Impact of Ghost Introgression on Coalescent-based Species Tree Inference and Estimation of Divergence Time

The species studied in any evolutionary investigation generally constitute a very small proportion of all the species currently existing or that have gone extinct. It is therefore likely that introgression, which is widespread across the tree of life, involves "ghosts," i.e., unsampled, unknown, or extinct lineages. However, the impact of ghost introgression on estimations of species trees has been rarely studied and is thus poorly understood. In this study, we use mathematical analysis and simulations to examine the robustness of species tree methods based on a multispecies coalescent model under gene flow sourcing from an extant or ghost lineage. We found that very low levels of extant or ghost introgression can result in anomalous gene trees (AGTs) on three-taxon rooted trees if accompanied by strong incomplete lineage sorting (ILS). In contrast, even massive introgression, with more than half of the recipient genome descending from the donor lineage, may not necessarily lead to AGTs. In cases involving an ingroup lineage (defined as one that diverged no earlier than the most basal species under investigation) acting as the donor of introgression, the time of root divergence among the investigated species was either underestimated or remained unaffected, but for the cases of outgroup ghost lineages acting as donors, the divergence time was generally overestimated. Under many conditions of ingroup introgression, the stronger the ILS was, the higher was the accuracy of estimating the time of root divergence, although the topology of the species tree is more prone to be biased by the effect of introgression.

Recent Progress on Methods for Estimating and Updating Large Phylogenies

With the increased availability of sequence data and even of fully sequenced and assembled genomes, phylogeny estimation of very large trees (even of hundreds of thousands of sequences) is now a goal for some biologists. Yet, the construction of these phylogenies is a complex pipeline presenting analytical and computational challenges, especially when the number of sequences is very large. In the last few years, new methods have been developed that aim to enable highly accurate phylogeny estimations on these large datasets, including divide-and-conquer techniques for multiple sequence alignment and/or tree estimation, methods that can estimate species trees from multi-locus datasets while addressing heterogeneity due to biological processes (e.g., incomplete lineage sorting and gene duplication and loss), and methods to add sequences into large gene trees or species trees. Here we present some of these recent advances and discuss opportunities for future improvements.

How to Tackle Phylogenetic Discordance in Recent and Rapidly Radiating Groups? Developing a Workflow Using Loricaria (Asteraceae) as an Example

A major challenge in phylogenetics and -genomics is to resolve young rapidly radiating groups. The fast succession of species increases the probability of incomplete lineage sorting (ILS), and different topologies of the gene trees are expected, leading to gene tree discordance, i.e., not all gene trees represent the species tree. Phylogenetic discordance is common in phylogenomic datasets, and apart from ILS, additional sources include hybridization, whole-genome duplication, and methodological artifacts. Despite a high degree of gene tree discordance, species trees are often well supported and the sources of discordance are not further addressed in phylogenomic studies, which can eventually lead to incorrect phylogenetic hypotheses, especially in rapidly radiating groups. We chose the high-Andean Asteraceae genus Loricaria to shed light on the potential sources of phylogenetic discordance and generated a phylogenetic hypothesis. By accounting for paralogy during gene tree inference, we generated a species tree based on hundreds of nuclear loci, using Hyb-Seq, and a plastome phylogeny obtained from off-target reads during target enrichment. We observed a high degree of gene tree discordance, which we found implausible at first sight, because the genus did not show evidence of hybridization in previous studies. We used various phylogenomic analyses (trees and networks) as well as the D-statistics to test for ILS and hybridization, which we developed into a workflow on how to tackle phylogenetic discordance in recent radiations. We found strong evidence for ILS and hybridization within the genus Loricaria. Low genetic differentiation was evident between species located in different Andean cordilleras, which could be indicative of substantial introgression between populations, promoted during Pleistocene glaciations, when alpine habitats shifted creating opportunities for secondary contact and hybridization.

Mahonia vs. Berberis Unloaded: Generic Delimitation and Infrafamilial Classification of Berberidaceae Based on Plastid Phylogenomics

