The role of multiple Pleistocene refugia in promoting diversification in the Pacific Northwest.

Pleistocene glacial cycles drastically changed the distributions of taxa endemic to temperate rainforests in the Pacific Northwest, with many experiencing reduced habitat suitability during glacial periods. In this study, we investigate whether glacial cycles promoted intraspecific divergence and whether subsequent range changes led to secondary contact and gene flow. For seven invertebrate species endemic to the PNW, we estimated species distribution models (SDMs) and projected them onto current and historical climate conditions to assess how habitat suitability changed during glacial cycles. Using single nucleotide polymorphism (SNP) data from these species, we assessed population genetic structure and used a machine-learning approach to compare models with and without gene flow between populations upon secondary contact after the last glacial maximum (LGM). Finally, we estimated divergence times and rates of gene flow between populations. SDMs suggest that there was less suitable habitat in the North Cascades and Northern Rocky Mountains during glacial compared to interglacial periods, resulting in reduced habitat suitability and increased habitat fragmentation during the LGM. Our genomic data identify population structure in all taxa, and support gene flow upon secondary contact in five of the seven taxa. Parameter estimates suggest that population divergences date to the later Pleistocene for most populations. Our results support a role of refugial dynamics in driving intraspecific divergence in the Cascades Range. In these invertebrates, population structure often does not correspond to current biogeographic or environmental barriers. Rather, population structure may reflect refugial lineages that have since expanded their ranges, often leading to secondary contact between once isolated lineages.

Genomic evidence of an ancient inland temperate rainforest in the Pacific Northwest of North America.

The disjunct temperate rainforests of the Pacific Northwest of North America (PNW) are characterized by late-successional dominant tree species Thuja plicata (western redcedar) and Tsuga heterophylla (western hemlock). The demographic histories of these species, along with the PNW rainforest ecosystem in its entirety, have been heavily impacted by geological and climatic changes the PNW has experienced over the last 5 million years, including mountain orogeny and repeated Pleistocene glaciations. These environmental events have ultimately shaped the history of these species, with inland populations potentially being extirpated during the Pleistocene glaciations. Here, we collect genomic data for both species across their ranges to test multiple demographic models, each reflecting a different phylogeographical hypothesis on how the ecosystem-dominating species may have responded to dramatic climatic change. Our results indicate that inland and coastal populations in both species diverged ~2.5 million years ago in the early Pleistocene and experienced decreases in population size during glacial cycles, with subsequent population expansion. Importantly, we found evidence for gene flow between coastal and inland populations during the mid-Holocene. It is likely that intermittent migration in these species during this time has prevented allopatric speciation via genetic drift alone. In conclusion, our results from combining genomic data and demographic inference procedures establish that populations of the ecosystem dominants Thuja plicata and Tsuga heterophylla persisted in refugia located in both the coastal and inland regions of the PNW throughout the Pleistocene, with populations expanding and contracting in response to glacial cycles with occasional gene flow.

Analysis of Paralogs in Target Enrichment Data Pinpoints Multiple Ancient Polyploidy Events in Alchemilla s.l. (Rosaceae).

Target enrichment is becoming increasingly popular for phylogenomic studies. Although baits for enrichment are typically designed to target single-copy genes, paralogs are often recovered with increased sequencing depth, sometimes from a significant proportion of loci, especially in groups experiencing whole-genome duplication (WGD) events. Common approaches for processing paralogs in target enrichment data sets include random selection, manual pruning, and mainly, the removal of entire genes that show any evidence of paralogy. These approaches are prone to errors in orthology inference or removing large numbers of genes. By removing entire genes, valuable information that could be used to detect and place WGD events is discarded. Here, we used an automated approach for orthology inference in a target enrichment data set of 68 species of Alchemilla s.l. (Rosaceae), a widely distributed clade of plants primarily from temperate climate regions. Previous molecular phylogenetic studies and chromosome numbers both suggested ancient WGDs in the group. However, both the phylogenetic location and putative parental lineages of these WGD events remain unknown. By taking paralogs into consideration and inferring orthologs from target enrichment data, we identified four nodes in the backbone of Alchemilla s.l. with an elevated proportion of gene duplication. Furthermore, using a gene-tree reconciliation approach, we established the autopolyploid origin of the entire Alchemilla s.l. and the nested allopolyploid origin of four major clades within the group. Here, we showed the utility of automated tree-based orthology inference methods, previously designed for genomic or transcriptomic data sets, to study complex scenarios of polyploidy and reticulate evolution from target enrichment data sets.[Alchemilla; allopolyploidy; autopolyploidy; gene tree discordance; orthology inference; paralogs; Rosaceae; target enrichment; whole genome duplication.].

