level phylogeny of Solanum is built using unpublished and publicly available Sanger sequences comprising 60% of all accepted species (742 spp.) and nine regions (ITS, waxy, and seven plastid markers). The robustness of this topology is tested by examining a full plastome dataset with 140 species and a nuclear target-capture dataset with 39 species of Solanum (Angiosperms353 probe set). RESULTS: While the taxonomic framework of Solanum remained stable, gene tree conflicts and discordance between phylogenetic trees generated from the target-capture and plastome datasets were observed. The latter correspond to regions with short internodal branches, and network analysis and polytomy tests suggest the backbone is composed of three polytomies found at different evolutionary depths. The strongest area of discordance, near the crown node of Solanum, could potentially represent a hard polytomy. CONCLUSIONS: We argue that incomplete lineage sorting due to rapid diversification is the most likely cause for these polytomies, and that embracing the uncertainty that underlies them is crucial to understand the evolution of large and rapidly radiating lineages.