The early-diverging eudicot family Berberidaceae is composed of a morphologically diverse assemblage of disjunctly distributed genera long praised for their great horticultural and medicinal values. However, despite century-long studies, generic delimitation of Berberidaceae remains controversial and its tribal classification has never been formally proposed under a rigorous phylogenetic context. Currently, the number of accepted genera in Berberidaceae ranges consecutively from 13 to 19, depending on whether to define Berberis, Jeffersonia, and Podophyllum broadly, or to segregate these three genera further and recognize Alloberberis, Mahonia, and Moranothamnus, Plagiorhegma, and Dysosma, Diphylleia, and Sinopodophyllum, respectively. To resolve Berberidaceae’s taxonomic disputes, we newly assembled 23 plastomes and, together with 85 plastomes from the GenBank, completed the generic sampling of the family. With 4 problematic and 14 redundant plastome sequences excluded, robust phylogenomic relationships were reconstructed based on 93 plastomes representing all 19 genera of Berberidaceae and three outgroups. Maximum likelihood phylogenomic relationships corroborated with divergence time estimation support the recognition of three subfamilies Berberidoideae, Nandinoideae, and Podophylloideae, with tribes Berberideae and Ranzanieae, Leonticeae and Nandineae, and Podophylleae, Achlydeae, Bongardieae tr. nov., Epimedieae, and Jeffersonieae tr. nov. in the former three subfamilies, respectively. By applying specifically stated criteria, our phylogenomic data also support the classification of 19 genera, recognizing Alloberberis, Mahonia, and Moranothamnus, Plagiorhegma, and Diphylleia, Dysosma, and Sinopodophyllum that are morphologically and evolutionarily distinct from Berberis, Jeffersonia, and Podophyllum, respectively. Comparison of plastome structures across Berberidaceae confirms inverted repeat expansion in the tribe Berberideae and reveals substantial length variation in accD gene caused by repeated sequences in Berberidoideae. Comparison of plastome tree with previous studies and nuclear ribosomal DNA (nrDNA) phylogeny also reveals considerable conflicts at different phylogenetic levels, suggesting that incomplete lineage sorting and/or hybridization had occurred throughout the evolutionary history of Berberidaceae and that Alloberberis and Moranothamnus could have resulted from reciprocal hybridization between Berberis and Mahonia in ancient times prior to the radiations of the latter two genera.

Herzog Vindicated: Integrative Taxonomy Reveals that Trichostomum brachydontium (Pottiaceae, Bryophyta) Comprises Several Species

The morphologically variable moss Trichostomum brachydontium is very common in south and west Europe, particularly under Mediterranean and Atlantic climates. A morphological study was conducted alongside a molecular phylogenetic study based on nr ITS and cp rbcL regions in order to assess if T. brachydontium is an exceptionally polymorphic species as evidenced by the number of described infraspecific taxa from the last century or, alternatively, if it includes more than one species, and if so, to find the valid name for them. Phylogenetic analyses of both nuclear and chloroplast datasets show that there are four well-supported clades. While the ITS based tree is in good agreement with the morphological data, there are a few inconsistencies with reference to the rbcL tree; this may be explained by incomplete lineage sorting or by hybridization. The morphological survey revealed well-defined discriminate differences between the four phylogenetic lineages. The taxonomic conclusions include the recognition of four species: T. brachydontium s.s., T. herzogii (a new name proposed for var. cuspidatum), T. littorale and T. meridionale (a new name proposed for var. densum). Lectotypes are designated for T. brachydontium and T. littorale. Our results underline the ongoing need of integrative studies to examine further the underestimated diversity of the T. brachydontium complex in other regions.

Complete mitochondrial genome of Manis pentadactyla pentadactyla (Mammalia: Pholidota), an endemic subspecies of Chinese pangolin: mitogenome characterisation and phylogenetic implications

The Chinese pangolin Manis pentadactyla is critically endangered because of over-exploitation and illegal trafficking and includes three subspecies. However, the taxonomic status of the three subspecies of the Chinese pangolin has not been well resolved, which impedes regional conservation and illegal trade traces. In this study, the complete mitogenome sequence of M. p. pentadactyla, an endemic subspecies of the Chinese pangolin in Taiwan, was determined. The complete mitogenome of M. p. pentadactyla is 16,570 base pairs (bp) in length with 13 protein-coding genes (PCG), 23 transfer RNAs (tRNAs), two ribosomal RNAs and a 1164 bp control region. The overall base composition of the genome showed a slight A + T bias (59.9%), positive AT skew (0.1515) and negative GC skew (-0.3406), which is similar to that of other pangolins. All PCGs started with a typical ATN codon and all tRNAs were typical cloverleaf-shaped secondary structures, except for tRNA-Ser(GCU). Phylogenetic analysis indicated a monophyletic relationship for M. p. pentadactyla and M. p. aurita and was monophyletic for M. p. pentadactyla, but paraphyletic for M. p. aurita. The paraphyly of M. p. aurita resulted from an incomplete lineage sorting. This study enriched the mitogenome database of the Chinese pangolin and the molecular information obtained should be very useful for future research on mitogenome evolution and genetic diversification in M. pentadactyla.

The Chloranthus sessilifolius genome provides insight into early diversification of angiosperms

Most extant angiosperms belong to Mesangiospermae, which comprises eudicots, monocots, magnoliids, Chloranthales and Ceratophyllales. However, phylogenetic relationships between these five lineages remain unclear. Here, we report the high-quality genome of a member of the Chloranthales lineage (Chloranthus sessilifolius). We detect only one whole genome duplication within this species and find that polyploidization events in different Mesangiospermae lineage are mutually independent. We also find that the members of all floral development-related gene lineages are present in C. sessilifolius despite its extremely simplified flower. The AP1 and PI genes, however, show a weak floral tissue-specialized expression. Our phylogenomic analyses suggest that Chloranthales and magnoliids are sister groups, and both are together sister to the clade comprising Ceratophyllales and eudicots, while the monocot lineage is sister to all other Mesangiospermae. Our findings suggest that in addition to hybridization, incomplete lineage sorting may largely account for phylogenetic inconsistencies between the observed gene trees.