Phylogenomic analyses in Phrymaceae reveal extensive gene tree discordance in relationships among major clades.

PREMISE: Phylogenomic datasets using genomes and transcriptomes provide rich opportunities beyond resolving bifurcating phylogenetic relationships. Monkeyflower (Phrymaceae) is a model system for evolutionary ecology. However, it lacks a well-supported phylogeny as a basis for a stable taxonomy and for macroevolutionary comparisons. METHODS: We sampled 24 genomes and transcriptomes in Phrymaceae and closely related families, including eight newly sequenced transcriptomes. We reconstructed the phylogeny using IQ-TREE and ASTRAL, evaluated gene tree discordance using PhyParts, Quartet Sampling, and a cloudogram, and carried out reticulation analyses using PhyloNet and HyDe. We searched for whole genome duplication (WGD) events using chromosome numbers, synonymous distances, and gene duplication events as evidence. RESULTS: Most gene trees support the monophyly of Phrymaceae and each of its tribes. Most gene trees also support tribe Mimuleae being sister to Phrymeae + Diplaceae + Leucocarpeae, with extensive gene tree discordance among the latter three. Despite the discordance, the monophyly of Mimulus s.l. is rejected, and no individual reticulation event among the Phrymaceae tribes is well-supported. Reticulation likely occurred among Erythranthe bicolor and closely related species. No ancient WGD was detected in Phrymaceae. Instead, small-scale duplications are among potential drivers of macroevolutionary diversification of Phrymaceae. CONCLUSIONS: We show that analysis of reticulate evolution is sensitive to taxon sampling and methods used. We also demonstrate that phylogenomic datasets using genomes and transcriptomes present rich opportunities to investigate gene family evolution and genome duplication events involved in lineage diversification and adaptation.

Phylogeny, classification, and character evolution of tribe Citharexyleae (Verbenaceae).

PREMISE: As a family of Neotropical origin and primarily Neotropical distribution, the Verbenaceae are a good but understudied system with which to understand Neotropical evolution. Tribe Citharexyleae comprises three genera: Baillonia, Citharexylum-one of the largest genera in Verbenaceae-and Rehdera. A molecular phylogenetic approach was taken to resolve intergeneric relationships in Citharexyleae and infrageneric relationships in Citharexylum. The phylogeny is used to elucidate character evolution in a widespread, morphologically diverse Neotropical genus. METHODS: Seven plastid regions, two nuclear ribosomal spacers, and six low-copy nuclear loci were analyzed for 64 species of Citharexyleae. Phylogenetic analyses were conducted using maximum likelihood, Bayesian inference, and multispecies coalescent approaches. Habit, presence or absence of thorns, inflorescence architecture, flower color, fruit color, and geography were examined to identify diagnostic character states for clades within Citharexylum. RESULTS: Rehdera is resolved as sister to Citharexylum, and Baillonia nested within Citharexylum. Two species, C. oleinum and C. tetramerum, are not closely related to tribe Citharexyleae, but may be related to members of tribe Duranteae instead. Seven clades within Citharexylum are inferred, each characterized by a combination of geography, fruit color and/or maturation, and inflorescence architecture. There is evidence of correlated evolution between habit, axillary inflorescences, and flower number per inflorescence. Shrubs with reduced inflorescences have evolved repeatedly. CONCLUSIONS: A subgeneric classification for Citharexylum is proposed. Although suites of associated traits are found, character morphology has been labile throughout Citharexylum's evolutionary history. Morphological diversity may be related to adaptation to differing mesic and xeric habitats.

Predicting plant conservation priorities on a global scale.

The conservation status of most plant species is currently unknown, despite the fundamental role of plants in ecosystem health. To facilitate the costly process of conservation assessment, we developed a predictive protocol using a machine-learning approach to predict conservation status of over 150,000 land plant species. Our study uses open-source geographic, environmental, and morphological trait data, making this the largest assessment of conservation risk to date and the only global assessment for plants. Our results indicate that a large number of unassessed species are likely at risk and identify several geographic regions with the highest need of conservation efforts, many of which are not currently recognized as regions of global concern. By providing conservation-relevant predictions at multiple spatial and taxonomic scales, predictive frameworks such as the one developed here fill a pressing need for biodiversity science.

Three keys to the radiation of angiosperms into freezing environments.

Early flowering plants are thought to have been woody species restricted to warm habitats. This lineage has since radiated into almost every climate, with manifold growth forms. As angiosperms spread and climate changed, they evolved mechanisms to cope with episodic freezing. To explore the evolution of traits underpinning the ability to persist in freezing conditions, we assembled a large species-level database of growth habit (woody or herbaceous; 49,064 species), as well as leaf phenology (evergreen or deciduous), diameter of hydraulic conduits (that is, xylem vessels and tracheids) and climate occupancies (exposure to freezing). To model the evolution of species' traits and climate occupancies, we combined these data with an unparalleled dated molecular phylogeny (32,223 species) for land plants. Here we show that woody clades successfully moved into freezing-prone environments by either possessing transport networks of small safe conduits and/or shutting down hydraulic function by dropping leaves during freezing. Herbaceous species largely avoided freezing periods by senescing cheaply constructed aboveground tissue. Growth habit has long been considered labile, but we find that growth habit was less labile than climate occupancy. Additionally, freezing environments were largely filled by lineages that had already become herbs or, when remaining woody, already had small conduits (that is, the trait evolved before the climate occupancy). By contrast, most deciduous woody lineages had an evolutionary shift to seasonally shedding their leaves only after exposure to freezing (that is, the climate occupancy evolved before the trait). For angiosperms to inhabit novel cold environments they had to gain new structural and functional trait solutions; our results suggest that many of these solutions were probably acquired before their foray into the cold.

Extensive allopolyploidy in the neotropical genus Lachemilla (Rosaceae) revealed by PCR-based target enrichment of the nuclear ribosomal DNA cistron and plastid phylogenomics.

PREMISE OF THE STUDY: Polyploidy has been long recognized as an important force in plant evolution. Previous studies had suggested widespread occurrence of polyploidy and the allopolyploid origin of several species in the diverse neotropical genus Lachemilla (Rosaceae). Nonetheless, this evidence has relied mostly on patterns of cytonuclear discordance, and direct evidence from nuclear allelic markers is still needed. METHODS: Here we used PCR target enrichment in combination with high throughput sequencing to obtain multiple copies of the nuclear ribosomal (nr) DNA cistron and 45 regions of the plastid genome (cpDNA) from 219 accessions representing 48 species of Lachemilla and to explore the allopolyploid origin of species in this group. KEY RESULTS: We were able to identify multiple nrDNA ribotypes and establish clear evidence of allopolyploidy in 33 species of Lachemilla, showing that this condition is common and widespread in the genus. Additionally, we found evidence for three autopolyploid species. We also established multiple, independent origins of several allopolyploid species. Finally, based solely on the cpDNA phylogeny, we identified that the monotypic genus Farinopsis is the sister group of Lachemilla and allied genera within subtribe Fragariinae. CONCLUSIONS: Our study demonstrates the utility of the nuclear ribosomal DNA cistron to detect allopolyploidy when concerted evolution of this region is not complete. Additionally, with a robust chloroplast phylogeny in place, the direction of hybridization events can be established, and multiple, independent origins of allopolyploid species can be identified.

Increasing phylogenetic stochasticity at high elevations on summits across a remote North American wilderness.

PREMISE: At the intersection of ecology and evolutionary biology, community phylogenetics can provide insights into overarching biodiversity patterns, particularly in remote and understudied ecosystems. To understand community assembly of the high alpine flora in the Sawtooth National Forest, USA, we analyzed phylogenetic structure within and between nine summit communities. METHODS: We used high-throughput sequencing to supplement existing data and infer a nearly completely sampled community phylogeny of the alpine vascular flora. We calculated mean nearest taxon distance (MNTD) and mean pairwise distance (MPD) to quantify phylogenetic divergence within summits, and assessed whether maximum elevation explains phylogenetic structure. To evaluate similarities between summits, we quantified phylogenetic turnover, taking into consideration microhabitats (talus vs. meadows). RESULTS: We found different patterns of community phylogenetic structure within the six most species-rich orders, but across all vascular plants phylogenetic structure was largely not different from random. There was a significant negative correlation between elevation and tree-wide phylogenetic diversity (MPD) within summits: overdispersion degraded as elevation increased. Between summits, we found high phylogenetic turnover driven by greater niche heterogeneity on summits with alpine meadows. CONCLUSIONS: Our results provide further evidence that stochastic processes may also play an important role in the assembly of vascular plant communities in high alpine habitats at regional scales. However, order-specific patterns suggest that adaptations are still important for assembly of specific sectors of the plant tree of life. Further studies quantifying functional diversity will be important in disentangling the interplay of eco-evolutionary processes that likely shape broad community phylogenetic patterns in extreme environments.