A phylogeny of the genus Limia (Teleostei: Poeciliidae) suggests a single-lake radiation nested in a Caribbean-wide allopatric speciation scenario

Objective The Caribbean is an important global biodiversity hotspot. Adaptive radiations there lead to many speciation events within a limited period and hence are particularly prominent biodiversity generators. A prime example are freshwater fish of the genus Limia, endemic to the Greater Antilles. Within Hispaniola, nine species have been described from a single isolated site, Lake Miragoâne, pointing towards extraordinary sympatric speciation. This study examines the evolutionary history of the Limia species in Lake Miragoâne, relative to their congeners throughout the Caribbean. Results For 12 Limia species, we obtained almost complete sequences of the mitochondrial cytochrome b gene, a well-established marker for lower-level taxonomic relationships. We included sequences of six further Limia species from GenBank (total N = 18 species). Our phylogenies are in concordance with other published phylogenies of Limia. There is strong support that the species found in Lake Miragoâne in Haiti are monophyletic, confirming a recent local radiation. Within Lake Miragoâne, speciation is likely extremely recent, leading to incomplete lineage sorting in the mtDNA. Future studies using multiple unlinked genetic markers are needed to disentangle the relationships within the Lake Miragoâne clade.

Sage Insights Into the Phylogeny of Salvia: Dealing With Sources of Discordance Within and Across Genomes

Next-generation sequencing technologies have facilitated new phylogenomic approaches to help clarify previously intractable relationships while simultaneously highlighting the pervasive nature of incongruence within and among genomes that can complicate definitive taxonomic conclusions. Salvia L., with ∼1,000 species, makes up nearly 15% of the species diversity in the mint family and has attracted great interest from biologists across subdisciplines. Despite the great progress that has been achieved in discerning the placement of Salvia within Lamiaceae and in clarifying its infrageneric relationships through plastid, nuclear ribosomal, and nuclear single-copy genes, the incomplete resolution has left open major questions regarding the phylogenetic relationships among and within the subgenera, as well as to what extent the infrageneric relationships differ across genomes. We expanded a previously published anchored hybrid enrichment dataset of 35 exemplars of Salvia to 179 terminals. We also reconstructed nearly complete plastomes for these samples from off-target reads. We used these data to examine the concordance and discordance among the nuclear loci and between the nuclear and plastid genomes in detail, elucidating both broad-scale and species-level relationships within Salvia. We found that despite the widespread gene tree discordance, nuclear phylogenies reconstructed using concatenated, coalescent, and network-based approaches recover a common backbone topology. Moreover, all subgenera, except for Audibertia, are strongly supported as monophyletic in all analyses. The plastome genealogy is largely resolved and is congruent with the nuclear backbone. However, multiple analyses suggest that incomplete lineage sorting does not fully explain the gene tree discordance. Instead, horizontal gene flow has been important in both the deep and more recent history of Salvia. Our results provide a robust species tree of Salvia across phylogenetic scales and genomes. Future comparative analyses in the genus will need to account for the impacts of hybridization/introgression and incomplete lineage sorting in topology and divergence time estimation.

Interrogating Genomic Data in the Phylogenetic Placement of Treeshrews Reveals Potential Sources of Conflict

The position of some taxa on the Tree of Life remains controversial despite the increase in genomic data used to infer phylogenies. While analyzing large datasets alleviates stochastic errors, it does not prevent systematic errors in inference, caused by both biological (e.g., incomplete lineage sorting, hybridization) and methodological (e.g., incorrect modeling, erroneous orthology assessments) factors. In our study, we systematically investigated factors that could result in these controversies, using the treeshrew (Scandentia, Mammalia) as a study case. Recent studies have narrowed the phylogenetic position of treeshrews to three competing hypotheses: sister to primates and flying lemurs (Primatomorpha), sister to rodents and lagomorphs (Glires), or sister to a clade comprising all of these. We sampled 50 mammal species including three treeshrews, a selection of taxa from the potential sister groups, and outgroups. Using a large diverse set of loci, we assessed support for the alternative phylogenetic position of treeshrews. A plurality of loci support treeshrews as sister to rodents and lagomorphs; however, only a few loci exhibit strong support for any hypothesis. Surprisingly, we found that a subset of loci that strongly support the monophyly of Primates, support treeshrews as sister to primates and flying lemurs. The overall small magnitude of differences in phylogenetic signal among the alternative hypotheses suggests that these three groups diversified nearly simultaneously. However, with our large dataset and approach to examining support, we provide evidence for the hypothesis of treeshrews as sister to rodents and lagomorphs, while demonstrating why support for alternate hypotheses has been seen in prior work. We also suggest that locus selection can unwittingly bias results.