Phylogenomic analyses reveal a deep history of hybridization and polyploidy in the Neotropical genus Lachemilla (Rosaceae).

Hybridization, incomplete lineage sorting, and phylogenetic error produce similar incongruence patterns, representing a great challenge for phylogenetic reconstruction. Here, we use sequence capture data and multiple species tree and species network approaches to resolve the backbone phylogeny of the Neotropical genus Lachemilla, while distinguishing among sources of incongruence. We used 396 nuclear loci and nearly complete plastome sequences from 27 species to clarify the relationships among the major groups of Lachemilla, and explored multiple sources of conflict between gene trees and species trees inferred with a plurality of approaches. All phylogenetic methods recovered the four major groups previously proposed for Lachemilla, but species tree methods recovered different topologies for relationships between these four clades. Species network analyses revealed that one major clade, Orbiculate, is likely of ancient hybrid origin, representing one of the main sources of incongruence among the species trees. Additionally, we found evidence for a potential whole genome duplication event shared by Lachemilla and allied genera. Lachemilla shows clear evidence of ancient and recent hybridization throughout the evolutionary history of the group. Also, we show the necessity to use phylogenetic network approaches that can simultaneously accommodate incomplete lineage sorting and gene flow when studying groups that show patterns of reticulation.

Combining allele frequency and tree-based approaches improves phylogeographic inference from natural history collections.

Model selection approaches in phylogeography have allowed researchers to evaluate the support for competing demographic histories, which provides a mode of inference and a measure of uncertainty in understanding climatic and spatial influences on intraspecific diversity. Here, to rank all models in the comparison set and determine what proportion of the total support the top-ranked model garners, we conduct model selection using two analytical approaches-allele frequency-based, implemented in fastsimcoal2, and gene tree-based, implemented in phrapl. We then expand this model selection framework by including an assessment of absolute fit of the models to the data. For this, we utilize DNA isolated from existing natural history collections that span the distribution of red alder (Alnus rubra) in the Pacific Northwest of North America to generate genomic data for the evaluation of 13 demographic scenarios. The quality of DNA recovered from herbarium specimen leaf tissue was assessed for its utility and effectiveness in demographic model selection, specifically in the two approaches mentioned. We present strong support for the use of herbarium tissue in the generation of genomic DNA, albeit with the inclusion of additional quality control checks prior to library preparation and analyses with multiple approaches that incorporate various data. Analyses with allele frequency spectra and gene trees predominantly support A. rubra having experienced an ancient vicariance event with intermittent and frequent gene flow between the disjunct populations. Additionally, the data consistently fit the most frequently selected model, corroborating the model selection techniques. Finally, these results suggest that the A. rubra disjunct populations do not represent separate species.

Incongruence in molecular species delimitation schemes: What to do when adding more data is difficult.

Using multiple, independent approaches to molecular species delimitation is advocated to accommodate limitations and assumptions of a single approach. Incongruence in delimitation schemes is a potential by-product of employing multiple methods on the same data, and little attention has been paid to its reconciliation. Instead, a particular scheme is prioritized, and/or molecular delimitations are coupled with additional, independent lines of evidence that mitigate incongruence. We advocate that incongruence within a line of evidence should be accounted for before comparing across lines of evidence that can themselves be incongruent. Additionally, it is not uncommon for empiricists working in nonmodel systems to be data-limited, generating some concern for the adequacy of available data to address the question of interest. With conservation and management decisions often hinging on the status of species, it seems prudent to understand the capabilities of approaches we use given the data we have. Here, we apply two molecular species delimitation approaches, spedeSTEM and BPP, to the Castilleja ambigua (Orobanchaceae) species complex, a relatively young plant lineage in western North America. Upon finding incongruence in our delimitation, we employed a post hoc simulation study to examine the power of these approaches to delimit species. Given the data we collected, we find that spedeSTEM lacks the power to delimit while BPP is capable, thus allowing us to address incongruence before proceeding in delimitation. We suggest post hoc simulation studies like this compliment empirical delimitation and serve as a means of exploring conflict within a line of evidence and dealing with it appropriately.

Impact of whole-genome duplication events on diversification rates in angiosperms.

PREMISE OF THE STUDY: Polyploidy or whole-genome duplication (WGD) pervades the evolutionary history of angiosperms. Despite extensive progress in our understanding of WGD, the role of these events in promoting diversification is still not well understood. We seek to clarify the possible association between WGD and diversification rates in flowering plants. METHODS: Using a previously published phylogeny spanning all land plants (31,749 tips) and WGD events inferred from analyses of the 1000 Plants (1KP) transcriptome data, we analyzed the association of WGDs and diversification rates following numerous WGD events across the angiosperms. We used a stepwise AIC approach (MEDUSA), a Bayesian mixture model approach (BAMM), and state-dependent diversification analyses (MuSSE) to investigate patterns of diversification. Sister-clade comparisons were used to investigate species richness after WGDs. KEY RESULTS: Based on the density of 1KP taxon sampling, 106 WGDs were unambiguously placed on the angiosperm phylogeny. We identified 334-530 shifts in diversification rates. We found that 61 WGD events were tightly linked to changes in diversification rates, and state-dependent diversification analyses indicated higher speciation rates for subsequent rounds of WGD. Additionally, 70 of 99 WGD events showed an increase in species richness compared to the sister clade. CONCLUSIONS: Forty-six of the 106 WGDs analyzed appear to be closely associated with upshifts in the rate of diversification in angiosperms. Shifts in diversification do not appear more likely than random within a four-node lag phase following a WGD; however, younger WGD events are more likely to be followed by an upshift in diversification than older WGD events.

Demographic model selection using random forests and the site frequency spectrum.

Phylogeographic data sets have grown from tens to thousands of loci in recent years, but extant statistical methods do not take full advantage of these large data sets. For example, approximate Bayesian computation (ABC) is a commonly used method for the explicit comparison of alternate demographic histories, but it is limited by the "curse of dimensionality" and issues related to the simulation and summarization of data when applied to next-generation sequencing (NGS) data sets. We implement here several improvements to overcome these difficulties. We use a Random Forest (RF) classifier for model selection to circumvent the curse of dimensionality and apply a binned representation of the multidimensional site frequency spectrum (mSFS) to address issues related to the simulation and summarization of large SNP data sets. We evaluate the performance of these improvements using simulation and find low overall error rates (~7%). We then apply the approach to data from Haplotrema vancouverense, a land snail endemic to the Pacific Northwest of North America. Fifteen demographic models were compared, and our results support a model of recent dispersal from coastal to inland rainforests. Our results demonstrate that binning is an effective strategy for the construction of a mSFS and imply that the statistical power of RF when applied to demographic model selection is at least comparable to traditional ABC algorithms. Importantly, by combining these strategies, large sets of models with differing numbers of populations can be evaluated.

Identifying cryptic diversity with predictive phylogeography.

Identifying units of biological diversity is a major goal of organismal biology. An increasing literature has focused on the importance of cryptic diversity, defined as the presence of deeply diverged lineages within a single species. While most discoveries of cryptic lineages proceed on a taxon-by-taxon basis, rapid assessments of biodiversity are needed to inform conservation policy and decision-making. Here, we introduce a predictive framework for phylogeography that allows rapidly identifying cryptic diversity. Our approach proceeds by collecting environmental, taxonomic and genetic data from codistributed taxa with known phylogeographic histories. We define these taxa as a reference set, and categorize them as either harbouring or lacking cryptic diversity. We then build a random forest classifier that allows us to predict which other taxa endemic to the same biome are likely to contain cryptic diversity. We apply this framework to data from two sets of disjunct ecosystems known to harbour taxa with cryptic diversity: the mesic temperate forests of the Pacific Northwest of North America and the arid lands of Southwestern North America. The predictive approach presented here is accurate, with prediction accuracies placed between 65% and 98.79% depending of the ecosystem. This seems to indicate that our method can be successfully used to address ecosystem-level questions about cryptic diversity. Further, our application for the prediction of the cryptic/non-cryptic nature of unknown species is easily applicable and provides results that agree with recent discoveries from those systems. Our results demonstrate that the transition of phylogeography from a descriptive to a predictive discipline is possible and effective.

Oligocene niche shift, Miocene diversification - cold tolerance and accelerated speciation rates in the St. John's Worts (Hypericum, Hypericaceae).

BACKGROUND: Our aim is to understand the evolution of species-rich plant groups that shifted from tropical into cold/temperate biomes. It is well known that climate affects evolutionary processes, such as how fast species diversify, species range shifts, and species distributions. Many plant lineages may have gone extinct in the Northern Hemisphere due to Late Eocene climate cooling, while some tropical lineages may have adapted to temperate conditions and radiated; the hyper-diverse and geographically widespread genus Hypericum is one of these. RESULTS: To investigate the effect of macroecological niche shifts on evolutionary success we combine historical biogeography with analyses of diversification dynamics and climatic niche shifts in a phylogenetic framework. Hypericum evolved cold tolerance c. 30 million years ago, and successfully colonized all ice-free continents, where today ~500 species exist. The other members of Hypericaceae stayed in their tropical habitats and evolved into ~120 species. We identified a 15-20 million year lag between the initial change in temperature preference in Hypericum and subsequent diversification rate shifts in the Miocene. CONCLUSIONS: Contrary to the dramatic niche shift early in the evolution of Hypericum most extant species occur in temperate climates including high elevations in the tropics. These cold/temperate niches are a distinctive characteristic of Hypericum. We conclude that the initial release from an evolutionary constraint (from tropical to temperate climates) is an important novelty in Hypericum. However, the initial shift in the adaptive landscape into colder climates appears to be a precondition, and may not be directly related to increased diversification rates. Instead, subsequent events of mountain formation and further climate cooling may better explain distribution patterns and species-richness in Hypericum. These findings exemplify important macroevolutionary patterns of plant diversification during large-scale global climate change.

Nested radiations and the pulse of angiosperm diversification: increased diversification rates often follow whole genome duplications.

Our growing understanding of the plant tree of life provides a novel opportunity to uncover the major drivers of angiosperm diversity. Using a time-calibrated phylogeny, we characterized hot and cold spots of lineage diversification across the angiosperm tree of life by modeling evolutionary diversification using stepwise AIC (MEDUSA). We also tested the whole-genome duplication (WGD) radiation lag-time model, which postulates that increases in diversification tend to lag behind established WGD events. Diversification rates have been incredibly heterogeneous throughout the evolutionary history of angiosperms and reveal a pattern of 'nested radiations' - increases in net diversification nested within other radiations. This pattern in turn generates a negative relationship between clade age and diversity across both families and orders. We suggest that stochastically changing diversification rates across the phylogeny explain these patterns. Finally, we demonstrate significant statistical support for the WGD radiation lag-time model. Across angiosperms, nested shifts in diversification led to an overall increasing rate of net diversification and declining relative extinction rates through time. These diversification shifts are only rarely perfectly associated with WGD events, but commonly follow them after a lag period.

Shifts in diversification rates linked to biogeographic movement into new areas: An example of a recent radiation in the Andes.

PREMISE OF THE STUDY: Clade-specific bursts in diversification are often associated with the evolution of key innovations. However, in groups with no obvious morphological innovations, observed upticks in diversification rates have also been attributed to the colonization of a new geographic environment. In this study, we explore the systematics, diversification dynamics, and historical biogeography of the plant clade Rhinantheae in the Orobanchaceae, with a special focus on the Andean clade of the genus Bartsia. METHODS: We sampled taxa from across Rhinantheae, including a representative sample of Andean Bartsia species. Using standard phylogenetic methods, we reconstructed evolutionary relationships, inferred divergence times among the clades of Rhinantheae, elucidated their biogeographic history, and investigated diversification dynamics. KEY RESULTS: We confirmed that the South American Bartsia species form a highly supported monophyletic group. The median crown age of Rhinantheae was determined to be ca. 30 Myr, and Europe played an important role in the biogeographic history of the lineages. South America was first reconstructed in the biogeographic analyses around 9 Myr ago, and with a median age of 2.59 Myr, this clade shows a significant uptick in diversification. CONCLUSIONS: Increased net diversification of the South American clade corresponds to biogeographic movement into the New World. This movement happened at a time when the Andes were reaching the necessary elevation to host an alpine environment. Although a specific route could not be identified with certainty, we provide plausible hypotheses to how the group colonized the New World.